David A. Relman, MD is the Thomas C. and Joan M. Merigan Professor in Medicine, and Professor of Microbiology & Immunology, and Senior Fellow at the Freeman Spogli Institute for International Studies at Stanford University. He is also Chief of Infectious Diseases at the Veterans Affairs Palo Alto Health Care System in Palo Alto, California. Dr. Relman?s research focus is the human indigenous microbiota and on diversity, assembly, stability, and resilience of the human microbial ecosystem, as well as the identification of previously-unrecognized microbial agents of disease. He has advised the U.S. Government on emerging infectious diseases, human-microbe interactions, and future biological threats. He is a member of the Board of Scientific Counselors at the National Center for Biotechnology Information (NCBI), NIH, a member of the Intelligence Community Studies Board at the National Academies of Science, Engineering and Medicine, and Past President of the Infectious Diseases Society of America. He is a Fellow of the American Academy of Microbiology, and a Member of the National Academy of Medicine.

Clinical Focus

  • Infectious Disease

Administrative Appointments

  • Member, Center for Computational, Evolutionary and Human Genomics (2018 - Present)
  • Faculty Fellow, Center for Innovation in Global Health, Stanford University (2015 - Present)
  • Senior Fellow, Freeman Spogli Institute for International Studies, Stanford University (2013 - Present)
  • Chief, Infectious Diseases Section, VA Palo Alto Health Care System (2002 - Present)
  • Chair, Administrative Panel on Biosafety, Stanford University (2001 - 2018)
  • Co-Director, Center for International Security and Cooperation (CISAC), Stanford University (2013 - 2017)

Honors & Awards

  • NIH Transformative Research Award, NIH (2013)
  • Member, Institute of Medicine, National Academies of Science (2011-)
  • Fellow, American Association for the Advancement of Science (2010)
  • Thomas C. and Joan M. Merigan Professor, Stanford University (2009-)
  • Distinguished Clinical Scientist Award, Doris Duke Charitable Foundation (2006)
  • NIH Director's Pioneer Award, NIH (2006)
  • Kinyoun Lecturer, NIAID/NIH (2005)
  • Fellow, American Academy of Microbiology (2003)
  • Senior Scholar Award in Global Infectious Diseases, Ellison Medical Foundation (2002)
  • Squibb Award, Infectious Diseases Society of America (2001)

Boards, Advisory Committees, Professional Organizations

  • Member, Intelligence Community Studies Board, National Academies of Science, Engineering, and Medicine (2016 - Present)
  • Member (Chair, 2018-2019), Board of Scientific Counselors, National Center for Biotechnology Information, NIH (2015 - Present)
  • Member, Working Group on Biodefense President?s Council of Advisors on Science and Technology, The White House (2015 - 2016)
  • Member, Working Group on Antimicrobial Resistance, President?s Council of Advisors on Science and Technology (2014 - 2014)
  • Member, Committee on Science, Technology and Law, National Academy of Science (2012 - 2017)
  • President, Infectious Diseases Society of America (2012 - 2013)
  • Chair, Forum on Microbial Threats, Institute of Medicine (National Academies of Science) (2007 - 2017)
  • Member, Science, Technology, and Engineering Advisory Panel, Lawrence Livermore National Laboratory (2007 - 2017)
  • Member, National Science Advisory Board for Biosecurity (2005 - 2014)

Professional Education

  • Fellowship:Stanford University School of Medicine Registrar (1988) CA
  • Board Certification: Infectious Disease, American Board of Internal Medicine (1988)
  • Fellowship:Massachusetts General Hospital (1986) MA
  • Internship:Massachusetts General Hospital (1986) MA
  • Board Certification: Internal Medicine, American Board of Internal Medicine (1985)
  • Residency:Massachusetts General Hospital (1985) MA
  • Medical Education:Harvard Medical School (1982) MA
  • S.B., M.I.T., Biology (1977)
  • M.D., Harvard Medical School, Medicine (1982)

Community and International Work

  • Chief Medical Officer, Rock Medicine, Haight-Ashbury Medical Clinics


    Free medical care at concerts and large public events

    Partnering Organization(s)

    Haight-Ashbury Medical Clinics



    Ongoing Project


    Opportunities for Student Involvement


Research & Scholarship

Current Research and Scholarly Interests

My primary research focus is the human indigenous microbiota (microbiome), and in particular, the nature and mechanisms of variation in patterns of microbial diversity within the human body as a function of time (microbial succession), space (biogeography within the host landscape), and in response to perturbation, e.g., antibiotics (community robustness and resilience). One of the goals of this work is to define the role of the human microbiome in health and disease. We are particularly interested in measuring and understanding resilience in the human microbial ecosystem. Our work includes the human oral cavity, gut, and female reproductive tract, as well as an analysis of microbial diversity in marine mammals. This research integrates theory and methods from ecology, population biology, environmental microbiology, genomics and clinical medicine.

During the past few decades, my research directions have also included pathogen discovery and the development of new strategies for identifying previously-unrecognized microbial agents of disease. This work has included the use of host gene expression response patterns to recognize and understand early stages of systemic infection. Currently, we are examining genomic patterns of host response in dengue fever and in cases of undiagnosed febrile illness, for diagnostic and prognostic purposes, as well as to understand better disease mechanism.


2017-18 Courses

Stanford Advisees


All Publications

  • A spatial gradient of bacterial diversity in the human oral cavity shaped by salivary flow. Nature communications Proctor, D. M., Fukuyama, J. A., Loomer, P. M., Armitage, G. C., Lee, S. A., Davis, N. M., Ryder, M. I., Holmes, S. P., Relman, D. A. 2018; 9 (1): 681


    Spatial and temporal patterns in microbial communities provide insights into the forces that shape them, their functions and roles in health and disease. Here, we used spatial and ecological statistics to analyze the role that saliva plays in structuring bacterial communities of the human mouth using >9000 dental and mucosal samples. We show that regardless of tissue type (teeth, alveolar mucosa, keratinized gingiva, or buccal mucosa), surface-associated bacterial communities vary along an ecological gradient from the front to the back of the mouth, and that on exposed tooth surfaces, the gradient is pronounced on lingual compared to buccal surfaces. Furthermore, our data suggest that this gradient is attenuated in individuals with low salivary flow due to Sjögren's syndrome. Taken together, our findings imply that salivary flow influences the spatial organization of microbial communities and that biogeographical patterns may be useful for understanding host physiological processes and for predicting disease.

    View details for DOI 10.1038/s41467-018-02900-1

    View details for PubMedID 29445174

    View details for PubMedCentralID PMC5813034

  • Clostridium difficile, Aging, and the Gut: Can Microbiome Rejuvenation Keep Us Young and Healthy? The Journal of infectious diseases Fischer, N., Relman, D. A. 2018; 217 (2): 174?76

    View details for DOI 10.1093/infdis/jix417

    View details for PubMedID 28968708

  • Enterotypes in the landscape of gut microbial community composition. Nature microbiology Costea, P. I., Hildebrand, F., Arumugam, M., Bäckhed, F., Blaser, M. J., Bushman, F. D., de Vos, W. M., Ehrlich, S. D., Fraser, C. M., Hattori, M., Huttenhower, C., Jeffery, I. B., Knights, D., Lewis, J. D., Ley, R. E., Ochman, H., O'Toole, P. W., Quince, C., Relman, D. A., Shanahan, F., Sunagawa, S., Wang, J., Weinstock, G. M., Wu, G. D., Zeller, G., Zhao, L., Raes, J., Knight, R., Bork, P. 2018; 3 (1): 8?16


    Population stratification is a useful approach for a better understanding of complex biological problems in human health and wellbeing. The proposal that such stratification applies to the human gut microbiome, in the form of distinct community composition types termed enterotypes, has been met with both excitement and controversy. In view of accumulated data and re-analyses since the original work, we revisit the concept of enterotypes, discuss different methods of dividing up the landscape of possible microbiome configurations, and put these concepts into functional, ecological and medical contexts. As enterotypes are of use in describing the gut microbial community landscape and may become relevant in clinical practice, we aim to reconcile differing views and encourage a balanced application of the concept.

    View details for DOI 10.1038/s41564-017-0072-8

    View details for PubMedID 29255284

    View details for PubMedCentralID PMC5832044

  • Role of priority effects in the early-life assembly of the gut microbiota. Nature reviews. Gastroenterology & hepatology Sprockett, D., Fukami, T., Relman, D. A. 2018; 15 (4): 197?205


    Understanding how microbial communities develop is essential for predicting and directing their future states. Ecological theory suggests that community development is often influenced by priority effects, in which the order and timing of species arrival determine how species affect one another. Priority effects can have long-lasting consequences, particularly if species arrival history varies during the early stage of community development, but their importance to the human gut microbiota and host health remains largely unknown. Here, we explore how priority effects might influence microbial communities in the gastrointestinal tract during early childhood and how the strength of priority effects can be estimated from the composition of the microbial species pool. We also discuss factors that alter microbial transmission, such as delivery mode, diet and parenting behaviours such as breastfeeding, which can influence the likelihood of priority effects. An improved knowledge of priority effects has the potential to inform microorganism-based therapies, such as prebiotics and probiotics, which are aimed at guiding the microbiota towards a healthy state.

    View details for DOI 10.1038/nrgastro.2017.173

    View details for PubMedID 29362469

  • The Landscape Ecology and Microbiota of the Human Nose, Mouth, and Throat CELL HOST & MICROBE Proctor, D. M., Relman, D. A. 2017; 21 (4): 421-432


    Landscape ecology examines the relationships between the spatial arrangement of different landforms and the processes that give rise to spatial and temporal patterns in local community structure. The spatial ecology of the microbial communities that inhabit the human body-in particular, those of the nose, mouth, and throat-deserves greater attention. Important questions include what defines the size of a population (i.e., "patch") in a given body site, what defines the boundaries of distinct patches within a single body site, and where and over what spatial scales within a body site are gradients detected. This Review looks at the landscape ecology of the upper respiratory tract and mouth and seeks greater clarity about the physiological factors-whether immunological, chemical, or physical-that govern microbial community composition and function and the ecological traits that underlie health and disease.

    View details for DOI 10.1016/j.chom.2017.03.011

    View details for Web of Science ID 000398896100004

    View details for PubMedID 28407480

  • A geographically-diverse collection of 418 human gut microbiome pathway genome databases SCIENTIFIC DATA Hahn, A. S., Altman, T., Konwar, K. M., Hanson, N. W., Kim, D., Relman, D. A., Dill, D. L., Hallam, S. J. 2017; 4


    Advances in high-throughput sequencing are reshaping how we perceive microbial communities inhabiting the human body, with implications for therapeutic interventions. Several large-scale datasets derived from hundreds of human microbiome samples sourced from multiple studies are now publicly available. However, idiosyncratic data processing methods between studies introduce systematic differences that confound comparative analyses. To overcome these challenges, we developed GutCyc, a compendium of environmental pathway genome databases (ePGDBs) constructed from 418 assembled human microbiome datasets using MetaPathways, enabling reproducible functional metagenomic annotation. We also generated metabolic network reconstructions for each metagenome using the Pathway Tools software, empowering researchers and clinicians interested in visualizing and interpreting metabolic pathways encoded by the human gut microbiome. For the first time, GutCyc provides consistent annotations and metabolic pathway predictions, making possible comparative community analyses between health and disease states in inflammatory bowel disease, Crohn's disease, and type 2 diabetes. GutCyc data products are searchable online, or may be downloaded and explored locally using MetaPathways and Pathway Tools.

    View details for DOI 10.1038/sdata.2017.35

    View details for Web of Science ID 000400150900001

    View details for PubMedID 28398290

    View details for PubMedCentralID PMC5387927

  • Candidatus Mycoplasma girerdii replicates, diversifies, and co-occurs with Trichomonas vaginalis in the oral cavity of a premature infant. Scientific reports Costello, E. K., Sun, C. L., Carlisle, E. M., Morowitz, M. J., Banfield, J. F., Relman, D. A. 2017; 7 (1): 3764


    Genital mycoplasmas, which can be vertically transmitted, have been implicated in preterm birth, neonatal infections, and chronic lung disease of prematurity. Our prior work uncovered 16S rRNA genes belonging to a novel, as-yet-uncultivated mycoplasma (lineage 'Mnola') in the oral cavity of a premature neonate. Here, we characterize the organism's associated community, growth status, metabolic potential, and population diversity. Sequencing of genomic DNA from the infant's saliva yielded 1.44 Gbp of high-quality, non-human read data, from which we recovered three essentially complete (including 'Mnola') and three partial draft genomes (including Trichomonas vaginalis). The completed 629,409-bp 'Mnola' genome (Candidatus Mycoplasma girerdii str. UC-B3) was distinct at the strain level from its closest relative, vaginally-derived Ca. M. girerdii str. VCU-M1, which is also associated with T. vaginalis. Replication rate measurements indicated growth of str. UC-B3 within the infant. Genes encoding surface-associated proteins and restriction-modification systems were especially diverse within and between strains. In UC-B3, the population genetic underpinnings of phase variable expression were evident in vivo. Unique among mycoplasmas, Ca. M. girerdii encodes pyruvate-ferredoxin oxidoreductase and may be sensitive to metronidazole. This study reveals a metabolically unique mycoplasma colonizing a premature neonate, and establishes the value of genome-resolved metagenomics in tracking phase variation.

    View details for DOI 10.1038/s41598-017-03821-7

    View details for PubMedID 28630471

    View details for PubMedCentralID PMC5476646

  • The Human Virome - Implications for Clinical Practice in Transplantation Medicine. Journal of clinical microbiology Tan, S. K., Relman, D. A., Pinsky, B. A. 2017


    Advances in DNA sequencing technology have provided an unprecedented opportunity to study the human virome. Transplant recipients and other immunocompromised hosts are at particular risk for developing virus-related pathology; thus, the impact of the virome on health and disease may be even more relevant in this population. Here we discuss technical considerations in studying the human virome, the current literature on the virome in transplant recipients, and near future applications of sequence-based findings that can further our understanding of viruses in transplantation medicine.

    View details for DOI 10.1128/JCM.00489-17

    View details for PubMedID 28724557

  • On Defining Global Catastrophic Biological Risks. Health security Palmer, M. J., Tiu, B. C., Weissenbach, A. S., Relman, D. A. 2017; 15 (4): 347?48

    View details for DOI 10.1089/hs.2017.0057

    View details for PubMedID 28737976

    View details for PubMedCentralID PMC5576069

  • Multidomain analyses of a longitudinal human microbiome intestinal cleanout perturbation experiment. PLoS computational biology Fukuyama, J., Rumker, L., Sankaran, K., Jeganathan, P., Dethlefsen, L., Relman, D. A., Holmes, S. P. 2017; 13 (8): e1005706


    Our work focuses on the stability, resilience, and response to perturbation of the bacterial communities in the human gut. Informative flash flood-like disturbances that eliminate most gastrointestinal biomass can be induced using a clinically-relevant iso-osmotic agent. We designed and executed such a disturbance in human volunteers using a dense longitudinal sampling scheme extending before and after induced diarrhea. This experiment has enabled a careful multidomain analysis of a controlled perturbation of the human gut microbiota with a new level of resolution. These new longitudinal multidomain data were analyzed using recently developed statistical methods that demonstrate improvements over current practices. By imposing sparsity constraints we have enhanced the interpretability of the analyses and by employing a new adaptive generalized principal components analysis, incorporated modulated phylogenetic information and enhanced interpretation through scoring of the portions of the tree most influenced by the perturbation. Our analyses leverage the taxa-sample duality in the data to show how the gut microbiota recovers following this perturbation. Through a holistic approach that integrates phylogenetic, metagenomic and abundance information, we elucidate patterns of taxonomic and functional change that characterize the community recovery process across individuals. We provide complete code and illustrations of new sparse statistical methods for high-dimensional, longitudinal multidomain data that provide greater interpretability than existing methods.

    View details for DOI 10.1371/journal.pcbi.1005706

    View details for PubMedID 28821012

    View details for PubMedCentralID PMC5576755

  • Long-term taxonomic and functional divergence from donor bacterial strains following fecal microbiota transplantation in immunocompromised patients. PloS one Moss, E. L., Falconer, S. B., Tkachenko, E., Wang, M., Systrom, H., Mahabamunuge, J., Relman, D. A., Hohmann, E. L., Bhatt, A. S. 2017; 12 (8): e0182585


    Immunocompromised individuals are at high risk of developing Clostridium difficile-associated disease (CDAD). Fecal microbiota transplantation (FMT) is a highly effective therapy for refractory or recurrent CDAD and, despite safety concerns, has recently been offered to immunocompromised patients. We investigated the genomics of bacterial composition following FMT in immunocompromised patients over a 1-year period. Metagenomic, strain and gene-level bacterial dynamics were characterized in two CDAD-affected hematopoietic stem cell (HCT) recipients following FMT. We found alterations in gene content, including loss of virulence and antibiotic resistance genes. These alterations were accompanied by long-term bacterial divergence at the species and strain levels. Our findings suggest limited durability of the specific bacterial consortium introduced with FMT and indicate that alterations of the functional potential of the microbiome are more complex than can be inferred by taxonomic information alone. Our observation that FMT alone cannot induce long-term donor-like alterations of the microbiota of HCT recipients suggests that FMT cannot indefinitely supersede environmental and/or host factors in shaping bacterial composition.

    View details for DOI 10.1371/journal.pone.0182585

    View details for PubMedID 28827811

    View details for PubMedCentralID PMC5565110

  • Replication and refinement of a vaginal microbial signature of preterm birth in two racially distinct cohorts of US women. Proceedings of the National Academy of Sciences of the United States of America Callahan, B. J., DiGiulio, D. B., Goltsman, D. S., Sun, C. L., Costello, E. K., Jeganathan, P., Biggio, J. R., Wong, R. J., Druzin, M. L., Shaw, G. M., Stevenson, D. K., Holmes, S. P., Relman, D. A. 2017


    Preterm birth (PTB) is the leading cause of neonatal morbidity and mortality. Previous studies have suggested that the maternal vaginal microbiota contributes to the pathophysiology of PTB, but conflicting results in recent years have raised doubts. We conducted a study of PTB compared with term birth in two cohorts of pregnant women: one predominantly Caucasian (n = 39) at low risk for PTB, the second predominantly African American and at high-risk (n = 96). We profiled the taxonomic composition of 2,179 vaginal swabs collected prospectively and weekly during gestation using 16S rRNA gene sequencing. Previously proposed associations between PTB and lower Lactobacillus and higher Gardnerella abundances replicated in the low-risk cohort, but not in the high-risk cohort. High-resolution bioinformatics enabled taxonomic assignment to the species and subspecies levels, revealing that Lactobacillus crispatus was associated with low risk of PTB in both cohorts, while Lactobacillus iners was not, and that a subspecies clade of Gardnerella vaginalis explained the genus association with PTB. Patterns of cooccurrence between L. crispatus and Gardnerella were highly exclusive, while Gardnerella and L. iners often coexisted at high frequencies. We argue that the vaginal microbiota is better represented by the quantitative frequencies of these key taxa than by classifying communities into five community state types. Our findings extend and corroborate the association between the vaginal microbiota and PTB, demonstrate the benefits of high-resolution statistical bioinformatics in clinical microbiome studies, and suggest that previous conflicting results may reflect the different risk profile of women of black race.

    View details for DOI 10.1073/pnas.1705899114

    View details for PubMedID 28847941

  • Childhood tuberculosis is associated with decreased abundance of T cell gene transcripts and impaired T cell function. PloS one Hemingway, C., Berk, M., Anderson, S. T., Wright, V. J., Hamilton, S., Eleftherohorinou, H., Kaforou, M., Goldgof, G. M., Hickman, K., Kampmann, B., Schoeman, J., Eley, B., Beatty, D., Pienaar, S., Nicol, M. P., Griffiths, M. J., Waddell, S. J., Newton, S. M., Coin, L. J., Relman, D. A., Montana, G., Levin, M. 2017; 12 (11): e0185973


    The WHO estimates around a million children contract tuberculosis (TB) annually with over 80 000 deaths from dissemination of infection outside of the lungs. The insidious onset and association with skin test anergy suggests failure of the immune system to both recognise and respond to infection. To understand the immune mechanisms, we studied genome-wide whole blood RNA expression in children with TB meningitis (TBM). Findings were validated in a second cohort of children with TBM and pulmonary TB (PTB), and functional T-cell responses studied in a third cohort of children with TBM, other extrapulmonary TB (EPTB) and PTB. The predominant RNA transcriptional response in children with TBM was decreased abundance of multiple genes, with 140/204 (68%) of all differentially regulated genes showing reduced abundance compared to healthy controls. Findings were validated in a second cohort with concordance of the direction of differential expression in both TBM (r2 = 0.78 p = 2x10-16) and PTB patients (r2 = 0.71 p = 2x10-16) when compared to a second group of healthy controls. Although the direction of expression of these significant genes was similar in the PTB patients, the magnitude of differential transcript abundance was less in PTB than in TBM. The majority of genes were involved in activation of leucocytes (p = 2.67E-11) and T-cell receptor signalling (p = 6.56E-07). Less abundant gene expression in immune cells was associated with a functional defect in T-cell proliferation that recovered after full TB treatment (p<0.0003). Multiple genes involved in T-cell activation show decreased abundance in children with acute TB, who also have impaired functional T-cell responses. Our data suggest that childhood TB is associated with an acquired immune defect, potentially resulting in failure to contain the pathogen. Elucidation of the mechanism causing the immune paresis may identify new treatment and prevention strategies.

    View details for DOI 10.1371/journal.pone.0185973

    View details for PubMedID 29140996

    View details for PubMedCentralID PMC5687722

  • Microbiota: A high-pressure situation for bacteria. Nature Relman, D. A. 2017; 551 (7682): 571?72

    View details for DOI 10.1038/nature24760

    View details for PubMedID 29143820

  • Novel Microbial Diversity and Functional Potential in the Marine Mammal Oral Microbiome. Current biology : CB Dudek, N. K., Sun, C. L., Burstein, D., Kantor, R. S., Aliaga Goltsman, D. S., Bik, E. M., Thomas, B. C., Banfield, J. F., Relman, D. A. 2017; 27 (24): 3752?62.e6


    The vast majority of bacterial diversity lies within phylum-level lineages called "candidate phyla," which lack isolated representatives and are poorly understood. These bacteria are surprisingly abundant in the oral cavity of marine mammals. We employed a genome-resolved metagenomic approach to recover and characterize genomes and functional potential from microbes in the oral gingival sulcus of two bottlenose dolphins (Tursiops truncatus). We detected organisms from 24 known bacterial phyla and one archaeal phylum. We also recovered genomes from two deep-branching, previously uncharacterized phylum-level lineages (here named "Candidatus Delphibacteria" and "Candidatus Fertabacteria"). The Delphibacteria lineage is found in both managed and wild dolphins; its metabolic profile suggests a capacity for denitrification and a possible role in dolphin health. We uncovered a rich diversity of predicted Cas9 proteins, including the two longest predicted Cas9 proteins to date. Notably, we identified the first type II CRISPR-Cas systems encoded by members of the Candidate Phyla Radiation. Using their spacer sequences, we subsequently identified and assembled a complete Saccharibacteria phage genome. These findings underscore the immense microbial diversity and functional potential that await discovery in previously unexplored environments.

    View details for DOI 10.1016/j.cub.2017.10.040

    View details for PubMedID 29153320

  • Risky Business: Meeting the Structural Needs of Transdisciplinary Science. The Journal of pediatrics Wise, P. H., Shaw, G. M., Druzin, M. L., Darmstadt, G. L., Quaintance, C., Mäkinen, E., Relman, D. A., Quake, S. R., Butte, A. J., Angst, M. S., Muglia, L. J., Macones, G., Driscoll, D., Ober, C., Simpson, J. L., Katz, M., Howse, J., Stevenson, D. K. 2017; 191: 255?58

    View details for DOI 10.1016/j.jpeds.2017.08.072

    View details for PubMedID 29173314

  • Cathelicidin Insufficiency in Patients with Fatal Leptospirosis. PLoS pathogens Lindow, J. C., Wunder, E. A., Popper, S. J., Min, J., Mannam, P., Srivastava, A., Yao, Y., Hacker, K. P., Raddassi, K., Lee, P. J., Montgomery, R. R., Shaw, A. C., Hagan, J. E., Araújo, G. C., Nery, N., Relman, D. A., Kim, C. C., Reis, M. G., Ko, A. I. 2016; 12 (11)


    Leptospirosis causes significant morbidity and mortality worldwide; however, the role of the host immune response in disease progression and high case fatality (>10-50%) is poorly understood. We conducted a multi-parameter investigation of patients with acute leptospirosis to identify mechanisms associated with case fatality. Whole blood transcriptional profiling of 16 hospitalized Brazilian patients with acute leptospirosis (13 survivors, 3 deceased) revealed fatal cases had lower expression of the antimicrobial peptide, cathelicidin, and chemokines, but more abundant pro-inflammatory cytokine receptors. In contrast, survivors generated strong adaptive immune signatures, including transcripts relevant to antigen presentation and immunoglobulin production. In an independent cohort (23 survivors, 22 deceased), fatal cases had higher bacterial loads (P = 0.0004) and lower anti-Leptospira antibody titers (P = 0.02) at the time of hospitalization, independent of the duration of illness. Low serum cathelicidin and RANTES levels during acute illness were independent risk factors for higher bacterial loads (P = 0.005) and death (P = 0.04), respectively. To investigate the mechanism of cathelicidin in patients surviving acute disease, we administered LL-37, the active peptide of cathelicidin, in a hamster model of lethal leptospirosis and found it significantly decreased bacterial loads and increased survival. Our findings indicate that the host immune response plays a central role in severe leptospirosis disease progression. While drawn from a limited study size, significant conclusions include that poor clinical outcomes are associated with high systemic bacterial loads, and a decreased antibody response. Furthermore, our data identified a key role for the antimicrobial peptide, cathelicidin, in mounting an effective bactericidal response against the pathogen, which represents a valuable new therapeutic approach for leptospirosis.

    View details for DOI 10.1371/journal.ppat.1005943

    View details for PubMedID 27812211

    View details for PubMedCentralID PMC5094754

  • A microbial perspective of human developmental biology NATURE Charbonneau, M. R., Blanton, L. V., DiGiulio, D. B., Relman, D. A., Lebrilla, C. B., Mills, D. A., Gordon, J. I. 2016; 535 (7610): 48-55


    When most people think of human development, they tend to consider only human cells and organs. Yet there is another facet that involves human-associated microbial communities. A microbial perspective of human development provides opportunities to refine our definitions of healthy prenatal and postnatal growth and to develop innovative strategies for disease prevention and treatment. Given the dramatic changes in lifestyles and disease patterns that are occurring with globalization, we issue a call for the establishment of 'human microbial observatories' designed to examine microbial community development in birth cohorts representing populations with diverse anthropological characteristics, including those undergoing rapid change.

    View details for DOI 10.1038/nature18845

    View details for Web of Science ID 000379015600027

    View details for PubMedID 27383979

  • Early Transcriptional Signatures of the Immune Response to a Live Attenuated Tetravalent Dengue Vaccine Candidate in Non-human Primates PLOS NEGLECTED TROPICAL DISEASES Strouts, F. R., Popper, S. J., Partidos, C. D., Stinchcomb, D. T., Osorio, J. E., Relman, D. A. 2016; 10 (5)


    The development of a vaccine against dengue faces unique challenges, including the complexity of the immune responses to the four antigenically distinct serotypes. Genome-wide transcriptional profiling provides insight into the pathways and molecular features that underlie responses to immune system stimulation, and may facilitate predictions of immune protection.In this study, we measured early transcriptional responses in the peripheral blood of cynomolgus macaques following vaccination with a live, attenuated tetravalent dengue vaccine candidate, TDV, which is based on a DENV-2 backbone. Different doses and routes of vaccine administration were used, and viral load and neutralizing antibody titers were measured at different time-points following vaccination. All 30 vaccinated animals developed a neutralizing antibody response to each of the four dengue serotypes, and only 3 of these animals had detectable serum viral RNA after challenge with wild-type dengue virus (DENV), suggesting protection of vaccinated animals to DENV infection. The vaccine induced statistically significant changes in 595 gene transcripts on days 1, 3, 5 and 7 as compared with baseline and placebo-treated animals. Genes involved in the type I interferon (IFN) response, including IFI44, DDX58, MX1 and OASL, exhibited the highest fold-change in transcript abundance, and this response was strongest following double dose and subcutaneous (versus intradermal) vaccine administration. In addition, modules of genes involved in antigen presentation, dendritic cell activation, and T cell activation and signaling were enriched following vaccination. Increased abundance of gene transcripts related to T cell activation on day 5, and the type I IFN response on day 7, were significantly correlated with the development of high neutralizing antibody titers on day 30.These results suggest that early transcriptional responses may be predictive of development of adaptive immunity to TDV vaccination in cynomolgus macaques, and will inform studies of human responses to dengue vaccines.

    View details for DOI 10.1371/journal.pntd.0004731

    View details for Web of Science ID 000377769300067

    View details for PubMedID 27214236

    View details for PubMedCentralID PMC4877054

  • Marine mammals harbor unique microbiotas shaped by and yet distinct from the sea. Nature communications Bik, E. M., Costello, E. K., Switzer, A. D., Callahan, B. J., Holmes, S. P., Wells, R. S., Carlin, K. P., Jensen, E. D., Venn-Watson, S., Relman, D. A. 2016; 7: 10516-?


    Marine mammals play crucial ecological roles in the oceans, but little is known about their microbiotas. Here we study the bacterial communities in 337 samples from 5 body sites in 48 healthy dolphins and 18 healthy sea lions, as well as those of adjacent seawater and other hosts. The bacterial taxonomic compositions are distinct from those of other mammals, dietary fish and seawater, are highly diverse and vary according to body site and host species. Dolphins harbour 30 bacterial phyla, with 25 of them in the mouth, several abundant but poorly characterized Tenericutes species in gastric fluid and a surprisingly paucity of Bacteroidetes in distal gut. About 70% of near-full length bacterial 16S ribosomal RNA sequences from dolphins are unique. Host habitat, diet and phylogeny all contribute to variation in marine mammal distal gut microbiota composition. Our findings help elucidate the factors structuring marine mammal microbiotas and may enhance monitoring of marine mammal health.

    View details for DOI 10.1038/ncomms10516

    View details for PubMedID 26839246

    View details for PubMedCentralID PMC4742810

  • A new view of the tree of life. Nature microbiology Hug, L. A., Baker, B. J., Anantharaman, K., Brown, C. T., Probst, A. J., Castelle, C. J., Butterfield, C. N., Hernsdorf, A. W., Amano, Y., Ise, K., Suzuki, Y., Dudek, N., Relman, D. A., Finstad, K. M., Amundson, R., Thomas, B. C., Banfield, J. F. 2016; 1: 16048-?


    The tree of life is one of the most important organizing principles in biology(1). Gene surveys suggest the existence of an enormous number of branches(2), but even an approximation of the full scale of the tree has remained elusive. Recent depictions of the tree of life have focused either on the nature of deep evolutionary relationships(3-5) or on the known, well-classified diversity of life with an emphasis on eukaryotes(6). These approaches overlook the dramatic change in our understanding of life's diversity resulting from genomic sampling of previously unexamined environments. New methods to generate genome sequences illuminate the identity of organisms and their metabolic capacities, placing them in community and ecosystem contexts(7,8). Here, we use new genomic data from over 1,000 uncultivated and little known organisms, together with published sequences, to infer a dramatically expanded version of the tree of life, with Bacteria, Archaea and Eukarya included. The depiction is both a global overview and a snapshot of the diversity within each major lineage. The results reveal the dominance of bacterial diversification and underline the importance of organisms lacking isolated representatives, with substantial evolution concentrated in a major radiation of such organisms. This tree highlights major lineages currently underrepresented in biogeochemical models and identifies radiations that are probably important for future evolutionary analyses.

    View details for DOI 10.1038/nmicrobiol.2016.48

    View details for PubMedID 27572647

  • REPRODUCIBLE RESEARCH WORKFLOW IN R FOR THE ANALYSIS OF PERSONALIZED HUMAN MICROBIOME DATA. Pacific Symposium on Biocomputing. Pacific Symposium on Biocomputing Callahan, B., Proctor, D., Relman, D., Fukuyama, J., Holmes, S. 2016; 21: 183-194


    This article presents a reproducible research workflow for amplicon-based microbiome studies in personalized medicine created using Bioconductor packages and the knitr markdown interface.We show that sometimes a multiplicity of choices and lack of consistent documentation at each stage of the sequential processing pipeline used for the analysis of microbiome data can lead to spurious results. We propose its replacement with reproducible and documented analysis using R packages dada2, knitr, and phyloseq. This workflow implements both key stages of amplicon analysis: the initial filtering and denoising steps needed to construct taxonomic feature tables from error-containing sequencing reads (dada2), and the exploratory and inferential analysis of those feature tables and associated sample metadata (phyloseq). This workow facilitates reproducible interrogation of the full set of choices required in microbiome studies. We present several examples in which we leverage existing packages for analysis in a way that allows easy sharing and modification by others, and give pointers to articles that depend on this reproducible workflow for the study of longitudinal and spatial series analyses of the vaginal microbiome in pregnancy and the oral microbiome in humans with healthy dentition and intra-oral tissues.

    View details for PubMedID 26776185

  • Variation in Taxonomic Composition of the Fecal Microbiota in an Inbred Mouse Strain across Individuals and Time PLOS ONE Hoy, Y. E., Bik, E. M., Lawley, T. D., Holmes, S. P., Monack, D. M., Theriot, J. A., Relman, D. A. 2015; 10 (11)
  • Temporal and spatial variation of the human microbiota during pregnancy PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA DiGiulio, D. B., Callahan, B. J., McMurdie, P. J., Costello, E. K., Lyell, D. J., Robaczewska, A., Sun, C. L., Goltsman, D. S., Wong, R. J., Shaw, G., Stevenson, D. K., Holmes, S. P., Relman, D. A. 2015; 112 (35): 11060-11065


    Despite the critical role of the human microbiota in health, our understanding of microbiota compositional dynamics during and after pregnancy is incomplete. We conducted a case-control study of 49 pregnant women, 15 of whom delivered preterm. From 40 of these women, we analyzed bacterial taxonomic composition of 3,767 specimens collected prospectively and weekly during gestation and monthly after delivery from the vagina, distal gut, saliva, and tooth/gum. Linear mixed-effects modeling, medoid-based clustering, and Markov chain modeling were used to analyze community temporal trends, community structure, and vaginal community state transitions. Microbiota community taxonomic composition and diversity remained remarkably stable at all four body sites during pregnancy (P > 0.05 for trends over time). Prevalence of a Lactobacillus-poor vaginal community state type (CST 4) was inversely correlated with gestational age at delivery (P = 0.0039). Risk for preterm birth was more pronounced for subjects with CST 4 accompanied by elevated Gardnerella or Ureaplasma abundances. This finding was validated with a set of 246 vaginal specimens from nine women (four of whom delivered preterm). Most women experienced a postdelivery disturbance in the vaginal community characterized by a decrease in Lactobacillus species and an increase in diverse anaerobes such as Peptoniphilus, Prevotella, and Anaerococcus species. This disturbance was unrelated to gestational age at delivery and persisted for up to 1 y. These findings have important implications for predicting premature labor, a major global health problem, and for understanding the potential impact of a persistent, altered postpartum microbiota on maternal health, including outcomes of pregnancies following short interpregnancy intervals.

    View details for DOI 10.1073/pnas.1502875112

    View details for Web of Science ID 000360383200068

  • Gain-of-function experiments: time for a real debate. Nature reviews. Microbiology Duprex, W. P., Fouchier, R. A., Imperiale, M. J., Lipsitch, M., Relman, D. A. 2015; 13 (1): 58-64


    According to the WHO, dual use research of concern (DURC) is "life sciences research that is intended for benefit, but which might easily be misapplied to do harm". Recent studies, particularly those on influenza viruses, have led to renewed attention on DURC, as there is an ongoing debate over whether the benefits of gain-of-function (GOF) experiments that result in an increase in the transmission and/or pathogenicity of potential pandemic pathogens (PPPs) are outweighed by concerns over biosecurity and biosafety. In this Viewpoint article, proponents and opponents of GOF experiments discuss the benefits and risks associated with these studies, as well as the implications of the current debate for the scientific community and the general public, and suggest how the current discussion should move forward.

    View details for DOI 10.1038/nrmicro3405

    View details for PubMedID 25482289

  • Variation in Taxonomic Composition of the Fecal Microbiota in an Inbred Mouse Strain across Individuals and Time. PloS one Hoy, Y. E., Bik, E. M., Lawley, T. D., Holmes, S. P., Monack, D. M., Theriot, J. A., Relman, D. A. 2015; 10 (11)


    Genetics, diet, and other environmental exposures are thought to be major factors in the development and composition of the intestinal microbiota of animals. However, the relative contributions of these factors in adult animals, as well as variation with time in a variety of important settings, are still not fully understood. We studied a population of inbred, female mice fed the same diet and housed under the same conditions. We collected fecal samples from 46 individual mice over two weeks, sampling four of these mice for periods as long as 236 days for a total of 190 samples, and determined the phylogenetic composition of their microbial communities after analyzing 1,849,990 high-quality pyrosequencing reads of the 16S rRNA gene V3 region. Even under these controlled conditions, we found significant inter-individual variation in community composition, as well as variation within an individual over time, including increases in alpha diversity during the first 2 months of co-habitation. Some variation was explained by mouse membership in different cage and vendor shipment groups. The differences among individual mice from the same shipment group and cage were still significant. Overall, we found that 23% of the variation in intestinal microbiota composition was explained by changes within the fecal microbiota of a mouse over time, 12% was explained by persistent differences among individual mice, 14% by cage, and 18% by shipment group. Our findings suggest that the microbiota of controlled populations of inbred laboratory animals may not be as uniform as previously thought, that animal rearing and handling may account for some variation, and that as yet unidentified factors may explain additional components of variation in the composition of the microbiota within populations and individuals over time. These findings have implications for the design and interpretation of experiments involving laboratory animals.

    View details for DOI 10.1371/journal.pone.0142825

    View details for PubMedID 26565698

  • Antibiotics and the gut microbiota JOURNAL OF CLINICAL INVESTIGATION Modi, S. R., Collins, J. J., Relman, D. A. 2014; 124 (10): 4212-4218

    View details for DOI 10.1172/JCI72333

    View details for Web of Science ID 000342649900015

  • Antibiotics and the gut microbiota. journal of clinical investigation Modi, S. R., Collins, J. J., Relman, D. A. 2014; 124 (10): 4212-4218


    Antibiotics have been a cornerstone of innovation in the fields of public health, agriculture, and medicine. However, recent studies have shed new light on the collateral damage they impart on the indigenous host-associated communities. These drugs have been found to alter the taxonomic, genomic, and functional capacity of the human gut microbiota, with effects that are rapid and sometimes persistent. Broad-spectrum antibiotics reduce bacterial diversity while expanding and collapsing membership of specific indigenous taxa. Furthermore, antibiotic treatment selects for resistant bacteria, increases opportunities for horizontal gene transfer, and enables intrusion of pathogenic organisms through depletion of occupied natural niches, with profound implications for the emergence of resistance. Because these pervasive alterations can be viewed as an uncoupling of mutualistic host-microbe relationships, it is valuable to reconsider antimicrobial therapies in the context of an ecological framework. Understanding the biology of competitive exclusion, interspecies protection, and gene flow of adaptive functions in the gut environment may inform the design of new strategies that treat infections while preserving the ecology of our beneficial constituents.

    View details for DOI 10.1172/JCI72333

    View details for PubMedID 25271726

  • Human microbiome science: vision for the future, Bethesda, MD, July 24 to 26, 2013 MICROBIOME Ravel, J., Blaser, M. J., Braun, J., Brown, E., Bushman, F. D., Chang, E. B., Davies, J., Dewey, K. G., Dinan, T., Dominguez-Bello, M., Erdman, S. E., Finlay, B. B., Garrett, W. S., Huffnagle, G. B., Huttenhower, C., Jansson, J., Jeffery, I. B., Jobin, C., Khoruts, A., Kong, H. H., Lampe, J. W., Ley, R. E., Littman, D. R., Mazmanian, S. K., Mills, D. A., Neish, A. S., Petrof, E., Relman, D. A., Rhodes, R., Turnbaugh, P. J., Young, V. B., Knight, R., White, O. 2014; 2
  • Population health: immaturity in the gut microbial community. Nature Costello, E. K., Relman, D. A. 2014; 510 (7505): 344-345

    View details for DOI 10.1038/nature13347

    View details for PubMedID 24896185

  • Unrest at home: diarrheal disease and microbiota disturbance. Genome biology Bik, E. M., Relman, D. A. 2014; 15 (6): 120-?


    Diarrhea and malnutrition, two intertwined worldwide problems, are both associated with lower diversity of the intestinal microbiota in children in low-income countries.

    View details for DOI 10.1186/gb4182

    View details for PubMedID 25002208

  • Nasal Microenvironments and Interspecific Interactions Influence Nasal Microbiota Complexity and S. aureus Carriage. Cell host & microbe Yan, M., Pamp, S. J., Fukuyama, J., Hwang, P. H., Cho, D., Holmes, S., Relman, D. A. 2013; 14 (6): 631-640


    The indigenous microbiota of the nasal cavity plays important roles in human health and disease. Patterns of spatial variation in microbiota composition may help explain Staphylococcus aureus colonization and reveal interspecies and species-host interactions. To assess the biogeography of the nasal microbiota, we sampled healthy subjects, representing both S. aureus carriers and noncarriers at three nasal sites (anterior naris, middle meatus, and sphenoethmoidal recess). Phylogenetic compositional and sparse linear discriminant analyses revealed communities that differed according to site epithelium type and S. aureus culture-based carriage status. Corynebacterium accolens and C. pseudodiphtheriticum were identified as the most important microbial community determinants of S. aureus carriage, and competitive interactions were only evident at sites with ciliated pseudostratified columnar epithelium. In vitro cocultivation experiments provided supporting evidence of interactions among these species. These results highlight spatial variation in nasal microbial communities and differences in community composition between S. aureus carriers and noncarriers.

    View details for DOI 10.1016/j.chom.2013.11.005

    View details for PubMedID 24331461

  • Genetically dictated change in host mucus carbohydrate landscape exerts a diet-dependent effect on the gut microbiota PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Kashyap, P. C., Marcobal, A., Ursell, L. K., Smits, S. A., Sonnenburg, E. D., Costello, E. K., Higginbottom, S. K., Domino, S. E., Holmes, S. P., Relman, D. A., Knight, R., Gordon, J. I., Sonnenburg, J. L. 2013; 110 (42): 17059-17064


    We investigate how host mucus glycan composition interacts with dietary carbohydrate content to influence the composition and expressed functions of a human gut community. The humanized gnotobiotic mice mimic humans with a nonsecretor phenotype due to knockout of their ?1-2 fucosyltransferase (Fut2) gene. The fecal microbiota of Fut2(-) mice that lack fucosylated host glycans show decreased alpha diversity relative to Fut2(+) mice and exhibit significant differences in community composition. A glucose-rich plant polysaccharide-deficient (PD) diet exerted a strong effect on the microbiota membership but eliminated the effect of Fut2 genotype. Additionally fecal metabolites predicted host genotype in mice on a polysaccharide-rich standard diet but not on a PD diet. A more detailed mechanistic analysis of these interactions involved colonization of gnotobiotic Fut2(+) and Fut2(-) mice with Bacteroides thetaiotaomicron, a prominent member of the human gut microbiota known to adaptively forage host mucosal glycans when dietary polysaccharides are absent. Within Fut2(-) mice, the B. thetaiotaomicron fucose catabolic pathway was markedly down-regulated, whereas BT4241-4247, an operon responsive to terminal ?-galactose, the precursor that accumulates in the Fut2(-) mice, was significantly up-regulated. These changes in B. thetaiotaomicron gene expression were only evident in mice fed a PD diet, wherein B. thetaiotaomicron relies on host mucus consumption. Furthermore, up-regulation of the BT4241-4247 operon was also seen in humanized Fut2(-) mice. Together, these data demonstrate that differences in host genotype that affect the carbohydrate landscape of the distal gut interact with diet to alter the composition and function of resident microbes in a diet-dependent manner.

    View details for DOI 10.1073/pnas.1306070110

    View details for Web of Science ID 000325634200076

    View details for PubMedID 24062455

  • Identification of Lactobacillus strains with probiotic features from the bottlenose dolphin (Tursiops truncatus) JOURNAL OF APPLIED MICROBIOLOGY Diaz, M. A., Bik, E. M., Carlin, K. P., Venn-Watson, S. K., Jensen, E. D., Jones, S. E., Gaston, E. P., Relman, D. A., Versalovic, J. 2013; 115 (4): 1037-1051


    In order to develop complementary health management strategies for marine mammals, we used culture-based and culture-independent approaches to identify gastrointestinal lactobacilli of the common bottlenose dolphin, Tursiops truncatus.We screened 307 bacterial isolates from oral and rectal swabs, milk, and gastric fluid, collected from 38 dolphins in the U.S. Navy Marine Mammal Program, for potentially beneficial features. We focused our search on lactobacilli and evaluated their ability to modulate TNF secretion by host cells and inhibit growth of pathogens. We recovered Lactobacillus salivarius strains which secreted factors that stimulated TNF production by human monocytoid cells. These L. salivarius isolates inhibited growth of selected marine mammal and human bacterial pathogens. In addition, we identified a novel Lactobacillus species by culture and direct sequencing with 96.3% 16S rDNA sequence similarity to Lactobacillus ceti.Dolphin-derived L. salivarius isolates possess features making them candidate probiotics for clinical studies in marine mammals.This is the first study to isolate lactobacilli from dolphins, including a new strain of L. salivarius, with potential for veterinary probiotic applications. The isolation and identification of novel Lactobacillus spp. and other indigenous microbes from bottlenose dolphins will enable the study of the biology of symbiotic members of the dolphin microbiota and facilitate the understanding of the microbiomes of these unique animals. This article is protected by copyright. All rights reserved.

    View details for DOI 10.1111/jam.12305

    View details for Web of Science ID 000325012200013

    View details for PubMedID 23855505

  • Metagenomics, Infectious Disease Diagnostics, and Outbreak Investigations Sequence First, Ask Questions Later? JAMA-JOURNAL OF THE AMERICAN MEDICAL ASSOCIATION Relman, D. A. 2013; 309 (14): 1531-1532

    View details for Web of Science ID 000317271700034

    View details for PubMedID 23571595

  • Type I Interferon Suppresses Type II Interferon-Triggered Human Anti-Mycobacterial Responses SCIENCE Teles, R. M., Graeber, T. G., Krutzik, S. R., Montoya, D., Schenk, M., Lee, D. J., Komisopoulou, E., Kelly-Scumpia, K., Chun, R., Iyer, S. S., Sarno, E. N., Rea, T. H., Hewison, M., Adams, J. S., Popper, S. J., Relman, D. A., Stenger, S., Bloom, B. R., Cheng, G., Modlin, R. L. 2013; 339 (6126): 1448-1453


    Type I interferons (IFN-? and IFN-?) are important for protection against many viral infections, whereas type II interferon (IFN-?) is essential for host defense against some bacterial and parasitic pathogens. Study of IFN responses in human leprosy revealed an inverse correlation between IFN-? and IFN-? gene expression programs. IFN-? and its downstream vitamin D-dependent antimicrobial genes were preferentially expressed in self-healing tuberculoid lesions and mediated antimicrobial activity against the pathogen Mycobacterium leprae in vitro. In contrast, IFN-? and its downstream genes, including interleukin-10 (IL-10), were induced in monocytes by M. leprae in vitro and preferentially expressed in disseminated and progressive lepromatous lesions. The IFN-?-induced macrophage vitamin D-dependent antimicrobial peptide response was inhibited by IFN-? and by IL-10, suggesting that the differential production of IFNs contributes to protection versus pathogenesis in some human bacterial infections.

    View details for DOI 10.1126/science.1233665

    View details for Web of Science ID 000316740700047

    View details for PubMedID 23449998

    View details for PubMedCentralID PMC3653587

  • The Increasingly Compelling Moral Responsibilities of Life Scientists HASTINGS CENTER REPORT Relman, D. A. 2013; 43 (2): 34-35

    View details for DOI 10.1002/hast.156

    View details for Web of Science ID 000316274100014

    View details for PubMedID 23494700

  • Undernutrition-Looking Within for Answers SCIENCE Relman, D. A. 2013; 339 (6119): 530-532
  • Distinct Distal Gut Microbiome Diversity and Composition in Healthy Children from Bangladesh and the United States PLOS ONE Lin, A., Bik, E. M., Costello, E. K., Dethlefsen, L., Haque, R., Relman, D. A., Singh, U. 2013; 8 (1)

    View details for DOI 10.1371/journal.pone.0053838

    View details for Web of Science ID 000314019100034

    View details for PubMedID 23349750

  • Distinct distal gut microbiome diversity and composition in healthy children from Bangladesh and the United States. PloS one Lin, A., Bik, E. M., Costello, E. K., Dethlefsen, L., Haque, R., Relman, D. A., Singh, U. 2013; 8 (1)


    Our current understanding of the composition and stability of the human distal gut microbiota is based largely on studies of infants and adults living in developed countries. In contrast, little is known about the gut microbiota and its variation over time in older children and adolescents, especially in developing countries.We compared the diversity, composition, and temporal stability of the fecal microbiota of healthy children, ages 9 to 14 years, living in an urban slum in Bangladesh with that of children of the same age range in an upper-middle class suburban community in the United States. We analyzed >8,000 near full-length 16S rRNA gene sequences and over 845,000 pyrosequencing reads of the 16S rRNA V1-V3 region. The distal gut of Bangladeshi children harbored significantly greater bacterial diversity than that of U.S. children, including novel lineages from several bacterial phyla. Bangladeshi and U.S. children had distinct fecal bacterial community membership and structure; the microbiota of Bangladeshi children was enriched in Prevotella, Butyrivibrio, and Oscillospira and depleted in Bacteroides relative to U.S. children (although similar to Bangladeshi adults). Furthermore, community membership and structure in Bangladeshi children was significantly less stable month-to-month than U.S. children.Together, these results suggest that differing environmental or genetic factors may shape the microbiota of healthy children in the two countries. Further investigation is necessary to understand the mechanisms and factors that underlie these differences, and to incorporate these findings into new strategies for the prevention and treatment of childhood and adolescent diseases.

    View details for DOI 10.1371/journal.pone.0053838

    View details for PubMedID 23349750

  • Probiotics, prebiotics, and the host microbiome: the science of translation ANNALS REPORTS Petschow, B., Dore, J., Hibberd, P., Dinan, T., Reid, G., Blaser, M., Cani, P. D., Degnan, F. H., Foster, J., Gibson, G., Hutton, J., Klaenhammer, T. R., Ley, R., Nieuwdorp, M., Pot, B., Relman, D., Serazin, A., Sanders, M. E. 2013; 1306: 1-17


    Recent advances in our understanding of the community structure and function of the human microbiome have implications for the potential role of probiotics and prebiotics in promoting human health. A group of experts recently met to review the latest advances in microbiota/microbiome research and discuss the implications for development of probiotics and prebiotics, primarily as they relate to effects mediated via the intestine. The goals of the meeting were to share recent advances in research on the microbiota, microbiome, probiotics, and prebiotics, and to discuss these findings in the contexts of regulatory barriers, evolving healthcare environments, and potential effects on a variety of health topics, including the development of obesity and diabetes; the long-term consequences of exposure to antibiotics early in life to the gastrointestinal (GI) microbiota; lactose intolerance; and the relationship between the GI microbiota and the central nervous system, with implications for depression, cognition, satiety, and mental health for people living in developed and developing countries. This report provides an overview of these discussions.

    View details for DOI 10.1111/nyas.12303

    View details for Web of Science ID 000329607800001

    View details for PubMedID 24266656

  • Microbiome assembly across multiple body sites in low-birthweight infants. mBio Costello, E. K., Carlisle, E. M., Bik, E. M., Morowitz, M. J., Relman, D. A. 2013; 4 (6): e00782-13


    ABSTRACT The purpose of this study was to evaluate the composition and richness of bacterial communities associated with low-birthweight (LBW) infants in relation to host body site, individual, and age. Bacterial 16S rRNA genes from saliva samples, skin swabs, and stool samples collected on postnatal days 8, 10, 12, 15, 18, and 21 from six LBW (five premature) infants were amplified, pyrosequenced, and analyzed within a comparative framework that included analogous data from normal-birthweight (NBW) infants and healthy adults. We found that body site was the primary determinant of bacterial community composition in the LBW infants. However, site specificity depended on postnatal age: saliva and stool compositions diverged over time but were not significantly different until the babies were 15 days old. This divergence was primarily driven by progressive temporal turnover in the distal gut, which proceeded at a rate similar to that of age-matched NBW infants. Neonatal skin was the most adult-like in microbiota composition, while saliva and stool remained the least so. Compositional variation among infants was marked and depended on body site and age. Only the smallest, most premature infant received antibiotics during the study period; this heralded a coexpansion of Pseudomonas aeruginosa and a novel Mycoplasma sp. in the oral cavity of this vaginally delivered, intubated patient. We conclude that concurrent molecular surveillance of multiple body sites in LBW neonates reveals a delayed compositional differentiation of the oral cavity and distal gut microbiota and, in the case of one infant, an abundant, uncultivated oral Mycoplasma sp., recently detected in human vaginal samples. IMPORTANCE Complications of premature birth are the most common cause of neonatal mortality. Colonization by the indigenous microbiota, which begins at delivery, may predispose some high-risk newborns to invasive infection or necrotizing enterocolitis (NEC), and protect others, yet neonatal microbiome dynamics are poorly understood. Here, we present the first cultivation-independent time series tracking microbiota assembly across multiple body sites in a synchronous cohort of hospitalized low-birthweight (LBW) neonates. We take advantage of archived samples and publically available sequence data and compare our LBW infant findings to those from normal-birthweight (NBW) infants and healthy adults. Our results suggest potential windows of opportunity for the dispersal of microbes within and between hosts and support recent findings of substantial baseline spatiotemporal variation in microbiota composition among high-risk newborns.

    View details for DOI 10.1128/mBio.00782-13

    View details for PubMedID 24169577

    View details for PubMedCentralID PMC3809564

  • Microbiome Assembly across Multiple Body Sites in Low-Birthweight Infants. mBio Costello, E. K., Carlisle, E. M., Bik, E. M., Morowitz, M. J., Relman, D. A. 2013; 4 (6)


    ABSTRACT The purpose of this study was to evaluate the composition and richness of bacterial communities associated with low-birthweight (LBW) infants in relation to host body site, individual, and age. Bacterial 16S rRNA genes from saliva samples, skin swabs, and stool samples collected on postnatal days 8, 10, 12, 15, 18, and 21 from six LBW (five premature) infants were amplified, pyrosequenced, and analyzed within a comparative framework that included analogous data from normal-birthweight (NBW) infants and healthy adults. We found that body site was the primary determinant of bacterial community composition in the LBW infants. However, site specificity depended on postnatal age: saliva and stool compositions diverged over time but were not significantly different until the babies were 15 days old. This divergence was primarily driven by progressive temporal turnover in the distal gut, which proceeded at a rate similar to that of age-matched NBW infants. Neonatal skin was the most adult-like in microbiota composition, while saliva and stool remained the least so. Compositional variation among infants was marked and depended on body site and age. Only the smallest, most premature infant received antibiotics during the study period; this heralded a coexpansion of Pseudomonas aeruginosa and a novel Mycoplasma sp. in the oral cavity of this vaginally delivered, intubated patient. We conclude that concurrent molecular surveillance of multiple body sites in LBW neonates reveals a delayed compositional differentiation of the oral cavity and distal gut microbiota and, in the case of one infant, an abundant, uncultivated oral Mycoplasma sp., recently detected in human vaginal samples. IMPORTANCE Complications of premature birth are the most common cause of neonatal mortality. Colonization by the indigenous microbiota, which begins at delivery, may predispose some high-risk newborns to invasive infection or necrotizing enterocolitis (NEC), and protect others, yet neonatal microbiome dynamics are poorly understood. Here, we present the first cultivation-independent time series tracking microbiota assembly across multiple body sites in a synchronous cohort of hospitalized low-birthweight (LBW) neonates. We take advantage of archived samples and publically available sequence data and compare our LBW infant findings to those from normal-birthweight (NBW) infants and healthy adults. Our results suggest potential windows of opportunity for the dispersal of microbes within and between hosts and support recent findings of substantial baseline spatiotemporal variation in microbiota composition among high-risk newborns.

    View details for DOI 10.1128/mBio.00782-13

    View details for PubMedID 24169577

  • Time series community genomics analysis reveals rapid shifts in bacterial species, strains, and phage during infant gut colonization GENOME RESEARCH Sharon, I., Morowitz, M. J., Thomas, B. C., Costello, E. K., Relman, D. A., Banfield, J. F. 2013; 23 (1): 111-120


    The gastrointestinal microbiome undergoes shifts in species and strain abundances, yet dynamics involving closely related microorganisms remain largely unknown because most methods cannot resolve them. We developed new metagenomic methods and utilized them to track species and strain level variations in microbial communities in 11 fecal samples collected from a premature infant during the first month of life. Ninety six percent of the sequencing reads were assembled into scaffolds of >500 bp in length that could be assigned to organisms at the strain level. Six essentially complete (?99%) and two near-complete genomes were assembled for bacteria that comprised as little as 1% of the community, as well as nine partial genomes of bacteria representing as little as 0.05%. In addition, three viral genomes were assembled and assigned to their hosts. The relative abundance of three Staphylococcus epidermidis strains, as well as three phages that infect them, changed dramatically over time. Genes possibly related to these shifts include those for resistance to antibiotics, heavy metals, and phage. At the species level, we observed the decline of an early-colonizing Propionibacterium acnes strain similar to SK137 and the proliferation of novel Propionibacterium and Peptoniphilus species late in colonization. The Propionibacterium species differed in their ability to metabolize carbon compounds such as inositol and sialic acid, indicating that shifts in species composition likely impact the metabolic potential of the community. These results highlight the benefit of reconstructing complete genomes from metagenomic data and demonstrate methods for achieving this goal.

    View details for DOI 10.1101/gr.142315.112

    View details for Web of Science ID 000312963400010

    View details for PubMedID 22936250

  • Restoration of the gut microbial habitat as a disease therapy NATURE BIOTECHNOLOGY Relman, D. A. 2013; 31 (1): 35-37

    View details for DOI 10.1038/nbt.2475

    View details for Web of Science ID 000313563600019

    View details for PubMedID 23302933

  • Temporal Dynamics of the Transcriptional Response to Dengue Virus Infection in Nicaraguan Children PLOS NEGLECTED TROPICAL DISEASES Popper, S. J., Gordon, A., Liu, M., Balmaseda, A., Harris, E., Relman, D. A. 2012; 6 (12)


    Dengue is the most prevalent mosquito-borne human illness worldwide. The ability to predict disease severity during the earliest days of the illness is a long-sought, but unachieved goal.We examined human genome-wide transcript abundance patterns in daily peripheral blood mononuclear cell (PBMC) samples from 41 children hospitalized with dengue virus (DENV) infection in Nicaragua, as well as 8 healthy control subjects. Nine patients had primary dengue fever (DF1), 11 had dengue fever with serologic evidence of prior DENV infection, i.e., secondary dengue fever (DF2), 12 had dengue hemorrhagic fever (DHF), and 9 had dengue shock syndrome (DSS). We identified 2,092 genes for which transcript abundance differed significantly between patients on days 3-6 of fever and healthy subjects (FDR<1%). Prior DENV infection explained the greatest amount of variation in gene expression among patients. The number of differentially expressed genes was greatest on fever day 3 in patients with DF1, while the number in patients with DF2 or DHF/DSS was greatest on day 5. Genes associated with the mitotic cell cycle and B cell differentiation were expressed at higher levels, and genes associated with signal transduction and cell adhesion were expressed at lower levels, in patients versus healthy controls. On fever day 3, a set of interferon-stimulated gene transcripts was less abundant in patients who subsequently developed DSS than in other patient groups (p<0.05, ranksum). Patients who later developed DSS also had higher levels of transcripts on day 3 associated with mitochondrial function (p<0.01, ranksum). These day 3 transcript abundance findings were not evident on subsequent fever days.In conclusion, we identified differences in the timing and magnitude of human gene transcript abundance changes in DENV patients that were associated with serologic evidence of prior infection and with disease severity. Some of these differential features may predict the outcome of DENV infection.

    View details for DOI 10.1371/journal.pntd.0001966

    View details for Web of Science ID 000312910200031

    View details for PubMedID 23285306

    View details for PubMedCentralID PMC3527342

  • Comparisons of clustered regularly interspaced short palindromic repeats and viromes in human saliva reveal bacterial adaptations to salivary viruses ENVIRONMENTAL MICROBIOLOGY Pride, D. T., Salzman, J., Relman, D. A. 2012; 14 (9): 2564-2576


    Explorations of human microbiota have provided substantial insight into microbial community composition; however, little is known about interactions between various microbial components in human ecosystems. In response to the powerful impact of viral predation, bacteria have acquired potent defences, including an adaptive immune response based on the clustered regularly interspaced short palindromic repeats (CRISPRs)/Cas system. To improve our understanding of the interactions between bacteria and their viruses in humans, we analysed 13?977 streptococcal CRISPR sequences and compared them with 2?588?172 virome reads in the saliva of four human subjects over 17 months. We found a diverse array of viruses and CRISPR spacers, many of which were specific to each subject and time point. There were numerous viral sequences matching CRISPR spacers; these matches were highly specific for salivary viruses. We determined that spacers and viruses coexist at the same time, which suggests that streptococcal CRISPR/Cas systems are under constant pressure from salivary viruses. CRISPRs in some subjects were just as likely to match viral sequences from other subjects as they were to match viruses from the same subject. Because interactions between bacteria and viruses help to determine the structure of bacterial communities, CRISPR-virus analyses are likely to provide insight into the forces shaping the human microbiome.

    View details for DOI 10.1111/j.1462-2920.2012.02775.x

    View details for Web of Science ID 000308300600025

    View details for PubMedID 22583485

  • The human microbiome: ecosystem resilience and health NUTRITION REVIEWS Relman, D. A. 2012; 70: S2-S9


    Given the importance of the microbiome for human health, both the stability and the response to disturbance of this microbial ecosystem are crucial issues. Yet, the current understanding of these factors is insufficient. Early data suggest there is relative stability in the microbiome of adults in the absence of gross perturbation, and that long-term stability of the human indigenous microbial communities is maintained not by inertia but by the action of restorative forces within a dynamic system. After brief exposures to some antibiotics, there is an immediate and substantial perturbation and at least a partial recovery of taxonomic composition. Responses to antibiotics are individualized and are influenced by prior experience with the same antibiotic. These findings suggest that the human microbiome has properties of resilience. Besides serving to reveal critical underlying functional attributes, microbial interactions, and keystone species within the indigenous microbiota, the response to disturbance may have value in predicting future instability and disease and in managing the human microbial ecosystem.

    View details for DOI 10.1111/j.1753-4887.2012.00489.x

    View details for Web of Science ID 000307107200002

    View details for PubMedID 22861804

  • Gut Immune Maturation Depends on Colonization with a Host-Specific Microbiota CELL Chung, H., Pamp, S. J., Hill, J. A., Surana, N. K., Edelman, S. M., Troy, E. B., Reading, N. C., Villablanca, E. J., Wang, S., Mora, J. R., Umesaki, Y., Mathis, D., Benoist, C., Relman, D. A., Kasper, D. L. 2012; 149 (7): 1578-1593


    Gut microbial induction of host immune maturation exemplifies host-microbe mutualism. We colonized germ-free (GF) mice with mouse microbiota (MMb) or human microbiota (HMb) to determine whether small intestinal immune maturation depends on a coevolved host-specific microbiota. Gut bacterial numbers and phylum abundance were similar in MMb and HMb mice, but bacterial species differed, especially the Firmicutes. HMb mouse intestines had low levels of CD4(+) and CD8(+) T cells, few proliferating T cells, few dendritic cells, and low antimicrobial peptide expression--all characteristics of GF mice. Rat microbiota also failed to fully expand intestinal T cell numbers in mice. Colonizing GF or HMb mice with mouse-segmented filamentous bacteria (SFB) partially restored T cell numbers, suggesting that SFB and other MMb organisms are required for full immune maturation in mice. Importantly, MMb conferred better protection against Salmonella infection than HMb. A host-specific microbiota appears to be critical for a healthy immune system.

    View details for DOI 10.1016/j.cell.2012.04.037

    View details for Web of Science ID 000305753800022

    View details for PubMedID 22726443

    View details for PubMedCentralID PMC3442780

  • MICROBIOLOGY Learning about who we are NATURE Relman, D. A. 2012; 486 (7402): 194-195

    View details for Web of Science ID 000305189000019

    View details for PubMedID 22699602

  • The Application of Ecological Theory Toward an Understanding of the Human Microbiome SCIENCE Costello, E. K., Stagaman, K., Dethlefsen, L., Bohannan, B. J., Relman, D. A. 2012; 336 (6086): 1255-1262


    The human-microbial ecosystem plays a variety of important roles in human health and disease. Each person can be viewed as an island-like "patch" of habitat occupied by microbial assemblages formed by the fundamental processes of community ecology: dispersal, local diversification, environmental selection, and ecological drift. Community assembly theory, and metacommunity theory in particular, provides a framework for understanding the ecological dynamics of the human microbiome, such as compositional variability within and between hosts. We explore three core scenarios of human microbiome assembly: development in infants, representing assembly in previously unoccupied habitats; recovery from antibiotics, representing assembly after disturbance; and invasion by pathogens, representing assembly in the context of invasive species. Judicious application of ecological theory may lead to improved strategies for restoring and maintaining the microbiota and the crucial health-associated ecosystem services that it provides.

    View details for DOI 10.1126/science.1224203

    View details for Web of Science ID 000304905300034

    View details for PubMedID 22674335

  • Microbiota-Targeted Therapies: An Ecological Perspective SCIENCE TRANSLATIONAL MEDICINE Lemon, K. P., Armitage, G. C., Relman, D. A., Fischbach, M. A. 2012; 4 (137)


    The connection between disease and the disruption of homeostatic interactions between the host and its microbiota is now well established. Drug developers and clinicians are starting to rely more heavily on therapies that directly target the microbiota and on the ecology of the microbiota to understand the outcomes of these treatments. The effects of those microbiota-targeted therapies that alter community composition range in scale from eliminating individual strains of a single species (for example, with antibacterial conjugate vaccines) to replacing the entire community with a new intact microbiota (for example, by fecal transplantation). Secondary infections linked to antibiotic use provide a cautionary tale of the unintended consequences of perturbing a microbial species network and highlight the need for new narrow-spectrum antibiotics with rapid companion diagnostics. Insights into microbial ecology will also benefit the development of probiotics, whose therapeutic prospects will depend on rigorous clinical testing. Future probiotics may take the form of a consortium of long-term community residents: "a fecal transplant in a capsule." The efficacy of microbiota-targeted therapies will need to be assessed using new diagnostic tools that measure community function rather than composition, including the temporal response of a microbial community to a defined perturbation such as an antibiotic or probiotic.

    View details for DOI 10.1126/scitranslmed.3004183

    View details for Web of Science ID 000305075700012

    View details for PubMedID 22674555

  • Single-cell sequencing provides clues about the host interactions of segmented filamentous bacteria (SFB) GENOME RESEARCH Pamp, S. J., Harrington, E. D., Quake, S. R., Relman, D. A., Blainey, P. C. 2012; 22 (6): 1107-1119


    Segmented filamentous bacteria (SFB) are host-specific intestinal symbionts that comprise a distinct clade within the Clostridiaceae, designated Candidatus Arthromitus. SFB display a unique life cycle within the host, involving differentiation into multiple cell types. The latter include filaments that attach intimately to intestinal epithelial cells, and from which "holdfasts" and spores develop. SFB induce a multifaceted immune response, leading to host protection from intestinal pathogens. Cultivation resistance has hindered characterization of these enigmatic bacteria. In the present study, we isolated five SFB filaments from a mouse using a microfluidic device equipped with laser tweezers, generated genome sequences from each, and compared these sequences with each other, as well as to recently published SFB genome sequences. Based on the resulting analyses, SFB appear to be dependent on the host for a variety of essential nutrients. SFB have a relatively high abundance of predicted proteins devoted to cell cycle control and to envelope biogenesis, and have a group of SFB-specific autolysins and a dynamin-like protein. Among the five filament genomes, an average of 8.6% of predicted proteins were novel, including a family of secreted SFB-specific proteins. Four ADP-ribosyltransferase (ADPRT) sequence types, and a myosin-cross-reactive antigen (MCRA) protein were discovered; we hypothesize that they are involved in modulation of host responses. The presence of polymorphisms among mouse SFB genomes suggests the evolution of distinct SFB lineages. Overall, our results reveal several aspects of SFB adaptation to the mammalian intestinal tract.

    View details for DOI 10.1101/gr.131482.111

    View details for Web of Science ID 000304728100012

    View details for PubMedID 22434425

  • Creating a Mammalian-Transmissible A/H5N1 Influenza Virus: Social Contracts, Prudence, and Alternative Perspectives JOURNAL OF INFECTIOUS DISEASES Osterholm, M. T., Relman, D. A. 2012; 205 (11): 1636-1638

    View details for DOI 10.1093/infdis/jis259

    View details for Web of Science ID 000304065600006

    View details for PubMedID 22454472

  • Presumed Guilt in the Anthrax Case Response SCIENCE Relman, D. A. 2012; 336 (6082): 669-670
  • Evidence of a robust resident bacteriophage population revealed through analysis of the human salivary virome ISME JOURNAL Pride, D. T., Salzman, J., Haynes, M., Rohwer, F., Davis-Long, C., White, R. A., Loomer, P., Armitage, G. C., Relman, D. A. 2012; 6 (5): 915-926


    Viruses are the most abundant known infectious agents on the planet and are significant drivers of diversity in a variety of ecosystems. Although there have been numerous studies of viral communities, few have focused on viruses within the indigenous human microbiota. We analyzed 2?267?695 virome reads from viral particles and compared them with 263?516 bacterial 16S rRNA gene sequences from the saliva of five healthy human subjects over a 2- to 3-month period, in order to improve our understanding of the role viruses have in the complex oral ecosystem. Our data reveal viral communities in human saliva dominated by bacteriophages whose constituents are temporally distinct. The preponderance of shared homologs between the salivary viral communities in two unrelated subjects in the same household suggests that environmental factors are determinants of community membership. When comparing salivary viromes to those from human stool and the respiratory tract, each group was distinct, further indicating that habitat is of substantial importance in shaping human viromes. Compared with coexisting bacteria, there was concordance among certain predicted host-virus pairings such as Veillonella and Streptococcus, whereas there was discordance among others such as Actinomyces. We identified 122?728 virulence factor homologs, suggesting that salivary viruses may serve as reservoirs for pathogenic gene function in the oral environment. That the vast majority of human oral viruses are bacteriophages whose putative gene function signifies some have a prominent role in lysogeny, suggests these viruses may have an important role in helping shape the microbial diversity in the human oral cavity.

    View details for DOI 10.1038/ismej.2011.169

    View details for Web of Science ID 000302950700002

    View details for PubMedID 22158393

  • Public health and biosecurity. Adaptations of avian flu virus are a cause for concern. Science Berns, K. I., Casadevall, A., Cohen, M. L., Ehrlich, S. A., Enquist, L. W., Fitch, J. P., Franz, D. R., Fraser-Liggett, C. M., Grant, C. M., Imperiale, M. J., Kanabrocki, J., Keim, P. S., Lemon, S. M., Levy, S. B., Lumpkin, J. R., Miller, J. F., Murch, R., Nance, M. E., Osterholm, M. T., Relman, D. A., Roth, J. A., Vidaver, A. K. 2012; 335 (6069): 660-661

    View details for DOI 10.1126/science.1217994

    View details for PubMedID 22294736

  • Policy: Adaptations of avian flu virus are a cause for concern. Nature Berns, K. I., Casadevall, A., Cohen, M. L., Ehrlich, S. A., Enquist, L. W., Fitch, J. P., Franz, D. R., Fraser-Liggett, C. M., Grant, C. M., Imperiale, M. J., Kanabrocki, J., Keim, P. S., Lemon, S. M., Levy, S. B., Lumpkin, J. R., Miller, J. F., Murch, R., Nance, M. E., Osterholm, M. T., Relman, D. A., Roth, J. A., Vidaver, A. K. 2012; 482 (7384): 153-154

    View details for DOI 10.1038/482153a

    View details for PubMedID 22294204

  • American Anthrax Fear, Crime, and the Investigation of the Nation's Deadliest Bioterror Attack (Book Review) SCIENCE Book Review Authored by: Relman, D. A. 2012; 335 (6068): 540-541
  • Comparisons of distance methods for combining covariates and abundances in microbiome studies. Pacific Symposium on Biocomputing. Pacific Symposium on Biocomputing Fukuyama, J., McMurdie, P. J., Dethlefsen, L., Relman, D. A., Holmes, S. 2012: 213-224


    This article compares different methods for combining abundance data, phylogenetic trees and clinical covariates in a nonparametric setting. In particular we study the output from the principal coordinates analysis on UNIFRAC and WEIGHTED UNIFRAC distances and the output from a double principal coordinate analyses DPCOA using distances computed on the phylogenetic tree. We also present power comparisons for some of the standard tests of phylogenetic signal between different types of samples. These methods are compared both on simulated and real data sets. Our study shows that DPCoA is less robust to outliers, and more robust to small noisy fluctuations around zero.

    View details for PubMedID 22174277

  • Modulation of the Host Interferon Response and ISGylation Pathway by B. pertussis Filamentous Hemagglutinin PLOS ONE Dieterich, C., Relman, D. A. 2011; 6 (11)


    Bordetella pertussis filamentous hemagglutinin (FHA) is a surface-associated and secreted protein that serves as a crucial adherence factor, and displays immunomodulatory activity in human peripheral blood mononuclear cells (PBMCs). In order to appreciate more fully the role of secreted FHA in pathogenesis, we analyzed FHA-induced changes in genome-wide transcript abundance in human PBMCs. Among the 683 known unique genes with greater than 3-fold change in transcript abundance following FHA treatment, 125 (18.3%) were identified as interferon (IFN)-regulated. Among the latter group were genes encoding several members of the IFN type I response, as well as 3 key components of the ISGylation pathway. Using real-time RT-PCR, we confirmed FHA-associated increases in transcript abundance for the genes encoding ubiquitin-like protein, ISG15, and its specific protease USP18. Western-blot analysis demonstrated the presence of both, free ISG15 and several ISGylated conjugates in FHA-stimulated PBMC lysates, but not in unstimulated cells. Intracellular FACS analysis provided evidence that monocytes and a natural killer-enriched cell population were the primary producers of ISG15 in PBMCs after FHA stimulation. Our data reveal previously-unrecognized effects of B. pertussis FHA on host IFN and ISGylation responses, and suggest previously-unsuspected mechanisms by which FHA may alter the outcome of the host-pathogen interaction.

    View details for DOI 10.1371/journal.pone.0027535

    View details for Web of Science ID 000298168100012

    View details for PubMedID 22140447

  • Transcriptional Profiling of the Iron Starvation Response in Bordetella pertussis Provides New Insights into Siderophore Utilization and Virulence Gene Expression JOURNAL OF BACTERIOLOGY Brickman, T. J., Cummings, C. A., Liew, S., Relman, D. A., Armstrong, S. K. 2011; 193 (18): 4798-4812


    Serological studies of patients with pertussis and the identification of antigenic Bordetella pertussis proteins support the hypothesis that B. pertussis perceives an iron starvation cue and expresses multiple iron source utilization systems in its natural human host environment. Furthermore, previous studies using a murine respiratory tract infection model showed that several of these B. pertussis iron systems are required for colonization and persistence and are differentially expressed over the course of infection. The present study examined genome-wide changes in B. pertussis gene transcript abundance in response to iron starvation in vitro. In addition to known iron source utilization genes, we identified a previously uncharacterized iron-repressed cytoplasmic membrane transporter system, fbpABC, that is required for the utilization of multiple structurally distinct siderophores including alcaligin, enterobactin, ferrichrome, and desferrioxamine B. Expression of type III secretion system genes was also found to be upregulated during iron starvation in both B. pertussis strain Tohama I and Bordetella bronchiseptica strain RB50. In a survey of type III secretion system protein production by an assortment of B. pertussis laboratory-adapted and low-passage clinical isolate strains, iron limitation increased the production and secretion of the type III secretion system-specific translocation apparatus tip protein Bsp22 in all Bvg-proficient strains. These results indicate that iron starvation in the infected host is an important environmental cue influencing not only Bordetella iron transport gene expression but also the expression of other important virulence-associated genes.

    View details for DOI 10.1128/JB.05136-11

    View details for Web of Science ID 000294261700025

    View details for PubMedID 21742863

  • Microbial Genomics and Infectious Diseases NEW ENGLAND JOURNAL OF MEDICINE Relman, D. A. 2011; 365 (4): 347-357

    View details for Web of Science ID 000293172200013

    View details for PubMedID 21793746

  • Myocardial depressant effects of interleukin 6 in meningococcal sepsis are regulated by p38 mitogen-activated protein kinase CRITICAL CARE MEDICINE Pathan, N., Franklin, J. L., Eleftherohorinou, H., Wright, V. J., Hemingway, C. A., Waddell, S. J., Griffiths, M., Dennis, J. L., Relman, D. A., Harding, S. E., Levin, M. 2011; 39 (7): 1692-1711


    Myocardial failure, leading to inotrope-unresponsive shock, is the predominant cause of death in meningococcal and other forms of septic shock. Proinflammatory cytokines released in septic shock are known to have myocardial depressant effects. We previously showed that interleukin 6 is a major myocardial depressant factor in children with meningococcal septicemia. In the current study, we aimed to investigate the mechanisms by which interleukin 6 induces myocardial failure in meningococcal sepsis and to identify potential novel therapeutic targets.Laboratory-based study.University hospital and laboratories.Children with a clinical diagnosis of meningococcal septic shock.We studied interleukin 6-induced signaling events, both in vitro using isolated rat ventricular cardiac myocytes as a model of myocardial contractility and in whole blood from children with meningococcal sepsis.None.We demonstrated involvement of Janus kinase 2, phosphatidylinositol 3-kinase, Akt, and p38 mitogen-activated protein kinase in interleukin 6-induced negative inotropy in isolated cardiac myocytes. Inhibition of p38 mitogen-activated protein kinase not only reversed interleukin 6-induced myocardial depression in both rat and human myocytes, but restored inotrope responsiveness. Cardiomyocytes transduced with dominant-negative p38 mitogen-activated protein kinase showed no interleukin 6-induced myocardial depression. To investigate p38 mitogen-activated protein kinase in vivo, we profiled global RNA expression patterns in peripheral blood of children with meningococcal septicemia. Transcripts for genes mapping to the p38 mitogen-activated protein kinase pathway showed significantly altered levels of abundance with a high proportion of genes of this pathway affected.Our findings demonstrate an integral role of the p38 mitogen-activated protein kinase pathway in interleukin 6-mediated cardiac contractile dysfunction and inotrope insensitivity. Dysregulation of the p38 mitogen-activated protein kinase pathway in meningococcal septicemia suggests that this pathway may be an important target for novel therapies to reverse myocardial dysfunction in patients with meningococcal septic shock who are not responsive to inotropic support.

    View details for DOI 10.1097/CCM.0b013e3182186d27

    View details for Web of Science ID 000291721800013

    View details for PubMedID 21494108

  • Minimum information about a marker gene sequence (MIMARKS) and minimum information about any (x) sequence (MIxS) specifications NATURE BIOTECHNOLOGY Yilmaz, P., Kottmann, R., Field, D., Knight, R., Cole, J. R., Amaral-Zettler, L., Gilbert, J. A., Karsch-Mizrachi, I., Johnston, A., Cochrane, G., Vaughan, R., Hunter, C., Park, J., Morrison, N., Rocca-Serra, P., Sterk, P., Arumugam, M., Bailey, M., Baumgartner, L., Birren, B. W., Blaser, M. J., Bonazzi, V., Booth, T., Bork, P., Bushman, F. D., Buttigieg, P. L., Chain, P. S., Charlson, E., Costello, E. K., Huot-Creasy, H., Dawyndt, P., DeSantis, T., Fierer, N., Fuhrman, J. A., Gallery, R. E., Gevers, D., Gibbs, R. A., Gil, I. S., Gonzalez, A., Gordon, J. I., Guralnick, R., Hankeln, W., Highlander, S., Hugenholtz, P., Jansson, J., Kau, A. L., Kelley, S. T., Kennedy, J., Knights, D., Koren, O., Kuczynski, J., Kyrpides, N., Larsen, R., Lauber, C. L., Legg, T., Ley, R. E., Lozupone, C. A., Ludwig, W., Lyons, D., Maguire, E., Methe, B. A., Meyer, F., Muegge, B., Nakielny, S., Nelson, K. E., Nemergut, D., Neufeld, J. D., Newbold, L. K., Oliver, A. E., Pace, N. R., Palanisamy, G., Peplies, J., Petrosino, J., Proctor, L., Pruesse, E., Quast, C., Raes, J., Ratnasingham, S., Ravel, J., Relman, D. A., Assunta-Sansone, S., Schloss, P. D., Schriml, L., Sinha, R., Smith, M. I., Sodergren, E., Spor, A., Stombaugh, J., Tiedje, J. M., Ward, D. V., Weinstock, G. M., Wendel, D., White, O., Whiteley, A., Wilke, A., Wortman, J. R., Yatsunenko, T., Gloeckner, F. O. 2011; 29 (5): 415-420


    Here we present a standard developed by the Genomic Standards Consortium (GSC) for reporting marker gene sequences--the minimum information about a marker gene sequence (MIMARKS). We also introduce a system for describing the environment from which a biological sample originates. The 'environmental packages' apply to any genome sequence of known origin and can be used in combination with MIMARKS and other GSC checklists. Finally, to establish a unified standard for describing sequence data and to provide a single point of entry for the scientific community to access and learn about GSC checklists, we present the minimum information about any (x) sequence (MIxS). Adoption of MIxS will enhance our ability to analyze natural genetic diversity documented by massive DNA sequencing efforts from myriad ecosystems in our ever-changing biosphere.

    View details for DOI 10.1038/nbt.1823

    View details for Web of Science ID 000290301700021

    View details for PubMedID 21552244

    View details for PubMedCentralID PMC3367316

  • Metagenomic detection of phage-encoded platelet-binding factors in the human oral cavity PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Willner, D., Furlan, M., Schmieder, R., Grasis, J. A., Pride, D. T., Relman, D. A., Angly, F. E., McDole, T., Mariella, R. P., Rohwer, F., Haynes, M. 2011; 108: 4547-4553


    The human oropharynx is a reservoir for many potential pathogens, including streptococcal species that cause endocarditis. Although oropharyngeal microbes have been well described, viral communities are essentially uncharacterized. We conducted a metagenomic study to determine the composition of oropharyngeal DNA viral communities (both phage and eukaryotic viruses) in healthy individuals and to evaluate oropharyngeal swabs as a rapid method for viral detection. Viral DNA was extracted from 19 pooled oropharyngeal swabs and sequenced. Viral communities consisted almost exclusively of phage, and complete genomes of several phage were recovered, including Escherichia coli phage T3, Propionibacterium acnes phage PA6, and Streptococcus mitis phage SM1. Phage relative abundances changed dramatically depending on whether samples were chloroform treated or filtered to remove microbial contamination. pblA and pblB genes of phage SM1 were detected in the metagenomes. pblA and pblB mediate the attachment of S. mitis to platelets and play a significant role in S. mitis virulence in the endocardium, but have never previously been detected in the oral cavity. These genes were also identified in salivary metagenomes from three individuals at three time points and in individual saliva samples by PCR. Additionally, we demonstrate that phage SM1 can be induced by commonly ingested substances. Our results indicate that the oral cavity is a reservoir for pblA and pblB genes and for phage SM1 itself. Further studies will determine the association between pblA and pblB genes in the oral cavity and the risk of endocarditis.

    View details for DOI 10.1073/pnas.1000089107

    View details for Web of Science ID 000288451300006

    View details for PubMedID 20547834

  • Incomplete recovery and individualized responses of the human distal gut microbiota to repeated antibiotic perturbation PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Dethlefsen, L., Relman, D. A. 2011; 108: 4554-4561


    The indigenous human microbiota is essential to the health of the host. Although the microbiota can be affected by many features of modern life, we know little about its responses to disturbance, especially repeated disturbances, and how these changes compare with baseline temporal variation. We examined the distal gut microbiota of three individuals over 10 mo that spanned two courses of the antibiotic ciprofloxacin, analyzing more than 1.7 million bacterial 16S rRNA hypervariable region sequences from 52 to 56 samples per subject. Interindividual variation was the major source of variability between samples. Day-to-day temporal variability was evident but constrained around an average community composition that was stable over several months in the absence of deliberate perturbation. The effect of ciprofloxacin on the gut microbiota was profound and rapid, with a loss of diversity and a shift in community composition occurring within 3-4 d of drug initiation. By 1 wk after the end of each course, communities began to return to their initial state, but the return was often incomplete. Although broadly similar, community changes after ciprofloxacin varied among subjects and between the two courses within subjects. In all subjects, the composition of the gut microbiota stabilized by the end of the experiment but was altered from its initial state. As with other ecosystems, the human distal gut microbiome at baseline is a dynamic regimen with a stable average state. Antibiotic perturbation may cause a shift to an alternative stable state, the full consequences of which remain unknown.

    View details for DOI 10.1073/pnas.1000087107

    View details for Web of Science ID 000288451300007

    View details for PubMedID 20847294

    View details for PubMedCentralID PMC3063582

  • Transforming Growth Factor-beta Signaling Pathway in Patients With Kawasaki Disease CIRCULATION-CARDIOVASCULAR GENETICS Shimizu, C., Jain, S., Davila, S., Hibberd, M. L., Lin, K. O., Molkara, D., Frazer, J. R., Sun, S., Baker, A. L., Newburger, J. W., Rowley, A. H., Shulman, S. T., Davila, S., Burgner, D., Breunis, W. B., Kuijpers, T. W., Wright, V. J., Levin, M., Eleftherohorinou, H., Coin, L., Popper, S. J., Relman, D. A., Fury, W., Lin, C., Mellis, S., Tremoulet, A. H., Burns, J. C. 2011; 4 (1): 16-U99
  • Transforming growth factor-beta signaling pathway in patients with Kawasaki disease. Circulation. Cardiovascular genetics Shimizu, C., Jain, S., Davila, S., Hibberd, M. L., Lin, K. O., Molkara, D., Frazer, J. R., Sun, S., Baker, A. L., Newburger, J. W., Rowley, A. H., Shulman, S. T., Burgner, D., Breunis, W. B., Kuijpers, T. W., Wright, V. J., Levin, M., Eleftherohorinou, H., Coin, L., Popper, S. J., Relman, D. A., Fury, W., Lin, C., Mellis, S., Tremoulet, A. H., Burns, J. C. 2011; 4 (1): 16-25


    Transforming growth factor (TGF)-? is a multifunctional peptide that is important in T-cell activation and cardiovascular remodeling, both of which are important features of Kawasaki disease (KD). We postulated that variation in TGF-? signaling might be important in KD susceptibility and disease outcome.We investigated genetic variation in 15 genes belonging to the TGF-? pathway in a total of 771 KD subjects of mainly European descent from the United States, the United Kingdom, Australia, and the Netherlands. We analyzed transcript abundance patterns using microarray and reverse transcriptase-polymerase chain reaction for these same genes, and measured TGF-?2 protein levels in plasma. Genetic variants in TGFB2, TGFBR2, and SMAD3 and their haplotypes were consistently and reproducibly associated with KD susceptibility, coronary artery aneurysm formation, aortic root dilatation, and intravenous immunoglobulin treatment response in different cohorts. A SMAD3 haplotype associated with KD susceptibility replicated in 2 independent cohorts and an intronic single nucleotide polymorphism in a separate haplotype block was also strongly associated (A/G, rs4776338) (P=0.000022; odds ratio, 1.50; 95% confidence interval, 1.25 to 1.81). Pathway analysis using all 15 genes further confirmed the importance of the TGF-? pathway in KD pathogenesis. Whole-blood transcript abundance for these genes and TGF-?2 plasma protein levels changed dynamically over the course of the illness.These studies suggest that genetic variation in the TGF-? pathway influences KD susceptibility, disease outcome, and response to therapy, and that aortic root and coronary artery Z scores can be used for phenotype/genotype analyses. Analysis of transcript abundance and protein levels further support the importance of this pathway in KD pathogenesis.

    View details for DOI 10.1161/CIRCGENETICS.110.940858

    View details for PubMedID 21127203

    View details for PubMedCentralID PMC3073847

  • Strain-resolved community genomic analysis of gut microbial colonization in a premature infant PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Morowitz, M. J., Denef, V. J., Costello, E. K., Thomas, B. C., Poroyko, V., Relman, D. A., Banfield, J. F. 2011; 108 (3): 1128-1133


    The intestinal microbiome is a critical determinant of human health. Alterations in its composition have been correlated with chronic disorders, such as obesity and inflammatory bowel disease in adults, and may be associated with neonatal necrotizing enterocolitis in premature infants. Increasing evidence suggests that strain-level genomic variation may underpin distinct ecological trajectories within mixed populations, yet there have been few strain-resolved analyses of genotype-phenotype connections in the context of the human ecosystem. Here, we document strain-level genomic divergence during the first 3 wk of life within the fecal microbiota of an infant born at 28-wk gestation. We observed three compositional phases during colonization, and reconstructed and intensively curated population genomic datasets from the third phase. The relative abundance of two Citrobacter strains sharing ~99% nucleotide identity changed significantly over time within a community dominated by a nearly clonal Serratia population and harboring a lower abundance Enterococcus population and multiple plasmids and bacteriophage. Modeling of Citrobacter strain abundance suggests differences in growth rates and host colonization patterns. We identified genotypic variation potentially responsible for divergent strain ecologies, including hotspots of sequence variation in regulatory genes and intergenic regions, and in genes involved in transport, flagellar biosynthesis, substrate metabolism, and host colonization, as well as differences in the complements of these genes. Our results demonstrate that a community genomic approach can elucidate gut microbial colonization at the resolution required to discern medically relevant strain and species population dynamics, and hence improve our ability to diagnose and treat microbial community-mediated disorders.

    View details for DOI 10.1073/pnas.1010992108

    View details for Web of Science ID 000286310300046

    View details for PubMedID 21191099

  • Stunned Silence: Gene Expression Programs in Human Cells Infected with Monkeypox or Vaccinia Virus PLOS ONE Rubins, K. H., Hensley, L. E., Relman, D. A., Brown, P. O. 2011; 6 (1)


    Poxviruses use an arsenal of molecular weapons to evade detection and disarm host immune responses. We used DNA microarrays to investigate the gene expression responses to infection by monkeypox virus (MPV), an emerging human pathogen, and Vaccinia virus (VAC), a widely used model and vaccine organism, in primary human macrophages, primary human fibroblasts and HeLa cells. Even as the overwhelmingly infected cells approached their demise, with extensive cytopathic changes, their gene expression programs appeared almost oblivious to poxvirus infection. Although killed (gamma-irradiated) MPV potently induced a transcriptional program characteristic of the interferon response, no such response was observed during infection with either live MPV or VAC. Moreover, while the gene expression response of infected cells to stimulation with ionomycin plus phorbol 12-myristate 13-acetate (PMA), or poly (I-C) was largely unimpaired by infection with MPV, a cluster of pro-inflammatory genes were a notable exception. Poly(I-C) induction of genes involved in alerting the innate immune system to the infectious threat, including TNF-alpha, IL-1 alpha and beta, CCL5 and IL-6, were suppressed by infection with live MPV. Thus, MPV selectively inhibits expression of genes with critical roles in cell-signaling pathways that activate innate immune responses, as part of its strategy for stealthy infection.

    View details for DOI 10.1371/journal.pone.0015615

    View details for Web of Science ID 000286519500024

    View details for PubMedID 21267444

  • Analysis of streptococcal CRISPRs from human saliva reveals substantial sequence diversity within and between subjects over time GENOME RESEARCH Pride, D. T., Sun, C. L., Salzman, J., Rao, N., Loomer, P., Armitage, G. C., Banfield, J. F., Relman, D. A. 2011; 21 (1): 126-136


    Viruses may play an important role in the evolution of human microbial communities. Clustered regularly interspaced short palindromic repeats (CRISPRs) provide bacteria and archaea with adaptive immunity to previously encountered viruses. Little is known about CRISPR composition in members of human microbial communities, the relative rate of CRISPR locus change, or how CRISPR loci differ between the microbiota of different individuals. We collected saliva from four periodontally healthy human subjects over an 11- to 17-mo time period and analyzed CRISPR sequences with corresponding streptococcal repeats in order to improve our understanding of the predominant features of oral streptococcal adaptive immune repertoires. We analyzed a total of 6859 CRISPR bearing reads and 427,917 bacterial 16S rRNA gene sequences. We found a core (ranging from 7% to 22%) of shared CRISPR spacers that remained stable over time within each subject, but nearly a third of CRISPR spacers varied between time points. We document high spacer diversity within each subject, suggesting constant addition of new CRISPR spacers. No greater than 2% of CRISPR spacers were shared between subjects, suggesting that each individual was exposed to different virus populations. We detect changes in CRISPR spacer sequence diversity over time that may be attributable to locus diversification or to changes in streptococcal population structure, yet the composition of the populations within subjects remained relatively stable. The individual-specific and traceable character of CRISPR spacer complements could potentially open the way for expansion of the domain of personalized medicine to the oral microbiome, where lineages may be tracked as a function of health and other factors.

    View details for DOI 10.1101/gr.111732.110

    View details for Web of Science ID 000285868300013

    View details for PubMedID 21149389

  • SmashCell: a software framework for the analysis of single-cell amplified genome sequences BIOINFORMATICS Harrington, E. D., Arumugam, M., Raes, J., Bork, P., Relman, D. A. 2010; 26 (23): 2979-2980


    Recent advances in single-cell manipulation technology, whole genome amplification and high-throughput sequencing have now made it possible to sequence the genome of an individual cell. The bioinformatic analysis of these genomes, however, is far more complicated than the analysis of those generated using traditional, culture-based methods. In order to simplify this analysis, we have developed SmashCell (Simple Metagenomics Analysis SHell-for sequences from single Cells). It is designed to automate the main steps in microbial genome analysis-assembly, gene prediction, functional annotation-in a way that allows parameter and algorithm exploration at each step in the process. It also manages the data created by these analyses and provides visualization methods for rapid analysis of the results.The SmashCell source code and a comprehensive manual are available at data are available at Bioinformatics online.

    View details for DOI 10.1093/bioinformatics/btq564

    View details for Web of Science ID 000284430900009

    View details for PubMedID 20966005

    View details for PubMedCentralID PMC2982155

  • Microbial invasion of the amniotic cavity in preeclampsia as assessed by cultivation and sequence-based methods JOURNAL OF PERINATAL MEDICINE DiGiulio, D. B., Gervasi, M., Romero, R., Mazaki-Tovi, S., Vaisbuch, E., Kusanovic, J. P., Seok, K. S., Gomez, R., Mittal, P., Gotsch, F., Chaiworapongsa, T., Oyarzun, E., Kim, C. J., Relman, D. A. 2010; 38 (5): 503-513


    Infection has been implicated in the pathogenesis of preeclampsia, yet the association between microbial invasion of the amniotic cavity (MIAC) and preeclampsia has not been determined. The aim of this study was to determine the prevalence, and microbial diversity associated with MIAC, as well as the nature of the host response to MIAC in patients with preeclampsia.Amniotic fluid (AF) from 62 subjects with preeclampsia, not in labor, was analyzed with both cultivation and molecular methods. Broad-range and group-specific PCR assays targeting small subunit ribosomal DNA, or other gene sequences, from bacteria, fungi and archaea were used. Results were correlated with measurements of host inflammatory response, including AF white blood cell count and AF concentrations of glucose, interleukin-6 (IL-6) and MMP-8.1) The rate of MIAC in preeclampsia was 1.6% (1/62) based on cultivation techniques, 8% (5/62) based on PCR, and 9.6% (6/62) based on the combined results of both methods; 2) among the six patients diagnosed with MIAC, three had a positive PCR for Sneathia/Leptotrichia spp.; and 3) patients with MIAC were more likely to have evidence of an inflammatory response in the amniotic cavity than those without MIAC, as determined by a higher median AF IL-6 [1.65 ng/mL interquartile range (IQR): 0.35-4.62 vs. 0.22 ng/mL IQR: 0.12-0.51; P=0.002).The prevalence of MIAC in preeclampsia is low, suggesting that intra-amniotic infection plays only a limited role in preeclampsia. However, the unexpectedly high number of positive AF specimens for Sneathia/Leptotrichia warrants further investigation.

    View details for DOI 10.1515/JPM.2010.078

    View details for Web of Science ID 000281566600009

    View details for PubMedID 20482470

    View details for PubMedCentralID PMC3325506

  • Microbial invasion of the amniotic cavity in pregnancies with small-for-gestational-age fetuses JOURNAL OF PERINATAL MEDICINE DiGiulio, D. B., Gervasi, M. T., Romero, R., Vaisbuch, E., Mazaki-Tovi, S., Kusanovic, J. P., Seok, K. S., Gomez, R., Mittal, P., Gotsch, F., Chaiworapongsa, T., Oyarzun, E., Kim, C. J., Relman, D. A. 2010; 38 (5): 495-502


    Microbial invasion of the amniotic cavity (MIAC) has been detected in women with preterm labor, preterm prelabor rupture of membranes (PROM), and in patients at term with PROM or in spontaneous labor. Intrauterine infection is recognized as a potential cause of fetal growth restriction; yet, the frequency of MIAC in pregnancies with small-for-gestational-age (SGA) fetuses is unknown. The aim of this study was to determine the frequency, diversity and relative abundance of microbes in amniotic fluid (AF) of women with an SGA neonate using a combination of culture and molecular methods.AF from 52 subjects with an SGA neonate was analyzed with both cultivation and molecular methods in a retrospective cohort study. Broad-range and group-specific PCR assays targeted small subunit rDNA, or other gene sequences, from bacteria, fungi and archaea. Results of microbiologic studies were correlated with indices of the host inflammatory response.1) All AF samples (n=52) were negative for microorganisms based on cultivation techniques, whereas 6% (3/52) were positive based on PCR; and 2) intra-amniotic inflammation was detected in one of the three patients with a positive PCR result, as compared with three patients (6.1%) of the 49 with both a negative culture and a negative PCR (P=0.2).MIAC is detected by PCR in some patients with an SGA fetus who were not in labor at the time of AF collection.

    View details for DOI 10.1515/JPM.2010.076

    View details for Web of Science ID 000281566600008

    View details for PubMedID 20482466

    View details for PubMedCentralID PMC2962935

  • Bacterial diversity in the oral cavity of 10 healthy individuals ISME JOURNAL Bik, E. M., Long, C. D., Armitage, G. C., Loomer, P., Emerson, J., Mongodin, E. F., Nelson, K. E., Gill, S. R., Fraser-Liggett, C. M., Relman, D. A. 2010; 4 (8): 962-974


    The composition of the oral microbiota from 10 individuals with healthy oral tissues was determined using culture-independent techniques. From each individual, 26 specimens, each from different oral sites at a single point in time, were collected and pooled. An 11th pool was constructed using portions of the subgingival specimens from all 10 individuals. The 16S ribosomal RNA gene was amplified using broad-range bacterial primers, and clone libraries from the individual and subgingival pools were constructed. From a total of 11,368 high-quality, nonchimeric, near full-length sequences, 247 species-level phylotypes (using a 99% sequence identity threshold) and 9 bacterial phyla were identified. At least 15 bacterial genera were conserved among all 10 individuals, with significant interindividual differences at the species and strain level. Comparisons of these oral bacterial sequences with near full-length sequences found previously in the large intestines and feces of other healthy individuals suggest that the mouth and intestinal tract harbor distinct sets of bacteria. Co-occurrence analysis showed significant segregation of taxa when community membership was examined at the level of genus, but not at the level of species, suggesting that ecologically significant, competitive interactions are more apparent at a broader taxonomic level than species. This study is one of the more comprehensive, high-resolution analyses of bacterial diversity within the healthy human mouth to date, and highlights the value of tools from macroecology for enhancing our understanding of bacterial ecology in human health.

    View details for DOI 10.1038/ismej.2010.30

    View details for Web of Science ID 000280592600002

    View details for PubMedID 20336157

    View details for PubMedCentralID PMC2941673

  • Genetic technologies. Synthetic "life," ethics, national security, and public discourse. Science Cho, M. K., Relman, D. A. 2010; 329 (5987): 38-39

    View details for DOI 10.1126/science.1193749

    View details for PubMedID 20595601

  • Preterm premature rupture of the membranes: current approaches to evaluation and management AMERICAN JOURNAL OF REPRODUCTIVE IMMUNOLOGY DiGiulio, D. B., Romero, R., Kusanovic, J. P., Gomez, R., Kim, C. J., Seok, K. S., Gotsch, F., Mazaki-Tovi, S., Vaisbuch, E., Sanders, K., Bik, E. M., Chaiworapongsa, T., Oyarzun, E., Relman, D. A. 2010; 64 (1): 38-57


    The role played by microbial invasion of the amniotic cavity (MIAC) in preterm pre-labor rupture of membranes (pPROM) is inadequately characterized, in part because of reliance on cultivation-based methods.Amniotic fluid from 204 subjects with pPROM was analyzed with both cultivation and molecular methods in a retrospective cohort study. Broad-range and group-specific polymerase chain reaction (PCR) assays targeted small subunit ribosomal DNA (rDNA), or other gene sequences, from bacteria, fungi, and archaea. Results were correlated with measurements of host inflammation, as well as pregnancy and perinatal outcomes.The prevalence of MIAC was 34% (70/204) by culture, 45% (92/204) by PCR, and 50% (101/204) by both methods combined. The number of bacterial species revealed by PCR (44 species-level phylotypes) was greater than that by culture (14 species) and included as-yet uncultivated taxa. Some taxa detected by PCR have been previously associated with the gastrointestinal tract (e.g., Coprobacillus sp.), the mouth (e.g., Rothia dentocariosa), or the vagina in the setting of bacterial vaginosis (e.g., Atopobium vaginae). The relative risk for histologic chorioamnionitis was 2.1 for a positive PCR [95% confidence interval (CI), 1.4-3.0] and 2.0 for a positive culture (95% CI, 1.4-2.7). Bacterial rDNA abundance exhibited a dose relationship with gestational age at delivery (R(2) = 0.26; P < 0.01). A positive PCR was associated with lower mean birthweight, and with higher rates of respiratory distress syndrome and necrotizing enterocolitis (P < 0.05 for each outcome).MIAC in pPROM is more common than previously recognized and is associated in some cases with uncultivated taxa, some of which are typically associated with the gastrointestinal tract. The detection of MIAC by molecular methods has clinical significance.

    View details for DOI 10.1111/j.1600-0897.2010.00830.x

    View details for Web of Science ID 000278395800008

    View details for PubMedID 20331587

    View details for PubMedCentralID PMC2907911

  • The biological century: coming to terms with risk in the life sciences NATURE IMMUNOLOGY Relman, D. A. 2010; 11 (4): 275-278

    View details for DOI 10.1038/ni0410-275

    View details for Web of Science ID 000275849500002

    View details for PubMedID 20300132

  • Dissecting Interferon-Induced Transcriptional Programs in Human Peripheral Blood Cells PLOS ONE Waddell, S. J., Popper, S. J., Rubins, K. H., Griffiths, M. J., Brown, P. O., Levin, M., Relman, D. A. 2010; 5 (3)


    Interferons are key modulators of the immune system, and are central to the control of many diseases. The response of immune cells to stimuli in complex populations is the product of direct and indirect effects, and of homotypic and heterotypic cell interactions. Dissecting the global transcriptional profiles of immune cell populations may provide insights into this regulatory interplay. The host transcriptional response may also be useful in discriminating between disease states, and in understanding pathophysiology. The transcriptional programs of cell populations in health therefore provide a paradigm for deconvoluting disease-associated gene expression profiles.We used human cDNA microarrays to (1) compare the gene expression programs in human peripheral blood mononuclear cells (PBMCs) elicited by 6 major mediators of the immune response: interferons alpha, beta, omega and gamma, IL12 and TNFalpha; and (2) characterize the transcriptional responses of purified immune cell populations (CD4+ and CD8+ T cells, B cells, NK cells and monocytes) to IFNgamma stimulation. We defined a highly stereotyped response to type I interferons, while responses to IFNgamma and IL12 were largely restricted to a subset of type I interferon-inducible genes. TNFalpha stimulation resulted in a distinct pattern of gene expression. Cell type-specific transcriptional programs were identified, highlighting the pronounced response of monocytes to IFNgamma, and emergent properties associated with IFN-mediated activation of mixed cell populations. This information provides a detailed view of cellular activation by immune mediators, and contributes an interpretive framework for the definition of host immune responses in a variety of disease settings.

    View details for DOI 10.1371/journal.pone.0009753

    View details for Web of Science ID 000275894300006

    View details for PubMedID 20339534

    View details for PubMedCentralID PMC2842296

  • Microbial threat lists: obstacles in the quest for biosecurity? Nature reviews. Microbiology Casadevall, A., Relman, D. A. 2010; 8 (2): 149-154


    Anxiety about threats from the microbial world and about the deliberate misuse of microorganisms has led to efforts to define and control these dangers using lists and regulations. One list with tremendous legal implications and a potentially huge impact on research is the Select Agents and Toxins List, which was created by the US Government to limit the possession of and access to particular microorganisms and toxins. In this article, in addition to highlighting general problems with taxonomy-based, microorganism-centric lists, we discuss our view that such lists may have the paradoxical effect of increasing the societal vulnerability to biological attack and natural epidemics by interfering with the sharing of microbial samples and hindering research on vaccines and therapeutics.

    View details for DOI 10.1038/nrmicro2299

    View details for PubMedID 20065941

  • SCIENCE AND SOCIETY Microbial threat lists: obstacles in the quest for biosecurity? NATURE REVIEWS MICROBIOLOGY Casadevall, A., Relman, D. A. 2010; 8 (2): 149-154
  • 2020 visions NATURE Norvig, P., Relman, D. A., Goldstein, D. B., Kammen, D. M., Weinberger, D. R., Aiello, L. C., Church, G., Hennessy, J. L., Sachs, J., Burrows, A., Pisano, G. P., Goldstein, J. R., Anastas, P., Klausner, R., Baltimore, D., Montgomery, D. R., Baer, T. M., Bigelow, N. P., Holt, R. D., Nicholson, J. K. 2010; 463 (7277): 26-32

    View details for DOI 10.1038/463026a

    View details for Web of Science ID 000273344900016

    View details for PubMedID 20054379

  • A molecular investigation of the microbial diversity and burden in preterm PROM reveals a high rate of infection with a broad range of organisms including gastrointestinal tract microbiota 30th Annual Clinical Meeting of the Society-for-Maternal-Fetal-Medicine Digiulio, D., Romero, R., Kusanovic, J. P., Gomez, R., Kim, C. J., Seok, K., Gotsch, F., Mazaki-Tovi, S., Vaisbuch, E., Sanders, K., Bik, E., Chaiworapongsa, T., Relman, D. MOSBY-ELSEVIER. 2009: S198?S199
  • Gene Transcript Abundance Profiles Distinguish Kawasaki Disease from Adenovirus Infection Joint Meeting of the Pediatric-Academic-Societies/Asian-Society-for-Pediatric-Research Popper, S. J., Watson, V. E., Shimizu, C., Kanegaye, J. T., Burns, J. C., Relman, D. A. OXFORD UNIV PRESS INC. 2009: 657?66


    Acute Kawasaki disease (KD) is difficult to distinguish from other illnesses that involve acute rash or fever, in part because the etiologic agent(s) and pathophysiology remain poorly characterized. As a result, diagnosis and critical therapies may be delayed.We used DNA microarrays to identify possible diagnostic features of KD. We compared gene expression patterns in the blood of 23 children with acute KD and 18 age-matched febrile children with 3 illnesses that resemble KD.Genes associated with platelet and neutrophil activation were expressed at higher levels in patients with KD than in patients with acute adenovirus infections or systemic adverse drug reactions, but levels in patients with KD were not higher than those in patients with scarlet fever. Genes associated with B cell activation were also expressed at higher levels in patients with KD than in control subjects. A striking absence of interferon-stimulated gene expression in patients with KD was confirmed in an independent cohort of patients with KD. Using a set of 38 gene transcripts, we successfully predicted the diagnosis for 21 of 23 patients with KD and 7 of 8 patients with adenovirus infection.These findings provide insight into the molecular features that distinguish KD from other febrile illnesses and support the feasibility of developing novel diagnostic reagents for KD based on the host response.

    View details for DOI 10.1086/603538

    View details for Web of Science ID 000268009700024

    View details for PubMedID 19583510

    View details for PubMedCentralID PMC2878183

  • Majority Rules? Tallying the Microbial Census in an Abscess by Means of Molecular Methods CLINICAL INFECTIOUS DISEASES DiGiulio, D. B., Relman, D. A. 2009; 48 (9): 1179-1181

    View details for DOI 10.1086/597579

    View details for Web of Science ID 000264897300002

    View details for PubMedID 19335163

  • Natural-host animal models indicate functional interchangeability between the filamentous haemagglutinins of Bordetella pertussis and Bordetella bronchiseptica and reveal a role for the mature C-terminal domain, but not the RGD motif, during infection MOLECULAR MICROBIOLOGY Julio, S. M., Inatsuka, C. S., Mazar, J., Dieterich, C., Relman, D. A., Cotter, P. A. 2009; 71 (6): 1574-1590


    Bacteria of the Bordetella genus cause respiratory tract infections. Both broad host range (e.g. Bordetella bronchiseptica) and human-adapted (e.g. Bordetella pertussis) strains produce a surface-exposed and secreted protein called filamentous haemagglutinin (FHA) that functions in adherence and immunomodulation. Previous studies using B. pertussis and cultured mammalian cells identified several FHA domains with potential roles in host cell interactions, including an Arg-Gly-Asp (RGD) triplet that was reported to bind integrins on epithelial cells and monocytes to activate host signalling pathways. We show here that, in contrast to our previous report, the fhaB genes of B. pertussis and B. bronchiseptica are functionally interchangeable, at least with regard to the various in vitro and in vivo assays investigated. This result is significant because it indicates that information obtained studying FHA using B. bronchiseptica and natural-host animal models should apply to B. pertussis FHA as well. We also show that the C-terminus of mature FHA, which we name the MCD, mediates adherence to epithelial and macrophage-like cells and is required for colonization of the rat respiratory tract and modulation of the inflammatory response in mouse lungs. We could not, however, detect a role for the RGD in any of these processes.

    View details for DOI 10.1111/j.1365-2958.2009.06623.x

    View details for Web of Science ID 000264187700017

    View details for PubMedID 19220744

  • Cross-talk in the gut GENOME BIOLOGY Dinalo, J. E., Relman, D. A. 2009; 10 (1)


    Modulation of host signaling by the products of microbial activity in the gut may affect weight gain and fat formation.

    View details for DOI 10.1186/gb-2009-10-1-203

    View details for Web of Science ID 000263823200004

    View details for PubMedID 19216729

  • CHARACTERIZATION OF THE GENE EXPRESSION PROGRAMS ASSOCIATED WITH DISEASE SEVERITY IN ACUTE PEDIATRIC DENGUE INFECTION 57th Annual Meeting of the American-Society-of-Tropical-Medicine-and-Hygiene Popper, S. J., Gordon, A., Liu, M., Vargas, M. J., Perry, C., Balmaseda, A., Rocha, C., Harris, E., Relman, D. A. AMER SOC TROP MED & HYGIENE. 2008: 5?5
  • Modulation of the NF-kappa B Pathway by Bordetella pertussis Filamentous Hemagglutinin PLOS ONE Abramson, T., Kedem, H., Relman, D. A. 2008; 3 (11)


    Filamentous hemagglutinin (FHA) is a cell-associated and secreted adhesin produced by Bordetella pertussis with pro-apoptotic and pro-inflammatory activity in host cells. Given the importance of the NF-kappaB transcription factor family in these host cell responses, we examined the effect of FHA on NF-kappaB activation in macrophages and bronchial epithelial cells, both of which are relevant cell types during natural infection.Exposure to FHA of primary human monocytes and transformed U-937 macrophages, but not BEAS-2B epithelial cells, resulted in early activation of the NF-kappaB pathway, as manifested by the degradation of cytosolic IkappaB alpha, by NF-kappaB DNA binding, and by the subsequent secretion of NF-kappaB-regulated inflammatory cytokines. However, exposure of macrophages and human monocytes to FHA for two hours or more resulted in the accumulation of cytosolic IkappaB alpha, and the failure of TNF-alpha to activate NF-kappaB. Proteasome activity was attenuated following exposure of cells to FHA for 2 hours, as was the nuclear translocation of RelA in BEAS-2B cells.These results reveal a complex temporal dynamic, and suggest that despite short term effects to the contrary, longer exposures of host cells to this secreted adhesin may block NF-kappaB activation, and perhaps lead to a compromised immune response to this bacterial pathogen.

    View details for DOI 10.1371/journal.pone.0003825

    View details for Web of Science ID 000265451500006

    View details for PubMedID 19043589

  • The Pervasive Effects of an Antibiotic on the Human Gut Microbiota, as Revealed by Deep 16S rRNA Sequencing PLOS BIOLOGY Dethlefsen, L., Huse, S., Sogin, M. L., Relman, D. A. 2008; 6 (11): 2383-2400


    The human intestinal microbiota is essential to the health of the host and plays a role in nutrition, development, metabolism, pathogen resistance, and regulation of immune responses. Antibiotics may disrupt these coevolved interactions, leading to acute or chronic disease in some individuals. Our understanding of antibiotic-associated disturbance of the microbiota has been limited by the poor sensitivity, inadequate resolution, and significant cost of current research methods. The use of pyrosequencing technology to generate large numbers of 16S rDNA sequence tags circumvents these limitations and has been shown to reveal previously unexplored aspects of the "rare biosphere." We investigated the distal gut bacterial communities of three healthy humans before and after treatment with ciprofloxacin, obtaining more than 7,000 full-length rRNA sequences and over 900,000 pyrosequencing reads from two hypervariable regions of the rRNA gene. A companion paper in PLoS Genetics (see Huse et al., doi: 10.1371/journal.pgen.1000255) shows that the taxonomic information obtained with these methods is concordant. Pyrosequencing of the V6 and V3 variable regions identified 3,300-5,700 taxa that collectively accounted for over 99% of the variable region sequence tags that could be obtained from these samples. Ciprofloxacin treatment influenced the abundance of about a third of the bacterial taxa in the gut, decreasing the taxonomic richness, diversity, and evenness of the community. However, the magnitude of this effect varied among individuals, and some taxa showed interindividual variation in the response to ciprofloxacin. While differences of community composition between individuals were the largest source of variability between samples, we found that two unrelated individuals shared a surprising degree of community similarity. In all three individuals, the taxonomic composition of the community closely resembled its pretreatment state by 4 weeks after the end of treatment, but several taxa failed to recover within 6 months. These pervasive effects of ciprofloxacin on community composition contrast with the reports by participants of normal intestinal function and with prior assumptions of only modest effects of ciprofloxacin on the intestinal microbiota. These observations support the hypothesis of functional redundancy in the human gut microbiota. The rapid return to the pretreatment community composition is indicative of factors promoting community resilience, the nature of which deserves future investigation.

    View details for DOI 10.1371/journal.pbio.0060280

    View details for Web of Science ID 000261187900010

    View details for PubMedID 19018661

    View details for PubMedCentralID PMC2586385

  • Exploring Microbial Diversity and Taxonomy Using SSU rRNA Hypervariable Tag Sequencing PLOS GENETICS Huse, S. M., Dethlefsen, L., Huber, J. A., Welch, D. M., Relman, D. A., Sogin, M. L. 2008; 4 (11)


    Massively parallel pyrosequencing of hypervariable regions from small subunit ribosomal RNA (SSU rRNA) genes can sample a microbial community two or three orders of magnitude more deeply per dollar and per hour than capillary sequencing of full-length SSU rRNA. As with full-length rRNA surveys, each sequence read is a tag surrogate for a single microbe. However, rather than assigning taxonomy by creating gene trees de novo that include all experimental sequences and certain reference taxa, we compare the hypervariable region tags to an extensive database of rRNA sequences and assign taxonomy based on the best match in a Global Alignment for Sequence Taxonomy (GAST) process. The resulting taxonomic census provides information on both composition and diversity of the microbial community. To determine the effectiveness of using only hypervariable region tags for assessing microbial community membership, we compared the taxonomy assigned to the V3 and V6 hypervariable regions with the taxonomy assigned to full-length SSU rRNA sequences isolated from both the human gut and a deep-sea hydrothermal vent. The hypervariable region tags and full-length rRNA sequences provided equivalent taxonomy and measures of relative abundance of microbial communities, even for tags up to 15% divergent from their nearest reference match. The greater sampling depth per dollar afforded by massively parallel pyrosequencing reveals many more members of the "rare biosphere" than does capillary sequencing of the full-length gene. In addition, tag sequencing eliminates cloning bias and the sequences are short enough to be completely sequenced in a single read, maximizing the number of organisms sampled in a run while minimizing chimera formation. This technique allows the cost-effective exploration of changes in microbial community structure, including the rare biosphere, over space and time and can be applied immediately to initiatives, such as the Human Microbiome Project.

    View details for DOI 10.1371/journal.pgen.1000255

    View details for Web of Science ID 000261481000010

    View details for PubMedID 19023400

  • 'Til death do us part': coming to terms with symbiotic relationships - Foreword NATURE REVIEWS MICROBIOLOGY Relman, D. A. 2008; 6 (10): 721-724


    Symbiotic interactions of microorganisms are widespread in nature, and support fundamentally important processes in diverse areas of biology that range from health and disease to ecology and the environment. Here, David Relman discusses the selection of articles in this Focus issue, which reflects the exciting advances in our understanding of intimate partnerships between organisms and their environments.

    View details for DOI 10.1038/nrmicro1990

    View details for Web of Science ID 000259217200020

    View details for PubMedID 19086265

  • Microbial Prevalence, Diversity and Abundance in Amniotic Fluid During Preterm Labor: A Molecular and Culture-Based Investigation PLOS ONE DiGiulio, D. B., Romero, R., Amogan, H. P., Kusanovic, J. P., Bik, E. M., Gotsch, F., Kim, C. J., Erez, O., Edwin, S., Relman, D. A. 2008; 3 (8)


    Preterm delivery causes substantial neonatal mortality and morbidity. Unrecognized intra-amniotic infections caused by cultivation-resistant microbes may play a role. Molecular methods can detect, characterize and quantify microbes independently of traditional culture techniques. However, molecular studies that define the diversity and abundance of microbes invading the amniotic cavity, and evaluate their clinical significance within a causal framework, are lacking.In parallel with culture, we used broad-range end-point and real-time PCR assays to amplify, identify and quantify ribosomal DNA (rDNA) of bacteria, fungi and archaea from amniotic fluid of 166 women in preterm labor with intact membranes. We sequenced up to 24 rRNA clones per positive specimen and assigned taxonomic designations to approximately the species level. Microbial prevalence, diversity and abundance were correlated with host inflammation and with gestational and neonatal outcomes. Study subjects who delivered at term served as controls. The combined use of molecular and culture methods revealed a greater prevalence (15% of subjects) and diversity (18 taxa) of microbes in amniotic fluid than did culture alone (9.6% of subjects; 11 taxa). The taxa detected only by PCR included a related group of fastidious bacteria, comprised of Sneathia sanguinegens, Leptotrichia amnionii and an unassigned, uncultivated, and previously-uncharacterized bacterium; one or more members of this group were detected in 25% of positive specimens. A positive PCR was associated with histologic chorioamnionitis (adjusted odds ratio [OR] 20; 95% CI, 2.4 to 172), and funisitis (adjusted OR 18; 95% CI, 3.1 to 99). The positive predictive value of PCR for preterm delivery was 100 percent. A temporal association between a positive PCR and delivery was supported by a shortened amniocentesis-to-delivery interval (adjusted hazard ratio 4.6; 95% CI, 2.2 to 9.5). A dose-response association was demonstrated between bacterial rDNA abundance and gestational age at delivery (r(2) = 0.42; P<0.002).The amniotic cavity of women in preterm labor harbors DNA from a greater diversity of microbes than previously suspected, including as-yet uncultivated, previously-uncharacterized taxa. The strength, temporality and gradient with which these microbial sequence types are associated with preterm delivery support a causal relationship.

    View details for DOI 10.1371/journal.pone.0003056

    View details for Web of Science ID 000264796300003

    View details for PubMedID 18725970

    View details for PubMedCentralID PMC2516597

  • Comparative Analysis of Viral Gene Expression Programs during Poxvirus Infection: A Transcriptional Map of the Vaccinia and Monkeypox Genomes PLOS ONE Rubins, K. H., Hensley, L. E., Bell, G. W., Wang, C., Lefkowitz, E. J., Brown, P. O., Relman, D. A. 2008; 3 (7)


    Poxviruses engage in a complex and intricate dialogue with host cells as part of their strategy for replication. However, relatively little molecular detail is available with which to understand the mechanisms behind this dialogue.We designed a specialized microarray that contains probes specific to all predicted ORFs in the Monkeypox Zaire (MPXV) and Vaccinia Western Reserve (VACV) genomes, as well as >18,000 human genes, and used this tool to characterize MPXV and VACV gene expression responses in vitro during the course of primary infection of human monocytes, primary human fibroblasts and HeLa cells. The two viral transcriptomes show distinct features of temporal regulation and species-specific gene expression, and provide an early foundation for understanding global gene expression responses during poxvirus infection.The results provide a temporal map of the transcriptome of each virus during infection, enabling us to compare viral gene expression across species, and classify expression patterns of previously uncharacterized ORFs.

    View details for DOI 10.1371/journal.pone.0002628

    View details for Web of Science ID 000264065800030

    View details for PubMedID 18612436

  • Learning to appreciate our differences JOURNAL OF INFECTIOUS DISEASES Relman, D. A. 2008; 198 (1): 4-5

    View details for DOI 10.1086/588672

    View details for Web of Science ID 000256632800002

    View details for PubMedID 18454681

  • Microbiology in the post-genomic era NATURE REVIEWS MICROBIOLOGY Medini, D., Serruto, D., Parkhill, J., Relman, D. A., Donati, C., Moxon, R., Falkow, S., Rappuoli, R. 2008; 6 (6): 419-430


    Genomics has revolutionized every aspect of microbiology. Now, 13 years after the first bacterial genome was sequenced, it is important to pause and consider what has changed in microbiology research as a consequence of genomics. In this article, we review the evolving field of bacterial typing and the genomic technologies that enable comparative analysis of multiple genomes and the metagenomes of complex microbial environments, and address the implications of the genomic era for the future of microbiology.

    View details for DOI 10.1038/nrmicro1901

    View details for Web of Science ID 000255953300009

    View details for PubMedID 18475305

  • Linking microbial phylogeny to metabolic activity at the single-cell level by using enhanced element labeling-catalyzed reporter deposition fluorescence in situ hybridization (EL-FISH) and NanoSIMS APPLIED AND ENVIRONMENTAL MICROBIOLOGY Behrens, S., Loesekann, T., Pett-Ridge, J., Weber, P. K., Ng, W., Stevenson, B. S., Hutcheon, I. D., Relman, D. A., Spormann, A. M. 2008; 74 (10): 3143-3150


    To examine phylogenetic identity and metabolic activity of individual cells in complex microbial communities, we developed a method which combines rRNA-based in situ hybridization with stable isotope imaging based on nanometer-scale secondary-ion mass spectrometry (NanoSIMS). Fluorine or bromine atoms were introduced into cells via 16S rRNA-targeted probes, which enabled phylogenetic identification of individual cells by NanoSIMS imaging. To overcome the natural fluorine and bromine backgrounds, we modified the current catalyzed reporter deposition fluorescence in situ hybridization (FISH) technique by using halogen-containing fluorescently labeled tyramides as substrates for the enzymatic tyramide deposition. Thereby, we obtained an enhanced element labeling of microbial cells by FISH (EL-FISH). The relative cellular abundance of fluorine or bromine after EL-FISH exceeded natural background concentrations by up to 180-fold and allowed us to distinguish target from non-target cells in NanoSIMS fluorine or bromine images. The method was optimized on single cells of axenic Escherichia coli and Vibrio cholerae cultures. EL-FISH/NanoSIMS was then applied to study interrelationships in a dual-species consortium consisting of a filamentous cyanobacterium and a heterotrophic alphaproteobacterium. We also evaluated the method on complex microbial aggregates obtained from human oral biofilms. In both samples, we found evidence for metabolic interactions by visualizing the fate of substrates labeled with (13)C-carbon and (15)N-nitrogen, while individual cells were identified simultaneously by halogen labeling via EL-FISH. Our novel approach will facilitate further studies of the ecophysiology of known and uncultured microorganisms in complex environments and communities.

    View details for DOI 10.1128/AEM.00191-08

    View details for Web of Science ID 000256074900028

    View details for PubMedID 18359832

  • Host transmission of Salmonella enterica serovar Typhimurium is controlled by virulence factors and indigenous intestinal microbiota INFECTION AND IMMUNITY Lawley, T. D., Bouley, D. A., Hoy, Y. E., Gerke, C., Relman, D. A., Monack, D. M. 2008; 76 (1): 403-416


    Transmission is an essential stage of a pathogen's life cycle and remains poorly understood. We describe here a model in which persistently infected 129X1/SvJ mice provide a natural model of Salmonella enterica serovar Typhimurium transmission. In this model only a subset of the infected mice, termed supershedders, shed high levels (>10(8) CFU/g) of Salmonella serovar Typhimurium in their feces and, as a result, rapidly transmit infection. While most Salmonella serovar Typhimurium-infected mice show signs of intestinal inflammation, only supershedder mice develop colitis. Development of the supershedder phenotype depends on the virulence determinants Salmonella pathogenicity islands 1 and 2, and it is characterized by mucosal invasion and, importantly, high luminal abundance of Salmonella serovar Typhimurium within the colon. Immunosuppression of infected mice does not induce the supershedder phenotype, demonstrating that the immune response is not the main determinant of Salmonella serovar Typhimurium levels within the colon. In contrast, treatment of mice with antibiotics that alter the health-associated indigenous intestinal microbiota rapidly induces the supershedder phenotype in infected mice and predisposes uninfected mice to the supershedder phenotype for several days. These results demonstrate that the intestinal microbiota plays a critical role in controlling Salmonella serovar Typhimurium infection, disease, and transmissibility. This novel model should facilitate the study of host, pathogen, and intestinal microbiota factors that contribute to infectious disease transmission.

    View details for DOI 10.1128/IAI.01189-07

    View details for Web of Science ID 000252126000043

    View details for PubMedID 17967858

  • Building a better virus trap TRENDS IN BIOTECHNOLOGY Long, C. D., Turner-Shelef, K., Relman, D. A. 2007; 25 (12): 535-538


    The concept of ecological 'traps' is based in theory from ecology and conservation biology that has now found application to infectious diseases with a study from Paul Turner's group. This study is important because it offers a mathematical model of ecological traps, applies this model to viruses, and tests the model in a bacteria-phage system. Although there will be technical hurdles to overcome, this concept might lead to benefits for both health and industry.

    View details for DOI 10.1016/j.tibtech.2007.08.013

    View details for Web of Science ID 000251738200001

    View details for PubMedID 17997180

  • An ecological and evolutionary perspective on human-microbe mutualism and disease NATURE Dethlefsen, L., McFall-Ngai, M., Relman, D. A. 2007; 449 (7164): 811-818


    The microbial communities of humans are characteristic and complex mixtures of microorganisms that have co-evolved with their human hosts. The species that make up these communities vary between hosts as a result of restricted migration of microorganisms between hosts and strong ecological interactions within hosts, as well as host variability in terms of diet, genotype and colonization history. The shared evolutionary fate of humans and their symbiotic bacteria has selected for mutualistic interactions that are essential for human health, and ecological or genetic changes that uncouple this shared fate can result in disease. In this way, looking to ecological and evolutionary principles might provide new strategies for restoring and maintaining human health.

    View details for DOI 10.1038/nature06245

    View details for Web of Science ID 000250230600030

    View details for PubMedID 17943117

  • Dissecting biological "dark matter" with single-cell genetic analysis of rare and uncultivated TM7 microbes from the human mouth PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Marcy, Y., Ouverney, C., Bik, E. M., Loesekann, T., Ivanova, N., Martin, H. G., Szeto, E., Platt, D., Hugenholtz, P., Relman, D. A., Quake, S. R. 2007; 104 (29): 11889-11894


    We have developed a microfluidic device that allows the isolation and genome amplification of individual microbial cells, thereby enabling organism-level genomic analysis of complex microbial ecosystems without the need for culture. This device was used to perform a directed survey of the human subgingival crevice and to isolate bacteria having rod-like morphology. Several isolated microbes had a 16S rRNA sequence that placed them in candidate phylum TM7, which has no cultivated or sequenced members. Genome amplification from individual TM7 cells allowed us to sequence and assemble >1,000 genes, providing insight into the physiology of members of this phylum. This approach enables single-cell genetic analysis of any uncultivated minority member of a microbial community.

    View details for DOI 10.1073/pnas.0704662104

    View details for Web of Science ID 000248199200007

    View details for PubMedID 17620602

  • Development of the human infant intestinal microbiota PLOS BIOLOGY Palmer, C., Bik, E. M., DiGiulio, D. B., Relman, D. A., Brown, P. O. 2007; 5 (7): 1556-1573


    Almost immediately after a human being is born, so too is a new microbial ecosystem, one that resides in that person's gastrointestinal tract. Although it is a universal and integral part of human biology, the temporal progression of this process, the sources of the microbes that make up the ecosystem, how and why it varies from one infant to another, and how the composition of this ecosystem influences human physiology, development, and disease are still poorly understood. As a step toward systematically investigating these questions, we designed a microarray to detect and quantitate the small subunit ribosomal RNA (SSU rRNA) gene sequences of most currently recognized species and taxonomic groups of bacteria. We used this microarray, along with sequencing of cloned libraries of PCR-amplified SSU rDNA, to profile the microbial communities in an average of 26 stool samples each from 14 healthy, full-term human infants, including a pair of dizygotic twins, beginning with the first stool after birth and continuing at defined intervals throughout the first year of life. To investigate possible origins of the infant microbiota, we also profiled vaginal and milk samples from most of the mothers, and stool samples from all of the mothers, most of the fathers, and two siblings. The composition and temporal patterns of the microbial communities varied widely from baby to baby. Despite considerable temporal variation, the distinct features of each baby's microbial community were recognizable for intervals of weeks to months. The strikingly parallel temporal patterns of the twins suggested that incidental environmental exposures play a major role in determining the distinctive characteristics of the microbial community in each baby. By the end of the first year of life, the idiosyncratic microbial ecosystems in each baby, although still distinct, had converged toward a profile characteristic of the adult gastrointestinal tract.

    View details for Web of Science ID 000249124400020

    View details for PubMedID 17594176

  • Phase variation and microevolution at homopolymeric tracts in Bordetella pertussis BMC GENOMICS Gogol, E. B., Cummings, C. A., Burns, R. C., Relman, D. A. 2007; 8


    Bordetella pertussis, the causative agent of whooping cough, is a highly clonal pathogen of the respiratory tract. Its lack of genetic diversity, relative to many bacterial pathogens, could limit its ability to adapt to a hostile and changing host environment. This limitation might be overcome by phase variation, as observed for other mucosal pathogens. One of the most common mechanisms of phase variation is reversible expansion or contraction of homopolymeric tracts (HPTs).The genomes of B. pertussis and the two closely related species, B. bronchiseptica and B. parapertussis, were screened for homopolymeric tracts longer than expected on the basis of chance, given their nucleotide compositions. Sixty-nine such HPTs were found in total among the three genomes, 74% of which were polymorphic among the three species. Nine HPTs were genotyped in a collection of 90 geographically and temporally diverse B. pertussis strains using the polymerase chain reaction/ligase detection reaction (PCR/LDR) assay. Six HPTs were polymorphic in this collection of B. pertussis strains. Of note, one of these polymorphic HPTs was found in the fimX promoter, where a single base insertion variant was present in seven strains, all of which were isolated prior to introduction of the pertussis vaccine. Transcript abundance of fimX was found to be 3.8-fold lower in strains carrying the longer allele. HPTs in three other genes, tcfA, bapC, and BP3651, varied widely in composition across the strain collection and displayed allelic polymorphism within single cultures.Allelic polymorphism at homopolymeric tracts is common within the B. pertussis genome. Phase variability may be an important mechanism in B. pertussis for evasion of the immune system and adaptation to different niches in the human host. High sensitivity and specificity make the PCR/LDR assay a powerful tool for investigating allelic variation at HPTs. Using this method, allelic diversity and phase variation were demonstrated at several B. pertussis loci.

    View details for DOI 10.1186/1471-2164-8-122

    View details for Web of Science ID 000247181800001

    View details for PubMedID 17509142

  • Patterns of host genome-wide gene transcript abundance in the peripheral blood of patients with acute dengue hemorrhagic fever JOURNAL OF INFECTIOUS DISEASES Simmons, C. P., Popper, S., Dolocek, C., Chau, T. N., Griffiths, M., Dung, N. T., Long, T. H., Hoang, D. M., Chau, N. V., Thao, L. T., Hien, T. T., Relman, D. A., Farrar, J. 2007; 195 (8): 1097-1107


    Responses by peripheral blood leukocytes may contribute to the pathogenesis of dengue hemorrhagic fever (DHF). We used DNA microarrays to reveal transcriptional patterns in the blood of 14 adults with DHF. Acute DHF was defined by an abundance of transcripts from cell cycle- and endoplasmic reticulum (ER)-related genes, suggesting a proliferative response accompanied by ER stress. Transcript-abundance levels for immunoresponse-associated genes, including cell surface markers, immunoglobulin, and innate response elements, were also elevated. Twenty-four genes were identified for which transcript abundance distinguished patients with dengue shock syndrome (DSS) from those without DSS. All the gene transcripts associated with DSS, many of which are induced by type I interferons, were less abundant in patients with DSS than in those without DSS. To our knowledge, these data provide the first snapshot of gene-expression patterns in peripheral blood during acute dengue and suggest that DSS is associated with attenuation of selected aspects of the innate host response.

    View details for DOI 10.1086/512162

    View details for Web of Science ID 000245405100005

    View details for PubMedID 17357045

  • The role of microbes in Crohn's disease CLINICAL INFECTIOUS DISEASES Eckburg, P. B., Relman, D. A. 2007; 44 (2): 256-262


    Despite decades of research, the etiology of Crohn's disease (CD) remains unknown. Its pathogenesis may involve a complex interplay between host genetics, immune dysfunction, and microbial or environmental factors. Microorganisms, including pathogens and members of the indigenous microbiota, may initiate or propagate the inflammatory process in CD. The pathogenesis of CD has been difficult to study, owing to the broad spectrum of typically nonspecific clinical manifestations, the complexity of environmental and genetic factors, the lack of an accurate model of disease, and the limitations of microbiological methods. A more useful and relevant paradigm for the etiology of CD might be based on the idea of a pathogenic microbial community profile and might emphasize the role of interactive sets of microbes, rather than the role of individual organisms. We review how microbes may participate in the pathogenesis of CD and how they may inappropriately activate the mucosal immune system in genetically predisposed individuals.

    View details for Web of Science ID 000242955700016

    View details for PubMedID 17173227

  • The temporal program of peripheral blood gene expression in the response of nonhuman primates to Ebola hemorrhagic fever GENOME BIOLOGY Rubins, K. H., Hensley, L. E., Wahl-Jensen, V., DiCaprio, K. M., Young, H. A., Reed, D. S., Jahrling, P. B., Brown, P. O., Relman, D. A., Geisbert, T. W. 2007; 8 (8)


    Infection with Ebola virus (EBOV) causes a fulminant and often fatal hemorrhagic fever. In order to improve our understanding of EBOV pathogenesis and EBOV-host interactions, we examined the molecular features of EBOV infection in vivo.Using high-density cDNA microarrays, we analyzed genome-wide host expression patterns in sequential blood samples from nonhuman primates infected with EBOV. The temporal program of gene expression was strikingly similar between animals. Of particular interest were features of the data that reflect the interferon response, cytokine signaling, and apoptosis. Transcript levels for tumor necrosis factor-alpha converting enzyme (TACE)/alpha-disintegrin and metalloproteinase (ADAM)-17 increased during days 4 to 6 after infection. In addition, the serum concentration of cleaved Ebola glycoprotein (GP2 delta) was elevated in late-stage EBOV infected animals. Of note, we were able to detect changes in gene expression of more than 300 genes before symptoms appeared.These results provide the first genome-wide ex vivo analysis of the host response to systemic filovirus infection and disease. These data may elucidate mechanisms of viral pathogenesis and host defense, and may suggest targets for diagnostic and therapeutic development.

    View details for DOI 10.1186/gb-2007-8-8-r174

    View details for Web of Science ID 000253938500026

    View details for PubMedID 17725815

  • The importance of individuals and scale: moving towards single cell microbiology ENVIRONMENTAL MICROBIOLOGY Dethlefsen, L., Relman, D. A. 2007; 9 (1): 8-10

    View details for Web of Science ID 000243294900008

    View details for PubMedID 17227405

  • Gene-expression patterns reveal underlying biological processes in Kawasaki disease GENOME BIOLOGY Popper, S. J., Shimizu, C., Shike, H., Kanegaye, J. T., Newburger, J. W., Sundel, R. P., Brown, P. O., Burns, J. C., Relman, D. A. 2007; 8 (12)


    Kawasaki disease (KD) is an acute self-limited vasculitis and the leading cause of acquired heart disease in children in developed countries. No etiologic agent(s) has been identified, and the processes that mediate formation of coronary artery aneurysms and abatement of fever following treatment with intravenous immunoglobulin (IVIG) remain poorly understood.In an initial survey, we used DNA microarrays to examine patterns of gene expression in peripheral whole blood from 20 children with KD; each was sampled during the acute, subacute, and convalescent phases of the illness. Acute KD was characterized by increased relative abundance of gene transcripts associated with innate immune and proinflammatory responses and decreased abundance of transcripts associated with natural killer cells and CD8+ lymphocytes. There was significant temporal variation in transcript levels during the acute disease phase and stabilization thereafter. We confirmed these temporal patterns in a second cohort of 64 patients, and identified additional inter-individual differences in transcript abundance. Notably, higher levels of transcripts of the gene for carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) were associated with an increased percentage of unsegmented neutrophils, fewer days of illness, higher levels of C-reactive protein, and subsequent non-response to IVIG; this last association was confirmed by quantitative reverse transcription PCR in a third cohort of 33 patients, and was independent of day of illness.Acute KD is characterized by dynamic and variable gene-expression programs that highlight the importance of neutrophil activation state and apoptosis in KD pathogenesis. Our findings also support the feasibility of extracting biomarkers associated with clinical prognosis from gene-expression profiles of individuals with systemic inflammatory illnesses.

    View details for DOI 10.1186/gb-2007-8-12-r261

    View details for Web of Science ID 000253451800009

    View details for PubMedID 18067656

  • Differential NF-kappaB responses induced by Bordetella pertussis Filamentous-Hemagglutinin (FHA) in macrophages and bronchial epithelial cells 7th Annual Meeting of the Federation-of-Clinical-Immunology-Societies Abramson, T., Relman, D., Kedem, H. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2007: S37?S37
  • How bacterial communities expand functional repertoires PLOS BIOLOGY Versalovic, J., Relman, D. 2006; 4 (12): 2193-2195

    View details for DOI 10.1371/journal.pbio.0040430

    View details for Web of Science ID 000242789100003

    View details for PubMedID 17238278

  • Characterization of a highly conserved island in the otherwise divergent Bordetella holmesii and Bordetella pertussis genomes JOURNAL OF BACTERIOLOGY Diavatopoulos, D. A., Cummings, C. A., van der Heide, H. G., van Gent, M., Liew, S., Relman, D. A., Mooi, F. R. 2006; 188 (24): 8385-8394


    The recently discovered pathogen Bordetella holmesii has been isolated from the airways and blood of diseased humans. Genetic events contributing to the emergence of B. holmesii are not understood, and its phylogenetic position among the bordetellae remains unclear. To address these questions, B. holmesii strains were analyzed by comparative genomic hybridization (CGH) to a Bordetella pertussis microarray and by multilocus sequence typing. Both methods indicated substantial sequence divergence between B. pertussis and B. holmesii. However, CGH identified a putative pathogenicity island of 66 kb that is highly conserved between these species and contains several IS481 elements that may have been laterally transferred from B. pertussis to B. holmesii. This island contains, among other genes, a functional, iron-regulated locus encoding the biosynthesis, export, and uptake of the siderophore alcaligin. The acquisition of this genomic island by B. holmesii may have significantly contributed to its emergence as a human pathogen. Horizontal gene transfer between B. pertussis and B. holmesii may also explain the unusually high sequence identity of their 16S rRNA genes.

    View details for DOI 10.1128/JB.01081-06

    View details for Web of Science ID 000242798100008

    View details for PubMedID 17041054

  • Gene expression programs in adults with acute dengue infections Popper, S., Simmons, C. P., Dolecek, C., Chau, T. N., Griffiths, M., Dung, N. T., Long, T. H., Hoang, D. M., Van Vinh Chau, N., Thao, L. T., Hien, T. T., Relman, D. A., Farrar, J. AMER SOC TROP MED & HYGIENE. 2006: 290?90
  • Growth phase- and nutrient limitation-associated transcript abundance regulation in Bordetella pertussis INFECTION AND IMMUNITY Nakarnura, M. M., Liew, S., Cummings, C. A., Brinig, M. M., Dieterich, C., Relman, D. A. 2006; 74 (10): 5537-5548


    To survive in a host environment, microbial pathogens must sense local conditions, including nutrient availability, and adjust their growth state and virulence functions accordingly. No comprehensive investigation of growth phase-related gene regulation in Bordetella pertussis has been reported previously. We characterized changes in genome-wide transcript abundance of B. pertussis as a function of growth phase and availability of glutamate, a key nutrient for this organism. Using a Bordetella DNA microarray, we discovered significant changes in transcript abundance for 861 array elements during the transition from log phase to stationary phase, including declining transcript levels of many virulence factor genes. The responses to glutamate depletion exhibited similarities to the responses induced by exit from log phase, including decreased virulence factor transcript levels. However, only 23% of array elements that showed at least a fourfold growth phase-associated difference in transcript abundance also exhibited glutamate depletion-associated changes, suggesting that nutrient limitation may be one of several interacting factors affecting gene regulation during stationary phase. Transcript abundance patterns of a Bvg+ phase-locked mutant revealed that the BvgAS two-component regulatory system is a key determinant of growth phase- and nutrient limitation-related transcriptional control. Several adhesin genes exhibited lower transcript abundance during stationary phase and under glutamate restriction conditions. The predicted bacterial phenotype was confirmed: adherence to bronchoepithelial cells decreased 3.3- and 4.4-fold at stationary phase and with glutamate deprivation, respectively. Growth phase and nutrient availability may serve as cues by which B. pertussis regulates virulence according to the stage of infection or the location within the human airway.

    View details for DOI 10.1128/IAI.00781-06

    View details for Web of Science ID 000240967900013

    View details for PubMedID 16988229

  • A brave new world in the life sciences BULLETIN OF THE ATOMIC SCIENTISTS Choffnes, E. R., Lemon, S. M., Relman, D. A. 2006; 62 (5): 26-33
  • Assembly of the human intestinal microbiota TRENDS IN ECOLOGY & EVOLUTION Dethlefsen, L., Eckburg, P. B., Bik, E. M., Relman, D. A. 2006; 21 (9): 517-523


    Complex microbial ecosystems occupy the skin, mucosa and alimentary tract of all mammals, including humans. Recent advances have highlighted the tremendous diversity of these microbial communities and their importance to host physiology, but questions remain about the ecological processes that establish and maintain the microbiota throughout life. The prevailing view, that the gastrointestinal microbiota of adult humans is a climax community comprised of the superior competitors for a stable set of niches, does not account for all of the experimental data. We argue here that the unique history of each community and intrinsic temporal dynamics also influence the structure of human intestinal communities.

    View details for DOI 10.1016/j.tree.2006.06.013

    View details for Web of Science ID 000240633900011

    View details for PubMedID 16820245

  • Metagenomic analysis of the human distal gut microbiome SCIENCE Gill, S. R., Pop, M., DeBoy, R. T., Eckburg, P. B., Turnbaugh, P. J., Samuel, B. S., Gordon, J. I., Relman, D. A., Fraser-Liggett, C. M., Nelson, K. E. 2006; 312 (5778): 1355-1359


    The human intestinal microbiota is composed of 10(13) to 10(14) microorganisms whose collective genome ("microbiome") contains at least 100 times as many genes as our own genome. We analyzed approximately 78 million base pairs of unique DNA sequence and 2062 polymerase chain reaction-amplified 16S ribosomal DNA sequences obtained from the fecal DNAs of two healthy adults. Using metabolic function analyses of identified genes, we compared our human genome with the average content of previously sequenced microbial genomes. Our microbiome has significantly enriched metabolism of glycans, amino acids, and xenobiotics; methanogenesis; and 2-methyl-d-erythritol 4-phosphate pathway-mediated biosynthesis of vitamins and isoprenoids. Thus, humans are superorganisms whose metabolism represents an amalgamation of microbial and human attributes.

    View details for DOI 10.1126/science.1124234

    View details for Web of Science ID 000237961600048

    View details for PubMedID 16741115

  • Early days: genomics and human responses to infection CURRENT OPINION IN MICROBIOLOGY Liu, M., Popper, S. J., Rubins, K. H., Relman, D. A. 2006; 9 (3): 312-319


    DNA microarray-based gene transcript-profiling of the responses of primates to infection has begun to yield new insights into host-pathogen interactions; this approach, however, remains plagued by challenges and complexities that have yet to be adequately addressed. The rapidly changing nature over time of acute infectious diseases in a host, and the genetic diversity of microbial pathogens present unique problems for the design and interpretation of functional-genomic studies in this field. In addition, there are the more common problems related to heterogeneity within clinical samples, the complex, non-standardized confounding variables associated with human subjects and the complexities posed by the analysis and validation of highly parallel data. Whereas various approaches have been developed to address each of these issues, there are significant limitations that remain to be overcome. The resolution of these problems should lead to a better understanding of the dialogue between the host and pathogen.

    View details for DOI 10.1016/j.mib.2006.04.006

    View details for Web of Science ID 000238795100013

    View details for PubMedID 16679048

  • Significant gene order and expression differences in Bordetella pertussis despite limited gene content variation JOURNAL OF BACTERIOLOGY Brinig, M. M., Cummings, C. A., Sanden, G. N., Stefanelli, P., Lawrence, A., Relman, D. A. 2006; 188 (7): 2375-2382


    Bordetella pertussis, an obligate human pathogen and the agent of whooping cough, is a clonal species, despite the dynamic selection pressures imposed by host immunity and vaccine usage. Because the generation of variation is critical for species evolution, we employed a variety of approaches to examine features of B. pertussis genetic variation. We found a high level of conservation of gene content among 137 B. pertussis strains with different geographical, temporal, and epidemiological associations, using comparative genomic hybridization. The limited number of regions of difference were frequently located adjacent to copies of the insertion element IS481, which is present in high numbers in the B. pertussis chromosome. This repeated sequence appears to provide targets for homologous recombination, resulting in deletion of intervening sequences. Using subtractive hybridization, we searched for previously undetected genes in diverse clinical isolates but did not detect any new genes, indicating that gene acquisition is rare in B. pertussis. In contrast, we found evidence of altered gene order in the several strains that were examined and again found an association of IS481 with sites of rearrangement. Finally, we compared whole-genome expression profiles of different strains and found significant changes in transcript abundance, even in the same strain after as few as 12 laboratory passages. This combination of approaches provides a detailed picture of a pathogenic species with little gene loss or gain but with the capacity to generate variation by rearranging its chromosome and altering gene expression. These findings have broad implications for host adaptation by microbial pathogens.

    View details for DOI 10.1128/JB.188.7.2375-2382.2006

    View details for Web of Science ID 000236403300009

    View details for PubMedID 16547023

  • In search of biosecurity SCIENCE Relman, D. A., Choffnes, E., Lemon, S. M. 2006; 311 (5769): 1835-1835

    View details for DOI 10.1126/science.1127725

    View details for Web of Science ID 000236387800001

    View details for PubMedID 16574824

  • Species- and strain-specific control of a complex, flexible regulon by Bordetella BvgAS JOURNAL OF BACTERIOLOGY Cummings, C. A., Bootsma, H. J., Relman, D. A., Miller, J. F. 2006; 188 (5): 1775-1785


    The Bordetella master virulence regulatory system, BvgAS, controls a spectrum of gene expression states, including the virulent Bvg(+) phase, the avirulent Bvg(-) phase, and at least one Bvg-intermediate (Bvg(i)) phase. We set out to define the species- and strain-specific features of this regulon based on global gene expression profiling. Rather than functioning as a switch, Bvg controls a remarkable continuum of gene expression states, with hundreds of genes maximally expressed in intermediate phases between the Bvg(+) and Bvg(-) poles. Comparative analysis of Bvg regulation in B. pertussis and B. bronchiseptica revealed a relatively conserved Bvg(+) phase transcriptional program and identified previously uncharacterized candidate virulence factors. In contrast, control of Bvg(-)- and Bvg(i)-phase genes diverged substantially between species; regulation of metabolic, transporter, and motility loci indicated an increased capacity in B. bronchiseptica, compared to B. pertussis, for ex vivo adaptation. Strain comparisons also demonstrated variation in gene expression patterns within species. Among the genes with the greatest variability in patterns of expression, predicted promoter sequences were nearly identical. Our data suggest that the complement of transcriptional regulators is largely responsible for transcriptional diversity. In support of this hypothesis, many putative transcriptional regulators that were Bvg regulated in B. bronchiseptica were deleted, inactivated, or unregulated by BvgAS in B. pertussis. We propose the concept of a "flexible regulon." This flexible regulon may prove to be important for pathogen evolution and the diversification of host range specificity.

    View details for DOI 10.1128/JB.188.5.1775-1785.2006

    View details for Web of Science ID 000235819200011

    View details for PubMedID 16484188

    View details for PubMedCentralID PMC1426559

  • Molecular analysis of the bacterial microbiota in the human stomach PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Bik, E. M., Eckburg, P. B., Gill, S. R., Nelson, K. E., Purdom, E. A., Francois, F., Perez-Perez, G., Blaser, M. J., Relman, D. A. 2006; 103 (3): 732-737


    The microbiota of the human stomach and the influence of Helicobacter pylori colonization on its composition remain largely unknown. We characterized bacterial diversity within the human gastric mucosa by using a small subunit 16S rDNA clone library approach and analyzed 1,833 sequences generated by broad-range bacterial PCR from 23 gastric endoscopic biopsy samples. A diverse community of 128 phylotypes was identified, featuring diversity at this site greater than previously described. The majority of sequences were assigned to the Proteobacteria, Firmicutes, Actinobacteria, Bacteroidetes, and Fusobacteria phyla. Ten percent of the phylotypes were previously uncharacterized, including a Deinococcus-related organism, relatives of which have been found in extreme environments but not reported before in humans. The gastric clone libraries from 19 subjects contained H. pylori rDNA; however, only 12 of these subjects tested positive for H. pylori by conventional laboratory methods. Statistical analysis revealed a large degree of intersubject variability of the gastric ecosystem. The presence of H. pylori did not affect the composition of the gastric community. This gastric bacterial rDNA data set was significantly different from sequence collections of the human mouth and esophagus described in other studies, indicating that the human stomach may be home to a distinct microbial eco-system. The gastric microbiota may play important, as-yet-undiscovered roles in human health and disease.

    View details for DOI 10.1073/pnas.0506655103

    View details for Web of Science ID 000234727800042

    View details for PubMedID 16407106

  • Bioterrorism - Preparing to fight the next war NEW ENGLAND JOURNAL OF MEDICINE Relman, D. A. 2006; 354 (2): 113-115

    View details for Web of Science ID 000234528600002

    View details for PubMedID 16407505

  • Rapid quantitative profiling of complex microbial populations NUCLEIC ACIDS RESEARCH PALMER, C., Bik, E. M., Eisen, M. B., Eckburg, P. B., Sana, T. R., Wolber, P. K., Relman, D. A., Brown, P. O. 2006; 34 (1)


    Diverse and complex microbial ecosystems are found in virtually every environment on earth, yet we know very little about their composition and ecology. Comprehensive identification and quantification of the constituents of these microbial communities--a 'census'--is an essential foundation for understanding their biology. To address this problem, we developed, tested and optimized a DNA oligonucleotide microarray composed of 10,462 small subunit (SSU) ribosomal DNA (rDNA) probes (7167 unique sequences) selected to provide quantitative information on the taxonomic composition of diverse microbial populations. Using our optimized experimental approach, this microarray enabled detection and quantification of individual bacterial species present at fractional abundances of <0.1% in complex synthetic mixtures. The estimates of bacterial species abundance obtained using this microarray are similar to those obtained by phylogenetic analysis of SSU rDNA sequences from the same samples--the current 'gold standard' method for profiling microbial communities. Furthermore, probes designed to represent higher order taxonomic groups of bacterial species reliably detected microbes for which there were no species-specific probes. This simple, rapid microarray procedure can be used to explore and systematically characterize complex microbial communities, such as those found within the human body.

    View details for DOI 10.1093/nar/gnj007

    View details for Web of Science ID 000234782300005

    View details for PubMedID 16407321

  • Genomic features of Bordetella parapertussis clades with distinct host species specificity GENOME BIOLOGY Brinig, M. M., Register, K. B., Ackermann, M. R., Relman, D. A. 2006; 7 (9)


    The respiratory pathogen Bordetella parapertussis is a valuable model in which to study the complex phenotype of host specificity because of its unique two-species host range. One subset of strains, including the sequenced representative, causes whooping cough in humans, while other strains infect only sheep. The disease process in sheep is not well understood, nor are the genetic and transcriptional differences that might provide the basis for host specificity among ovine and human strains.We found 40 previously unknown genomic regions in an ovine strain of B. parapertussis using subtractive hybridization, including unique lipopolysaccharide genes. A microarray survey of the gene contents of 71 human and ovine strains revealed further differences, with 47 regions of difference distinguishing the host-restricted subgroups. In addition, sheep and human strains displayed distinct whole-genome transcript abundance profiles. We developed an animal model in which sheep were inoculated with a sheep strain, human strain, or mixture of the two. We found that the ovine strain persisted in the nasal cavity for 12 to 14 days, while the human strain colonized at lower levels and was no longer detected by 7 days post-inoculation. The ovine strain induced less granulocyte infiltration of the nasal mucosa.Several factors may play a role in determining host range of B. parapertussis. Human- and ovine-associated strains have differences in content and sequence of genes encoding proteins that mediate host-pathogen contact, such as lipopolysaccharide and fimbriae, as well as variation in regulation of toxins, type III secretion genes, and other virulence-associated genes.

    View details for DOI 10.1186/gb-2006-7-9-r81

    View details for Web of Science ID 000242490400007

    View details for PubMedID 16956413

  • Bordetella pertussis, the causative agent of whooping cough, evolved from a distinct, human-associated lineage of B-bronchiseptica PLOS PATHOGENS Diavatopoulos, D. A., Cummings, C. A., Schouls, L. M., Brinig, M. M., Relman, D. A., Mooi, F. R. 2005; 1 (4): 373-383


    Bordetella pertussis, B. bronchiseptica, B. parapertussis(hu), and B. parapertussis(ov) are closely related respiratory pathogens that infect mammalian species. B. pertussis and B. parapertussis(hu) are exclusively human pathogens and cause whooping cough, or pertussis, a disease that has resurged despite vaccination. Although it most often infects animals, infrequently B. bronchiseptica is isolated from humans, and these infections are thought to be zoonotic. B. pertussis and B. parapertussis(hu) are assumed to have evolved from a B. bronchiseptica-like ancestor independently. To determine the phylogenetic relationships among these species, housekeeping and virulence genes were sequenced, comparative genomic hybridizations were performed using DNA microarrays, and the distribution of insertion sequence elements was determined, using a collection of 132 strains. This multifaceted approach distinguished four complexes, representing B. pertussis, B. parapertussis(hu), and two distinct B. bronchiseptica subpopulations, designated complexes I and IV. Of the two B. bronchiseptica complexes, complex IV was more closely related to B. pertussis. Of interest, while only 32% of the complex I strains were isolated from humans, 80% of the complex IV strains were human isolates. Comparative genomic hybridization analysis identified the absence of the pertussis toxin locus and dermonecrotic toxin gene, as well as a polymorphic lipopolysaccharide biosynthesis locus, as associated with adaptation of complex IV strains to the human host. Lipopolysaccharide structural diversity among these strains was confirmed by gel electrophoresis. Thus, complex IV strains may comprise a human-associated lineage of B. bronchiseptica from which B. pertussis evolved. These findings will facilitate the study of pathogen host-adaptation. Our results shed light on the origins of the disease pertussis and suggest that the association of B. pertussis with humans may be more ancient than previously assumed.

    View details for DOI 10.1371/journal.ooat.0010045

    View details for Web of Science ID 000202894000009

    View details for PubMedID 16389302

  • Timing in collection of stool samples - Response SCIENCE Eckburg, P. B., Bik, E. M., Bernstein, C. N., Dethlefsen, L., Purdom, E., Sargent, M., Gills, S. R., Nelson, K. E., Relman, D. A. 2005; 310 (5751): 1118-1119
  • Acquired predisposition to mycobacterial disease due to autoantibodies to IFN-gamma JOURNAL OF CLINICAL INVESTIGATION Kampmann, B., Hemingway, C., Stephens, A., Davidson, R., Goodsall, A., Anderson, S., Nicol, M., Scholvinck, E., Relman, D., Waddell, S., Langford, P., Sheehan, B., Semple, L., Wilkinson, K. A., Wilkinson, R. J., Ress, S., Hibberd, M., Levin, M. 2005; 115 (9): 2480-2488


    Genetic defects in the IFN-gamma response pathway cause unique susceptibility to intracellular pathogens, particularly mycobacteria, but are rare and do not explain mycobacterial disease in the majority of affected patients. We postulated that acquired defects in macrophage activation by IFN-gamma may cause a similar immunological phenotype and thus explain the occurrence of disseminated intracellular infections in some patients without identifiable immune deficiency. Macrophage activation in response to IFN-gamma and IFN-gamma production were studied in whole blood and PBMCs of 3 patients with severe, unexplained nontuberculous mycobacterial infection. In all 3 patients, IFN-gamma was undetectable following mitogen stimulation of whole blood, but significant quantities were detectable in the supernatants of PBMCs when stimulated in the absence of the patients' own plasma. The patients' plasma inhibited the ability of IFN-gamma to increase production of TNF-alpha by both autologous and normal donor PBMCs, and recovery of exogenous IFN-gamma from the patients' plasma was greatly reduced. Using affinity chromatography, surface-enhanced laser desorption/ionization mass spectrometry, and sequencing, we isolated an IFN-gamma-neutralizing factor from the patients' plasma and showed it to be an autoantibody against IFN-gamma. The purified anti-IFN-gamma antibody was shown to be functional first in blocking the upregulation of TNF-alpha production in response to endotoxin; second in blocking induction of IFN-gamma-inducible genes (according to results of high-density cDNA microarrays); and third in inhibiting upregulation of HLA class II expression on PBMCs. Acquired defects in the IFN-gamma pathway may explain unusual susceptibility to intracellular pathogens in other patients without underlying, genetically determined immunological defects.

    View details for DOI 10.1172/JCI19316

    View details for Web of Science ID 000231708500025

    View details for PubMedID 16127458

  • Diversity of the human intestinal microbial flora SCIENCE Eckburg, P. B., Bik, E. M., Bernstein, C. N., Purdom, E., Dethlefsen, L., Sargent, M., Gill, S. R., Nelson, K. E., Relman, D. A. 2005; 308 (5728): 1635-1638


    The human endogenous intestinal microflora is an essential "organ" in providing nourishment, regulating epithelial development, and instructing innate immunity; yet, surprisingly, basic features remain poorly described. We examined 13,355 prokaryotic ribosomal RNA gene sequences from multiple colonic mucosal sites and feces of healthy subjects to improve our understanding of gut microbial diversity. A majority of the bacterial sequences corresponded to uncultivated species and novel microorganisms. We discovered significant intersubject variability and differences between stool and mucosa community composition. Characterization of this immensely diverse ecosystem is the first step in elucidating its role in health and disease.

    View details for DOI 10.1126/science.1110591

    View details for Web of Science ID 000229827000059

    View details for PubMedID 15831718

  • Genomewide analysis of the host response to malaria in Kenyan children JOURNAL OF INFECTIOUS DISEASES Griffiths, M. J., Shafi, M. J., Popper, S. J., Hemingway, C. A., Kortok, M. M., Wathen, A., Rockett, K. A., Mott, R., Levin, M., Newton, C. R., Marsh, K., Relman, D. A., Kwiatkowski, D. P. 2005; 191 (10): 1599-1611


    Malaria is a global problem, and there is a critical need for further understanding of the disease process. When malarial parasites invade and develop within the bloodstream, they stimulate a profound host response whose main clinical sign is fever. To explore this response, we measured host gene expression in whole blood from Kenyan children hospitalized with either acute malaria or other febrile illnesses. Genomewide analysis of expression identified 2 principal gene-expression profiles related to neutrophil and erythroid activity. In addition to these general acute responses, a third gene-expression profile was associated with host parasitemia; mediators of erythrophagocytosis and cellular stress were notable components of this response. The delineation of subjects on the basis of patterns of gene expression provides a molecular perspective of the host response to malaria and further functional insight into the underlying processes of pathogenesis.

    View details for Web of Science ID 000228465000005

    View details for PubMedID 15838786

  • VCAM-1 expression on CD8(+) cells correlates with enhanced anti-HIV suppressing activity JOURNAL OF IMMUNOLOGY DIAZ, L. S., Foster, H., Stone, M. R., Fujimura, S., Relman, D. A., Levy, J. A. 2005; 174 (3): 1574-1579


    CD8(+) cells from HIV-infected individuals showing the CD8(+) cell noncytotoxic antiviral response unexpectedly revealed mRNA for VCAM-1, a cell surface molecule found on endothelial cells. Uninfected subjects had undetectable levels of VCAM-1 mRNA in their CD8(+) cells. Flow cytometry analysis showed that up to 12% of the CD8(+) cells from HIV-positive individuals expressed VCAM-1 compared with 0.8% of the CD8(+) cells of HIV-negative individuals. Enrichment of the CD8(+)VCAM-1(+) cell population and subsequent coculture with CD4(+) cells acutely infected with HIV-1 showed that the VCAM-1(+)CD8(+) cells were able to suppress viral replication with 50% less input cells than the unseparated CD8(+) cell population. This study demonstrates, for the first time, the expression of VCAM-1 on CD8(+) cells. Moreover, the CD8(+)VCAM-1(+) cells show enhanced CD8(+) cell noncytotoxic antiviral response activity that could have clinical importance in HIV infection.

    View details for Web of Science ID 000226571300054

    View details for PubMedID 15661918

  • Exploring the potential of variola virus infection of cynomolgus macaques as a model for human smallpox PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Jahrling, P. B., Hensley, L. E., Martinez, M. J., LEDUC, J. W., Rubins, K. H., Relman, D. A., HUGGINS, J. W. 2004; 101 (42): 15196-15200


    Smallpox virus (variola) poses a significant threat as an agent of bioterrorism. To mitigate this risk, antiviral drugs and an improved vaccine are urgently needed. Satisfactory demonstration of protective efficacy against authentic variola will require development of an animal model in which variola produces a disease course with features consistent with human smallpox. Toward this end, cynomolgus macaques were exposed to several variola strains through aerosol and/or i.v. routes. Two strains, Harper and India 7124, produced uniform acute lethality when inoculated i.v. in high doses (10(9) plaque-forming units). Lower doses resulted in less fulminant, systemic disease and lower mortality. Animals that died had profound leukocytosis, thrombocytopenia, and elevated serum creatinine levels. After inoculation, variola was disseminated by means of a monocytic cell-associated viremia. Distribution of viral antigens by immunohistochemistry correlated with the presence of replicating viral particles demonstrated by electron microscopy and pathology in the lymphoid tissues, skin, oral mucosa, gastrointestinal tract, reproductive system, and liver. These particles resembled those seen in human smallpox. High viral burdens in target tissues were associated with organ dysfunction and multisystem failure. Evidence of coagulation cascade activation (D dimers) corroborated histologic evidence of hemorrhagic diathesis. Depletion of T cell-dependent areas of lymphoid tissues occurred, probably as a consequence of bystander apoptotic mechanisms initiated by infected macrophages. Elaboration of cytokines, including IL-6 and IFN-gamma, contribute to a cytokine storm formerly known as "toxemia." A more precise understanding of disease pathogenesis should provide targets for therapeutic intervention, to be used alone or in combination with inhibitors of variola virus replication.

    View details for DOI 10.1073/pnas.0405954101

    View details for Web of Science ID 000224688700040

    View details for PubMedID 15477589

  • The host response to smallpox: Analysis of the gene expression program in peripheral blood cells in a nonhuman primate model PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Rubins, K. H., Hensley, L. E., JAHRLING, P. B., Whitney, A. R., Geisbert, T. W., HUGGINS, J. W., Owen, A., LEDUC, J. W., Brown, P. O., Relman, D. A. 2004; 101 (42): 15190-15195


    Smallpox has played an unparalleled role in human history and remains a significant potential threat to public health. Despite the historical significance of this disease, we know little about the underlying pathophysiology or the virulence mechanisms of the causative agent, variola virus. To improve our understanding of variola pathogenesis and variola-host interactions, we examined the molecular and cellular features of hemorrhagic smallpox in cynomolgus macaques. We used cDNA microarrays to analyze host gene expression patterns in sequential blood samples from each of 22 infected animals. Variola infection elicited striking and temporally coordinated patterns of gene expression in peripheral blood. Of particular interest were features that appear to represent an IFN response, cell proliferation, immunoglobulin gene expression, viral dose-dependent gene expression patterns, and viral modulation of the host immune response. The virtual absence of a tumor necrosis factor alpha/NF-kappaB-activated transcriptional program in the face of an overwhelming systemic infection suggests that variola gene products may ablate this response. These results provide a detailed picture of the host transcriptional response during smallpox infection, and may help guide the development of diagnostic, therapeutic, and prophylactic strategies.

    View details for DOI 10.1073/pnas.0405759101

    View details for Web of Science ID 000224688700039

    View details for PubMedID 15477590

  • Methanogenic Archaea and human periodontal disease PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Lepp, P. W., Brinig, M. M., Ouverney, C. C., Palm, K., Armitage, G. C., Relman, D. A. 2004; 101 (16): 6176-6181


    Archaea have been isolated from the human colon, vagina, and oral cavity, but have not been established as causes of human disease. In this study, we reveal a relationship between the severity of periodontal disease and the relative abundance of archaeal small subunit ribosomal RNA genes (SSU rDNA) in the subgingival crevice by using quantitative PCR. Furthermore, the relative abundance of archaeal small subunit rDNA decreased at treated sites in association with clinical improvement. Archaea were harbored by 36% of periodontitis patients and were restricted to subgingival sites with periodontal disease. The presence of archaeal cells at these sites was confirmed by fluorescent in situ hybridization. The archaeal community at diseased sites was dominated by a Methanobrevibacter oralis-like phylotype and a distinct Methanobrevibacter subpopulation related to archaea that inhabit the gut of numerous animals. We hypothesize that methanogens participate in syntrophic relationships in the subgingival crevice that promote colonization by secondary fermenters during periodontitis. Because they are potential alternative syntrophic partners, our finding of larger Treponema populations sites without archaea provides further support for this hypothesis.

    View details for DOI 10.1073/pnas.0308766101

    View details for Web of Science ID 000220978000089

    View details for PubMedID 15067114

  • Bordetella species are distinguished by patterns of substantial gene loss and host adaptation JOURNAL OF BACTERIOLOGY Cummings, C. A., Brinig, M. M., Lepp, P. W., van de Pas, S., Relman, D. A. 2004; 186 (5): 1484-1492


    Pathogens of the bacterial genus Bordetella cause respiratory disease in humans and animals. Although virulence and host specificity vary across the genus, the genetic determinants of this diversity remain unidentified. To identify genes that may underlie key phenotypic differences between these species and clarify their evolutionary relationships, we performed a comparative analysis of genome content in 42 Bordetella strains by hybridization of genomic DNA to a microarray representing the genomes of three Bordetella species and by subtractive hybridization. Here we show that B. pertussis and B. parapertussis are predominantly differentiated from B. bronchiseptica by large, species-specific regions of difference, many of which encode or direct synthesis of surface structures, including lipopolysaccharide O antigen, which may be important determinants of host specificity. The species also exhibit sequence diversity at a number of surface protein-encoding loci, including the fimbrial major subunit gene, fim2. Gene loss, rather than gene acquisition, accompanied by the proliferation of transposons, has played a fundamental role in the evolution of the pathogenic bordetellae and may represent a conserved evolutionary mechanism among other groups of microbial pathogens.

    View details for Web of Science ID 000189117900030

    View details for PubMedID 14973121

  • Bordetella pertussis infection of primary human monocytes alters HLA-DR expression INFECTION AND IMMUNITY Shumilla, J. A., Lacaille, V., Hornell, T. M., Huang, J., Narasimhan, S., Relman, D. A., Mellins, E. D. 2004; 72 (3): 1450-1462


    Bordetella pertussis is the causative agent of whooping cough, a potentially lethal respiratory disease in children. In immunocompetent individuals, B. pertussis infection elicits an effective adaptive immune response driven by activated CD4(+) T cells. However, live B. pertussis persists in the host for 3 to 4 weeks prior to clearance. Thus, B. pertussis appears to have evolved short-term mechanisms for immune system evasion. We investigated the effects of B. pertussis wild-type strain BP338 on antigen presentation in primary human monocytes. BP338 infection reduced cell surface expression of HLA-DR and CD86 but not that of major histocompatibility complex class I proteins. This change in cell surface HLA-DR expression reflected intracellular redistribution of HLA-DR. The proportion of peptide-loaded molecules was unchanged in infected cells, suggesting that intracellular retention occurred after peptide loading. Although B. pertussis infection of monocytes induced rapid and robust expression of interleukin-10 (IL-10), HLA-DR redistribution did not appear to be explained by increased IL-10 levels. BP338-infected monocytes exhibited reduced synthesis of HLA-DR dimers. Interestingly, those HLA-DR proteins that were generated appeared to be longer-lived than HLA-DR in uninfected monocytes. BP338 infection also prevented gamma interferon (IFN-gamma) induction of HLA-DR protein synthesis. Using mutant strains of B. pertussis, we found that reduction in HLA-DR surface expression was due in part to the presence of pertussis toxin whereas the inhibition of IFN-gamma induction of HLA-DR could not be linked to any of the virulence factors tested. These data demonstrate that B. pertussis utilizes several mechanisms to modulate HLA-DR expression.

    View details for DOI 10.1128/IAI.72.3.145-1462.2004

    View details for Web of Science ID 000189270800029

    View details for PubMedID 14977950

  • Role of interleukin 6 in myocardial dysfunction of meningococcal septic shock LANCET Pathan, N., Hemingway, C. A., Alizadeh, A. A., Stephens, A. C., Boldrick, J. C., Oragui, E. E., McCabe, C., Welch, S. B., Whitney, A., O'Gara, P., Nadel, S., Relman, D. A., Harding, S. E., Levin, M. 2004; 363 (9404): 203-209


    Myocardial failure has a central role in the complex pathophysiology of septic shock and contributes to organ failure and death. During the sepsis-induced inflammatory process, specific factors are released that depress myocardial contractile function. We aimed to identify these mediators of myocardial depression in meningococcal septic shock.We combined gene-expression profiling with protein and cellular methods to identify a serum factor causing cardiac dysfunction in meningococcal septic shock. We identified genes that were significantly upregulated in blood after exposure to meningococci. We then selected for further analysis those genes whose protein products had properties of a myocardial depressant factor--specifically a 12-25 kDa heat-stable protein that is released into serum shortly after onset of meningococcal infection.We identified 174 significantly upregulated genes in meningococcus-infected blood: six encoded proteins that were of the predicted size and had characteristics of a myocardial depressant factor. Of these, interleukin 6 caused significant myocardial depression in vitro. Removal of interleukin 6 from serum samples of patients with meningococcaemia and from supernatants of inflammatory cells stimulated by meningococci in vitro abolished the negative inotropic activity. Furthermore, concentrations in serum of interleukin 6 strongly predicted degree of myocardial dysfunction and severity of disease in children with meningococcal septic shock.Interleukin 6 is a mediator of myocardial depression in meningococcal disease. This cytokine and its downstream mediators could be a target for future treatment strategies.

    View details for Web of Science ID 000188243900010

    View details for PubMedID 14738793

  • Public health - Understanding threats to scientific openness SCIENCE Petro, J. B., Relman, D. A. 2003; 302 (5652): 1898-1898

    View details for Web of Science ID 000187174000034

    View details for PubMedID 14671279

  • Shedding light on microbial detection NEW ENGLAND JOURNAL OF MEDICINE Relman, D. A. 2003; 349 (22): 2162-2163

    View details for Web of Science ID 000186779700014

    View details for PubMedID 14645646

  • Single-cell enumeration of an uncultivated TM7 subgroup in the human subgingival crevice APPLIED AND ENVIRONMENTAL MICROBIOLOGY Ouverney, C. C., Armitage, G. C., Relman, D. A. 2003; 69 (10): 6294-6298


    Specific oligonucleotide hybridization conditions were established for single-cell enumeration of uncultivated TM7 and IO25 bacteria by using clones expressing heterologous 16S rRNA. In situ analysis of human subgingival crevice specimens revealed that a greater proportion of samples from sites of chronic periodontitis than from healthy sites contained TM7 subgroup IO25. In addition, IO25 bacterial cells from periodontitis site samples were more abundant and fourfold longer than IO25 cells from healthy site samples.

    View details for DOI 10.1128/AEM.69.10.6294-6298.2003

    View details for Web of Science ID 000185881300072

    View details for PubMedID 14532094

  • Cultivation of Tropheryma whipplei from cerebrospinal fluid JOURNAL OF INFECTIOUS DISEASES Maiwald, M., Von Herbay, A., Fredricks, D. N., Ouverney, C. C., Kosek, J. C., Relman, D. A. 2003; 188 (6): 801-808


    Whipple disease (WD) is a systemic disorder caused by the bacterium Tropheryma whipplei. Since the recognition of a bacterial etiology in 1961, many attempts have been made to cultivate this bacterium in vitro. It was eventually isolated, in 2000, from an infected heart valve, in coculture with human fibroblasts. Here we report the isolation of 2 new strains of T. whipplei from cerebrospinal fluid (CSF) of 2 patients with intestinal WD but no neurological signs or symptoms. One culture-positive specimen was obtained before treatment; the other was obtained 12 months after discontinuation of therapy, at a time of intestinal remission. In both cases, 15 passages of the cultures were completed over 17 months. Bacterial growth was measured by quantitative polymerase chain reaction, which suggested a generation time of 4 days. Staining with YO-PRO nucleic-acid dye showed characteristic rod-shaped bacteria arranged in chains. Fluorescent in situ hybridization with a T. whipplei-specific oligonucleotide probe, a broad-range bacterial probe, and a nonspecific nucleic-acid stain indicated that all visible bacteria were T. whipplei. Scanning electron microscopy and transmission electron microscopy showed both intracellular and extracellular bacteria. This first isolation of T. whipplei from CSF provides clear evidence of viable bacteria in the central nervous system in individuals with WD, even after prolonged antibiotic therapy.

    View details for Web of Science ID 000185215600002

    View details for PubMedID 12964110

  • Lethal invasive cestodiasis in immunosuppressed patients 74th Annual Meeting of the American-Society-of-Parasitologists Olson, P. D., Yoder, K., Fajardo, L. F., Marty, A. M., van de Pas, S., Olivier, C., Relman, D. A. UNIV CHICAGO PRESS. 2003: 1962?66


    Using both traditional methods and broad-range 18S ribosomal DNA (rDNA) polymerase chain reaction, we examined 2 cases of lethal cestodiasis, in which the disease agent had been poorly identified or misidentified. In one case, involving a patient with AIDS, we identified the human dwarf tapeworm, Hymenolepis nana, as a cause of aberrant metastatic larval disease. In the second case with similar pathologic abnormalities, involving a patient with Hodgkin disease, we identified a larval cestode with a previously uncharacterized 18S rDNA sequence. A prior report of this case nearly 30 years ago, based on tissue examination, had suggested that the parasite was a sparganum.

    View details for Web of Science ID 000183722600017

    View details for PubMedID 12792874

  • Smallpox vaccination - Reply NEW ENGLAND JOURNAL OF MEDICINE Rubins, K., Relman, D. A. 2003; 348 (19): 1925-1925
  • Analysis of conserved non-rRNA genes of Tropheryma whipplei SYSTEMATIC AND APPLIED MICROBIOLOGY Maiwald, M., Lepp, P. W., Relman, D. A. 2003; 26 (1): 3-12


    The causative agent of Whipple's disease, Tropheryma whipplei, is a slow-growing bacterium that remains poorly-understood. Genetic characterization of this organism has relied heavily upon rRNA sequence analysis. Pending completion of a complete genome sequencing effort, we have characterized several conserved non-rRNA genes from T. whipplei directly from infected tissue using broad-range PCR and a genome-walking strategy. Our goals were to evaluate its phylogenetic relationships, and to find ways to expand the strain typing scheme, based on rDNA sequence comparisons. The genes coding for the ATP synthase beta subunit (atpD), elongation factor Tu (tuf), heat shock protein GroEL (groEL), beta subunit of DNA-dependent RNA polymerase (rpoB), and RNase P RNA (rnpB) were analyzed, as well as the regions upstream and downstream of the rRNA operon. Phylogenetic analyses with all non-rRNA marker molecules consistently placed T. whipplei within the class, Actinobacteria. The arrangement of genes in the atpD and rpoB chromosomal regions was also consistent with other actinomycete genomes. Tandem sequence repeats were found upstream and downstream of the rRNA operon, and downstream of the groEL gene. These chromosomal sites and the 16S-23S rRNA intergenic spacer regions were examined in the specimens of 11 patients, and a unique combination of tandem repeat numbers and spacer polymorphisms was found in each patient. These data provide the basis for a more discriminatory typing method for T. whipplei.

    View details for Web of Science ID 000182851700001

    View details for PubMedID 12747404

  • Prevalence of bacteria of division TM7 in human subgingival plaque and their association with disease APPLIED AND ENVIRONMENTAL MICROBIOLOGY Brinig, M. M., Lepp, P. W., Ouverney, C. C., Armitage, G. C., Relman, D. A. 2003; 69 (3): 1687-1694


    Members of the uncultivated bacterial division TM7 have been detected in the human mouth, but little information is available regarding their prevalence and diversity at this site. Human subgingival plaque samples from healthy sites and sites exhibiting various stages of periodontal disease were analyzed for the presence of TM7 bacteria. TM7 ribosomal DNA (rDNA) was found in 96% of the samples, and it accounted for approximately 0.3%, on average, of all bacterial rDNA in the samples as determined by real-time quantitative PCR. Two new phylotypes of this division were identified, and members of the division were found to exhibit filamentous morphology by fluorescence in situ hybridization. The abundance of TM7 rDNA relative to total bacterial rDNA was higher in sites with mild periodontitis (0.54% +/- 0.1%) than in either healthy sites (0.21% +/- 0.05%, P < 0.01) or sites with severe periodontitis (0.29% +/- 0.06%, P < 0.05). One division subgroup, the I025 phylotype, was detected in 1 of 18 healthy samples and 38 of 58 disease samples. These data suggest that this phylotype, and the TM7 bacterial division in general, may play a role in the multifactorial process leading to periodontitis.

    View details for DOI 10.1128/AEM.69.3.1687-1694.2003

    View details for Web of Science ID 000181435600047

    View details for PubMedID 12620860

  • Engineered antibodies to treat whooping cough. 225th National Meeting of the American-Chemical-Society Maynard, J., Relman, D., Iverson, B., Merkel, T., Georgiou, G. AMER CHEMICAL SOC. 2003: U196?U196
  • Sequencing and analysis of the genome of the Whipple's disease bacterium Tropheryma whipplei LANCET Bentley, S. D., Maiwald, M., Murphy, L. D., Pallen, M. J., Yeats, C. A., Dover, L. G., Norbertczak, H. T., Besra, G. S., Quail, M. A., Harris, D. E., Von Herbay, A., Goble, A., Rutter, S., Squares, R., Squares, S., Barrell, B. G., Parkhill, J., Relman, D. A. 2003; 361 (9358): 637-644


    Whipple's disease is a rare multisystem chronic infection, involving the intestinal tract as well as various other organs. The causative agent, Tropheryma whipplei, is a Gram-positive bacterium about which little is known. Our aim was to investigate the biology of this organism by generating and analysing the complete DNA sequence of its genome.We isolated and propagated T whipplei strain TW08/27 from the cerebrospinal fluid of a patient diagnosed with Whipple's disease. We generated the complete sequence of the genome by the whole genome shotgun method, and analysed it with a combination of automatic and manual bioinformatic techniques.Sequencing revealed a condensed 925938 bp genome with a lack of key biosynthetic pathways and a reduced capacity for energy metabolism. A family of large surface proteins was identified, some associated with large amounts of non-coding repetitive DNA, and an unexpected degree of sequence variation.The genome reduction and lack of metabolic capabilities point to a host-restricted lifestyle for the organism. The sequence variation indicates both known and novel mechanisms for the elaboration and variation of surface structures, and suggests that immune evasion and host interaction play an important part in the lifestyle of this persistent bacterial pathogen.

    View details for Web of Science ID 000181129500012

    View details for PubMedID 12606174

  • Individuality and variation in gene expression patterns in human blood PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Whitney, A. R., Diehn, M., Popper, S. J., Alizadeh, A. A., Boldrick, J. C., Relman, D. A., Brown, P. O. 2003; 100 (4): 1896-1901


    The nature and extent of interindividual and temporal variation in gene expression patterns in specific cells and tissues is an important and relatively unexplored issue in human biology. We surveyed variation in gene expression patterns in peripheral blood from 75 healthy volunteers by using cDNA microarrays. Characterization of the variation in gene expression in healthy tissue is an essential foundation for the recognition and interpretation of the changes in these patterns associated with infections and other diseases, and peripheral blood was selected because it is a uniquely accessible tissue in which to examine this variation in patients or healthy volunteers in a clinical setting. Specific features of interindividual variation in gene expression patterns in peripheral blood could be traced to variation in the relative proportions of specific blood cell subsets; other features were correlated with gender, age, and the time of day at which the sample was taken. An analysis of multiple sequential samples from the same individuals allowed us to discern donor-specific patterns of gene expression. These data help to define human individuality and provide a database with which disease-associated gene expression patterns can be compared.

    View details for DOI 10.1073/pnas.252784499

    View details for Web of Science ID 000181073000082

    View details for PubMedID 12578971

    View details for PubMedCentralID PMC149930

  • The complexity of the inflammatory response to meningococcal sepsis revealed by gene expression profiling using cDNA microarrays 32nd Critical Care Congress of the Society-of-Critical-Care-Medicine Pathan, N., Hemingway, C., Levin, M., Whitney, A., Popper, S., Relman, D. LIPPINCOTT WILLIAMS & WILKINS. 2003: A47?A47
  • Archaea and their potential role in human disease INFECTION AND IMMUNITY Eckburg, P. B., Lepp, P. W., Relman, D. A. 2003; 71 (2): 591-596
  • Progression of the lesion at the site of inoculation after smallpox vaccination NEW ENGLAND JOURNAL OF MEDICINE Rubins, K., Relman, D. A. 2003; 348 (5): 414-414
  • Images in clinical medicine. Progression of the lesion at the site of inoculation after smallpox vaccination. New England journal of medicine Rubins, K., Relman, D. A. 2003; 348 (5): 414-?

    View details for PubMedID 12556544

  • Individuality and variation in gene expression patterns in human blood. Proc Natl Acad Sci (USA) Relman DA, Whitney AR, Diehn M, Popper SJ, Alizadeh AA, Boldrick JC, Brown PO. 2003: 1896-1901
  • Role of phosphatidylinositol 3-kinase in the binding of Bordetella pertussis to human monocytes CELLULAR MICROBIOLOGY Ishibashi, Y., Yoshimura, K., Nishikawa, A., Claus, S., Laudanna, C., Relman, D. A. 2002; 4 (12): 825-833


    Bordetella pertussis, the causative agent of whooping cough, adheres to human monocytes by means of filamentous haemagglutinin (FHA), a bacterial surface protein that is recognized by complement receptor type 3 (CR3, alphaMbeta2 integrin). Previous work has shown that an FHA Arg-Gly-Asp (RGD, residues 1097-1099) site interacts with a complex composed of leucocyte response integrin (LRI, alphavbeta3 integrin) and integrin-associated protein (IAP, CD47) on human monocytes, resulting in enhancement of CR3-mediated bacterial binding. However, the pathway that mediates alphavbeta3-alphaMbeta2 integrin signalling remains to be characterized. Here we describe the involvement of phosphatidylinositol 3-kinase (PI3-K) in this pathway. Wortmannin and LY294002, inhibitors of PI3-K, reduced alphavbeta3/IAP-upregulated, CR3-associated bacterial binding to human monocytes. B. pertussis infection of human monocytes resulted in a marked recruitment of cellular PI3-K to the sites of B. pertussis contact. In contrast, cells infected with an isogenic strain carrying a G1098A mutation at the FHA RGD site did not show any recruitment of PI3-K. We found that ligation of FHA by alphavbeta3/IAP induced RGD-dependent tyrosine phosphorylation of a 60 kDa protein, which associated with IAP and PI3-K in human monocytes. These results suggest that PI3-K and a tyrosine phosphorylated 60 kDa protein may be involved in this biologically important integrin signalling pathway.

    View details for Web of Science ID 000179648500005

    View details for PubMedID 12464013

  • New technologies, human-microbe interactions, and the search for previously unrecognized pathogens 3rd Pfizer Symposium on Immunopathogenesis of Infections and Inflammatory Diseases of the Brain Relman, D. A. OXFORD UNIV PRESS INC. 2002: S254?S258


    Evidence suggests that a significant number of clinically important microbial pathogens remain unrecognized. Observations from the natural world, from patterns of disease in human populations, from the bedside, and from the clinical laboratory all contribute to this body of evidence. A variety of acute and chronic neurologic syndromes illustrate this point; despite features of infection, most cases of aseptic meningitis, encephalitis, and cerebral vasculitis cannot be assigned a microbiologic diagnosis. The development and clinical application of molecular methods have led to the discovery of novel members of the endogenous normal flora as well as putative disease agents. Current challenges include the establishment of criteria for disease causation and further characterization of the human microbiome during states of health. These challenges and the goal of understanding microbial contributions to inflammatory disease may be addressed effectively through the thoughtful integration of modern technologies and clinical insight.

    View details for Web of Science ID 000179587500019

    View details for PubMedID 12424706

  • Molecular identification of cyanobacteria associated with stromatolites from distinct geographical locations Conference on Astrobiology Neilan, B. A., Burns, B. P., Relman, D. A., Lowe, D. R. MARY ANN LIEBERT INC. 2002: 271?80


    Modern stromatolites represent a significant resource for studying microbial ecology and evolution. A preliminary investigation was undertaken employing specific genetic probes to characterize the cyanobacteria responsible for stromatolite construction in a range of environments, including microbial mats found in Australia not previously examined with molecular methods. Isolates of cyanobacteria were collected from stromatolites in thermal springs, hypersaline lakes, and oceanic fringes on two continents. A polymerase chain reaction specific for DNA of cyanobacterial 16S rRNA was developed, the resulting products of the DNA amplification reaction were sequenced, and the data were used to infer relatedness between the isolates studied and other members of the cyanobacterial radiation. Complete sequence was generated for the region from position 27 to 408 for 13 strains of cyanobacteria associated with stromatolites. All stromatolite-derived sequences were most closely related to cyanobacteria, as indicated by local sequence alignment. It was possible to correlate genetic identity with morphological nomenclatures and to expand the phylogeny of benthic cyanobacteria. These inferences were also expanded to temporal variation in the dominant resident cyanobacterial species based on sampling of surface and core sinter laminations. Under the methods employed, only one cyanobacterial strain was detected in each sample, suggesting the possible dominance of a specific clonal population of cyanobacteria at any one time in the biota of the samples tested. The data indicate that internal core samples of a stromatolite at least 10 years old can be successfully analyzed by DNA-based methods to identify preserved cyanobacteria.

    View details for Web of Science ID 000182464000005

    View details for PubMedID 12530237

  • Genome-wide responses of a pathogenic bacterium to its host JOURNAL OF CLINICAL INVESTIGATION Relman, D. A. 2002; 110 (8): 1071-1073

    View details for DOI 10.1172/JCI200216944

    View details for Web of Science ID 000178793700004

    View details for PubMedID 12393841

  • Mining the natural world for new pathogens AMERICAN JOURNAL OF TROPICAL MEDICINE AND HYGIENE Relman, D. A. 2002; 67 (2): 133-134

    View details for Web of Science ID 000178479700001

    View details for PubMedID 12389934

  • Smallpox research activities: US Interagency collaboration, 2001 EMERGING INFECTIOUS DISEASES LEDUC, J. W., Damon, I., Meegan, J. M., Relman, D. A., Huggins, J., JAHRLING, P. B. 2002; 8 (7): 743-745

    View details for Web of Science ID 000176394200022

    View details for PubMedID 12095449

  • Genomics and microbiology - Microbial forensics - "Cross-examining pathogens" SCIENCE Cummings, C. A., Relman, D. A. 2002; 296 (5575): 1976-?

    View details for DOI 10.1126/science.1073125

    View details for Web of Science ID 000176273300036

    View details for PubMedID 12004075

  • Sequence variability in the first internal transcribed spacer region within and among Cyclospora species is consistent with polyparasitism (vol 31, 1475, 2001) INTERNATIONAL JOURNAL FOR PARASITOLOGY Olivier, C., van de Pas, S., Lepp, P. W., Yoder, K., Relman, D. A. 2002; 32 (4): 489-489
  • The periodontal-systemic connection: Molecular mechanisms of host pathogen interactions in cardovascular disease and spontaneous pre term birth. Genco, C., Kinane, D. F., Ozinsky, A., Offenbacher, S., Relman, D. SAGE PUBLICATIONS INC. 2002: A45?A45
  • Identification of Cardiobacterium hominis by broad-range bacterial polymerase chain reaction analysis in a case of culture-negative endocarditis ARCHIVES OF INTERNAL MEDICINE Nikkari, S., Gotoff, R., Bourbeau, P. P., BROWN, R. E., Kamal, N. R., Relman, D. A. 2002; 162 (4): 477-479


    Culture-negative bacterial endocarditis may be attributed to fastidious microorganisms, prior institution of antibiotic treatment, or both. We describe a case of culture-negative endocarditis in which a modified Steiner stain revealed bacterial structures in the resected heart valve material. Prompted by this finding, broad-range polymerase chain reaction (PCR) amplification of small-subunit ribosomal DNA (16S rDNA) was performed, and Cardiobacterium hominis sequences were detected. This case demonstrates the usefulness of both the Steiner stain and broad-range direct molecular amplification as supplemental diagnostic tools in identification of otherwise unexplained infections.

    View details for Web of Science ID 000173854800017

    View details for PubMedID 11863484

  • Human herpesvirus 8 and sarcoidosis CLINICAL INFECTIOUS DISEASES Fredricks, D. N., Martin, T. M., Edwards, A. O., ROSENBAUM, J. T., Relman, D. A. 2002; 34 (4): 559-560

    View details for Web of Science ID 000173458700026

    View details for PubMedID 11797191

  • The human body as microbial observatory NATURE GENETICS Relman, D. A. 2002; 30 (2): 131-133

    View details for Web of Science ID 000173708700005

    View details for PubMedID 11818955

  • Broad-range bacterial detection and the analysis of unexplained death and critical illness EMERGING INFECTIOUS DISEASES Nikkari, S., Lopez, F. A., Lepp, P. W., Cieslak, P. R., Ladd-Wilson, S., Passaro, D., Danila, R., Relman, D. A. 2002; 8 (2): 188-194


    Broad-range rDNA polymerase chain reaction (PCR) provides an alternative, cultivation-independent approach for identifying pathogens. In 1995, the Centers for Disease Control and Prevention initiated population-based surveillance for unexplained life-threatening infections (Unexplained Death and Critical Illness Project [UNEX]). To address the causes of UNEX cases, we examined 59 specimens from 46 cases by using broad-range bacterial 16S rDNA PCR and phylogenetic analysis of amplified sequences. Specimens from eight cases yielded sequences from Neisseria meningitidis (cerebrospinal fluid from two patients with meningitis), Streptococcus pneumoniae (cerebrospinal fluid from one patient with meningitis2 and pleural fluid from two patients with pneumonia), or Stenotrophomonas maltophilia (bone marrow aspirate from one patient with pneumonia). Streptococcus pneumoniae rDNA sequence microheterogeneity was found in one pleural fluid specimen, suggesting the presence of multiple strains. In conclusion, known bacterial pathogens cause some critical illnesses and deaths that fail to be explained with traditional diagnostic methods.

    View details for Web of Science ID 000173757800013

    View details for PubMedID 11897072

  • Surveillance for unexplained deaths and critical illnesses due to possibly infectious causes, United States, 1995-1998 EMERGING INFECTIOUS DISEASES Hajjeh, R. A., Relman, D., Cieslak, P. R., Sofair, A. N., Passaro, D., Flood, J., Johnson, J., Hacker, J. K., Shieh, W. J., Hendry, R. M., Nikkari, S., Ladd-Wilson, S., Hadler, J., Rainbow, J., Tappero, J. W., Woods, C. W., Conn, L., Reagan, S., Zaki, S., Perkins, B. A. 2002; 8 (2): 145-?


    Population-based surveillance for unexplained death and critical illness possibly due to infectious causes (UNEX) was conducted in four U.S. Emerging Infections Program sites (population 7.7 million) from May 1, 1995, to December 31, 1998, to define the incidence, epidemiologic features, and etiology of this syndrome. A case was defined as death or critical illness in a hospitalized, previously healthy person, 1 to 49 years of age, with infection hallmarks but no cause identified after routine testing. A total of 137 cases were identified (incidence rate 0.5 per 100,000 per year). Patients' median age was 20 years, 72 (53%) were female, 112 (82%) were white, and 41 (30%) died. The most common clinical presentations were neurologic (29%), respiratory (27%), and cardiac (21%). Infectious causes were identified for 34 cases (28% of the 122 cases with clinical specimens); 23 (68%) were diagnosed by reference serologic tests, and 11 (32%) by polymerase chain reaction-based methods. The UNEX network model would improve U.S. diagnostic capacities and preparedness for emerging infections.

    View details for Web of Science ID 000173757800005

    View details for PubMedID 11897065

  • Stereotyped and specific gene expression programs in human innate immune responses to bacteria PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Boldrick, J. C., Alizadeh, A. A., Diehn, M., Dudoit, S., Liu, C. L., Belcher, C. E., Botstein, D., Staudt, L. M., Brown, P. O., Relman, D. A. 2002; 99 (2): 972-977


    The innate immune response is crucial for defense against microbial pathogens. To investigate the molecular choreography of this response, we carried out a systematic examination of the gene expression program in human peripheral blood mononuclear cells responding to bacteria and bacterial products. We found a remarkably stereotyped program of gene expression induced by bacterial lipopolysaccharide and diverse killed bacteria. An intricately choreographed expression program devoted to communication between cells was a prominent feature of the response. Other features suggested a molecular program for commitment of antigen-presenting cells to antigens captured in the context of bacterial infection. Despite the striking similarities, there were qualitative and quantitative differences in the responses to different bacteria. Modulation of this host-response program by bacterial virulence mechanisms was an important source of variation in the response to different bacteria.

    View details for Web of Science ID 000173450100078

    View details for PubMedID 11805339

    View details for PubMedCentralID PMC117415

  • Sequence variability in the first internal transcribed spacer region within and among Cyclospora species is consistent with polyparasitism INTERNATIONAL JOURNAL FOR PARASITOLOGY Olivier, C., van de Pas, S., Lepp, P. W., Yoder, K., Relman, D. A. 2001; 31 (13): 1475-1487


    Cyclospora cayetanensis is a coccidian parasite which causes severe gastroenteritis in humans. Molecular information on this newly emerging pathogen is scarce. Our objectives were to assess genetic variation within and between human-associated C. cayetanensis and baboon-associated Cyclospora papionis by examining the internal transcribed spacer (ITS) region of the ribosomal RNA operon, and to develop an efficient polymerase chain reaction- (PCR)-based method to distinguish C. cayetanensis from other closely related organisms. For these purposes, we studied C. cayetanensis ITS-1 nucleotide variability in 24 human faecal samples from five geographic locations and C. papionis ITS-1 variability in four baboon faecal samples from Tanzania. In addition, a continuous sequence encompassing ITS-1, 5.8S rDNA and ITS-2 was determined from two C. cayetanensis samples. The results indicate that C. cayetanensis and C. papionis have distinct ITS-1 sequences, but identical 5.8S rDNA sequences. ITS-1 is highly variable within and between samples, but variability does not correlate with geographic origin of the samples. Despite this variability, conserved species-specific ITS-1 sequences were identified and a single-round, C. cayetanensis-specific PCR-based assay with a sensitivity of one to ten oocysts was developed. This consistent and remarkable diversity among Cyclospora spp. ITS-1 sequences argues for polyparasitism and simultaneous transmission of multiple strains.

    View details for Web of Science ID 000171912800010

    View details for PubMedID 11595235

  • Bioterrorism preparedness: What practitioners need to know INFECTIONS IN MEDICINE Relman, D. A., Olson, J. E. 2001; 18 (11): 497-?
  • Does blood of healthy subjects contain bacterial ribosomal DNA? JOURNAL OF CLINICAL MICROBIOLOGY Nikkari, S., McLaughlin, I. J., Bi, W. L., Dodge, D. E., Relman, D. A. 2001; 39 (5): 1956-1959


    Real-time PCR methods with primers and a probe targeting conserved regions of the bacterial 16S ribosomal DNA (rDNA) revealed a larger amount of rDNA in blood specimens from healthy individuals than in matched reagent controls. However, the origins and identities of these blood-associated bacterial rDNA sequences remain obscure.

    View details for Web of Science ID 000168527900048

    View details for PubMedID 11326021

  • Invasion of human respiratory epithelial cells by Bordetella pertussis: Possible role for a filamentous hemagglutinin Arg-Gly-Asp sequence and alpha 5 beta 1 integrin MICROBIAL PATHOGENESIS Ishibashi, Y., Relman, D. A., Nishikawa, A. 2001; 30 (5): 279-288


    Bordetella pertussis, the agent of whooping cough, is capable of invading human respiratory epithelial cells. In this study, we investigated the mechanisms by which B. pertussis invades the human lung epithelial cell line A549 and normal human bronchial epithelial (NHBE) cells. In vitro adhesion and invasion assays using both cell types with a virulent B. pertussis strain and its isogenic mutants revealed profound defects in a mutant deficient in filamentous hemagglutinin (FHA) expression. In addition, a mutant in which an FHA Arg-Gly-Asp (RGD) site had been changed to Arg-Ala-Asp had significantly diminished invasiveness, although its adhesiveness was comparable to that of the parental strain. Furthermore, a synthetic RGD-containing hexapeptide inhibited invasion of both cell types by the virulent strain. These results demonstrate that an RGD sequence of FHA is involved in B. pertussis invasion of epithelial cells in vitro. Monoclonal antibodies directed against human alpha5beta1 integrin, but not other integrins, blocked invasion, indicating that this integrin is involved in B. pertussis invasion. Taken together, these findings suggest that B. pertussis FHA may promote invasion of human respiratory epithelial cells through the interaction of its RGD sequence with host cell alpha5beta1 integrin.

    View details for DOI 10.1006/mpat.2001.0432

    View details for Web of Science ID 000169167200003

    View details for PubMedID 11373122

  • The meaning and impact of the human genome sequence for microbiology TRENDS IN MICROBIOLOGY Relman, D. A., FALKOW, S. 2001; 9 (5): 206-208


    The characterization of life is immeasurably enhanced by determination of complete genome sequences. For organisms that engage in intimate interactions with others, the genome sequence from one participant, and associated tools, provide unique insight into its partner. We discuss how the human genome sequence will further our understanding of microbial pathogens and commensals, and vice versa. We also propose criteria for implicating a host gene in microbial pathogenesis, and urge consideration of a'second human genome project'.

    View details for Web of Science ID 000168719300008

    View details for PubMedID 11336835

  • Localization of Tropheryma whippelii rRNA in tissues from patients with Whipple's disease JOURNAL OF INFECTIOUS DISEASES Fredricks, D. N., Relman, D. A. 2001; 183 (8): 1229-1237


    Whipple's disease is caused by a cultivation-resistant bacterium, Tropheryma whippelii. Ultrastructural studies of intestinal biopsy specimens from patients with Whipple's disease have shown that intracellular and extracellular bacteria are present, but the preferred site of growth is unknown. Tissue sections from 8 patients with Whipple's disease and from 19 healthy control subjects were analyzed by use of fluorescence in situ hybridization and laser scanning confocal microscopy, to determine the location of rRNA that would indicate the presence of metabolically active bacteria. T. whippelii rRNA was most prevalent near the tips of intestinal villi, in the lamina propria, just basal to epithelial cells. Most of the bacterial rRNA signal appeared to be located between cells and did not colocalize with the human intracellular protein vimentin. The location of bacterial rRNA in tissues from patients with Whipple's disease provides evidence that bacteria are growing outside cells and suggests that T. whippelii is not an obligate intracellular pathogen.

    View details for Web of Science ID 000167674600008

    View details for PubMedID 11262205

  • Proinflammatory and proapoptotic activities associated with Bordetella pertussis filamentous hemagglutinin INFECTION AND IMMUNITY Abramson, T., Kedem, H., Relman, D. A. 2001; 69 (4): 2650-2658


    Filamentous hemagglutinin (FHA) is a dominant cell surface-associated Bordetella pertussis adhesin. Recognition that this protein is secreted in significant amounts and that bacterial adhesins may have other activities, prompted an assessment of FHA effects on human macrophages. Incubation of human macrophage-like U937 cells with preparations of FHA resulted in dose-dependent cytotoxicity, with death of 95% of treated cells after 24 h. Based on the use of four independent methods, death of these cells could be largely attributed to apoptosis. FHA-associated apoptosis was also observed in THP-1 macrophage-like cells, fresh human peripheral blood monocyte-derived macrophages (MDM), and BEAS-2B human bronchial epithelial cells. Infection of MDM with wild-type B. pertussis resulted in apoptosis within 6 h, while infection with an FHA-deficient derivative strain was only 50% as effective. FHA-associated cytotoxicity was preceded by host cell secretion of tumor necrosis factor alpha (TNF-alpha), a potential proapoptotic factor. However, pretreatment of cells with a neutralizing anti-TNF-alpha monoclonal antibody inhibited only 16% of the FHA-associated apoptosis. On the other hand, a blocking monoclonal antibody directed against TNF-alpha receptor 1 inhibited FHA-associated apoptosis by 47.7% (P = 0.0001), suggesting that this receptor may play a role in the death pathway activated by FHA. Our in vitro data indicate that secreted and cell-associated FHA elicits proinflammatory and proapoptotic responses in human monocyte-like cells, MDM, and bronchial epithelial cells and suggest a previously unrecognized role for this prominent virulence factor in the B. pertussis-host interaction.

    View details for Web of Science ID 000167616500084

    View details for PubMedID 11254631

  • Comparing functional genomic datasets: lessons from DNA. microarray analyses of host-pathogen interactions CURRENT OPINION IN MICROBIOLOGY Diehn, M., Relman, D. A. 2001; 4 (1): 95-101


    Functional genomic technologies such as high density DNA microarrays allow biologists to study the structure and behavior of thousands of genes in a single experiment. One of the fields in which microarrays have had an increasingly important impact is host-pathogen interactions. Early investigations in this area over the past two years not only emphasize the utility of this approach, but also highlight the stereotyped gene expression responses of different host cells to diverse infectious stimuli, and the potential value of broad dataset comparisons in revealing fundamental features of innate immunity. The comparative analysis of recently published datasets involving human gene expression responses to two bacterial respiratory pathogens illustrates many of these points. Comparisons between these large, highly parallel sets of experimental observations also emphasize important technical and experimental design issues as future challenges.

    View details for Web of Science ID 000166840200015

    View details for PubMedID 11173041

  • Whipple's disease and Tropheryma whippelii: Secrets slowly revealed CLINICAL INFECTIOUS DISEASES Maiwald, M., Relman, D. A. 2001; 32 (3): 457-463


    Whipple's disease was described in 1907 and was designated "intestinal lipodystrophy," despite the detection of bacteria in 1 specimen. This finding was later substantiated by the success of antibiotic therapy, which resulted in dramatic clinical responses, and by use of electron microscopy, which detected monomorphic bacilli in affected tissues. Many attempts at culture failed, and these bacteria were characterized as actinomycetes for the first time by means of broad-range 16S rDNA amplification and molecular phylogenetic methods. The name "Tropheryma whippelii" was proposed for this bacterium. Whipple's disease is a systemic disease that affects many organ systems, producing protean manifestations. This article summarizes recent developments with regard to this topic as well as unanswered questions regarding the pathogenesis and acquisition of infection, the biology and ecology of the organism, the clinical spectrum of disease, diagnosis of the disease, and therapy.

    View details for Web of Science ID 000166674300016

    View details for PubMedID 11170954

  • Tropheryma whippelii DNA is rare in the intestinal mucosa of patients without other evidence of Whipple disease ANNALS OF INTERNAL MEDICINE Maiwald, M., Von Herbay, A., Persing, D. H., Mitchell, P. S., Abdelmalek, M. F., Thorvilson, J. N., Fredricks, D. N., Relman, D. A. 2001; 134 (2): 115-119


    Little is known about the pathogenesis of Whipple disease, the reservoirs of Tropheryma whippelii, and the proportion of persons harboring the bacterium without "classic" intestinal abnormalities.To assess the presence of T. whippelii in patients undergoing upper endoscopy for a variety of indications.Prospective and routine diagnostic examination of patients.Three academic medical centers in California; Minnesota; and Heidelberg, Germany.342 patients undergoing endoscopy for evaluation of dyspepsia or possible peptic ulcer (group A, 173 patients), malabsorption (group B, 37 patients), or clinical suspicion of Whipple disease (group C, 132 patients).Small-intestinal biopsy specimens were tested by polymerase chain reaction for T. whippelii DNA and examined for histopathologic abnormalities.All patients with negative histologic findings also had negative results for T. whippelii DNA.T. whippelii occurs only rarely in intestinal mucosa that lacks histopathologic evidence of Whipple disease. The human small intestinal mucosa is an unlikely reservoir for this organism.

    View details for Web of Science ID 000166356000004

    View details for PubMedID 11177314

  • The transcriptional responses of respiratory epithelial cells to Bordetella pertussis reveal host defensive and pathogen counter-defensive strategies PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Belcher, C. E., Drenkow, J., Kehoe, B., Gingeras, T. R., McNamara, N., Lemjabbar, H., Basbaum, C., Relman, D. A. 2000; 97 (25): 13847-13852


    Bordetella pertussis, the causative agent of whooping cough, has many well-studied virulence factors and a characteristic clinical presentation. Despite this information, it is not clear how B. pertussis interaction with host cells leads to disease. In this study, we examined the interaction of B. pertussis with a human bronchial epithelial cell line (BEAS-2B) and measured host transcriptional profiles by using high-density DNA microarrays. The early transcriptional response to this pathogen is dominated by altered expression of cytokines, DNA-binding proteins, and NFkappaB-regulated genes. This previously unrecognized response to B. pertussis was modified in similar but nonidentical fashions by the antiinflammatory agents dexamethasone and sodium salicylate. Cytokine protein expression was confirmed, as was neutrophil chemoattraction. We show that B. pertussis induces mucin gene transcription by BEAS-2B cells then counters this defense by using mucin as a binding substrate. A set of genes is described for which the catalytic activity of pertussis toxin is both necessary and sufficient to regulate transcription. Host genomic transcriptional profiling, in combination with functional assays to evaluate subsequent biological events, provides insight into the complex interaction of host and pathogen.

    View details for Web of Science ID 000165728800071

    View details for PubMedID 11087813

  • Using DNA microarrays to study host-microbe interactions EMERGING INFECTIOUS DISEASES Cummings, C. A., Relman, D. A. 2000; 6 (5): 513-525


    Complete genomic sequences of microbial pathogens and hosts offer sophisticated new strategies for studying host-pathogen interactions. DNA microarrays exploit primary sequence data to measure transcript levels and detect sequence polymorphisms, for every gene, simultaneously. The design and construction of a DNA microarray for any given microbial genome are straightforward. By monitoring microbial gene expression, one can predict the functions of uncharacterized genes, probe the physiologic adaptations made under various environmental conditions, identify virulence-associated genes, and test the effects of drugs. Similarly, by using host gene microarrays, one can explore host response at the level of gene expression and provide a molecular description of the events that follow infection. Host profiling might also identify gene expression signatures unique for each pathogen, thus providing a novel tool for diagnosis, prognosis, and clinical management of infectious disease.

    View details for Web of Science ID 000089785300011

    View details for PubMedID 10998383

  • Exploring gene expression signatures of host responses to infection Boldrick, J. C., Belcher, C. E., Alizadeh, A. A., Liu, C. L., Diehn, M., Brown, P. O., Relman, D. A. OXFORD UNIV PRESS INC. 2000: 218?18
  • Collaborative multidisciplinary workshop report: Detection, culture, serology, and antimicrobial susceptibility testing of Chlamydia pneumoniae JOURNAL OF INFECTIOUS DISEASES Tompkins, L. S., SCHACHTER, J., Boman, J., Dowell, S., Gaydos, C. A., Levison, M. E., MAASS, M., Madico, G., Orfila, J., Ouchi, K., Peeling, R. W., Taylor-Robinson, D., STAMM, W. E., Wang, S. P., Blasi, F., Relman, D. 2000; 181: S460-S461

    View details for Web of Science ID 000088023900019

    View details for PubMedID 10839740

  • Organization, structure, and variability of the rRNA operon of the Whipple's disease bacterium (Tropheryma whippelii) JOURNAL OF BACTERIOLOGY Maiwald, M., Von Herbay, A., Lepp, P. W., Relman, D. A. 2000; 182 (11): 3292-3297


    Whipple's disease is a systemic disorder associated with a cultivation-resistant, poorly characterized actinomycete, Tropheryma whippelii. We determined a nearly complete rRNA operon sequence of T. whippelii from specimens from 3 patients with Whipple's disease, as well as partial operon sequences from 43 patients. Variability was observed in the 16S-23S rRNA spacer sequences, leading to the description of five distinct sequence types. One specimen contained two spacer sequence types, raising the possibility of a double infection. Secondary structure models for the primary rRNA transcript and mature rRNAs revealed rare or unique features.

    View details for Web of Science ID 000086988000042

    View details for PubMedID 10809715

  • Rhinosporidium seeberi: A human pathogen from a novel group of aquatic protistan parasites EMERGING INFECTIOUS DISEASES Fredricks, D. N., Jolley, J. A., Lepp, P. W., Kosek, J. C., Relman, D. A. 2000; 6 (3): 273-282


    Rhinosporidium seeberi, a microorganism that can infect the mucosal surfaces of humans and animals, has been classified as a fungus on the basis of morphologic and histochemical characteristics. Using consensus polymerase chain reaction (PCR), we amplified a portion of the R. seeberi 18S rRNA gene directly from infected tissue. Analysis of the aligned sequence and inference of phylogenetic relationships showed that R. seeberi is a protist from a novel clade of parasites that infect fish and amphibians. Fluorescence in situ hybridization and R. seeberi- specific PCR showed that this unique 18S rRNA sequence is also present in other tissues infected with R. seeberi. Our data support the R. seeberi phylogeny recently suggested by another group. R. seeberi is not a classic fungus, but rather the first known human pathogen from the DRIPs clade, a novel clade of aquatic protistan parasites (Ichthyosporea).

    View details for Web of Science ID 000087321300007

    View details for PubMedID 10827117

  • Molecular characterization of Bordetella bronchiseptica filamentous haemagglutinin and its secretion machinery MICROBIOLOGY-SGM Jacob-Dubuisson, F., Kehoe, B., Willery, E., Reveneau, N., Locht, C., Relman, D. A. 2000; 146: 1211-1221


    Two closely related pathogens, Bordetella pertussis and Bordetella bronchiseptica, share a number of virulence factors. Filamentous haemagglutinin (FHA) is widely regarded as the dominant adhesin of B. pertussis, and its multiple binding activities have been well characterized. This large protein is produced and secreted at high levels by B. pertussis and significantly lower levels by B. bronchiseptica strains. FHA secretion is mediated by a single outer-membrane accessory protein, FhaC. The genes encoding FHA and FhaC in B. bronchiseptica were characterized by sequencing and functional analyses and are highly similar to those of B. pertussis. The most distinctive feature of B. bronchiseptica FHA is additional repeats in the N-terminal portion of the predicted protein. Interestingly, a point mutation in the fhaB promoter region of the B. bronchiseptica GP1 isolate, relative to other isolates, was found to be detrimental to promoter activity and to FHA production. FhaC and the N-terminal secretion domain of FHA of B. bronchiseptica were fully functional for secretion in B. pertussis. Thus, the different levels of FHA secretion by these Bordetella species might reflect differences in physiology, composition and structure of cell envelope, or differential protein degradation. Characterization of FHA expression and function may provide clues as to the basis of host species tropism, tissue localization and receptor recognition.

    View details for Web of Science ID 000087035400023

    View details for PubMedID 10832649

  • How the host 'sees' pathogens: global gene expression responses to infection CURRENT OPINION IN IMMUNOLOGY Manger, I. D., Relman, D. A. 2000; 12 (2): 215-218


    Innate immune responses to pathogens are believed to be patterned and stereotyped. Adaptive responses display variety but in relatively few types of products and with limited numbers of mechanisms. Is this apparent disparity between microbial pathogen diversity and a restricted set of host responses an accurate picture of infection or is it the result of a limited collection of analytic tools? DNA microarray technology permits one to address simple descriptive questions about global gene expression inside cells. In particular, it offers an opportunity to examine the relationship between host and pathogen in much greater detail than has been possible previously. One can now ask, firstly, how a host cell or organism 'sees' a microbial pathogen from the viewpoint of gene expression responses and, secondly, at what level it is able to discriminate between different agents. Other potential insights to be reaped include the identification of microbial determinants of the host response, the temporal features of the 'conversation' between host and pathogen, novel strategies for therapeutic and prophylactic intervention and prognostic markers of outcome.

    View details for Web of Science ID 000085786300015

    View details for PubMedID 10712949

  • A role for mucin in Bordetella pertussis pathogenesis is revealed in the transcriptional profile of infected human bronchial epithelial cells Belcher, C., Drenkow, J., Gingeras, T., McNamara, N., Basbaum, C., Relman, D. NATURE PUBLISHING GROUP. 2000: 257A?257A
  • Biologic weapons: What infectious disease practitioners need to know INFECTIONS IN MEDICINE Olson, J. E., Relman, D. A. 2000; 17 (1): 29-?
  • Bacterial diversity within the human subgingival crevice PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Kroes, I., Lepp, P. W., Relman, D. A. 1999; 96 (25): 14547-14552


    Molecular, sequence-based environmental surveys of microorganisms have revealed a large degree of previously uncharacterized diversity. However, nearly all studies of the human endogenous bacterial flora have relied on cultivation and biochemical characterization of the resident organisms. We used molecular methods to characterize the breadth of bacterial diversity within the human subgingival crevice by comparing 264 small subunit rDNA sequences from 21 clone libraries created with products amplified directly from subgingival plaque, with sequences obtained from bacteria that were cultivated from the same specimen, as well as with sequences available in public databases. The majority (52.5%) of the directly amplified 16S rRNA sequences were <99% identical to sequences within public databases. In contrast, only 21.4% of the sequences recovered from cultivated bacteria showed this degree of variability. The 16S rDNA sequences recovered by direct amplification were also more deeply divergent; 13.5% of the amplified sequences were more than 5% nonidentical to any known sequence, a level of dissimilarity that is often found between members of different genera. None of the cultivated sequences exhibited this degree of sequence dissimilarity. Finally, direct amplification of 16S rDNA yielded a more diverse view of the subgingival bacterial flora than did cultivation. Our data suggest that a significant proportion of the resident human bacterial flora remain poorly characterized, even within this well studied and familiar microbial environment.

    View details for Web of Science ID 000084149700073

    View details for PubMedID 10588742

  • Absence of Kaposi's sarcoma-associated herpesvirus DNA angiomatosis-peliosis lesions 33rd Annual Meeting of the American-Society-of-Dermatopathology Relman, D. A., Fredricks, D. N., Yoder, K. E., Mirowski, G., Berger, T., Koehler, J. E. UNIV CHICAGO PRESS. 1999: 1386?89


    Bartonella henselae and B. quintana induce an unusual vascular proliferative tissue response known as bacillary angiomatosis (BA) and bacillary peliosis (BP) in some human hosts. The mechanisms of Bartonella-associated vascular proliferation remain unclear. Although host factors probably play a role, microbial coinfection has not been ruled out. Because of the vascular proliferative characteristics noted in both Kaposi's sarcoma (KS) and BA and occasional colocalization of KS and BA, the possibility was explored that KS-associated herpesvirus (KSHV) might be associated with BA lesions. Tissues with BA and positive and negative control tissues were tested for the presence of KSHV DNA by a sensitive polymerase chain reaction assay. Only 1 of 10 BA tissues, a splenic biopsy, was positive in this assay; this tissue was from a patient with concomitant KS of the skin. Thus, KSHV is probably not involved in the vascular proliferative response seen in BA-BP.

    View details for Web of Science ID 000083019500065

    View details for PubMedID 10479179

  • Application of polymerase chain reaction to the diagnosis of infectious diseases CLINICAL INFECTIOUS DISEASES Fredricks, D. N., Relman, D. A. 1999; 29 (3): 475-486

    View details for Web of Science ID 000082703600001

    View details for PubMedID 10530433

  • The search for unrecognized pathogens SCIENCE Relman, D. A. 1999; 284 (5418): 1308-1310


    The distribution and diversity of microorganisms in the world are far greater than have been previously appreciated. Molecular, cultivation-independent methods have played a key role in this insight. To what extent do humans remain ignorant of microbial diversity within the human body and the settings in which microorganisms cause human disease? In addition to implicating microbial agents in nontraditional infectious diseases, the use of methods such as broad-range polymerase chain reaction, representational difference analysis, expression library screening, and host gene expression profiling may force a reassessment of the concepts of microbial disease causation.

    View details for Web of Science ID 000080430600040

    View details for PubMedID 10334977

  • Molecular characterization of Cyclospora-like organisms from baboons 35th Annual Meeting of the Infectious-Diseases-Society-of-America Lopez, F. A., Manglicmot, J., Schmidt, T. M., Yeh, C., Smith, H. V., Relman, D. A. UNIV CHICAGO PRESS. 1999: 670?76


    Cyclospora organisms are intestinal pathogens of humans that are increasingly recognized in many parts of the world; yet, the reservoirs and host range remain poorly defined. Analysis of 18S ribosomal DNA (rDNA) suggests that the human-associated Cyclospora species (Cyc-hu) is most closely related to the Eimeria species, which are host species-specific. Recently, oocysts identical to those of Cyc-hu were detected in baboon fecal specimens from Tanzania. The 18S rDNA from 3 of these baboon-associated oocyst specimens was amplified and sequenced. Phylogenetic analysis indicated that these baboon-associated Cyclospora-like organisms (Cyc-bab) are nearly identical to each other and are distinct from Cyc-hu (1.6%-1.7% dissimilar); however, these Cyc-bab organisms are the closest known relatives of Cyc-hu. Together, these primate-associated cyclosporans constitute a coherent clade within the diverse group of Eimeria species. These findings raise important questions about the evolutionary relationships of the eimeriids and Cyc-hu host range and should lead to improved polymerase chain reaction-based diagnostics.

    View details for Web of Science ID 000079566700017

    View details for PubMedID 9952374

  • Paraffin removal from tissue sections for digestion and PCR analysis BIOTECHNIQUES Fredricks, D. N., Relman, D. A. 1999; 26 (2): 198-?

    View details for Web of Science ID 000078631700004

    View details for PubMedID 10023524

  • Molecular approaches for identification of infectious agents in Wegener's granulomatosis and other vasculitides. Current opinion in rheumatology Nikkari, S., Relman, D. A. 1999; 11 (1): 11-16


    The primary symptoms of many vasculitides resemble those of infectious diseases. Patients with Wegener's granulomatosis usually seek medical care for respiratory tract symptoms resembling those caused by infection or allergy. In addition, vasculitis is a well-documented manifestation of infection by some known microbial agents. There have been relatively few controlled studies, however, seeking to identify infectious agents as the triggering factors in systemic vasculitides. Molecular methods offer powerful approaches for the identification of infectious agents in diseases of previously unknown origin. These methods include broad-range amplification of microbial nucleic acid sequences and comparative or subtractive methods, such as differential display and representational difference analysis. Host gene expression profiles (using DNA-chip technology) may also provide clues as to the possible infectious cause of an idiopathic disease. Furthermore, the application of molecular methods may reveal pathologic mechanisms and novel therapeutic strategies for the vasculitides.

    View details for PubMedID 9894625

  • Filamentous hemagglutinin of Bordetella bronchiseptica is required for efficient establishment of tracheal colonization INFECTION AND IMMUNITY Cotter, P. A., Yuk, M. H., Mattoo, S., Akerley, B. J., Boschwitz, J., Relman, D. A., Miller, J. F. 1998; 66 (12): 5921-5929


    Adherence to ciliated respiratory epithelial cells is considered a critical early step in Bordetella pathogenesis. For Bordetella pertussis, the etiologic agent of whooping cough, several factors have been shown to mediate adherence to cells and cell lines in vitro. These putative adhesins include filamentous hemagglutinin (FHA), fimbriae, pertactin, and pertussis toxin. Determining the precise roles of each of these factors in vivo, however, has been difficult, due in part to the lack of natural-host animal models for use with B. pertussis. Using the closely related species Bordetella bronchiseptica, and by constructing both deletion mutation and ectopic expression mutants, we have shown that FHA is both necessary and sufficient for mediating adherence to a rat lung epithelial (L2) cell line. Using a rat model of respiratory infection, we have shown that FHA is absolutely required, but not sufficient, for tracheal colonization in healthy, unanesthetized animals. FHA was not required for initial tracheal colonization in anesthetized animals, however, suggesting that its role in establishment may be dedicated to overcoming the clearance action of the mucociliary escalator.

    View details for Web of Science ID 000077179600044

    View details for PubMedID 9826374

    View details for PubMedCentralID PMC108750

  • Improved amplification of microbial DNA from blood cultures by removal of the PCR inhibitor sodium polyanetholesulfonate JOURNAL OF CLINICAL MICROBIOLOGY Fredricks, D. N., Relman, D. A. 1998; 36 (10): 2810-2816


    Molecular methods are increasingly used to identify microbes in clinical samples. A common technical problem with PCR is failed amplification due to the presence of PCR inhibitors. Initial attempts at amplification of the bacterial 16S rRNA gene from inoculated blood culture media failed for this reason. The inhibitor persisted, despite numerous attempts to purify the DNA, and was identified as sodium polyanetholesulfonate (SPS), a common additive to blood culture media. Like DNA, SPS is a high-molecular-weight polyanion that is soluble in water but insoluble in alcohol. Accordingly, SPS tends to copurify with DNA. An extraction method was designed for purification of DNA from blood culture media and removal of SPS. Blood culture media containing human blood and spiked with Escherichia coli was subjected to an organic extraction procedure with benzyl alcohol, and removal of SPS was documented spectrophotometrically. Successful amplification of the extracted E. coli 16S rRNA gene was achieved by adding 5 microliter of undiluted processed sample DNA to a 50-microliter PCR mixture. When using other purification methods, the inhibitory effect of SPS could be overcome only by dilution of these samples. By our extraction technique, even uninoculated blood culture media were found to contain bacterial DNA when they were subjected to broad-range 16S rRNA gene consensus PCR. We conclude that the blood culture additive SPS is a potent inhibitor of PCR, is resistant to removal by traditional DNA purification methods, but can be removed by a benzyl alcohol extraction protocol that results in improved PCR performance.

    View details for Web of Science ID 000075896800002

    View details for PubMedID 9738025

  • PCR analysis of T-whippelii DNA in a case of Whipple's disease: Effect of antibiotics and correlation with histology AMERICAN JOURNAL OF GASTROENTEROLOGY Petrides, P. E., Muller-Hocker, J., Fredricks, D. N., Relman, D. A. 1998; 93 (9): 1579-1582


    A 58-yr-old man developed severe weight loss, arthralgias, and diarrhea. Endoscopic examination of the stomach and duodenum revealed thickened folds of duodenal mucosa. Biopsy of the gastric mucosa was negative, whereas duodenal biopsy revealed blunted epithelial villi and PAS-positive foamy macrophages within the lamina propria. Bacilli typical of those associated with Whipple's disease were found by electron microscopy. The diagnosis was confirmed by polymerase chain reaction (PCR) assay, which detected a portion of the 16S ribosomal RNA gene sequence corresponding to the Whipple bacillus (Tropheryma whippelii) in duodenum, stomach, and liver biopsies before therapy. T. whippelii DNA was eliminated from all tissues tested within 3 months of starting antibiotic treatment, but the histological improvement lagged behind the clinical and molecular evidence of improvement.

    View details for Web of Science ID 000075746400043

    View details for PubMedID 9732952

  • Explaining the unexplained in clinical infectious diseases: Looking forward International Conference on Emerging Infectious Diseases Perkins, B. A., Relman, D. CENTER DISEASE CONTROL. 1998: 395?97

    View details for Web of Science ID 000075433800012

    View details for PubMedID 9716953

  • Detection and identification of previously unrecognized microbial pathogens International Conference on Emerging Infectious Diseases Relman, D. A. CENTER DISEASE CONTROL. 1998: 382?89


    Features of a number of important but poorly explained human clinical syndromes strongly indicate a microbial etiology. In these syndromes, the failure of cultivation-dependent microbial detection methods reveals our ignorance of microbial growth requirements. Sequence-based molecular methods, however, offer alternative approaches for microbial identification directly from host specimens found in the setting of unexplained acute illnesses, chronic inflammatory disease, and from anatomic sites that contain commensal microflora. The rapid expansion of genome sequence databases and advances in biotechnology present opportunities and challenges: identification of consensus sequences from which reliable, specific phylogenetic information can be inferred for all taxonomic groups of pathogens, broad-range pathogen identification on the basis of virulence-associated gene families, and use of host gene expression response profiles as specific signatures of microbial infection.

    View details for Web of Science ID 000075433800010

    View details for PubMedID 9716951

  • Molecular and cellular microbiology: new tools of the trade CURRENT OPINION IN MICROBIOLOGY Relman, D. A., Wright, A. 1998; 1 (3): 337-339
  • The 'emergence' of Bartonella and the development of molecular discovery methods for microbial pathogens Conference on the Challenge of Infectious Diseases - the Eurpoean Perspective Relman, D. A. VAN ZUIDEN COMMUNICATIONS. 1998: 249?55

    View details for Web of Science ID 000075358200009

    View details for PubMedID 9718924

  • Are all Bartonella henselae strains created equal? Clinical infectious diseases Relman, D. A. 1998; 26 (6): 1300-1301

    View details for PubMedID 9636851

  • Culture-negative endocarditis caused by Bartonella henselae JOURNAL OF PEDIATRICS Baorto, E., Payne, R. M., Slater, L. N., LOPEZ, F., Relman, D. A., Min, K. W., StGeme, J. W. 1998; 132 (6): 1051-1054


    A 4-year-old girl presented with clinical evidence of infective endocarditis involving her aortic valve, but blood cultures were sterile. Serologic studies and analysis of resected valve by immunohistochemistry and polymerase chain reaction established the diagnosis of Bartonella henselae endocarditis. Clinicians should be aware that B. henselae can cause apparent culture-negative endocarditis in children.

    View details for Web of Science ID 000074065100030

    View details for PubMedID 9627605

  • Diagnostic utility of the polymerase chain reaction in 2 cases of suspected Whipple disease ARCHIVES OF INTERNAL MEDICINE Tasken, K., Schulz, T., Elgjo, K., Skullerud, K., Relman, D., Brubakk, O. 1998; 158 (7): 801-803


    We describe 2 patients with a diagnosis of Whipple disease in whom the usual antibiotic therapy failed. A polymerase chain reaction-based test was used to identify the recently described Whipple bacillus, Tropheryma whippelii. In one case, the diagnosis was confirmed, whereas in the second case, which had been histologically diagnosed as Whipple disease of the brain, the process was identified as a monocyte-derived histiocytosis. In conclusion, Whipple disease can be distinguished from other diseases with similar histological features with the use of a polymerase chain reaction-based test.

    View details for Web of Science ID 000073083800015

    View details for PubMedID 9554687

  • Cyclospora INFECTIOUS DISEASE CLINICS OF NORTH AMERICA Soave, R., Herwaldt, B. L., Relman, D. A. 1998; 12 (1): 1-?


    Although Cyclospora infection has been documented in humans worldwide since at least 1977, it is only in the past 2 years that this organism has come into prominence as a result of major foodborne outbreaks in the United States and Canada. Cyclospora causes significant gastrointestinal disease in immunocompetent and immunocompromised hosts and can be successfully treated with trimethoprim-sulfamethoxazole. The infection is under-recognized because our methods for diagnosis are rudimentary and insensitive. The mechanisms by which the parasite causes disease, the range of animal hosts, and the natural reservoir are unknown. Cyclospora is a unique coccidian parasite that has just begun to emerge; as yet, we have no clue as to where it comes from or where it hides.

    View details for Web of Science ID 000072123000002

    View details for PubMedID 9494825

  • Radiologic features of a fatal platyhelminth (tapeworm) infection in an AIDS patient AMERICAN JOURNAL OF ROENTGENOLOGY Stark, P., Relman, D. A., Santamaria-Fries, M., Fajardo, L. F. 1998; 170 (1): 136-138

    View details for Web of Science ID 000071081000037

    View details for PubMedID 9423618

  • Infectious agents and the etiology of chronic idiopathic diseases. Current clinical topics in infectious diseases Fredricks, D. N., Relman, D. A. 1998; 18: 180-200


    At the end of the nineteenth century, the field of microbiology was born, and the infectious nature of many previously unexplained diseases was illuminated as powerful new technology was applied. At the end of the twentieth century, the etiology of myriad chronic diseases remains unexplained. We have argued that many of these diseases have clinical, epidemiological, and pathological features that suggest a role for microbes in their pathogenesis. Although definitive evidence of microbial disease causation is lacking, we believe that new technologies, such as sequence-based microbial identification, will successfully be applied to many of these chronic idiopathic diseases in the near future. As novel pathogens and previously described pathogens are revealed as the causative agents for some of these conditions, new diagnostic, preventive, and therapeutic modalities may emerge, transforming some diseases from idiopathic and chronic, to infectious and curable.

    View details for PubMedID 9779355

  • Molecular and cellular microbiology: new tools of the trade. Current opinion in microbiology Relman, D. A., Wright, A. 1998; 1 (3): 337?39

    View details for PubMedID 10066502

  • Bordetella bronchiseptica expresses the fimbrial structural subunit gene fimA JOURNAL OF BACTERIOLOGY BOSCHWITZ, J. S., VANDERHEIDE, H. G., Mooi, F. R., Relman, D. A. 1997; 179 (24): 7882-7885


    The differential host species specificities of Bordetella pertussis, B. parapertussis, and B. bronchiseptica might be explained by polymorphisms in adherence factor genes. We have found that B. parapertussis and B. bronchiseptica, unlike B. pertussis, contain a full-length gene for the fimbrial subunit FimA. B. bronchiseptica expresses fimA in a BvgAS-dependent fashion.

    View details for Web of Science ID A1997YL26600037

    View details for PubMedID 9401052

  • Cultivation of Whipple bacillus: the irony and the ecstasy LANCET Fredricks, D. N., Relman, D. A. 1997; 350 (9087): 1262-1263

    View details for Web of Science ID A1997YD68900002

    View details for PubMedID 9357400

  • Editorial: The Whipple bacillus lives (ex vivo)! JOURNAL OF INFECTIOUS DISEASES Relman, D. A. 1997; 176 (3): 752-754

    View details for Web of Science ID A1997XT81900028

    View details for PubMedID 9291325

  • Bordetella pertussis infection of human monocytes inhibits antigen-dependent CD4 T cell proliferation Antibody Workshop - The Role of Humoral Immunity in the Treatment and Prevention of Emerging and Extant Infectious Diseases BOSCHWITZ, J. S., Batanghari, J. W., Kedem, H., Relman, D. A. OXFORD UNIV PRESS INC. 1997: 678?86


    Human monocytes and macrophages bind Bordetella pertussis through multiple specific receptor-ligand interactions; however, the effect of these interactions on monocyte and macrophage function is not well understood. In an in vitro system, B. pertussis infection of human monocytes significantly impaired T cell proliferation to exogenous antigen at MOIs as low as 1.0. B. pertussis isogenic mutant strains deficient in filamentous hemagglutinin or adenylate cyclase toxin were incapable of proliferation inhibition, suggesting that these virulence-associated factors are essential for this activity. B. pertussis-induced monocyte death alone did not explain these results, nor did differences in intracellular survival. In addition, B. pertussis infection did not significantly alter monocyte phagocytosis of complement-opsonized latex particles, indicating that B. pertussis infection does not globally impair monocyte functions in this system. These results suggest that B. pertussis may be capable of subverting cellular immune defenses in an infected host.

    View details for Web of Science ID A1997XT81900018

    View details for PubMedID 9291315

  • Tropheryma whippelii and arthritides: A more frequent association than thought? Georgescu, L., Raman, C., Relman, D., Paget, S. WILEY-BLACKWELL. 1997: 684?84
  • Polymerase chain reaction-based detection of Tropheryma whippelii in central nervous system Whipple's disease ANNALS OF NEUROLOGY Lynch, T., Odel, J., Fredericks, D. N., Louis, E. D., Forman, S., Rotterdam, H., Fahn, S., Relman, D. A. 1997; 42 (1): 120-124


    Whipple's disease of the central nervous system (CNS) may be associated with normal intestinal histology as a result of minimal or patchy involvement. The diagnosis is difficult and is frequently made post mortem. We studied 6 patients with clinically suspected CNS Whipple's disease; 2 had oculomasticatury myorhythmia (OMM) fitting criteria for a diagnosis of definite CNS Whipple's disease. One of the 2 had duodenal histology highly suggestive of Whipple's disease the other 5 patients had normal duodenal histology. DNA was extracted from paraffin-embedded duodenal tissues in all patients and frozen pontine tissue in 1. Two primer pairs (W3F-W4R, W3F-W2R) were used in separate polymerase chain reactions (PCRs) to amplify fragments of Tropberyma whippelii 16S rDNA from these tissue samples. PCR amplicons were detected only in the duodenal tissues from the 2 patients with OMM. The sequences of these amplicons were identical to the corresponding region of the previously published Tropheryma whippelii 16S rDNA sequence. PCR-based assays of intestinal or brain tissue may be of value for confirming, and possibly refuting, a clinical diagnosis of CNS Whipple's disease in a patient with any combination of dementia, supranuclear gaze palsy, hypothalamic manifestations, myoclonus, seizures, ataxia, or OMM, especially when tissue histology is unrevealing.

    View details for Web of Science ID A1997XK10600018

    View details for PubMedID 9225695

  • Emerging infections and newly-recognised pathogens NETHERLANDS JOURNAL OF MEDICINE Relman, D. A. 1997; 50 (5): 216-220


    Clinicians and microbiologists have for many years relied on growth and characterisation of micro-organisms in the laboratory as the major method for their detection and identification, but reliance upon microbial growth in the laboratory has probably significantly limited our ability to recognise important pathogenic micro-organisms. The traditional methods are often slow, non-specific and insensitive, and sometimes discriminate poorly among microbial species and strains. It is now known that the evolutionary ancestry and interrelationships of all living organisms can be reliably inferred from sequences in their genetic material. Highly conserved sequences characterise broad phylogenetic groups and variable sequences allow specific identification. Sequence-based methods combined with DNA amplification methods, such as the polymerase chain reaction (PCR), have led to powerful molecular identification techniques such as consensus nucleic acid amplification and representational difference analysis. These methods allow one to detect and isolate informative gene sequences from occult microbial pathogens in human tissues. Sequence-based methods are often quicker, more sensitive and more specific than traditional methods not only in detecting known microbial pathogens, but also in identifying previously-uncharacterised micro-organisms. Widespread, organised use of these methods will reveal new emerging microbial pathogens, implicate microbes in the aetiology of poorly-understood chronic inflammatory diseases and significantly expand our understanding of microbial diversity.

    View details for Web of Science ID A1997XA07300006

    View details for PubMedID 9175403

  • Diagnosis and monitoring of whipple disease by polymerase chain reaction ANNALS OF INTERNAL MEDICINE Ramzan, N. N., Loftus, E., Burgart, L. J., Rooney, M., Batts, K. P., Wiesner, R. H., Fredricks, D. N., Relman, D. A., Persing, D. H. 1997; 126 (7): 520-?


    Whipple disease is a chronic, multisystem disorder associated with infection with Tropheryma whippelii, an organism that has not yet been grown on artificial media. In some cases, the diagnosis of Whipple disease is uncertain if it is based on histology alone. Although antibiotic regimens of various durations have been used, the disease recurs in about one third of cases. No test for cure is available.To develop a test that is more sensitive and specific than histologic examination to diagnose Whipple disease and monitor the effects of antibiotic therapy.Retrospective, laboratory-based evaluations of stored tissue specimens.30 patients with clinically diagnosed, histologically confirmed Whipple disease and 8 patients in whom Whipple disease was clinically suspected but who did not have definitive histologic evidence.Pretreatment and post-treatment biopsy specimens of the small bowel and lymph node were tested by polymerase chain reaction for the presence of T. whippeli DNA.Results on PCR were positive in 29 of the 30 specimens from patients with histologically confirmed disease (sensitivity, 96.6%; specificity, 100%) and in 7 of the 8 specimens from patients in whom disease was clinically suspected. Small-bowel biopsy specimens were obtained after treatment from 17 patients (median duration of follow-up, 119 months); specimens from 12 of these patients had positive results on PCR. When these cases were correlated with therapeutic outcome, it was found that 7 of the 12 patients had clinical relapse during subsequent follow-up or had never responded to treatment (positive predictive value, 58% [95% CI, 28% to 85%]). In contrast, none of the 5 patients whose post-treatment biopsy specimens had negative results on PCR had relapse (negative predictive value, 100% [CI, 48% to 100%]; P = 0.044). No correlation was found between post-treatment histology and clinical outcome (P > 0.2).Polymerase chain reaction is highly sensitive and specific when used to confirm the diagnosis of Whipple disease, to identify inconclusive and suspicious cases, and to monitor response to therapy. A negative result on PCR may predict a low likelihood of clinical relapse; a positive test result that remains positive despite therapy may be associated with a poor clinical outcome. Histopathologic evaluation of post-treatment specimens does not predict clinical cure or relapse.

    View details for Web of Science ID A1997WQ19500013

    View details for PubMedID 9092317

  • Limited role for PCR-based diagnosis of Whipple's disease from peripheral blood mononuclear cells LANCET Marth, T., Fredricks, D., Strober, W., Relman, D. A. 1996; 348 (9019): 66-67

    View details for Web of Science ID A1996UV92300061

    View details for PubMedID 8691962

  • Lethal infection by a previously unrecognised metazoan parasite LANCET SantamariaFries, M., Fajardo, L. F., Sogin, M. L., Olson, P. D., Relman, D. A. 1996; 347 (9018): 1797-1801


    New microbial pathogens or variant clinical manifestations of known organisms may be first found in immunodeficient patients. An HIV-infected man developed a rapidly-enlarging abdominal mass, suggestive of a neoplasm, that subsequently invaded his liver and caused death. Initial studies showed unusual tissue morphology that could not be matched with any known disease process.Tissues obtained from biopsy at laparotomy and necropsy were studied by light microscopy, immunohistochemistry, electron microscopy, and broad-range ribosomal DNA-amplification and sequence analysis.Tissue lesions were characterised by peculiar cytoplasmic sacs containing minute cells with very prominent nucleoli. The pathological process was recognised as a parasitic infection, although its features were different from those of any known eukaryotic pathogen. Phylogenetic analysis of a 357 bp 18S rDNA sequence amplified directly from the involved tissue indicated that the causative agent was a previously-uncharacterised cestode.Fatal disease produced by this newly recognised cestode may not be limited to immunodeficient hosts. Awareness of this metazoan infection may allow early diagnosis-by morphology and DNA sequence analysis--and perhaps successful treatment of subsequent cases.

    View details for Web of Science ID A1996UU46900011

    View details for PubMedID 8667924

  • Molecular diagnosis and monitoring of Whipple's disease. Ramzan, N. N., Loftus, E. V., Burgart, L., Rooney, M., Batts, K., Wiesner, R. H., Fredricks, D. N., Relman, D. A., Persing, D. H. W B SAUNDERS CO-ELSEVIER INC. 1996: A998?A998
  • Advances in the diagnosis of uveitis Relman, D. A. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 1996: 52?52
  • Molecular phylogenetic analysis of Cyclospora, the human intestinal pathogen, suggests that it is closely related to Eimeria species JOURNAL OF INFECTIOUS DISEASES Relman, D. A., Schmidt, T. M., Gajadhar, A., Sogin, M., Cross, J., Yoder, K., SETHABUTR, O., Echeverria, P. 1996; 173 (2): 440-445


    A coccidian organism assigned to the genus Cyclospora has been increasingly recognized in association with prolonged diarrhea in humans throughout the world. Confusion surrounds the taxonomy of this fastidious organism, despite the availability of morphology and sporulation characteristics. The small subunit rRNA coding region from cyclosporan oocysts purified from a human fecal specimen was amplified and sequenced. The same sequence was present in specimens from 8 other patients with cyclosporan oocysts but absent in specimens from asymptomatic subjects and from cryptosporidiosis patients. Phylogenetic analysis of rDNA sequences reveals that the human-associated Cyclospora is closely related to members of the Eimeria genus. These results allow predictions concerning Cyclospora host specificity, life cycle, and epidemiology as well as the development of a specific polymerase chain reaction-based diagnostic assay.

    View details for Web of Science ID A1996TT66500022

    View details for PubMedID 8568307

  • Sequence-based identification of microbial pathogens: A reconsideration of Koch's postulates CLINICAL MICROBIOLOGY REVIEWS Fredricks, D. N., Relman, D. A. 1996; 9 (1): 18-?


    Over 100 years ago, Robert Koch introduced his ideas about how to prove a causal relationship between a microorganism and a disease. Koch's postulates created a scientific standard for causal evidence that established the credibility of microbes as pathogens and led to the development of modern microbiology. In more recent times, Koch's postulates have evolved to accommodate a broader understanding of the host-parasite relationship as well as experimental advances. Techniques such as in situ hybridization, PCR, and representational difference analysis reveal previously uncharacterized, fastidious or uncultivated, microbial pathogens that resist the application of Koch's original postulates, but they also provide new approaches for proving disease causation. In particular, the increasing reliance on sequence-based methods for microbial identification requires a reassessment of the original postulates and the rationale that guided Koch and later revisionists. Recent investigations of Whipple's disease, human ehrlichiosis, hepatitis C, hantavirus pulmonary syndrome, and Kaposi's sarcoma illustrate some of these issues. A set of molecular guidelines for establishing disease causation with sequence-based technology is proposed, and the importance of the scientific concordance of evidence in supporting causal associations is emphasized.

    View details for Web of Science ID A1996TP77400002

    View details for PubMedID 8665474

  • A molecular approach to bacterial diversity in the human mouth. Kroes, I., Morton, A., Mickelsen, P. A., Relman, D. A. LIPPINCOTT WILLIAMS & WILKINS. 1996: A126?A126
  • Bordetella pertussis and monocyte integrin signaling. Claus, S. V., Laudanna, C., Relman, D. A. LIPPINCOTT WILLIAMS & WILKINS. 1996: A110?A110

    View details for Web of Science ID A1995QF77000010

    View details for PubMedID 7529896

  • Brief report: uveitis caused by Tropheryma whippelii (Whipple's bacillus) New England journal of medicine RICKMAN, L. S., Freeman, W. R., Green, W. R., Feldman, S. T., Sullivan, J., Russack, V., Relman, D. A. 1995; 332 (6): 363-366

    View details for PubMedID 7529892



    The use of conventional methods to detect a possible infectious cause of Kawasaki disease (KD) has been unsuccessful. Using the polymerase chain reaction and DNA hybridization techniques, we have sought evidence that a known or new herpesvirus, parvovirus, or bacterial pathogen is related etiologically to KD. Peripheral blood DNA from acute KD patients was subjected to amplification and dot-blot hybridization to detect the presence of herpesvirus DNA, and acute KD peripheral blood and serum DNA were subjected to dot-blot hybridization for the presence of parvoviral DNA. All samples were negative for both herpesvirus and parvovirus DNA. In addition, we analyzed buffy-coat white blood cell DNA, synovial fluid DNA, and frozen autopsy and formalin-fixed, paraffin-embedded myocardial tissue DNA from KD patients for the presence of highly conserved bacterial 16S ribosomal RNA gene sequences with the polymerase chain reaction, and all were negative. These results argue against a direct pathogenic role for herpesviruses, parvoviruses, and bacteria in KD. This approach to the detection of highly conserved genomic sequences among broad groups of microorganisms can be adapted for the detection of other groups of microorganisms and may yet prove useful in the search for an etiologic agent of KD.

    View details for Web of Science ID A1994PP80100003

    View details for PubMedID 7877872



    Haemophilus parainfluenzae is both a human oropharyngeal commensal bacterium and a cause of serious invasive disease. The fastidious growth characteristics of this organism and the poor specificity of traditional methods for species identification are likely to have led to inaccuracies in the diagnosis of infections caused by H. parainfluenzae and related organisms. We report a case of H. parainfluenzae endocarditis in which confusion related to microbial identification was resolved by the analysis of 16S ribosomal RNA sequences. Rapid identification was facilitated by amplification of 16S ribosomal DNA directly from cultured cells with use of the polymerase chain reaction and by direct DNA sequence determination of the amplified product. This procedure is potentially useful for the identification of fastidious bacterial pathogens by reference laboratories.

    View details for Web of Science ID A1994PK85700006

    View details for PubMedID 7528552



    Bordetella pertussis, the causative agent of whooping cough, adheres to human monocytes/macrophages by means of a bacterial surface-associated protein, filamentous hemagglutinin (FHA) and the leukocyte integrin, complement receptor 3 (CR3, alpha M beta 2, CD11b/CD18). We show that an FHA Arg-Gly-Asp site induces enhanced B. pertussis binding to monocytes, and that this enhancement is blocked by antibodies directed against CR3. Enhancement requires a monocyte signal transduction complex, composed of leukocyte response integrin (alpha? beta 3) and integrin-associated protein (CD47). This complex is known to upregulate CR3 binding activity. Thus, a bacterial pathogen enhances its own attachment to host cells by coopting a host cell signaling pathway.

    View details for Web of Science ID A1994PJ70300006

    View details for PubMedID 7931059

  • Bacillary angiomatosis and Rochalimaea species. Current clinical topics in infectious diseases Relman, D. A. 1994; 14: 205-219

    View details for PubMedID 8086116


    View details for Web of Science ID A1994BA88R00016

    View details for PubMedID 7520119



    Clinicians have long been aware of human diseases that are associated with visible but uncultured microorganisms. Without the ability to cultivate these organisms, they have remained unidentified. Environmental microbiologists have also discovered on the basis of recent advances in the field of molecular phylogeny that culture-based methods for detecting microorganisms are biased and insensitive. A culture-independent experimental approach is described for the identification of microbial pathogens. This approach incorporates fundamental aspects of 16S rRNA-based molecular phylogeny as well as nucleic acid amplification technology. From its application to Whipple's disease, one can speculate as to the potential insights a highly sensitive, culture-independent method may provide into the diversity and natural ecology of human microbial pathogens.

    View details for Web of Science ID A1993LH88000001

    View details for PubMedID 7685802


    View details for Web of Science ID A1993LB49500019

    View details for PubMedID 7682656



    The combination of enzymatic nucleic acid amplification techniques with 16S rRNA-based molecular phylogeny has brought about a new approach to the identification of microbial pathogens that can not be cultivated in the laboratory. The applications of this experimental approach to bacillary angiomatosis and to Whipple's disease have revealed the presence of two previously uncharacterized organisms. These results suggest the existence of a far greater microbial diversity among human pathogens than has been so far appreciated with culture-dependent methods. PCR-based studies of aquatic environmental microbial communities have already reached similar conclusions. As a result, new and provocative questions are raised concerning the association of amplified 16S rRNA sequences with diseased tissue. The answers must await the results of further investigations and the expansion of sequence data bases.

    View details for Web of Science ID A1992KD52000003

    View details for PubMedID 1285351

  • Localization of Mycobacterium avium-intracellulare within a skin lesion of bacillary angiomatosis in a patient with AIDS. Diagnostic molecular pathology SAGERMAN, P. M., Relman, D. A., Niroomand, F., Niedt, G. W. 1992; 1 (3): 212-216


    We report a 39-year-old man who had AIDS and who presented with an unusual cutaneous vascular lesion, which was clinically thought to be Kaposi's sarcoma. Histologically, the lesion was characterized by capillary proliferation and a mixed inflammatory infiltrate that included numerous histiocytes. The lesion was found to contain slender intracellular acid-fast bacilli, as well as plump extracellular Warthin-Starry-positive bacilli. The acid-fast bacilli were confirmed to be Mycobacterium avium-intracellulare by subsequent positive blood cultures for this organism. To further investigate the lesion, polymerase chain reaction DNA amplification and sequencing was performed, and the lesion was found to contain DNA sequences identical to those previously established for the agent of bacillary angiomatosis. The lesion is thought to represent a lesion of bacillary angiomatosis with secondary involvement by M. avium-intracellulare.

    View details for PubMedID 1285277



    Whipple's disease is a systemic disorder known for 85 years to be associated with an uncultured, and therefore unidentified, bacillus.We used a molecular genetic approach to identify this organism. The bacterial 16S ribosomal RNA (rRNA) sequence was amplified directly from tissues of five unrelated patients with Whipple's disease by means of the polymerase chain reaction, first with broad-range primers and then with specific primers. We determined and analyzed the nucleotide sequence of the amplification products.A unique 1321-base bacterial 16S rRNA sequence was amplified from duodenal tissue of one patient. This sequence indicated the presence of a previously uncharacterized organism. We then detected this sequence in tissues from all 5 patients with Whipple's disease, but in none of those from 10 patients without the disorder. According to phylogenetic analysis, this bacterium is a gram-positive actinomycete that is not closely related to any known genus.We have identified the uncultured bacillus associated with Whipple's disease. The phylogenetic relations of this bacterium, its distinct morphologic characteristics, and the unusual features of the disease are sufficient grounds for naming this bacillus Tropheryma whippelii gen. nov. sp. nov. Our findings also provide a basis for a specific diagnostic test for this organism.

    View details for Web of Science ID A1992JF65500001

    View details for PubMedID 1377787



    Bacillary angiomatosis (BA) and chronic bartonellosis are bacterial infections of humans which result in an unusual vascular proliferative tissue response. In order to determine their phylogenetic relationships, we have determined greater than 95% of the 16S rRNA sequences for these two organisms by amplification directly from infected BA tissue and from a Bartonella bacilliformis lyophilized culture. The BA agent and B. bacilliformis are closely related alpha-proteobacteria (98.5%), although the BA agent is more closely related to Rochalimaea quintana (99.1%). Contrary to previous belief, the BA agent is distinct from, and less closely related to, the cat scratch bacillus (Afipia felis) (90.7%). We propose a novel secondary structure in a hypervariable region of the 16S rRNA which is useful for alignment of primary sequences and which may be useful for design of nucleic acid probes.

    View details for Web of Science ID A1992JC20400009

    View details for PubMedID 1378524

  • THE AGENT OF BACILLARY ANGIOMATOSIS - REPLY NEW ENGLAND JOURNAL OF MEDICINE Relman, D. A., Loutit, J. S., Schmidt, T. M., FALKOW, S., Tompkins, L. S. 1991; 324 (21): 1513-1513

    View details for Web of Science ID A1991FM46000024

    View details for PubMedID 2023615



    Bacillary angiomatosis is an infectious disease causing proliferation of small blood vessels in the skin and visceral organs of patients with human immunodeficiency virus infection and other immunocompromised hosts. The agent is often visualized in tissue sections of lesions with Warthin-Starry staining, but the bacillus has not been successfully cultured or identified. This bacillus may also cause cat scratch disease.In attempting to identify this organism, we used the polymerase chain reaction. We used oligonucleotide primers complementary to the 16S ribosomal RNA genes of eubacteria to amplify 16S ribosomal gene fragments directly from tissue samples of bacillary angiomatosis. The DNA sequence of these fragments was determined and analyzed for phylogenetic relatedness to other known organisms. Normal tissues were studied in parallel.Tissue from three unrelated patients with bacillary angiomatosis yielded a unique 16S gene sequence. A sequence obtained from a fourth patient with bacillary angiomatosis differed from the sequence found in the other three patients at only 4 of 241 base positions. No related 16S gene fragment was detected in the normal tissues. These 16S sequences associated with bacillary angiomatosis belong to a previously uncharacterized microorganism, most closely related to Rochalimaea quintana.The cause of bacillary angiomatosis is a previously uncharacterized rickettsia-like organism, closely related to R. quintana. This method for the identification of an uncultured pathogen may be applicable to other infectious diseases of unknown cause.

    View details for Web of Science ID A1990EK69600001

    View details for PubMedID 2233945



    During the course of whooping cough, Bordetella pertussis interacts with alveolar macrophages and other leukocytes on the respiratory epithelium. We report here mechanisms by which these bacteria adhere to human macrophages in vitro. Whole bacteria adhere by means of two proteins, filamentous hemagglutinin (FHA) and pertussis toxin, either of which is sufficient to mediate adherence. FHA interacts with two classes of molecules on macrophages, galactose-containing glycoconjugates and the integrin CR3 (alpha M beta 2, CD11b/CD18). The interaction between CR3 and FHA involves recognition of the Arg-Gly-Asp (RGD) sequence at positions 1097-1099 in FHA. This study demonstrates that bacterial adherence can be based on the interaction of a bacterial adhesin RGD sequence with an integrin and that bacterial adhesins can have multiple binding sites characteristic of eukaryotic extracellular matrix proteins.

    View details for Web of Science ID A1990DM15600025

    View details for PubMedID 2364431



    The nucleotide sequence of the structural gene for filamentous haemagglutinin (FHA), fhaB, a crucial adherence factor for Bordetella pertussis, has been determined. Its 10774 nucleotides are far more than necessary to encode the 220 kD biologically active, mature polypeptide product, suggesting a role for co- or post-translational processing. Fusion proteins derived from various portions of the fhaB open reading frame (ORF) were used to generate polyclonal antisera. Western immunoblot analysis of purified FHA and Bordetella sp. whole cell extracts with these antisera indicated that the 220 kD product is encoded by the 5' portion of the ORF and that the smaller polypeptide species are breakdown products of this polypeptide. These data, as well as N-terminal amino acid sequencing of the major polypeptide species, suggest a scheme for the proteolytic processing of an FHA precursor polypeptide.

    View details for Web of Science ID A1990DF11700011

    View details for PubMedID 2388559



    Filamentous hemagglutinin (FHA) is a cell surface protein of Bordetella pertussis which functions as an adhesin for this organism. It is a component of many new acellular pertussis vaccines. The proposed role of FHA in immunity to pertussis is based on animal studies which have produced some conflicting results. To clarify this situation, we reexamined the protective activity of FHA in an adult mouse respiratory infection model. Four-week-old BALB/c mice were immunized with one or two doses of 4 or 8 micrograms of FHA and then aerosol challenged with B. pertussis Tohama I. In control mice receiving tetanus toxoid, the CFU in the lungs increased from 10(5) immediately following infection to greater than 10(6) by days 5 and 9 after challenge. Mice immunized with FHA by the intraperitoneal or intramuscular route had significantly reduced bacterial colonization in the lungs. A decrease in colonization of the trachea was also observed in FHA-immunized mice. Evaluation of antibody responses in these mice revealed high titers of immunoglobulin G (IgG) and IgM to FHA in sera and of IgG to FHA in lung lavage fluids. No IgA to FHA was detected. BALB/c mice were also passively immunized intravenously with either goat or rat antibodies to FHA and then aerosol challenged 24 h later, when anti-FHA antibodies were detected in the respiratory tract. Lung and tracheal colonization was markedly reduced in mice immunized with FHA-specific antibodies compared with those receiving control antibodies. In additional studies, the role of FHA in the colonization of the mouse respiratory tract was evaluated by using strain BP101, an FHA mutant of B. pertussis. FHA was important in the initial colonization of the mouse trachea, but was not required for colonization of the trachea later in the infection. FHA was not a factor in colonization of the lungs. Collectively, these experiments demonstrate (i) that systemic immunization with FHA can provide significant protection against B. pertussis infection in both the lower and upper respiratory tract of mice as defined by the lungs and trachea, respectively; (ii) that this protection is mediated primarily by serum antibodies to FHA, which transudate into respiratory secretions; and (iii) that FHA is an important upper respiratory tract colonization factor. These studies provide further evidence for the role of FHA in pertussis pathogenesis and immunity.

    View details for Web of Science ID A1990CE82000002

    View details for PubMedID 2294058



    Filamentous hemagglutinin is a surface-associated adherence protein of Bordetella pertussis, which is a component of some new acellular pertussis vaccines. The nucleotide sequence of an open reading frame that encompasses the filamentous hemagglutinin structural gene, fhaB, suggests that proteolytic processing is necessary to generate the mature 220-kDa filamentous hemagglutinin product. An Arg-Gly-Asp (RGD) tripeptide is found within filamentous hemagglutinin that may be involved in its adherence properties. An internal in-frame deletion in fhaB, encompassing the RGD region, causes loss of B. pertussis-binding to ciliated eukaryotic cells, confirming a potential role for this protein in host-cell binding and infection.

    View details for Web of Science ID A1989U231400025

    View details for PubMedID 2539596


    View details for Web of Science ID A1987G858500048

    View details for PubMedID 3559282

  • MECHANICAL INSTABILITY OF OXY-FORM OF SICKLE HEMOGLOBIN NATURE Asakura, T., AGARWAL, P. L., Relman, D. A., McCray, J. A., Chance, B., Schwartz, E., Friedman, S., Lubin, B. 1973; 244 (5416): 437-438

    View details for Web of Science ID A1973Q428000034

    View details for PubMedID 4582496

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