A framework for human microbiome research

A variety of microbial communities and their genes (the microbiome) exist throughout the human body, with fundamental roles in human health and disease. The National Institutes of Health (NIH)-funded Human Microbiome Project Consortium has established a population-scale framework to develop metagenomic protocols, resulting in a broad range of quality-controlled resources and data including standardized methods for creating, processing and interpreting distinct types of high-throughput metagenomic data available to the scientific community. Here we present resources from a population of 242 healthy adults sampled at 15 or 18 body sites up to three times, which have generated 5,177 microbial taxonomic profiles from 16S ribosomal RNA genes and over 3.5 terabases of metagenomic sequence so far. In parallel, approximately 800 reference strains isolated from the human body have been sequenced. Collectively, these data represent the largest resource describing the abundance and variety of the human microbiome, while providing a framework for current and future studies

Structure, function and diversity of the healthy human microbiome

Author Posting. © The Authors, 2012. This article is posted here by permission of Nature Publishing Group. The definitive version was published in Nature 486 (2012): 207-214, doi:10.1038/nature11234.Studies of the human microbiome have revealed that even healthy individuals differ remarkably in the microbes that occupy habitats such as the gut, skin and vagina. Much of this diversity remains unexplained, although diet, environment, host genetics and early microbial exposure have all been implicated. Accordingly, to characterize the ecology of human-associated microbial communities, the Human Microbiome Project has analysed the largest cohort and set of distinct, clinically relevant body habitats so far. We found the diversity and abundance of each habitat’s signature microbes to vary widely even among healthy subjects, with strong niche specialization both within and among individuals. The project encountered an estimated 81–99% of the genera, enzyme families and community configurations occupied by the healthy Western microbiome. Metagenomic carriage of metabolic pathways was stable among individuals despite variation in community structure, and ethnic/racial background proved to be one of the strongest associations of both pathways and microbes with clinical metadata. These results thus delineate the range of structural and functional configurations normal in the microbial communities of a healthy population, enabling future characterization of the epidemiology, ecology and translational applications of the human microbiome.This research was supported in part by National Institutes of Health grants U54HG004969 to B.W.B.; U54HG003273 to R.A.G.; U54HG004973 to R.A.G., S.K.H. and J.F.P.; U54HG003067 to E.S.Lander; U54AI084844 to K.E.N.; N01AI30071 to R.L.Strausberg; U54HG004968 to G.M.W.; U01HG004866 to O.R.W.; U54HG003079 to R.K.W.; R01HG005969 to C.H.; R01HG004872 to R.K.; R01HG004885 to M.P.; R01HG005975 to P.D.S.; R01HG004908 to Y.Y.; R01HG004900 to M.K.Cho and P. Sankar; R01HG005171 to D.E.H.; R01HG004853 to A.L.M.; R01HG004856 to R.R.; R01HG004877 to R.R.S. and R.F.; R01HG005172 to P. Spicer.; R01HG004857 to M.P.; R01HG004906 to T.M.S.; R21HG005811 to E.A.V.; M.J.B. was supported by UH2AR057506; G.A.B. was supported by UH2AI083263 and UH3AI083263 (G.A.B., C. N. Cornelissen, L. K. Eaves and J. F. Strauss); S.M.H. was supported by UH3DK083993 (V. B. Young, E. B. Chang, F. Meyer, T. M. S., M. L. Sogin, J. M. Tiedje); K.P.R. was supported by UH2DK083990 (J. V.); J.A.S. and H.H.K. were supported by UH2AR057504 and UH3AR057504 (J.A.S.); DP2OD001500 to K.M.A.; N01HG62088 to the Coriell Institute for Medical Research; U01DE016937 to F.E.D.; S.K.H. was supported by RC1DE0202098 and R01DE021574 (S.K.H. and H. Li); J.I. was supported by R21CA139193 (J.I. and D. S. Michaud); K.P.L. was supported by P30DE020751 (D. J. Smith); Army Research Office grant W911NF-11-1-0473 to C.H.; National Science Foundation grants NSF DBI-1053486 to C.H. and NSF IIS-0812111 to M.P.; The Office of Science of the US Department of Energy under Contract No. DE-AC02-05CH11231 for P.S. C.; LANL Laboratory-Directed Research and Development grant 20100034DR and the US Defense Threat Reduction Agency grants B104153I and B084531I to P.S.C.; Research Foundation - Flanders (FWO) grant to K.F. and J.Raes; R.K. is an HHMI Early Career Scientist; Gordon&BettyMoore Foundation funding and institutional funding fromthe J. David Gladstone Institutes to K.S.P.; A.M.S. was supported by fellowships provided by the Rackham Graduate School and the NIH Molecular Mechanisms in Microbial Pathogenesis Training Grant T32AI007528; a Crohn’s and Colitis Foundation of Canada Grant in Aid of Research to E.A.V.; 2010 IBM Faculty Award to K.C.W.; analysis of the HMPdata was performed using National Energy Research Scientific Computing resources, the BluBioU Computational Resource at Rice University

A multimodal cell census and atlas of the mammalian primary motor cortex

ABSTRACT We report the generation of a multimodal cell census and atlas of the mammalian primary motor cortex (MOp or M1) as the initial product of the BRAIN Initiative Cell Census Network (BICCN). This was achieved by coordinated large-scale analyses of single-cell transcriptomes, chromatin accessibility, DNA methylomes, spatially resolved single-cell transcriptomes, morphological and electrophysiological properties, and cellular resolution input-output mapping, integrated through cross-modal computational analysis. Together, our results advance the collective knowledge and understanding of brain cell type organization: First, our study reveals a unified molecular genetic landscape of cortical cell types that congruently integrates their transcriptome, open chromatin and DNA methylation maps. Second, cross-species analysis achieves a unified taxonomy of transcriptomic types and their hierarchical organization that are conserved from mouse to marmoset and human. Third, cross-modal analysis provides compelling evidence for the epigenomic, transcriptomic, and gene regulatory basis of neuronal phenotypes such as their physiological and anatomical properties, demonstrating the biological validity and genomic underpinning of neuron types and subtypes. Fourth, in situ single-cell transcriptomics provides a spatially-resolved cell type atlas of the motor cortex. Fifth, integrated transcriptomic, epigenomic and anatomical analyses reveal the correspondence between neural circuits and transcriptomic cell types. We further present an extensive genetic toolset for targeting and fate mapping glutamatergic projection neuron types toward linking their developmental trajectory to their circuit function. Together, our results establish a unified and mechanistic framework of neuronal cell type organization that integrates multi-layered molecular genetic and spatial information with multi-faceted phenotypic properties

The PmpD production profile along development.

<p><i>C</i>. <i>psittaci</i>-infected HeLa cells were fixed at 12, 18, 24, 32 and 48 hpi, and double-stained with chlamydial LPS-specific antibody (FITC-conjugated, green) and PmpD-specific antibody (Alexa Fluor 568-conjugated, red). At 12 and 18 hpi (A), only colored merged images are shown under normal and penicillin-induced persistence culture conditions. At 24 and 48 hpi (B), single channel images are shown in black and white (2 left-most columns), while merged images and insets thereof are shown in color (2 right-most columns). Staining patterns at 32 hpi (not shown) were similar to those at 48 hpi. Bar = 2 μm for the 12 and 18 hpi times, and 24 and 48 hpi insets (i.e. top row and right-most column) and 10 μm for all remaining images.</p

PmpA and PmpH localize to the chlamydial inner and outer membranes, to the inclusion membrane and to small putative outer membrane vesicles.

<p><i>C</i>. <i>psittaci</i> infected HeLa cells were fixed at late times (24 and 48 hpi) and stained with primary PmpA- or PmpH-specific antibody and secondary gold conjugated goat anti-guinea pig antibody. Localization of PmpA and PmpH in the chlamydial inner (IM) and outer (OM) membranes (top row, long arrows), in the inclusion membrane (middle row, short arrows) and in putative OMVs (bottom row, arrowheads) is shown as indicated. Bars = 0.1 μm.</p

PmpA, B, D and H production differs under normal <i>C</i>. <i>psittaci</i> and during penicillin-induced stress culture conditions.

<p><i>C</i>. <i>psittaci</i> infected HeLa cells were fixed at 12, 18, 24, 32 and 48 hpi, and double-stained with a chlamydial LPS-specific antibody and Pmp-specific antibody. For each Pmp subtype, the percentage of positive inclusions was determined by a macro based on co-localization of the two antigens. The results shown here did not take into account smaller inclusions that are formed at 24, 32 and 48 hpi. The data are expressed as box plots: the box represents the 25^th-75^th percentiles, the median is depicted by a bar across the box and the whiskers on each box represent minimum and maximum value. Outliers are depicted by dots. Statistically significant differences (P < 0.05) are indicated with an asterix.</p

Guinea pig polyclonal antibodies against PmpA, B, D and H are specific for their respective immunizing antigens.

<p>(A) The specificity of polyclonal antibodies raised against rPmpA (anti-PmpA), rPmpB (anti-PmpB), rPmpD (anti-PmpD) and rPmpH (anti-PmpH) was verified by immunoblotting using (A) partially purified recombinant PmpA, B, D, E1, G3 and H as well as (B) density gradient purified EBs of <i>C</i>. <i>psittaci</i> Cal10. The calculated molecular masses of recombinant PmpA, PmpB, PmpD, PmpE1, PmpH and PmpG3 are 92 kDa, 74 kDa, 95 kDa, 74 kDa, 88 kDa and 60 kDa, respectively. The observed molecular masses were 75 kDa, 74 kDa, 95 kDa, 70 kDa, 75 kDa and 60 kDa, respectively. (B) The calculated and observed molecular masses of the protein bands detected in EBs are shown.</p

<i>pmp</i> genes and gene products of <i>C</i>. <i>psittaci</i> Cal10.

<p><i>pmp</i> genes and gene products of <i>C</i>. <i>psittaci</i> Cal10.</p