Metagenomics is expanding our knowledge of the gene content, functional significance, and genetic variability in natural microbial communities. Still, there exists limited information concerning the regulation and dynamics of genes in the environment. We report here global analysis of expressed genes in a naturally occurring microbial community. We first adapted RNA amplification technologies to produce large amounts of cDNA from small quantities of total microbial community RNA. The fidelity of the RNA amplification procedure was validated with Prochlorococcus cultures and then applied to a microbial assemblage collected in the oligotrophic Pacific Ocean. Microbial community cDNAs were analyzed by pyrosequencing and compared with microbial community genomic DNA sequences determined from the same sample. Pyrosequencingbased estimates of microbial community gene expression compared favorably to independent assessments of individual gene expression using quantitative PCR. Genes associated with key metabolic pathways in open ocean microbial species-including genes involved in photosynthesis, carbon fixation, and nitrogen acquisition-and a number of genes encoding hypothetical proteins were highly represented in the cDNA pool. Genes present in the variable regions of Prochlorococcus genomes were among the most highly expressed, suggesting these encode proteins central to cellular processes in specific genotypes. Although many transcripts detected were highly similar to genes previously detected in ocean metagenomic surveys, a significant fraction (Ϸ50%) were unique. Thus, microbial community transcriptomic analyses revealed not only indigenous gene-and taxon-specific expression patterns but also gene categories undetected in previous DNA-based metagenomic surveys.bacterial communities ͉ metagenomics ͉ metatranscriptomics ͉ marine ͉ cDNA

Distinct partitioning has been observed in the composition and diversity of bacterial communities inhabiting the surface and overlying seawater of three coral species infected with black band disease (BBD) on the southern Caribbean island of Curaçao, Netherlands Antilles. PCR amplification and sequencing of bacterial 16S rRNA genes (rDNA) with universally conserved primers have identified over 524 unique bacterial sequences affiliated with 12 bacterial divisions. The molecular sequences exhibited less than 5% similarity in bacterial community composition between seawater and the healthy, black band diseased, and dead coral surfaces. The BBD bacterial mat rapidly migrates across and kills the coral tissue. Clone libraries constructed from the BBD mat were comprised of eight bacterial divisions and 13% unknowns. Several sequences representing bacteria previously found in other marine and terrestrial organisms (including humans) were isolated from the infected coral surfaces, including Clostridium spp., Arcobacter spp., Campylobacter spp., Cytophaga fermentans, Cytophaga columnaris, and Trichodesmium tenue.Infectious disease in scleractinian corals has emerged as one of the primary causes of the accelerating global destruction of coral reef ecosystems (22,25,27,56,74). Black band disease (BBD) is one of the most widespread and destructive of these coral infections (see review in reference 50). The diagnostic symptom of BBD is the development of a narrow 0.1-to 7-cmwide ring-shaped black to red microbial mat that migrates from top to bottom across massive coral colonies, killing healthy coral tissue at rates of as much as 1 cm per day (47, 53). BBD preferentially affects corals such as Montastrea annularis, Montastrea cavernosa, and Diploria strigosa (6,15,53). These species, known as framework building corals, form large structures that become the dominant physical elements of reefs. As a result, coral mortality caused by BBD is a potent force in restructuring coral reef ecosystems (15,36).There is considerable controversy as to whether BBD is caused by physical and chemical environmental stresses or is an infectious disease or both (50, 56). However, an impediment to determining the cause of BBD has been the lack of information about the diversity and distribution of microbial populations that inhabit normal healthy coral tissue and the BBD bacterial mat. It is known from studies of infectious disease in marine and terrestrial invertebrates, fish, and mammals (including humans) that pathogens are most effectively studied within an ecological context of interactions among microbes, their hosts, and the environmental conditions in which they live (25, 54). Accurate diagnosis and eventual treatment and prevention of BBD will therefore require a basic knowledge of the composition and distribution of the microbial communities associated with healthy as well as diseased organisms. This type of community-based comparative analysis of the microorganisms associated with infectious diseases in corals has not previously been...

BackgroundPeriodontitis is a polymicrobial biofilm-induced inflammatory disease that affects 743 million people worldwide. The current model to explain periodontitis progression proposes that changes in the relative abundance of members of the oral microbiome lead to dysbiosis in the host-microbiome crosstalk and then to inflammation and bone loss. Using combined metagenome/metatranscriptome analysis of the subgingival microbiome in progressing and non-progressing sites, we have characterized the distinct molecular signatures of periodontitis progression.MethodsMetatranscriptome analysis was conducted on samples from subgingival biofilms from progressing and stable sites from periodontitis patients. Community-wide expression profiles were obtained using Next Generation Sequencing (Illumina). Sequences were aligned using ‘bowtie2’ against a constructed oral microbiome database. Differential expression analysis was performed using the non-parametric algorithm implemented on the R package ‘NOISeqBio’. We summarized global functional activities of the oral microbial community by set enrichment analysis based on the Gene Ontology (GO) orthology.ResultsGene ontology enrichment analysis showed an over-representation in the baseline of active sites of terms related to cell motility, lipid A and peptidoglycan biosynthesis, and transport of iron, potassium, and amino acids. Periodontal pathogens (Tannerella forsythia and Porphyromonas gingivalis) upregulated different TonB-dependent receptors, peptidases, proteases, aerotolerance genes, iron transport genes, hemolysins, and CRISPR-associated genes. Surprisingly, organisms that have not been usually associated with the disease (Streptococcus oralis, Streptococcus mutans, Streptococcus intermedius, Streptococcus mitis, Veillonella parvula, and Pseudomonas fluorenscens) were highly active transcribing putative virulence factors. We detected patterns of activities associated with progression of clinical traits. Among those we found that the profiles of expression of cobalamin biosynthesis, proteolysis, and potassium transport were associated with the evolution towards disease.ConclusionsWe identified metabolic changes in the microbial community associated with the initial stages of dysbiosis. Regardless of the overall composition of the community, certain metabolic signatures are consistent with disease progression. Our results suggest that the whole community, and not just a handful of oral pathogens, is responsible for an increase in virulence that leads to progression.Trial registrationNCT01489839, 6 December 2011.Electronic supplementary materialThe online version of this article (doi:10.1186/s13073-015-0153-3) contains supplementary material, which is available to authorized users.

To better understand the temporal and spatial dynamics of Prochlorococcus populations, and how these populations co-vary with the physical environment, we followed monthly changes in the abundance of five ecotypes-two high-light adapted and three low-light adapted-over a 5-year period in coordination with the Bermuda Atlantic Time Series (BATS) and Hawaii Ocean Time-series (HOT) programs. Ecotype abundance displayed weak seasonal fluctuations at HOT and strong seasonal fluctuations at BATS. Furthermore, stable 'layered' depth distributions, where different Prochlorococcus ecotypes reached maximum abundance at different depths, were maintained consistently for 5 years at HOT. Layered distributions were also observed at BATS, although winter deep mixing events disrupted these patterns each year and produced large variations in ecotype abundance. Interestingly, the layered ecotype distributions were regularly reestablished each year after deep mixing subsided at BATS. In addition, Prochlorococcus ecotypes each responded differently to the strong seasonal changes in light, temperature and mixing at BATS, resulting in a reproducible annual succession of ecotype blooms. Patterns of ecotype abundance, in combination with physiological assays of cultured isolates, confirmed that the low-light adapted eNATL could be distinguished from other low-light adapted ecotypes based on its ability to withstand temporary exposure to high-intensity light, a characteristic stress of the surface mixed layer. Finally, total Prochlorococcus and Synechococcus dynamics were compared with similar time series data collected a decade earlier at each location. The two data sets were remarkably similar-testimony to the resilience of these complex dynamic systems on decadal time scales.

Despite increasing knowledge on phylogenetic composition of the human microbiome, our understanding of the in situ activities of the organisms in the community and their interactions with each other and with the environment remains limited. Characterizing gene expression profiles of the human microbiome is essential for linking the role of different members of the bacterial communities in health and disease. The oral microbiome is one of the most complex microbial communities in the human body and under certain circumstances, not completely understood, the healthy microbial community undergoes a transformation toward a pathogenic state that gives rise to periodontitis, a polymicrobial inflammatory disease. We report here the in situ genome-wide transcriptome of the subgingival microbiome in six periodontally healthy individuals and seven individuals with periodontitis. The overall picture of metabolic activities showed that iron acquisition, lipopolysaccharide synthesis and flagellar synthesis were major activities defining disease. Unexpectedly, the vast majority of virulence factors upregulated in subjects with periodontitis came from organisms that are not considered major periodontal pathogens. One of the organisms whose gene expression profile was characterized was the uncultured candidate division TM7, showing an upregulation of putative virulence factors in the diseased community. These data enhance understanding of the core activities that are characteristic of periodontal disease as well as the role that individual organisms in the subgingival community play in periodontitis.