Genome-Centric Dynamics Shape the Diversity of Oral Bacterial Populations.

Two major viewpoints have been put forward for how microbial populations change, differing in whether adaptation is driven principally by gene-centric or genome-centric processes. Longitudinal sampling at microbially relevant timescales, i.e., days to weeks, is critical for distinguishing these mechanisms. Because of its significance for both microbial ecology and human health and its accessibility and high level of curation, we used the oral microbiota to study bacterial intrapopulation genome dynamics. Metagenomes were generated by shotgun sequencing of total community DNA from the healthy tongues of 17 volunteers at four to seven time points obtained over intervals of days to weeks. We obtained 390 high-quality metagenome-assembled genomes (MAGs) defining population genomes from 55 genera. The vast majority of genes in each MAG were tightly linked over the 2-week sampling window, indicating that the majority of the population's genomes were temporally stable at the MAG level. MAG-defined populations were composed of up to 5 strains, as determined by single-nucleotide-variant frequencies. Although most were stable over time, individual strains carrying over 100 distinct genes that rose from low abundance to dominance in a population over a period of days were detected. These results indicate a genome-wide as opposed to a gene-level process of population change. We infer that genome-wide selection of ecotypes is the dominant mode of adaptation in the oral populations over short timescales. IMPORTANCE The oral microbiome represents a microbial community of critical relevance to human health. Recent studies have documented the diversity and dynamics of different bacteria to reveal a rich, stable ecosystem characterized by strain-level dynamics. However, bacterial populations and their genomes are neither monolithic nor static; their genomes are constantly evolving to lose, gain, or alter their functional potential. To better understand how microbial genomes change in complex communities, we used culture-independent approaches to reconstruct the genomes (MAGs) for bacterial populations that approximated different species, in 17 healthy donors' mouths over a 2-week window. Our results underscored the importance of strain-level dynamics, which agrees with and expands on the conclusions of previous research. Altogether, these observations reveal patterns of genomic dynamics among strains of oral bacteria occurring over a matter of days.

Exploring the protist microbiome: The diversity of bacterial communities associated with Arcella spp. (Tubulina: Amoebozoa).

Research on protist-bacteria interactions is increasingly relevant as these associations are now known to play important roles in ecosystem and human health. Free-living amoebae are abundant in all environments and are frequent hosts for bacterial endosymbionts including pathogenic bacteria. However, to date, only a small fraction of these symbionts have been identified, while the structure and composition of the total symbiotic bacterial communities still remains largely unknown. Here, we use the testate amoeba Arcella spp. as model organisms to investigate the specificity and diversity of Arcella-associated microbial communities. High-throughputamplicon sequencing from the V4 region of the 16S rRNA gene revealed high diversity in the bacterial communities associated with the wild Arcella spp. To investigate the specificity of the associated bacterial community with greater precision, we investigated the bacterial communities of two lab-cultured Arcella species, A. hemispherica and A. intermedia, grown in two different media types. Our results suggest that Arcella-bacteria associations are species-specific, and that the associated bacterial community of lab-cultured Arcella spp. remains distinct from that of the surrounding media. Further, each host Arcella species could be distinguished based on its bacterial composition. Our findings provide insight into the understanding of eukaryotic-bacterial symbiosis.

The curious consistency of carbon biosignatures over billions of years of Earth-life coevolution.

The oldest and most wide-ranging signal of biological activity (biosignature) on our planet is the carbon isotope composition of organic materials preserved in rocks. These biosignatures preserve the long-term evolution of the microorganism-hosted metabolic machinery responsible for producing deviations in the isotopic compositions of inorganic and organic carbon. Despite billions of years of ecosystem turnover, evolutionary innovation, organismic complexification, and geological events, the organic carbon that is a residuum of the global marine biosphere in the rock record tells an essentially static story. The ~25‰ mean deviation between inorganic and organic 13C/12C values has remained remarkably unchanged over >3.5 billion years. The bulk of this record is conventionally attributed to early-evolved, RuBisCO-mediated CO2 fixation that, in extant oxygenic phototrophs, produces comparable isotopic effects and dominates modern primary production. However, billions of years of environmental transition, for example, in the progressive oxygenation of the Earth's atmosphere, would be expected to have accompanied shifts in the predominant RuBisCO forms as well as enzyme-level adaptive responses in RuBisCO CO2-specificity. These factors would also be expected to result in preserved isotopic signatures deviating from those produced by extant RuBisCO in oxygenic phototrophs. Why does the bulk carbon isotope record not reflect these expected environmental transitions and evolutionary innovations? Here, we discuss this apparent discrepancy and highlight the need for greater quantitative understanding of carbon isotope fractionation behavior in extant metabolic pathways. We propose novel, laboratory-based approaches to reconstructing ancestral states of carbon metabolisms and associated enzymes that can constrain isotopic biosignature production in ancient biological systems. Together, these strategies are crucial for integrating the complementary toolsets of biological and geological sciences and for interpretation of the oldest record of life on Earth.

Metapangenomics of the oral microbiome provides insights into habitat adaptation and cultivar diversity.

BACKGROUND: The increasing availability of microbial genomes and environmental shotgun metagenomes provides unprecedented access to the genomic differences within related bacteria. The human oral microbiome with its diverse habitats and abundant, relatively well-characterized microbial inhabitants presents an opportunity to investigate bacterial population structures at an ecosystem scale. RESULTS: Here, we employ a metapangenomic approach that combines public genomes with Human Microbiome Project (HMP) metagenomes to study the diversity of microbial residents of three oral habitats: tongue dorsum, buccal mucosa, and supragingival plaque. For two exemplar taxa, Haemophilus parainfluenzae and the genus Rothia, metapangenomes reveal distinct genomic groups based on shared genome content. H. parainfluenzae genomes separate into three distinct subgroups with differential abundance between oral habitats. Functional enrichment analyses identify an operon encoding oxaloacetate decarboxylase as diagnostic for the tongue-abundant subgroup. For the genus Rothia, grouping by shared genome content recapitulates species-level taxonomy and habitat preferences. However, while most R. mucilaginosa are restricted to the tongue as expected, two genomes represent a cryptic population of R. mucilaginosa in many buccal mucosa samples. For both H. parainfluenzae and the genus Rothia, we identify not only limitations in the ability of cultivated organisms to represent populations in their native environment, but also specifically which cultivar gene sequences are absent or ubiquitous. CONCLUSIONS: Our findings provide insights into population structure and biogeography in the mouth and form specific hypotheses about habitat adaptation. These results illustrate the power of combining metagenomes and pangenomes to investigate the ecology and evolution of bacteria across analytical scales.

The saccharibacterium TM7x elicits differential responses across its host range.

Host range is a fundamental component of symbiotic interactions, yet it remains poorly characterized for the prevalent yet enigmatic subcategory of bacteria/bacteria symbioses. The recently characterized obligate bacterial epibiont Candidatus Nanosynbacter lyticus TM7x with its bacterial host Actinomyces odontolyticus XH001 offers an ideal system to study such a novel relationship. In this study, the host range of TM7x was investigated by coculturing TM7x with various related Actinomyces strains and characterizing their growth dynamics from initial infection through subsequent co-passages. Of the twenty-seven tested Actinomyces, thirteen strains, including XH001, could host TM7x, and further classified into "permissive" and "nonpermissive" based on their varying initial responses to TM7x. Ten permissive strains exhibited growth/crash/recovery phases following TM7x infection, with crash timing and extent dependent on initial TM7x dosage. Meanwhile, three nonpermissive strains hosted TM7x without a growth-crash phase despite high TM7x dosage. The physical association of TM7x with all hosts, including nonpermissive strains, was confirmed by microscopy. Comparative genomic analyses revealed distinguishing genomic features between permissive and nonpermissive hosts. Our results expand the concept of host range beyond a binary to a wider spectrum, and the varying susceptibility of Actinomyces strains to TM7x underscores how small genetic differences between hosts can underly divergent selective trajectories.

An inter-island comparison of Darwin's finches reveals the impact of habitat, host phylogeny, and island on the gut microbiome.

Darwin's finch species in the Galapagos Archipelago are an iconic adaptive radiation that offer a natural experiment to test for the various factors that influence gut microbiome composition. The island of Floreana has the longest history of human settlement within the archipelago and offers an opportunity to compare island and habitat effects on Darwin's finch microbiomes. In this study, we compare gut microbiomes in Darwin's finch species on Floreana Island to test for effects of host phylogeny, habitat (lowlands, highlands), and island (Floreana, Santa Cruz). We used 16S rRNA Illumina sequencing of fecal samples to assess the gut microbiome composition of Darwin's finches, complemented by analyses of stable isotope values and foraging data to provide ecological context to the patterns observed. Overall bacterial composition of the gut microbiome demonstrated co-phylogeny with Floreana hosts, recapitulated the effect of habitat and diet, and showed differences across islands. The finch phylogeny uniquely explained more variation in the microbiome than did foraging data. Finally, there were interaction effects for island × habitat, whereby the same Darwin's finch species sampled on two islands differed in microbiome for highland samples (highland finches also had different diets across islands) but not lowland samples (lowland finches across islands had comparable diet). Together, these results corroborate the influence of phylogeny, age, diet, and sampling location on microbiome composition and emphasize the necessity for comprehensive sampling given the multiple factors that influence the gut microbiome in Darwin's finches, and by extension, in animals broadly.

Host phylogeny, diet, and habitat differentiate the gut microbiomes of Darwin's finches on Santa Cruz Island.

Darwin's finches are an iconic example of an adaptive radiation with well-characterized evolutionary history, dietary preferences, and biogeography, offering an unparalleled opportunity to disentangle effects of evolutionary history on host microbiome from other factors like diet and habitat. Here, we characterize the gut microbiome in Darwin's finches, comparing nine species that occupy diverse ecological niches on Santa Cruz island. The finch phylogeny showed moderate congruence with the microbiome, which was comprised mostly of the bacterial phyla Firmicutes, Actinobacteria, and Proteobacteria. Diet, as measured with stable isotope values and foraging observations, also correlated with microbiome differentiation. Additionally, each gut microbial community could easily be classified by the habitat of origin independent of host species. Altogether, these findings are consistent with a model of microbiome assembly in which environmental filtering via diet and habitat are primary determinants of the bacterial taxa present with lesser influence from the evolutionary history between finch species.

Correction: Longitudinal microbiome profiling reveals impermanence of probiotic bacteria in domestic pigeons.

[This corrects the article DOI: 10.1371/journal.pone.0217804.].

Longitudinal microbiome profiling reveals impermanence of probiotic bacteria in domestic pigeons.

Probiotics are bacterial species or assemblages that are applied to animals and plants with the intention of altering the microbiome in a beneficial way. Probiotics have been linked to positive health effects such as faster disease recovery times in humans and increased weight gain in poultry. Pigeon fanciers often feed their show pigeons probiotics with the intention of increasing flight performance. The objective of our study was to determine the effect of two different probiotics, alone and in combination, on the fecal microbiome of Birmingham Roller pigeons. We sequenced fecal samples from 20 pigeons divided into three probiotic treatments, including prior to, during, and after treatment. Pre-treatment and control group samples were dominated by Actinobacteria, Firmicutes, Proteobacteria, and Cyanobacteria. Administration of a probiotic pellet containing Enterococcus faecium and Lactobacillus acidophilus resulted in increase in average relative abundance of Lactobacillus spp. from 4.7 ± 2.0% to 93.0 ± 5.3%. No significant effects of Enterococcus spp. were detected. Probiotic-induced shifts in the microbiome composition were temporary and disappeared within 2 days of probiotic cessation. Administration of a probiotic powder in drinking water that contained Enterococcus faecium and three Lactobacillus species had minimal effect on the microbiome. We conclude that supplementing Birmingham roller pigeons with the probiotic pellets, but not the probiotic powder, temporarily changed the microbiome composition. A next step is to experimentally test the effect of these changes in microbiome composition on host health and physical performance.

Isotope discrimination by form IC RubisCO from Ralstonia eutropha and Rhodobacter sphaeroides, metabolically versatile members of 'Proteobacteria' from aquatic and soil habitats.

RubisCO, the CO2 fixing enzyme of the Calvin-Benson-Bassham (CBB) cycle, is responsible for the majority of carbon fixation on Earth. RubisCO fixes 12 CO2 faster than 13 CO2 resulting in 13 C-depleted biomass, enabling the use of δ13 C values to trace CBB activity in contemporary and ancient environments. Enzymatic fractionation is expressed as an ε value, and is routinely used in modelling, for example, the global carbon cycle and climate change, and for interpreting trophic interactions. Although values for spinach RubisCO (ε = ~29‰) have routinely been used in such efforts, there are five different forms of RubisCO utilized by diverse photolithoautotrophs and chemolithoautotrophs and ε values, now known for four forms (IA, B, D and II), vary substantially with ε = 11‰ to 27‰. Given the importance of ε values in δ13 C evaluation, we measured enzymatic fractionation of the fifth form, form IC RubisCO, which is found widely in aquatic and terrestrial environments. Values were determined for two model organisms, the 'Proteobacteria' Ralstonia eutropha (ε = 19.0‰) and Rhodobacter sphaeroides (ε = 22.4‰). It is apparent from these measurements that all RubisCO forms measured to date discriminate less than commonly assumed based on spinach, and that enzyme ε values must be considered when interpreting and modelling variability of δ13 C values in nature.

Diverse Legionella-Like Bacteria Associated with Testate Amoebae of the Genus Arcella (Arcellinida: Amoebozoa).

Diverse species of Legionella and Legionella-like amoebal pathogens (LLAPs) have been identified as intracellular bacteria in many amoeboid protists. There are, however, other amoeboid groups such as testate amoeba for which we know little about their potential to host such bacteria. In this study, we assessed the occurrence and diversity of Legionella spp. in cultures and environmental isolates of freshwater arcellinid testate amoebae species, Arcella hemispherica, Arcella intermedia, and Arcella vulgaris, via 16S rRNA gene sequence analyses and fluorescent in situ hybridization (FISH). Analysis of the 16S rRNA gene sequences indicated that A. hemispherica, A. intermedia, and A. vulgaris host Legionella-like bacteria with 94-98% identity to other Legionella spp. based on NCBI BLAST search. Phylogenetic analysis placed Legionella-like Arcella-associated bacteria (LLAB) in three different clusters within a tree containing all other members of Legionella and LLAPs. The intracellular localization of the Legionella within Arcella hosts was confirmed using FISH with a Legionella-specific probe. This study demonstrates that the host range of Legionella and Legionella-like bacteria in the Amoebozoa extends beyond members of "naked" amoebae species, with members of the testate amoebae potentially serving an ecological role in the dispersal, protection, and replication of Legionella spp. in natural environments.

Intrahost Genetic Diversity of Bacterial Symbionts Exhibits Evidence of Mixed Infections and Recombinant Haplotypes.

Even the simplest microbial-eukaryotic mutualisms are comprised of entire populations of symbionts at the level of the host individual. Early work suggested that these intrahost populations maintain low genetic diversity as a result of transmission bottlenecks or to avoid competition between symbiont genotypes. However, the amount of genetic diversity among symbionts within a single host remains largely unexplored. To address this, we investigated the chemosynthetic symbiosis between the bivalve Solemya velum and its intracellular bacterial symbionts, which exhibits evidence of both vertical and horizontal transmission. Intrahost symbiont populations were sequenced to high coverage (200-1,000×). Analyses of nucleotide diversity revealed that the symbiont genome sequences were largely homogeneous within individual host specimens, consistent with vertical transmission, except for particular regions that were polymorphic in ∼20% of host specimens. These variant sites were also found segregating in other host individuals from the same population, colocalized to several regions of the genome, and consistently co-occurred on the same short read pairs (derived from the same chromosome). These results strongly suggest that these variant haplotypes originated through recombination events, potentially during prior mixed infections or in the external environment, rather than as novel mutations within symbiont populations. This abundant genetic diversity could have a profound influence on symbiont evolution as it provides the opportunity for selection to limit the extent of reductive genome evolution commonly seen in obligate intracellular bacteria and to enable the evolution of adaptive genotypes.

Mixed transmission modes and dynamic genome evolution in an obligate animal-bacterial symbiosis.

Reliable transmission of symbionts between host generations facilitates the evolution of beneficial and pathogenic associations. Although transmission mode is typically characterized as either vertical or horizontal, the prevalence of intermediate transmission modes, and their impact on symbiont genome evolution, are understudied. Here, we use population genomics to explore mixed transmission modes of chemosynthetic bacterial symbionts in the bivalve Solemya velum. Despite strong evidence for symbiont inheritance through host oocytes, whole-genome analyses revealed signatures of frequent horizontal transmission, including discordant mitochondrial-symbiont genealogies, widespread recombination and a dynamic symbiont genome structure consistent with evolutionary patterns of horizontally transmitted associations. Population-level analyses thus provide a tractable means of ascertaining the fidelity of vertical versus horizontal transmission. Our data support the strong influence horizontal transmission can have on symbiont genome evolution, and shed light on the dynamic evolutionary pressures shaping symbiotic bacterial genomes.

The genome of the intracellular bacterium of the coastal bivalve, Solemya velum: a blueprint for thriving in and out of symbiosis.

BACKGROUND: Symbioses between chemoautotrophic bacteria and marine invertebrates are rare examples of living systems that are virtually independent of photosynthetic primary production. These associations have evolved multiple times in marine habitats, such as deep-sea hydrothermal vents and reducing sediments, characterized by steep gradients of oxygen and reduced chemicals. Due to difficulties associated with maintaining these symbioses in the laboratory and culturing the symbiotic bacteria, studies of chemosynthetic symbioses rely heavily on culture independent methods. The symbiosis between the coastal bivalve, Solemya velum, and its intracellular symbiont is a model for chemosynthetic symbioses given its accessibility in intertidal environments and the ability to maintain it under laboratory conditions. To better understand this symbiosis, the genome of the S. velum endosymbiont was sequenced. RESULTS: Relative to the genomes of obligate symbiotic bacteria, which commonly undergo erosion and reduction, the S. velum symbiont genome was large (2.7 Mb), GC-rich (51%), and contained a large number (78) of mobile genetic elements. Comparative genomics identified sets of genes specific to the chemosynthetic lifestyle and necessary to sustain the symbiosis. In addition, a number of inferred metabolic pathways and cellular processes, including heterotrophy, branched electron transport, and motility, suggested that besides the ability to function as an endosymbiont, the bacterium may have the capacity to live outside the host. CONCLUSIONS: The physiological dexterity indicated by the genome substantially improves our understanding of the genetic and metabolic capabilities of the S. velum symbiont and the breadth of niches the partners may inhabit during their lifecycle.

Evidence for horizontal transmission from multilocus phylogeny of deep-sea mussel (Mytilidae) symbionts.

Many invertebrates at deep-sea hydrothermal vents depend upon bacterial symbionts for nutrition, and thus the mechanism of symbiont transmission, vertical (via the egg or sperm) or horizontal (from environment or contemporary hosts) is critically important. Under a strict maternal transmission model, symbiont and host mitochondrial genomes pass through the same individuals leading to congruent host-symbiont phylogenies. In contrast, horizontally transmitted symbionts are environmentally acquired, leading to incongruent host-symbiont phylogenies. Each of these transmission strategies is predicted to have different consequences for symbiont ecology and genome evolution. Deep-sea mussels (Bathymodiolinae) are globally distributed at deep-sea hydrothermal vents, depend upon chemoautotrophic symbionts for their survival, and are hypothesized to transmit their symbionts horizontally. This study explored bathymodioline symbiont ecology through quantification of symbionts at two hydrothermal vent sites and symbiont evolution using functional gene phylogenies. These phylogenies revealed a dramatically more complex evolutionary history than 16S ribosomal RNA phylogenies, suggesting that horizontal gene transfer may have played an important role in symbiont gene evolution. Tests of the strict maternal transmission hypothesis found that host-symbiont lineages were significantly decoupled across multiple genes. These findings expand our understanding of symbiont ecology and evolution, and provide the strongest evidence yet for horizontal transmission of bathymodioline symbionts.

Characterizing the plasticity of nitrogen metabolism by the host and symbionts of the hydrothermal vent chemoautotrophic symbioses Ridgeia piscesae.

Chemoautotrophic symbionts of deep sea hydrothermal vent tubeworms are known to provide their hosts with all their primary nutrition. While studies have examined how chemoautotrophic symbionts provide the association with nitrogen, fewer have examined if symbiont nitrogen metabolism varies as a function of environmental conditions. Ridgeia piscesae tubeworms flourish at Northeastern Pacific vents, occupy a range of microhabitats, and exhibit a high degree of morphological plasticity [e.g. long-skinny (LS) and short-fat (SF) phenotypes] that may relate to environmental conditions. This plasticity affords an opportunity to examine whether symbiont nitrogen metabolism varies among host phenotypes. LS and SF R. piscesae were recovered from the Axial and Main Endeavour Field hydrothermal vents. Nitrate and ammonium were quantified in Ridgeia blood, and the expression of key nitrogen metabolism genes, as well as stable nitrogen isotope ratios, was quantified in host branchial plume and symbiont-containing tissues. Nitrate and ammonium were abundant in the blood of both phenotypes though environmental ammonium concentrations were, paradoxically, lowest among individuals with the highest blood ammonium. Assimilatory nitrate reductase transcripts were always below detection, though in both LS and SF R. piscesae symbionts, we observed elevated expression of dissimilatory nitrate reductase genes, as well as symbiont and host ammonium assimilation genes. Site-specific differences in expression, along with tissue stable isotope analyses, suggest that LS and SF Ridgeia symbionts are engaged in both dissimilatory nitrate reduction and ammonia assimilation to varying degrees. As such, it appears that environmental conditions -not host phenotype-primarily dictates symbiont nitrogen metabolism.

The methanol dehydrogenase gene, mxaF, as a functional and phylogenetic marker for proteobacterial methanotrophs in natural environments.

The mxaF gene, coding for the large (α) subunit of methanol dehydrogenase, is highly conserved among distantly related methylotrophic species in the Alpha-, Beta- and Gammaproteobacteria. It is ubiquitous in methanotrophs, in contrast to other methanotroph-specific genes such as the pmoA and mmoX genes, which are absent in some methanotrophic proteobacterial genera. This study examined the potential for using the mxaF gene as a functional and phylogenetic marker for methanotrophs. mxaF and 16S rRNA gene phylogenies were constructed based on over 100 database sequences of known proteobacterial methanotrophs and other methylotrophs to assess their evolutionary histories. Topology tests revealed that mxaF and 16S rDNA genes of methanotrophs do not show congruent evolutionary histories, with incongruencies in methanotrophic taxa in the Methylococcaceae, Methylocystaceae, and Beijerinckiacea. However, known methanotrophs generally formed coherent clades based on mxaF gene sequences, allowing for phylogenetic discrimination of major taxa. This feature highlights the mxaF gene's usefulness as a biomarker in studying the molecular diversity of proteobacterial methanotrophs in nature. To verify this, PCR-directed assays targeting this gene were used to detect novel methanotrophs from diverse environments including soil, peatland, hydrothermal vent mussel tissues, and methanotroph isolates. The placement of the majority of environmental mxaF gene sequences in distinct methanotroph-specific clades (Methylocystaceae and Methylococcaceae) detected in this study supports the use of mxaF as a biomarker for methanotrophic proteobacteria.

Metatranscriptomic analysis of sulfur oxidation genes in the endosymbiont of solemya velum.

Thioautotrophic endosymbionts in the Domain Bacteria mediate key sulfur transformations in marine reducing environments. However, the molecular pathways underlying symbiont metabolism and the extent to which these pathways are expressed in situ are poorly characterized for almost all symbioses. This is largely due to the difficulty of culturing symbionts apart from their hosts. Here, we use pyrosequencing of community RNA transcripts (i.e., the metatranscriptome) to characterize enzymes of dissimilatory sulfur metabolism in the model symbiosis between the coastal bivalve Solemya velum and its intracellular thioautotrophic symbionts. High-throughput sequencing of total RNA from the symbiont-containing gill of a single host individual generated 1.6 million sequence reads (500 Mbp). Of these, 43,735 matched Bacteria protein-coding genes in BLASTX searches of the NCBI database. The taxonomic identities of the matched genes indicated relatedness to diverse species of sulfur-oxidizing Gammaproteobacteria, including other thioautotrophic symbionts and the purple sulfur bacterium Allochromatium vinosum. Manual querying of these data identified 28 genes from diverse pathways of sulfur energy metabolism, including the dissimilatory sulfite reductase (Dsr) pathway for sulfur oxidation to sulfite, the APS pathway for sulfite oxidation, and the Sox pathway for thiosulfate oxidation. In total, reads matching sulfur energy metabolism genes represented 7% of the Bacteria mRNA pool. Together, these data highlight the dominance of thioautotrophy in the context of symbiont community metabolism, identify the likely pathways mediating sulfur oxidation, and illustrate the utility of metatranscriptome sequencing for characterizing community gene transcription of uncultured symbionts.

Phylogenetic characterization of episymbiotic bacteria hosted by a hydrothermal vent limpet (lepetodrilidae, vetigastropoda).

Marine invertebrates hosting chemosynthetic bacterial symbionts are known from multiple phyla and represent remarkable diversity in form and function. The deep-sea hydrothermal vent limpet Lepetodrilus fucensis from the Juan de Fuca Ridge complex hosts a gill symbiosis of particular interest because it displays a morphology unique among molluscs: filamentous bacteria are found partially embedded in the host's gill epithelium and extend into the fluids circulating across the lamellae. Our objective was to investigate the phylogenetic affiliation of the limpet's primary gill symbionts for comparison with previously characterized bacteria. Comparative 16S rRNA sequence analysis identified one γ- and three ε-Proteobacteria as candidate symbionts. We used fluorescence in situ hybridization (FISH) to test which of these four candidates occur with the limpet's symbiotic gill bacteria. The γ-proteobacterial probes consistently hybridized to the entire area where symbiotic bacteria were found, but fluorescence signal from the ε-proteobacterial probes was generally absent. Amplification of the γ-proteobacterial 16S rRNA gene using a specific forward primer yielded a sequence similar to that of limpets collected from different ridge sections. In total, direct amplification or FISH identified a single γ-proteobacterial lineage from the gills of 23 specimens from vents separated by a distance up to about 200 km and collected over the course of 2 years, suggesting a highly specific and widespread symbiosis. Thus, we report the first filamentous γ-proteobacterial gill symbiont hosted by a mollusc.

Evidence for a core gut microbiota in the zebrafish.

Experimental analysis of gut microbial communities and their interactions with vertebrate hosts is conducted predominantly in domesticated animals that have been maintained in laboratory facilities for many generations. These animal models are useful for studying coevolved relationships between host and microbiota only if the microbial communities that occur in animals in lab facilities are representative of those that occur in nature. We performed 16S rRNA gene sequence-based comparisons of gut bacterial communities in zebrafish collected recently from their natural habitat and those reared for generations in lab facilities in different geographic locations. Patterns of gut microbiota structure in domesticated zebrafish varied across different lab facilities in correlation with historical connections between those facilities. However, gut microbiota membership in domesticated and recently caught zebrafish was strikingly similar, with a shared core gut microbiota. The zebrafish intestinal habitat therefore selects for specific bacterial taxa despite radical differences in host provenance and domestication status.