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Mobile Type VI secretion system loci of the gut Bacteroidales display extensive intra-ecosystem transfer, multi-species spread and geographical clustering.

ABSTRACT: The human gut microbiota is a dense microbial ecosystem with extensive opportunities for bacterial contact-dependent processes such as conjugation and Type VI secretion system (T6SS)-dependent antagonism. In the gut Bacteroidales, two distinct genetic architectures of T6SS loci, GA1 and GA2, are contained on Integrative and Conjugative Elements (ICE). Despite intense interest in the T6SSs of the gut Bacteroidales, there is only a superficial understanding of their evolutionary patterns, and of their dissemination among Bacteroidales species in human gut communities. Here, we combine extensive genomic and metagenomic analyses to better understand their ecological and evolutionary dynamics. We identify new genetic subtypes, document extensive intrapersonal transfer of these ICE to Bacteroidales species within human gut microbiomes, and most importantly, reveal frequent population fixation of these newly armed strains in multiple species within a person. We further show the distribution of each of the distinct T6SSs in human populations and show there is geographical clustering. We reveal that the GA1 T6SS ICE integrates at a minimal recombination site leading to their integration throughout genomes and their frequent interruption of genes, whereas the GA2 T6SS ICE integrate at one of three different tRNA genes. The exclusion of concurrent GA1 and GA2 T6SSs in individual strains is associated with intact T6SS loci and with an ICE-encoded gene. By performing a comprehensive analysis of mobile genetic elements (MGE) in co-resident Bacteroidales species in numerous human gut communities, we identify 74 MGE that transferred to multiple Bacteroidales species within individual gut microbiomes. We further show that only three other MGE demonstrate multi-species spread in human gut microbiomes to the degree demonstrated by the GA1 and GA2 ICE. These data underscore the ubiquity and dissemination of mobile T6SS loci within Bacteroidales communities and across human populations.

SUBMITTER: Garcia-Bayona L

PROVIDER: S-EPMC8102008 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Type VI secretion systems (T6SSs) are contact-dependent antagonistic systems employed by Gram negative bacteria to intoxicate other bacteria or eukaryotic cells. T6SSs were recently discovered in a few Bacteroidetes strains, thereby extending the presence of these systems beyond Proteobacteria. The present study was designed to analyze in a global nature the diversity, abundance, and properties of T6SSs in the Bacteroidales, the most predominant Gram negative bacterial order of the human gut.By performing extensive bioinformatics analyses and creating hidden Markov models for Bacteroidales Tss proteins, we identified 130 T6SS loci in 205 human gut Bacteroidales genomes. Of the 13 core T6SS proteins of Proteobacteria, human gut Bacteroidales T6SS loci encode orthologs of nine, and an additional five other core proteins not present in Proteobacterial T6SSs. The Bacteroidales T6SS loci segregate into three distinct genetic architectures with extensive DNA identity between loci of a given genetic architecture. We found that divergent DNA regions of a genetic architecture encode numerous types of effector and immunity proteins and likely include new classes of these proteins. TheT6SS loci of genetic architecture 1 are contained on highly similar integrative conjugative elements (ICEs), as are the T6SS loci of genetic architecture 2, whereas the T6SS loci of genetic architecture 3 are not and are confined to Bacteroides fragilis. Using collections of co-resident Bacteroidales strains from human subjects, we provide evidence for the transfer of genetic architecture 1 T6SS loci among co-resident Bacteroidales species in the human gut. However, we also found that established ecosystems can harbor strains with distinct T6SS of all genetic architectures.This is the first study to comprehensively analyze of the presence and diversity of T6SS loci within an order of bacteria and to analyze T6SSs of bacteria from a natural community. These studies demonstrate that more than half of our gut Bacteroidales, equivalent to about ¼ of the bacteria of this ecosystem, encode T6SSs. The data reveal several novel properties of these systems and suggest that antagonism between or distributed defense among these abundant intestinal bacteria may be common in established human gut communities.

Project description:UnlabelledThe genome sequences of intestinal Bacteroidales strains reveal evidence of extensive horizontal gene transfer. In vitro studies of Bacteroides and other bacteria have addressed mechanisms of conjugative transfer and some phenotypic outcomes of these DNA acquisitions in the recipient, such as the acquisition of antibiotic resistance. However, few studies have addressed the horizontal transfer of genetic elements between bacterial species coresident in natural microbial communities, especially microbial ecosystems of humans. Here, we examine the genomes of Bacteroidales species from two human adults to identify genetic elements that were likely transferred among these Bacteroidales while they were coresident in the intestine. Using seven coresident Bacteroidales species from one individual and eight from another, we identified five large chromosomal regions, each present in a minimum of three of the coresident strains at near 100% DNA identity. These five regions are not found in any other sequenced Bacteroidetes genome at this level of identity and are likely all integrative conjugative elements (ICEs). Such highly similar and unique regions occur in only 0.4% of phylogenetically representative mock communities, providing strong evidence that these five regions were transferred between coresident strains in these subjects. In addition to the requisite proteins necessary for transfer, these elements encode proteins predicted to increase fitness, including orphan DNA methylases that may alter gene expression, fimbriae synthesis proteins that may facilitate attachment and the utilization of new substrates, putative secreted antimicrobial molecules, and a predicted type VI secretion system (T6SS), which may confer a competitive ecological advantage to these strains in their complex microbial ecosystem.ImportanceBy analyzing Bacteroidales strains coresident in the gut microbiota of two human adults, we provide strong evidence for extensive interspecies and interfamily transfer of integrative conjugative elements within the intestinal microbiota of individual humans. In the recipient strain, we show that the conjugative elements themselves can be modified by the transposition of insertion sequences and retroelements from the recipient's genome, with subsequent transfer of these modified elements to other members of the microbiota. These data suggest that the genomes of our gut bacteria are substantially modified by other, coresident members of the ecosystem, resulting in highly personalized Bacteroidales strains likely unique to that individual. The genetic content of these ICEs suggests that their transfer from successful adapted members of an ecosystem confers beneficial properties to the recipient, increasing its fitness and allowing it to better compete within its particular personalized gut microbial ecosystem.

Project description:Three distinct genetic architectures (GAs) of Type VI secretion systems (T6SSs) have been described in gut Bacteroidales species, each with unique genes and characteristics. Unlike the GA3 T6SSs, potent antagonism has not yet been demonstrated for the GA1 or GA2 T6SSs. We previously showed that the GA2 T6SS loci are contained on integrative and conjugative elements and that there are five subtypes. Collectively, GA2 are the most prevalent Bacteroidales T6SSs in the human populations analyzed. In this study, we provide a comprehensive bioinformatic analysis of the three variable regions of GA2 T6SS loci, which encode toxic effector and immunity proteins. In total, we identified 63 distinct effectors encoded within 31 nonredundant GA2 loci, 18 of which do not have described motifs or predicted functions. We provide experimental evidence for toxin activity for four different GA2 effectors, showing that each functions only when present in the periplasm, and experimentally confirm their cognate immunity proteins. Our data demonstrate that each GA2 locus encodes at least three distinct effectors with targets in both the cytoplasm and the periplasm. The data also suggest that the effectors of a given locus are loaded onto the tube by different mechanisms, which may allow all three effectors encoded within a single GA2 locus with distinct antibacterial activity to be loaded onto a single T6 tube, increasing the antagonistic effect. IMPORTANCE Humans are colonized with many gut Bacteroidales species at high density, allowing for extensive opportunities for contact-dependent antagonism. To begin to understand the antagonistic potential of the GA2 T6SSs of the gut Bacteroidales, we performed bioinformatic and experimental analyses of the three divergent regions containing the toxin effector and immunity genes. We show that each GA2 T6SS locus encodes at least three distinct toxic effectors including toxins linked to Rhs and Hcp with cytoplasmic targets, and unlinked effectors with targets in the periplasm. The diversity and modality of effectors exceeds that of the GA1 or GA3 T6SS loci (M. J. Coyne, K. G. Roelofs, and L. E. Comstock, BMC Genomics 17:58, 2016, https://doi.org/10.1186/s12864-016-2377-z) and suggests that these T6SSs have the potential to be potent antibacterial weapons in the human gut.

Project description:Bacteroidales use type VI secretion systems (T6SS) to competitively colonize and persist in the colon. We identify a horizontally transferred T6SS with Ntox15 family nuclease effector (Tde1) that mediates interbacterial antagonism among Bacteroidales, including several derived from a single human donor. Expression of cognate (Tdi1) or orphan immunity proteins in acquired interbacterial defense systems protects against Tde1-dependent attack. We find that immunity protein interaction induces a large effector conformational change in Tde nucleases, disrupting the active site and altering the DNA-binding site. Crystallographic snapshots of isolated Tde1, the Tde1/Tdi1 complex, and homologs from Phocaeicola vulgatus (Tde2/Tdi2) illustrate a conserved mechanism of immunity inserting into the central core of Tde, splitting the nuclease fold into two subdomains. The Tde/Tdi interface and immunity mechanism are distinct from all other polymorphic toxin-immunity interactions of known structure. Bacteroidales abundance has been linked to inflammatory bowel disease activity in prior studies, and we demonstrate that Tde and T6SS structural genes are each enriched in fecal metagenomes from ulcerative colitis subjects. Genetically mobile Tde1-encoding T6SS in Bacteroidales mediate competitive growth and may be involved in inflammatory bowel disease. Broad immunity is conferred by Tdi1 homologs through a fold-disrupting mechanism unique among polymorphic effector-immunity pairs of known structure. IMPORTANCE Bacteroidales are related to inflammatory bowel disease severity and progression. We identify type VI secretion system (T6SS) nuclease effectors (Tde) which are enriched in ulcerative colitis and horizontally transferred on mobile genetic elements. Tde-encoding T6SSs mediate interbacterial competition. Orphan and cognate immunity proteins (Tdi) prevent intoxication by multiple Tde through a new mechanism among polymorphic toxin systems. Tdi inserts into the effector central core, splitting Ntox15 into two subdomains and disrupting the active site. This mechanism may allow for evolutionary diversification of the Tde/Tdi interface as observed in colicin nuclease-immunity interactions, promoting broad neutralization of Tde by orphan Tdi. Tde-dependent T6SS interbacterial antagonism may contribute to Bacteroidales diversity in the context of ulcerative colitis.

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Mobile Type VI secretion system loci of the gut Bacteroidales display extensive intra-ecosystem transfer, multi-species spread and geographical clustering.

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