Tip extension and simultaneous multiple fission in a filamentous bacterium.

Organisms display an immense variety of shapes, sizes, and reproductive strategies. At microscopic scales, bacterial cell morphology and growth dynamics are adaptive traits that influence the spatial organization of microbial communities. In one such community-the human dental plaque biofilm-a network of filamentous Corynebacterium matruchotii cells forms the core of bacterial consortia known as hedgehogs, but the processes that generate these structures are unclear. Here, using live-cell time-lapse microscopy and fluorescent D-amino acids to track peptidoglycan biosynthesis, we report an extraordinary example of simultaneous multiple division within the domain Bacteria. We show that C. matruchotii cells elongate at one pole through tip extension, similar to the growth strategy of soil-dwelling Streptomyces bacteria. Filaments elongate rapidly, at rates more than five times greater than other closely related bacterial species. Following elongation, many septa form simultaneously, and each cell divides into 3 to 14 daughter cells, depending on the length of the mother filament. The daughter cells then nucleate outgrowth of new thinner vegetative filaments, generating the classic "whip handle" morphology of this taxon. Our results expand the known diversity of bacterial cell cycles and help explain how this filamentous bacterium can compete for space, access nutrients, and form important interspecies interactions within dental plaque.

Spatial ecology of Haemophilus and Aggregatibacter in the human oral cavity.

Haemophilus and Aggregatibacter are two of the most common bacterial genera in the human oral cavity, encompassing both commensals and pathogens of substantial ecological and medical significance. In this study, we conducted a metapangenomic analysis of oral Haemophilus and Aggregatibacter species to uncover genomic diversity, phylogenetic relationships, and habitat specialization within the human oral cavity. Using three metrics-pangenomic gene content, phylogenomics, and average nucleotide identity (ANI)-we first identified distinct species and sub-species groups among these genera. Mapping of metagenomic reads then revealed clear patterns of habitat specialization, such as Aggregatibacter species predominantly in dental plaque, a distinctive Haemophilus parainfluenzae sub-species group on the tongue dorsum, and H. sp. HMT-036 predominantly in keratinized gingiva and buccal mucosa. In addition, we found that supragingival plaque samples contained predominantly only one out of the three taxa, H. parainfluenzae, Aggregatibacter aphrophilus, and A. sp. HMT-458, suggesting independent niches or a competitive relationship. Functional analyses revealed the presence of key metabolic genes, such as oxaloacetate decarboxylase, correlated with habitat specialization, suggesting metabolic versatility as a driving force. Additionally, heme synthesis distinguishes H. sp. HMT-036 from closely related Haemophilus haemolyticus, suggesting that the availability of micronutrients, particularly iron, was important in the evolutionary ecology of these species. Overall, our study exemplifies the power of metapangenomics to identify factors that may affect ecological interactions within microbial communities, including genomic diversity, habitat specialization, and metabolic versatility. IMPORTANCE: Understanding the microbial ecology of the mouth is essential for comprehending human physiology. This study employs metapangenomics to reveal that various Haemophilus and Aggregatibacter species exhibit distinct ecological preferences within the oral cavity of healthy individuals, thereby supporting the site-specialist hypothesis. Additionally, it was observed that the gene pool of different Haemophilus species correlates with their ecological niches. These findings shed light on the significance of key metabolic functions in shaping microbial distribution patterns and interspecies interactions in the oral ecosystem.

The oral microbiome: diversity, biogeography and human health.

The human oral microbiota is highly diverse and has a complex ecology, comprising bacteria, microeukaryotes, archaea and viruses. These communities have elaborate and highly structured biogeography that shapes metabolic exchange on a local scale and results from the diverse microenvironments present in the oral cavity. The oral microbiota also interfaces with the immune system of the human host and has an important role in not only the health of the oral cavity but also systemic health. In this Review, we highlight recent advances including novel insights into the biogeography of several oral niches at the species level, as well as the ecological role of candidate phyla radiation bacteria and non-bacterial members of the oral microbiome. In addition, we summarize the relationship between the oral microbiota and the pathology of oral diseases and systemic diseases. Together, these advances move the field towards a more holistic understanding of the oral microbiota and its role in health, which in turn opens the door to the study of novel preventive and therapeutic strategies.

Phages are unrecognized players in the ecology of the oral pathogen Porphyromonas gingivalis.

BACKGROUND: Porphyromonas gingivalis (hereafter "Pg") is an oral pathogen that has been hypothesized to act as a keystone driver of inflammation and periodontal disease. Although Pg is most readily recovered from individuals with actively progressing periodontal disease, healthy individuals and those with stable non-progressing disease are also colonized by Pg. Insights into the factors shaping the striking strain-level variation in Pg, and its variable associations with disease, are needed to achieve a more mechanistic understanding of periodontal disease and its progression. One of the key forces often shaping strain-level diversity in microbial communities is infection of bacteria by their viral (phage) predators and symbionts. Surprisingly, although Pg has been the subject of study for over 40 years, essentially nothing is known of its phages, and the prevailing paradigm is that phages are not important in the ecology of Pg. RESULTS: Here we systematically addressed the question of whether Pg are infected by phages-and we found that they are. We found that prophages are common in Pg, they are genomically diverse, and they encode genes that have the potential to alter Pg physiology and interactions. We found that phages represent unrecognized targets of the prevalent CRISPR-Cas defense systems in Pg, and that Pg strains encode numerous additional mechanistically diverse candidate anti-phage defense systems. We also found that phages and candidate anti-phage defense system elements together are major contributors to strain-level diversity and the species pangenome of this oral pathogen. Finally, we demonstrate that prophages harbored by a model Pg strain are active in culture, producing extracellular viral particles in broth cultures. CONCLUSION: This work definitively establishes that phages are a major unrecognized force shaping the ecology and intra-species strain-level diversity of the well-studied oral pathogen Pg. The foundational phage sequence datasets and model systems that we establish here add to the rich context of all that is already known about Pg, and point to numerous avenues of future inquiry that promise to shed new light on fundamental features of phage impacts on human health and disease broadly. Video Abstract.

Site-specialization of human oral Gemella species.

Gemella species are core members of the human oral microbiome in healthy subjects and are regarded as commensals, although they can cause opportunistic infections. Our objective was to evaluate the site-specialization of Gemella species among various habitats within the mouth by combining pangenomics and metagenomics. With pangenomics, we identified genome relationships and categorized genes as core and accessory to each species. With metagenomics, we identified the primary oral habitat of individual genomes. Our results establish that the genomes of three species, G. haemolysans, G. sanguinis and G. morbillorum, are abundant and prevalent in human mouths at different oral sites: G. haemolysans on buccal mucosa and keratinized gingiva; G. sanguinis on tongue dorsum, throat, and tonsils; and G. morbillorum in dental plaque. The gene-level basis of site-specificity was investigated by identifying genes that were core to Gemella genomes at a specific oral site but absent from other Gemella genomes. The riboflavin biosynthesis pathway was present in G. haemolysans genomes associated with buccal mucosa but absent from the rest of the genomes. Overall, metapangenomics show that Gemella species have clear ecological preferences in the oral cavity of healthy humans and provides an approach to identifying gene-level drivers of site specificity.

Mucin glycans drive oral microbial community composition and function.

Human microbiome composition is closely tied to health, but how the host manages its microbial inhabitants remains unclear. One important, but understudied, factor is the natural host environment: mucus, which contains gel-forming glycoproteins (mucins) that display hundreds of glycan structures with potential regulatory function. Leveraging a tractable culture-based system to study how mucins influence oral microbial communities, we found that mucin glycans enable the coexistence of diverse microbes, while resisting disease-associated compositional shifts. Mucins from tissues with unique glycosylation differentially tuned microbial composition, as did isolated mucin glycan libraries, uncovering the importance of specific glycan patterns in microbiome modulation. We found that mucins shape microbial communities in several ways: serving as nutrients to support metabolic diversity, organizing spatial structure through reduced aggregation, and possibly limiting antagonism between competing taxa. Overall, this work identifies mucin glycans as a natural host mechanism and potential therapeutic intervention to maintain healthy microbial communities.

Site Specialization of Human Oral Veillonella Species.

Veillonella species are abundant members of the human oral microbiome with multiple interspecies commensal relationships. Examining the distribution patterns of Veillonella species across the oral cavity is fundamental to understanding their oral ecology. In this study, we used a combination of pangenomic analysis and oral metagenomic information to clarify Veillonella taxonomy and to test the site specialist hypothesis for the Veillonella genus, which contends that most oral bacterial species are adapted to live at specific oral sites. Using isolate genome sequences combined with shotgun metagenomic sequence data, we showed that Veillonella species have clear, differential site specificity: Veillonella parvula showed strong preference for supra- and subgingival plaque, while closely related V. dispar, as well as more distantly related V. atypica, preferred the tongue dorsum, tonsils, throat, and hard palate. In addition, the provisionally named Veillonella sp. Human Microbial Taxon 780 showed strong site specificity for keratinized gingiva. Using comparative genomic analysis, we identified genes associated with thiamine biosynthesis and the reductive pentose phosphate cycle that may enable Veillonella species to occupy their respective habitats. IMPORTANCE Understanding the microbial ecology of the mouth is fundamental for understanding human physiology. In this study, metapangenomics demonstrated that different Veillonella species have clear ecological preferences in the oral cavity of healthy humans, validating the site specialist hypothesis. Furthermore, the gene pool of different Veillonella species was found to be reflective of their ecology, illuminating the potential role of vitamins and carbohydrates in determining Veillonella distribution patterns and interspecies interactions.

Intelectin-1 binds and alters the localization of the mucus barrier-modifying bacterium Akkermansia muciniphila.

Intelectin-1 (ITLN1) is a lectin secreted by intestinal epithelial cells (IECs) and upregulated in human ulcerative colitis (UC). We investigated how ITLN1 production is regulated in IECs and the biological effects of ITLN1 at the host-microbiota interface using mouse models. Our data show that ITLN1 upregulation in IECs from UC patients is a consequence of activating the unfolded protein response. Analysis of microbes coated by ITLN1 in vivo revealed a restricted subset of microorganisms, including the mucolytic bacterium Akkermansia muciniphila. Mice overexpressing intestinal ITLN1 exhibited decreased inner colonic mucus layer thickness and closer apposition of A. muciniphila to the epithelial cell surface, similar to alterations reported in UC. The changes in the inner mucus layer were microbiota and A. muciniphila dependent and associated with enhanced sensitivity to chemically induced and T cell-mediated colitis. We conclude that by determining the localization of a select group of bacteria to the mucus layer, ITLN1 modifies this critical barrier. Together, these findings may explain the impact of ITLN1 dysregulation on UC pathogenesis.

Corncob structures in dental plaque reveal microhabitat taxon specificity.

BACKGROUND: The human mouth is a natural laboratory for studying how bacterial communities differ across habitats. Different bacteria colonize different surfaces in the mouth-teeth, tongue dorsum, and keratinized and non-keratinized epithelia-despite the short physical distance between these habitats and their connection through saliva. We sought to determine whether more tightly defined microhabitats might have more tightly defined sets of resident bacteria. A microhabitat may be characterized, for example, as the space adjacent to a particular species of bacterium. Corncob structures of dental plaque, consisting of coccoid bacteria bound to filaments of Corynebacterium cells, present an opportunity to analyze the community structure of one such well-defined microhabitat within a complex natural biofilm. Here, we investigate by fluorescence in situ hybridization and spectral imaging the composition of the cocci decorating the filaments. RESULTS: The range of taxa observed in corncobs was limited to a small subset of the taxa present in dental plaque. Among four major groups of dental plaque streptococci, two were the major constituents of corncobs, including one that was the most abundant Streptococcus species in corncobs despite being relatively rare in dental plaque overall. Images showed both Streptococcus types in corncobs in all individual donors, suggesting that the taxa have different ecological roles or that mechanisms exist for stabilizing the persistence of functionally redundant taxa in the population. Direct taxon-taxon interactions were observed not only between the Streptococcus cells and the central corncob filament but also between Streptococcus cells and the limited subset of other plaque bacteria detected in the corncobs, indicating species ensembles involving these taxa as well. CONCLUSIONS: The spatial organization we observed in corncobs suggests that each of the microbial participants can interact with multiple, albeit limited, potential partners, a feature that may encourage the long-term stability of the community. Additionally, our results suggest the general principle that a precisely defined microhabitat will be inhabited by a small and well-defined set of microbial taxa. Thus, our results are important for understanding the structure and organizing principles of natural biofilms and lay the groundwork for future work to modulate and control biofilms for human health. Video Abstract.

Site-tropism of streptococci in the oral microbiome.

A detailed understanding of where bacteria localize is necessary to advance microbial ecology and microbiome-based therapeutics. The site-specialist hypothesis predicts that most microbes in the human oral cavity have a primary habitat type within the mouth where they are most abundant. We asked whether this hypothesis accurately describes the distribution of the members of the genus Streptococcus, a clinically relevant taxon that dominates most oral sites. Prior analysis of 16S rRNA gene sequencing data indicated that some oral Streptococcus clades are site-specialists while others may be generalists. However, within complex microbial populations composed of numerous closely related species and strains, such as the oral streptococci, genome-scale analysis is necessary to provide the resolution to discriminate closely related taxa with distinct functional roles. Here, we assess whether individual species within this genus are specialists using publicly available genomic sequence data that provide species-level resolution. We chose a set of high-quality representative genomes for human oral Streptococcus species. Onto these genomes, we mapped shotgun metagenomic sequencing reads from supragingival plaque, tongue dorsum, and other sites in the oral cavity. We found that every abundant Streptococcus species in the healthy human oral cavity showed strong site-tropism and that even closely related species such as S. mitis, S. oralis, and S. infantis specialized in different sites. These findings indicate that closely related bacteria can have distinct habitat distributions in the absence of dispersal limitation and under similar environmental conditions and immune regimes. Substantial overlap between the core genes of these three species suggests that site-specialization is determined by subtle differences in genomic content.