Mark Welch
Jessica L.
Mark Welch
Jessica L.
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PreprintMassive multiplication of genome and ribosomes in dormant cells (akinetes) of Aphanizomenon ovalisporum (Cyanobacteria)( 2011-08) Sukenik, Assaf ; Kaplan-Levy, Ruth N. ; Mark Welch, Jessica L. ; Post, Anton F.Akinetes are dormancy cells commonly found among filamentous cyanobacteria, many of which are toxic and/or nuisance, bloom-forming species. Development of akinetes from vegetative cells is a process that involves morphological and biochemical modifications. Here we applied a single cell approach to quantify genome and ribosome content of akinetes and vegetative cells in Aphanizomenon ovalisporum (Cyanobacteria). Vegetative cells of A. ovalisporum were naturally polyploid and contained on average 8 genome copies per cell. However, the chromosomal content of akinetes increased up to 450 copies, with an average value of 119 genome copies per akinete, 15 fold higher that in vegetative cells. Based on fluorescence in situ hybridization with a probe targeting 16S rRNA and detection with confocal laser scanning microscopy we conclude that ribosomes accumulated in akinetes to a higher level than that found in vegetative cells. We further present evidence that this massive accumulation of nucleic acids in akinetes is likely supported by phosphate supplied from inorganic polyphosphate bodies that were abundantly present in vegetative cells, but notably absent from akinetes. These results are interpreted in the context of cellular investments for proliferation following long term dormancy, as the high nucleic acid content would provide the basis for extended survival, rapid resumption of metabolic activity and cell division upon germination.
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ArticleIndividuality, stability, and variability of the plaque microbiome(Frontiers Media, 2016-04-22) Utter, Daniel R. ; Mark Welch, Jessica L. ; Borisy, Gary G.Dental plaque is a bacterial biofilm composed of a characteristic set of organisms. Relatively little information from cultivation-independent, high-throughput analyses has been published on the temporal dynamics of the dental plaque microbiome. We used Minimum Entropy Decomposition, an information theory-based approach similar to oligotyping that provides single-nucleotide resolution, to analyze a previously published time series data set and investigate the dynamics of the plaque microbiome at various analytic and taxonomic levels. At both the genus and 97% Operational Taxonomic Unit (OTU) levels of resolution, the range of variation within each individual overlapped that of other individuals in the data set. When analyzed at the oligotype level, however, the overlap largely disappeared, showing that single-nucleotide resolution enables differentiation of individuals from one another without ambiguity. The overwhelming majority of the plaque community in all samples was made up of bacteria from a moderate number of plaque-typical genera, indicating that the overall community framework is shared among individuals. Each of these genera fluctuated in abundance around a stable mean that varied between individuals, with some genera having higher inter-individual variability than others. Thus, at the genus level, differences between individuals lay not in the identity of the major genera but in consistently differing proportions of these genera from mouth to mouth. However, at the oligotype level, we detected oligotype “fingerprints,” a highly individual-specific set of persistently abundant oligotypes fluctuating around a stable mean over time. For example, within the genus Corynebacterium, more than a dozen oligotypes were detectable in each individual, of which a different subset reached high abundance in any given person. This pattern suggests that each mouth contains a subtly different community of organisms. We also compared the Chinese plaque community characterized here to previously characterized Western plaque communities, as represented by analyses of data emerging from the Human Microbiome Project, and found no major differences between Chinese and Western supragingival plaque. In conclusion, we found the plaque microbiome to be highly individualized at the oligotype level and characterized by stability of community membership, with variability in the relative abundance of community members between individuals and over time.
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ArticleDynamics of tongue microbial communities with single-nucleotide resolution using oligotyping(Frontiers Media, 2014-11-07) Mark Welch, Jessica L. ; Utter, Daniel R. ; Rossetti, Blair J. ; Mark Welch, David B. ; Eren, A. Murat ; Borisy, Gary G.The human mouth is an excellent system to study the dynamics of microbial communities and their interactions with their host. We employed oligotyping to analyze, with single-nucleotide resolution, oral microbial 16S ribosomal RNA (rRNA) gene sequence data from a time course sampled from the tongue of two individuals, and we interpret our results in the context of oligotypes that we previously identified in the oral data from the Human Microbiome Project. Our previous work established that many of these oligotypes had dramatically different distributions between individuals and across oral habitats, suggesting that they represented functionally different organisms. Here we demonstrate the presence of a consistent tongue microbiome but with rapidly fluctuating proportions of the characteristic taxa. In some cases closely related oligotypes representing strains or variants within a single species displayed fluctuating relative abundances over time, while in other cases an initially dominant oligotype was replaced by another oligotype of the same species. We use this high temporal and taxonomic level of resolution to detect correlated changes in oligotype abundance that could indicate which taxa likely interact synergistically or occupy similar habitats, and which likely interact antagonistically or prefer distinct habitats. For example, we found a strong correlation in abundance over time between two oligotypes from different families of Gamma Proteobacteria, suggesting a close functional or ecological relationship between them. In summary, the tongue is colonized by a microbial community of moderate complexity whose proportional abundance fluctuates widely on time scales of days. The drivers and functional consequences of these community dynamics are not known, but we expect they will prove tractable to future, targeted studies employing taxonomically resolved analysis of high-throughput sequencing data sampled at appropriate temporal intervals and spatial scales.
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ArticleSite specialization of human oral veillonella species(American Society for Microbiology, 2023-01-25) Giacomini, Jonathan J. ; Torres-Morales, Julian ; Dewhirst, Floyd E. ; Borisy, Gary G. ; Mark Welch, Jessica L.Veillonella species are abundant members of the human oral microbiome with multiple interspecies commensal relationships. Examining the distribution patterns ofspecies 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 clarifytaxonomy and to test the site specialist hypothesis for thegenus, 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 thatspecies 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 namedsp. 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 enablespecies to occupy their respective habitats.Understanding the microbial ecology of the mouth is fundamental for understanding human physiology. In this study, metapangenomics demonstrated that differentspecies have clear ecological preferences in the oral cavity of healthy humans, validating the site specialist hypothesis. Furthermore, the gene pool of differentspecies was found to be reflective of their ecology, illuminating the potential role of vitamins and carbohydrates in determiningdistribution patterns and interspecies interactions.
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ArticlePreservation of three-dimensional spatial structure in the gut microbiome(Public Library of Science, 2017-11-27) Hasegawa, Yuko ; Mark Welch, Jessica L. ; Rossetti, Blair J. ; Borisy, Gary G.Preservation of three-dimensional structure in the gut is necessary in order to analyze the spatial organization of the gut microbiota and gut luminal contents. In this study, we evaluated preparation methods for mouse gut with the goal of preserving micron-scale spatial structure while performing fluorescence imaging assays. Our evaluation of embedding methods showed that commonly used media such as Tissue-Tek Optimal Cutting Temperature (OCT) compound, paraffin, and polyester waxes resulted in redistribution of luminal contents. By contrast, a hydrophilic methacrylate resin, Technovit H8100, preserved three-dimensional organization. Our mouse intestinal preparation protocol optimized using the Technovit H8100 embedding method was compatible with microbial fluorescence in situ hybridization (FISH) and other labeling techniques, including immunostaining and staining with both wheat germ agglutinin (WGA) and 4', 6-diamidino-2-phenylindole (DAPI). Mucus could be visualized whether the sample was fixed with paraformaldehyde (PFA) or with Carnoy’s fixative. The protocol optimized in this study enabled simultaneous visualization of micron-scale spatial patterns formed by microbial cells in the mouse intestines along with biogeographical landmarks such as host-derived mucus and food particles.