978 research outputs found

    Investigating Eco-evolutionary Interactions between Hosts and Members of Their Gut Microbiota

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    Evolutionary and ecological interactions between hosts and their associated microbial communities, their microbiota, and between members of these communities are vital to understand. Microbial communities are widespread across diverse host taxa and hosts receive a variety of well-documented benefits from their microbial communities. Despite the importance of understanding eco-evolutionary dynamics for colonization outcomes and the benefits these communities provide to their hosts, our current knowledge in this area remains incomplete. For example, we do not know the full extent of coevolution and specific relationships between hosts and microbes, and between the microbes themselves, across host taxa. Questions remain about how host taxonomy, ecology and physiology, and other present microbes influence microbial community membership and function, host and microbe evolution, and specificity in colonization of hosts. I present several studies that aim to shed further light on these eco-evolutionary topics utilizing insect pollinators, with a particular focus on bumble bees, and their gut microbial communities

    Developing Manduca sexta as a model for microbiome research

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    Evolutionary, ecological and biotechnological perspectives on plasmids resident in the human gut mobile metagenome

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    Numerous mobile genetic elements (MGE) are associated with the human gut microbiota and collectively referred to as the gut mobile metagenome. The role of this flexible gene pool in development and functioning of the gut microbial community remains largely unexplored, yet recent evidence suggests that at least some MGE comprising this fraction of the gut microbiome reflect the co-evolution of host and microbe in the gastro-intestinal tract. In conjunction, the high level of novel gene content typical of MGE coupled with their predicted high diversity, suggests that the mobile metagenome constitutes an immense and largely unexplored gene-space likely to encode many novel activities with potential biotechnological or pharmaceutical value, as well as being important to the development and functioning of the gut microbiota. Of the various types of MGE that comprise the gut mobile metagenome, plasmids are of particular importance since these elements are often capable of autonomous transfer between disparate bacterial species, and are known to encode accessory functions that increase bacterial fitness in a given environment facilitating bacterial adaptation. In this article current knowledge regarding plasmids resident in the human gut mobile metagenome is reviewed, and available strategies to access and characterize this portion of the gut microbiome are described. The relative merits of these methods and their present as well as prospective impact on our understanding of the human gut microbiota is discussed

    Abundance, distribution and functional characterisation of gut-associated Type II toxin-antitoxin systems

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    Prokaryotic toxin-antitoxin (TA) systems (also known as addiction modules), are ubiquitous genetic modules first discovered due to their role in stabilising vertical transmission of plasmids. Generally they are two-gene systems encoding a stable toxin (Tx) and an unstable antitoxin (ATx). Loss of the TA module leads to rapid ATx degradation and depletion, leaving the Tx free to interact with cellular targets and inhibit growth. For plasmid encoded TA systems, this leads to the death of plasmid free daughter cells, ensuring plasmid maintenance in a population, and gives rise to the term "addiction module". More recently, the expansion in microbial genome data has highlighted the prevalence and diversity of TA systems, and demonstrated that they are common features of many bacterial chromosomes. In addition, metagenomic surveys have pointed to the enrichment of some TA families in particular microbial ecosystems; a prime example from surveys of the human gut microbiome and RelBE TA family. Collectively, these observations indicate a wider role for TA modules in bacterial function, with numerous roles for TA systems now hypothesised. These include: i) Stabilisation of TA associated chromosomal DNA during vertical transmission; ii) Formation of "persister" cells resistant to environmental stresses, and; iii) Population level resistance to bacteriophage attack. Additionally, some Tx components have shown activity in eukaryotic cells, raising the potential for a role in prokaryote-eukaryote interaction. Here we undertook a systematic study of Type II TA systems, to provide a comprehensive assessment of their distribution and relative abundance, to confirm activity of prevalent TA systems, and to understand putative roles these may play in gut associated bacteria and the gut microbiome. A comparative genomic and metagenomic analysis of 3919 bacterial chromosomes, 4580 plasmids, 711 bacteriophage genomes, and 781 metagenomes encompassing 16 distinct habitats was conducted using all known Type II TA systems present in the Toxin Antitoxin Database (~10,100 TA genes ~1:1 Tx:ATx). Of the 817 Type II TA system homologues found in human gut datasets, 686 were observed to have significantly higher relative abundance in the human gut microbiome over other microbial ecosystems. In parallel to these in silico findings, PCR and qPCR surveys of microbiomes from 65 stool samples obtained from healthy volunteers, as well as those with polyps or colorectal cancer, were undertaken. This demonstrated a higher ATx presence than Tx or complete module, however no differences in Tx copy number between health groups was seen. To confirm the activity of the most abundant TA system homologues identified in sequence surveys, ORFs were amplified from gut metagenomic DNA, and individual Tx or ATx cloned under the control of inducible promoters. Induction of Tx expression under normal growth conditions resulted in bacterial growth inhibition, while live dead staining showed entry into a viable but non-cultivatable state, commensurate with TA function. Experiments simulating environmental stresses encountered during colonisation of the GI tract (starvation, low pH, bile), indicated that expression of these TA systems could increase cell survival when carbon or nitrogen availability was limited (starvation). Since antibiotics are also commonly encountered by gut associated-bacteria (both as residents of the GI tract and during colonisation of other body sties) a role for gut associated TA systems in facilitating survival during antibiotic exposure was also explored. This revealed an increased number of cells surviving two hours post-treatment with ÎČ-lactams when Tx genes were expressed, and in keeping with an impact on cell growth. To test the hypothesis that TA systems may stabilize associated regions of DNA, the composition of gene neighbourhoods surrounding TA systems were also explored. ORFs surrounding TA system homologues identified in metagenomic and genomic datasets were identified using the Metagene annotator, and ORF functions predicted based on searches of the Clusters of Orthlogous Groups (COG) database. This revealed significant increases in ORFs with functions related to replication/recombination/repair and those with unknown functions. It also identified a decrease in the proportion of ORFs encoding functions such as carbohydrate and lipid transport and metabolism in regions surrounding TA systems, suggesting involvement with stabilization of mobile elements. Finally, we explored the potential for gut associated TA systems to modulate phage-microbe, and host-microbe interactions. In the case of phage-host interactions, TA systems have previously been shown to function as mediators of phage resistance at the population level, by directing cells towards a dormant state which prevents phage replication, and permits a sub-set of cells to survive phage attack. Our findings indicated the potential for gut associated TA systems to provide some degree of protection during particular host-phage interactions, but specific modules did not provide universal protection against phage. In the case of host-microbe interaction, some Type II TA system Tx components have been shown to be functional in cultured eukaryotic cells, promoting apoptosis when introduced and expressed in these cell types. However, no studies to date have examined the potential for bacterially expressed TA systems to influence eukaryotic cell health in co-culture models. To investigate this, we assessed the impact of bacterial TA system expression on the health of the intestinal epithelial cell line Caco-2 in co-culture systems specifically focusing on cell apoptosis and necrosis whilst in the presence of Escherichia coli expressing p22-RelBE

    Antibiotic Disruption of Oral Microbiota Dysregulates the Osteoimmune Response and Alveolar Bone Homeostasis in the Healthy Periodontium

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    Problem: A balanced relationship between the host and oral microbiota supports periodontal health and alveolar bone homeostasis. Antibiotic perturbation of the gut microbiota critically regulates the osteoimmune response at non-oral skeletal sites. However, the impact of antibiotics on the oral microbiota and osteoimmune mechanisms regulating alveolar bone homeostasis are unknown. Considering that periodontitis driven bone loss is caused by dysbiotic shifts in the oral microbiome, antibiotic disruption of the oral microbiota may have deleterious effects on alveolar bone homeostasis. Approach: Drinking water of sex-matched C57BL/6T specific-pathogen-free (SPF) mice was supplemented with minocycline (MINO) or vehicle (VEH) control treatment from age 6 to 12 weeks. SPF mice were euthanized at age 12 weeks to assess immediate effects and at age 18 weeks to evaluate sustained minocycline effects. 16S rDNA analysis was performed to evaluate bacterial load and phylum level alterations in the oral microbiome. Micro-CT was utilized to assess linear alveolar bone loss in the maxilla and cortical/trabecular bone microarchitecture in the mandible. qRT-PCR analysis was carried out to assess pro-osteoclastic and pro-inflammatory genes in the mandible bone marrow (MBM) and gingiva. TRAP+ osteoclastic cell outcomes in alveolar bone were evaluated by in situ and in vitro approaches. Flow cytometric analysis of immune cells was performed in MBM and cervical lymph nodes (CLNs). In a separate experiment, drinking water of male C57BL/6T germ-free (GF) mice was supplemented with MINO or VEH treatment from age 6 to 12 weeks. Results: MINO treatment increased overall bacterial load and induced phylum level alterations in the oral bacteriome of 12-week-old male SPF mice. Disruption of phylum level bacterial communities were sustained in 18-week-old male SPF mice. The effects of MINO treatment on the oral microbiota were sex-dependent as no alterations were seen in female mice. MINO treatment induced linear alveolar bone loss in both male and female SPF mice at the age 12 weeks and these effects persisted at age 18 weeks. Validating that MINO-induced catabolic effects on alveolar bone is dependent on the oral microbiota, no differences were found in linear alveolar bone loss in MINO vs. VEH treated male GF mice. Cortical bone thickness was decreased in the mandible in response to MINO treatment. Osteoclast cell size and bone interface were increased in maxillary alveolar bone sections from MINO vs. VEH treated male SPF mice. Exogenous MINO stimulation in MBM cultures derived from naĂŻve 12-week-old male SPF mice increased osteoclast size and number of nuclei. Intriguingly, these findings suggest that MINO-induced pro-osteoclastic effects could be in part independent of the microbiota. Pro-inflammatory plasmacytoid dendritic cells (DCs) were upregulated within MBM and CLNs of MINO vs. VEH treated male SPF mice. Paralleling the plasmacytoid DCs, MINO treatment profoundly increased TH1 and TH17 cell populations in the MBM and CLNs. Conclusion: The current investigation reveals that MINO disruption of oral microbiota induces a pro-inflammatory immune response, which upregulates osteoclastogenesis, and drives alveolar bone loss. This novel research shows that oral MINO therapy, a commonly prescribed antibiotic treatment, may have detrimental clinical effects on alveolar bone in the healthy periodontium

    Metabolic Feedback Inhibition Influences Metabolite Secretion by the Human Gut Symbiont Bacteroides thetaiotaomicron

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    Microbial metabolism and trophic interactions between microbes give rise to complex multispecies communities in microbe-host systems. Bacteroides thetaiotaomicron (B. theta) is a human gut symbiont thought to play an important role in maintaining host health. Untargeted nuclear magnetic resonance metabolomics revealed B. theta secretes specific organic acids and amino acids in defined minimal medium. Physiological concentrations of acetate and formate found in the human intestinal tract were shown to cause dose-dependent changes in secretion of metabolites known to play roles in host nutrition and pathogenesis. While secretion fluxes varied, biomass yield was unchanged, suggesting feedback inhibition does not affect metabolic bioenergetics but instead redirects carbon and energy to CO2 and H2. Flux balance analysis modeling showed increased flux through CO2-producing reactions under glucose-limiting growth conditions. The metabolic dynamics observed for B. theta, a keystone symbiont organism, underscores the need for metabolic modeling to complement genomic predictions of microbial metabolism to infer mechanisms of microbemicrobe and microbe-host interactions

    Gut microbiota functions: metabolism of nutrients and other food components

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    The diverse microbial community that inhabits the human gut has an extensive metabolic repertoire that is distinct from, but complements the activity of mammalian enzymes in the liver and gut mucosa and includes functions essential for host digestion. As such, the gut microbiota is a key factor in shaping the biochemical profile of the diet and, therefore, its impact on host health and disease. The important role that the gut microbiota appears to play in human metabolism and health has stimulated research into the identification of specific microorganisms involved in different processes, and the elucidation of metabolic pathways, particularly those associated with metabolism of dietary components and some host-generated substances. In the first part of the review, we discuss the main gut microorganisms, particularly bacteria, and microbial pathways associated with the metabolism of dietary carbohydrates (to short chain fatty acids and gases), proteins, plant polyphenols, bile acids, and vitamins. The second part of the review focuses on the methodologies, existing and novel, that can be employed to explore gut microbial pathways of metabolism. These include mathematical models, omics techniques, isolated microbes, and enzyme assays

    Commensal microbiota modulates larval foraging behaviour, development rate and pupal production in Bactrocera tryoni

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    Project Raising Q-fly Sterile Insect Technique to World Standard (HG14033) is funded by the Hort Frontiers Fruit Fly Fund, part of the Hort Frontiers strategic partnership initiative developed by Hort Innovation, with co-investment from Macquarie University and contributions from the Australian Government. BN is supported by an international Research Training Program (iRTP) scholarship from Macquarie University (NSW, Australia).Peer reviewedPublisher PD

    Microbiome function predicts amphibian chytridiomycosis disease dynamics

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    [Background] The fungal pathogenBatrachochytrium dendrobatidis (Bd) threatens amphibian biodiversity and ecosystem stability worldwide. Amphibian skin microbial community structure has been linked to the clinical outcome of Bd infections, yet its overall functional importance is poorly understood. [Methods] Microbiome taxonomic and functional profiles were assessed using high-throughput bacterial 16S rRNA and fungal ITS2 gene sequencing, bacterial shotgun metagenomics and skin mucosal metabolomics. We sampled 56 wild midwife toads (Alytes obstetricans) from montane populations exhibiting Bd epizootic or enzootic disease dynamics. In addition, to assess whether disease-specific microbiome profiles were linked to microbe-mediated protection or Bd-induced perturbation, we performed a laboratory Bd challenge experiment whereby 40 young adult A. obstetricans were exposed to Bd or a control sham infection. We measured temporal changes in the microbiome as well as functional profiles of Bd-exposed and control animals at peak infection. [Results] Microbiome community structure and function differed in wild populations based on infection history and in experimental control versus Bd-exposed animals. Bd exposure in the laboratory resulted in dynamic changes in microbiome community structure and functional differences, with infection clearance in all but one infected animal. Sphingobacterium, Stenotrophomonas and an unclassified Commamonadaceae were associated with wild epizootic dynamics and also had reduced abundance in laboratory Bd-exposed animals that cleared infection, indicating a negative association with Bd resistance. This was further supported by microbe-metabolite integration which identified functionally relevant taxa driving disease outcome, of which Sphingobacterium and Bd were most influential in wild epizootic dynamics. The strong correlation between microbial taxonomic community composition and skin metabolome in the laboratory and field is inconsistent with microbial functional redundancy, indicating that differences in microbial taxonomy drive functional variation. Shotgun metagenomic analyses support these findings, with similar disease-associated patterns in beta diversity. Analysis of differentially abundant bacterial genes and pathways indicated that bacterial environmental sensing and Bd resource competition are likely to be important in driving infection outcomes. [Conclusions] Bd infection drives altered microbiome taxonomic and functional profiles across laboratory and field environments. Our application of multi-omics analyses in experimental and field settings robustly predicts Bd disease dynamics and identifies novel candidate biomarkers of infection. [MediaObject not available: see fulltext.]K.A.B. was funded by a CASE studentship from NERC, NERC Biomolecular Analysis Facility grant (NBAF939) and an E.P. Abraham Junior Research Fellowship from St Hilda’s College, University of Oxford. M.C.F and T.W.J.G. were funded by NERC award NE/E006701/1 and the Biodiversa project RACE: Risk Assessment of Chytridiomycosis to European Amphibian Biodiversity. T.W.J.G was also funded by Research England and NERC NE/S000062/1. D.S.S. and A.L. received funding through the project People, Pollution, and Pathogens financed through the call “Mountains as Sentinels of Change” by the Belmont-Forum (ANR-15-MASC-0001 - P3, DFG-SCHM3059/6-1, NERC-1633948, NSFC-41661144004). D.S.S. holds the AXA Chair for Functional Mountain Ecology funded by the AXA Research Fund through the project GloMEc and M.C.F. is a fellow in the CIFAR ‘Fungal Kingdoms’ Program
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