Dataset Information

Identification of Simplified Microbial Communities That Inhibit Clostridioides difficile Infection through Dilution/Extinction.

ABSTRACT: The gastrointestinal microbiome plays an important role in limiting susceptibility to infection with Clostridioides difficile To better understand the ecology of bacteria important for C. difficile colonization resistance, we developed an experimental platform to simplify complex communities of fecal bacteria through dilution and rapidly screen for their ability to resist C. difficile colonization after challenge, as measured by >100-fold reduction in levels of C. difficile in challenged communities. We screened 76 simplified communities diluted from cultures of six fecal donors and identified 24 simplified communities that inhibited C. difficile colonization in vitro Sequencing revealed that simplified communities were composed of 19 to 67 operational taxonomic units (OTUs) and could be partitioned into four distinct community types. One simplified community could be further simplified from 56 to 28 OTUs through dilution and retain the ability to inhibit C. difficile We tested the efficacy of seven simplified communities in a humanized microbiota mouse model. We found that four communities were able to significantly reduce the severity of the initial C. difficile infection and limit susceptibility to disease relapse. Analysis of fecal microbiomes from treated mice demonstrated that simplified communities accelerated recovery of indigenous bacteria and led to stable engraftment of 19 to 22 OTUs from simplified communities. Overall, the insights gained through the identification and characterization of these simplified communities increase our understanding of the microbial dynamics of C. difficile infection and recovery.IMPORTANCE Clostridioides difficile is the leading cause of antibiotic-associated diarrhea and a significant health care burden. Fecal microbiota transplantation is highly effective at treating recurrent C. difficile disease; however, uncertainties about the undefined composition of fecal material and potential long-term unintended health consequences remain. These concerns have motivated studies to identify new communities of microbes with a simpler composition that will be effective at treating disease. This work describes a platform for rapidly identifying and screening new simplified communities for efficacy in treating C. difficile infection. Four new simplified communities of microbes with potential for development of new therapies to treat C. difficile disease are identified. While this platform was developed and validated to model infection with C. difficile, the underlying principles described in the paper could be easily modified to develop therapeutics to treat other gastrointestinal diseases.

SUBMITTER: Auchtung JM

PROVIDER: S-EPMC7392540 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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Identification of Simplified Microbial Communities That Inhibit Clostridioides difficile Infection through Dilution/Extinction.

Auchtung Jennifer M JM Preisner Eva C EC Collins James J Lerma Armando I AI Britton Robert A RA

mSphere 20200729 4

The gastrointestinal microbiome plays an important role in limiting susceptibility to infection with Clostridioides difficile To better understand the ecology of bacteria important for C. difficile colonization resistance, we developed an experimental platform to simplify complex communities of fecal bacteria through dilution and rapidly screen for their ability to resist C. difficile colonization after challenge, as measured by >100-fold reduction in levels of C. diffici ...[more]

PMID: 32727857

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Project description:Clostridioides difficile infection (CDI) creates an imbalance in the intestinal microbiota due to the interaction of the components making up this ecosystem, but little is known about the impact of this disease on other microbial members. This work has thus been aimed at evaluating the taxonomic composition, potential gene-associated functions, virulence factors, and antimicrobial resistance profiles of gut microbiomes. A total of 48 DNA samples obtained from patients with health care facility-acquired (HCFO) and community-onset (CO) diarrhea were distributed in the following four groups according to CDI status: HCFO/+ (n = 13), HCFO/- (n = 8), CO/+ (n = 13), and CO/- (n = 14). These samples were subjected to shotgun metagenomics sequencing. Although the CDI groups' microbiota had microbiome alterations, the greatest imbalance was observed in the in the HCFO+/- groups, with an increase in common pathogens and phage populations, as well as a decrease in beneficial microorganisms that leads to a negative impact on some intestinal homeostasis-related metabolic processes. A reduction in the relative abundance of butyrate metabolism-associated genes was also detected in the HCFO groups (P < 0.01), with an increase in some virulence factors and antibiotic-resistance markers. A set of 51 differentially abundant species in the groups with potential association to CDI enabled its characterization, leading to their spatial separation by onset. Strong correlations between phages and some archaeal and bacterial phyla were identified. This highlighted the need to study the microbiota's various components since their imbalance is multifactorial, with some pathogens contributing to a greater or lesser extent because of their interaction with the ecosystem they inhabit. IMPORTANCE Clostridioides difficile infection represents a serious public health problem in different countries due to its high morbi-mortality and the high costs it represents for health care systems. Studies have shown the impact of this infection on intestinal microbiome homeostasis, mainly on bacterial populations. Our research provides evidence of the impact of CDI at both the compositional (bacteria, archaea, and viruses), and functional levels, allowing us to understand that the alterations of the microbiota occur systemically and are caused by multiple perturbations generated by different members of the microbiota as well as by some pathogens that take advantage of the imbalance to proliferate. Likewise, the 51 differentially abundant species in the study groups with potential association to CDI found in this study could help us envisage future treatments against this and other inflammatory diseases, improving future therapeutic options for patients.

Project description:The ability of the anaerobic gastrointestinal pathogen Clostridioides difficile to survive outside the host relies on the formation of dormant endospores. Spore formation is contingent on the activation of a conserved transcription factor, Spo0A, by phosphorylation. Multiple kinases and phosphatases regulate Spo0A activity in other spore-forming organisms; however, these factors are not well conserved in C. difficile. Previously, we discovered that deletion of a predicted histidine kinase, CD1492, increases sporulation, indicating that CD1492 inhibits C. difficile spore formation. In this study, we investigate the functions of additional predicted orphan histidine kinases CD2492, CD1579, and CD1949, which are hypothesized to regulate Spo0A phosphorylation. Disruption of CD2492 also increased sporulation frequency, similarly to the CD1492 mutant and in contrast to a previous study. A CD1492 CD2492 mutant phenocopied the sporulation and gene expression patterns of the single mutants, suggesting that these proteins function in the same genetic pathway to repress sporulation. Deletion of CD1579 variably increased sporulation frequency; however, knockdown of CD1949 expression did not influence sporulation. We provide evidence that CD1492, CD2492, and CD1579 function as phosphatases, as mutation of the conserved histidine residue for phosphate transfer abolished CD2492 function, and expression of the CD1492 or CD2492 histidine site-directed mutants or the wild-type CD1579 allele in a parent strain resulted in a dominant-negative hypersporulation phenotype. Altogether, at least three predicted histidine kinases, CD1492, CD2492, and CD1579 (herein, PtpA, PtpB and PtpC), repress C. difficile sporulation initiation by regulating activity of Spo0A. IMPORTANCE The formation of inactive spores is critical for the long-term survival of the gastrointestinal pathogen Clostridioides difficile. The onset of sporulation is controlled by the master regulator of sporulation, Spo0A, which is activated by phosphorylation. Multiple kinases and phosphatases control Spo0A phosphorylation; however, this regulatory pathway is not defined in C. difficile. We show that two predicted histidine kinase proteins, CD1492 (PtpA) and CD2492 (PtpB), function in the same regulatory pathway to repress sporulation by preventing Spo0A phosphorylation. We show that another predicted histidine kinase protein, CD1579 (PtpC), also represses sporulation and present evidence that a fourth predicted histidine kinase protein, CD1949, does not impact sporulation. These results support the idea that C. difficile inhibits sporulation initiation through multiple phosphatases.

Dataset Information

Identification of Simplified Microbial Communities That Inhibit Clostridioides difficile Infection through Dilution/Extinction.

Publications

Identification of Simplified Microbial Communities That Inhibit Clostridioides difficile Infection through Dilution/Extinction.

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