Phytobacter diazotrophicus from Intestine of Caenorhabditis elegans Confers Colonization-Resistance against Bacillus nematocida Using Flagellin (FliC) as an Inhibition Factor

Symbiotic microorganisms in the intestinal tract can influence the general fitness of their hosts and contribute to protecting them against invading pathogens. In this study, we obtained isolate Phytobacter diazotrophicus SCO41 from the gut of free-living nematode Caenorhabditis elegans that displayed strong colonization-resistance against invading biocontrol bacterium Bacillus nematocida B16. The colonization-resistance phenotype was found to be mediated by a 37-kDa extracellular protein that was identified as flagellin (FliC). With the help of genome information, the fliC gene was cloned and heterologously expressed in E. coli. It could be shown that the B. nematocida B16 grows in chains rather than in planktonic form in the presence of FliC. Scanning Electronic Microscopy results showed that protein FliC-treated B16 bacterial cells are thinner and longer than normal cells. Localization experiments confirmed that the protein FliC is localized in both the cytoplasm and the cell membrane of B16 strain, in the latter especially at the position of cell division. ZDOCK analysis showed that FliC could bind with serine/threonine protein kinase, membrane protein insertase YidC and redox membrane protein CydB. It was inferred that FliC interferes with cell division of B. nematocidal B16, therefore inhibiting its colonization of C. elegans intestines in vivo. The isolation of P. diazotrophicus as part of the gut microbiome of C. elegans not only provides interesting insights about the lifestyle of this nitrogen-fixing bacterium, but also reveals how the composition of the natural gut microbiota of nematodes can affect biological control efforts by protecting the host from its natural enemies.

A Computational Method to Dissect Colonization Resistance of the Gut Microbiota against Pathogens

The indigenous gut microbes have co-evolved with their hosts for millions of years. Those gut microbes play a crucial role in host health and disease. In particular, they protect the host against incursion by exogenous and often harmful microorganisms, a mechanism known as colonization resistance (CR). Yet, identifying the exact microbes responsible for the gut microbiota-mediated CR against a particular pathogen remains a fundamental challenge in microbiome research. Here, we develop a computational method --- Generalized Microbe-Phenotype Triangulation (GMPT) to systematically identify causal microbes that directly influence the microbiota-mediated CR against a pathogen. We systematically validate GMPT using a classical population dynamics model in community ecology, and then apply it to microbiome data from two mouse studies on C. difficile infection. The developed method will not only significantly advance our understanding of CR mechanisms but also pave the way for the rational design of microbiome-based therapies for preventing and treating enteric infections.

Mechanism of the Gut Microbiota Colonization Resistance and Enteric Pathogen Infection

The mammalian gut microbial community, known as the gut microbiota, comprises trillions of bacteria, which co-evolved with the host and has an important role in a variety of host functions that include nutrient acquisition, metabolism, and immunity development, and more importantly, it plays a critical role in the protection of the host from enteric infections associated with exogenous pathogens or indigenous pathobiont outgrowth that may result from healthy gut microbial community disruption. Microbiota evolves complex mechanisms to restrain pathogen growth, which included nutrient competition, competitive metabolic interactions, niche exclusion, and induction of host immune response, which are collectively termed colonization resistance. On the other hand, pathogens have also developed counterstrategies to expand their population and enhance their virulence to cope with the gut microbiota colonization resistance and cause infection. This review summarizes the available literature on the complex relationship occurring between the intestinal microbiota and enteric pathogens, describing how the gut microbiota can mediate colonization resistance against bacterial enteric infections and how bacterial enteropathogens can overcome this resistance as well as how the understanding of this complex interaction can inform future therapies against infectious diseases.

130. Design and Preclinical Characterization of SER-155, an Investigational Cultivated Microbiome Therapeutic to Restore Colonization Resistance and Prevent Infection in Patients Undergoing Hematopoietic Stem Cell Transplantation

Background During allogeneic hematopoietic stem cell transplant (HSCT), the diversity and stability of the GI microbiome is disrupted, increasing the risk of domination by pathogens associated with bacteremia, aGvHD, and mortality. SER-155 is an investigational, oral microbiome therapeutic composed of cultivated spores and vegetative bacterial strains rationally designed to reduce the risk of bacteremia and aGvHD in HSCT recipients by decolonizing potential pathogens and restoring GI colonization resistance. SER-155 was evaluated in vitro for key pharmacological properties associated with colonization resistance, and in vivo to assess its ability to restore colonization resistance by reducing Enterococcus and Enterobacteriaceae carriage. Methods The design of SER-155 leveraged genomic data from interventional and observational human datasets to include taxa associated with reduced risk of infection and aGvHD in HSCT. Strains of interest were phenotyped, and over 50 candidate consortia containing different combinations of over 150 species were designed and tested in vitro and in vivo. In vivo, candidate compositions were evaluated in mouse models of vancomycin-resistant Enterococcus faecium (VRE) and carbapenem-resistant Klebsiella pneumoniae (CRE) colonization. Results Oral administration of SER-155 led to a 2-3 Log10 reduction in VRE and CRE titers compared to untreated mice (Figure 1). In vitro, the carbon source utilization profile of VRE, CRE, and SER-155 strains were assessed using a panel of 85 carbon sources. All 56 carbon sources used by CRE or VRE for anaerobic growth were also utilized by SER-155 strains, supporting a model in which nutrient competition may contribute to reducing CRE and VRE carriage and restoring colonization resistance. Figure 1. SER-155 Efficacy in Mouse Models of VRE and CRE Colonization. The titers of VRE or CRE were quantified in fecal pellets by plating on selective agar at the indicated time-points. The median A) VRE and B) CRE CFU per gram of feces was calculated for each group and plotted on the line graph (n=6-10 per group). L.O.D., limit of detection. Data were analyzed using the Mann-Whitney t-test and significance was determined as a p-value of p< 0.05*, p<0.01**, p<0.001***, p<0.0001****. Conclusion SER-155 is an investigational cultivated microbiome therapeutic intended to reduce the risk of infection by engrafting human-commensal bacterial strains in adults undergoing allogeneic HSCT. Preclinical assessments in vitro and in vivo support the ability of SER-155 to reduce VRE and CRE carriage and restore colonization resistance in the gut. A Phase 1b study evaluating SER-155 in allogeneic HSCT patients is being planned. Disclosures Elizabeth Halvorsen, PhD, Seres Therapeutics (Employee, Shareholder) Marin Vulic, PhD, Seres Therapeutics (Employee) Edward J. O’Brien, PhD, Seres Therapeutics (Employee, Shareholder) Jessica Byrant, PhD, Seres Therapeutics (Employee, Shareholder) Mary-Jane Lombardo, PhD, Seres Therapeutics (Employee, Shareholder) Christopher Ford, PhD, Seres Therapeutics (Employee, Shareholder) Matt Henn, PhD, Seres Therapeutics (Employee, Shareholder)

Citrobacter amalonaticus Inhibits the Growth of Citrobacter rodentium in the Gut Lumen

Gut bacterial infections involve three-way interactions between virulence factors, the host immune responses, and the microbiome. While the microbiome erects colonization resistance barriers, pathogens employ virulence factors to overcome them.

1,3-Diethylbenzimidazolium Triodide in Pharmacotherapy of Chronic Compensated Tonsillopharyngitis

The article presents the results of a clinical and bacteriological assessment of the pharmacological efficacy of 1,3-diethylbenzimidazolium triiodide in chronic compensated tonsillopharyngitis. As a result of a 10-day course of treatment, an improvement in the clinical status of patients was achieved, as well as a significant positive effect on the composition of the pharyngeal microbiocenosis. In particular, the content of Staphylococcus aureus, β-hemolytic streptococci, Escherichia signifi cantly decreased, and Enterobacteria, non-fermenting bacteria, and Streptococci pneumonia completely disappeared from the microbial focus. However, the conducted 10-day treatment did not lead to the complete disappearance of α-hemolytic and non-hemolytic streptococci from the microbial population. This may be explained by the reparative effect of Stellanin® on the lymphoid formations of the pharynx, contributing to the restoration of colonization resistance.