Our current Merit Award focuses on factors contributing to virulence in Clostridium difficile, and a number of surface-layer proteins (SLPs) have been identified, that mediate bacterial adhesion to host intestinal epithelial cells. We now propose to significantly expand on these studies with the ultimate goal of translating our findings to a safe, easily utilizable, probiotic-based intervention for prevention of C. difficile infection (CDI).
Three Specific Aims are proposed. First, we will define mechanistic bases of CD adherence to host epithelia mediated by surface-layer proteins (SLPs). CD mutagenesis and animal models of CDI will be exploited for these studies. Second, we will characterize non-SLP factors involved in CD colonization. Both bacterial and host- response proteins involved in, and contributing to, CD colonization as well as colonization resistance will be studied. Third, we will translate our bench-research findings to a clinically relevant outcome by developing a new adherence-based CDI intervention. This will involve construction of a targeted CD colonization inhibitor, by engineering a probiotic bacterium (Lactobacillus acidophilus) to express the CD colonization protein SlpA. We will validate the probiotic developed above by testing it against frequently isolated single-episode and relapse strains of CD recovered from surveillance studies performed at our VA hospital. For all studies in this proposal, we will use both the hamster and mouse models of acute CDI and CD colonization. We will also incorporate state-of-the-art methodologies including automated mass spectrometry and live-animal bioluminescence imaging to track the fate of ingested CD spores.

Public Health Relevance

Over 400,000 cases of Clostridium difficile infection (CDI) occur annually in the USA, imposing a burden of >$3 billion on the healthcare system. Risk factors for CDI include hospitalization, age >65, co-morbid conditions, and antibiotic/PPI use - all of which are immediately relevant to patients treated in VA hospitals. Upto 11% of veterans also relapse with multiple recurrences of the disease. Severity of initial infection as well as relapses are also much higher with the newly emerged hypervirulent strains of C. difficile that are now common in VA hospitals. The studies proposed in this application are focused on understanding how C. difficile colonizes the human gut, and to validate a safe, cost-effective, easily tolerated intervention to prevent this establishment. The proposed work is thus consistent with the Merit Review mechanism, and is directly relevant to Veterans health.

National Institute of Health (NIH)
Veterans Affairs (VA)
Non-HHS Research Projects (I01)
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Gastroenterology (GAST)
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Southern Arizona VA Health Care System
United States
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Roxas, Jennifer Lising; Monasky, Ross Calvin; Roxas, Bryan Angelo P et al. (2018) Enteropathogenic Escherichia coli EspH-Mediated Rho GTPase Inhibition Results in Desmosomal Perturbations. Cell Mol Gastroenterol Hepatol 6:163-180
Soavelomandroso, Anna P; Gaudin, Fran├žoise; Hoys, Sandra et al. (2017) Biofilm Structures in a Mono-Associated Mouse Model of Clostridium difficile Infection. Front Microbiol 8:2086
Merrigan, Michelle M; Venugopal, Anilrudh; Roxas, Jennifer L et al. (2013) Surface-layer protein A (SlpA) is a major contributor to host-cell adherence of Clostridium difficile. PLoS One 8:e78404
Roxas, Jennifer Lising; Wilbur, John Scott; Zhang, Xiangfeng et al. (2012) The enteropathogenic Escherichia coli-secreted protein EspZ inhibits host cell apoptosis. Infect Immun 80:3850-7
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McQuade, Rebecca; Roxas, Bryan; Viswanathan, V K et al. (2012) Clostridium difficile clinical isolates exhibit variable susceptibility and proteome alterations upon exposure to mammalian cationic antimicrobial peptides. Anaerobe 18:614-20