Specific Enzymes Found to Restrict the Growth of C difficile

A new study has found that bile salt hydrolases (BSH) enzymes can restrict the growth of the deadly colitis-inducing bacterium Clostridioides difficile (C diff).

The study, published in Nature Microbiology, examined the diversity of BSHs in the gut commensals Lactobacillaceae, a class of bacteria commonly used as probiotics. The dysregulation of BSH activity and bile acid (BA) synthesis have been proven to increase the risk of obesity, cancer, inflammatory bowel disease, and colonization resistance against C diff and other inflammatory pathogens.

“Susceptibility to CDI [C diff infection] is predominantly driven by antibiotic-induced perturbations to the intestinal microbiota resulting in a loss of colonization resistance,” the study authors wrote. “Strong evidence suggests that microbial BA metabolism is an important mechanism of colonization resistance against C difficile, as antibiotic usage depletes BSHs, increasing conjugated BAs and decreasing secondary BAs.”

First, the investigators identified a 3-residue selectivity loop that could predict the glycine versus taurine substrate preferences of BSH enzymes. Guided by this structural model, the investigators deployed a set of selective BSH enzymes with varying substrate preferences to hinder C diff spore germination and colonization. Lastly, they found an additional use for BSHs as enzymes capable of shaping microbial conjugated bile acids (MCBAs) in vivo.

To define the diversity of BSH enzymes within the Lactobacillaceae genus, the investigators assembled 3712 genomes from 274 Lactobacillaceae. BSH sequences that were highly similar were grouped into 84 clusters, and a representative sequence from each cluster were used to construct a phylogenetic tree.

Most of the BSH clusters derived from vertebrate-adapted species, with ~40% human-associated. With this cocktail of Lactobacillaceae BSHs, the investigators worked to change the bile acid pool enough to alter C diff colonization. They used human stool samples collected from patients at risk of CDI, as well as a mouse model of CDI.

In both the human and mouse models, pre-treatment with BSH cocktails stunted C diff colonization. In the BSH-treated mice, there were elevated levels of MCBAs in the gut microbiota, leading the investigators to examine whether MCBAs were also capable of inhibiting C diff germination and growth.

In vitro, the presence of MCBAs halted growth at multiple stages of the C diff life cycle. “This is more evidence that BSHs are driving changes in the bile acid pool—including making MCBAs—that could serve to inhibit C diff,” said Casey Theriot, PhD, an associate professor of infectious disease at North Carolina State University. “We’ve uncovered a new function for BSH enzymes.”

Theriot and fellow investigators believe this work is a step toward new probiotics, customized to protect against specific bacterial infections and gastrointestinal diseases.