Strain-Dependent Inhibition of Clostridioides difficile by Commensal <i>Clostridia</i> Carrying the Bile Acid-Inducible ( <i>bai</i> ) Operon

Abstract

<i>Clostridioides difficile</i> is one of the leading causes of antibiotic-associated diarrhea. Gut microbiota-derived secondary bile acids and commensal <i>Clostridia</i> that carry the bile acid-inducible (<i>bai</i>) operon are associated with protection from <i>C. difficile</i> infection (CDI), although the mechanism is not known. In this study, we hypothesized that commensal <i>Clostridia</i> are important for providing colonization resistance against <i>C. difficile</i> due to their ability to produce secondary bile acids, as well as potentially competing against <i>C. difficile</i> for similar nutrients. To test this hypothesis, we examined the abilities of four commensal <i>Clostridia</i> carrying the <i>bai</i> operon (<i>Clostridium scindens</i> VPI 12708, <i>C. scindens</i> ATCC 35704, <i>Clostridium hiranonis</i>, and <i>Clostridium hylemonae</i>) to convert cholate (CA) to deoxycholate (DCA) <i>in vitro,</i> and we determined whether the amount of DCA produced was sufficient to inhibit the growth of a clinically relevant <i>C. difficile</i> strain. We also investigated the competitive relationships between these commensals and <i>C. difficile</i> using an <i>in vitro</i> coculture system. We found that inhibition of <i>C. difficile</i> growth by commensal <i>Clostridia</i> supplemented with CA was strain dependent, correlated with the production of ∼2 mM DCA, and increased the expression of <i>bai</i> operon genes. We also found that <i>C. difficile</i> was able to outcompete all four commensal <i>Clostridia</i> in an <i>in vitro</i> coculture system. These studies are instrumental in understanding the relationship between commensal <i>Clostridia</i> and <i>C. difficile</i> in the gut, which is vital for designing targeted bacterial therapeutics. Future studies dissecting the regulation of the <i>bai</i> operon <i>in vitro</i> and <i>in vivo</i> and how this affects CDI will be important.<b>IMPORTANCE</b> Commensal <i>Clostridia</i> carrying the <i>bai</i> operon, such as <i>C. scindens,</i> have been associated with protection against CDI; however, the mechanism for this protection is unknown. Herein, we show four commensal <i>Clostridia</i> that carry the <i>bai</i> operon and affect <i>C. difficile</i> growth in a strain-dependent manner, with and without the addition of cholate. Inhibition of <i>C. difficile</i> by commensals correlated with the efficient conversion of cholate to deoxycholate, a secondary bile acid that inhibits <i>C. difficile</i> germination, growth, and toxin production. Competition studies also revealed that <i>C. difficile</i> was able to outcompete the commensals in an <i>in vitro</i> coculture system. These studies are instrumental in understanding the relationship between commensal <i>Clostridia</i> and <i>C. difficile</i> in the gut, which is vital for designing targeted bacterial therapeutics.