Clostridium (C.) difficile-infection (CDI), a nosocomial gastrointestinal disorder, is of growing concern due to its rapid rise in recent years. Antibiotic therapy of CDI is associated with disrupted metabolic function and altered gut microbiota. The use of probiotics as an adjunct is being studied extensively due to their potential to modulate metabolic functions and the gut microbiota. In the present study, we assessed the ability of several single strain probiotics and a probiotic mixture to change the metabolic functions of normal and C. difficile-infected fecal samples. The production of short-chain fatty acids (SCFAs), hydrogen sulfide (H 2 S), and ammonia was measured, and changes in microbial composition were assessed by 16S rRNA gene amplicon sequencing. The C. difficile-infection in fecal samples resulted in a significant decrease (p < 0.05) in SCFA and H 2 S production, with a lower microbial alpha diversity. All probiotic treatments were associated with significantly increased (p < 0.05) levels of SCFAs and restored H 2 S levels. Probiotics showed no effect on microbial composition of either normal or C. difficile-infected fecal samples. These findings indicate that probiotics may be useful to improve the metabolic dysregulation associated with C. difficile infection.Probiotics, defined as live microorganisms that impart beneficial effects on the host when given in adequate quantities [11], have shown beneficial effects in the GI tract such as improving metabolic function [12][13][14][15], counteracting infections [16][17][18], regulating immune function, decreasing GI disorder symptoms [16,[19][20][21], and potentially lowering the risk of developing colon cancer [22]. Most of the probiotics utilized to date are usually from the Lactobacilli, Bifidobacteria, and yeast (Saccharomyces) groups. The efficacy and proposed mechanisms of action of these microbes in regulating intestinal microbiota functions are generally strain-specific. Some probiotic strains are thought to produce antimicrobial metabolites such as bacteriocins, to lower the pH by generating hydrogen peroxide and SCFAs, or to restrict pathogenic growth by competing for essential nutrients and adherence onto the gut mucosal barrier [18,[23][24][25][26][27]. Several probiotics may reduce CDI-associated diarrhea and prevent primary CDI formation using some of the abovementioned mechanisms, but perhaps predominantly by inhibiting the adhesion of C. difficile in the intestine [28,29]. In the case of Saccharomyces (S.) boulardii, the mechanism was shown to involve the proteolytic hydrolysis of the CD enterotoxins A and B [30]. Although the Lactobacillus (L.) rhamnosus GG strain and S. boulardii have been studied the most in the context of CDI-associated diarrhea [9,31], several other strains such as Bifidobacterium (B.) longum and L. acidophilus CL1285 have also shown efficacy against antibiotic-associated diarrhea [32][33][34][35]. Furthermore, strains such as L. plantarum 299v have been shown to enhance microbial function in CDI patients recei...