Assessing the Clinical Value of Faecal Bile Acid Profiling to Predict Recurrence in Primary Clostridioides Difficile Infection

Benjamin H. Mullish; Laura Martinez-Gili; Elena Chekmeneva; Gonçalo D. S. Correia; Matthew R. Lewis; Verena Horneffer-Van Der Sluis; Lauren A. Roberts; Julie A. K. McDonald; Alexandros Pechlivanis; Julian R. F. Walters; Emma L. McClure; Julian R. Marchesi; Jessica R. Allegretti


Aliment Pharmacol Ther. 2022;56(11):1556-1569. 

In This Article

Abstract and Introduction


Background: Factors influencing recurrence risk in primary Clostridioides difficile infection (CDI) are poorly understood, and tools predicting recurrence are lacking. Perturbations in bile acids (BAs) contribute to CDI pathogenesis and may be relevant to primary disease prognosis.

Aims: To define stool BA dynamics in patients with primary CDI and to explore signatures predicting recurrence

Methods: Weekly stool samples were collected from patients with primary CDI from the last day of anti-CDI therapy until recurrence or, otherwise, through 8 weeks post-completion. Ultra-high performance liquid chromatography-mass spectrometry was used to profile BAs. Stool bile salt hydrolase (BSH) activity was measured to determine primary BA bacterial deconjugation capacity. Multivariate and univariate models were used to define differential BA trajectories in patients with recurrence versus those without, and to assess faecal BAs as predictive markers for recurrence.

Results: Twenty (36%) of 56 patients (median age: 57, 64% male) had recurrence; 80% of recurrences occurred within the first 9 days post-antibiotic treatment. Principal component analysis of stool BA profiles demonstrated clustering by recurrence status and post-treatment timepoint. Longitudinal faecal BA trajectories showed recovery of secondary BAs and their derivatives only in patients without recurrence. BSH activity increased over time only among non-relapsing patients (β = 0.056; likelihood ratio test p = 0.018). A joint longitudinal-survival model identified five stool BAs with area under the receiver operating characteristic curve >0.73 for predicting recurrence within 9 days post-CDI treatment.

Conclusions: Gut BA metabolism dynamics differ in primary CDI patients between those developing recurrence and those who do not. Individual BAs show promise as potential novel biomarkers to predict CDI recurrence.


Clostridioides difficile infection (CDI) continues to present a considerable global disease burden, with an estimated annual incidence of 462,100 cases in the United States on latest assessment, and this trend expected to increase in line with predicted worldwide growth of antimicrobial resistance.[1] While a growing proportion of cases appear to be community-acquired,[2] CDI remains the major cause of nosocomial gastrointestinal infection,[3] leading to increased hospitalisation time[4] and a higher burden of clinical complications and mortality.[5] Furthermore, CDI is associated with considerable healthcare expenditure, equating to $1.5 billion annually within the United States.[6]

Recurrent CDI remains a major clinical challenge. A key clinical dilemma in the management of CDI patients is prediction of the risk of recurrence caused by either re-exposure to C. difficile or reactivation of dormant spores within vulnerable patients. The rate of recurrence within 8 weeks following treatment for a primary episode of CDI is 15%–25%, and rises as high as 40%–60% for patients experiencing further recurrences.[7,8] Updated CDI clinical guidelines recognise that the risk of recurrence in primary CDI patients may influence the preferred management approach and the need for preventative strategies, such as the potential benefit for bezlotoxumab in primary CDI patients with higher recurrence risk compared to those with lower risk.[9,10] However, at present, limited tools exist for the prediction of CDI recurrence,[11] with risk of recurrence estimation based on the use of broad clinical criteria (including age, immunocompromise, and severity of CDI at diagnosis[9,12]). The dynamic assessment of biomarkers could be important not only to stratify low- and high-risk patients but also to further understand the mechanisms underlying recurrence.

One such biological area of interest relates to the contribution of gut microbiota-bile acid (BA) interactions to the pathogenesis of CDI.[13] Prior antibiotic exposure is well established as the major risk factor for CDI[14] and entails a loss of bacterial microbiome community members possessing bile-metabolising enzymes (including bile salt hydrolases [BSHs] and 7-α-dehydroxylase).[15–18] BSHs are widely distributed amongst bacteria resident in the gut, and their action (in removing the taurine or glycine groups of primary BAs secreted into the gut) is the key rate-limiting step for microbiota-mediated 7α/β-dehydroxylation within the gastrointestinal tract.[19] Predicted BSH gene abundance has previously been demonstrated by our group to be reduced in stool in patients with recurrent CDI compared to those with primary CDI or control patients,[15] but no comparison of BSH activity dynamics between primary CDI patients experiencing recurrence vs no recurrence after initial treatment has been described previously. Regarding specific BAs and CDI risk, the antibiotic-exposed gut develops enrichment in primary BAs (including taurocholic acid [TCA], a major pro-germinant trigger to C. difficile[20]), and loss of secondary BAs, which have established roles in restricting the growth of C. difficile and its toxin activity[21–23] and in modulating regulatory T-cell activity.[24] In addition, CDI is also characterised by reduced activity in the farnesoid X receptor–fibroblast growth factor axis, important for BA homeostasis.[25] Faecal BA profiles have shown promise for differentiating non-CDI diarrhoea from CDI-related diarrhoea.[26]

Our recent analysis of clinical factors in a cohort of patients experiencing a first episode of uncomplicated CDI demonstrated that primary diagnosis of CDI via toxin enzyme immunoassay (EIA) and treatment with metronidazole were both factors increasing the risk of recurrence.[27] Extending upon this work, we here present a longitudinal analysis of faecal BA profiles after anti-CDI therapy cessation until recurrence or until 8 weeks post-therapy of patients within this cohort, with the joint aims of better delineating differences in BA dynamics in patients with primary CDI who recur or not, and in identifying potential biomarkers that may predict future CDI recurrence.