Metabolic Profiling of Human In-patient Stools: The Influence of C. difficile Infection and Recurrence

BACKGROUND: Initial C. difficile infection (CDI) and recurrence is dependent on host susceptibility. Hospital patients have a diverse range of other comorbidities and concomitant medications that cannot be duplicated in animal models. We hypothesize that the intersection of the luminal biome and host is represented by the metabolites. Their contribution facilitates CDI clearance or recurrence. METHODS: Discarded stool samples from 4 groups of in-patients (UC IRB 2014–8646) were used. Case controls (CC,n = 20) have diarrhea but are CDI-. Initial CDI+ (I, n = 20) clear infection. Initial CDI+ who become recurrent (IR, n = 18) have samples from initial infection. Known CDI+ Recurrent (KR, n = 10) have samples from second to fourth CDI. Stools were kept at 4°C until aliquoted, weighed and frozen at -80°C. We lyophilized each aliquot and performed metabolome analysis on 50 mg of dry sample. The NMR-Based Metabolomics Core used 1D NOESY and 1D CPMG NMR experiments to generate raw metabolic concentrations. Metabolite IDs were validated with 2D 1H-1H TOCSY and 2D 1H-13C HSQC experiments. RESULTS: 46 metabolites could be definitively identified and compared. Analysis of raw metabolite mM concentrations in CDI+ stools demonstrated more acetate (median ± median absolute deviation) (CC: 1.63 ± 1.54, I: 5.20 ± 3.21, IR: 4.21 ± 1.46, KR: 2.4 ± 2.15) and butyrate (CC: .1195 ± 0.1185, I: 0.4992 ± 0.3721, IR: 0.4196 ± 0.3832, KR: 0.2279 ± 0.2109). This suggests inability of host’s colon to productively use these SCFA. Levels of stool propionate appear to be linked to the recurrent environment (CC: 0.7620 ± 0.6562, I: 1.5162 ± 0.9189, IR: 0.6928 ± 0.5369, KR: 0.4490 ± 0.3406). Tryptophan catabolism has been linked to the CDI neutrophil response in mice. We detected decreased tryptophan in KR stool as well as lower numbers of circulating neutrophils (KR: 0.007 ± 0.007, CC: 0.0229 ± 0.0199, I: 0.0163 ± 0.0163, IR: 0.0246 ± 0.0201). This suggests altered host luminal metabolism can lead to biosystem changes CONCLUSION: CDI in the human biosystem is complex with contribution of host comorbidities and medications. The luminal interface between biome and host is the metabolome. NMR metabolomics of human in-patient stools can provide a more relevant understanding of what conditions are necessary for CDI, its clearance and recurrence. Defeat of this costly infection needs alternative approaches. DISCLOSURES: M. B. Yacyshyn, Merck: Grant Investigator, Research grant; B. Yacyshyn, Merck: Grant Investigator and Speaker’s Bureau, Research grant and Speaker honorarium