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BackgroundWithin eight weeks of primary Clostridioides difficile infection (CDI), as many as 30% of patients develop recurrent disease with the associated risks of multiple relapses, morbidity, and economic burden. There are no validated biomarkers predictive of recurrence during primary infection. This study demonstrates the potential of a simple test for identifying hospitalized CDI patients at low risk for disease recurrence.MethodsForty-six hospitalized CDI patients were enrolled at Emory University Hospitals. Serum and MENSA samples prepared during weeks 1, 2, and 4 following symptom-onset were measured for antibodies specific for ten C. difficile antigens.ResultsAmong the 46 C. difficile-infected patients, nine (19.5%) experienced recurrence within eight weeks of primary infection. Among the 37 non-recurrent patients, 23 had anti-C. difficile MENSA antibodies specific for any of the three toxin antigens: TcdB-CROP, TcdBvir-CROP, and/or CDTb. Positive MENSA responses occurred within the first week post-symptom onset, including six patients who never seroconverted. A similar trend was observed in serum responses, but they peaked later and identified fewer patients (19/37). In contrast, none of the patients who subsequently recurred after hospitalization produced antibodies specific for the three C. difficile toxin antigens. IgA antibodies for the toxin antigens demonstrated the greatest predictive power for protection from recurrence.DiscussionThe development of IgG and/or IgA antibodies for three C. difficile toxins in serum and/or MENSA has prognostic potential. These immunoassays measure nascent immune responses that reduce the likelihood of recurrence. Early identification of patients at-risk for recurrence can reduce costs and morbidity.

BACKGROUNDClostridioides difficile infections (CDI) have been a challenging and increasing serious concern in recent years. While early and accurate diagnosis is crucial, available assays have frustrating limitationsOBJECTIVEDevelop a simple, blood-based immunoassay to accurately diagnose patients suffering from active CDI.MATERIALS AND METHODSUninfected controls (n=95) and CDI patients (n=167) were recruited from Atlanta area hospitals. Blood samples were collected from patients within twelve days of a positive CDI test and processed to yield serum and PBMCs cultured to yield medium enriched for newly synthesized antibodies (MENSA). Multiplex immunoassays measured Ig responses to ten recombinant C. difficile antigens.RESULTSSixty-six percent of CDI patients produced measurable responses to C. difficile antigens in their serum or MENSA within twelve days of a positive CDI test. Fifty-two of the 167 CDI patients (31%) were detectable in both serum and MENSA, but 32/167 (19%) were detectable only in MENSA, and 27/167 (16%) were detectable only in serum.DISCUSSIONWe describe the results of a multiplex immunoassay for the diagnosis of ongoing CDI in hospitalized patients. Our assay resolved patients into four categories: MENSA-positive only, serum-positive only, MENSA- and serum-positive, and MENSA- and serum-negative. The MENSA positive-only patients accounted for 30% and may be attributed to nascent antibody secretion in MENSA prior to seroconversion. Conversely, the serum positive-only subset may have been more advanced in their disease course. Immunocompromise and misdiagnosis may have contributed to the 34% of CDI patients who were not identified using MENSA or serum immunoassays.IMPORTANCEWhile there was considerable overlap between patients identified through MENSA and serum, both methods detected additional, unique patients. The combined use of both MENSA and serum to detect CDI patients resulted in the greatest identification of CDI patients. Together, longitudinal analysis of MENSA and serum will provide a more accurate evaluation of successful host humoral immune responses in CDI patients.