Infection, Disease Control and Prevention

Disease induced by Clostridium difficile infection (CDI) is generally viewed as "monomicrobial" being dominated by the virulence factors of CDI alone. However, co-infections may occur but their significance in CDI is unknown. Fecal specimens from pediatric patients (2-18 years) were screened using BioFire FilmArray GI Panel which detects 22 enteric pathogens. Of 357 patients, 88% had antibiotic-associated diarrhea. Based on toxin PCR, 50% were diagnosed with non-recurrent CDI (nCDI), 8% with recurrent CDI (rCDI), and 30% were C. difficile toxin negative (AAD). Patients without GI symptoms served as controls. FilmArray identified additional pathogens in 31.1% of patients with primary CDI; 64.5% with rCDI; 49.5% with AAD; and 11.9% controls. Enteropathogenic E. coli (EPEC) and rotavirus were significant co-infections in rCDI compared with nCDI (p<0.05). In a murine co-infection model, rotavirus improved clinical symptoms; whereas, co-infection with Citrobacter rodentium, a model of EPEC, resulted in greater disease and mortality than singly infected mice (p<0.05). Four weeks post-infection, co-infected mice showed significant intestinal inflammation that was not present in singly infected mice (p<0.05), which correlated with prolonged bacterial shedding and toxin production. Mortality in co-infected mice was associated with reductions in early response chemokines involved in the recruitment of protective innate immune cells. Administration of innate cytokine IL-22 protected co-infected mice from death compared to controls (p<0.05). Taken together, co-infections can exert differential clinical outcomes in CDI. Notably, co-infection with EPEC may place CDI patients at greater risk of disease recurrence because of pathogen-induced impairment in protective innate immunity against C. Difficile.