Epidemiological and genetic characterization of Clostridium butyricum cultured from neonatal cases of necrotizing enterocolitis in China

OBJECTIVE: Laboratory-based characterization and traceback of Clostridium butyricum isolates linked to outbreak cases of neonatal necrotizing enterocolitis (NEC) in a hospital in China. METHODS: In total, 37 samples were collected during the NEC outbreak. Classical bacteriological methods were applied to isolate and identify Clostridium spp. Meanwhile, 24 samples collected after an outbreak were similarly tested. All Clostridium isolates were identified to species level as either C. butyricum or C. sporogenes. These isolates were subsequently subtyped using pulsed-field gel electrophoresis (PFGE). Genomic DNA was purified from 2 representative C. butyricum isolates and sequenced to completion. RESULTS: Of 37 samples collected during the NEC outbreak, 17 (45.95%) were positive for Clostridium spp. One species, C. butyricum, was cultured from 10 samples. Another species cultured from 2 other samples was identified as C. sporogenes. Both of these species were cocultured from 5 samples. Pulsotyping showed that the 15 C. butyricum and the 7 C. sporogenes isolates produced indistinguishable DNA profiles. No NEC cases were reported after disinfection following the outbreak, and all samples collected after the outbreak were negative for Clostridium spp. Whole-genome sequencing (WGS) indicated that sialidase, hemolysin, and enterotoxin virulence factors were located on the chromosomes of 2 C. butyricum isolates. CONCLUSIONS: The outbreak of NEC was epidemiologically linked to C. butyricum contamination within the hospital. This is the first report of an NEC outbreak associated with C. butyricum infection in China.