Abstract:

Objective To investigate the virulence of Clostridium difficile clinical isolates, and conduct an epidemiologic study by PCR-ribotyping for the strains. Methods The unformed stool samples from hospitalized patients with diarrhea were collected, and were inoculated onto CDMN selective culture media after pretreatment with dehydrated alcohol for the culture of Clostridium difficile. Isolates were identified by Gram staining, oxygen tolerance test and agglutination assay. Bacterial genome DNA was extracted from the Clostridium difficile strains, and toxin gene tcdA and tcdB were amplified by PCR with specific primers. Meanwhile, PCR-ribotyping was performed through amplification of the genomic 16S-23S rDNA intergenic spacer region sequence, followed by electrophoresis to distinguish different ribotypes according to the polymorphism of the bands. Results Forty-four Clostridium difficile strains were divided into 3 toxin types: A+B+strains, A-B+strains and A-B-strains, which accounted for 57%, 34% and 9%, respectively. All the strains belonged to 18 ribotypes, mainly ribotype R8 (20%) and ribotype R4 (18%). Conclusion In the study, the Clostridium difficile clinical isolates were mainly A+B+ strains, and there existed relatively predominant ribotypes, but no evidence suggested nosocomial outbreak of Clostridium difficile infection.

Key words: Clostridium difficile, infection, toxin, ribotype