Molecular analysis and genotyping of pathogenicity locus in Clostridioides difficile strains isolated from patients in Tehran hospitals during the years 2007-2010.

BACKGROUND & AIMS: Clostridioides difficile (C. difficile) has been identified as the leading cause of antibiotic associated diarrhea (AAD). Co-carriage of an intact pathogenicity locus (PaLoc) with binary toxin genes in C. difficile strains seems to be linked with severe disease outcomes in the infected patients. Epidemiology of C. difficile infection (CDI) in hospital setting and knowledge about their genetic context help us to decrease the morbidity, mortality, and costs associated with Clostridioides difficile infection. In the present study was aimed to characterize genetic diversity of PaLoc among different C. difficile strains isolated from hospitalized patients and carriage of cytolethal distending toxin gene (cdt) in different hospitals.

METHOD

C. difficile strains were isolated from stool samples of inpatients referred to a reference laboratory from different hospitals and also outpatients with diarrhea, during 2008-2011. DNA was extracted from pure culture of the bacterium and PCR was performed for tcdA, tcdB, tcdE, tcdC, tcdD, and cdu2 genes. Carriage of two binary toxin genes cdtA, cdtB was also determined in these strains. To find clonal strains, similarity of genotypes and integrity of PaLoc among the isolates was compared in each hospital.

RESULTS

The intact PaLoc was found most frequently among the isolates in the outpatients (19/51, 37.2%, Group I), while incomplete PaLoc found mostly in patients who were hospitalized in the infectious diseases and internal diagnosis wards. tcdA and tcdB genes were detected in different combinations among the studied strains. These strains showed tcdAB, tcdAB, and tcdAB genotypes in a frequency of 76.4% (39/51), 7.8% (4/51), and 17.6% (9/51), respectively. Analysis of gene composition of the PaLoc showed 19 distinct genotypes among the 51 strains. Accordingly, 38 strains were classified mainly into 6 regular groups, while the remaining strains showed heterogeneous patterns. tcdC/tcdD constituted the most common genotypic group among the strains with partial PaLoc (7/51, 13.7%). A hypertoxigenic genotype, tcdC/tcdA/tcdB, was detected in 2 strains (2/51, 3.9%). The intact genotype was also detected in a C. difficile isolate from outpatients. Cdt encoding genes toxins was observed in low numbers of the strains (7/52, 13.5%). All of cdtAB strains were belonged to PaLoc group 1 (intact genotype). Statistical analyses showed no correlation between particular genotypes and special wards of the hospitals (p value>0.05).

CONCLUSION

Collectively, our results showed diversity of C. difficile strains in most wards of the studied hospitals. Diversity of PaLoc genotypes in the strains that isolated from the same wards proposed endogenous routes of the infection, as common cause of CDI in these patients.