Xufen Xia,1,* Tao Lv,2,* Lisi Zheng,2 Yuhong Zhao,3 Ping Shen,2 Danhua Zhu,2 Yunbo Chen2 

1Department of Laboratory Medicine, Tongde Hospital of Zhejiang Province, Hangzhou, Zhejiang, People’s Republic of China; 2State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, People’s Republic of China; 3Department of Blood Transfusion, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Yunbo Chen, Email chenyunbo@aliyun.com

Background: Clostridioides difficile sequence type (ST) 81, mainly associated with ribotype (RT) 369, is a TcdA-negative and TcdB-positive genotype and a common ST found in China. Furthermore, ST81 strains are reported with highest resistance rates to many antimicrobial agents. However, given the potential for C. difficile ST81 transmission, research into the epidemiological characteristics of this type of ST remain limited.
Methods: We conducted a genomic epidemiology study addressing the genetic characteristics of C. difficile ST81 in five tertiary hospitals covering different regions in China between January 2010 and January 2021. Clinical toxigenic C. difficile strains were identified, typed by multi-locus sequence typing (MLST), and phylogenetic analysis, antimicrobial resistant gene (AMR) identification were performed after all these strains were conducted by whole genome sequencing (WGS).
Results: In total, 108 clinical C. difficile strains of ST81 were isolated and successfully analyzed by WGS, which showed that the percentage of isolates with AMRs was common in this type of ST. Furthermore, two types of transposons, Tn916 and Tn6189, were also detected. We found that all C. difficile ST81 genomes were closely related as pairwise core-genomic SNP (cgSNP) distance between the strains was on average 13 cgSNPs (range, 0– 425 cgSNPs). Notably, these isolates were split into two sub-lineages (SL I and SL II) by Bayesian analysis, which suggested that both sub-lineages emerged independently. It is noted that some AMRs (such as clbA, dfrF, and cfrB) and Tn916 were only detected in SL I.
Conclusion: C. difficile ST81 is among the common STs in this study. Two independent sub-lineages of C. difficile ST81 strains are found. Furthermore, the presence of a high number of AMR genes and multiple mobile elements indicate a potential risk for transmission of C. difficile ST81. Based on these results, a robust surveillance system is crucial for identifying outbreaks, tracking infection trends, and implementing timely interventions.

Keywords: Clostridioides difficile infection, C. difficile ST81, antimicrobial resistance, whole genome sequencing, transmission