Clostridioides difficile hypervirulent strain ST1 isolated from clinical stool specimens obtained from three Provinces in South Africa.

Objectives: Clostridioides difficile infection is a serious healthcare-associated infection linked to antimicrobial use. The severity of the disease can be associated with hypervirulent ribotypes such as RT027. The study aimed to investigate the molecular epidemiology and genomic characteristics of C. difficile isolates from private and public healthcare settings in South Africa.

Methods: One hundred clinical stool specimens were cultured on cycloserine-cefoxitin-fructose agar. Conventional multiplex polymerase chain reaction (M-PCR) assays were conducted for isolate identification and detection of toxin genes. Genomic characteristics of the isolates were determined using whole genome sequencing (WGS) and data was analysed using pubMLST, EnteroBase, Pathogenwatch and CARD.

Results: One hundred clinically presumptive C. difficile positive stool specimens were collected, of which 62 % (62/100) were confirmed as C. difficile by M-PCR assay. Among the 62 identified C. difficile isolates, 97 % (60/62) were toxigenic, with the most dominant toxin profile being A  B  CDT  according to the M-PCR assay. The results showed that 93 % (40/43) of the WGS analysed C. difficile strains clustered into clades 1 to 5. These 40 strains were categorized into 16 sequence types (STs), with ST1 (clade 2) being the most prevalent, representing 45 % (18/40), this strain is an RT027-associated strain previously epidemic hypervirulent strain. One major cluster (n = 18) comprising ST1 strains was identified in Gauteng Province and all the isolates associated with this cluster showed the same resistome (antimicrobial resistance genes and mutations: CDD-1, aac (6')-Ie-aph (2″)-Ia, PnimB and Thr82Ile). The study also identified one strain as ST11, this strain is well known for its zoonotic potential, and two strains were identified as ST37 known as an epidemic strain. Strains from public healthcare settings exhibited genetic similarity, while those from private settings showed greater genetic diversity.

Conclusion: The study reported, for the first time, hypervirulent strains ST1 in Africa and ST11 in South Africa, with a minimum spanning tree indicating an ongoing ST1 outbreak.