Phenotypic and genotypic acyclovir resistance surveillance of genital herpes simplex virus 2 in South Africa.

Acyclovir (ACV) is currently included in the syndromic management algorithm for genital ulcer disease in South Africa, and is the recommended first-line treatment for herpes simplex virus 2 (HSV-2). In the majority of cases, HSV-2 resistance to ACV is due to amino acid changes within the viral thymidine kinase (TK). Phenotypic and genotypic ACV resistance surveillance of HSV-2 derived from genital ulcer disease swab specimens was conducted at a primary healthcare facility in Johannesburg between 2018 and 2020. The objectives of this surveillance were to identify ACV resistance-associated mutations and polymorphisms in HSV-2 TK, and to determine the phenotypic ACV resistance profiles of the corresponding clinical HSV-2 isolates. Genotypic analysis of TK from 67 HSV-2 positive genital ulcer swabs revealed 48 specimens with TK mutations, conferring 113 nucleotide changes. No resistance-associated mutations were found, however, we identified nine known natural polymorphisms (R26H, A27T, S29A, G39E, N78D, L140F, T159I, R220K and R284S) and five amino acid changes of unknown significance (R18C, G39K, M70R, P75S and L263P). Phenotypic susceptibility testing of 52 cultivable HSV-2 isolates revealed all to be susceptible to ACV with IC50 values of <2 μg/ml. The five amino acid changes of unknown significance identified by genotypic testing were not correlated to phenotypic ACV resistance, and therefore grouped as natural polymorphisms. We did not detect any unknown or resistance-associated mutations in specimens that could not be phenotypically tested for ACV resistance. Our findings will supplement existing databases of HSV antiviral resistance-associated mutations and polymorphisms that could be used for genotypic ACV resistance screening.