Molecular characteristics of Brucella melitensis isolates from humans in Qinghai Province, China

Abstract

Background The prevalence of human brucellosis in Qinghai Province has been increasing rapidly, with confirmed cases distributed across 31 counties. However, the epidemiology of brucellosis transmission has not been fully elucidated. To characterize the two isolated strains, multiple locus variable-number tandem repeats analysis (MLVA) and whole-genome single-nucleotide polymorphism (SNP)-based approaches were employed. Methods Blood samples were isolated from two males that were confirmed B. melitensis positive following MLVA. Genomic DNA was extracted from these samples, and whole-genome sequencing (WGS) was performed. Next, SNP-based phylogenetic analysis was performed to compare the two strains to 94 B. melitensis strains (complete genome and draft genome) retrieved from online databases. Results The two Brucella isolates were identified as B. melitensis biovar 3 (QH2019001 and QH2019005) following conventional biotyping and were found to have differences in their variable number tandem repeats (VNTRs) using MLVA-16. Phylogenetic examination assigned the 96 strains to five genotype groups, with QH2019001 and QH2019005 assigned to the same group, but different subgroups. Moreover, the QH2019005 strain was assigned to a new subgenotype, IIj, within genotype II. These findings were then combined to determine the geographic origin of the two Brucella strains. Conclusions Utilizing a whole-genome SNP-based approach enabled differences between the two B. melitensis strains to be more clearly resolved, and facilitated the elucidation of their different evolutionary histories. This approach also revealed that QH2019005 is a member of a new subgenotype (IIj) with an ancient origin in the eastern Mediterranean Sea.