Geographic distribution and genetic diversity of Brucella melitensis from Yunnan province, China

Human brucellosis is a re-emerging disease in Yunnan, China, that poses serious threats to public health. However, the species/biovars, geographic distribution, and genetic diversity profile of circulating strains remain unclear. In this study, bacteriology procedures, AMOS-PCR, agglutination tests with anti-A and anti-M monospecific sera, and multiple-locus variable-number tandem-repeat analysis (MLVA) genotyping approaches were applied to investigate the genetic diversity profile of the strains. A total of 514 B. melitensis strains were isolated and identified from 2017 to 2023, of which 498 were in biovar (bv.) 3 and 16 were in bv. 1; the strains were distributed in 12 cities/prefectures and 67 counties. These data show that human B. melitensis infection has become a serious public health challenge. Moreover, 514 strains were sorted into three MLVA-11 clusters (I-III): those strains harboring 15 MLVA-11 genotypes, those strains belonging to the Eastern Mediterranean lineage, and those strains exhibiting high genetic diversity. MLVA-16 was used to divide 514 strains into eight (a-h) clades that contained 208 MLVA-16 genotypes (GTs), of which 96 shared GTs comprising 402 strains, with a cluster rate of 78.21%, suggesting that the disease was potentially dominated by a cluster epidemic; however, further genome sequence analysis is necessary to uncover the epidemic pattern of the disease. The remaining 112 GTs were unique (single strain), suggesting they were linked to sporadic cases rather than a cluster epidemic. A nationwide MLVA-16 comparison showed that 81 genotypes were shared by strains from the present study and strains from 29 other provinces, implying that strains from the nationally recognized B. melitensis genotypes were continuously spread. However, further genome-based analysis is necessary to identify the transmission chain and source of infection. Another MLVA-16 comparison with global datasets showed that the strains in this study had genotypes that were completely identical to strains from Asia (Afghanistan, Kazakhstan, Mongolia, Syria, Turkey, Saudi Arabia, Vietnam, Iraq and India) and Europe (Portugal, Spain, and Greece), indicating that these strains belonged to internationally recognized lineages. Further genome analysis is needed to identify the phylogenetic relationships of these strains. The present study describes a comprehensive scenario involving bio-typing and genetic diversity characteristics of B. melitensis strains, which can provide vital clues to tailor surveillance and control measures for human brucellosis.