Phylogenetic Analysis of Brucella melitensis Strains Isolated from Humans Using 16S rRNA Sequencing and Multiple Locus Variable Number of Tandem Repeats Analysis-16.

Background: Brucellosis is the most important public health problem worldwide, and the annual incidence of the disease in humans is 2.1 million. The Brucella genome is highly conserved, with over 90% similarity among species. The aim of this study was to perform species-level identification of Brucella spp. strains isolated from humans diagnosed with brucellosis and to further investigate the phylogenetic relationships using multiple locus variable number of tandem repeats analysis (MLVA)-16 and 16S rRNA sequencing analysis. Materials and Methods: Brucella spp. was isolated from the blood cultures of 54 patients who tested positive for brucellosis through serological examinations. Real-time PCR was used to identify the isolates in species, and the genus level of Brucella was confirmed with 16S rRNA. All isolates were subjected to phylogenetic analysis using variable number of tandem repeat analysis with multiple loci. Results: Subsequent analysis via real-time PCR confirmed these isolates to be of the Brucella melitensis species. The 16S rRNA sequence analysis showed 100% homogeneity among the isolates. MLVA revealed the formation of five different genotypic groups. While two groups were formed based on the 16S rRNA sequence analysis, five groups were formed in the MLVA. Conclusions: The study concluded that 16S rRNA sequence analysis alone did not provide sufficient discrimination for phylogenetic analysis but served as a supportive method for identification. MLVA exhibited higher phylogenetic power. The widespread isolation of B. melitensis from human brucellosis cases highlights the importance of controlling brucellosis in small ruminants to prevent human infections.

Genotyping of Brucella isolates from animals and humans by Multiple-Locus Variable-number Tandem Repeat Analysis (MLVA).

This study investigates country-wide genotype variations through the genotyping of Brucella strains isolated from domestic ruminants and humans. The Brucella spp. isolated from samples taken from animals and humans were first identified as B. abortus and B. melitensis by real-time PCR, and the MLVA-16 approach was then used for the genotyping of the identified isolates. For the study, 416 Brucella spp. were isolated from aborted fetus samples examined between 2018 and 2021, and 74 Brucella spp. from infected humans. Of the 74 human isolates analyzed, 1.3% were identified as B. abortus and 98.7% (73/74) as B. melitensis. The MLVA-16 typing method revealed 30 clonal groups for B. abortus and 37 clonal groups for B. melitensis from which the dominant genotypes and similarities with human isolates in Türkiye were determined.