Genetic Diversity and Phylogenetic Relationship of Clinical Isolates of Brucella melitensis Based on Gene Polymorphism of β Subunit of RNA Polymerase (rpoB) Gene in Iran

Abstract

Background: The prevalence of Brucella infections in animals and humans has indicated the important need for different regional/local reference laboratories to use valid species-determining approaches to facilitate and compare data exchange. The purpose of current study was to evaluate the RNA Polymerase Beta Subunit (rpoB) as a molecular marker in Brucella species differentiation and to determine the genotype of Brucella melitensis species using single-nucleotide polymorphism (SNP) analysis. Materials & Methods: In this study, blood and cerebrospinal fluid (CSF) samples were taken from 108 patients with brucellosis. After culturing the samples insupplemented Brucella agar, eleven isolates of Brucella bacteria were isolated and identified by classical and molecular biotyping methods. Then the complete sequence of their rpoB gene was multiplied and sequenced. Sequencing results were analyzed by Mega6 program. Results: According to the results, the rpoB gene was able to differentiate between Brucella species and other bacteria.Moreover, the rpoB typing grouped the majority of Iranian isolates in the rpoB type 2, while only one strain belonged to the rpoB type 1. Among the 10 isolates of rpoB type 2, there are six different isolates with only one unique type-2 SNPs in codon 985, which gives rise to new genotype 2 variants. Conclusion: Our results shown a high discriminative power of rpoB gene among B. melitensis strains from some regions of Iran, which leads to accurate genotype and identification of these bacteria.

Investigation of Mutations in the Rifampin-Resistance-Determining Region of the rpoB Gene of Brucella melitensis by Gene Analysis

“…These findings are in agreement with those of a study showing similar missense mutations at the same location (i.e., the codon 985) of the rpoB gene in three B. melitensis type 2 strains (25,27). Other rpoB missense mutations of the genotype 2 of B. melitensis have also been observed in other studies (25)(26)(27)30). The genotype 1 of B. melitensis, harboring the rpoB gene type 1, was identified in a single isolate from Iranian capital, Tehran (accession number: MK790248).…”

“…According to our results, rpoB typing showed that most of the assessed Iranian Brucella isolates belonged to the rpoB type 2. These findings are in agreement with those of a study showing similar missense mutations at the same location (i.e., the codon 985) of the rpoB gene in three B. melitensis type 2 strains (25,27). Other rpoB missense mutations of the genotype 2 of B. melitensis have also been observed in other studies (25)(26)(27)30).…”

“…Moreover, rpoB genotyping allows the identification of new bacterial species and facilitates the analysis of bacterial communities (26,27). The frequency and nature of rpoB gene mutations vary significantly among different bacterial species (23,24).…”

Importance of brucellosis control programs of livestock on the improvement of one health

“…The first species-specific multiplex PCR assay, named AMOS-PCR, for the differentiation of Brucella was designed according to the polymorphism arising from species-specific localisation of the insertion sequence IS711 in the Brucella chromosome (Bricker and Halling 1995). For Brucella identification, further different genes were targeted such as bcsp31 (Bounaadja et al 2009), 16S-23S rDNA Interspacer (Keid et al 2007), recA gene (Scholz et al 2008), and RNA Polymerase Beta Subunit (rpoB) (Bazrgari et al 2020) also have been used. Another multiplex PCR assay (Bruce-ladder) also has been described for simple and rapid one-step Brucella identification and differentiation of most Brucella species and the vaccine strains B.melitensis Rev.1, B.abortus strain 19 (S19), and B.abortus RB51 (Lopez-Goñi et al 2008).…”

“… 2008 ), and RNA Polymerase Beta Subunit ( rpoB ) (Bazrgari et al. 2020 ) also have been used. Another multiplex PCR assay (Bruce-ladder) also has been described for simple and rapid one-step Brucella identification and differentiation of most Brucella species and the vaccine strains B.melitensis Rev.1, B.abortus strain 19 (S19), and B.abortus RB51(López-Goñi et al.…”