SEQUENCING AND COMPARATIVE ANALYSIS OF FLAGELLIN GENES FLIA AND FLIB IN BOVINE CLOSTRIDIUM CHAUVOEI ISOLATES

Abstract

Background: Clostridium chauvoei is the etiological agent of blackleg as an endogenous infection in cattle. Flagella have been known to play a critical role in the protective immunity of animals to clostridial infections. C.chauvoei has two copies of fliC gene, namely fliA and fliB. OBJECTIVES: The aim of this study was the determination and nucleotide sequence analysis of both copies of fliC genes in vaccinal strain and Iranian C. chauvoei isolates. METHODS: Six specific primers for amplification of fliA, fliB, and flagellin (fliC) genes were designed by Oligo software. Polymerase chain reaction was performed to amplify a fragment of 700 bp for both copies of flagellin (fliA and fliB) genes. The nucleotide percentage identity and divergence among isolates were deduced using BlAST and MegAlign softwares. RESULTS: It was found that divergence in fliB was more than fliA by sequence alignment analysis. Six highly conserve regions, thirty-one SNPs and 13 amino acid polymorphisms were found in fliC gene (between fliA and fliB sequences) of Iranian C. chauvoei isolates. In comparative analysis, genomic similarity of the fliA and fliB genes between the vaccinal strain and examined field isolates was proved to be as high as 97.3 % and 98.2%, respectively. CONCLUSIONS: The fliC copies were identified as excellent biomarkers to study the molecular epidemiology and strain diversity among C. chauvoei isolates. The existence of genetic variation between two alleles of fliC gene in C. chauvoei is reported for the first time in Iran. In spite of some genetic variations, the immunologic cross protection test showed a high protection power of the local vaccine (produced by Razi Institute) against homologous and heterologous challenge.