Molecular cloning and characterisation of the methionine sulphoxide reductase A (msrA) gene locus in Campylobacter lari organisms

Abstract

The methionine sulphoxide reductase A (msrA) gene and its adjacent genetic loci from urease-negative (UN) Campylobacter lari RM2100 and urease-positive thermophilic Campylobacter (UPTC) CF89–12 strains appear to be composed of a msrA structure gene (507 base pairs [bp]) and another five-gene cluster (approximately 6300 bp) in the same strand and direction. A primer pair (F1/R4-msrA) for polymerase chain reaction (PCR) amplification was designed to generate a product of approximately 900 bp of the msrA gene, including its adjacent genetic loci for the thermophilic Campylobacter organisms and generate an amplicon with 16 C. lari isolates (n=4 for UN C. lari; n=12 for UPTC). Following direct nucleotide sequencing, sequence analysis and nucleotide sequence alignment analysis, the putative full-length msrA gene from the 16 C. lari isolates showed high nucleotide sequence similarities (91.8–100%) to each other and relatively low similarity (69.3–71.8%) to three reference C. jejuni and C. coli strains. In addition, the msrA gene was transcribed in both the UPTC CF89–12 and NCTC12893 cells using reverse transcription PCR. An immunoreactively positive signal was identified in the UPTC CF89–12 and NCTC12893 cells with anti-UPTC MsrA synthetic peptide antibodies.

Key Words:

• Campylobacter
• DNA sequencing
• Polymerase chain reaction