[Identification and phylogenetic analysis on the aroA gene of Haemophilus parasuis].

ABSTRACT

OBJECTIVE: We clarified the genetic structure and phylogenetic relationship of the aroA gene in Haemophilus parasuis, a gene encoding 5'-enolpyruvylshikimate-3-phosphate synthetase. METHODS: We used PCR and bacterial genome walking techniques to determine the aroA gene locus in the genome of H. parasuis serovar 5. Then we sequenced the aroA gene of H. parasuis reference strains and local isolates. All aroA gene sequences from H. parasuis, other members of the family Pasteurellaceae, Escherichia coli and Salmonella typhimurium were analyzed for phylogenetic relationship. RESULTS: An approximately 3.7 kb fragment containing the whole aroA gene was obtained from the genomic DNA of H. parasuis serovar 5. Sequence analysis showed that the size of the whole aroA gene was 1314 bp, encoding a 437 aa product with molecular weight 47.9 kDa. The sequence immediately upstream of the aroA gene shared high similarity with phosphoenolpyruvate carboxylase gene. The whole aroA gene fragment could be amplified from all strains of H. parasuis serovars with a size of 1476 bp. The nucleotide sequence similarity was above 97.7% in H. parasuis serovars. The similarity was 70.6%-78.9% between H. parasuis serovar 5 and other members of Pasteurellaceae. The similarity between H. parasuis serovar 5 and E. coli or S. typhimurium was 66.4% and 67.2% respectively. CONCLUSION: No obvious difference was found in the aroA gene sequence of H. parasuis reference strains (15 serovars) and 3 local isolates. The nucleotide sequence of the aroA gene was well-conserved within Gram-negative organisms.