Cloning and Sequence Analysis of aha1 Gene Encoding Major Adhesin Protein from Aeromonas sp. Isolated from Aquaculture Animals with Haemorrhagic Septicemia

摘要

Nine bacterial strains (LA1, LA2, LA4, JA1, CA3, CA4, BA1, BA18 and EA9) were isolated from different aquaculture animals with haemorrhagic septicemia and then they were identified as A. hydrophila (LA4, JA1, CA3 and BA1), A. sobria (LA1 and EA9), A. caviae (BA18) and A. veronii (CA4 and LA2), respectively by morphological and biochemical characterization. All isolates were found to be pathogenic to experimental zebrafish (Danio rerio) by artificial infection test. The outer membrane protein Aha1 is a major adhesin of A. hydrophila and also highly conserved in different serotypes of A. hydrophila. In order to ascertain the conservation of aha1 protein among mesophilic motile aeromonads, full length aha1 gene from all isolates was detected, cloned and sequenced. As the results show, the aha1 genes were amplified in all strains and the ORF size of the aha1 gene from A. hydrophila and other phenotypic species of aeromonas isolates was 1,068 and 1,038 bp, respectively. Four Anhui A. hydrophila isolates and six A. hydrophila reference strains formed a cluster together with 91.4-99.7% nucleotide identity and 91.9-99.7% amino acid identity of the aha1 gene. Five Anhui other phenotypic species isolates formed another cluster, they shared 79.5-81.1% nucleotide identity and 79.6-81.6% amino acid identity of the aha1 gene compared with A. hydrophila and the major sequence variations were observed between amino acids 85-134, 176- 227, 243-263, 280-295 and 321-336.