Detection and characterization of the hemolysin genes in Aeromonas hydrophila and Aeromonas sobria by multiplex PCR.
A multiplex PCR assay was designed to amplify the Aeromonas hydrophila and A. veronii bv. sobria hemolysin and aerolysin genes. The assay was evaluated by using 121 clinical isolates and 7 reference strains of Aeromonas spp., and these were divided into five genotypes on the basis of the results of the multiplex PCR. The five genotypes were characterized as type 1 for those carrying the ahh1 gene only (36% of isolates), type 2 for those carrying the asa1 gene only (8.5% of isolates), type 3 for those carrying both the ahh1 and the asa1 genes (4% of isolates), type 4 for those carrying the ahh1 gene and the A. hydrophila aerA (aerolysin) gene (37.5% of isolates), and type 5 for those in which no hemolysin genes were detected (14% of isolates). The most common single hemolysin gene carried among all the Aeromonas isolates examined was ahh1, with 99 of 128 (77%) of isolates testing positive for this gene either alone or in combination with other hemolysin genes. Phenotypic expression of toxins was evaluated in a Vero cell culture cytotoxicity assay. These results indicated that there is a statistically significant correlation between the cytotoxin titers and the hemolysin genotype. Isolates belonging to genotype 4 (carrying both the ahh1 gene and the aerolysin and hemolysin aerA genes) expressed higher cytotoxin titers than isolates of the other genotypes (P < 0.001). These isolates were more cytotoxic in cell culture and may have greater clinical significance.
Wang
Gehua
G
National Laboratory for Enteric Pathogens. National Laboratory for Bacteriology, National Microbiology Laboratory, Winnipeg, Manitoba, Canada. Gahua_Wang@hc-sc.gc.ca
Clark
Clifford G
CG
Liu
Chenyi
C
Pucknell
Chad
C
Munro
Cindy K
CK
Kruk
Tamara M A C
TM
Caldeira
Richard
R
Woodward
David L
DL
Rodgers
Frank G
FG
D016980:Aeromonas hydrophila / Q000235:genetics*; D000818:Animals; D001426:Bacterial Proteins / Q000032:analysis* / Q000235:genetics; D001427:Bacterial Toxins / Q000032:analysis / Q000235:genetics; D001431:Bacteriological Techniques; D002522:Chlorocebus aethiops; D006460:Hemolysin Proteins / Q000032:analysis* / Q000235:genetics; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D052899:Pore Forming Cytotoxic Proteins; D014709:Vero Cells
D001426:Bacterial Proteins; D001427:Bacterial Toxins; D006460:Hemolysin Proteins; D052899:Pore Forming Cytotoxic Proteins; C032676:aerolysin