Increased sensitivity in the detection and quantitation of virulence factors of Aeromonas hydrophila and Aeromonas caviae using quantitative PCR

Aeromonas spp., opportunistic pathogens of humans and fish, are ubiquitous in the aquatic environment. Aeromonas hydrophila and Aeromonas caviae have increasingly been isolated from patients with gastroenteritis and superficial wound infections and the former also has been associated with hemorrhagic septicemia in fish. This is the first study to report Aeromonas occurrence and frequency of virulence factors in the aeromonad coastal population and individual isolates in Southern California. In a previous study on the coastal waters of Southern California, we examined Aeromonas levels and associated toxins, hemolytic/cytolytic enterotoxins and serine protease activators, using a most probable number-single PCR assay. This study quantitated copy number of the virulence factors using real-time PCR with TaqMan probes that had a detection limit of 1 to 10 copies. However, variability was greater amongst replicates when copy numbers decreased below 100. We found 22.4% of dilutions that gave negative results by the MPN-single PCR assay became positive using real time. The frequency of the hemolysin/cyctolytic enterotoxins in Aeromonas coastal population ranged from 1 copy in 100 cells to 1 copy in 10,000 cells that carry the trait. The low prevalence of the toxin genes within this aeromonad coastal water population suggests that this microbe might not be a human health risk. However, in individual Aeromonas isolates, 35% and 37% contained the toxin and activator genes, respectively. A higher frequency of the virulence factors was present in warmer months than cooler months. The health risks of bathers from Aeromonas wound infection may still be of concern if these isolates are prevalent in the aeromonad community during warm months.