PCR detection of potentially pathogenic aeromonads in raw and cold-smoked freshwater fish.
OBJECTIVE
Development of a PCR assay for detection of aeromonads carrying the hlyA and/or aerA genes in fish.


RESULTS
The protocol involves an overnight selective enrichment step in tryptic soy broth yeast extract containing 10 microg ml(-1) of ampicillin followed by extraction of DNA and PCR amplification of two haemolysin genes that contribute to the virulence of Aer. hydrophila. This procedure can detect initial populations of 1-10 cfu g(-1) within 24 h in artificially contaminated samples. In naturally contaminated fish, both genes were detected in 13 out of 14 fresh fish lots (aeromonads levels between < 1 and 5.42 log cfu g(-1)) and in 4 out of 16 lots of vacuum-packed cold-smoked fish (aeromonads levels between < 1 and 3.37 log cfu g(-1)). Before enrichment, dominant species were Aer. hydrophila HG1 (aerA+hlyA+), Aer. bestiarum HG2 (aerA+hlyA+) and Aer. caviae HG4 (aerA-hlyA-). After enrichment, Aer. hydrophila HG1 (aerA+hlyA+) was dominant.


CONCLUSIONS
Fresh fish and even smoked fish carry hlyA+ and/or aerA+ aeromonads that can be detected by PCR within 24 h.


CONCLUSIONS
The PCR assay described offers considerable potential as a rapid method with specificity, sensitivity and simplicity.
González-Rodríguez
M N
MN
Department of Food Hygiene and Food Technology, University of León, 24071-León, Spain.
Santos
J A
JA
Otero
A
A
García-López
M L
ML
D000333:Aeromonas / Q000145:classification / Q000235:genetics / Q000302:isolation & purification* / Q000472:pathogenicity*; D000818:Animals; D003080:Cold Temperature; D015169:Colony Count, Microbial; D004269:DNA, Bacterial / Q000032:analysis; D005393:Fish Diseases / Q000382:microbiology; D005511:Food Handling / Q000379:methods; D005618:Fresh Water; D016905:Gram-Negative Bacterial Infections / Q000382:microbiology / Q000662:veterinary*; D006460:Hemolysin Proteins / Q000235:genetics; D017686:Oncorhynchus mykiss / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D020317:Salmo salar / Q000382:microbiology*; D012680:Sensitivity and Specificity; D014618:Vacuum; D014774:Virulence
D004269:DNA, Bacterial; D006460:Hemolysin Proteins