Detection of Yersinia ruckeri in rainbow trout blood by use of the polymerase chain reaction.
We evaluated a polymerase chain reaction (PCR) method for detecting Yersinia ruckeri, the bacterial pathogen causing enteric redmouth disease (ERM), in blood of rainbow trout Oncorhynchus mykiss. Identification of the PCR product was confirmed by Southern blot hybridization with a 32P-labeled oligonucleotide probe matching a sequence within the small subunit ribosomal RNA gene of Y. ruckeri. Following a 1 h immersion of rainbow trout in water with 4.5 x 10(6) colony-forming units of Y. ruckeri l(-1), the PCR was positive for all blood samples from 1 h (first sample) to 5 d and was negative from 9 to 30 d (last sample). Fish in this experiment did not show signs of disease, probably because they had been vaccinated against Y. ruckeri. To test this method with naturally infected fish, 42 rainbow trout from hatcheries were examined. Four of these fish had clinical signs of ERM and were infected with Y. ruckeri based on bacteriological culture. The PCR method detected Y. ruckeri in blood, intestine, liver, and trunk kidney from the 4 fish with ERM and from 5 additional rainbow trout that were bacteriologically negative for Y. ruckeri. Three of 5 rainbow trout from streams receiving effluent from hatcheries were positive for Y. ruckeri when tested with PCR, although there was no growth of Y. ruckeri on culture plates inoculated with the same samples. Samples were successfully stored for 1 wk in lysis buffer at 25 degrees C. This study demonstrated that a non-lethal blood sample can be used with PCR to detect Y. ruckeri.
Altinok
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Department of Fisheries and Allied Aquacultures, Auburn University, Alabama 36849, USA.
Grizzle
J M
JM
Liu
Z
Z
D000818:Animals; D017756:Aquaculture; D001431:Bacteriological Techniques / Q000662:veterinary; D015139:Blotting, Southern / Q000662:veterinary; D004269:DNA, Bacterial / Q000302:isolation & purification; D005393:Fish Diseases / Q000175:diagnosis* / Q000382:microbiology; D017686:Oncorhynchus mykiss / Q000097:blood / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D013048:Specimen Handling / Q000662:veterinary; D015007:Yersinia / Q000302:isolation & purification*; D015009:Yersinia Infections / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*
D004269:DNA, Bacterial