Detection of Renibacterium salmoninarum in chinook salmon, Oncorhynchus tshawytscha (Walbaum), using quantitative PCR.
A quantitative polymerase chain reaction (QPCR) assay has been developed to detect varying levels of Renibacterium salmoninarum, the causative agent of bacterial kidney disease. This assay allows for the direct enumeration of bacterial DNA or RNA copy number within tissues and body fluids. The assay can be applied non-lethally and can be used to determine whether R. salmoninarum is transcriptionally active. The presence of R. salmoninarum in kidney tissues from 430 chinook salmon collected from five Idaho Fish and Game operated hatcheries was initially evaluated using the widely employed enzyme-linked immunosorbent assay (ELISA) with two sets of Kirkegaard and Perry Laboratories polyclonal antibodies, 'mother batches' 1 and 2. The same tissue samples were then analysed using the novel QPCR assay and the results compared. At moderate to high levels of infection [optical density (OD > 0.5)], ELISA values and estimated DNA copy number were highly correlated (r(2) > 0.80), although correlation to specific antibody batches varied. However, lower ELISA values (OD < 0.5) observed with either antibody batch did not correlate well with the QPCR assay (R(2) <or= 0.43). Negative controls, run concurrently with the PCR assay, did not indicate extraneous DNA contamination in the PCR samples.
Powell
M
M
Center for Salmonid and Freshwater Species at Risk, Hagerman Fish Culture Experiment Station, Hagerman, ID 83332, USA. mpowell@uidaho.edu
Overturf
K
K
Hogge
C
C
Johnson
K
K
D000193:Actinomycetales Infections / Q000235:genetics / Q000662:veterinary*; D000818:Animals; D017756:Aquaculture / Q000379:methods*; D017931:DNA Primers; D004797:Enzyme-Linked Immunosorbent Assay; D005393:Fish Diseases / Q000235:genetics* / Q000382:microbiology*; D007668:Kidney / Q000382:microbiology; D008835:Micrococcaceae / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D012474:Salmon
D017931:DNA Primers