DAO 75:239-249 (2007)  -  doi:10.3354/dao075239

Development and validation of an RNA- and DNA-based quantitative PCR assay for determination of Kudoa thyrsites infection levels in Atlantic salmon Salmo salar

Valerie A. Funk1,*, Monique Raap2, Ken Sojonky3, Simon Jones2, John Robinson3, Christy Falkenberg2, Kristina M. Miller2

1BC Centre for Aquatic Health Sciences, 621 Island Highway, PO Box 277, Campbell River, British Columbia V9W 5B1, Canada
2Pacific Biological Station, Department of Fisheries and Oceans Canada, 3190 Hammond Bay Road, Nanaimo, British Columbia V9T 6N7, Canada
3Animal Health Centre, Ministry of Agriculture, Fisheries and Food, 1767 Angus Campbell Road, Abbotsford, British Columbia V3G 2M3, Canada

ABSTRACT: Quantitative PCR (QPCR) methods targeting the 18S rDNA gene (DNA QPCR) and cathepsin L mRNA (RNA QPCR) from Kudoa thyrsites (Gilchrist) were developed and compared with histology for determination of K. thyrsites infection levels in Atlantic salmon Salmo salar L. Both QPCR tests were specific, reproducible and sensitive down to 3 copies. DNA QPCR was able to detect lower K. thyrsites infection levels than those detected by RNA QPCR and histology. The higher sensitivity of the DNA-based test compared with the RNA-based test appeared to be biological in nature and suggested that when infection levels were low, there were fewer copies of cathepsin L mRNA than 18S rDNA genes. However, all 3 diagnostic methods were highly correlated. Regression analyses comparing DNA QPCR and histology data from 2 distinct groups of fish showed that the relationship between these 2 diagnostic methods was reproducible. A logistic regression analysis comparing diagnostic data with a visual assessment of post-mortem flesh quality indicated that histology was the single best predictor of flesh quality, followed by DNA QPCR and then RNA QPCR.


KEY WORDS: Kudoa thyrsites · Atlantic salmon · Diagnostics · QPCR · RNA test · DNA test · Histology · Myoliquefaction


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