An enzyme linked immunosorbent assay for the detection of Kudoa thyrsites in Atlantic salmon Salmo salar

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

“…The amount of muscle cross-sectional area observed for the histological detection of Kudoa thyrsites plasmodia in Group 1 fish averaged 435 mm 2 , 3 times that observed in previous studies (Dawson-Coates et al 2003, Taylor & Jones 2005 and over twice that observed for the assessment of fish in Groups 2 and 3 in the present study. This surface area represents 9 thin section measuring 7 × 7 mm, and yet this increased effort resulted in only 27% of fish in Group 1 being identified as positive for K. thyrsites compared with 74% diagnosed as positive using the DNA QPCR test.…”

“…There is no definitive test for Kudoa thyrsites or for other pathogens such as Myxobolus cerebralis and Toxoplasma spp., which can be present at low levels and with unpredictable distribution patterns, so that the accuracy of a diagnostic method is linked to test sensitivity, the abundance of the target used for detection and the sampling regimen (Esteban-Redondo et al 1999, Jauregui et al 2001, Dawson-Coates et al 2003, Cavender et al 2004, Taylor & Jones 2005. Evidence was reported herein demonstrating that all 3 diagnostic tests, coupled to the sampling regimens, accurately quantified K. thyrsites infection levels.…”

“…However, histology is labour intensive, requires multiple large pieces of tissue to provide accurate results at low infection levels and cannot be used as a reliable method for detection of plasmodia until approximately 900 degree days post-exposure (Moran et al 1999b). A newly developed ELISA technique has proven to be both sensitive and accurate but is not readily available to all laboratories owing to the requirement of polyclonal antiserum (Taylor & Jones 2005).…”

“…Several methods have been developed for the quantitation of Kuoda thyrsites infections in Atlantic salmon, including histology, spore counts and ELISA (St-Hilaire et al 1997a,b, Moran et al 1999b, DawsonCoates et al 2003, Taylor & Jones 2005. Histological examination of muscle for the presence of plasmodia has been the standard technique for determining K. thyrsites infection levels, and this measure has been shown to correlate well with post-mortem flesh quality (Dawson-Coates et al 2003).…”

“…However, histology is labour intensive, requires multiple large pieces of tissue to provide accurate results at low infection levels and cannot be used as a reliable method for detection of plasmodia until approximately 900 degree days post-exposure (Moran et al 1999b). A newly developed ELISA technique has proven to be both sensitive and accurate but is not readily available to all laboratories owing to the requirement of polyclonal antiserum (Taylor & Jones 2005).QPCR is rapidly replacing traditional diagnostics as the method of choice owing to its sensitivity, specificity, decreased labour, and increased sample throughput as well as its availability and reproducibility among laboratories. QPCR assays have already been successfully developed for the following fish pathogens: Piscirickettsia salmonis, infectious haematopoietic necrosis virus, Edwardsiella ictaturi, Flavo- ), (B) log 10 -transformed DNA QPCR data (relative copy number of K. thyrsites 18S rDNA), (C) log 10 transformed RNA QPCR data (relative copy number of K. thyrsites cathepsin L mRNA).…”