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Diseases of Aquatic Organisms

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DAO 128:215-224 (2018)  -  DOI: https://doi.org/10.3354/dao03221

Analytical and diagnostic performance of a qPCR assay for Ichthyophonus spp. compared to the tissue culture ‘gold standard’

Vanessa C. Lowe1,*, Paul K. Hershberger2, Carolyn S. Friedman3

1Resource Assessment and Conservation Engineering Division, Alaska Fisheries Science Center, National Marine Fisheries Service, National Oceanic and Atmospheric Administration, 7600 Sand Point Way NE, Seattle, WA 98115, USA
2Marrowstone Marine Field Station, Western Fisheries Research Center, U.S. Geological Survey, 616 Marrowstone Point Rd., Nordland, WA 98358, USA
3School of Aquatic and Fishery Sciences, College of the Environment, University of Washington, 1122 NE Boat St, Seattle, WA 98105, USA
*Corresponding author:

ABSTRACT: Parasites of the genus Ichthyophonus infect many fish species and have a non-uniform distribution within host tissues. Due in part to this uneven distribution, the comparative sensitivity and accuracy of using molecular-based detection methods versus culture to estimate parasite prevalence is under debate. We evaluated the analytical and diagnostic performance of an existing qPCR assay in comparison to the ‘gold standard’ culture method using Pacific herring Clupea pallasii with known exposure history. We determined that the assay is suitable for use in this host, and diagnostic specificity was consistently high (>98%) in both heart and liver tissues. Diagnostic sensitivity could not be fully assessed due to low infection rates, but our results suggest that qPCR is not as sensitive as culture under all circumstances. Diagnostic sensitivity of qPCR relative to culture is likely affected by the amount of sample processed. The prevalence values estimated by the 2 methods were not significantly different when sample amounts were equal (heart tissue), but when the assayed sample amounts were unequal (liver tissue), the culture method detected a significantly higher prevalence of the parasite than qPCR. Further, culture of liver also detected significantly more Ichthyophonus infections than culture of heart, suggesting that the density and distribution of parasites in tissues also plays a role in assay sensitivity. This sensitivity issue would be most problematic for fish with light infections. Although qPCR does not detect the presence of a live organism, DNA-based pathogen detection methods provide the opportunity for alternate testing strategies when culture is not possible.


KEY WORDS: Ichthyophonus · qPCR · Validation · Sensitivity · Specificity · Pacific herring · Clupea pallasii


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Cite this article as: Lowe VC, Hershberger PK, Friedman CS (2018) Analytical and diagnostic performance of a qPCR assay for Ichthyophonus spp. compared to the tissue culture ‘gold standard’. Dis Aquat Org 128:215-224. https://doi.org/10.3354/dao03221

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