Diagnostic accuracy of pathogen detection depends upon the selection of suitable tests. Problems can arise when the selected diagnostic test gives false-positive or false-negative results, which can affect control measures, with consequences for the population health. The aim of this study was to compare sensitivity of different diagnostic methods IHC, PCR and qPCR detecting Tetracapsuloides bryosalmonae, the causative agent of proliferative kidney disease in salmonid fish and as a consequence differences in disease prevalence. We analysed tissue from 388 salmonid specimens sampled from a recirculating system and rivers in the Czech Republic. Overall prevalence of T. bryosalmonae was extremely high at 92.0%, based on positive results of at least one of the above-mentioned screening methods. IHC resulted in a much lower detection rate (30.2%) than both PCR methods (qPCR32: 65.4%, PCR: 81.9%). While qPCR32 produced a good match with IHC (60.8%), all other methods differed significantly (p < .001) in the proportion of samples determined positive. Both PCR methods showed similar sensitivity, though specificity (i.e., the proportion of non-diseased fish classified correctly) differed significantly (p < .05). Sample preservation method significantly (p < .05) influenced the results of PCR, with a much lower DNA yield extracted from paraffin-embedded samples. Use of different methods that differ in diagnostic sensitivity and specificity resulted in random and systematic diagnosis errors, illustrating the importance of interpreting the results of each method carefully.

Department of Ecology and Diseases of Zoo Animals Game Fish and Bees Faculty of Veterinary Hygiene and Ecology University of Veterinary and Pharmaceutical Sciences Brno Brno Czech Republic

Department of Infectious Diseases and Preventive Medicine Veterinary Research Institute Brno Czech Republic

Department of Pathobiology Centre for Fish and Wildlife Health University of Bern Bern Switzerland

Department of Zoology Fisheries Hydrobiology and Apiculture Mendel University in Brno Brno Czech Republic

Institute of Vertebrate Biology Czech Academy of Sciences Brno Czech Republic

Zobrazit více v PubMed

Abd‐Elfattah, A., Kumar, G., Soliman, H., & El‐Matbouli, M. (2014). Persistence of PubMed DOI PMC

Adams, A., Richards, R. H., & de Mateo, M. M. (1992). Development of monoclonal antibodies to PK ‘X’, the causative agent of proliferative kidney disease. Journal of Fish Diseases, 15(6), 515–521. 10.1111/j.1365-2761.1992.tb00683.x DOI

Bettge, K., Segner, H., Burki, R., Schmidt‐Posthaus, H., & Wahli, T. (2009). Proliferative kidney disease (PKD) of rainbow trout: Temperature‐and time‐related changes of PubMed

Bettge, K., Wahli, T., Segner, H., & Schmidt‐Posthaus, H. (2009). Proliferative kidney disease in rainbow trout: Time‐and temperature‐related renal pathology and parasite distribution. Diseases of Aquatic Organisms, 83(1), 67–76. 10.3354/dao01989 PubMed DOI

Bonin, S., Petrera, F., Niccolini, B., & Stanta, G. (2003). PCR analysis in archival postmortem tissues. Molecular Pathology, 56(3), 184. 10.1136/mp.56.3.184 PubMed DOI PMC

Bramwell, N. H., & Burns, B. F. (1988). The effects of fixative type and fixation time on the quantity and quality of extractable DNA for hybridization studies on lymphoid tissue. Experimental Hematology, 16(8), 730–732. PubMed

Canning, E. U., Curry, A., Feist, S. W., Longshaw, M., & Okamura, B. (2000). A new class and order of myxozoans to accommodate parasites of bryozoans with ultrastructural observations on PubMed DOI

Castagnaro, M., Marin, M., Ghittino, C., & Hedrick, R. P. (1991). Lectin histochemistry and ultrastructure of rainbow trout DOI

Chilmonczyk, S., Monge, D., & De Kinkelin, P. (2002). Proliferative kidney disease: Cellular aspects of the rainbow trout DOI

Clifton‐Hadley, R. S., Bucke, D., & Richards, R. H. (1986). Economic importance of proliferative kidney disease of salmonid fish in England and Wales. Veterinary Record, 119(12), 305–306. 10.1136/vr.119.12.305 PubMed DOI

Clifton‐Hadley, R. S., Richards, R. H., & Bucke, D. (1986). Proliferative kidney disease (PKD) in rainbow trout DOI

Feist, S. W., Longshaw, M., Canning, E. U., & Okamura, B. (2001). Induction of proliferative kidney disease (PKD) in rainbow trout PubMed DOI

Ferguson, H. W. (1981). The effects of water temperature on the development of proliferative kidney disease in rainbow trout, DOI

Ferguson, H. W., & Ball, H. J. (1979). Epidemiological aspects of proliferative kidney disease amongst rainbow trout DOI

Ferguson, H. W., & Needham, E. A. (1978). Proliferative kidney disease in rainbow trout DOI

Gay, M., Okamura, B., & De Kinkelin, P. (2001). Evidence that infectious stages of PubMed DOI

Grabner, D. S., & El‐Matbouli, M. (2010). PubMed DOI

Hedrick, R. P., MacConnell, E., & De Kinkelin, P. (1993). Proliferative kidney disease of salmonid fish. Annual Review of Fish Diseases, 3, 277–290. 10.1016/0959-8030(93)90039-E DOI

Hedrick, R. P., Marin, M., Castagnaro, M., Monge, D., & De Kinkelin, P. (1992). Rapid lectin‐based staining procedure for the detection of the myxosporean causing proliferative kidney disease in salmonid fish. Diseases of Aquatic Organisms, 13(2), 129–132. 10.3354/dao013129 DOI

Henderson, M., & Okamura, B. (2004). The phylogeography of salmonid proliferative kidney disease in Europe and North America. Proceedings of the Royal Society of London. Series B: Biological Sciences, 271(1549), 1729–1736. 10.1098/rspb.2004.2677 PubMed DOI PMC

Kent, M. L., & Hedrick, R. P. (1985). PKX, the causative agent of proliferative kidney disease (PKD) in Pacific salmonid fishes and its affinities with the myxozoa 1. The Journal of Protozoology, 32(2), 254–260. 10.1111/j.1550-7408.1985.tb03047.x PubMed DOI

Kent, M. L., Khattra, J., Hervio, D. M. L., & Devlin, R. H. (1998). Ribosomal DNA sequence analysis of isolates of the PKX myxosporean and their relationship to members of the genus Sphaerospora. Journal of Aquatic and Animal Health, 10, 12–21. 10.1577/1548-8667(1998)010<0012:RDSAOI>2.0.CO;2 DOI

Klontz, G. W., & Chacko, A. J. (1983). Methods to detect the organism causing proliferative kidney disease in salmonids. Bulletin of the European Association of Fish Pathologists, 3, 33–36.

Kopp, R., Palíková, M., Papežíková, I., Mareš, J., Navrátil, S., Pikula, J., & Pohanka, M. (2018). Oxidative stress response of rainbow trout ( DOI

Lewisch, E., Unfer, G., Pinter, K., Bechter, T., & El‐Matbouli, M. (2018). Distribution and prevalence of PubMed

Longshaw, M., Feist, S. W., Canning, E. U., & Okamura, B. (1999). First identification of PKX in bryozoans from the United Kingdom‐molecular evidence. Bulletin‐European Association of Fish Pathologists, 19, 146–148.

Longshaw, M., Le Deuff, R. M., Harris, A. F., & Feist, S. W. (2002). Development of proliferative kidney disease in rainbow trout, DOI

Morris, D. C., Morris, D. J., & Adams, A. (2002). Molecular evidence of release of DOI

Nowak, B., Mueffling, T. V., Caspari, K., & Hartung, J. (2006). Validation of a method for the detection of virulent PubMed DOI

Okamura, B., Anderson, C. L., Longshaw, M., Feist, S. W., & Canning, E. U. (2001). Patterns of occurrence and 18S rDNA sequence variation of PKX ( PubMed

Palikova, M., Papezikova, I., Markova, Z., Navratil, S., Mares, J., Mares, L., Vojtek, L., Hyrsl, P., Jelinkova, E., & Schmidt‐Posthaus, H. (2017). Proliferative kidney disease in rainbow trout ( PubMed

Pojezdal, Ľ., Adamek, M., Syrová, E., Steinhagen, D., Minářová, H., Papežíková, I., Seidlová, V., Reschová, S., & Palíková, M. (2020). Health surveillance of wild brown trout ( PubMed PMC

Ribeiro, C. N. M., Peres, L. C., & Pina‐Neto, J. M. (2004). DNA extraction and quantification from touch and scrape preparations obtained from autopsy liver cells. Brazilian journal of medical and biological research, 37(5), 635–642. PubMed

Rüssmann, H., Kempf, V. A., Koletzko, S., Heesemann, J., & Autenrieth, I. B. (2001). Comparison of fluorescent in situ hybridization and conventional culturing for detection of PubMed DOI PMC

Schmidt‐Posthaus, H., Bettge, K., Forster, U., Segner, H., & Wahli, T. (2012). Kidney pathology and parasite intensity in rainbow trout PubMed DOI

Sengüven, B., Baris, E., Oygur, T., & Berktas, M. (2014). Comparison of methods for the extraction of DNA from formalin‐fixed, paraffin‐embedded archival tissues. International Journal of Medical Sciences, 11(5), 494. 10.7150/ijms.8842 PubMed DOI PMC

Skovgaard, A., & Buchmann, K. (2012). PubMed DOI

Sterud, E., Forseth, T., Ugedal, O., Poppe, T. T., Jørgensen, A., Bruheim, T., Fjeldstad, H.‐P., & Mo, T. A. (2007). Severe mortality in wild Atlantic salmon PubMed

Suresh, K., & Smith, H. (2004). Comparison of methods for detecting PubMed DOI

Syrová, E., Palíková, M., Mendel, J., Seidlová, V., Papežíková, I., Schmidt‐Posthaus, H., Somerlíková, K., Minářová, H., Mareš, L., Mikulíková, I., Pikula, J., & Mareš, J. (2020). Field study indicating susceptibility differences between salmonid species and their lineages to proliferative kidney disease. Journal of Fish Diseases, 43(10), 1201–1211. 10.1111/jfd.13221 PubMed DOI

Tops, S., Lockwood, W., & Okamura, B. (2006). Temperature‐driven proliferation of PubMed DOI

Vasemägi, A., Nousiainen, I., Saura, A., Vähä, J. P., Valjus, J., & Huusko, A. (2017). First record of proliferative kidney disease agent PubMed DOI

Whyte, P., Mc Gill, K., Collins, J. D., & Gormley, E. (2002). The prevalence and PCR detection of salmonella contamination in raw poultry. Veterinary Microbiology, 89(1), 53–60. 10.1016/S0378-1135(02)00160-8 PubMed DOI

Nephrocalcinosis in farmed salmonids: diagnostic challenges associated with low performance and sporadic mortality