title: Comparative evaluation of molecular diagnostic tests for Nucleospora salmonis and prevalence in migrating juvenile salmonids from the Snake River, USA.
abstract: Nucleospora salmonis is an intranuclear microsporidian that primarily infects lymphoblast cells and contributes to chronic lymphoblastosis and a leukemia-like condition in a range of salmonid species. The primary goal of this study was to evaluate the prevalence of N. salmonis in out-migrating juvenile hatchery and wild Chinook salmon Oncorhynchus tshawytscha and steelhead O. mykiss from the Snake River in the U.S. Pacific Northwest. To achieve this goal, we first addressed the following concerns about current molecular diagnostic tests for N. salmonis: (1) nonspecific amplification patterns by the published nested polymerase chain reaction (nPCR) test, (2) incomplete validation of the published quantitative PCR (qPCR) test, and (3) whether N. salmonis can be detected reliably from nonlethal samples. Here, we present an optimized nPCR protocol that eliminates nonspecific amplification. During validation of the published qPCR test, our laboratory developed a second qPCR test that targeted a different gene sequence and used different probe chemistry for comparison purposes. We simultaneously evaluated the two different qPCR tests for N. salmonis and foundthat both assays were highly specific, sensitive, and repeatable. The nPCR and qPCR tests had good overall concordance when DNA samples derived from both apparently healthy and clinically diseased hatchery rainbow trout were tested. Finally, we demonstrated that gill snips were a suitable tissue for nonlethal detection of N. salmonis DNA in juvenile salmonids. Monitoring of juvenile salmonid fish in the Snake River over a 3-year period revealed low prevalence of N. salmonis in hatchery and wild Chinook salmon and wild steelhead but significantly higher prevalence in hatchery-derived steelhead. Routine monitoring of N. salmonis is not performed for all hatchery steelhead populations. At present, the possible contribution of this pathogen to delayed mortality of steelhead has not been determined.
authors.affiliation: U.S. Geological Survey, Western Fisheries Research Center 6505 Northeast 65th Street, Seattle, Washington 98115, USA.
authors.forename: Samantha
authors.initials: S
authors.lastname: Badil
authors.forename: Diane G
authors.initials: DG
authors.lastname: Elliott
authors.forename: Tomofumi
authors.initials: T
authors.lastname: Kurobe
authors.forename: Ronald P
authors.initials: RP
authors.lastname: Hedrick
authors.forename: Kathy
authors.initials: K
authors.lastname: Clemens
authors.forename: Marilyn
authors.initials: M
authors.lastname: Blair
authors.forename: Maureen K
authors.initials: MK
authors.lastname: Purcell
mesh_terms: D025041:Animal Migration; D000818:Animals; D021901:DNA, Intergenic; D005393:Fish Diseases / Q000175:diagnosis*; D005880:Gills / Q000382:microbiology; D016814:Microsporidia / Q000302:isolation & purification*; D009181:Mycoses / Q000453:epidemiology / Q000382:microbiology / Q000662:veterinary*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary; D015995:Prevalence; D015203:Reproducibility of Results; D045483:Rivers; D012487:Salmonidae; D013997:Time Factors; D014481:United States / Q000453:epidemiology
chemical_list: D021901:DNA, Intergenic