Molecular assays for detecting Aphanomyces invadans in ulcerative mycotic fish lesions.
The pathogenic oomycete Aphanomyces invadans is the primary etiological agent in ulcerative mycosis, an ulcerative skin disease caused by a fungus-like agent of wild and cultured fish. We developed sensitive PCR and fluorescent peptide nucleic acid in situ hybridization (FISH) assays to detect A. invadans. Laboratory-challenged killifish (Fundulus heteroclitus) were first tested to optimize and validate the assays. Skin ulcers of Atlantic menhaden (Brevoortia tyrannus) from populations found in the Pamlico and Neuse River estuaries in North Carolina were then surveyed. Results from both assays indicated that all of the lesioned menhaden (n = 50) collected in September 2004 were positive for A. invadans. Neither the FISH assay nor the PCR assay cross-reacted with other closely related oomycetes. These results provided strong evidence that A. invadans is the primary oomycete pathogen in ulcerative mycosis and demonstrated the utility of the assays. The FISH assay is the first molecular assay to provide unambiguous visual confirmation that hyphae in the ulcerated lesions were exclusively A. invadans.
Vandersea
Mark W
MW
Center for Coastal Fisheries and Habitat Research, National Ocean Service, National Oceanic Atmospheric Administration, 101 Pivers Island Rd., Beaufort, NC 28516-9722, USA. Mark.W.Vandersea@noaa.gov
Litaker
R Wayne
RW
Yonnish
Bryan
B
Sosa
Emilio
E
Landsberg
Jan H
JH
Pullinger
Chris
C
Moon-Butzin
Paula
P
Green
Jason
J
Morris
James A
JA
Kator
Howard
H
Noga
Edward J
EJ
Tester
Patricia A
PA
D000818:Animals; D044744:Aphanomyces / Q000235:genetics* / Q000302:isolation & purification* / Q000472:pathogenicity; D001483:Base Sequence; D004271:DNA, Fungal / Q000235:genetics; D005393:Fish Diseases / Q000382:microbiology*; D005399:Fishes; D017404:In Situ Hybridization, Fluorescence / Q000379:methods / Q000662:veterinary*; D008969:Molecular Sequence Data; D009181:Mycoses / Q000382:microbiology / Q000662:veterinary*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D012883:Skin Ulcer / Q000382:microbiology / Q000662:veterinary; D013045:Species Specificity
D004271:DNA, Fungal