title: Molecular detection and genotyping of Aphanomyces astaci directly from preserved crayfish samples uncovers the Norwegian crayfish plague disease history.
abstract: Aphanomyces astaci causes crayfish plague in European freshwater crayfish, but most historical epizootics lack agent isolation and identification. Although declared as crayfish plague outbreaks by the Norwegian Competent Authorities, only presumptive diagnoses without agent isolation exist from Norwegian epizootics until 2005. Molecular methods now allow both A. astaci detection and genotype determination from preserved samples. We therefore aimed to (1) investigate molecularly if A. astaci was involved in a selection of mass-mortality events in Norwegian noble crayfish populations from 1971 to 2004, and (2) determine the eventually involved A. astaci genotype groups both from these historical and also more recent mass-mortality events. DNA was extracted directly from presumptively infected crayfish tissues, and screened by A. astaci specific qPCR. A representative selection of positive samples was confirmed by ITS-sequencing. Finally, genotype determination was performed with microsatellite markers that distinguish all known A. astaci genotype groups. The molecular examination detected A. astaci in crayfish materials from all examined mass-mortality events. The first event in 1971-1974 was caused by the A. astaci genotype group A, presumably the first genotype group that entered Europe more than 150 years ago. All later outbreaks were caused by the A. astaci genotype group B which was introduced to Europe by importation of signal crayfish in the 1960s. The results suggest that molecular methods can verify the involvement of A. astaci in the vast majority of observed crayfish mass mortalities in Europe whenever preserved materials exist. Moreover, microsatellite genotyping can reveal at least parts of the underlying epidemiology.
authors.affiliation: Norwegian Veterinary Institute, P.O. Box 750, Sentrum, Ullevålsveien 68, N-0106 Oslo, Norway; University of Oslo, Department of Biosciences, P.O. Box 1066, Blindern, NO-0316 Oslo, Norway. Electronic address: trude.vralstad@vetinst.no.
authors.forename: Trude
authors.initials: T
authors.lastname: Vrålstad
authors.affiliation: Norwegian Veterinary Institute, P.O. Box 750, Sentrum, Ullevålsveien 68, N-0106 Oslo, Norway; University of Oslo, Department of Biosciences, P.O. Box 1066, Blindern, NO-0316 Oslo, Norway.
authors.forename: David A
authors.initials: DA
authors.lastname: Strand
authors.affiliation: Universite de Poitiers, UFR Sciences Fondamentales et Appliquées Laboratoire Ecologie et Biologie des Interactions - UMR CNRS 7267, Equipe Ecologie Evolution Symbiose - Batiment B8-B35, 6, rue Michel Brunet, TSA 51106, F-86073 Poitiers cedex 9, France.
authors.forename: Frédéric
authors.initials: F
authors.lastname: Grandjean
authors.affiliation: Norwegian Veterinary Institute, P.O. Box 750, Sentrum, Ullevålsveien 68, N-0106 Oslo, Norway.
authors.forename: Agnar
authors.initials: A
authors.lastname: Kvellestad
authors.affiliation: Norwegian Veterinary Institute, P.O. Box 750, Sentrum, Ullevålsveien 68, N-0106 Oslo, Norway.
authors.forename: Tore
authors.initials: T
authors.lastname: Håstein
authors.affiliation: Norwegian Veterinary Institute, P.O. Box 750, Sentrum, Ullevålsveien 68, N-0106 Oslo, Norway.
authors.forename: Ann Kristin
authors.initials: AK
authors.lastname: Knutsen
authors.affiliation: Norwegian Institute for Nature Research (NINA), Fakkelgården, 2624 Lillehammer, Norway.
authors.forename: Trond
authors.initials: T
authors.lastname: Taugbøl
authors.affiliation: Norwegian Veterinary Institute, P.O. Box 750, Sentrum, Ullevålsveien 68, N-0106 Oslo, Norway.
authors.forename: Ida
authors.initials: I
authors.lastname: Skaar
mesh_terms: D000818:Animals; D044744:Aphanomyces / Q000145:classification / Q000235:genetics* / Q000472:pathogenicity; D003400:Astacoidea / Q000469:parasitology*; D004247:DNA / Q000235:genetics*; D005838:Genotype; D060005:Genotyping Techniques; D049673:History, 20th Century; D049674:History, 21st Century; D018895:Microsatellite Repeats; D009664:Norway; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D004247:DNA
keywords: Aphanomyces astaci; Astacus astacus; Diagnostics; Genotyping; Microsatellites