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Novel combinations of mitochondrial DNA (CO1) and internal transcribed spacers of nuclear ribosomal DNA (ITS) were detected among Gyrodactylus parasites on brown trout (Salmo trutta L.), rainbow trout (Oncorhynchus mykiss (Walbaum)), and Ohrid trout (Salmo letnica (Karaman)) from salmonid farms in Poland and Macedonia. Some clones differed from standard ITS only by ≤ 4 nucleotides, but they belonged to a mtDNA clade that differed from the Northern European lineages of G. salaris by d MCL = 0.266 ± 0.108 (maximum composite likelihood distance). The divergence of d MCL = 0.013 ± 0.005 within the alien mtDNA clade suggested introgression from an unknown maternal ancestor into the G. salaris Malmberg genome 137 to 57 kyr ago (or, less probably, repeated introgression). A comparable modern hybrid was detected based on permanently heterozygous ITS (28 bp/1264 = 2.2%) in a clone that is widespread throughout Finnish rainbow trout farms. This was a F1 hybrid of maternal G. pomeraniae Kuusela, Ziętara et Lumme (on roach, Rutilus rutilus (L.)) and G. lavareti Malmberg (on whitefish, Coregonus lavaretus (L.)). The mtDNA of the parental species differed by d MCL = 0.290 ± 0.130. The observations emphasize that both nuclear and maternally-inherited DNA markers are necessary to characterize sexually-produced lineages or clones of Gyrodactylus. The hybridization events demonstrated predict incongruence of mitochondrial vs. nuclear gene trees, i.e., reticulate evolution in species trees.
Three previously undescribed species of wageneri group of Gyrodactylus Nordmann, 1832 (subgenus Limnonephrotus, Gyrodactylidae, Monogenoidea) related to G. lavareti Malmberg, 1957 are described here. G. pomeraniae sp. nov. was found on roach (Rutilus rutilus) in Poland and Belgium, G. ouluensis sp. nov. on roach in Finland and G. salvelini sp. nov. on Arctic charr (Salvelinus alpinus) in the Lake Inari, Finland. A molecular redescription of G. lavareti on Coregonus lavaretus is also presented, and G. bliccensis on Alburnus alburnus from river Morava, Czech Republic is included in the phylogenetic analysis. In addition, a hybrid clone of maternal G. pomeraniae sp. nov. and paternal G. lavareti found on farmed rainbow trout (Oncorhynchus mykiss) is characterized. The molecular species description was based on the complete CO1 gene of the mitochondrial DNA, and on phylogenetic comparison of the internal transcribed spacer segment (ITS1-5.8S rDNA-ITS2) of nuclear ribosomal DNA. The species hosted by cyprinids were basal in the phylogeny rooted by numerous relatives of wageneri-species group.
The Gyrodactylus fauna of 274 fish taken from ten salmonid farms in Poland was sampled in 2006. Four fish species were investigated: rainbow trout Oncorhynchus mykiss, brown trout Salmo trutta (morphs fario, lacustris, and trutta), grayling Thymallus thymallus and huchen Hucho hucho. No parasites were observed on huchen. No indications of gyrodactylosis were observed, but an unexpected parasite species diversity was found. A molecular species identification by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) of ITS1 + 5.8S + ITS2 was utilized, with addition of morphometric methods. The most frequent parasite was a new record in Poland, G. teuchis. It was present in two molecular forms on brown trout and rainbow trout, which also carried G. derjavinoides and G. truttae. Three molecular forms of G. salaris/G. thymalli were found, the standard type ITS only on grayling. A heterozygous (or heterogenic) G. salaris type described earlier in Denmark was found in seven farms on rainbow trout, and a complementary homozygous clone which differs from the standard by three nucleotides, in two farms. This homozygous form has not been recorded earlier. The PCR-RFLP results were confirmed by sequencing ITS segment from representative specimens of each type and comparing them with all available salmonid-specific Gyrodactylus sequences in GenBank. The Polish fauna with seven different Gyrodactylus clones separated by PCR-RFLP was the most diverse reported in fish farms in any country so far.
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