Abstract:
:Three species of blood fluke from the genus Cardicola are known to parasitize and cause disease in Bluefin Tunas--C. forsteri, C. orientalis, and C. opisthorchis. Although initially believed to be separated by geography and host specificity, recent identification of at least two Cardicola spp. concurrently present within all three Bluefin species has raised questions concerning pathogenicity, relative abundance, and distribution of these parasites within Bluefin populations. Here, we present sensitive and differential real-time qPCR nucleic acid detection of these Cardicola spp. by targeting the ITS2 region of the parasite rDNA for PCR amplification. A limit of sensitivity of 1-5 genome copy equivelents was achieved for each of the three Cardicola species tested without cross-species or host genomic amplification. Similar sensitivity was further achieved in the presence of up to 20 ng/μL non-target host gDNA using SYBR Green chemistry alone, or in the presence of up to 160 ng/μL host gDNA through the utilization of a TaqMan probe common-reporter detection system. These methods were subsequently used to positively identify both C. forsteri and C. orientalis DNA in preserved samples of serum, gill, and heart from ranched Southern Bluefin Tuna Thunnus maccoyii. Both methods were more sensitive for positively and differentially identifying the presence of Cardicola spp. than either histological or heart-flush microscopy techniques previously employed, and also possess the ability to be applied in non-lethal blood sampling of these highly valued fish. This is the first report for rapid and differential molecular quantitative detection of Cardicola, and opens the potential for effective monitoring of infection in cultured bluefin populations. Further, it is anticipated that the use of SYBR Green for melt-curve analyses in conjunction with a common-reporter TaqMan assay will present a flexible, accurate, and cost-effective approach for differential detection of a variety of other pathogens in future.
journal_name
Mol Biochem Parasitoljournal_title
Molecular and biochemical parasitologyauthors
Polinski M,Hamilton DB,Nowak B,Bridle Adoi
10.1016/j.molbiopara.2013.07.002subject
Has Abstractpub_date
2013-09-01 00:00:00pages
7-15issue
1eissn
0166-6851issn
1872-9428pii
S0166-6851(13)00114-Xjournal_volume
191pub_type
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journal_title:Molecular and biochemical parasitology
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journal_title:Molecular and biochemical parasitology
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journal_title:Molecular and biochemical parasitology
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journal_title:Molecular and biochemical parasitology
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journal_title:Molecular and biochemical parasitology
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journal_title:Molecular and biochemical parasitology
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章,评审
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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journal_title:Molecular and biochemical parasitology
pub_type: 杂志文章
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journal_title:Molecular and biochemical parasitology
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更新日期:2001-11-01 00:00:00