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用于检测和定量多种麻痹性贝类毒素产生物种的qPCR检测方法

qPCR Assays for the Detection and Quantification of Multiple Paralytic Shellfish Toxin-Producing Species of .

作者信息

Ruvindy Rendy, Bolch Christopher J, MacKenzie Lincoln, Smith Kirsty F, Murray Shauna A

机构信息

Climate Change Cluster, University of Technology Sydney, Sydney, NSW, Australia.

Institute for Marine and Antarctic Studies, University of Tasmania, Launceston, TAS, Australia.

出版信息

Front Microbiol. 2018 Dec 18;9:3153. doi: 10.3389/fmicb.2018.03153. eCollection 2018.

Abstract

Paralytic shellfish toxin producing dinoflagellates have negatively impacted the shellfish aquaculture industry worldwide, including in Australia and New Zealand. Morphologically identical cryptic species of dinoflagellates that may differ in toxicity, in particular, species of the former species complex, co-occur in Australia, as they do in multiple regions in Asia and Europe. To understand the dynamics and the ecological drivers of the growth of each species in the field, accurate quantification at the species level is crucial. We have developed the first quantitative polymerase chain reaction (qPCR) primers for , and new primers targeting , and . We showed that our new primers for are more specific than previously published primer pairs. These assays can be used to quantify planktonic cells and cysts in the water column and in sediment samples with limits of detection of 2 cells/L for the and assays, 2 cells/L and 1 cyst/mg sediment for the assay, and 1 cells/L for the assay, and efficiencies of >90%. We utilized these assays to discriminate and quantify co-occurring , and in samples from the east coast of Tasmania, Australia.

摘要

产生麻痹性贝类毒素的甲藻对全球贝类养殖业产生了负面影响,包括在澳大利亚和新西兰。形态相同但毒性可能不同的甲藻隐种,特别是前一个物种复合体中的物种,在澳大利亚共同出现,在亚洲和欧洲的多个地区也是如此。为了了解野外每个物种生长的动态和生态驱动因素,在物种水平上进行准确的定量至关重要。我们开发了第一种针对[具体物种1]的定量聚合酶链反应(qPCR)引物,以及针对[具体物种2]、[具体物种3]的新引物。我们表明,我们针对[具体物种1]的新引物比以前发表的引物对更具特异性。这些检测方法可用于定量水柱和沉积物样本中的浮游细胞和孢囊,[具体物种1]检测的检测限为2个细胞/升,[具体物种2]检测的检测限为2个细胞/升和1个孢囊/毫克沉积物,[具体物种3]检测的检测限为1个细胞/升,效率均大于90%。我们利用这些检测方法对来自澳大利亚塔斯马尼亚东海岸样本中共存的[具体物种1]、[具体物种2]和[具体物种3]进行了区分和定量。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/594f/6305576/5329c77e7b02/fmicb-09-03153-g001.jpg

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