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首页> 外文期刊>Vector borne and zoonotic diseases >Pyrosequencing Using SL and 5S rRNA as Molecular Markers for Identifying Zoonotic Filarial Nematodes in Blood Samples and Mosquitoes
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Pyrosequencing Using SL and 5S rRNA as Molecular Markers for Identifying Zoonotic Filarial Nematodes in Blood Samples and Mosquitoes

机译:使用SL和5S rRNA作为分子标记的焦磷酸测序鉴定血液样品和蚊子中的人畜共患的线虫线虫

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Backgrounds: Lymphatic filariasis is principally caused by Wuchereria bancrofti, and Brugia malayi. The other two filarial nematode species, Brugia pahangi and Dirofilaria immitis, possibly cause human zoonotic diseases. Methods: We propose the development of a PCR assay linked with DNA pyrosequencing as a rapid tool to identify W. bancrofti, B. malayi, B. pahangi, and D. immitis in blood samples and mosquitoes. Primers targeting the fragment of the 5S ribosomal RNA and spliced leader sequences were newly designed and developed to identify these four filarial nematodes. Analytical sensitivity and specificity were evaluated. Results: Pyrosequencing determination of nucleotide variations within 36 nucleotides for B. malayi and B. pahangi, and 32 nucleotides for W. bancrofti and D. immitis is sufficient for differentiation of those filarial nematodes, and for detection of intraspecies genetic variation of B. malayi. This analysis could detect a single B. malayi, B. pahangi, W. bancrofti, and D. immitis microfilaria in blood samples. Conclusions: Overall, the PCR-linked pyrosequencing-based method was faster than direct sequencing and less expensive than real-time PCR or direct sequencing. This is the possibility of choice that can be applied in a high-throughput platform for identification and surveillance of reservoirs and vectors infected with lymphatic filaria in endemic areas.
机译:背景:淋巴丝虫病主要由Wuchereria bancrofti和Brugia malayi引起。其他两种丝虫线虫,Bugia pahangi和Dirofilaria炎,可能引起人畜共患疾病。方法:我们建议开发一种与DNA焦磷酸测序相关的PCR检测方法,作为鉴定血液样本和蚊子中班氏杆菌,马拉地衣芽孢杆菌,帕汉氏芽孢杆菌和D.炎症的快速工具。新设计并开发了针对5S核糖体RNA片段和剪接前导序列的引物,以鉴定这四个丝状线虫。分析敏感性和特异性进行了评估。结果:焦磷酸测序法测定了马来芽孢杆菌和彭亨氏芽孢杆菌36个核苷酸内的核苷酸变异,班氏菌和班氏梭菌感染的32个核苷酸内的核苷酸变异足以区分那些丝状线虫,并检测马来芽孢杆菌的种内遗传变异。 。这项分析可以检测血样中的单个B. malayi,B。pahangi,W。bancrofti和D. Immitis微丝aria。结论:总的来说,基于PCR的焦磷酸测序方法比直接测序更快,并且比实时PCR或直接测序便宜。这是可以选择在高通量平台上进行识别和监视流行地区淋巴丝虫感染的宿主和载体的可能性。

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