副猪绿脓杆菌、缩耳支原体和滑膜支原体多重定量PCR检测方法的建立

IF 3.9 3区 生物学 Q2 MICROBIOLOGY
MicrobiologyOpen Pub Date : 2023-05-15 DOI:10.1002/mbo3.1353
Simone Scherrer, Sarah Schmitt, Fenja Rademacher, Peter Kuhnert, Giovanni Ghielmetti, Sophie Peterhans, Roger Stephan
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引用次数: 1

摘要

副猪绿脓杆菌、缩耳支原体和滑膜支原体是导致多浆液炎、多关节炎、脑膜炎、肺炎和败血症的重要猪病原体,对养猪业造成重大经济损失。设计了一种新的多重定量聚合酶链反应(qPCR),一方面用于检测副猪螺旋体及其毒力标记物vtaA,以区分高毒力和非毒力菌株;另一方面,建立了以16S核糖体RNA基因为靶点的水合棘球蚴和水合棘球蚴荧光探针检测和鉴定方法。基于15个已知副猪螺旋体血清型的参考菌株,以及猪螺旋体螺旋体ATCC 17981T和猪螺旋体螺旋体螺旋体NCTC 10167T型菌株构建qPCR。用21株副猪分枝杆菌、26株猪分枝杆菌和3株猪滑膜分枝杆菌实地分离株进一步评价了新的qPCR。此外,还进行了一项包括42头病猪不同临床标本的初步研究。该方法的特异性为100%,无交叉反应性,不检测其他猪致病菌。结果表明,新qPCR的灵敏度在11-180个基因组当量(GE)之间,对水滑支原体和水滑支原体,对副猪支原体和vtaA的灵敏度在140-1200个基因组当量(GE)之间。截止阈值周期为35。该方法灵敏度高、特异性强,有望成为兽医诊断实验室检测和鉴定副猪螺旋体及其毒力标记物vtaA、猪鼻支原体和猪鼻支原体的有效分子检测工具。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Development of a new multiplex quantitative PCR for the detection of Glaesserella parasuis, Mycoplasma hyorhinis, and Mycoplasma hyosynoviae

Development of a new multiplex quantitative PCR for the detection of Glaesserella parasuis, Mycoplasma hyorhinis, and Mycoplasma hyosynoviae

Glaesserella parasuis, Mycoplasma hyorhinis, and Mycoplasma hyosynoviae are important porcine pathogens responsible for polyserositis, polyarthritis, meningitis, pneumonia, and septicemia causing significant economic losses in the swine industry. A new multiplex quantitative polymerase chain reaction (qPCR) was designed on one hand for the detection of G. parasuis and the virulence marker vtaA to distinguish between highly virulent and non-virulent strains. On the other hand, fluorescent probes were established for the detection and identification of both M. hyorhinis and M. hyosynoviae targeting 16S ribosomal RNA genes. The development of the qPCR was based on reference strains of 15 known serovars of G. parasuis, as well as on the type strains M. hyorhinis ATCC 17981T and M. hyosynoviae NCTC 10167T. The new qPCR was further evaluated using 21 G. parasuis, 26 M. hyorhinis, and 3 M. hyosynoviae field isolates. Moreover, a pilot study including different clinical specimens of 42 diseased pigs was performed. The specificity of the assay was 100% without cross-reactivity or detection of other bacterial swine pathogens. The sensitivity of the new qPCR was demonstrated to be between 11–180 genome equivalents (GE) of DNA for M. hyosynoviae and M. hyorhinis, and 140–1200 GE for G. parasuis and vtaA. The cut-off threshold cycle was found to be at 35. The developed sensitive and specific qPCR assay has the potential to become a useful molecular tool, which could be implemented in veterinary diagnostic laboratories for the detection and identification of G. parasuis, its virulence marker vtaA, M. hyorhinis, and M. hyosynoviae.

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来源期刊
MicrobiologyOpen
MicrobiologyOpen MICROBIOLOGY-
CiteScore
8.00
自引率
0.00%
发文量
78
审稿时长
20 weeks
期刊介绍: MicrobiologyOpen is a peer reviewed, fully open access, broad-scope, and interdisciplinary journal delivering rapid decisions and fast publication of microbial science, a field which is undergoing a profound and exciting evolution in this post-genomic era. The journal aims to serve the research community by providing a vehicle for authors wishing to publish quality research in both fundamental and applied microbiology. Our goal is to publish articles that stimulate discussion and debate, as well as add to our knowledge base and further the understanding of microbial interactions and microbial processes. MicrobiologyOpen gives prompt and equal consideration to articles reporting theoretical, experimental, applied, and descriptive work in all aspects of bacteriology, virology, mycology and protistology, including, but not limited to: - agriculture - antimicrobial resistance - astrobiology - biochemistry - biotechnology - cell and molecular biology - clinical microbiology - computational, systems, and synthetic microbiology - environmental science - evolutionary biology, ecology, and systematics - food science and technology - genetics and genomics - geobiology and earth science - host-microbe interactions - infectious diseases - natural products discovery - pharmaceutical and medicinal chemistry - physiology - plant pathology - veterinary microbiology We will consider submissions across unicellular and cell-cluster organisms: prokaryotes (bacteria, archaea) and eukaryotes (fungi, protists, microalgae, lichens), as well as viruses and prions infecting or interacting with microorganisms, plants and animals, including genetic, biochemical, biophysical, bioinformatic and structural analyses. The journal features Original Articles (including full Research articles, Method articles, and Short Communications), Commentaries, Reviews, and Editorials. Original papers must report well-conducted research with conclusions supported by the data presented in the article. We also support confirmatory research and aim to work with authors to meet reviewer expectations. MicrobiologyOpen publishes articles submitted directly to the journal and those referred from other Wiley journals.
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