Rapid detection of Listeria monocytogenes sequence type 121 strains using a novel multiplex PCR assay
Rapid detection of Listeria monocytogenes sequence type 121 strains using a novel multiplex PCR assay
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使用新型多重 PCR 检测快速检测单核细胞增生李斯特氏菌序列 121 型菌株
DOI：
10.1016/j.lwt.2019.108474
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发表时间：
2019-12-01
影响因子：
6
通讯作者：
Wu, Qingping
中科院分区：
文献类型：
--
作者：
Chen, Moutong;Cheng, Jianheng;Wu, Qingping
单核细胞增生李斯特菌是一种危险的食源性病原体，常见于食品和食品生产环境中。L.在食品生产环境中的单核细胞增多症序列类型（ST）121是具有挑战性的问题之一。本研究的目的是建立一种多重PCR方法用于L.单核细胞增生性ST 121菌株。通过比较基因组学方法共筛选出50个ST 121特异性候选基因。利用ST 121菌株特异性引物（LM6179_0609和LM6179_0171）和L.单核细胞生成素特异性引物（prfA）。该多重PCR方法的特异性得到了其它乳酸链球菌的有力验证。单核细胞增多症和非L.单核细胞增生菌株。多重PCR的检测限为253 fg/μ L基因组DNA。该多重PCR方法可避免L.在一些实施方案中，微生物以不同比例存在于单核细胞增生症ST 9、ST 8、ST 87和ST 155中，这些是食品中常见的污染物。此外，该多重PCR方法可成功检测2.5 × 10（4）~ 2.5 × 10（0）cfu/10 mL的L.在4-12 h富集后，人工污染牛奶中的单核细胞增多症ST 121。本研究建立的多重PCR方法为特异、快速、准确地检测L.单核细胞增生性ST 121菌株。
Listeria monocytogenes is a hazardous foodborne causative agent, commonly found in foods and food production environments. The persistent contamination of L. monocytogenes sequence type (ST) 121 in food production environments is one of the challenging issues. The objective of this study was to develop a multiplex PCR for identification of L. monocytogenes ST121 strains. A total of 50 specific candidate genes of ST121 were screened by the comparative genomics approach. A novel multiplex PCR for ST121 identification, which comprised of ST121 strain-specific primers (LM6179_0609 and LM6179_0171) and L. monocytogenes-specific primers (prfA), was designed. The specificity of this multiplex PCR was robustly verified by other STs of L. monocytogenes and non-L. monocytogenes strains. The detection limit of the multiplex PCR was 253 fg/mu L genomic DNA. This multiplex PCR could avoid the interference of L. monocytogenes ST9, ST8, ST87, and ST155 in different ratios, which are common contaminants in food items. In addition, this multiplex PCR method could successfully detect 2.5 x 10(4)-2.5 x 10(0) cfu/10 mL of L. monocytogenes ST121 in artificially contaminated milk after 4-12 h enrichment. These results suggested that the newly developed multiplex PCR in this study offers a promising approach for specific, rapid, and accurate detection of L. monocytogenes ST121 strains.