Advanced Medical and Dental Institute, Universiti Sains Malaysia, Bertam, Penang, Malaysia.
Institute of Experimental Pathology (ZMBE), University of Münster, Münster, Germany.
Am J Trop Med Hyg. 2019 Jun;100(6):1328-1334. doi: 10.4269/ajtmh.18-0525.
The diarrheal disease "cholera" is caused by , and is primarily confined to endemic regions, mostly in Africa and Asia. It is punctuated by outbreaks and creates severe challenges to public health. The disease-causing strains are most-often members of serogroups O1 and O139. PCR-based methods allow rapid diagnosis of these pathogens, including the identification of their biotypes. However, this necessitates the selection of specific target sequences to differentiate even the closely related biotypes of . Oligonucleotides for selective amplification of small RNA (sRNA) genes that are specific to these subtypes were designed. The resulting multiplex PCR assay was validated using cultures (i.e., 19 and 22 non- isolates) and spiked stool samples. The validation using cultures and spiked stool suspensions revealed detection limits of 10-100 pg DNA per reaction and 1.5 cells/mL suspension, respectively. The multiplex PCR assay that targets sRNA genes for amplification enables the sensitive and specific detection, as well as the differentiation of -O1 classical, O1 El Tor, and O139 biotypes. Most importantly, the assay enables fast and cheaper diagnosis compared with classic culture-based methods.
腹泻病“霍乱”由 引起，主要局限于流行地区，主要在非洲和亚洲。它由爆发引起，并对公共卫生造成严重挑战。致病菌株大多是 O1 和 O139 血清群的成员。基于 PCR 的方法可快速诊断这些病原体，包括鉴定其生物型。然而，这需要选择特定的靶序列来区分甚至是密切相关的 生物型。设计了用于选择性扩增特定于这些 亚型的小 RNA(sRNA)基因的寡核苷酸。使用 培养物（即 19 个和 22 个非分离株）和加标粪便样本对所得多重 PCR 检测进行了验证。使用 培养物和加标粪便悬液进行的验证显示，每个反应的检测限分别为 10-100 pg DNA 和 1.5 个细胞/mL 悬浮液。针对 sRNA 基因进行扩增的多重 PCR 检测可实现灵敏、特异的检测以及 -O1 经典型、O1 El Tor 型和 O139 生物型的区分。最重要的是，与经典的基于培养的方法相比，该检测可实现更快和更廉价的诊断。
