Laboratory of Medical Zoology, Department of Microbiology, University of Massachusetts, Amherst, Massachusetts, United States of America.
NH Citizens Health Initiative, Institute for Health Policy and Practice, University of New Hampshire, Concord, New Hampshire, United States of America.
PLoS One. 2022 Jun 1;17(6):e0269266. doi: 10.1371/journal.pone.0269266. eCollection 2022.
Borrelia burgdorferi is an important tickborne human pathogen comprising several strains based on nucleotide sequence of the outer surface protein C (ospC) gene. Detection and characterization of different ospC genotypes is vital for research on B. burgdorferi and the risk it poses to humans. Here we present a novel, multiplex assay based on Luminex xMAP technology for the detection of B. burgdorferi ospC genotypes. The assay has five major steps: amplification of the ospC gene, hydrolyzation of surplus primers and nucleotides, incorporation of biotinylated nucleotides into the template DNA, hybridization to Luminex microspheres, and detection of fluorescent signals corresponding to each ospC genotype. We validated the protocol by comparing results obtained from our method against results from an established ospC genotyping method. This protocol can be used for the characterization of ospC genotypes in B. burgdorferi infected ticks, reservoir hosts, and/or clinical samples.
伯氏疏螺旋体是一种重要的蜱传人类病原体，由基于外表面蛋白 C（ospC）基因的核苷酸序列的几种菌株组成。不同 ospC 基因型的检测和特征对于伯氏疏螺旋体的研究及其对人类的风险至关重要。在这里，我们提出了一种基于 Luminex xMAP 技术的新型多重检测方法，用于检测伯氏疏螺旋体 ospC 基因型。该检测方法有五个主要步骤：ospC 基因的扩增、多余引物和核苷酸的水解、将生物素化核苷酸掺入模板 DNA、与 Luminex 微球杂交，以及检测与每个 ospC 基因型相对应的荧光信号。我们通过将我们的方法的结果与已建立的 ospC 基因分型方法的结果进行比较来验证该方案。该方案可用于鉴定感染蜱、储存宿主和/或临床样本的伯氏疏螺旋体 ospC 基因型。
