Institute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China; Key Laboratory of Livestock Disease Prevention of Guangdong Province, Guangzhou 510640, China; Scientific Observation and Experiment Station of Veterinary Drugs and Diagnostic Techniques of Guangdong Province, Ministry of Agriculture, Guanzhou, 510640, China.
Vocational and Technical College, Inner Mongolia Agricultural University, Inner Mongolia 014109, China.
J Virol Methods. 2019 Apr;266:121-125. doi: 10.1016/j.jviromet.2018.12.018. Epub 2019 Jan 9.
This study reports the findings of a high-resolution melting (HRM) curve analysis combined with PCR technique (PCR-HRM) to differentiate between Muscovy duck parvovirus (MDPV) and goose parvovirus (GPV). A degenerate primer set was designed based on the VP3 gene of MDPV and GPV. The PCR HRM assay was able to discriminate between MDPVs and GPVs by differences in melting curve shapes and melting temperatures. A total of forty-five clinical samples, passaged in the allantoic cavity of Muscovy duck eggs, were detected by the PCR-HRM assay. Among the 12 positive samples, two were identified as MDPV and two as GPV with high genotype confidence percentage (GCP) values. Seven positive samples had low GCP values for the melting curve analysis and were identified as co-infection samples. One of the 12 positive samples, designed GDNX strain, was identified as a variant strain with a divergent melting curve profile. To assess the capability of PCR-HRM assay to distinguish MDPV and GPV, fifty-two field samples were collected and examined. Seven samples were positive for MDPV and/or GPV. Thus, this developed assay was useful for discrimination of MDPVs and GPVs and can also be suitable for detecting co-infection samples.
本研究报告了一种高分辨率熔解（HRM）曲线分析与聚合酶链反应（PCR）技术相结合的方法，用于区分麝香鸭细小病毒（MDPV）和鹅细小病毒（GPV）。根据 MDPV 和 GPV 的 VP3 基因设计了一套简并引物。PCR-HRM 检测法能够通过熔解曲线形状和熔解温度的差异区分 MDPV 和 GPV。共检测了 45 份经鸭胚尿囊腔传代的临床样本。通过 PCR-HRM 检测，12 份阳性样本中，有 2 份被鉴定为 MDPV，2 份被鉴定为 GPV，其基因型置信百分比（GCP）值较高。7 份阳性样本的熔解曲线分析 GCP 值较低，被鉴定为混合感染样本。12 份阳性样本中的 1 份，即 GDNX 株，被鉴定为熔解曲线特征不同的变异株。为评估 PCR-HRM 检测法区分 MDPV 和 GPV 的能力，收集了 52 份田间样本进行检测。有 7 份样本为 MDPV 和/或 GPV 阳性。因此，该方法可用于区分 MDPV 和 GPV，也可用于检测混合感染样本。
