Simultaneous differentiation and diagnosis of goose parvovirus and astrovirus in clinical samples with duplex SYBR Green I real-time PCR. (August 2020)

• Record Type: Journal Article
• Authors: Yang, Kan-kan; Xu, Liang; Liang, Yue-qiao; Yin, Dong-dong; Tu, Jian; Song, Xiang-jun; Shao, Ying; Liu, Hong-mei; Qi, Ke-zong
• Is Part Of: Molecular and cellular probes. Volume 52(2020)
• Publication Date: 2020-08
• Subjects: Goose parvovirus -- Goose astrovirus -- SYBR Green I -- Duplex qPCR -- Detection; Molecular probes -- Diagnostic use -- Periodicals; Pathology, Cellular -- Technique -- Periodicals; Cell Biology -- Periodicals; Molecular Biology -- Periodicals; Sondes moléculaires -- Utilisation diagnostique -- Périodiques; Cytopathologie -- Technique -- Périodiques; 572
• DOI: 10.1016/j.mcp.2020.101561
• Languages: English
• ISSNs: 0890-8508

Abstract

Abstract: Two pairs of primers were designed to bind conserved genomic regions of goose parvovirus (GPV) and goose astrovirus (GAstV) to establish a simple, sensitive, and highly specific duplex quantitative PCR (qPCR) method to simultaneously detect the two viruses. The duplex qPCR can distinguish GPV (melting point: 82.1 °C) and GAstV (melting point: 79.8 °C) by the peaks of their individual melting curves. Mixed testing with other waterfowl viruses produced no nonspecific peaks. The established standard curves showed good linear relationships (R 2 > 0.997) and the limits of detection (LOD) for GPV and GAstV were 5.74 × 10 1 and 6.58 × 10 1 copies/μL, respectively. Both intra- and inter-assay coefficients of variation were <2%, indicating that the method has good repeatability. Twenty tissue samples from diseased geese were examined with the duplex qPCR assay and conventional PCR. Duplex qPCR showed positive rates of 25% for GPV and 45% for GAstV, and the positive rate for GPV and GAstV coinfection was 15%, slightly higher than the results for conventional PCR. These results indicated that this duplex qPCR method is highly sensitive, specific, and reproducible, and is suitable for epidemiological studies to effectively control the transmission of GPV and GAstV. Highlights: We established a duplex SYBR green-based real-time qPCR assay to detect GPV and GAstV. The melt curve temperature of GPV and GAstV was 82.1 °C and 79.8 °C. This method has the advantages of high sensitivity, good specificity and reproducibility.