Development of One-Step Real-Time Reverse Transcription Polymerase Chain Reaction Assays for Rapid Detection of Porcine Group C Rotaviruses

Abstract

Abstract. Although the widespread occurrence of porcine group C rotaviruses (GCRV) is assumed, precise prevalence remains largely unknown because of the absence of reliable, specific, and rapid diagnostic methods. To detect and quantify porcine GCRV, the authors evaluated and optimized SYBR Green and TaqMan real-time reverse transcription polymerase chain reaction (RT-PCR) assays and applied them to 108 piglet fecal samples. Using serially diluted standard RNA transcripts of porcine GCRV VP6 gene, both SYBR Green and TaqMan real-time RT-PCR assays detected as few as 1 3 10 1 genome copies/ml (correlation coefficiency .0.99), whereas conventional RT-PCR detected 1.0 3 10 3 copies/ml. In addition, the conventional assay detected porcine GCRV in 24% (26/108) of fecal samples, whereas the detection rates of both SYBR Green and TaqMan assays were 72% (78 of 108) and 64% (70 of 108), respectively. The current study indicated that both real-time RT-PCR assays were reliable, specific, and rapid methods for the detection of porcine GCRV in porcine fecal samples.

The prevalence and genetic diversity of group A rotaviruses on pig farms in the Mekong Delta region of Vietnam

“…Similar high prevalence of ARoVs has been reported from Thailand (22.3%) (Khamrin et al, 2007), Brazil (35.3%) (Rácz et al, 2000), and South Korea (38%) (Chun et al, 2010); whereas other countries have reported much lower prevalence in swine populations (e.g. 4% in Germany; 9.2% in Canada; 3% in Argentina) (Wieler et al, 2001; Morin et al, 1983; Parra et al, 2008).…”

Genetic diversity of the VP7, VP4 and VP6 genes of Korean porcine group C rotaviruses

“…In our previous reports using RT-nested PCR and real-time PCR targeting the VP6 gene of RVCs (Chun et al, 2010;Jeong et al, 2009), porcine RVC infections were found to be widespread in Korean piglets with diarrhea. However, the VP7, VP4 or VP6 genotypes of the infecting Korean porcine RVCs remained largely unknown.…”

Dual priming oligonucleotide (DPO)-based real-time RT-PCR assay for accurate differentiation of four major viruses causing porcine viral diarrhea

“…Currently, polymerase chain reaction (PCR)-based methods have been proven convenient and highly sensitive for detecting porcine diarrhea-associated viruses [20,21]. Of them real-time SYBR Green PCR method that combines conventional PCR and fluorescent signal detection with advantages of high sensitivity, good repeatability, and intuitive results is widely used for epidemiological investigation and pathogen identification [22][23][24]. Moreover, the closed-tube detection of SYBR Green-based real-time PCR assay can effectively prevent aerosol pollution and reduce the incidence of false positives.…”