Development of a one-step multiplex RT-qPCR method for rapid detection of bovine diarrhea viruses DOI Creative Commons
Dequan Yang, Liyuan Ma,

Zhongping Yang

et al.

Frontiers in Cellular and Infection Microbiology, Journal Year: 2025, Volume and Issue: 14

Published: Jan. 28, 2025

Introduction Viral calf diarrhea poses a significant challenge to the cattle industry worldwide due its high morbidity and mortality rates, leading substantial economic losses. The clinical symptoms associated with various pathogens often overlap, complicating accurate diagnosis; thus, there is an urgent need for rapid precise diagnostic methods improve prevention treatment efforts. In this study, we developed one-step multiplex reverse-transcription quantitative real-time polymerase chain reaction (mRT-qPCR) that enables simultaneous detection of three key viral responsible diarrhea: bovine kobuvirus (BKoV), astrovirus (BoAstV), torovirus (BToV). However, development distinguish these viruses helpful early detection, disease surveillance, control diarrhea. Methods Specific primers minor groove binder (MGB)-based probes were designed targeting 3D region BKoV, ORF1 BoAstV, N BToV. sensitivity, specificity, reproducibility ability evaluated mRT-qPCR. Further, 80 fecal samples subjected mRT-qPCR, results verified using conventional PCR (RT-PCR) or sequencing methods. Results This novel method demonstrated sensitivity specificity,achieving limit 24 copies/mL each pathogen. Furthermore, assay exhibited excellent reproducibility, coefficients variation below 1.5%, strong linear correlation (R 2 > 0.996), amplification efficiency between 90% 110%. Validation from both diarrheic non-diarrheic across four farms in Shanghai showed degree concordance RT-PCR, positive rates BToV at 28.75%, 8.75%, 3.75%, respectively, highlighting predominance BKoV BoAstV. Notably, study represents first identification region. Discussion mRT-qPCR robust, rapid, simple tool identifying diarrhea, facilitating effective measures are vital future sustainability industry.

Language: Английский

Development of a one-step multiplex RT-qPCR method for rapid detection of bovine diarrhea viruses DOI Creative Commons
Dequan Yang, Liyuan Ma,

Zhongping Yang

et al.

Frontiers in Cellular and Infection Microbiology, Journal Year: 2025, Volume and Issue: 14

Published: Jan. 28, 2025

Introduction Viral calf diarrhea poses a significant challenge to the cattle industry worldwide due its high morbidity and mortality rates, leading substantial economic losses. The clinical symptoms associated with various pathogens often overlap, complicating accurate diagnosis; thus, there is an urgent need for rapid precise diagnostic methods improve prevention treatment efforts. In this study, we developed one-step multiplex reverse-transcription quantitative real-time polymerase chain reaction (mRT-qPCR) that enables simultaneous detection of three key viral responsible diarrhea: bovine kobuvirus (BKoV), astrovirus (BoAstV), torovirus (BToV). However, development distinguish these viruses helpful early detection, disease surveillance, control diarrhea. Methods Specific primers minor groove binder (MGB)-based probes were designed targeting 3D region BKoV, ORF1 BoAstV, N BToV. sensitivity, specificity, reproducibility ability evaluated mRT-qPCR. Further, 80 fecal samples subjected mRT-qPCR, results verified using conventional PCR (RT-PCR) or sequencing methods. Results This novel method demonstrated sensitivity specificity,achieving limit 24 copies/mL each pathogen. Furthermore, assay exhibited excellent reproducibility, coefficients variation below 1.5%, strong linear correlation (R 2 > 0.996), amplification efficiency between 90% 110%. Validation from both diarrheic non-diarrheic across four farms in Shanghai showed degree concordance RT-PCR, positive rates BToV at 28.75%, 8.75%, 3.75%, respectively, highlighting predominance BKoV BoAstV. Notably, study represents first identification region. Discussion mRT-qPCR robust, rapid, simple tool identifying diarrhea, facilitating effective measures are vital future sustainability industry.

Language: Английский

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