Development and implementation of a TaqMan triplex real-time PCR assay for concurrent detection of pseudorabies virus, porcine teschovirus 1, and Streptococcus suis 2
IntroductionPorcine neurological disorders represent a prevalent clinical condition that leads to significant mortality and economic losses within the swine industry. Pseudorabies virus (PRV), porcine teschovirus 1 (PTV1), and Streptococcus suis 2 (SS2) are key viral and bacterial pathogens implicat...
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Frontiers Media S.A.
2025-06-01
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| Series: | Frontiers in Veterinary Science |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fvets.2025.1589175/full |
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| author | Ranran Lai Ranran Lai Chen Yang Chen Yang Lili Wu Lili Wu Lili Wu Weisheng Wu Lulu Li Wei Liu Zheng Yan Diankun Yu Shengzhi Ren Zhiqiang Hu Xiaowen Li Xiaowen Li Xiaowen Li Xiaowen Li |
| author_facet | Ranran Lai Ranran Lai Chen Yang Chen Yang Lili Wu Lili Wu Lili Wu Weisheng Wu Lulu Li Wei Liu Zheng Yan Diankun Yu Shengzhi Ren Zhiqiang Hu Xiaowen Li Xiaowen Li Xiaowen Li Xiaowen Li |
| author_sort | Ranran Lai |
| collection | DOAJ |
| description | IntroductionPorcine neurological disorders represent a prevalent clinical condition that leads to significant mortality and economic losses within the swine industry. Pseudorabies virus (PRV), porcine teschovirus 1 (PTV1), and Streptococcus suis 2 (SS2) are key viral and bacterial pathogens implicated in the manifestation of neurological symptoms in pig populations. The overlapping clinical presentations and pathological alterations associated with these pathogens pose challenges in their clinical differentiation. Therefore, it is essential to develop a diagnostic method with high sensitivity and specificity that can simultaneously detect and differentiate these viral and bacterial agents.Materials and methodsA triplex real-time PCR assay using TaqMan probes was developed to simultaneously detect PRV, PTV1, and SS2. To assess the efficacy of the established assay, 30 clinical samples of animals with nervous symptoms were used to compare the results obtained from the triplex real-time PCR assay with those obtained from commercial singleplex real-time PCR kits. Furthermore, a total of 282 samples were tested and analyzed to validate the utility of the assay.ResultsThe triplex real-time PCR assay exhibited high sensitivity, specificity, and repeatability, with a detection limit of 1.0 × 100 copy/μL. The triplex real-time PCR method and commercial singleplex real-time PCR kits showed complete concordance in detecting PRV, PTV1, and SS2. Clinical data indicated single infection rates of 8.16% for PRV, 26.95% for PTV1, and 7.80% for SS2. The observed co-infection rates were 7.45% for PRV + PTV1, 0.71% for PRV + SS2, 1.42% for PTV1 + SS2, and 1.77% for PRV + PTV1 + SS2, respectively.ConclusionThe triplex real-time PCR method developed in this study effectively distinguishes PRV, PTV1, and SS2 simultaneously, serving as a valuable diagnostic tool. This method is anticipated to play a crucial role in preventing and controlling infectious disease spread and supporting epidemiological investigations. |
| format | Article |
| id | doaj-art-f4a9b12eb31d4fc7920e11d91a1fbdda |
| institution | DOAJ |
| issn | 2297-1769 |
| language | English |
| publishDate | 2025-06-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Veterinary Science |
| spelling | doaj-art-f4a9b12eb31d4fc7920e11d91a1fbdda2025-08-20T03:21:35ZengFrontiers Media S.A.Frontiers in Veterinary Science2297-17692025-06-011210.3389/fvets.2025.15891751589175Development and implementation of a TaqMan triplex real-time PCR assay for concurrent detection of pseudorabies virus, porcine teschovirus 1, and Streptococcus suis 2Ranran Lai0Ranran Lai1Chen Yang2Chen Yang3Lili Wu4Lili Wu5Lili Wu6Weisheng Wu7Lulu Li8Wei Liu9Zheng Yan10Diankun Yu11Shengzhi Ren12Zhiqiang Hu13Xiaowen Li14Xiaowen Li15Xiaowen Li16Xiaowen Li17Shandong Engineering Research Center of Pig and Poultry Health Breeding and Important Infectious Disease Purification, Shandong New Hope Liuhe Group Co., Ltd., Qingdao, ChinaJuye Xinhao Agriculture and Animal Husbandry Co., Ltd., Heze, ChinaShandong Engineering Research Center of Pig and Poultry Health Breeding and Important Infectious Disease Purification, Shandong New Hope Liuhe Group Co., Ltd., Qingdao, ChinaJuye Xinhao Agriculture and Animal Husbandry Co., Ltd., Heze, ChinaShandong Engineering Research Center of Pig and Poultry Health Breeding and Important Infectious Disease Purification, Shandong New Hope Liuhe Group Co., Ltd., Qingdao, ChinaJuye Xinhao Agriculture and Animal Husbandry Co., Ltd., Heze, ChinaChina Agriculture Research System-Yangling Comprehensive Test Station, Yangling, ChinaKey Laboratory of Feed and Livestock and Poultry Products Quality & Safety Control, Ministry of Agriculture, New Hope Liuhe Co., Ltd., Chengdu, ChinaJuye Xinhao Agriculture and Animal Husbandry Co., Ltd., Heze, ChinaKey Laboratory of Feed and Livestock and Poultry Products Quality & Safety Control, Ministry of Agriculture, New Hope Liuhe Co., Ltd., Chengdu, ChinaJuye Xinhao Agriculture and Animal Husbandry Co., Ltd., Heze, ChinaKey Laboratory of Feed and Livestock and Poultry Products Quality & Safety Control, Ministry of Agriculture, New Hope Liuhe Co., Ltd., Chengdu, ChinaShandong Engineering Research Center of Pig and Poultry Health Breeding and Important Infectious Disease Purification, Shandong New Hope Liuhe Group Co., Ltd., Qingdao, ChinaKey Laboratory of Animal Epidemic Disease Detection and Prevention in Panxi District, College of Animal Science, Xichang University, Xichang, ChinaShandong Engineering Research Center of Pig and Poultry Health Breeding and Important Infectious Disease Purification, Shandong New Hope Liuhe Group Co., Ltd., Qingdao, ChinaJuye Xinhao Agriculture and Animal Husbandry Co., Ltd., Heze, ChinaChina Agriculture Research System-Yangling Comprehensive Test Station, Yangling, ChinaKey Laboratory of Feed and Livestock and Poultry Products Quality & Safety Control, Ministry of Agriculture, New Hope Liuhe Co., Ltd., Chengdu, ChinaIntroductionPorcine neurological disorders represent a prevalent clinical condition that leads to significant mortality and economic losses within the swine industry. Pseudorabies virus (PRV), porcine teschovirus 1 (PTV1), and Streptococcus suis 2 (SS2) are key viral and bacterial pathogens implicated in the manifestation of neurological symptoms in pig populations. The overlapping clinical presentations and pathological alterations associated with these pathogens pose challenges in their clinical differentiation. Therefore, it is essential to develop a diagnostic method with high sensitivity and specificity that can simultaneously detect and differentiate these viral and bacterial agents.Materials and methodsA triplex real-time PCR assay using TaqMan probes was developed to simultaneously detect PRV, PTV1, and SS2. To assess the efficacy of the established assay, 30 clinical samples of animals with nervous symptoms were used to compare the results obtained from the triplex real-time PCR assay with those obtained from commercial singleplex real-time PCR kits. Furthermore, a total of 282 samples were tested and analyzed to validate the utility of the assay.ResultsThe triplex real-time PCR assay exhibited high sensitivity, specificity, and repeatability, with a detection limit of 1.0 × 100 copy/μL. The triplex real-time PCR method and commercial singleplex real-time PCR kits showed complete concordance in detecting PRV, PTV1, and SS2. Clinical data indicated single infection rates of 8.16% for PRV, 26.95% for PTV1, and 7.80% for SS2. The observed co-infection rates were 7.45% for PRV + PTV1, 0.71% for PRV + SS2, 1.42% for PTV1 + SS2, and 1.77% for PRV + PTV1 + SS2, respectively.ConclusionThe triplex real-time PCR method developed in this study effectively distinguishes PRV, PTV1, and SS2 simultaneously, serving as a valuable diagnostic tool. This method is anticipated to play a crucial role in preventing and controlling infectious disease spread and supporting epidemiological investigations.https://www.frontiersin.org/articles/10.3389/fvets.2025.1589175/fulltriplex real-time PCRporcine nervous diseasesPRVPTV1SS2 |
| spellingShingle | Ranran Lai Ranran Lai Chen Yang Chen Yang Lili Wu Lili Wu Lili Wu Weisheng Wu Lulu Li Wei Liu Zheng Yan Diankun Yu Shengzhi Ren Zhiqiang Hu Xiaowen Li Xiaowen Li Xiaowen Li Xiaowen Li Development and implementation of a TaqMan triplex real-time PCR assay for concurrent detection of pseudorabies virus, porcine teschovirus 1, and Streptococcus suis 2 Frontiers in Veterinary Science triplex real-time PCR porcine nervous diseases PRV PTV1 SS2 |
| title | Development and implementation of a TaqMan triplex real-time PCR assay for concurrent detection of pseudorabies virus, porcine teschovirus 1, and Streptococcus suis 2 |
| title_full | Development and implementation of a TaqMan triplex real-time PCR assay for concurrent detection of pseudorabies virus, porcine teschovirus 1, and Streptococcus suis 2 |
| title_fullStr | Development and implementation of a TaqMan triplex real-time PCR assay for concurrent detection of pseudorabies virus, porcine teschovirus 1, and Streptococcus suis 2 |
| title_full_unstemmed | Development and implementation of a TaqMan triplex real-time PCR assay for concurrent detection of pseudorabies virus, porcine teschovirus 1, and Streptococcus suis 2 |
| title_short | Development and implementation of a TaqMan triplex real-time PCR assay for concurrent detection of pseudorabies virus, porcine teschovirus 1, and Streptococcus suis 2 |
| title_sort | development and implementation of a taqman triplex real time pcr assay for concurrent detection of pseudorabies virus porcine teschovirus 1 and streptococcus suis 2 |
| topic | triplex real-time PCR porcine nervous diseases PRV PTV1 SS2 |
| url | https://www.frontiersin.org/articles/10.3389/fvets.2025.1589175/full |
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