Development and application of reverse transcriptase droplet digital PCR technology for sensitive detection of BVDV-1 and BTV in bovine semen
Abstract Background Bovine viral diarrhoea virus genotype 1 (BVDV-1) and bluetongue virus (BTV) are potent viral pathogens that may be transmitted through semen, resulting in the spread of diseases via artificial insemination. Thus, establishing an early detection method for BVDV-1 and BTV infection...
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2025-01-01
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| Online Access: | https://doi.org/10.1186/s12917-025-04506-4 |
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| author | Zhichao Yu Linjun Chen Qiang Cui Han Yan Junyan Li Xiaoping Luo Yingying Li Xianghong Ju Yanhong Yong Namula Zhao Zhiguo Zhao |
| author_facet | Zhichao Yu Linjun Chen Qiang Cui Han Yan Junyan Li Xiaoping Luo Yingying Li Xianghong Ju Yanhong Yong Namula Zhao Zhiguo Zhao |
| author_sort | Zhichao Yu |
| collection | DOAJ |
| description | Abstract Background Bovine viral diarrhoea virus genotype 1 (BVDV-1) and bluetongue virus (BTV) are potent viral pathogens that may be transmitted through semen, resulting in the spread of diseases via artificial insemination. Thus, establishing an early detection method for BVDV-1 and BTV infection is important for the trading of semen. In this study, we developed two RT‒ddPCR methods to detect BVDV-1 and BTV, and each method was evaluated for repeatability, limit of detection and specificity. The sensitivity of these methods was compared with that of RT‒qPCR (WOAH) by analysing clinical samples. Results The RT‒ddPCR results revealed that both methods exhibited good repeatability at low concentrations, with detection limits of 1.05 copies/µL and 0.662 copies/µL per reaction for BVDV-1 and BTV, respectively; additionally, both methods exhibited high specificity and did not exhibit cross-reaction with other important semen-transmitted pathogens. Eighty bovine semen samples and twenty mixed semen samples were tested. The results revealed that the positivity rates of BVDV-1 and BTV RT‒ddPCR (25% and 23%, respectively) were greater than those of RT‒qPCR (19% and 18%, respectively). Conclusions RT‒ddPCR was highly sensitive for detecting low concentrations of BVDV-1 and BTV in clinical samples and could be a good supplement for qPCR testing. |
| format | Article |
| id | doaj-art-c81f7cc6fcdc44eb8804493962911973 |
| institution | Kabale University |
| issn | 1746-6148 |
| language | English |
| publishDate | 2025-01-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Veterinary Research |
| spelling | doaj-art-c81f7cc6fcdc44eb88044939629119732025-02-02T12:29:16ZengBMCBMC Veterinary Research1746-61482025-01-012111810.1186/s12917-025-04506-4Development and application of reverse transcriptase droplet digital PCR technology for sensitive detection of BVDV-1 and BTV in bovine semenZhichao Yu0Linjun Chen1Qiang Cui2Han Yan3Junyan Li4Xiaoping Luo5Yingying Li6Xianghong Ju7Yanhong Yong8Namula Zhao9Zhiguo Zhao10Department of Veterinary Medicine, College of Coastal Agricultural Sciences, Guangdong Ocean UniversityTechnology Center, Hohhot Customs DistrictTechnology Center, Hohhot Customs DistrictTechnology Center, Hohhot Customs DistrictKey Laboratory of Grass-Feeding Livestock Healthy Breeding and Livestock Product Quality Control, Veterinary Research Institute, Inner Mongolia Academy of Agricultural and Animal Husbandry SciencesKey Laboratory of Grass-Feeding Livestock Healthy Breeding and Livestock Product Quality Control, Veterinary Research Institute, Inner Mongolia Academy of Agricultural and Animal Husbandry SciencesAnimal and Plant Quarantine and Animal Disease Prevention and Control CenterDepartment of Veterinary Medicine, College of Coastal Agricultural Sciences, Guangdong Ocean UniversityDepartment of Veterinary Medicine, College of Coastal Agricultural Sciences, Guangdong Ocean UniversityDepartment of Veterinary Medicine, College of Coastal Agricultural Sciences, Guangdong Ocean UniversityTechnology Center, Hohhot Customs DistrictAbstract Background Bovine viral diarrhoea virus genotype 1 (BVDV-1) and bluetongue virus (BTV) are potent viral pathogens that may be transmitted through semen, resulting in the spread of diseases via artificial insemination. Thus, establishing an early detection method for BVDV-1 and BTV infection is important for the trading of semen. In this study, we developed two RT‒ddPCR methods to detect BVDV-1 and BTV, and each method was evaluated for repeatability, limit of detection and specificity. The sensitivity of these methods was compared with that of RT‒qPCR (WOAH) by analysing clinical samples. Results The RT‒ddPCR results revealed that both methods exhibited good repeatability at low concentrations, with detection limits of 1.05 copies/µL and 0.662 copies/µL per reaction for BVDV-1 and BTV, respectively; additionally, both methods exhibited high specificity and did not exhibit cross-reaction with other important semen-transmitted pathogens. Eighty bovine semen samples and twenty mixed semen samples were tested. The results revealed that the positivity rates of BVDV-1 and BTV RT‒ddPCR (25% and 23%, respectively) were greater than those of RT‒qPCR (19% and 18%, respectively). Conclusions RT‒ddPCR was highly sensitive for detecting low concentrations of BVDV-1 and BTV in clinical samples and could be a good supplement for qPCR testing.https://doi.org/10.1186/s12917-025-04506-4Bovine viral diarrhoea virus genotype 1Bluetongue virusDroplet digital PCRBovine semen |
| spellingShingle | Zhichao Yu Linjun Chen Qiang Cui Han Yan Junyan Li Xiaoping Luo Yingying Li Xianghong Ju Yanhong Yong Namula Zhao Zhiguo Zhao Development and application of reverse transcriptase droplet digital PCR technology for sensitive detection of BVDV-1 and BTV in bovine semen BMC Veterinary Research Bovine viral diarrhoea virus genotype 1 Bluetongue virus Droplet digital PCR Bovine semen |
| title | Development and application of reverse transcriptase droplet digital PCR technology for sensitive detection of BVDV-1 and BTV in bovine semen |
| title_full | Development and application of reverse transcriptase droplet digital PCR technology for sensitive detection of BVDV-1 and BTV in bovine semen |
| title_fullStr | Development and application of reverse transcriptase droplet digital PCR technology for sensitive detection of BVDV-1 and BTV in bovine semen |
| title_full_unstemmed | Development and application of reverse transcriptase droplet digital PCR technology for sensitive detection of BVDV-1 and BTV in bovine semen |
| title_short | Development and application of reverse transcriptase droplet digital PCR technology for sensitive detection of BVDV-1 and BTV in bovine semen |
| title_sort | development and application of reverse transcriptase droplet digital pcr technology for sensitive detection of bvdv 1 and btv in bovine semen |
| topic | Bovine viral diarrhoea virus genotype 1 Bluetongue virus Droplet digital PCR Bovine semen |
| url | https://doi.org/10.1186/s12917-025-04506-4 |
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