Development of a simple and highly sensitive method for the concentration and detection of African swine fever virus in oral fluids and its feasibility assessment on raised pigs in Northern Vietnam

Mai Thi, N.; Tran Thi Huong, G.; Dong Van, H.; Le Van, P.; Huynh Thi My, L.; Bui Tran Anh, D.; Uemura, R.; Yamazaki, Y.; Nguyen Thi, L.; Yamazaki, W.

doi:10.1101/2024.01.13.575502

bioRxiv · 10.1101/2024.01.13.575502

Development of a simple and highly sensitive method for the concentration and detection of African swine fever virus in oral fluids and its feasibility assessment on raised pigs in Northern Vietnam

Mai Thi, N.·Tran Thi Huong, G.·Dong Van, H.·Le Van, P.·Huynh Thi My, L.·Bui Tran Anh, D.·Uemura, R.·Yamazaki, Y.·Nguyen Thi, L.·Yamazaki, W.

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Abstract

BackgroundEarly detection and early slaughter through quarantine are essential to prevent the spread of the African swine fever virus (ASFV). Highly accurate testing is effective for early detection, but it is still difficult to establish a system, especially in the Global South. Pooled oral fluids tests have been used for simple pathogen monitoring, but compared to blood tests, the virus concentration in oral fluids is low, resulting in false negative and missing true positive cases. We collected oral fluids from raised pigs in northern Vietnam and attempted a highly sensitive ASFV survey using a newly developed virus concentration and detection method. ResultsIn a spike test, the developed method showed up to 100 times greater sensitivity than a reference method. To compare and evaluate the performance of the developed method, a total of 68 pooled oral fluid samples were collected, 63 from northern Vietnam and 5 from southern Japan. Using real-time PCR, 9/68 (13.2%) were positive by the reference method, and 23/68 (33.8%) by the developed method. Using real-time LAMP, 1/68 (1.5%) were positive by the reference method and 6/68 (8.8%) by the developed method. ConclusionsThe developed method improved the sensitivity of ASFV detection from oral fluids and enabled early diagnosis of pigs before the onset of the disease. The developed method has the potential to enable simple and highly sensitive diagnosis of ASF, which is essential for its early diagnosis and effective surveillance. HighlightO_LIA method has been developed to detect ASFV in pig oral fluids with up to 100 times greater sensitivity than a reference method. C_LIO_LIThe developed method combines innovative pretreatment in less than 60 min with conventional nucleic acid extraction with a column kit followed by real-time PCR or LAMP. C_LIO_LIApplied to feasibility assessment for ASF diagnosis on raised pigs, the developed method showed higher diagnostic sensitivity than the reference method. C_LIO_LIThe developed method can be a useful tool that contributes to early detection and surveillance of ASFV. C_LI

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Mai Thi, N., Tran Thi Huong, G., Dong Van, H., Le Van, P., Huynh Thi My, L., Bui Tran Anh, D., Uemura, R., Yamazaki, Y., Nguyen Thi, L., Yamazaki, W.. 2024-01-14. Development of a simple and highly sensitive method for the concentration and detection of African swine fever virus in oral fluids and its feasibility assessment on raised pigs in Northern Vietnam. https://doi.org/10.1101/2024.01.13.575502

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