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Title: Development of a reverse transcription loop-mediated isothermal amplification assay for the detection of vesicular stomatitis New Jersey virus: use of rapid molecular assay to differentiate between vesicular disease viruses

Author
item FOWLER, VERONICA - Pirbright Laboratory
item HOWSON, EMMA - Pirbright Laboratory
item MADI, MIKIDACHE - Pirbright Laboratory
item MIOULET, VALERIE - Pirbright Laboratory
item CAIUSI, CHIARA - Pirbright Laboratory
item Pauszek, Steven
item Rodriguez, Luis
item KING, DONALD - Pirbright Laboratory

Submitted to: Journal of Virological Methods
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/17/2016
Publication Date: 4/23/2016
Citation: Fowler, V.L., Howson, E.L., Madi, M., Mioulet, V., Caiusi, C., Pauszek, S.J., Rodriguez, L.L., King, D. 2016. Development of a reverse transcription loop-mediated isothermal amplification assay for the detection of vesicular stomatitis New Jersey virus: use of rapid molecular assay to differentiate between vesicular disease viruses. Journal of Virological Methods. 234:123-131. doi: 10.1016/j.jviromet.2016.04.012.

Interpretive Summary: Vesicular stomatitis (VS) is an important viral disease causing blisters in the mouth and feet of horses, cattle and swine. In cattle and swine the clinical signs resemble those of foot-and-mouth disease (FMD) a devastating viral disease of livestock that does not occur in the United States. Unlike FMD, VS does occur in the US causing sporadic outbreaks especially in southwestern and western States including Arizona, New Mexico, Texas and Colorado among others. There is a pressing need for rapid, sensitive and specific diagnostic assays that can differentiate VS from FMD in the field. A new type of test based on the detection of the virus genetic material termed RT-LAMP has been developed for vesicular diseases such as FMD and swine vesicular disease (SVD) but there is currently no RT-LAMP assay that can detect VS virus (VSV) nor are there any RT-LAMP assays which permit rapid discrimination between these ‘look-a-like’ diseases in a single test. This study describes the development of a novel RT-LAMP assay for the detection of VSV focusing on the New Jersey (VSNJV) serotype, which has caused most of the recent VS cases in the Americas. This RT-LAMP assay was combined with other RT-LAMP tests for detecting FMD and SVD. In addition these tests were adapted to be read using a disposable devices similar to those used for reading pregnancy tests. This assay was able to detect representative strains of VSNJV from various geographic origins and the limit of detection of the assay was similar whether testing was done alone or in combination with the other diseases. Furthermore, VSNJV could be reliably detected in clinical samples directly without the need for prior expensive and time consuming sample preparation. This test provides an approach that could be used as the basis for a rapid and low cost assay for differentiation of FMD from other vesicular diseases in the field.

Technical Abstract: Vesicular stomatitis (VS) is endemic in Central America and northern regions of South America. Sporadic outbreaks of VS can occur in cattle and pigs where the clinical presentation can be similar to foot-and-mouth disease (FMD). There is therefore a pressing need for rapid, sensitive and specific differential diagnostic assays that are suitable for decision making in the field. RT-LAMP assays have been developed for vesicular diseases such as FMD and swine vesicular disease (SVD) but there is currently no RT-LAMP assay that can detect VS virus (VSV), nor are there any multiplex RT-LAMP assays which permit rapid discrimination between these ‘look-a-like’ diseases in situ. This study describes the development of a novel RT-LAMP assay for the detection of VSV focusing on the New Jersey (VSNJV) serotype, which has caused most of the recent VS cases in the Americas. This RT-LAMP assay was combined in a multiplex format combining molecular lateral-flow devices for the discrimination between FMD and VS. This assay was able to detect representative VSNJV and the limit of detection of the singleplex and multiplex VSNJV RT-LAMP assays were equivalent to laboratory based real-time RT-PCR assays. A similar multiplex RT-LAMP assay was developed to discriminate between FMDV and SVDV, showing that FMDV, SVDV and VSNJV could be reliably detected within epithelial suspensions without the need for prior RNA extraction, providing an approach that could be used as the basis for a rapid and low cost assay for differentiation of FMD from other vesicular diseases in the field.