Application Of Recombinase Polymerase Amplification In Rapid Detection Of Peste Des Petites Ruminants Virus

Peste des petits ruminants is an acute contagious disease caused by peste des petits ruminants virus.Goats,sheep and wild ruminants are susceptible to epidemic diseases.In1942,the peste des petits ruminants epidemic was first reported in the Ivorian region of Africa,and then became endemic in most of Africa,the Middle East,South Asia,and parts of asia.In2007 and 2013,peste des petits ruminants were introduced into China successively.It has appeared in many provinces of the whole country and has brought enormous economic loss to animal husbandry of china.Developing and applying rapid detection technology with high sensitivity,high specificity and high throughput has become the urgent task to prevent and controll the disease.This paper introduces the existing detection methods of peste des petits ruminants,including the establishment of detection methods,sensitivity,specificity,detection objectives,advantages and disadvantages,and so on,which provides technical support for the prevention and control of Peste des petits ruminants.Peste des petits ruminants?PPR?,caused by Peste des petits ruminants virus?PPRV?,is one of the most important pathogens in small ruminants,and has tremendous negative economic impact on the sheep industry worldwide.Current detection of PPRV in clinical samples mainly relies on real-time RT-PCR.Particularly,samples collected from rural area require highly equipped laboratories for screening.A rapid point-of-care test,real-time reverse-transcription recombinase polymerase amplification assay?RT-RPA?,employing primers and exo probe,was thus developed to perform under the condition of 42°C 20 min,and the detection limit at 95%probability was 0.326 TCID₅₀/mL.All the four lineages of PPRV could be detected with no cross-reaction to other pathogens including measles virus?MEV?,goat pox virus?GPV?,canine distemper virus?CDV?,foot-and-mouth disease virus?FMDV?and Mycoplasma capricolum subsp.The assay performance was validated by testing138 field samples and compared to real-time RT-PCR.The results indicated a good consistence between RT-RPA and a reference real-time RT-PCR method?98.55%,R²=0.947?according to linear regression analysis.Compared to real-time RT-PCR,the sensitivity of RT-RPA was 100%,and the specificity was 97.8%?Kappa value=0.8134?.The developed RT-RPA assay offers a promising platform for simple,rapid,and reliable detection of PPRV in both laboratory and point-of-care facility,especially in the resource-limited settings.Peste des Petites ruminants?PPR?is a highly infectious disease of small ruminants and caused by Peste des petites ruminants virus?PPRV?.In China,PPR was reported first in 2007and re-emerged in 2013.In this study,primers and nfo-probes were designed on the basis of the blast results of 4 lineages,39 whole genome sequences of PPRV.The recombinase polymerase amplification with lateral flow dipstick?RPA-LFD?assay for detection of PPRV was established by optimizing reaction conditions.The detection end point for cell culture-derived virus of this assay was 2.89 TCID50 per reaction.Cross reactivity with other pathogens was not observed.Clinical evaluation by the 138 samples collected in 2014-2017showed that the performance of the RPA-LFD assay was comparable to that of PCR.The sensitivity and specificity of the RPA-LFD assay relatively to PCR were 91.5%and 98.9%,respectively.In conclusion,the RPA-LFD assay built in this study has the advantages of reading the results conveniently,fast,and easy operation,to provide technical assistance for the prevention and control of PPR.