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Abstract
Peste des petits ruminants (PPR), caused by small ruminant morbillivirus (SRMV), formerly called 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, real-time reverse-transcription recombinase polymerase amplification assay (RT-RPA), employing primers and exo probe, was thus developed to perform at 42 °C for 20 min, and the detection limit at 95% probability was 14.98 copies per reaction and 0.326 TCID50/mL based on plasmid copy number and tissue culture infectivity titre. All the four lineages of PPRV could be detected with no cross-reaction to other pathogens including measles virus (MeV), goatpox virus (GTPV), canine distemper virus (CDV), foot-and-mouth disease virus (FMDV) and Mycoplasma capricolum subsp. capripneumoniae (Mccp). The performance of real-time RT-RPA assay was validated by testing 138 field samples and compared to real-time RT-PCR. The results indicated an excellent diagnostic agreement between real-time RT-RPA and a reference real-time RT-PCR method with the kappa value of 0.968. Compared to real-time RT-PCR, the sensitivity of real-time RT-RPA was 100%, while the specificity was 97.80%. The developed RT-RPA assay offers a promising platform for simple, rapid, and reliable detection of PPRV, especially in the resource-limited settings.
Highlights
Peste des petits ruminants (PPR), caused by small ruminant morbillivirus (SRMV), formerly called peste des petits ruminants virus (PPRV), mainly affects sheep, goats and wild small ruminants, leading to high mortality[1]
To determine the sensitivity of PPRV real-time reverse transcription recombinase polymerase amplification (RT-recombinase polymerase amplification (RPA)), a dilution ranging from 2 × 104 to 0.2 TCID50/mL was tested for eight replicates
In order to identify the specificity of RT-RPA assay, PPRV, measles virus (MeV), goatpox virus (GTPV), canine distemper virus (CDV), foot-and-mouth disease virus (FMDV) and Mycoplasma capricolum subsp. capripneumoniae (Mccp) were involved in the specificity test
Summary
Peste des petits ruminants (PPR), caused by small ruminant morbillivirus (SRMV), formerly called peste des petits ruminants virus (PPRV), mainly affects sheep, goats and wild small ruminants, leading to high mortality[1]. This acute, highly contagious viral disease has severely impacted the farmer’s livelihood and led to a constantly feared threat to socioeconomic development of endemic regions, including Africa, the Middle East, Arabian Peninsula and Southern Asia, since its emergence in West Africa in 1942. We aim to develop real-time reverse transcription recombinase polymerase amplification (RT-RPA) for the rapid detection of PPRV from clinical samples and evaluate its efficacy in comparison with real-time PCR testing
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