Development of real-time reverse transcription recombinase polymerase amplification (RPA) for rapid detection of peste des petits ruminants virus in clinical samples and its comparison with real-time PCR test
Yuanli Li1,2, Lin Li1, Xiaoxu Fan1, Yanli Zou1, YongqiangZhang1, QinghuaWang1,
Chengyou Sun1, Shude Pan2, XiaodongWu1 & ZhiliangWang1
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 amplifcation assay (RT-RPA), employing primers and exo probe, was thus developed to
perform at 42°C for 20min, 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 feld 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 specifcity was 97.80%. The developed RT-RPA assay ofers a promising platform for
simple, rapid, and reliable detection of PPRV, especially in the resource-limited settings.
Aucun commentaire:
Enregistrer un commentaire