Vector-delivered artificial miRNA effectively inhibits Porcine epidemic diarrhea virus replication

Abstract

Background Porcine epidemic diarrhea virus (PEDV) is an α-coronavirus that causes highly contagious intestinal infectious disease, involving clinically characterized by diarrhea, dehydration, vomiting, and high mortality to suckling piglets. As a strategy for antiviral therapy, artificial microRNA (amiRNA) mediated suppression of viral replication has recently become increasingly important. In this study, we evaluated the advantages of using an amiRNA vector against PEDV. Methods In this study, we evaluated the advantages of using an amiRNA vector against PEDV. We designed two single amiRNA sequences for different conserved sequences of the PEDV S and N genes, and tested their inhibitory effects on PEDV in Vero cells. Results It was obvious from the CCK-8 results that the transient transfection of amiRNA was non-toxic to the cells. In addition, our results showed that the transient expression of two amiRNAs (amiRNA-349 and amiRNA-1447) significantly reduced the expression of viral RNA and protein in the cells. The TCID50 results showed that the release of virus particles into the culture supernatant was significantly reduced, with an effect as high as 90%. To avoid virus mutation escape, the above two single amiRNA sequences were tandem in this study (amiRNA-349 + 1447), enabling a single microRNA to be expressed simultaneously. The real-time PCR and Western blot results showed that the inhibitory effect was significantly enhanced in each of the different time periods. The TCID50 results showed that the release of virus particles in the culture supernatant was significantly reduced at the different time periods. Conclusions In summary, these results suggest that an RNAi based on amiRNA targeting the conserved region of the virus is an effective method to improve PEDV nucleic acid inhibitors and provide a novel treatment strategy for PEDV infection.

Regulatory Non-Coding RNAs during Porcine Viral Infections: Potential Targets for Antiviral Therapy

“…Viral infection could be effectively inhibited by designing amiRNAs that target viral mRNAs ( Table 2 ). The amiRNA-349 and amiRNA-1447 targeting the conserved regions of PEDV N and S genes, respectively, effectively inhibited PEDV infection [ 186 ]. amirGP5-370 and amirM-263-M-263 targeting of PRRSV GP5 or M proteins, respectively, effectively inhibited PRRSV infection [ 187 ].…”

“…However, how amiRNAs can achieve more efficient delivery still needs further study. amiRNA-1447 PEDV S PEDV [186] amiRGP5-370 PRRSV GP5 PRRSV [187] amiRM-263-M-263 PRRSV M PRRSV [187] amiRNA-Sn Sn PRRSV [188] amiRNA-CD163 CD163 PRRSV [188] amiR3UTR PRRSV 3 ′ UTR PRRSV [189]…”