DWV Infection in vitro Using Honey Bee Pupal Tissue

Abstract

The deformed wing virus (DWV) has been best characterized among honey bee viruses; however, very little is known regarding the mechanisms of viral infection and replication due to the lack of immortalized honey bee cell lines. To solve this problem, we established an in vitro system using honey bee pupal tissue to reconstruct DWV binding and entry into the host cell, followed by translation of the RNA genome and polyprotein processing using RNA-dependent RNA polymerase (RdRP) as a marker. Using this system, the P-domain of the virion subunit VP1 was found to be essential for DWV infection, but not for binding and entry into the cell. DWV efficiently infected the head tissue derived from early but not late pupa, suggesting that undifferentiated cells are targeted for viral infection. Furthermore, we found that inhibitors of mammalian picornavirus 3C-protease, rupintrivir and quercetin suppressed RdRP synthesis, indicating that this in vitro system is also useful for screening a compound to control viral infection. Our in vitro system may help to understand the mechanism of DWV infection in host cells.

DWV 3C Protease Uncovers the Diverse Catalytic Triad in Insect RNA Viruses

“…We previously showed that rupintrivir suppresses DWV replication in honey bee pupal cells based on the synthesis of RdRP ( 34 ). Thus, we tested the effect of ebselen on DWV replication.…”

“…Thus, ebselen (MW: 274.18) could be a potential lead compound for fragment-based drug discovery approaches to increase the specificity and potency to DWV 3C pro . Rupintrivir and quercetin ( 34 ) as well as ebselen inhibited DWV replication in cultured honey bee cells. Thus, these compounds and the other 3C pro inhibitors we identified in this study could be used to control DWV in honey bee colony.…”

A simple method for ex vivo honey bee cell culture capable of in vitro gene expression analysis

“…Difficulties are associated with maintaining these cell lines, which may be due to viral infection [ 15 , 17 , 18 ]. In contrast, primary cultured cells have been established from various tissues and organs in the honey bee, and these primary cell cultures are proven to be useful for viral studies and neuronal processes [ 15 , 39 , 40 ]. Although primary cell cultures and ex vivo cultured cells may only be maintained for a limited period of time under healthy conditions, these cells are available for examining gene expression and signaling pathways [ 2 , 5 ].…”