Identification of a BTV-Strain-Specific Single Gene That Increases Culicoides Vector Infection Rate.

Honorata M Ropiak,Simon Gubbins,Kerry Newbrook,John Flannery,Gillian D Pullinger,Marc Guimerà Busquets,Hannah Brown,Karin E Darpel,Paulina Rajko-Nenow,Carrie Batten,Simon King

doi:10.3390/v13091781

Abstract

Since the 2000s, the distribution of bluetongue virus (BTV) has changed, leading to numerous epidemics and economic losses in Europe. Previously, we found a BTV-4 field strain with a higher infection rate of a Culicoides vector than a BTV-1 field strain has. We reverse-engineered parental BTV-1 and BTV-4 strains and created BTV-1/BTV-4 reassortants to elucidate the influence of individual BTV segments on BTV replication in both C. sonorensis midges and in KC cells. Substitution of segment 2 (Seg-2) with Seg-2 from the rBTV-4 significantly increased vector infection rate in reassortant BTV-14S2 (30.4%) in comparison to reverse-engineered rBTV-1 (1.0%). Replacement of Seg-2, Seg-6 and Seg-7 with those from rBTV-1 in reassortant BTV-41S2S6S7 (2.9%) decreased vector infection rate in comparison to rBTV-4 (30.2%). However, triple-reassorted BTV-14S2S6S7 only replicated to comparatively low levels (3.0%), despite containing Seg-2, Seg-6 and Seg-7 from rBTV-4, indicating that vector infection rate is influenced by interactions of multiple segments and/or host-mediated amino acid substitutions within segments. Overall, these results demonstrated that we could utilize reverse-engineered viruses to identify the genetic basis influencing BTV replication within Culicoides vectors. However, BTV replication dynamics in KC cells were not suitable for predicting the replication ability of these virus strains in Culicoides midges.

Highlights

Since the 2000s, the distribution of bluetongue virus (BTV) has changed, leading to numerous epidemics and economic losses in Europe
Cloning was confirmed by Sanger sequencing and the inserts with the lowest number of nucleotide substitutions in comparison to the wtBTV strains were selected for the generation of reverse-engineered and reassortant BTV strains
We demonstrated that two reverse-engineered BTV strains, based on BTV1 and BTV-4 derived from the Moroccan outbreaks, retained their diverging infection rates of C. sonorensis first observed in wild-type isolates

Summary

Introduction

Since the 2000s, the distribution of bluetongue virus (BTV) has changed, leading to numerous epidemics and economic losses in Europe. Substitution of segment 2 (Seg-2) with Seg-2 from the rBTV-4 significantly increased vector infection rate in reassortant BTV-14S2 (30.4%) in comparison to reverse-engineered rBTV-1 (1.0%). Triple-reassorted BTV-14S2S6S7 only replicated to comparatively low levels (3.0%), despite containing Seg-2, Seg-6 and Seg-7 from rBTV-4, indicating that vector infection rate is influenced by interactions of multiple segments and/or host-mediated amino acid substitutions within segments. Overall, these results demonstrated that we could utilize reverse-engineered viruses to identify the genetic basis influencing BTV replication within Culicoides vectors. BTV strains vary greatly in their ability to infect and replicate, even in the same Culicoides species under controlled conditions [13,14,15]

Methods

Results

Discussion

Conclusion