Schmallenberg virus non-structural proteins NSs and NSm are not essential for experimental infection of biting midges.

The teratogenic orthobunyavirus Schmallenberg virus (SBV) is transmitted between its mammalian hosts by biting midges. The genome of circulating SBV, i.e., variants present in viremic ruminants or insect vectors, is very stable, while variants found in malformed ruminant fetuses display a high genetic variability. It was suggested that fetal infection provides an environment that favors viral mutations that enable immune escape in the unborn but at the cost of limiting the ability of the virus to spread further. To investigate infection and dissemination rates of different SBV variants in the insect vectors, we fed laboratory-reared with blood containing the prototype strain BH80/11-4 from a viremic cow or strain D281/12, which was isolated from the brain of a sheep fetus and harbors multiple mutations in all three genome segments. Furthermore, virus variants lacking NSs, NSm, or both non-structural proteins were included. Six days after feeding, virus replication was found in about 2% of the midges exposed to wild-type strain BH80/11-4. The absence of the non-structural proteins had no obvious effect on the oral susceptibility to virus infection, as after 6 days, 2.78% of the midges fed with the NSs-deletion mutant displayed viral loads higher than the respective day-0 group, 1.92% of the midges exposed to the NSm-deletion mutant, and 1.55% of midges exposed to the NSs/NSm-deletion mutant. In contrast, strain D281/12 did not replicate at all in the midges, supporting the assumption that SBV variants arising in infected fetuses are unable to enter the normal insect-mammalian host cycle.IMPORTANCEBiting midges are responsible for the transmission of Schmallenberg virus (SBV), a pathogen of veterinary importance that primarily infects ruminants. Although SBV has been extensively studied in the mammalian host, the virus-intrinsic factors allowing infection of and replication in biting midges are largely unknown. Therefore, we infected laboratory-reared midges with SBV variants by feeding them virus-containing blood. The SBV variants differed in their genome composition, as we used the prototype wild-type strain, a strain with multiple mutations that was isolated from the brain of a malformed fetus, and recombinants lacking either NSs or NSm or both of these non-structural proteins. While the non-structural proteins had no obvious effect, the variant from the malformed fetus did not replicate at all, indicating that virus variants with characteristic genomic mutations present in fetuses lose their ability to infect the insect vector and will be excluded from the natural transmission cycle.