Spatial and seasonal distribution of V1016G mutation in Aedes albopictus along a transect from coastal areas to the metropolitan city of Rome and beyond

Genotyping pyrethroid resistance-associated mutations in natural mosquito populations represents a powerful approach to detect early signs of resistance without the need of carrying out demanding phenotypic bioassays on live specimens. In Aedes albopictus, target-site resistance to pyrethroids is associated to mutation in the vgsc gene. In particular, mutation V1016G has been reported from 9 European countries, with frequencies up to 45% in Italian sites where phenotypic resistance was shown. The study aimed at the evaluation of spatial and temporal variation in frequency of V1016G mutation along a transect in Lazio region from the sea to the mountains encompassing Rome metropolitan area. Results from PCR-genotyping of 1352 specimens collected in 24 sites from June to December 2023 highlight spatial variability along the transect, with frequencies ranging from 13% to 38% in sites within Rome metropolitan area and from 4% to 48% in suburban sites towards the coast. Lowest frequencies (≤3%) were observed in rural areas north of Rome. Results from Generalized Linear Model suggest a statistically significant positive correlation between 1016G allele frequency and mean human population density (which could be taken as a proxy for pyrethroid usage). Overall, 1016G frequency did not significantly change from early to late 2023 season, with the exception of two sites where it increased from 7%-16% to 29%-49%. Interestingly, both sites are located within sporting clubs, where mosquitoes experience strong selective pressure due to intensive pyrethroid treatments regularly carried out to reduce nuisance. This trend was not observed in most of the sampling sites situated in public areas where mosquito adulticides are not routinely implemented and selective pressure is lower and irregular. Consistent results were observed in 5 of the 24 sites, which were sampled again in both early and late season 2024.Finally, 1016G frequencies recorded in 7 sites within urban Rome in 2023 were also compared with those recorded in the same sites in 2012, when they ranged between 6% and 34%. A significant increase in frequency (from <10% to 28%) was observed in two sites only.These data highlight widespread and heterogeneous presence of V1016G mutation in the study area, reaching worrisome frequencies in some sites. However, overall data indicate a limited increase in frequencies over 11 years, which could be possibly due to dispersal of wild-type mosquitoes from non-treated to treated areas and/or fitness costs associated with the 1016G allele. Specific laboratory bioassay on selected 1016V and 1016G homozygous colonies are ongoing to assess possible fitness costs of V1016G mutation.