Development and application of species ID and insecticide resistance assays, for monitoring sand fly Leishmania vectors in the Mediterranean basin and in the Middle East

Abstract

AbstractsO_ST_ABSBackgroundC_ST_ABSDevelopment of insecticide resistance (IR) in sand fly populations is an upcoming issue of public health concern, threatening leishmaniasis mitigation efforts by insecticide- based vector control. There is a major knowledge gap in the IR status of wild populations worldwide, possibly attributed to the unavailability of specialized tools, such as bioassay protocols, species baseline susceptibility to insecticides and molecular markers, to monitor such phenomena in sand flies. Methodology/ Principal findingsSeveral sand fly populations from (semi-)rural regions of Greece, Turkey and Iran were sampled and identified to species, showing populations structure in accordance with previously reported data. Genotyping of known pyrethroid resistance-associated loci revealed the occurrence of voltage-gated sodium channel (vgsc) mutations in all surveyed countries. Knock-down resistance (kdr) mutation L1014F was prevalent in Turkish regions and L1014F and L1014S were recorded for the first time in Iran and in Turkey and Greece, respectively, yet in low frequencies. Moreover, CDC bottle bioassays against pyrethroids in mixed species populations from Greece indicated full susceptibility, using though the mosquito discriminating doses. In parallel, we established a novel individual bioassay protocol and applied it comparatively among distinct Phlebotomus species populations, to detect any possible divergent species-specific response to insecticides. Indeed, a significantly different knock-down rate between P. simici and P. perfiliewi was observed upon exposure to deltamethrin. Conclusions/SignificanceIR in sand flies is increasingly reported in leishmaniasis endemic regions, highlighting the necessity to generate additional monitoring tools, that could be implemented in relevant eco-epidemiological settings, in the context of IR management. Our molecular and phenotypic data add to the IR map in a macroarea with otherwise limited data coverage. Author SummaryPhlebotomine sand flies vector leishmaniasis to human and animals, a neglected tropical disease of the (sub-)tropics, displaying an expanding epidemiological tendency towards previously non-endemic regions. As leishmaniasis eradication largely relies on vectors insecticidal control, regular monitoring of insecticide resistance (IR) is a core element of integrated vector management. IR data are limited in sand fly populations worldwide though, among others due to the unavailability of robust molecular diagnostic and phenotyping tools. Here, we aim to characterise the pyrethroid resistance (PyR) status of populations originating from countries of the Mediterranean basin and the Middle East; i.e. Greece, Turkey and Iran. PyR-associated mutations were detected in all three countries, while the populations from Greece exhibited susceptibility upon exposure to deltamethrin in CDC bioassays. We established an individual bioassay protocol to enable IR monitoring in settings with multi- species population structure, and applied it comparatively among three distinct Phlebotomus species, revealing that they respond differently to insecticide treatment. Our study will facilitate systematic surveillance and evidence-based sand fly control in the endemic regions.