Assessing the acoustic behaviour ofAnopheles gambiaes.l.dsxFmutants: Implications for Vector Control
Authors: Su Matthew P , Georgiades Marcos , Bagi Judit , Kyrou Kyros , Crisanti Andrea , Albert Joerg T 
Published: September 2020

AbstractBackgroundThe release of genetically modified mosquitoes which use gene-drive mechanisms to suppress reproduction in natural populations ofAnophelesmosquitoes is one of the scientifically most promising methods for malaria transmission control. However, many scientific, regulatory and ethical questions remain before transgenic mosquitoes can be utilised in the field. Mutations which reduce an individual’s reproductive success are likely to create strong selective pressures to evolve resistance. It is thus crucial that the targeted population collapses as rapidly and as completely as possible to reduce the available time for the emergence of drive-resistant mutations. At a behavioural level, this means that the gene-drive carrying mutants should be at least as (and ideally more) sexually attractive than the wildtype population they compete against. A key element in the copulatory negotiations ofAnophelesmosquitoes is their acoustic courtship. We therefore analysed sound emissions and acoustic preference in adoublesexmutant previously used to successfully collapse caged colonies ofAnopheles gambiae s.l..MethodsThe flight tones produced by the beating of their wings form the signals for acoustic mating communication inAnophelesspecies. We assessed the acoustic impact of the disruption of a female-specific isoform of thedoublesexgene (dsxF) on the wing beat frequency (WBF; measured asflight tone) of both males (XY) and females (XX) in homozygousdsxF-mutants (dsxF-/-), heterozygousdsxF-carriers (dsxF+/-) and G3 ‘wildtype’dsxF+controls (dsxF+/+). To exclude non-genetic influences, we controlled for temperature and measured wing lengths for all experimental animals. We used a phonotaxis assay to test the acoustic preferences of mutant and control mosquitoes.ResultsA previous study demonstrated an altered phenotype only for females homozygous for the disrupteddsxallele (dsxF-/-), who appear intersex. No phenotypic changes were observed for heterozygous carriers or males, suggesting that the female-specificdsxFallele is haplosufficient. We here identify significant, dose-dependent increases in the flight tones of bothdsxF-/-anddsxF+/-females when compared todsxF+/+control females. Flight tone frequencies in all three female genotypes remained significantly lower than in males, however. When tested experimentally, males showed stronger phonotactic responses to the flight tones of controldsxF+/+females. While flight tones fromdsxF+/-anddsxF-/-females also elicited positive phonotactic behaviour in males, this was significantly reduced compared to responses to control tones. We found no evidence of phonotactic behaviour in any female genotype tested. None of the male genotypes displayed any deviations from the control condition.ConclusionsA key prerequisite for copulation in anopheline mosquitoes is the phonotactic attraction of males towards female flight tones within large - spatially and acoustically crowded - mating swarms. Reductions in acoustic attractiveness of released mutant lines, as reported here for heterozygousdsxF+/-females, reduce the line’s mating efficiency, and could consequently reduce the efficacy of the associated population control effort. Assessments of caged populations may not successfully reproduce the challenges posed by natural mating scenarios. We propose to amend existing testing protocols in order to more faithfully reflect the competitive conditions between a mutant line and the wildtype population it is meant to interact with. This should also include novel tests of ‘acoustic fitness’. In line with previous studies, our findings confirm that disruption of the female-specific isoformdsxFhas no effect on males; for some phenotypic traits, such as female flight tones, however, the effects ofdsxFappear to be dose-dependent rather than haplosufficient.