Anopheles gambiae, responsible for the majority of malariadeaths annually, is a complex of seven species and severalchromosomal/molecular forms. The complexity of malariaepidemiology and control is due in part to An. gambiae’sremarkable genetic plasticity, enabling its adaptation toa range of human-influenced habitats. This leads to rapidecological speciation when reproductive isolation mechanismsdevelop [1–6]. Although reproductive isolation isessential for speciation, little is known about how it occursin sympatric populations of incipient species . We showthat in such a population of ‘‘M’’ and ‘‘S’’ molecular forms,a novel mechanism of sexual recognition (male-femaleflight-tone matching [7–9]) also confers the capability ofmate recognition, an essential precursor to assortativemating; frequency matching occurs more consistently insame-form pairs than in mixed-form pairs (p = 0.001).Furthermore, the key to frequency matching is ‘‘differencetones’’ produced in the nonlinear vibrations of the antennaby the combined flight tones of a pair of mosquitoes and detectedby the Johnston’s organ. By altering their wing-beatfrequencies to minimize these difference tones, mosquitoescan match flight-tone harmonic frequencies above theirauditory range. This is the first description of close-rangemating interactions in incipient An. gambiae species.