Abstract
Oviposition site selection is critical for mosquito population dynamics. Gravid mosquitoes rely on chemical cues to identify suitable breeding habitats. However, the sensory mechanisms governing this behavior in Anopheles stephensi remain poorly understood. Here, we examined the role of indole, a microbial volatile associated with aquatic environments, in oviposition site choice and assessed the involvement of sensory organs in its detection. In two-choice oviposition assays, water conditioned with first-instar larvae attracted gravid females (OAI = 0.56). In contrast, water from fourth-instar larvae was repellent (OAI = -0.20), consistent with avoidance of suboptimal, resource-depleted habitats. Single indole cue elicited strong oviposition attraction across a broad concentration range (0.5-50 µM), with no clear dose-response relationship. Surgical ablation of antennae and maxillary palps did not abolish indole-mediated preference but significantly reduced behavioral variability, suggesting that these structures modulate, rather than solely mediate, indole detection. Reanalysis of transcriptomes of antennae, maxillary palps, and legs in An. gambiae and An. coluzzii, along with quantitative RT-PCR in An. stephensi revealed the expression of chemosensory genes (including Obp1, Obp13, Obp25, Obp71, Or2, and Or10) in the legs, suggesting a potential role for leg chemosensation in oviposition decisions. These findings underscore the complexity of chemoperception in mosquito habitat assessment.