Stage-specific and cumulative effects of heat stress on male and female reproductive performance in Drosophila melanogaster

Frequent heat events, driven by climate change, pose significant threats to biodiversity, particularly for insects and other ectothermic species with limited thermoregulatory capacities. Beyond affecting survival, heat stress can severely impair reproductive fitness. Previous studies have typically examined the effects of heat events on single reproductive stages or sexes. However, the thermal sensitivity likely varies across sex-specific reproductive stages, from gonad development to post-mating gamete interactions. Here, we investigated the timing- and sex-specific effects of heat stress across key reproductive stages in Drosophila melanogaster. Using a full-factorial design, we exposed males and females to heat stress during development, as premating adults, and after mating to assess stage-specific and cumulative impacts on reproductive performance. Our results revealed the highest thermal sensitivity during post-mating sperm storage by females, where even brief heat stress critically reduced reproductive output. Pre-mating adult heat exposure also affected both sexes, but less severely, whilst developmental heat exposure had the least impact and mostly on males. These findings highlight the critical role of timing and sex-specific reproductive stages in mediating thermal fertility. Our study provides novel insights into the vulnerability of sex-specific reproductive stages to heat stress, offering essential knowledge for predicting more accurate population dynamics and persistence under increasingly variable thermal conditions driven by climate change.