Sex-specific sterility caused by extreme temperatures is likely to create cryptic changes to the operational sex ratio in Drosophila virilis

Climate change is increasing the frequency and severity of short-term heat shocks that threaten the persistence of natural populations. However, most work addressing the evolutionary consequences of anthropogenic environmental change has focused on natural selection, with less attention paid to the impacts on sexual selection. The conditions under which sexual selection operates is a topic of debate, but a generally observed pattern is that the operational sex ratio (OSR) of a population is key to determining both the extent of competition for fertilizations and the scope for mate choice (Weir et al. 2011). Therefore, if high temperatures affect the ratio of reproductive males to females in a population, this could influence sexual selection. Sub-lethal temperatures can sterilize individuals from a range of biological systems, including plants, insects, corals, birds, and mammals (reviewed in Walsh et al. 2019a). If high temperatures affect reproduction in one sex more than the other, this may create cryptic shifts in the OSR of a population (Petry et al. 2016). However, although fertility loss at high temperatures is generally thought to be more common in males than in females (Iossa 2019), very few studies measure fertility in both sexes under identical conditions (Walsh et al. 2019b). Where sensitivity to temperature has been observed to vary between the sexes (Janowitz and Fischer 2011; Zwoinska et al. 2020), the effect on population sex ratios has not been considered. Furthermore, natural selection, sexual selection, and population dynamics are more likely to be affected by biased sex ratios if sterility is long-lasting. However, to date, patterns of sexually dimorphic heat-induced sterility have not been shown over organisms’ reproductive life spans. Here, we aim to test whether heat stress differentially affects male and female fertility in the cosmopolitan fruit fly Drosophila virilis and if this creates cryptic bias in population sex ratios over time. Specifically, we hypothesize that pupal heat stress will significantly delay adult sexual maturation and that this will be more severe in males compared to females under identical conditions. To do this, we exposed pupal D. virilis to a sub-lethal heat shock of 38 C for 4 h to simulate the peak of a mid-day heat wave. We chose to heat pupae because they are immobile and cannot behaviorally escape heat stress in nature. We subsequently examined both complete sterility and pupal offspring production over an ecologically realistic lifespan in both males and females. We combine male and female time-series data to predict the effect of heat-induced sterility on the OSR, and discuss its potential consequences on sexual selection. Detailed methods are described in the Supplementary Materials. We found that the rate at which newly eclosed D. virilis become fertile is significantly influenced by the interaction between sex and temperature. While female fertility is not significantly affected by heat stress, male sexual maturation is significantly extended if they are exposed to 38 C as pupae (Cox proportional hazard test interaction term: hazard ratio (HR) 1⁄4 1.4866, v1 1⁄4 16:275, P<0.001; Figure 1A and B). Furthermore, we found that the proportion of individuals that never produced offspring was predicted by a significant interaction between sex and treatment, wherein males exposed to heat stress were more likely than controls or females in any heat treatment to be rendered permanently sterile (v1 1⁄4 5.657, P1⁄40.017; Supplementary Figure S1). This is a relatively small effect, showing that most males recovered fertility at some point during the experiment. We found that control males reached sexual maturity 7 days post eclosion, in line with previous observations. This results in an observed OSR for control males and females to stabilize at 0.5 from that point 7 days onwards (Figure 1c). In stressed males and females, however, the sterile males prevent the OSR from reaching 0.5 over the 17-day duration of our experiment. This results in an observed female bias in the sex ratio when flies are heated as pupae (Figure 1C). In males, pupal heat stress significantly reduced pupal offspring number by 58%