A combined approach to heat stress effect on male fertility in Nasonia vitripennis: from the physiological consequences on spermatogenesis to the reproductive adjustment of females mated with stressed males

Abstract

In recent years, several studies have shown a decline in reproductive success in males in both humans and wildlife. Research on male fertility has largely focused on vertebrates, although invertebrates constitute the vast majority of terrestrial biodiversity. The reduction of their reproductive capacities due to environmental stresses can have strong negative ecological impacts, and also dramatic consequences on world food production if it affects the reproductive success of biological control agents, such as parasitic wasps used to control crop pests. Here Nasonia vitripennis, a parasitic wasp of various fly species, was studied to test the effects of 24h-heat stress applied during the first pupal stage on male fertility. Results showed that only primary spermatocytes were present at the first pupal stage in all cysts of the testes. Heat stress caused a delay in spermatogenesis during development and a significant decrease in sperm stock at emergence. Females mated with these heat-stressed males showed a reduce sperm count stored in their spermatheca. Females did not appear to distinguish heat-stressed from control males and did not remate more frequently to compensate for the lack of sperm transferred. As a result, females mated with heat-stressed males produced a suboptimal lifetime offspring sex ratio compared to those mated with control males. This could further impact the population dynamics of this species. N. vitripennis appears to be an interesting biological model to study the mechanisms of subfertility and its consequence on female reproductive strategies and provides new research perspectives in both invertebrates and vertebrates.

Fig 1. Sperm stock in seminal vesicles of emerging N. vitripennis white pupa males stressed at 34, 36 & 38°C for 24 hours (median with lower and upper quartiles).

Different letters indicate a significant difference between treatments at α = 0.05 (25° C: N = 12, 34°C: N = 11, 36°C: N = 11 and 38°C: N = 10).

Fig 2. Histological examination of testes at the early pupal stage.

A) Morphology of germ cells in cysts from testes. GC = Germ Cell, CT = Cyst. Bar = 10μm. B) mRNA expression of non-differentiated and meiotic germ cells from the early pupal stage and legs.

Fig 3. Morphology of the reproductive tract of control and heat-stressed males.

A) Morphology of germ cells in cysts from control testes at the second pupal stage. B) Morphology of germ cells in cysts from heat-stressed testes at the second pupal stage (after heat stress). C) Morphology of germ cells in cysts from control testes at the final pupal stage. D) Morphology of germ cells in cysts from heat-stressed testes at the final pupal stage (72 hours after heat stress). D) Morphology of seminal vesicles from control testes at the final pupal stage. E) Morphology of seminal vesicles from heat-stressed testes at the final pupal stage (72 hours after heat stress). GC = Germ Cell, SPD = Spermatid, CT = Cyst, TP = Testis Pocket, SPZ = Spermatozoa. Bar = 10 μm

Fig 4. Total number of spermatozoa in female spermatheca after a single mating with either a control or a 36°C heat-stressed male (median with lower and upper quartiles).

** indicates a significant variation between heat-stressed and control males at α = 0.01 (Mann-Whitney, U = 0, N1 = 13, N2 = 13, P < 0.01).

Fig 5. Behavioral response of virgin females to odors of control male versus no male (χ²₁ = 3.2667, P = 0.041, N = 15) or control male versus 36°C heat-stressed male (χ²₁ = 0.2222, P = 0.64, N = 18) in a still-air Y-tube olfactometer.

* indicates a significant difference at α = 0.05.

Fig 6. Lifetime offspring sex ratio (proportion of females) produced by mated females with either a control or a heat-stressed male (median with lower and upper quartiles).

** indicates significant variation between female offspring of females mated with heat-stressed or control males at α = 0.01 (Mann-Whitney, U = 272, N1 = 20, N2 = 17, P < 0.01).