Curing Parthenogenesis-Inducing (PI) Wolbachia-Induced Reproductive Disorders in the Egg Parasitoid Telenomus remus

Wolbachia is an endosymbiotic bacterium widespread in invertebrates that causes various reproductive effects, including cytoplasmic incompatibility, feminization, male killing, and the induction of parthenogenesis (PI). PI-Wolbachia wRem converts Telenomus remus, an egg parasitoid of Spodoptera frugiperda, from arrhenotokous reproduction (male-producing) to thelytokous reproduction (female-producing). Long-term symbiosis between egg parasitoids and Wolbachia has been shown to lead to reproductive barriers and "female functional virginity," causing progressive and potentially irreversible sex ratio imbalances. However, whether such reproductive barriers occur in T. remus remains unknown, which has important implications for biological control programs utilizing this parasitoid. To address this question, we cured wRem using tetracycline and conducted crossing experiments with naturally uninfected strains (W-). The results indicated that the cured strain (Wcure) retained normal sexual reproductive capability, with self-crossing fertilization rates comparable to those of W- strains. However, first-generation hybridization between Wcure and W- strains produced strongly male-biased offspring (male proportion: 94.3% and 85.8% for W-♂ × Wcure♀ and Wcure♂ × W-♀, respectively), indicating substantial reproductive incompatibility. Notably, an asymmetric pattern was observed between reciprocal crosses. In second-generation hybridization experiments, hybrid females (W-/Wcure) mated with W- or Wcure males showed markedly recovered sex ratios (male proportion: 14.3% and 15.6%, respectively), although total offspring numbers remained lower than in self-crossing groups. These results suggest that the reproductive incompatibility in T. remus differs from female functional virginity and is more consistent with mitonuclear incompatibility arising from population divergence. The partial recovery in second-generation hybrids indicates that surviving F1 hybrid females likely represent individuals selected for compatibility, rather than exhibiting progressive deterioration of sexual function. These findings offer insights into Wolbachia's impact on parasitoid reproduction and highlight key considerations for biological control applications, underscoring the importance of evaluating reproductive barriers before deploying cured strains and preventing symbiont loss within populations.