Abstract: 【Aim】 The objective of this study is to reveal the impacts of Wolbachia on the egg hatching of hybrids of two sibling tea geometrids and the underlying mechanism. 【Methods】 A subset of Ectropis grisescens larvae were fed with fresh tea leaves soaked in 2.5 mg/mL tetracycline solution for 1 min to remove Wolbachia. E. grisescens populations without Wolbachia were established after being fed with tetracycline-treated tea leaves for three generations. E. obliqua adults were hybridized with E. grisescens adults with and without Wolbachia to produce hybrids for the investigation of the impacts of Wolbachia on the egg hatching. The iTRAQ technique was used to detect and analyze the differences in sperm proteins between E. grisescens with and without Wolbachia. 【Results】 When Wolbachia was removed from E. grisescens, the egg hatching rates of hybrids between E. grisescens and E. obliqua were significantly increased from 3.92% to 56.20%, indicating that Wolbachia induced cytoplasmic incompatibility. Results of differential sperm proteins analysis showed that a total of 128 proteins were confirmed to have significantly different expression levels in sperms of E. grisescens with and without Wolbachia. According to the KEGG database, 45 differential proteins were enriched in 106 pathways, including sphingolipid metabolism, salivary secretion, lysosome, sphingolipid signaling, sphingolipid biosynthesis, etc. Among them the expression levels of ceramidases and phosphatases related to ceramide synthesis in the sphingolipid metabolic pathway were significantly up-regulated. 【Conclusion】 The symbiotic bacterium, Wolbachia, mediates the prezygotic reproductive isolation between the two sibling tea geometrids through cytoplasmic incompatibility, leading to a significant decrease in the egg hatching rate of hybrids. The observed cytoplasmic incompatibility may be attributable to the modification of the male sperm proteins caused by the impacts of Wolbachia on the sphingolipid metabolism pathway.

Key words: Tea geometrids, sibling species; Wolbachia, hybrid eggs, cytoplasmic incompatibility, sperm protein