Molecular Cloning and Functional Analysis of CTL14 and PPO4: Their Roles in Development, Reproduction, and Antiviral Defense in Hyphantria cunea

Abstract

The insect innate immune system serves as the primary line of defense against pathogens, regulated by pattern recognition receptors (PRRs) such as C-type lectins (CTLs) and effector systems like prophenoloxidase (PPO). This study aims to provide molecular and functional characterization of the CTL14 and PPO4 genes from Hyphantria cunea. Cloning results revealed that CTL14 encodes 378 amino acids, whereas PPO4 encodes 681 amino acids. Expression analysis indicated that CTL14 is highly expressed during the fourth instar and within midgut tissues, while PPO4 expression is predominant during the first instar and in the epidermis. Functional assays demonstrated that silencing both genes significantly increased larval mortality following infection with the NPV virus using RNA interference (RNAi), with mortality rates reaching 75–100% by the tenth day. Furthermore, knockdown of these genes resulted in noticeable changes in larval physiology, manifested by altered feeding behavior and significantly reduced body weight gain. Conversely, the injection of CTL14 protein resulted in a marked increase in larval body mass accumulation compared to controls. These findings underscore the crucial roles of CTL14 and PPO4 as they are involved in both the antiviral defense system and the physiological development of H. cunea, thereby identifying them as potential novel targets for biological pest control strategies.