Prokaryotic-expressed V-ATPaseA dsRNA and its effects on growth and development in Lymantria dispar

Objective The objective of this study was to thoroughly investigate how the LdV-ATPaseA gene affects various physiological activities of Lymantria dispar, including molting, reproduction, growth, and development. Furthermore, we evaluated its potential as a green control target for Lymantria dispar RNA interference (RNAi) and provide reference for the development of new biopesticides for L. dispar.

Method The ORF sequence of V-ATPase was successfully cloned, and quantitative reverse transcription PCR (qRT-PCR) was used to measure gene expression levels across various developmental stages and tissues in L. dispar. Additionally, an in vitro prokaryotic expression system was developed to produce dsRNA targeting the LdV-ATPaseA gene, continuous feeding of 2nd and 3rd instar larvae of L. dispar. Subsequently, we conducted experimental statistics on indicators such as food intake, body mass, cumulative mass gain rate, mortality rate, pupation rate, pupal mass, egg production, and developmental duration.

Result The LdV-ATPaseA gene showed consistent expression levels across all developmental phases of L. dispar, with distinct tissue specific expression levels. The highest expression level was detected in the tests, with 30.80-fold of that in the control fat body. This was followed by ovary and epidermis, with 12.82-fold and 9.66-fold of that in the fat body, respectively. The recombinant expression of LdV-ATPaseA vector was successfully constructed to generate dsRNA which specifically induces V-ATPaseA gene silencing. Moreover, the dsRNA ingested by 2nd and 3rd instar L. dispar resulted in decreased food intake, reduced larval mass, decreased larval growth rate, decreased pupal mass of females and increased mortality. Additionally, the dsRNA ingested by 2nd instar L. dispar resulted larval and pupal life span was significantly longer, increased the pupal malformation rate, and reduced the pupation and egg production in female adults.

Conclusion The LdV-ATPaseA plays a crucial role in the molting, reproduction, growth, and development of L. dispar. These findings highlight its potential as an RNAi-based green control target and offer valuable insights for developing biopesticides against Lepidoptera pests.