Why is oral-induced RNAi inefficient in Diatraea saccharalis? A possible role for DsREase and other nucleases

• Published in: Pesticide biochemistry and physiology Vol. 186; p. 105166
• Main Authors: Abreu Reis, Manoely, Noriega, Daniel David, dos Santos Alves, Gessica, Ramos Coelho, Roberta, Grossi-de-Sa, Maria Fatima, Antonino, José Dijair
• Format: Journal Article
• Language: English
• Published: Elsevier Inc 01.08.2022
• Subjects: dsRNA stability; Nuclease; RNA interference; RNAi efficiency; Sugarcane borer; dsRNA stability; Sugarcane borer; RNA interference; Nuclease; RNAi efficiency
• ISSN: 0048-3575; 1095-9939
• DOI: 10.1016/j.pestbp.2022.105166

The efficiency of RNAi technology in insects varies considerably, particularly in lepidopterans. An important limiting factor of RNAi-mediated gene silencing is the degradation of dsRNA by insect nucleases before cellular uptake. To date, few studies have reported effective gene knockdown in the sugarcane borer Diatraea saccharalis. However, yielding contradictory results when using oral delivery. Further, the RNAi efficiency in D. saccharalis and presumed activity of gut nucleases remain poorly understood. Therefore, we investigated whether gene silencing was feasible via dsRNA feeding in D. saccharalis. Two different genes were tested, juvenile hormone esterase (DsJHE) and chitin synthase 1 (DsCHS1). Discrete knockdown was verified only for DsCHS1 with high dsRNA dosages and long exposure times. Neither mortality nor abnormal phenotypes were observed after treatment with any tested dsRNA. It was also verified that dsRNAs were quickly degraded when incubated with gut juice. Furthermore, we identified four possible nucleases that could reduce the knockdown efficiency in D. saccharalis. Three of them had the endonuclease_NS domain (DsNucleases), and one had the PIN domain (DsREase), with REase-like genes being scarcely represented in databanks. We further remark that DsNuclease1 and DsREase are highly expressed in the larval gut, and DsREase was upregulated as insects were fed with artificial diet (without dsRNA), and also when injected with dsRNA. Conversely, no nuclease was triggered when insects were fed with a sucrose droplet containing dsRNA. Thus, our findings suggest that nuclease activity within the gut is one of the possible reasons for the inefficiency of RNAi in D. saccharalis. Our data may shed light on the challenges to overcome when introducing RNAi as a strategy for controlling lepidopteran pests. [Display omitted] •The RNAi response is low and transient in D. saccharalis.•The gut juice content of D. saccharalis can fast degrade dsRNA.•D. saccharalis has 3 nucleases with endonuclease NS domain and one with PIN domain.•DsREase is upregulated in fed insects when compared to starved ones.•DsREase is upregulated when dsRNA is injected into insect hemolymph.