Abstract:

【Aim】 To screen the high efficient delivery nanocarrier of dsRNA used for RNAi in Laodelphax striatellus. 【Methods】 Chitosan, carbon quantum dot (CQD), and lipofectamine 2000 were used as representative nanoparticles (NPs). The dsRNA loading efficacy of each NP was measured by spectrophotometry. The membrane-bound trehalase gene of L. striatellus, LsStre, was selected as the target gene to test the RNAi efficiency mediated by the three NPs. The mRNA expression levels of LsStre in the 2nd instar nymphs of L. striatellus at 2 d after being fed with dsLsStre mediated by different NPs were quantified by using real-time quantitative PCR (qPCR) strategy, and the corrected mortality rates induced by dsRNAs in 6 d were also detected and calculated. The synergic effects of the three NPs to RNAi of LsStre were evaluated based on the corrected mortality rates of the 2nd instar nymphs after RNAi with dsLsStre mediated by different NPs compared with that caused by naked dsLsStre (the control). 【Results】 All the three NPs could load dsEgfp, with the loading efficiencies of the three NPs coupled with dsEgfp all over 95%. All the three NPs showed low toxicity to the 2nd instar nymphs of L. striatellus. The qPCR results demonstrated that compared to the control group fed with naked dsLsStre not encapsulated with nanoparticles (causing 46% inhibition rate on the expression level of LsStre), chitosan and CQD significantly increased the RNAi efficiency of LsStre via feeding delivery, causing 78% and 84% inhibition rate, respectively, on the expression level of

LsStre, but lipofectamine 2000 could not significantly enhance the RNAi efficiency of LsStre, causing 52% inhibition rate on the expression level of LsStre. Moreover, the feeding assays also showed that chitosan and CQD significantly improved the dsLsStre-mediated lethal effect on the 2nd instar nymphs of L. striatellus. At 6 d after being fed with chitosan and CQD NP-dsLsStre conjugates, the corrected mortality of the 2nd instar nymphs of L. striatellus reached 76% and 82%, respectively, and the synergism ratios (SRs) of chitosan-and CQD-mediated RNAi groups were 2.17 and 2.34, respectively, compared with the control fed with naked dsLsStre (causing 35% corrected mortality). Meanwhile, lipofectamine 2000 was the most inefficient carrier for inducing RNAi in L. striatellus (SR=1.09), and lipofectamine 2000-dsLsStre conjugates caused 38% mortality of the 2nd instar nymphs of L. striatellus. 【Conclusion】 Chitosan and CQD nanocarriers are

more efficient to deliver dsRNA than lipofectamine 2000 in L. striatellus. The results are helpful in elucidating the RNAi efficacy boosted by NPs and provide a theoretical basis and practical strategies for developing and screening novel nanocarriers to realize the eco-friendly pest control.

Key words: Laodelphax striatellus, trehalase, RNAi, nanoparticles, dsRNA, RNAi efficiency, synergism ratio