Characteristics of piRNA loci for triggering piRNA-induced gene silencing in hemipterans

RNA-interference-mediated biopesticide approaches for insect control are exclusively siRNA dependent and deploy long double-stranded RNAs (dsRNA) to trigger gene silencing. Two additional classes of small RNAs, the microRNAs, and piRNAs, are also present in all insects. Though piRNAs were initially considered gonad-specific, piRNAs in many insects are now known to have extensive roles in somatic tissues, where in addition to controlling the transposable elements (TEs), they regulate protein-coding genes. This offers an opportunity to exploit somatic piRNAs as a previously unexplored RNAi strategy, recently found effective in the whitefly Bemisia tabaci. The piRNA biogenesis involves selective recruitment of target RNAs originated from the piRNA cluster, into the pathway. This selective process was exploited in the whitefly study by attaching a known piRNA sequence to the sequence of the gene of interest so that piRNAs were produced from the entire fusion sequence. The objective of this study is to identify the ideal piRNA loci in Diaphorina citri (Kuwayama), the Asian citrus psyllid (ACP), and Bactericera cockerelli (Sulc.), the tomato or potato psyllid (PoP) to exploit the somatic piRNAs for RNAi. Using small RNA sequencing data, 5686 and 6357 endogenous piRNA-producing loci were identified bioinformatically from the ACP and PoP genomes, respectively. Characteristics of the loci, including relative abundance of phased and ping-pong piRNAs, strand bias, and presence within a piRNA cluster were assessed bioinformatically and by manual inspection, and two loci were selected from both species to generate fusion constructs and test for the efficacy of piRNA-triggered environmental RNAi in psyllids.