Molecular Identification of Cysteine and Trypsin Protease, Effect of Different Hosts on Protease Expression, and Rnai Mediated Silencing of Cysteine Protease Gene in the Sunn Pest

Abstract

Sunn pest, Eurygaster integriceps, is a serious pest of cereals in the wide area of the globe from Near and Middle East to East and South Europe and North Africa. This study described for the first time, identification of E. integriceps trypsin serine protease and cathepsin-L cysteine, transcripts involved in digestion, which might serve as targets for pest control management. A total of 478 and 500 base pair long putative trypsin and cysteine gene sequences were characterized and named Tryp and Cys, respectively. In addition, the tissue-specific relative gene expression levels of these genes as well as gluten hydrolase (Gl) were determined under different host kernels feeding conditions. Result showed that mRNA expression of Cys, Tryp, and Gl was significantly affected after feeding on various host plant species. Transcript levels of these genes were most abundant in the wheat-fed E. integriceps larvae compared to other hosts. The Cys transcript was detected exclusively in the gut, whereas the Gl and Tryp transcripts were detectable in both salivary glands and gut. Also possibility of Sunn pest gene silencing was studied by topical application of cysteine double-stranded RNA (dsRNA). The results indicated that topically applied dsRNA on fifth nymphal stage can penetrate the cuticle of the insect and induce RNA interference. The Cys gene mRNA transcript in the gut was reduced to 83.8% 2 days posttreatment. Also, it was found that dsRNA of Cys gene affected fifth nymphal stage development suggesting the involvement of this protease in the insect growth, development, and molting.

Proteases and nucleases involved in the biphasic digestion process of the brown marmorated stink bug, Halyomorpha halys (Hemiptera: Pentatomidae)

“…Consistent with findings for N. viridula (Lomate, & Bonning, 2016), cysteine protease activity was not detected in the salivary gland or saliva with cysteine protease-specific substrate (Figure 2). The higher serine protease activities in the H. halys salivary gland compared to those of the gut are consistent with the salivary serine protease activities in N. viridula and Podisus maculiventris, and trypsin expression in the Sunn pest, Eurygaster integriceps (Amiri, Bandani, & Alizadeh, 2016;Bell, Down, Edwards, Gatehouse, & Gatehouse, 2005;Lomate, & Bonning, 2016).…”

Differential expression of heat shock proteins and antioxidant enzymes in response to temperature, starvation, and parasitism in the Carob moth larvae, Ectomyelois ceratoniae (Lepidoptera: Pyralidae)

“…The sequences of the HSP70 and HSP90 cDNA were confirmed by a homology search of other HSP70 and HSP90 sequences known within the GenBank database of the National Center for Biotechnology Information (NCBI) website (http://www.ncbi.nlm.nih.gov/BLAST/). Phylogenetic trees were constructed by the neighbor-joining method with a Poisson correction model (1,000 bootstrap replications to check for repeatability of the results) using the MEGA 6.0 software [38].…”

Implementing the sterile insect technique with RNA interference – a review

“…eRNAi has been demonstrated to successfully achieve gene silencing in adults following ingestion of: (1) dsRNA‐expressing transgenic plants (Coleman et al., ; Tzin et al., ; Malik et al., ), (2) dsRNA‐expressing transgenic bacteria (Li et al., ; Taracena et al., ; Whitten et al., ), (3) dsRNA dissolved in solution (Coy et al., ; Ratzka et al., ; Shim et al., ), and (4) naked dsRNA overlaid on diet (Yi et al., ; Zheng et al., ). In addition, topical application to adults of dsRNA (Pridgeon et al., ; Killiny et al., ; Amiri et al., ) and infection with transgenic fungi (Chen et al., ) are reported. All methods have the potential for use in SIT development.…”