Insecticides are key weapons for the control of pests. Large scale use of insecticides is harmful to the ecosystem, which is made up of a wide range of species and environments. MicroRNAs (miRNAs) are a class of endogenous single-stranded noncoding small RNAs in length of 20-24 nucleotides (nt), which extensively regulate expression of genes at transcriptional and post-transcriptional levels. The current research on miRNA-induced insecticide resistance reveals that dysregulated miRNAs cause significant changes in detoxification genes, particularly cytochrome P450s. Meanwhile, insecticide-induced changes in miRNAs are related to the decline of honeybees and threatened the development of zebrafish and other animals. Additionally, miRNAs are involved in insecticide-induced cytotoxicity, and dysregulated miRNAs are associated with human occupational and environmental exposure to insecticides. Therefore, miRNAs are valuable novel biomarkers of insecticide exposure, and they are potential factors to explain the toxicological effects of insecticides.

MicroRNAs as Potential Biomarkers of Insecticide Exposure: A Review.

Variable RNA interference (RNAi) efficiencies limit RNAi-based pest management strategies for many pests. Previous efforts to understand mechanisms contributing to low RNAi efficiency indicate that double-stranded RNA (dsRNA) is degraded in the European corn borer (ECB), Ostrinia nubilalis, due to nuclease activity. To investigate the contribution of dsRNA-degrading endonucleases (dsRNases) and lepidopteran-specific RNAi efficiency-related nucleases (REases) to dsRNA instability and low RNAi efficiency in ECB, five complementary DNAs putatively encoding four dsRNases (OndsRNase1, 2, 3, and 4) and one REase (OnREase) were sequenced. Characterization of these transcripts revealed that substrate specificity might vary among the four dsRNases due to different amino acid combinations in the substrate-binding sites. Gene expression analysis indicated that OndsRNase2 and OnREase were highly expressed in the larval gut, and OndsRNase1 showed the highest expression in hemolymph, especially in older developmental stages. Transcript level analysis after dsRNA exposure revealed that expression of OnREase rapidly increased upon dsRNA ingestion or injection, whereas OndsRNase4 expression only increased after long-term ingestion of dsRNA. While the biological function of these nucleases remains to be verified, our results suggest that OnREase and OndsRNase2, and OndsRNase1 and OndsRNase4 may be responsible for degradation of dsRNAs in the ECB gut and hemolymph, respectively, thereby contributing to low RNAi efficiency.

https://www.mdpi.com/2075-4450/11/10/652/pdf

Molecular Characterizations of Double-Stranded RNA Degrading Nuclease Genes from Ostrinia nubilalis.

Bt Resistance: Characterization and Strategies for GM Crops Producing Bacillus thuringiensis Toxins

Insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) are used globally as bioinsecticides and in genetically engineered Bt crops to manage key insect pests. Transgenic Bt crops have been cumulatively planted on more than 1.5 billion hectares for more than 27 years, providing enhanced pest suppression, improved yields, increased farmer profits, and reduced environmental and health risks associated with the decreased use of conventional chemical insecticides. These benefits are reduced or lost when pests evolve resistance to Bt toxins in the field. Understanding the molecular genetic basis of resistance can be useful for monitoring, managing, and countering pest resistance to Bt crops. To date, 26 cases of field-evolved practical resistance and 18 cases of early warning of resistance to Bt crops have been reported, but the molecular genetic basis is known for only 9 of the 44 resistance cases to Bt crops. Here, we provide a comprehensive update on the molecular genetic mechanisms for these nine cases of field-evolved resistance to Bt crops and the numerous cases of laboratory-selected resistance to Bt toxins. We highlight lab-selected resistance involving regulation of Bt receptor genes and how recent research shows that regulatory pathways can be coopted to cause resistance. We further review recent technological advances in genomics and reverse genetics that have improved our ability to monitor and understand of the molecular genetic basis of pest resistance. The findings outlined here describe not only the complex genetic nature of Bt resistance, but also highlights new areas of exploration into the fundamental evolutionary basis of resistance mechanisms, which are important for sustaining the efficacy of currently available and future transgenic Bt crops. 

Mechanisms and molecular genetics of insect resistance to insecticidal proteins from Bacillus thuringiensis

Midgut de novo transcriptome analysis and gene expression profiling of Achaea janata larvae exposed with Bacillus thuringiensis (Bt)-based biopesticide formulation

Transgenic maize hybrids that express insecticidal Bacillus thuringiensis (Bt) crystalline (Cry) protein toxins effectively protect against the European corn borer, Ostrinia nubilalis, a devastating maize pest. Field monitoring and laboratory selections have detected varying levels of O. nubilalis resistance to Cry1Ab toxin. MicroRNAs (miRNAs) are short noncoding RNAs that are involved in post-transcriptional gene regulation. Their potential roles in the evolution of Bt resistance, however, remain largely unknown. Sequencing of small RNA libraries from the midgut of Cry1Ab-susceptible and resistant O. nubilalis larvae resulted in the discovery of 277 miRNAs, including 248 conserved and 29 novel. Comparative analyses of miRNA expression profiles between the laboratory strains predicted 26 and nine significantly up- and down-regulated transcripts, respectively, in the midgut of Cry1Ab resistant larvae. Amongst 15 differentially regulated miRNAs examined by quantitative real-time PCR, nine (60%) were validated as cosegregating with Cry1Ab resistance in a backcross progeny. Differentially expressed miRNAs were predicted to affect transcripts involved in cell membrane components with functions in metabolism and binding, and the putative Bt-resistance genes aminopeptidase N and cadherin. These results lay the foundation for future investigation of the potential role of miRNAs in the evolution of Bt resistance.

microRNA profiling between Bacillus thuringiensis Cry1Ab-susceptible and -resistant European corn borer, Ostrinia nubilalis (Hübner).

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microRNA profiling between Bacillus thuringiensis Cry1Ab-susceptible and -resistant European corn borer, Ostrinia nubilalis (Hübner).