Transcriptome Analysis and Knockdown of the Juvenile Hormone Esterase Gene Reveal Abnormal Feeding Behavior in the Sugarcane Giant Borer.

The sugarcane giant borer (SGB), Telchin licus licus, is a pest that has strong economic relevance for sugarcane producers. Due to the endophytic behavior of the larva, current methods of management are inefficient. A promising biotechnological management option has been proposed based on RNA interference (RNAi), a process that uses molecules of double-stranded RNA (dsRNA) to specifically knock down essential genes and reduce insect survival. The selection of suitable target genes is often supported by omic sciences. Studies have shown that genes related to feeding adaptation processes are good candidates to be targeted by RNAi for pest management. Among those genes, esterases are highlighted because of their impact on insect development. In this study, the objective was to evaluate the transcriptome responses of the SGB's gut in order to provide curated data of genes that could be used for pest management by RNAi in future studies. Further, we validated the function of an esterase-coding gene and its potential as a target for RNAi-based control. We sequenced the gut transcriptome of SGB larvae by Illumina HiSeq and evaluated its gene expression profiles in response to different diets (sugarcane stalk and artificial diet). We obtained differentially expressed genes (DEGs) involved in detoxification, digestion, and transport, which suggest a generalist mechanism of adaptation in SGB larvae. Among the DEGs, was identified and characterized a candidate juvenile hormone esterase gene (Tljhe). We knocked down the Tljhe gene by oral delivery of dsRNA molecules and evaluated gene expression in the gut. The survival and nutritional parameters of the larvae were measured along the developmental cycle of treated insects. We found that the gene Tljhe acts as a regulator of feeding behavior. The knockdown of Tljhe triggered a forced starvation state in late larval instars that significantly reduced the fitness of the larvae. However, the mechanism of action of this gene remains unclear, and the correlation between the expression of Tljhe and the levels of juvenile hormone (JH) metabolites in the hemolymph of the SGB must be assessed in future research.

Comparative gut transcriptome analysis of Diatraea saccharalis in response to the dietary source.

The sugarcane borer (Diatraea saccharalis, Fabricius, 1794) is a devastating pest that causes millions of dollars of losses each year to sugarcane producers by reducing sugar and ethanol yields. The control of this pest is difficult due to its endophytic behavior and rapid development. Pest management through biotechnological approaches has emerged in recent years as an alternative to currently applied methods. Genetic information about the target pests is often required to perform biotechnology-based management. The genomic and transcriptomic data for D. saccharalis are very limited. Herein, we report a tissue-specific transcriptome of D. saccharalis larvae and a differential expression analysis highlighting the physiological characteristics of this pest in response to two different diets: sugarcane and an artificial diet. Sequencing was performed on the Illumina HiSeq 2000 platform, and a de novo assembly was generated. A total of 27,626 protein-coding unigenes were identified, among which 1,934 sequences were differentially expressed between treatments. Processes such as defence, digestion, detoxification, signaling, and transport were highly represented among the differentially expressed genes (DEGs). Furthermore, seven aminopeptidase genes were identified as candidates to encode receptors of Cry proteins, which are toxins of Bacillus thuringiensis used to control lepidopteran pests. Since plant-insect interactions have produced a considerable number of adaptive responses in hosts and herbivorous insects, the success of phytophagous insects relies on their ability to overcome challenges such as the response to plant defences and the intake of nutrients. In this study, we identified metabolic pathways and specific genes involved in these processes. Thus, our data strongly contribute to the knowledge advancement of insect transcripts, which can be a source of target genes for pest management.

Transcriptome and gene expression analysis of three developmental stages of the coffee berry borer, Hypothenemus hampei.

Coffee production is a global industry valued at approximately 173 billion US dollars. One of the main challenges facing coffee production is the management of the coffee berry borer (CBB), Hypothenemus hampei, which is considered the primary arthropod pest of coffee worldwide. Current control strategies are inefficient for CBB management. Although biotechnological alternatives, including RNA interference (RNAi), have been proposed in recent years to control insect pests, characterizing the genetics of the target pest is essential for the successful application of these emerging technologies. In this study, we employed RNA-seq to obtain the transcriptome of three developmental stages of the CBB (larva, female and male) to increase our understanding of the CBB life cycle in relation to molecular features. The CBB transcriptome was sequenced using Illumina Hiseq and assembled de novo. Differential gene expression analysis was performed across the developmental stages. The final assembly produced 29,434 unigenes, of which 4,664 transcripts were differentially expressed. Genes linked to crucial physiological functions, such as digestion and detoxification, were determined to be tightly regulated between the reproductive and nonreproductive stages of CBB. The data obtained in this study help to elucidate the critical roles that several genes play as regulatory elements in CBB development.

Improved drought stress tolerance in Arabidopsis by CRISPR/dCas9 fusion with a Histone AcetylTransferase.

Drought episodes decrease plant growth and productivity, which in turn cause high economic losses. Plants naturally sense and respond to water stress by activating specific signalling pathways leading to physiological and developmental adaptations. Genetically engineering genes that belong to these pathways might improve the drought tolerance of plants. The abscisic acid (ABA)-responsive element binding protein 1/ABRE binding factor (AREB1/ABF2) is a key positive regulator of the drought stress response. We investigated whether the CRISPR activation (CRISPRa) system that targets AREB1 might contribute to improve drought stress tolerance in Arabidopsis. Arabidopsis histone acetyltransferase 1 (AtHAT1) promotes gene expression activation by switching chromatin to a relaxed state. Stable transgenic plants expressing chimeric dCas9HAT were first generated. Then, we showed that the CRISPRa dCas9HAT mechanism increased the promoter activity controlling the ß-glucuronidase (GUS) reporter gene. To activate the endogenous promoter of AREB1, the CRISPRa dCas9HAT system was set up, and resultant plants showed a dwarf phenotype. Our qRT-PCR experiments indicated that both AREB1 and RD29A, a gene positively regulated by AREB1, exhibited higher gene expression than the control plants. The plants generated here showed higher chlorophyll content and faster stomatal aperture under water deficit, in addition to a better survival rate after drought stress. Altogether, we report that CRISPRa dCas9HAT is a valuable biotechnological tool to improve drought stress tolerance through the positive regulation of AREB1.

Pyrosequencing the Midgut Transcriptome of the Banana Weevil Cosmopolites sordidus (Germar) (Coleoptera: Curculionidae) Reveals Multiple Protease-Like Transcripts.

The banana weevil Cosmopolites sordidus is an important and serious insect pest in most banana and plantain-growing areas of the world. In spite of the economic importance of this insect pest very little genomic and transcriptomic information exists for this species. In the present study, we characterized the midgut transcriptome of C. sordidus using massive 454-pyrosequencing. We generated over 590,000 sequencing reads that assembled into 30,840 contigs with more than 400 bp, representing a significant expansion of existing sequences available for this insect pest. Among them, 16,427 contigs contained one or more GO terms. In addition, 15,263 contigs were assigned an EC number. In-depth transcriptome analysis identified genes potentially involved in insecticide resistance, peritrophic membrane biosynthesis, immunity-related function and defense against pathogens, and Bacillus thuringiensis toxins binding proteins as well as multiple enzymes involved with protein digestion. This transcriptome will provide a valuable resource for understanding larval physiology and for identifying novel target sites and management approaches for this important insect pest.