The results demonstrate that continuous ingestion of dsRNA with EDTA or liposome-encapsulated dsRNA can prevent dsRNA from being degraded enzymatically and suggest great potential for using these formulations in dsRNA delivery to use RNAi as a functional genomics tool or for pest management of stink bugs. © 2018 Society of Chemical Industry.

Variability in RNA-interference (RNAi) efficacy among different insect orders poses a big hurdle in the development of RNAi-based pest control strategies. The activity of double-stranded ribonucleases (dsRNases) in the digestive canal of insects can be one of the critical factors affecting oral RNAi efficacy. Here, the involvement of these dsRNases in the southern green stinkbug Nezaraviridula was investigated. First, the full sequence of the only dsRNase (NvdsRNase) in the transcriptome of N. viridula was obtained, followed by an oral feeding bioassay to evaluate the effect of NvdsRNase-silencing on oral RNAi efficacy. The NvdsRNase was first silenced in nymphs by NvdsRNase-dsRNA injections, followed by exposure to an artificial diet containing a lethal αCop-specific dsRNA. A significantly higher mortality was observed in the NvdsRNase-silenced nymphs when placed on the dsαCop-containing diet (65%) than in the dsGFP injected and dsαCop fed control (46.67%). Additionally, an ex vivo dsRNA degradation assay showed a higher stability of dsRNA in the saliva and midgut juice of NvdsRNase-silenced adults. These results provide evidence for the involvement of NvdsRNase in the reduction of oral RNAi efficacy in N. viridula. This information will be useful in further improving potential RNAi-based strategies to control this pest.

BACKGROUND: The southern green stinkbug, Nezara viridula (Hemiptera: Pentatomidae), is an important emerging polyphagous pest infesting soybean in the United States, Brazil and Argentina. The indiscriminate use of synthetic insecticides to control stinkbugs has limited the effectiveness of current management strategies. Alternatively, RNA interference (RNAi) has emerged as a novel mode of action to control pests in an eco-friendly manner. RESULTS: Here, we assessed the potential of RNAi technology by oral delivery of double-stranded RNA (dsRNA) for the control of N. viridula. Initially, ten candidate genes were tested by microinjection assay to select the best target genes for oral delivery. Seven genes resulted in more than 90% mortality after microinjection. To evaluate RNAi efficacy by oral delivery of dsRNA, five genes were tested by feeding the insects on gene-specific dsRNA mixed with an artificial diet. Significant mortality of 43% and 45% was observed after 14 days of treatment with ds⊍Cop and dsvATPase A, respectively. To elucidate the lower RNAi efficacy via oral delivery of dsRNA, ex vivo dsRNA degradation in the saliva and the midgut juice was performed, which indicated that the reduced RNAi efficacy is accompanied by a rapid degradation of dsRNA by digestive secretions. CONCLUSION: This study proves that RNAi can be triggered by orally delivered dsRNA in N. viridula and can be exploited to control this economically important pest. The reduced stability of dsRNA in saliva and midgut that was observed indicates a need to further improve RNAi efficacy, for example by use of specific formulations.

Burgeoning the human population with its required food demand created a burden on ever-decreasing cultivated land and our food production systems. This situation prompted plant scientists to breed crops in a short duration with specific traits. Marker-assisted selection (MAS) has emerged as a potential tool to achieve desirable results in plants with the help of molecular markers and improves the traits of interest in a short duration. The MAS has comprehensively been used in plant breeding to characterize germplasm, diversity analysis, trait stacking, gene pyramiding, multi-trait introgression, and genetic purity of different cereals, pulses, oilseeds, and fiber crops, etc. Mapping studies pointed out several marker-trait associations from different crop species, which specifies the potential application of MAS in accelerating crop improvement. This chapter presents an overview of molecular markers, their genesis, and potential use in plant breeding.

Draft genomes of the fungal species Ambrosiella cleistominuta, Cercospora brassicicola, C. citrullina, Physcia stellaris, and Teratosphaeria pseudoeucalypti are presented. Physcia stellaris is an important lichen forming fungus and Ambrosiella cleistominuta is an ambrosia beetle symbiont. Cercospora brassicicola and C. citrullina are agriculturally relevant plant pathogens that cause leaf-spots in brassicaceous vegetables and cucurbits respectively. Teratosphaeria pseudoeucalypti causes severe leaf blight and defoliation of Eucalyptus trees. These genomes provide a valuable resource for understanding the molecular processes in these economically important fungi.