The challenge of RNAi-mediated control of hemipterans Olivier Christiaens and Guy Smagghe The post-transcriptional gene silencing mechanism RNA interference (RNAi) has potential as a crop protection strategy against important pest insects. Here we focus on Hemiptera pests, comprising some of the most devastating pest organisms as aphids, whiteflies, psyllids, bedbugs and kissing bugs. At first, a state-of-the-art overview is provided of the progress in RNAi in Hemiptera, as well as on the challenges when developing new RNAi-based pest control strategies against hemipteran pests, such as the delivery of dsRNA and degradation in the insect body. We also discuss the variability in RNAi efficiency as observed between species and experiments, and the factors potentially responsible for this phenomenon. Addresses Department of Crop Protection, Faculty of Bioscience Engineering, Ghent University, 9000 Ghent, Belgium Corresponding author: Smagghe, Guy (guy.smagghe@ugent.be) Current Opinion in Insect Science 2014, 6:15–21 This review comes from a themed issue on Pests and resistance Edited by Guy Smagghe and Luc Swevers For a complete overview see the Issue and the Editorial Available online 28th September 2014 http://dx.doi.org/10.1016/j.cois.2014.09.012 2214-5745/# 2014 Elsevier Ltd. All right reserved. Introduction Since its discovery at the end of the previous century, the post-translational gene silencing mechanism RNA inter- ference (RNAi) had a huge impact on biological sciences. It provided researchers with an easy tool for investigating gene functions. Given its high sequence-dependent specificity, RNAi also quickly drew attention of crop protectors who recognized it as a novel and safe strategy in the fight against important pest organisms [1]. In this paper, we discuss the challenges that accompany this search for an effective RNAi-based crop protection strategy against Hemiptera. These sucking pests include important pests in agriculture and human health. Good examples of plant-sucking pests are aphids, whiteflies, planthoppers and psyllids because they cause extensive damage in important crops such as vegetables, grains and rice; the damage is dual due to the herbivorous nature of the insects, but also because they are important vectors of devastating diseases and viruses. In medicinal entomology, bedbugs and other blood-sucking bugs as the kissing bug Rhodnius are of increasing important in human health. In addition, many of these hemipteran pest populations have developed high levels of resistance to the conventional and modern insecticide groups. In a first part, a state-of-the-art overview is given of what is known on RNAi in different Hemiptera as studied so far. In second, we discuss a number of challenges with an effect on the RNAi efficacy: for example dsRNA delivery in the insect body, dsRNA degradation, the role of the RNAi machinery and possible interference by viruses. Requirements for RNAi Upon entry into a target cell, long dsRNA is cleaved by a RNaseIII-enzyme called Dicer into small interfering RNA (siRNA) molecules of 20–25 nucleotides long. Sub- sequently, these siRNAs are assembled with an Argo- naute protein to form the RNA-induced silencing complex (RISC), which then targets and destroys the endogenous mRNA that is complementary to the guide strand in the RISC. A number of factors determine whether a strong silencing effect can be achieved. First, dsRNA has to be taken up into the target cell in order to elicit a response. Second, this dsRNA must be processed by the RNAi machinery within the cell, eventually creating secondary 20–25 bp siRNA molecules that can act as triggers for sequence- specific enzymatic degradation of endogenous mRNA. In order to achieve a strong silencing effect in whole tissues or entire organisms, it is required that the RNAi signal is spread throughout the body. This is called systemic RNAi [1]. Finally, amplification of this signal, as observed in Caenorhabditis elegans, greatly facilitates a strong and long- lasting silencing effect [2]. For more information on general experimental design for RNAi and on factors affecting RNAi efficacy in insects in general, we refer to [3  ]. The current review is mainly focusing on the challenges faced when searching for a hemipteran pest control application. RNAi in Hemiptera To date, RNAi has mainly been used as a research tool. Interestingly, these experiments suggest that there is a substantial amount of variability in RNAi efficiency be- tween different species. In general, several species belonging to Coleoptera, such as the red flour beetle Tribolium castaneum, Colorado potato beetle Leptinotarsa decemlineata and western corn rootworm Diabrotica virgi- fera, possess a robust and highly sensitive RNAi system [4,5  ,6]. In contrast, the success in Diptera and Lepi- doptera is rather variable [7,8]. Available online at www.sciencedirect.com ScienceDirect www.sciencedirect.com Current Opinion in Insect Science 2014, 6:15–21