COIS-75; NO. OF PAGES 8 Please cite this article in press as: Liu S, et al.: RNA virus discovery in insects, Curr Opin Insect Sci (2015), http://dx.doi.org/10.1016/j.cois.2014.12.005 RNA virus discovery in insects Sijun Liu, Yuting Chen and Bryony C Bonning The advent of next generation sequencing (NGS) technology has allowed for significant advances in insect RNA virus discovery, particularly for identification of covert viruses and for identification of previously undescribed virus groups. Recent research has revealed the complexity and diversity both of viral populations within an insect, and of different virus groups infecting insects. While appropriate verification of the ability of a given virus to infect a putative host is essential, this is frequently challenging for newly discovered viruses. In this chapter, we describe recently discovered RNA viruses of insects, optimization of NGS data analysis for virus discovery, and challenges associated with virus nomenclature. Addresses Department of Entomology, Iowa State University, Ames, IA 50011, USA Corresponding author: Liu, Sijun (sliu@iastate.edu) Current Opinion in Insect Science 2015, 8:xx–yy This review comes from a themed issue on Parasites/Parasitoids/ Biological Control Edited by Bryony C Bonning http://dx.doi.org/10.1016/j.cois.2014.12.005 2214-5745/# 2015 Published by Elsevier Inc. Introduction Insects as major agricultural pests and vectors of human, animal and plant diseases have tremendous negative impacts both on the economy and on human life. The pathogens of insects can be exploited for suppression of pestiferous insect populations, and insect viruses in par- ticular have great potential for use as biological control agents [1]. In the past decade, next generation sequenc- ing (NGS) or deep sequencing technology has fundamen- tally changed the approach to virus discovery, and has become a primary tool for virus discovery in animals including both humans [2  ,3] and arthropods [4  ,5]. NGS technology enables researchers to de novo assemble viral sequences without reference to viral sequences (nucleotides or proteins). NGS has been used to investi- gate arthropod viral sequence populations from insects and from insect-derived cell lines [6], as well as from insectivorous bat species [7,8]. As an additional benefit, assembly of RNA virus genomes from NGS data can result in more complete sequence for 5 0 -untranslated regions (UTR) than the use of 5 0 -RACE, which can be blocked by RNA secondary structure in the 5 0 UTR [5,9] (see also Carrillo-Tripp et al., this volume). In this review, we describe recently discovered insect RNA viruses and discuss limitations associated with the use of NGS for virus detection. We also discuss optimization of NGS data analysis to facilitate virus discovery. Metagenomic analyses for virus sequence discovery Metagenomic analysis, the study of genomic material recovered from environmental samples, has been used to identify organisms associated with economically im- portant insects, such as honey bees (Apis mellifera) [10]. Honey bees are known to be affected by more than 23 different viruses. A metagenomic analysis of RNA collected from 20 honey bee colonies in a large-scale migratory U.S. beekeeping operation resulted in identifi- cation of two dicistroviruses and two other new viruses distantly related to viruses belonging to Nodaviridae [11]. In a separate metagenomic analysis of Spanish honey bees, three RNA viruses, aphid lethal paralysis virus, Israeli acute paralysis virus and Lake Sinai virus were detected [12]. Deep sequencing of small RNA (sRNA) can also be used for discovery of viral sequences [6]. Investigation of sRNA sequenced from field-collected mosquitoes and chironomids (Diptera) to assess viral diversity resulted in the discovery of many viral sequences related to at least 10 viruses. The genome of only one picorna-like virus (Anopheline-associated C virus) was nearly complete however [13]. While metage- nomic analyses can be used to identify many viral sequences and provide an indication of viral diversity within insect populations, the majority of the assembled viral genomic sequences are partial or incomplete gen- omes. Novel viruses identified from NGS data analysis The rate of virus discovery has increased significantly with the adoption of NGS technologies with many new viruses and novel isolates of known viruses discovered by using this approach in recent years (Table 1). Viruses identified from pests of medical importance, such as flavivirus and others in the order Nidovirales of mosqui- toes, ticks and sandflies, were recently reviewed [4  ], and hence are not included here. Iflaviridae Iflaviruses are arthropod-infecting viruses with positive- sense, single-stranded RNA genomes (+ssRNA). Iflaviruses are commonly found in hymenopteran, lepidopteran, and Available online at www.sciencedirect.com ScienceDirect www.sciencedirect.com Current Opinion in Insect Science 2015, 8:1–8