Deep Sequencing of the Transcriptomes of Soybean Aphid and Associated Endosymbionts

Abstract

BackgroundThe soybean aphid has significantly impacted soybean production in the U.S. Transcriptomic analyses were conducted for further insight into leads for potential novel management strategies.Methodology/Principal FindingsTranscriptomic data were generated from whole aphids and from 2,000 aphid guts using an Illumina GAII sequencer. The sequence data were assembled de novo using the Velvet assembler. In addition to providing a general overview, we demonstrate (i) the use of the Multiple-k/Multiple-C method for de novo assembly of short read sequences, followed by BLAST annotation of contigs for increased transcript identification: From 400,000 contigs analyzed, 16,257 non-redundant BLAST hits were identified; (ii) analysis of species distributions of top non-redundant hits: 80% of BLAST hits (minimum e-value of 1.0-E3) were to the pea aphid or other aphid species, representing about half of the pea aphid genes; (iii) comparison of relative depth of sequence coverage to relative transcript abundance for genes with high (membrane alanyl aminopeptidase N) or low transcript abundance; (iv) analysis of the Buchnera transcriptome: Transcripts from 57.6% of the genes from Buchnera aphidicola were identified; (v) identification of Arsenophonus and Wolbachia as potential secondary endosymbionts; (vi) alignment of full length sequences from RNA-seq data for the putative salivary gland protein C002, the silencing of which has potential for aphid management, and the putative Bacillus thuringiensis Cry toxin receptors, aminopeptidase N and alkaline phosphatase.Conclusions/SignificanceThis study provides the most comprehensive data set to date for soybean aphid gene expression: This work also illustrates the utility of short-read transcriptome sequencing and the Multiple-k/Multiple-C method followed by BLAST annotation for rapid identification of target genes for organisms for which reference genome sequences are not available, and extends the utility to include the transcriptomes of endosymbionts.

Heritable symbionts in a world of varying temperature

“…For instance, extensive genome reduction in Buchnera is reflected in this symbiont producing just 5 heat shock proteins, a substantial decrease compared with the 75 produced by its free-living and more thermotolerant relative Escherichia coli (Bronikowski et al, 2001;Wilcox et al, 2003;Pérez-Brocal et al, 2006;Liu et al, 2012). More widely, accumulation of deleterious mutations in remaining genes (Moran, 1996) is reflected in weaker secondary and tertiary structure of proteins in Buchnera (van Ham et al, 2003), with the result that the function of proteins in obligate symbionts is disproportionately impaired at elevated temperatures compared with proteins encoded in the host genome.…”

Achievements and Challenges in Legume Breeding for Pest and Disease Resistance

“…This will be complemented by efforts to sequence the genomes of key pathogens and pests such as the genome sequences of Fusarium oxysporum , Verticillium albo-atrum (Klosterman et al, 2011), Botrytis cinerea (Staats and van Kan, 2012), Rhizoctonia solani (Wibberg et al, 2012), Phytophthora sojae (Tyler et al, 2006), Macrophomina phaseolina (Islam et al, 2012), Ralstonia solanacearum (Remenant et al, 2010), Pseudomonas syringae (Gardiner et al, 2013), Curtobacterium flaccumfaciens (Flanagan et al, 2013), Xylella fastidiosa , Meloidogyne incognita (Abad et al, 2008), Meloidogyne hapla , alfalfa mosaic virus (Cornelissen et al, 1983), cucumber mosaic virus (Owen et al, 1990), pea enation mosaic virus (Demler and de Zoeten, 1991), bean leafroll virus (Domier et al, 2002), pea seed-borne mosaic virus (Johansen et al, 1991) and the pea and soybean aphids (International Aphid Genomics Consortium, 2010;Liu et al, 2012). In addition, further in-depth studies on the biology of the pests and pathogens are required.…”

RNA virus discovery in insects

“…Insect viruses in which a permuted RdRP has previously been observed include tetraviruses [23] and Drosophila A virus [24]. Several new insect viruses with permuted RdRP have now been discovered including Aphis glycines virus (AGV) identified from the soybean aphid [25,26 ], Dezidougou virus, Santana virus [27 ] and Tanay virus [28] isolated from mosquitoes. The novel mosquito viruses with permuted RdRPs were classified into a proposed new taxon of insect-specific viruses, Negevirus, which currently has 6 members [27 ] and a possible new member [28].…”

“…The novel mosquito viruses with permuted RdRPs were classified into a proposed new taxon of insect-specific viruses, Negevirus, which currently has 6 members [27 ] and a possible new member [28]. Interestingly, the closest but still distant virus relative of Negeviruses is a plant virus, Citrus leprosis virus C (CiLV-C) [25], which encodes an RdRP with the more common A ! B !…”