Survival and Feeding Rates of Four Aphid Species (Hemiptera: Aphididae) on Various Sucrose Concentrations in Diets.

Different concentrations of sucrose were used to investigate how survival and feeding was affected in four species of aphids (Hemiptera: Aphididae). Seven sucrose concentrations were evaluated in feeding chambers fitted with parafilm membranes and infested with nymphs of Aphis glycines Matsumura, Diuraphis noxia Kurdjumov, Myzus persicae Sulzer, or Schizaphis graminum Rondani at 25 °C and a photoperiod of 14:10 (L:D) h. Survival on each diet was recorded 1, 3, 5, 7, 9, and 11 d. Diet volumes (µl) consumed and amounts of honeydew produced were then determined. Aphid survival differed significantly by concentration, time (d), and aphid species. Aphis glycines survival was highest (83.8%) on 30% sucrose although percent survival in the 70's occurred on concentrations ranging from 15-25%. Diuraphis noxia survival was highest on the 15 and 20% sucrose concentrations. Survival for Myzus persicae was optimal on 20% sucrose (92%) but did not differ that much (5%) on concentrations of 15-35%. Schizaphis graminum survival (93.0-93.6%) was highest on sucrose concentrations ranging from 20-30% sucrose. Myzus persicae and S. graminum, which feed on a wide-range of host plants, were overall more adapted to feeding on a wider range of sucrose concentrations than the more host-restricted aphid species, A. glycines. Diet consumption by A. glycines did not vary on the sucrose concentrations, but D. noxia and M. persicae exhibited increased consumption on diets that provided optimal survival. Results will aid in the design of short-term studies using sucrose-only diets to evaluate effects of bioactive materials on aphid survival for up to 11 d.

The genome of Diuraphis noxia, a global aphid pest of small grains.

BACKGROUND: The Russian wheat aphid, Diuraphis noxia Kurdjumov, is one of the most important pests of small grains throughout the temperate regions of the world. This phytotoxic aphid causes severe systemic damage symptoms in wheat, barley, and other small grains as a direct result of the salivary proteins it injects into the plant while feeding. RESULTS: We sequenced and de novo assembled the genome of D. noxia Biotype 2, the strain most virulent to resistance genes in wheat. The assembled genomic scaffolds span 393 MB, equivalent to 93% of its 421 MB genome, and contains 19,097 genes. D. noxia has the most AT-rich insect genome sequenced to date (70.9%), with a bimodal CpG(O/E) distribution and a complete set of methylation related genes. The D. noxia genome displays a widespread, extensive reduction in the number of genes per ortholog group, including defensive, detoxification, chemosensory, and sugar transporter groups in comparison to the Acyrthosiphon pisum genome, including a 65% reduction in chemoreceptor genes. Thirty of 34 known D. noxia salivary genes were found in this assembly. These genes exhibited less homology with those salivary genes commonly expressed in insect saliva, such as glucose dehydrogenase and trehalase, yet greater conservation among genes that are expressed in D. noxia saliva but not detected in the saliva of other insects. Genes involved in insecticide activity and endosymbiont-derived genes were also found, as well as genes involved in virus transmission, although D. noxia is not a viral vector. CONCLUSIONS: This genome is the second sequenced aphid genome, and the first of a phytotoxic insect. D. noxia's reduced gene content of may reflect the influence of phytotoxic feeding in shaping the D. noxia genome, and in turn in broadening its host range. The presence of methylation-related genes, including cytosine methylation, is consistent with other parthenogenetic and polyphenic insects. The D. noxia genome will provide an important contrast to the A. pisum genome and advance functional and comparative genomics of insects and other organisms.

Potential transmission of Pantoea spp. and Serratia marcescens (Enterobacteriales: Enterobacteriaceae) to plants by Lygus hesperus (Hemiptera: Miridae).

Lygus hesperus Knight (Hemiptera: Miridae) is a key agricultural pest in the western United States. In a recent study, proteins from Pantoea ananatis and Serratia marcescens (Enterobacteriales: Enterobacteriaceae) were identified in diet that was stylet probed and fed on by L. hesperus adults. P. ananatis and S. marcescens are ubiquitous bacteria that infect a wide range of crops. The objective of our study was to determine whether L. hesperus transfer P. ananatis and S. marcescens to food substrates during stylet-probing activities. Sucrose (5%) was spread under parafilm and exposed to adult L. hesperus for 24 h. Diet similarly prepared but not exposed to insects was used for controls. MacConkey agar was inoculated with stylet-probed or control diets and incubated at 25 degrees C. After 24 h, bacterial colonies were observed on agar that was inoculated with stylet-probed diet, but were not observed on agar inoculated with control diet. Isolated bacterial colonies were putatively identified as either Pantoea spp. or S. marcescens using the API 20e identification kit. These results indicate that L. hesperus is capable of vectoring P. ananatis and S. marcescens.

Variation in the salivary proteomes of differentially virulent greenbug (Schizaphis graminum Rondani) biotypes.

Greenbug (Schizaphis graminum Rondani) biotypes are classified by their differential virulence to wheat, barley, and sorghum varieties possessing greenbug resistance genes. Virulent greenbug biotypes exert phytotoxic effects upon their hosts during feeding, directly inducing physiological and metabolic alterations and accompanying foliar damage. Comparative analyses of the salivary proteomes of four differentially virulent greenbug biotypes C, E, G, and H showed significant proteomic divergence between biotypes. Thirty-two proteins were identified by LC-MS/MS; the most prevalent of which were three glucose dehydrogenase paralogs (GDH), lipophorin, complementary sex determiner, three proteins of unknown function, carbonic anhydrase, fibroblast growth factor receptor, and abnormal oocyte (ABO). Seven nucleotide-binding proteins were identified, including ABO which is involved in mRNA splicing. Quantitative variation among greenbug biotypes was detected in six proteins; two GDH paralogs, carbonic anhydrase, ABO, and two proteins of unknown function. Our findings reveal that the greenbug salivary proteome differs according to biotype and diverges substantially from those reported for other aphids. The proteomic profiles of greenbug biotypes suggest that interactions between aphid salivary proteins and the plant host result in suppression of plant defenses and cellular transport, and may manipulate transcriptional regulation in the plant host, ultimately allowing the aphid to maintain phloem ingestion. BIOLOGICAL SIGNIFICANCE: Greenbug (Schizaphis graminum Rondani, GB) is a major phytotoxic aphid pest of wheat, sorghum, and barley. Unlike non-phytotoxic aphids, GB directly damages its host, causing uniformly characteristic symptoms leading to host death. As saliva is the primary interface between the aphid and its plant host, saliva is also the primary aphid biotypic determinant, and differences in biotypic virulence are the result of biotypic variations in salivary content. This study analyzed the exuded saliva of four distinct Greenbug biotypes with a range of virulence to crop lines containing greenbug resistance traits in order to identify differences between salivary proteins of the examined biotypes. Our analyses confirmed that the salivary proteomes of the examined greenbug biotypes differ widely, identified 32 proteins of the greenbug salivary proteome, and found significant proteomic variation between six identified salivary proteins. The proteomic variation identified herein is likely the basis of biotypic virulence, and the proteins identified can serve as the basis for functional studies into both greenbug-induced phytotoxic damage and into the molecular basis of virulence in specific GB biotypes. This article is part of a Special Issue entitled: SI: Proteomics of non-model organisms.

Proteomic analysis of secreted saliva from Russian wheat aphid (Diuraphis noxia Kurd.) biotypes that differ in virulence to wheat.

Diuraphis noxia, Russian Wheat Aphid (RWA), biotypes are classified by their differential virulence to wheat varieties containing resistance genes. RWA salivary proteins, unlike those of most aphid species, cause foliar damage and physiological alterations in plants. A comparative proteomic analysis of secreted saliva from four differentially virulent RWA biotypes identified thirty-four individual proteins. The five major proteins were glucose dehydrogenase, lipophorin, chitinase, CiV16.8g1-like, and lava lamp. Fourteen proteins quantitatively varied among biotypes; trehalase, ß-N-acetylglucosaminidase (chitinase), two separate glucose dehydrogenases, calreticulin, aminopeptidase, acetylglucosaminyltransferase, hydroxymethylglutaryl-CoA lyase, acyltransferase, ficolin-3, lava lamp, retinaldehyde-binding protein, and two proteins of unknown function. Fifty-four percent of spectral counts were associated with glucose dehydrogenase, which is thought to detoxify plant defensive compounds. One-dimensional electrophoresis detected nine protein bands from 9 to 60 kDa that quantitatively differed. Two-dimensional electrophoresis identified six major gel zones with quantitative and qualitative variance in proteins. Our findings reveal that the salivary proteome of RWA, a phytotoxic aphid, differs considerably from those reported for nonphytotoxic aphids. The potential roles of proteins used in the general plant feeding processes of aphids and those that are potential phytotoxins related to aphid virulence are discussed.

Transgene constructs lacking transcription termination signal induce efficient silencing of endogenous targets in Arabidopsis.

Efficient gene silencing tools are extremely important for genomics and biotechnology applications. A number of aberrant RNA species induce silencing of homologous genes; however, only the inverted repeat RNA is widely utilized for targeted gene silencing in plants. Development of additional silencing constructs would not only provide an alternative technology for single targets, it may also serve as a simpler platform for multi-target silencing. The present study found that aberrantly terminated sense transcripts serve as efficient inducers of gene silencing. A construct initially designed to carry out homologous recombination into the Phytochrome A locus of Arabidopsis induced instead a high rate of Phytochrome A silencing, resulting in a strong mutant phenotype. Attempts to decipher the silencing determinant of the construct revealed that absence of a proper transcription termination signal contributed to the silencing efficiency of the construct. Furthermore, silencing of an additional gene, Phytochrome B, was conferred by a separate construct of a similar design. Proper placement of a transcription termination signal in the silencing construct abolished its silencing ability. Therefore, transcription of aberrantly terminated transcripts was deduced to be the cause of the elevated rate of silencing observed in transgenic lines. These transcripts could be produced either by direct transcription or read-through transcription. Since silencing resulted in strong mutant phenotypes in the majority of transgenic lines, terminator-less constructs are proposed to serve as efficient tools for knocking out endogenous targets.

Transgene-induced silencing of Arabidopsis phytochrome A gene via exonic methylation.

Transgene-induced promoter or enhancer methylation clearly retards gene activity. While exonic methylation of genes is frequently observed in the RNAi process, only sporadic evidence has demonstrated its definitive role in gene suppression. Here, we report the isolation of a transcriptionally suppressed epi-allele of the Arabidopsis thaliana phytochrome A gene (PHYA) termed phyA' that shows methylation only in symmetric CG sites resident in exonic regions. These exonic modifications confer a strong phyA mutant phenotype, characterized by elongated hypocotyls in seedlings grown under continuous far-red light. De-methylation of phyA' in the DNA methyl transferase I (met1) mutant background increased PHYA expression and restored the wild-type phenotype, confirming the pivotal role of exonic CG methylation in maintaining the altered epigenetic state. PHYA epimutation was apparently induced by a transgene locus; however, it is stably maintained following segregation. Chromatin immunoprecipitation assays revealed association with dimethyl histone H3 lysine 9 (H3K9me2), a heterochromatic marker, within the phyA' coding region. Therefore, transgene-induced exonic methylation can lead to chromatin alteration that affects gene expression, most likely through reduction in the transcription rate.