Hairpin Plum pox virus coat protein (hpPPV-CP) structure in ‘HoneySweet’ C5 plum provides PPV resistance when genetically engineered into plum (Prunus domestica) seedlings

The genetically engineered plum ‘HoneySweet’ (aka C5) has proven to be highly resistant to Plum pox virus (PPV) for over 10 years in field trials. The original vector used for transformation to develop ‘HoneySweet’ carried a single sense sequence of the full length PPV coat protein (ppv-cp) gene, yet DNA blot analyses indicated that there was an inserted copy of the ppv-cp that appeared to be an inverted repeat structure. Since the resistance mechanism of ‘HoneySweet’ was found to be based on post-transcriptional gene silencing (PTGS), it was hypothesized that the inverted repeat structure conferred the resistance to PPV in ‘HoneySweet’. Sequencing of the transgene insertions confirmed the presence of an inverted repeat of the PPV-CP sequence. We hypothesized that transcription from this structure produced a hairpin (hp) RNA that was responsible for PTGS of the transgene and the destruction of PPV viral RNA resulting in the high level of resistance to PPV infection. In order to confirm this hypothesis the hpPPV-CP insert was cloned from ‘HoneySweet’ and transferred into ‘Bluebyrd’ plum seedlings through Agrobacterium tumefaciens transformation of hypocotyl slices. The introduced DNA contained the CP inverted repeat flanked by 35S promoters on either end. Transgenic plum plants containing single or multiple copies of this hp insert were inoculated with PPV D isolated from Pennsylvania, USA. PPV infection was evaluated through three cycles of cold-induced dormancy (CID) by symptom expression and by two or more ELISA and PCR tests. Of the 18 plants evaluated, eight were always virusfree, five occasionally had weak or moderate infections, and five plants were clearly infected in multiple tests. While all plants of some clones were virus-free others had a mix of uninfected and mildly infected plants of the same clone. Most of the resistant plants contained a single copy of the hp construct. These data strongly support the hypothesis that the hp structure of the ppv-cp insert in ‘HoneySweet’ plum can confer PPV resistance.Keywords: Breeding, gene silencing, Rosaceae, shark

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.Peer reviewedBiochemistry and Molecular Biolog

An Assessment Model for Rating High-Threat Crop Pathogens

Natural, accidental, and deliberate introductions of nonindigenous crop pathogens have become increasingly recognized as threats to the U.S. economy. Given the large number of pathogens that could be introduced, development of rapid detection methods and control strategies for every potential agent would be extremely difficult and costly. Thus, to ensure the most effective direction of resources a list of high-threat pathogens is needed. We address development of a pathogen threat assessment model based on the analytic hierarchy process (AHP) that can be applied worldwide, using the United States as an illustrative example. Previously, the AHP has been shown to work well for strategic planning and risk assessment. Using the collective knowledge of subject matter expert panels incorporated into commercial decision-making software, 17 biological and economic criteria were determined and given weights for assessing the threat of accidental or deliberately introduced pathogens. The rating model can be applied by experts on particular crops to develop threat lists, especially those of high priority, based on the current knowledge of individual diseases

Transferring potyvirus coat protein genes through hybridization of transgenic plants to produce plum pox virus resistant plums (Prunus domestica L.)

International audienc