Changes in Disease Resistance Phenotypes Associated With Plant Physiological Age Are Not Caused by Variation in R Gene Transcript Abundance

Foliar late blight is one of the most important diseases of potato. Foliar blight resistance has been shown to change as a plant ages. In other pathosystems, resistance (R) gene transcript levels appear to be correlated to disease resistance. The cloning of the broad-spectrum, foliar blight resistance gene RB provided the opportunity to explore how foliar blight resistance and R-gene transcript levels vary with plant age. Plants of Solanum bulbocastanum PT29, from which RB, including the native promoter and other flanking regions, was cloned, and S. tuberosum cv. Dark Red Norland (nontransformed and RB-transformed) representing three different developmental stages were screened for resistance to late blight and RB transcript levels. Preflowering plants of all genotypes exhibited the highest levels of resistance, followed by postflowering and near-senescing plants. The RB transgene significantly affected resistance, enhancing resistance levels of all RB-containing lines, especially in younger plants. RB transgene transcripts were detected at all plant ages, despite weak correlation with disease resistance. Consistent transcript levels in plants of different physiological ages with variable levels of disease resistance demonstrate that changes in disease-resistance phenotypes associated with plant age cannot be attributed to changes in R-gene transcript abundance

Higher copy numbers of the potato RB transgene correspond to enhanced transcript and late blight resistance levels.

Late blight of potato ranks among the costliest of crop diseases worldwide. Host resistance offers the best means for controlling late blight, but previously deployed single resistance genes have been short-lived in their effectiveness. The foliar blight resistance gene RB, previously cloned from the wild potato Solanum bulbocastanum, has proven effective in greenhouse tests of transgenic cultivated potato. In this study, we examined the effects of the RB transgene on foliar late blight resistance in transgenic cultivated potato under field production conditions. In a two-year replicated trial, the RB transgene, under the control of its endogenous promoter, provided effective disease resistance in various genetic backgrounds, including commercially prominent potato cultivars, without fungicides. RB copy numbers and transcript levels were estimated with transgene-specific assays. Disease resistance was enhanced as copy numbers and transcript levels increased. The RB gene, like many other disease resistance genes, is constitutively transcribed at low levels. Transgenic potato lines with an estimated 15 copies of the RB transgene maintain high RB transcript levels and were ranked among the most resistant of 57 lines tested. We conclude that even in these ultra–high copy number lines, innate RNA silencing mechanisms have not been fully activated. Our findings suggest resistance-gene transcript levels may have to surpass a threshold before triggering RNA silencing. Strategies for the deployment of RB are discussed in light of the current research

Molecular phylogeny, diagnostics, and diversity of plant-parasitic nematodes of the genus Hemicycliophora (Nematoda: Hemicycliophoridae)

The genus Hemicycliophora (Nematoda: Hemicycliophoridae) contains 132 valid species of plant-parasitic nematodes, collectively known as ‘sheath nematodes’. Hemicycliophora spp. are characterized morphologically by a long stylet with rounded basal knobs and a cuticular sheath, present in juvenile and adult stages. Populations of 20 valid and 14 putative species of Hemicycliophora and Loofia from several countries were characterized morphologically using light (LM) and scanning electron microscopy (SEM) and molecularly using the D2-D3 segments of 28S rRNA and internal transcribed spacer (ITS) rRNA gene sequences. LM and SEM observations provided new details on the morphology of these species. PCR-restriction fragment length polymorphisms (PCR-RFLPs) of the D2-D3 of 28S rDNA were proposed for identification of the species. Phylogenetic relationships within populations of 36 species of the genus Hemicycliophora using 102 D2-D3 of 28S rDNA and 97 ITS rRNA gene sequences as inferred from Bayesian analysis are reconstructed and discussed. Ancestral state reconstructions of diagnostic characters (body and stylet length, number of body annuli, shape of vulval lip and tail), using maximum parsimony and Bayesian inference, revealed that none of the traits are individually reliable characters for classifying the studied sheath nematode. The Shimodaira–Hasegawa test rejected the validity of the genus Loofia. This is the most complete phylogenetic analysis of Hemicycliophora species conducted so far.Fil: Subbotin, Sergei A.. California Department of Food and Agriculture; Estados Unidos. Institute of Ecology and Evolution of the Russian Academy of Sciences; RusiaFil: Chitambar, John J.. California Department of Food and Agriculture; Estados UnidosFil: Chizhov, Vladimir N.. Institute of Ecology and Evolution of the Russian Academy of Sciences; RusiaFil: Stanley, Jason D.. Florida Department of Agriculture and Consumer Services; Estados UnidosFil: Inserra, Renato N.. Florida Department of Agriculture and Consumer Services; Estados UnidosFil: Doucet, Marcelo Edmundo. Universidad Nacional de Cordoba. Facultad de Cs.exactas Fisicas y Naturales. Centro de Zoologia Aplicada; Argentina. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Cordoba. Instituto de Diversidad y Ecologia Animal; ArgentinaFil: Mcclure, Michael. University Of Arizona; Estados UnidosFil: Ye, Weimin. North Carolina Department of Agriculture & Consumer Services; Estados UnidosFil: Yeates, George.Fil: Mollov, Dimitre S.. University Of Minnesota; Estados UnidosFil: Cantalapiedra Navarrete, Carolina. Consejo Superior de Investigaciones Científicas. Instituto de Agricultura Sostenible; EspañaFil: Vovlas, Nicola. Istituto per la Protezione delle Piante; ItaliaFil: Van Den Berg, Esther. ARC-Plant Protection Research Institute; SudáfricaFil: Castillo, Pablo. Consejo Superior de Investigaciones Científicas. Instituto de Agricultura Sostenible; Españ

Alfalfa virus S, a new species in the family Alphaflexiviridae.

A new species of the family Alphaflexiviridae provisionally named alfalfa virus S (AVS) was discovered in alfalfa samples originating from Sudan. A complete nucleotide sequence of the viral genome consisting of 8,349 nucleotides excluding the 3' poly(A) tail was determined by high throughput sequencing (HTS) on an Illumina platform. NCBI BLAST searches revealed that the virus shares the greatest degree of sequence identity with members of the family Alphaflexiviridae, genus Allexivirus. The AVS genome contains six computationally-predicted open reading frames (ORF) encoding viral replication protein, triple gene block protein 1 (TGB1), TGB2, TGB3-like protein, unknown 38.4 kDa protein resembling serine-rich 40 kDa protein characteristic for allexiviruses, and coat protein (CP). AVS lacks a clear 3' proximal ORF that encodes a nucleic acid-binding protein typical for allexiviruses. The identity of the virus was confirmed by RT-PCR with primers derived from the HTS-generated sequence, dot blot hybridization with DIG-labeled virus-specific RNA probes, and Western blot analysis with antibodies produced against a peptide derived from the CP sequence. Transmission electron microscopic observations of the infected tissues showed the presence of filamentous particles similar to allexiviruses in their length and appearance. To the best of our knowledge, this is the first report on the identification of a putative allexivirus in alfalfa (Medicago sativa). The genome sequence of AVS has been deposited in NCBI GenBank on 03/02/2016 as accession № KY696659

Genome characterization and complete sequence of a new badnavirus from Pandanus amaryllifolius

International audienceA new badnavirus was sequenced from fragrant pandan grass (Pandanus amaryllifolius) displaying mosaic and chlorosis on the leaves. The complete genome sequence was determined by high-throughput sequencing. The new badnavirus was tentatively named "pandanus mosaic associated virus" (PMaV). Similar to those of other members of the genus Badnavirus, the genome of PMaV consists of a circular DNA molecule of 7,481 bp with three open reading frames (ORF) potentially coding for three proteins. ORF3 encodes a polyprotein with conserved protein domains including zinc finger, trimeric dUT-Pase, aspartic protease, reverse transcriptase (RT), and RNase H domains. Pairwise comparisons of the highly conserved RT + RNase H region revealed the highest nucleotide (nt) sequence identity (70.71%) to taro bacilliform CH virus-Et17 (MG017324). In addition to PMaV, viral sequences corresponding to orchid fleck dichorhavirus (OFV) were detected in the same plant sample. The complete sequence of the OFV coding region shared >98% nt sequence identity with other isolates of OFV available in the GenBank database. Disease symptoms could not be attributed exclusively to PMaV or OFV, as both viruses were present in the pandan grass exhibiting mosaic and chlorosis

Phylogenetic relationship between alfalfa virus S (highlighted), classified allexiviruses and unassigned members of the family <i>Alphaflexiviridae</i>.

<p>The tree was built based on the available complete nucleotide sequences using MEGA 7 software (version 7.0.21) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0178222#pone.0178222.ref025" target="_blank">25</a>] and the Neighbor-Joining method. The optimal tree with the sum of branch length = 3.10976014 is shown. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (1000 replicates) are shown next to the branches.</p