Geographical gradient of the <em>eIF4E</em> alleles conferring resistance to potyviruses in pea (<em>Pisum</em>) germplasm

<div><p>Background</p><p>The eukaryotic translation initiation factor 4E was shown to be involved in resistance against several potyviruses in plants, including pea. We combined our knowledge of pea germplasm diversity with that of the <i>eIF4E</i> gene to identify novel genetic diversity.</p><p>Methodology/Principal findings</p><p>Germplasm of 2803 pea accessions was screened for <i>eIF4E</i> intron 3 length polymorphism, resulting in the detection of four <i>eIF4E^A-B-C-S</i> variants, whose distribution was geographically structured. The <i>eIF4E^A</i> variant conferring resistance to the P1 PSbMV pathotype was found in 53 accessions (1.9%), of which 15 were landraces from India, Afghanistan, Nepal, and 7 were from Ethiopia. A newly discovered variant, <i>eIF4E^B</i>, was present in 328 accessions (11.7%) from Ethiopia (29%), Afghanistan (23%), India (20%), Israel (25%) and China (39%). The <i>eIF4E^C</i> variant was detected in 91 accessions (3.2% of total) from India (20%), Afghanistan (33%), the Iberian Peninsula (22%) and the Balkans (9.3%). The <i>eIF4E^S</i> variant for susceptibility predominated as the wild type. Sequencing of 73 samples, identified 34 alleles at the whole gene, 26 at cDNA and 19 protein variants, respectively. Fifteen alleles were virologically tested and 9 alleles (<i>eIF4E^{A-1-2-3-4-5-6-7}</i>, <i>eIF4E^B-1</i>, <i>eIF4E^C-2</i>) conferred resistance to the P1 PSbMV pathotype.</p><p>Conclusions/Significance</p><p>This work identified novel <i>eIF4E</i> alleles within geographically structured pea germplasm and indicated their independent evolution from the susceptible <i>eIF4E^S1</i> allele. Despite high variation present in wild <i>Pisum</i> accessions, none of them possessed resistance alleles, supporting a hypothesis of distinct mode of evolution of resistance in wild as opposed to crop species. The Highlands of Central Asia, the northern regions of the Indian subcontinent, Eastern Africa and China were identified as important centers of pea diversity that correspond with the diversity of the pathogen. The series of alleles identified in this study provides the basis to study the co-evolution of potyviruses and the pea host.</p></div

Molecular study of turnip mosaic virus population in the Czech Republic

Turnip mosaic virus (TuMV) is the most important virus of brassica crops. In our study, we compare the genetic structure of two Czech TuMV populations sampled in the country's 25-year interval of virus presence. The 21 isolates, mainly infecting rutabaga and horseradish, were collected from four farms under organic production, and nearly complete genome sequences, 9 596-9 787 nt in length, were obtained using Sanger sequencing for all of them. The analysis of variability and polymorphism showed differences in genetic structure but the relative stability of both populations and moderate negative selection as a factor affecting the current TuMV population. The newly collected isolates are characterised by a relatively high frequency of intralineage recombinants; interlineage recombinants were not detected compared to the 25-year-old population. The phylogenetic analysis allowed the classification of all Czech isolates into world-B phylogroup, with the prevalence of isolates of subgroup B2. The spread of isolates belonging to the other phylogenetic groups posing higher phytopathological risk, which were present in the old population and some surrounding countries, was not found

Complete genome sequence of a novel cytorhabdovirus infecting elderberry (Sambucus nigra L.) in the Czech Republic

The genus Cytorhabdovirus includes plant viruses with an unsegmented, single-stranded, negative-sense RNA genome that infect various plant hosts. In this work, we report the detection of a new cytorhabdovirus infecting elderberry (Sambucus nigra L.). Total RNA was purified from infected leaves and, after ribodepletion, sequenced using an Illumina system. The RNA genome of viral isolate B15 is 12,622 nucleotides (nt) long, and that of isolate B42 is 12,621 nt long. A nearly complete sequence (12,592 nt) was also obtained for a third isolate (B160). The RNA genomes of all three isolates showed an organisation typical of cytorhabdoviruses, harbouring all six of the expected genes (3´ N-P-P3-M-G-L 5´), separated by intergenic regions. These isolates were closely related to each other (99.5-99.6% nt sequence identity) and showed the highest overall similarity to trichosanthes associated rhabdovirus 1 (63.5% identity) and Wuhan insect virus 5 (58% identity), and similar results were obtained when comparing individual coding sequences or proteins. Phylogenetic analysis confirmed that this elderberry virus, for which we propose the name "sambucus virus 1" belongs to the genus Cytorhabdovirus and fulfils the criteria to represent a novel species

Molecular Characterization of ‘<i>Candidatus</i> Phytoplasma prunorum’ in the Czech Republic and Susceptibility of Apricot Rootstocks to the Two Most Abundant Haplotypes

‘Candidatus Phytoplasma prunorum’ is one of the most destructive pathogens of Prunus species, where susceptible species render unproductive several years after infection. In epidemiology, the molecular characterization of phytoplasmas is based on sequence analysis of variable nonribosomal genes. In this study aceF, pnp, imp and secY genes were used for characterization of the ‘Ca. P. prunorum’ genotypes present in the Czech Republic. In total, 56 plant and 33 vector (Cacopsylla pruni) samples positive to ‘Ca. P. prunorum’ collected in seven localities were used in the study. Based on sequence analysis, four aceF, two pnp, six imp, and three secY genotypes were identified in analyzed samples. The most abundant in both plant and insect samples were the A6, P2, I4, and S2 genotypes. Most of the Czech ‘Ca. P. prunorum’ haplotypes clustered together in the haplotype network analysis. Next, two isolates representing the two most abundant Czech haplotypes (A6-P2-I4-S2 and A5-P2-I4-S2) were used in the susceptibility test of three apricot rootstock types (St. Julien A, M-VA-1, GF-305). Susceptibility was analyzed by phytoplasma quantification using quantitative real-time PCR and evaluation of symptom manifestation. Based on the results, the influence of the rootstock type on the phytoplasma titer and symptom manifestation was greater than of the phytoplasma isolate, while the year of analysis had no influence on the results. The results also showed that the phytoplasma titer is increasing in plant tissues during the vegetation period

Molecular Characterization of Mitogenome of <i>Cacopsylla picta</i> and <i>Cacopsylla melanoneura</i>, Two Vector Species of ‘<i>Candidatus</i> Phytoplasma mali’

The mitochondrial genomes of two vector psyllids of the ‘Candidatus Phytoplasma mali’, Cacopsylla picta and C. melanoneura, were sequenced using high-throughput sequencing on the Illumina platform. The main objective of the study was to describe their mitogenome and characterize their genetic variability and the potential changes in the context of the observed global warming. The four complete sequences for C. picta, 14,801 bp and 14,802 bp in length, two complete and one partial sequence for C. melanoneura, ranging from 14,879 bp to 14,881 bp in length, were obtained for the first time for these European apple psyllids. The detected intraspecies mtDNA identity was highly similar (99.85–99.98%), the identity’s similarity with other Cacopsylla species varied between 79.79 and 86.64%. The mitogenomes showed a typical mitochondrial DNA structure with 13 protein-coding genes, 2 rRNA genes and 22 tRNA genes; the presence of CGGA motif in the ND1-trnS2 junction was detected in both species. Phylogenetic analysis placed both species in close relationship with C. burckhardti within the Cacopsylla clade-I O group. The analysis of complete mitogenomes and of partial COI sequences of fifty-two Cacopsylla individuals showed a high homogeneity of genotypes over 15 years and among the different localities in the Czech Republic

Allele designation based on intron 3 length polymorphism, indicating size of obtained fragments with primer combinations A and B, number of repeat motives, intron 3 difference and total length.

<p>Allele designation based on intron 3 length polymorphism, indicating size of obtained fragments with primer combinations A and B, number of repeat motives, intron 3 difference and total length.</p

Geographical distribution of four <i>eIF4E</i> alleles expressed as percentage of total.

<p>Comparison of allele frequencies in 13 geographical regions, as detailed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0090394#pone-0090394-t002" target="_blank"><b>Table 2</b></a>.</p

Summarized amino acid exchanges of identified 19 <i>eIF4E</i> alleles.

<p>Asterix indicates the position of exchanges previously identified as crucial for resistance.</p

List of sequenced 73 accessions assigned to identified 19 protein alleles of four <i>eIF4E ^A-B-C-S</i> intron 3 length variants with indicated country of origin.

<p>List of sequenced 73 accessions assigned to identified 19 protein alleles of four <i>eIF4E ^A-B-C-S</i> intron 3 length variants with indicated country of origin.</p