Construction of a high-density genetic map and detection of a major QTL of resistance to powdery mildew (Erysiphe necator Sch.) in Caucasian grapes (Vitis vinifera L.)

Background Vitis vinifera L. is the most cultivated grapevine species worldwide. Erysiphe necator Sch., the causal agent of grape powdery mildew, is one of the main pathogens affecting viticulture. V. vinifera has little or no genetic resistances against E. necator and the grape industry is highly dependent on agrochemicals. Some Caucasian V. vinifera accessions have been reported to be resistant to E. necator and to have no genetic relationships to known sources of resistance to powdery mildew. The main purpose of this work was the study and mapping of the resistance to E. necator in the Caucasian grapes ‘Shavtsitska’ and ‘Tskhvedianis tetra’. Results The Caucasian varieties ‘Shavtsitska’ and ‘Tskhvedianis tetra’ showed a strong partial resistance to E. necator which segregated in two cross populations: the resistant genotypes delayed and limited the pathogen mycelium growth, sporulation intensity and number of conidia generated. A total of 184 seedlings of ‘Shavtsitska’ x ‘Glera’ population were genotyped through the Genotyping by Sequencing (GBS) technology and two high-density linkage maps were developed for the cross parents. The QTL analysis revealed a major resistance locus, explaining up to 80.15% of the phenotypic variance, on ‘Shavtsitska’ linkage group 13, which was associated with a reduced pathogen infection as well as an enhanced plant necrotic response. The genotyping of 105 Caucasian accessions with SSR markers flanking the QTL revealed that the resistant haplotype of ‘Shavtsitska’ was shared by ‘Tskhvedianis tetra’ and a total of 25 Caucasian grape varieties, suggesting a widespread presence of this resistance in the surveyed germplasm. The uncovered QTL was mapped in the region where the Ren1 locus of resistance to E. necator, identified in the V. vinifera ‘Kishmish vatkana’ and related grapes of Central Asia, is located. The genetic analysis conducted revealed that the Caucasian grapes in this study exhibit a resistant haplotype different from that of Central Asian grape accessions. Conclusions The QTL isolated in ‘Shavtsitska’ and present in the Caucasian V. vinifera varieties could be a new candidate gene of resistance to E. necator to use in breeding programmes. It co-localizes with the Ren1 locus but shows a different haplotype from that of grapevines of Central Asia. We therefore consider that the Caucasian resistance locus, named Ren1.2, contains a member of a cluster of R-genes, of which the region is rich, and to be linked with, or possibly allelic, to Ren1.

Variability of biological traits of genotypes in the ‘Talisman x Asma’ crossing population

Целью исследований было изучение биологических особенностей новых гибридных форм винограда и степень их изменчивости. Гибридные формы столового винограда получены путем гибридизации обработанной колхицином материнской формы Талисман. Скрещивания проведены в 2011 г. в направлении создания крупноягодных столовых сорто-форм среднепозднего и позднего срока созревания, предназначенных для длительного хранения. Установлено что к 2020 г. всего 77 % растений в популяции сеянцев вступили в пору плодоношения, из них у 21 сеянца определен обоеполый тип цветка и у 30 растений тип цветка определен как функционально женский. Продукционный период в популяции сеянцев варьирует от 119-150 дней; наибольшую группу составляют сеянцы со средним сроком созревания (135-145 дней) - 65 % растений. Коэффициент плодоношения в популяции сеянцев находится в пределах 0,49±0,09, а коэффициент плодоносности - 1,17±0,09. При этом коэффициент плодоношения варьирует в значительно большей степени, чем коэффициент плодоносности. Отмечено низкое среднее значение полученного фактического урожая в популяции сеянцев относительно сортов-родительских форм; однако данные вариационного анализа указывают на присутствие в данной популяции растений, характеризующиеся как очень низкой урожайностью, так и высокой (V = 67-69 %). В популяции сеянцев на естественном инфекционном фоне выделены генотипы с высокой устойчивостью к оидиуму (Erysiphe necator) - более 20%. На основе статистического анализа комплекса признаков выделена группа из 29 сеянцев для дальнейшего изучения. Предварительно выделены в качестве элитных форм Магарач №19-11-3-20 (для длительного хранения) и Магарач № 19-11-3-53, М.№19-11-4-17. The aim of the research was to study the biological characteristics of new hybrid forms of grapes and the degree of their variability. Hybrid forms of table grapes are obtained by hybridization of the treated with colchicine female parent of ‘Talisman’. Crossings were carried out in 2011 in the direction of creating large-berry table varieties of medium-late and late ripening, intended for long-term storage. It was established that by 2020 only 77% of the plants in the seedling population entered the season of fruiting, 21 seedlings of which had a bisexual flower type and 30 plants had a flower type defined as functionally female. The production period in the population of seedlings varies from 119 to 150 days; the largest group consists of seedlings with an average ripening period (135-145 days) - 65% of plants. The coefficient of fruiting in the population of seedlings is within the range of 0.49±0.09, and the coefficient of fertility is 1.17±0.09. At the same time, the fruiting coefficient varies to a much greater extent than the fertility coefficient. There was a low average value of actual yield obtained in the population of seedlings relative to the varieties - parental forms; however, the data of analysis of variance indicate the presence of plants in this population, characterized by both very low and high cropping capacity (V=67-69%). In the population of seedlings against a natural infectious background, genotypes with high resistance to powdery mildew ( Erysiphe necator ) - more than 20% were identified. On the basis of statistical analysis of the complex of characters, a group of 29 seedlings was selected for further study. The ‘Magarach No. 19-11-3-20’ (for long-term storage) and ‘Magarach No. 19-11-3-53’, ‘M. No. 19-11-4-17’ were preliminarily identified as elite forms.

Genome Sequence Resource for Erysiphe necator NAFU1, a Grapevine Powdery Mildew Isolate Identified in Shaanxi Province of China

Erysiphe necator is an economically important biotrophic fungal pathogen responsible for powdery mildew disease on grapevine. Currently, genome sequences are available for only a few Erysiphe necator isolates from USA. Based on the combination of Nanopore and Illumina sequencing technologies, we present here the complete genome assembly for an isolate of E. necator NAFU1 identified in China. We acquired a total of 15.93 Gb raw reads. These reads were processed into a 61.12 Mb genome assembly containing 73 contigs with the N50 of 2.06 Mb and a maximum length of 6.05 Mb. Combining the results of three gene-prediction modules, i.e. an evidence-based gene modeler (EVidenceModeler or EVM), an ab initio gene modeler, and a homology-based gene modeler, we predicted 7235 protein-coding genes in the assembled genome of E. necator NAFU1. This information will facilitate studies of genome evolution and pathogenicity mechanisms of E. necator and other powdery mildew species through comparative genome sequence analysis and other molecular genetic tools.

Complete Genome Sequence of a Novel Umbra-like Mycovirus From the Plant Pathogenic Fungus Phoma Matteucciicola

Here, a novel umbra-like mycovirus, ‘Phoma matteucciicola RNA virus 2’ (PmRV2), isolated from Phoma matteucciicola strain HNQH1 in Hainan province of China, was sequenced and analyzed. The complete genomic sequence of PmRV2 is 3,460 nucleotides (nts) with a GC content of 56.71%. Sequence analysis of PmRV2 indicated that the presence of two noncontiguous open reading frames (ORFs) encoding a hypothetical protein and a RNA-dependent RNA polymerase (RdRp), respectively. PmRV2 contains a metal-binding ‘GDN’ triad in Motif C of RdRp while most + ssRNA mycoviruses contained a ‘GDD’ motif in the same region. Additionally, a BLASTp search showed that the RdRp amino acid sequence of PmRV2 was most closely related to the RdRp of Macrophomina phaseolina umbra-like virus 1 (50.72% identity) and Erysiphe necator umbra-like virus 2 (44.84% identity). Phylogenetic analysis indicated that PmRV2 grouped together with Erysiphe necator umbra-like virus 2 (EnUlV2) within the recently proposed family of ‘Mycotombusviridae’.

The mitochondrial genome of the grape powdery mildew pathogen Erysiphe necator is intron rich and exhibits a distinct gene organization

Powdery mildews are notorious fungal plant pathogens but only limited information exists on their genomes. Here we present the mitochondrial genome of the grape powdery mildew fungus Erysiphe necator and a high-quality mitochondrial gene annotation generated through cloning and Sanger sequencing of full-length cDNA clones. The E. necator mitochondrial genome consists of a circular DNA sequence of 188,577 bp that harbors a core set of 14 protein-coding genes that are typically present in fungal mitochondrial genomes, along with genes encoding the small and large ribosomal subunits, a ribosomal protein S3, and 25 mitochondrial-encoded transfer RNAs (mt-tRNAs). Interestingly, it also exhibits a distinct gene organization with atypical bicistronic-like expression of the nad4L/nad5 and atp6/nad3 gene pairs, and contains a large number of 70 introns, making it one of the richest in introns mitochondrial genomes among fungi. Sixty-four intronic ORFs were also found, most of which encoded homing endonucleases of the LAGLIDADG or GIY-YIG families. Further comparative analysis of five E. necator isolates revealed 203 polymorphic sites, but only five were located within exons of the core mitochondrial genes. These results provide insights into the organization of mitochondrial genomes of powdery mildews and represent valuable resources for population genetic and evolutionary studies.

Detection of Erysiphe necator, the Causal Agent of Powdery Mildew on Grapevine, and Determination of their Mating Types in Southern Syria Using Some Molecular Markers

Powdery mildew caused by Erysiphe necator, is one of the most economically damaging diseases of grapevine throughout the world. This study aimed to identify the mating types of this pathogen in Syria using molecular markers. Three DNA extraction methods: Promega Kit, phenol chloroform isoamyl alcohol (PCI) and SDS were compared. The amount and quality of DNAs obtained by the SDS method were suitable for PCR amplification and other molecular assays. PCR amplification using specific primers (Uncin 144 and Uncin 511) was performed and the expected amplicon of 300-400 bp was obtained from 29 isolates of E. necator collected from different geographical locations and from different grapevine cultivars. Results obtained showed that E. necator may cause atypical symptoms similar to those of downy mildew, in some grapevine cultivars according to the environmental conditions and training system used. Isolates of E. necator were classified into four groups according to geographical locations and grapevine cultivar, based on the observed variation in banding pattern with E07 primer and the constructed phenogram by using UPGMA. RAPD analysis of 39 E. necator isolates using the primer E07 showed the presence of two mating types, and the frequency of each varied depending on the geographical location, with a predominance of the - mating type which did not produce a 1000 bp band using the PCR primer E07. Keywords: Powdery mildew, Erysiphe necator, RAPD, mating type, DNA extraction.

Construction of A High-Density Genetic Map And Detection of A Major QTL of Resistance To Powdery Mildew (Erysiphe Necator Sch.) In Caucasian Grapes (Vitis Vinifera L.)

BackgroundVitis vinifera L. is the most cultivated grapevine species worldwide. Erysiphe necator Sch., the causal agent of grape powdery mildew, is one of the main pathogens affecting viticulture. V. vinifera has little or no genetic resistance against E. necator and the grape industry is highly dependent on agrochemicals. Some Caucasian V. vinifera accessions have been reported to be resistant to E. necator and to have no genetic relationship to known sources of resistance to powdery mildew. The main purpose of this work was the study and mapping of the resistance to E. necator in the Caucasian grapes ‘Shavtsitska’ and ‘Tskhvedianis tetra’.ResultsThe Caucasian varieties ‘Shavtsitska’ and ‘Tskhvedianis tetra’ showed a strong partial resistance to E. necator which segregated in two cross populations: the resistant genotypes delayed and limited the pathogen mycelium growth, sporulation intensity and number of conidia generated. A total of 184 seedlings of ‘Shavtsitska’ x ‘Glera’ population were genotyped through the Genotyping by Sequencing (GBS) technology and two high-density linkage maps were developed for the cross parents. The QTL analysis revealed a major resistance locus, explaining up to 80.15% of the phenotypic variance, on ‘Shavtsitska’ linkage group 13, which was associated with a reduced pathogen infection as well as an enhanced plant necrotic response. The genotyping of 105 Caucasian accessions with SSR markers flanking the QTL revealed that the resistant haplotype of ‘Shavtsitska’ was shared by ‘Tskhvedianis tetra’ and a total of 24 Caucasian grape varieties, suggesting a widespread presence of this resistance in the surveyed germplasm. The uncovered QTL was mapped in the region where the Ren1 gene of resistance to E. necator, identified in the V. vinifera ‘Kishmish vatkana’ and related grapes of Central Asia, is located. The genetic analysis conducted revealed that the Caucasian grapes in this study exhibit a resistant haplotype different from that of Central Asian grape accessions.ConclusionsThe QTL isolated in ‘Shavtsitska’ and present in the Caucasian V. vinifera varieties could be a new candidate gene of resistance to E. necator to use in breeding programmes. It co-localizes with the Ren1 locus but shows a different haplotype from that of grapevines of Central Asia. We therefore consider this gene, named Ren1-2, a member of a cluster of R genes, of which the region is rich, and linked with, or possibly allelic, to the Ren1.

Flashes of UV-C light stimulate defences of Vitis vinifera L. cv. Chardonnay against Erysiphe necator in greenhouse and vineyard conditions

Using detached leaves, UV-C light in the form of 1-sec flashes has recently been shown to stimulate defences of several plants against different pathogenes better than 1 min exposures under greenhouse conditions. In the present work, the pathological tests were conducted using undetached leaves under greenhouse and vineyard conditions. In a first trial, two flashes of UV-C light were applied to plants of Vitis vinifera L. cv. Chardonnay grown under greenhouse conditions, at an interval of 10 days. Plants were inoculated with Erysiphe necator two days after the last light treatment. After 18 days of inoculation, the symptom severity on leaves was reduced by 60 % when compared with the untreated control. In a second trial, flashes of UV-C light were applied to grapevine Chardonnay plants under field conditions in the South-East of France, every 10 days from the 18th of April until the 10th of July 2019. The symptom severity resulting from natural contaminations by Erysiphe necator was reduced by 42 % in leaves on the 4th of July 2019 and by 65 % in clusters on the 25th of July 2019. In a third trial, we observed that UV-C light did not have any effect on net photosynthesis, maximal net photosynthesis, dark respiration, maximal quantum efficiency of photosystem II, the performance index of Strasser and, generally, any parameter derived from induction curves of maximal chlorophyll fluorescence. It was concluded that flashes of UV-C light have true potential for stimulating plant defences against Erysiphe necator under vineyard conditions and, therefore, help in reducing fungicide use.