High-density linkage mapping and QTL identification of black rot resistance towards marker-assisted breeding in grapevine

Today sustainability is a pivotal objective for viticulture, but it also presents us with new challenges. The decrease in treatments has in fact changed grapevine-pathogen interactions and dynamics causing the reaffirmation of diseases previously considered secondary. This is the case of black rot (BR, caused by Phyllosticta ampelicida), which is spreading with increasing pressure in warm-humid regions causing the loss of entire crops. The interinstitutional DAMAGE project between Edmund Mach Foundation (FEM) and the Institute for Grapevine Breeding (JKI-Geilweilerhof) aims to characterize BR resistance and to develop a toolkit of molecular markers (MM) to be routinely used in marker-assisted breeding for the introgression of this trait into mildew resistant backgrounds. The first fungal strain was isolated from infected leaves collected in Trentino-Italy and mixed with a second strain isolated in southwestern Germany. This inoculum has been used to screen a segregating population derived from 'Merzling' (V. rupestris × V. lincecumii, mid-resistant) × 'Teroldego' (V. vinifera, susceptible). BR resistance phenotyping consisted of a newly developed inoculation protocol on potted plants. The F1 individuals were genotyped with the Vitis18KSNP chip and a high-density genetic map has been constructed, following the integration of 190 informative SSRs. Moreover, explorative QTL analyzes have been conducted on phenotypic data of 2020 and 2021 seasons. Finally, upon the genomic interval characterization, new MM will be designed, tested and validated on various segregating populations with different genetic backgrounds

Grape production, technological parameters, and stilbenic compounds as affected by lime-induced chlorosis

Vitis vinifera L. cv. Merlot clone R3, grafted on 3309 C (lime-susceptible) rootstock, was grown in pots on a noncalcareous and a calcareous soil. The aim of the experiment was to check the effect of lime stress conditions on chlorosis, grape yield, technological parameters and stilbene (resveratrol, piceid, piceatannol, ε-viniferin) concentrations in grapes. Lime-induced chlorosis decreased grape yield per plant to a very high extent, as a result of a reduction of cluster and berry size. Technological grape parameters such as soluble solids, pH, anthocyanins, increased under lime stress over the control, whilst titratable acidity was not affected. All the tested stilbenes, being stress compounds, increased in the grapes of chlorotic vines.

Towards Marker-Assisted Breeding for Black Rot Bunch Resistance: Identification of a Major QTL in the Grapevine Cultivar 'Merzling'

Black rot (BR), caused by Guignardia bidwellii, is an emergent fungal disease threatening viticulture and affecting several mildew-tolerant varieties. However, its genetic bases are not fully dissected yet. For this purpose, a segregating population derived from the cross 'Merzling' (hybrid, resistant) × 'Teroldego' (V. vinifera, susceptible) was evaluated for BR resistance at the shoot and bunch level. The progeny was genotyped with the GrapeReSeq Illumina 20K SNPchip, and 7175 SNPs were combined with 194 SSRs to generate a high-density linkage map of 1677 cM. The QTL analysis based on shoot trials confirmed the previously identified Resistance to Guignardia bidwellii (Rgb)1 locus on chromosome 14, which explained up to 29.2% of the phenotypic variance, reducing the genomic interval from 2.4 to 0.7 Mb. Upstream of Rgb1, this study revealed a new QTL explaining up to 79.9% of the variance for bunch resistance, designated Rgb3. The physical region encompassing the two QTLs does not underlie annotated resistance (R)-genes. The Rgb1 locus resulted enriched in genes belonging to phloem dynamics and mitochondrial proton transfer, while Rgb3 presented a cluster of pathogenesis-related Germin-like protein genes, promoters of the programmed cell death. These outcomes suggest a strong involvement of mitochondrial oxidative burst and phloem occlusion in BR resistance mechanisms and provide new molecular tools for grapevine marker-assisted breeding

Mining grapevine downy mildew susceptibility genes: A resource for genomics-based breeding and tailored gene editing

Several pathogens continuously threaten viticulture worldwide. Until now, the investigation on resistance loci has been the main trend to understand the interaction between grapevine and the mildew causal agents. Dominantly inherited gene-based resistance has shown to be race-specific in some cases, to confer partial immunity, and to be potentially overcome within a few years since its introgression. Recently, on the footprint of research conducted in Arabidopsis, putative genes associated with downy mildew susceptibility have been discovered also in the grapevine genome. In this work, we deep-sequenced four putative susceptibility genes\u2014namely VvDMR6.1, VvDMR6.2, VvDLO1, VvDLO2\u2014in 190 genetically diverse grapevine genotypes to discover new sources of broad-spectrum and recessively inherited resistance. Identified Single Nucleotide Polymorphisms were screened in a bottleneck analysis from the genetic sequence to their impact on protein structure. Fifty-five genotypes showed at least one impacting mutation in one or more of the scouted genes. Haplotypes were inferred for each gene and two of them at the VvDMR6.2 gene were found significantly more represented in downy mildew resistant genotypes. The current results provide a resource for grapevine and plant genetics and could corroborate genomic-assisted breeding programs as well as tailored gene editing approaches for resistance to biotic stresses

Temporal and spatial change of exergy and ascendency in different benthic marine ecosystems

Holistic indicators such as exergy and ascendency have been widely employed to assess the health of ecosystems given by their structure, function and organization. In this study we calculate the exergy, specific exergy and ascendency for the microbenthic loop that represents a major sub-system within the marine food chain. The analysis of the microbenthic loop investigated in terms of organic matter, bacteria, microphytobenthos and meiofauna reflected changes occurring in the trophic state of benthic ecosystems and provided a tool for comparison between different environments. Temporal and spatial variability of the holistic indicators were evaluated using benthic measures collected at different times for different environments in the Mediterranean Sea. Exergy was strongly correlated with the organic contents of the sediments, and did not provide a useful description of the investigated system. In contrast, specific exergy resulted related to the microbenthic loop structure and complexity while Ascendency mostly reflected its activity and organization. Temporal analysis showed that in natural ecosystems specific exergy and ascendency showed convergence and follow similar seasonal trends. On the contrary in strongly eutrophicated systems an uncoupling between the two indicators occurred indicating a malfunctioning of the microbenthic loop that become strongly dissipative.http://www.sciencedirect.com/science/article/B6V2S-4CC7RP3-D/1/49cf1c6b40bddddc9a3bb98c5072e51

Rgb3: an organ-specific QTL for bunch resistance to black rot identified in the hybrid cultivar ‘Merzling’ Hands-on demo of the potential of Grapedia

Crops are continuously exposed to the onset of emerging diseases and the exploitation of resistance donors in breeding programs is a widely used valid strategy. In the last two decades black rot (BR), caused by the ascomycete Phyllosticta ampelicida (syn. Guignardia bidwellii), has begun to threaten European viticulture in humid continental areas due to the advent of higher temperatures which favor the pathogenic cycle of the fungus. For this reason, a program has been established for the dissection of BR resistance trait and its introgression both in Vitis vinifera varieties and mildew resistant hybrids. A preliminary study was carried out to improve germplasm screening through different approaches. Historical phenotypic and pedigree information of resistance donors were gathered to support the decision-making process in breeding. A new propagation and inoculation strategy was developed to optimize and fasten inoculation experiments. Isolates of P. ampelicida were genetically characterized and combined for the assessment of resistance independent of race-specificity. Finally, these improvements were employed first for the identification of new BR resistant parental lines and breeding selections, and then for the quantitative trait locus (QTL) analysis in a segregating population derived from the cross ‘Merzling’ (hybrid, resistant) × ‘Teroldego’ (V. vinifera, susceptible). The screening of this progeny, under green-house and field conditions, allowed the discrimination between two distinct organ-specific QTLs on chromosome 14. The previously identified Resistance to G. bidwellii (Rgb)1 locus was confirmed associated with leaf/shoot resistance, while upstream a new QTL designated Rgb3 was discovered linked to bunch resistance. Driven and inspired by the INTEGRAPE community effort that led to GRAPEDIA (GRAPEvine -omics encyclopDIA), all the available tools and resources have been exploited for the exploration of the physical region of the two QTLs, providing a hands-on demo of the potential of this portal

Highly dense linkage mapping and identification of a major QTL associated with BLACK ROT resistance in the grapevine cultivar ‘Merzling’

When the urgency of a sustainable transition comes to agriculture, one of the key areas of improvement involves the decrease of chemicals. Although vineyards cover less than 5% of agricultural land in Europe, viticulture is responsible for the use of more than 60% of all fungicides. The exploitation of grapevine varieties resistant to mildews is an efficient strategy already implemented in integrated/organic farming to reduce treatments. However, from the beginning of this century, European viticulture has been threatened by severe outbreaks of black rot (BR), an emergent and destructive disease caused by the ascomycete Phyllosticta ampelicida (sexual morph Guignardia bidwellii). These events introduced the urgent need for the introgression of BR resistance in mildew-tolerant genotypes. For this purpose, a set of parental lines and breeding selections of the Fondazione Edmund Mach has been screened for BR resistance in a growing chamber with in vivo produced spores using an optimized artificial infection protocol. Given the good performance of ‘Merzling’ (a complex genotype derived from V. vinifera and V. rupestris × V. aestivalis var. lincecumii), this cultivar was used for a cross with the susceptible variety ‘Teroldego’ (V. vinifera) and the segregating offspring was genetically characterized by means of the GrapeReSeq 18K Vitis SNP chip. Five phenotypic experiments were carried out under controlled conditions on leaves of potted plants, and three on bunches in the field. A dense genetic map was constructed combining 7,175 SNP with 194 SSR markers of a previous map. All QTL analyses revealed the presence of a strong major BR resistance locus on chromosome 14. It explains up to 45% of the trait variability (LOD 10.5) and spans a genomic region of 1.36 Mb. A specific SNP marker was found robustly associated with the resistance trait. No minor QTLs were detected. The genes underlying this region are currently under investigation via bioinformatic analysis, and microscopic inspections of disease progression are in place to understand the biological causes of the resistance trait. Finally, new molecular markers will be developed and validated on segregating populations with different genetic backgrounds, to be implemented in marker-assisted selection for BR resistance in grapevin

Genetics-assisted breeding for downy/powdery mildew and phylloxera resistance at fem

The genetics-assisted program for resistance to biotic stresses began at the Edmund Mach Foundation in 2007 and has developed on two lines. The first line was based on obtaining resistant materials with parents the historical varieties of Trentino. During the three-year period 2015–2017, 3 Teroldego X Merzling genotypes, 3 Marzemino X Merzling genotypes, 1 Nosiola X Bianca genotype and 1 Nosiola X Kulneany genotype were collected the data related to resistance to various fungal diseases both on the leaf and on the bunch. At harvest the plant production data and the must characteristics were recorded; the grapes were vinified and for each year the wines were subjected to sensorial analysis. 5 resistant selections were considered interesting for all the characteristics found and therefore the data for the registration to the National Register of Grapevine Varieties will be collected. The second line of research was based on the retrieval and both genotypic and phenotypic characterization of potentially parental lines acquired from foreign breeding programs and of wild materials. Once the crossings have been planned and obtained, the evaluation of the progeny takes place following a process of Marker-Assisted Selection optimized in order to maintain a compromise between efficiency and cost containment

Interactions between Mytilus galloprovincialis hemocytes and the bivalve pathogens Vibrio aestuarianus 01/032 and Vibrio splendidus LGP32

none9Marine bivalves can accumulate large numbers of bacteria, in particular Vibrio species, whose persistence in bivalve tissues largely depends on their sensitivity to the bactericidal activity of circulating hemocytes and hemolymph soluble factors. The interactions between vibrios and hemolymph have been investigated, in particular in bivalve species susceptible to infection by certain Vibrio spp. and strains. In this work, the effects of two bivalve pathogens, Vibrio splendidus LGP32 (V.s.) and Vibrio aestuarianus 01/032 (V.a.), isolated from oyster mortality outbreaks, on the hemocytes of Mytilus galloprovincialis were investigated. In vitro, V.s., but not V.a., induced a dramatic decrease in lysosomal membrane stability-LMS in the hemocytes; both vibrios induced a moderate lysozyme release, with V.s. > V.a.. The V.s.-induced decrease in LMS was mediated by activation of PI-3Kinase, as shown by use of different kinase inhibitors. TEM analysis showed rapid internalization of both vibrios; however, V.s. lead to cellular and lysosomal damage and was able to survive within the hemocytes, whereas significant killing of V.a. was observed. In vivo, in mussels challenged with either vibrio and sampled at 6, 24 and 96 h post-injection, transient decreases in hemocyte LMS and progressive increases in serum lysozyme activity were observed, with V.s. > V.a.. Moreover, whereas V.a. was efficiently cleared from hemolymph, V.s. showed significant growth, that was maximal at 24 h p.i. when lowest LMS values were recorded in the hemocytes. Both vibrios also induced significant decreases in LMS in the digestive gland, again with V.s. > V.a.. The results indicate distinct interactions between mussel hemocytes and the two vibrio strains tested. The effects of V.s. may be due to the capacity of this strain to interfere with the signaling pathways involved in hemocyte function, thus escaping the bactericidal activity of the host cell, as observed for certain mammalian pathogens. Although V.s. is considered not pathogenic to Mytilus, this vibrio strain can affect the lysosomal function at the cellular and tissue level, thus leading to stressful conditions.openBalbi, T; Fabbri, R; Cortese, K; Smerilli, A; Ciacci, C; Grande, C; Vezzulli, L; Pruzzo, C; Canesi, LBalbi, T; Fabbri, R; Cortese, K; Smerilli, A; Ciacci, C; Grande, C; Vezzulli, Luigi; Pruzzo, C; Canesi,

A 'Regent' pedigree update: ancestors, offspring and their confirmed resistance loci

'Regent' is the fungal resistant grapevine cultivar with the highest acreage in Germany and an important resistance donor in international breeding programs. It carries the resistance loci Rpv3.1 as well as Ren3 and Ren9 against downy and powdery mildew, respectively. As the parents of 'Chambourcin', the resistant paternal ancestor of 'Regent', did not coincide with the breeder's information, the germplasm repository of JKI Geilweilerhof was screened to find the missing ancestors. SSR marker analysis revealed that 'Joannes Seyve 11369' and 'Plantet' are the true parents of 'Chambourcin' and not 'Seyve Villard 12-417' and 'Chancellor'. Furthermore, the origin of the resistance loci Ren3 and Ren9 could be traced back to the genotypes 'Seibel 4614' and 'Munson'. Since the breeder Hermann Jaeger mentioned 'Munson' as a direct descendant of Vitis aestivalis Michx. var. linsecomii (Buckley) L. H. Bailey and Vitis rupestris Scheele, one of these wild species might have been the donor of the loci