A 3-year application of different mycorrhiza-based plant biostimulants distinctively modulates photosynthetic performance, leaf metabolism, and fruit quality in grapes (Vitis vinifera L.).

The use of microbial biostimulants in agriculture is recognized as a sustainable approach to promoting crop productivity and quality due to improved nutrient uptake, enhanced stress tolerance, and improved ability to cope with non-optimal environments. The present paper aimed to comparatively investigate the effect of seven different commercial mycorrhizal-based treatments in terms of yield, phytochemical components, and technological traits of Malvasia di Candia Aromatica grape (Vitis vinifera L.) plants. Metabolomic analysis and photosynthetic performance were first investigated in leaves to point out biochemical differences related to plant growth. Higher photosynthetic efficiency and better PSII functioning were found in biostimulant-treated vines, reflecting an overall decrease in photoinhibition compared to untreated plants. Untargeted metabolomics followed by multivariate statistics highlighted a robust reprogramming of primary (lipids) and secondary (alkaloids and terpenoids) metabolites in treated plants. The analysis of berry yield and chemical components exhibited significant differences depending on the biostimulant product. Generally, berries obtained from treated plants displayed improved contents of polyphenols and sugars, while yield remained unchanged. These results elucidated the significant role of microbial biostimulants in determining the quality of grape berries and eliciting biochemical changes in vines.

The Functional Profile and Antioxidant Capacity of Tomato Fruits Are Modulated by the Interaction between Microbial Biostimulants, Soil Properties, and Soil Nitrogen Status.

The application of microbial biostimulants to plants has revealed positive effects related to nutrients uptake, stress tolerance, root development and phenological growth. However, little information is available exploiting the potential synergistic biostimulant action of microbes on the functional quality of the yields. The current research elucidated the effect of single or coupled action of biostimulants, associated with either optimal or reduced nitrogen application, on the functional quality of tomato fruits. Chemical assays and untargeted metabolomics were applied to investigate Rhizoglomus irregulare and Funneliformis mosseae administration (both being arbuscular mycorrhiza, AMF), under optimal or low N input conditions, alone or coupled to Trichoderma atroviride application. The coupling of AMF and Trichoderma fungal inoculations resulted in a synergistic biostimulant effect on tomato fruits under sub-optimal fertility, revealing improved concentrations of carotenoid compounds-B-carotene (0.647 ± 0.243 mg/100 g), Z-carotene (0.021 ± 0.021 mg/100 g), 13-z-lycopene (0.145 ± 0.052 mg/100 g) and all-trans-lycopene (12.586 ± 1.511 mg/100 g), and increased values for total phenolic content (12.9 ± 2.9 mgGAE/g), total antioxidant activity (phosphomolybdenum, 0.9 ± 0.2 mmolTE/g), radical scavenging activity (DPPH, 3.4 ± 3.7 mgTE/g), reducing power (FRAP, 23.6 ± 6.3 mgTE/g and CUPRAC, 37.4 ± 7.6 mg TE/g), and enzyme inhibitory activity (AChE, 2.4 ± 0.1 mg GALAE/g), when compared to control. However, evidence of carotenoid and bioactive compounds were exclusively observed under the sub-optimal fertility and no significant differences could be observed between the biostimulant treatment and control under optimal fertility.

Plant Disease Models and Forecasting: Changes in Principles and Applications over the Last 50 Years.

This review gives a perspective of selected advances made since the middle of the 20th century in plant disease modeling, and the associated increase in the number of models published during that time frame. This progress can be mainly attributed to advances in (i) sensors and automatic environmental data collection technology, (ii) instrumentation and methods for studying botanical epidemiology, and (iii) data analytics and computer science. We review the evolution of techniques for developing data-based (empirical) models and process-based (mechanistic) models using the wheat rusts as a case study. We also describe the increased importance of knowledge about biological processes for plant disease modeling by using apple scab as a second case study. For both wheat rusts and apple scab, we describe how the models have evolved over the last 50 years by considering certain milestones that have been achieved in disease modeling. Finally, we describe how plant disease models are used as part of a multi-modeling approach to develop decision-making tools in the application of integrated pest management.

Development and Validation of a Mechanistic Model That Predicts Infection by Diaporthe ampelina, the Causal Agent of Phomopsis Cane and Leaf Spot of Grapevines.

Phomopsis cane and leaf spot (PCLS), known in Europe as "excoriose," is an important fungal disease of grapevines caused by Diaporthe spp., and most often by Diaporthe ampelina (synonym Phomopsis viticola). PCLS is re-emerging worldwide, likely due to climate change, changes in the management of downy mildew from calendar- to risk-based criteria that eliminate early-season (unnecessary) sprays, and the progressive reduction in the application of broad-spectrum fungicides. In this study, a mechanistic model for D. ampelina infection was developed based on published information. The model accounts for the following processes: (i) overwintering and maturation of pycnidia on affected canes; (ii) dispersal of alpha conidia to shoots and leaves; (iii) infection; and (iv) onset of disease symptoms. The model uses weather and host phenology to predict infection periods and disease progress during the season. Model output was validated against 11 independent PCLS epidemics that occurred in Italy (4 vineyards in 2019 and 2020) and Montenegro (3 vineyards in 2020). The model accurately predicted PCLS disease progress, with a concordance correlation coefficient (CCC) = 0.925 between observed and predicted data. A ROC analysis (AUROC>0.7) confirmed the ability of the model to predict the infection periods leading to an increase in PCLS severity in the field, indicating that growers could use the model to perform risk-based fungicide applications.

Nitrogen use efficiency, rhizosphere bacterial community, and root metabolome reprogramming due to maize seed treatment with microbial biostimulants.

Seed inoculation with beneficial microorganisms has gained importance as it has been proven to show biostimulant activity in plants, especially in terms of abiotic/biotic stress tolerance and plant growth promotion, representing a sustainable way to ensure yield stability under low input sustainable agriculture. Nevertheless, limited knowledge is available concerning the molecular and physiological processes underlying the root-inoculant symbiosis or plant response at the root system level. Our work aimed to integrate the interrelationship between agronomic traits, rhizosphere microbial population and metabolic processes in roots, following seed treatment with either arbuscular mycorrhizal fungi (AMF) or Plant Growth-Promoting Rhizobacteria (PGPR). To this aim, maize was grown under open field conditions with either optimal or reduced nitrogen availability. Both seed treatments increased nitrogen uptake efficiency under reduced nitrogen supply revealed some microbial community changes among treatments at root microbiome level and limited yield increases, while significant changes could be observed at metabolome level. Amino acid, lipid, flavone, lignan, and phenylpropanoid concentrations were mostly modulated. Integrative analysis of multi-omics datasets (Multiple Co-Inertia Analysis) highlighted a strong correlation between the metagenomics and the untargeted metabolomics datasets, suggesting a coordinate modulation of root physiological traits.

Development of an online pan-European Integrated Pest Management Resource Toolbox.

The IPM WORKSIPM Resource Toolbox (Toolbox) has been developed as an interactive, online repository of integrated pest management (IPM) resources. Populated with high priority resources for farmers and their advisors during the project, its structure enables additional resources added over time. The repository is a public interactive website, available to anyone looking to access, understand, and implement IPM. Built on an open-source content management system, the toolbox is designed to require minimal post-production site maintenance and support, while being easily expanded to integrate resources from future initiatives. To ensure an efficient but comprehensive website design, population, maintenance, a survey of target user needs was conducted. Different type of IPM stakeholders, both internal and external to the IPMworks project, ranked the key requirements for the Toolbox, such as practical information about diseases and pests' management and economic thresholds: 343 feedbacks and answers from a survey of 10 questions, carried out across Europe in four languages, provided the key elements and foundation for the IPM Resource Toolbox website development and specification. The Toolbox resources are explained in different languages, with images, divided by topics, with the possibility to find additional details and accessible by smartphone.

Evaluation of Sown Cover Crops and Spontaneous Weed Flora as a Potential Reservoir of Black-Foot Pathogens in Organic Viticulture.

(1) Background. An extensive survey of grapevine-sown cover crops and spontaneous weed flora was conducted from 2019 to 2020 in organic vineyards in six European countries (France, Italy, Romania, Slovenia, Spain, Switzerland). Our main objective was to detect and identify the presence of Cylindrocarpon-like asexual morphs species associated with black-foot disease on their roots. (2) Methods. Fungal isolations from root fragments were performed on culture media. Cylindrocarpon-like asexual morph species were identified by analyzing the DNA sequence data of the histone H3 (his3) gene region. In all, 685 plants belonging to different botanical families and genera were analyzed. Cylindrocarpon-like asexual morphs were recovered from 68 plants (9.9% of the total) and approximately 0.97% of the plated root fragments. (3) Results. Three fungal species (Dactylonectria alcacerensis, Dactylonectria torresensis, Ilyonectria robusta) were identified. Dactylonectria torresensis was the most frequent, and was isolated from many cover crop species in all six countries. A principal component analysis with the vineyard variables showed that seasonal temperatures and organic matter soil content correlated positively with Cylindrocarpon-like asexual morphs incidence. (4) Conclusions. The presence of Cylindrocarpon-like asexual morphs on roots of cover crops suggests that they can potentially act as alternative hosts for long-term survival or to increase inoculum levels in vineyard soils.

Dynamics of Diaporthe ampelina Conidia Released from Grape Canes that Overwintered in the Vineyard.

Phomopsis cane and leaf spot (PCLS) is an important disease of grapevines that is mainly caused by Diaporthe ampelina. Dispersal dynamics of D. ampelina spores were investigated in two vineyards, one in northern Italy and one in Montenegro, by using spore samplers that collected α- and ß-conidia from rain water running off from PCLS-affected canes. The canes were collected from each vineyard, deployed, and overwintered in the corresponding vineyards. In each of three years (2016, 2017, and 2018), conidial dispersal was investigated during one (Montenegro) or two (Italy) growing seasons following the deployment of the PCLS-affected canes. In the first growing season following cane deployment in both vineyards, α-conidia were mostly found in runoff water after grapevine bud break, especially in April and May, and ß-conidia were regularly found in numbers comparable to those of α-conidia, most frequently from June to September. In Italy, high numbers of α- and ß-conidia were also collected during the second growing season following cane deployment. The dispersal dynamics of α-conidia over time were described by a Gompertz equation using hydrothermal time (i.e., the accumulated effect of temperature on the maturation rate of pycnidia on days in which the number of hours of wetness was ≥6 or 9 h), with R2 and concordance correlation coefficient >0.9. Rain (≥0.2 mm) was a good predictor of conidial dispersal, with an overall accuracy of 0.97. These results increase our understanding of D. ampelina spore dispersal and should be integrated into warning systems for PCLS management.

Development and Evaluation of a Model that Predicts Grapevine Anthracnose Caused by Elsinoë ampelina.

Grapevine anthracnose caused by Elsinoë ampelina is a serious threat in many vineyards, and its control requires repeated application of fungicides, usually on a calendar basis. A better understanding of the pathogen life cycle would help growers manage anthracnose more safely and effectively. After conducting a systematic literature search of grape anthracnose, we used the retrieved information and data to develop a mechanistic model based on systems analysis. The model simulates production and maturation of primary inoculum, infection caused by both primary and secondary conidia, and lesion formation and production of secondary inoculum. The model was validated for its ability to predict first seasonal onset of anthracnose lesions by using 8 years of data collected at Auckland, New Zealand, and disease progress during the season by using 3 years of data collected at Frelighsburg, Canada. Overall, the model provided accurate predictions of infection occurrence, with 0.96 accuracy, 0.91 sensitivity, and 0.97 specificity. The model also showed good accuracy for predicting disease progress, with a concordance correlation coefficient between observed and predicted disease severities of 0.92, a root mean square error of 0.14, and a coefficient of residual mass of 0.06. Although the model failed to predict 10 of 110 real infection periods, these missed infections led to only mild disease symptoms. We therefore conclude that the model is reliable and can be used to reduce the costs of anthracnose management by improving the timing of fungicide applications.

Assessment of Resistance Components for Improved Phenotyping of Grapevine Varieties Resistant to Downy Mildew.

Grapevine varieties showing partial resistance to downy mildew, caused by Plasmopara viticola, are a promising alternative to fungicides for disease control. Resistant varieties are obtained through breeding programs aimed at incorporating Rpv loci controlling the quantitative resistance into genotypes characterized by valuable agronomic and wine quality traits by mean of crossing. Traditional phenotyping methods used in these breeding programs are mostly based on the assessment of the resistance level after artificial inoculation of leaf discs in bioassays, by using the visual score proposed in the 2nd Edition of the International Organization of Vine and Wine (OIV) Descriptor List for Grape Varieties and Vitis species (2009). In this work, the OIV score was compared with an alternative approach, not used for the grapevine-downy mildew pathosystem so far, based on the measurement of components of resistance (RCs); 15 grapevine resistant varieties were used in comparison with the susceptible variety 'Merlot'. OIV scores were significantly correlated with P. viticola infection frequency (IFR), the latent period for the downy mildew (DM) lesions to appear (LP50), and the number of sporangia produced per lesion (SPOR), so that when the OIV score increased (i.e., the resistance level increases), IFR and SPOR decreased, while LP50 increased. The relationship was linear for LP50, monomolecular for IFR and hyperbolic for SPOR. No significant correlation was found between OIV score and DM lesion size, sporangia produced per unit area of lesion, length of infectious period, and infection efficiency of the sporangia produced on DM lesions. The correlation between OIV score and area under the disease progress curve (AUDPC) calculated by using the RCs and a simulation model was significant and fit an inverse exponential function. Based on the results of this study, the measurement of the RCs to P. viticola in grapevine varieties by means of monocyclic, leaf disc bioassays, as well as their incorporation into a model able to simulate their effect on the polycyclic development of DM epidemics in vineyards, represents an improved method for phenotyping resistance level.

A network meta-analysis provides new insight into fungicide scheduling for the control of Botrytis cinerea in vineyards.

BACKGROUND: Control of Botrytis bunch rot (BBR) is currently based on the application of fungicides at four timings corresponding to specific growth stages of vines: end of flowering (A), pre-bunch closure (B), veraison (C) and before harvest (D). The current research provides a network meta-analysis of 116 studies conducted between 1963 and 2016 in nine countries, in which 14 strategies (based on combinations of 1, 2, 3, or 4 sprays applied in A, B, C, and/or D) were compared. RESULTS: When a one-spray strategy was applied, BBR control was more effective with sprays applied in A, C, or D than B. With a two-spray strategy, strategy AC provided similar control as strategy BC; strategy CD also provided good control. For a 3-spray strategy, the best disease control was consistently obtained with strategy ACD. Four-spray strategy ABCD provided the best control but often involved needless sprays so that the routine application of four sprays is not justified. CONCLUSIONS: Spraying at timing A seems to be very important for achieving efficient and flexible disease control. Flexibility is reduced by spraying at timing B rather than A. © 2018 Society of Chemical Industry.

A Mechanistic Model of Botrytis cinerea on Grapevines That Includes Weather, Vine Growth Stage, and the Main Infection Pathways.

A mechanistic model for Botrytis cinerea on grapevine was developed. The model, which accounts for conidia production on various inoculum sources and for multiple infection pathways, considers two infection periods. During the first period ("inflorescences clearly visible" to "berries groat-sized"), the model calculates: i) infection severity on inflorescences and young clusters caused by conidia (SEV1). During the second period ("majority of berries touching" to "berries ripe for harvest"), the model calculates: ii) infection severity of ripening berries by conidia (SEV2); and iii) severity of berry-to-berry infection caused by mycelium (SEV3). The model was validated in 21 epidemics (vineyard × year combinations) between 2009 and 2014 in Italy and France. A discriminant function analysis (DFA) was used to: i) evaluate the ability of the model to predict mild, intermediate, and severe epidemics; and ii) assess how SEV1, SEV2, and SEV3 contribute to epidemics. The model correctly classified the severity of 17 of 21 epidemics. Results from DFA were also used to calculate the daily probabilities that an ongoing epidemic would be mild, intermediate, or severe. SEV1 was the most influential variable in discriminating between mild and intermediate epidemics, whereas SEV2 and SEV3 were relevant for discriminating between intermediate and severe epidemics. The model represents an improvement of previous B. cinerea models in viticulture and could be useful for making decisions about Botrytis bunch rot control.

Production and release of asexual sporangia in Plasmopara viticola.

To study the influence of environmental conditions on sporulation of Plasmopara viticola lesions under vineyard's conditions, unsprayed vines were inspected every second or third day and the numbers of sporulating and nonsporulating lesions were counted in two North Italy vineyards in 2008 to 2010. Infected leaves were removed so that only fresh lesions were assessed at each field assessment. Sporulation was studied at two scales, across field assessments and across the seasonal population of lesions. Frequencies of sporulating lesions were positively correlated with the numbers of moist hours in the preceding dark period (i.e., the number of hours between 8:00 p.m. and 7:00 a.m. with relative humidity ≥80%, rainfall >0 mm, or wetness duration >30 min). In a receiver operating characteristic analysis, predicted sporulation based on the occurrence of ≥3 moist hours at night provided overall accuracy of 0.85. To study the time course of sporulation on lesions which were not washed by rainfall, numbers of sporangia produced per square millimeter of lesion were estimated on individual cohorts of lesions over the whole infectious period. The numbers of sporangia per square millimeter of lesion increased rapidly during the first 4 days after the beginning of sporulation and then tapered off prior to a halt; the time course of cumulative sporangia production by a lesion followed a monomolecular growth model (R(2) = 0.97). The total number of sporangia produced by a square millimeter of lesion increased as the maximum temperature decreased and moist hours in the dark increased. To study the release pattern of the sporangia, spore samplers were placed near grapevines with sporulating lesions. Airborne sporangia were caught in 91.2% of the days over a wide range of weather conditions, including rainless periods. The results of this study provide quantitative information on production of P. viticola sporangia that may help refine epidemiological models used as decision aids in grape disease management programs.

No indication of strict host associations in a widespread mycoparasite: grapevine powdery mildew (Erysiphe necator) is attacked by phylogenetically distant Ampelomyces strains in the field.

Pycnidial fungi belonging to the genus Ampelomyces are common intracellular mycoparasites of powdery mildews worldwide. Some strains have already been developed as commercial biocontrol agents (BCAs) of Erysiphe necator and other powdery mildew species infecting important crops. One of the basic, and still debated, questions concerning the tritrophic relationships between host plants, powdery mildew fungi, and Ampelomyces mycoparasites is whether Ampelomyces strains isolated from certain species of the Erysiphales are narrowly specialized to their original mycohosts or are generalist mycoparasites of many powdery mildew fungi. This is also important for the use of Ampelomyces strains as BCAs. To understand this relationship, the nuclear ribosomal DNA internal transcribed spacer (ITS) and partial actin gene (act1) sequences of 55 Ampelomyces strains from E. necator were analyzed together with those of 47 strains isolated from other powdery mildew species. These phylogenetic analyses distinguished five major clades and strains from E. necator that were present in all but one clade. This work was supplemented with the selection of nine inter-simple sequence repeat (ISSR) markers for strain-specific identification of Ampelomyces mycoparasites to monitor the environmental fate of strains applied as BCAs. The genetic distances among strains calculated based on ISSR patterns have also highlighted the genetic diversity of Ampelomyces mycoparasites naturally occurring in grapevine powdery mildew. Overall, this work showed that Ampelomyces strains isolated from E. necator are genetically diverse and there is no indication of strict mycohost associations in these strains. However, these results cannot rule out a certain degree of quantitative association between at least some of the Ampelomyces lineages identified in this work and their original mycohosts.

The role of rain in dispersal of the primary inoculum of Plasmopara viticola.

Although primary infection of grapevines by Plasmopara viticola requires splash dispersal of inoculum from soil to leaves, little is known about the role of rain in primary inoculum dispersal. Distribution of rain splashes from soil to grapevine canopy was evaluated over 20 rain periods (0.2 to 64.2 mm of rain) with splash samplers placed within the canopy. Samplers at 40, 80, and 140 cm above the soil caught 4.4, 0.03, and 0.003 drops/cm(2) of sampler area, respectively. Drops caught at 40 and 80 cm (1.5 cm in diameter) were larger than drops at 140 cm (1.3 cm). Leaf coverage by splashed drops, total drop number, and drop size increased with an increase in the maximum intensity of rain (mm/h) during any rain period. Any rainfall led to infection in potted grapevines placed outside on leaf litter containing oospores, if the litter contained germinated oospores at the time of rain; infection severity was unrelated to rain amount or intensity. Results from vineyards also indicate that any rain can carry P. viticola inoculum from soil to leaves and should be considered a splash event in disease prediction systems. Sampling for early disease detection should focus on the lower canopy, where the probability of splash impact is greatest.

Evaluation of a Warning System for Early-Season Control of Grapevine Powdery Mildew.

In several grape-growing areas of the world, including northern Italy, powdery mildew epidemics, caused by Erysiphe necator, are mainly triggered by the ascospores produced in overwintered chasmothecia. Growers in northern Italy usually control the disease with fixed-interval fungicide applications. A warning system was developed for early-season powdery mildew control based on (i) short-term weather forecasts, (ii) a model that simulates the severity of each E. necator ascosporic infection, and (iii) a mobile phone short-message system. This warning system was evaluated in six vineyards in northern Italy from 2006 to 2008, between bud break of vines and early berry development; an unsprayed control was compared with "low-risk" and "high-risk" model-driven sprays and a calendar-based "grower" spray program. Use of the warning system reduced disease severity on leaves and bunches compared with the unsprayed control and resulted in the same level of control of powdery mildew as the grower's spray program, with reduced fungicide applications and costs. On average, 5.7 sprays were applied following the grower's spray program (with an average cost of 221 €/ha/year); use of the warning system reduced fungicide applications by 36% (low-risk program, saving of 56 €/ha/year) or 75% (high-risk program, saving of 161 €/ha/year).

Dynamics of ascospore maturation and discharge in Erysiphe necator, the causal agent of grape powdery mildew.

Dynamics of ascocarp development, ascospore maturation, and dispersal in Erysiphe necator were studied over a 4-year period, from the time of ascocarp formation to the end of the ascosporic season at the end of June in the following spring. Naturally dispersed chasmothecia were collected from mid-August to late November (when leaf fall was complete); the different collections were used to form three to five cohorts of chasmothecia per year, with each cohort containing ascocarps formed in different periods. Chasmothecia were exposed to natural conditions in a vineyard and periodically sampled. Ascocarps were categorized as containing mature or immature ascospores, or as empty; mature ascospores inside chasmothecia were enumerated starting from late February. Ascospore discharge was determined using silicone-coated slides that were placed 3 to 4 cm from sections of the vine trunk holding the chasmothecia. Before complete leaf fall, 34% of the chasmothecia had mature ascospores, 48% had immature ascospores, and 18% were empty; in the same period, the trapped ascospores represented 56% of the total ascospores trapped in an ascosporic season (i.e., from late summer until the next spring or early summer). The number of viable chasmothecia diminished over time; 11 and 5% of chasmothecia had mature ascospores between complete leaf fall and bud break and after bud break, respectively. These ascocarps discharged ≈2 and 42% of the total ascospores, respectively. All the ascocarp cohorts released ascospores in autumn, survived the winter, and discharged viable ascospores in spring; neither ascospore numbers nor their pattern of temporal release was influenced by the time when chasmothecia were collected and exposed in the vineyard. Abundance of mature ascospores in chasmothecia was expressed as a function of degree-days (DD) (base 10°C) accumulated before and after bud break through a Gompertz equation (R² = 0.92). Based on this equation, 90% of the ascospores were mature when 153 DD (confidence interval, 100 to 210 DD) had accumulated after bud break. Most ascospores were trapped in periods with >2 mm of rain; however, a few ascospores were airborne with <2 mm of rain and, occasionally, in wet periods of ≥3.5 h not initiated by rain.