Osmotic Stress and ABA Affect Immune Response and Susceptibility of Grapevine Berries to Gray Mold by Priming Polyamine Accumulation

Abiotic factors inducing osmotic stress can affect plant immunity and resistance against pathogen attack. Although a number of studies have characterized grapevine responses to various forms of biotic and abiotic stresses, the relationships between osmotic stress response and susceptibility of mature berries to Botrytis cinerea still remain unknown. In this study, we investigated the effects of osmotic stress and abscisic acid (ABA) on defense responses of mature grapevine berries before and after B. cinerea infection. We focused on the possible involvement of polyamines in the interaction between osmotic stress response and susceptibility to B. cinerea. We showed that osmotic stress induced by PEG or sucrose, and exogenous ABA induce transient but low defense responses, including weak expression of PR genes and phytoalexin synthesis in mature berries. This was accompanied by an upregulation of NCED2 involved in ABA biosynthesis and a large production of free polyamines. However, osmotic stress followed by B. cinerea infection primed berries for enhanced accumulation of polyamines, but slowed down the defense responses and increased susceptibility to the pathogen. A weak increase of diamine- and polyamine-oxidase activities was also recorded in stressed berries, but declined after pathogen infection. The pretreatment of stressed berries with appropriate inhibitors of diamine- and polyamine-oxidases further increased polyamine level and greatly lowered defense responses, leading to higher susceptibility to B. cinerea. These results suggest that increased polyamine titer through low activation of their oxidative degradation in grape berries may contribute at least in part to the weakening of defense responses and subsequent disease susceptibility

Bacillus subtilis PTA-271 Counteracts Botryosphaeria Dieback in Grapevine, Triggering Immune Responses and Detoxification of Fungal Phytotoxins

Plant pathogens have evolved various strategies to enter hosts and cause diseases. Particularly Neofusicoccum parvum, a member of Botryosphaeria dieback consortium, can secrete the phytotoxins (-)-terremutin and (R)-mellein during grapevine colonization. The contribution of phytotoxins to Botryosphaeria dieback symptoms still remains unknown. Moreover, there are currently no efficient control strategies of this disease, and agro-environmental concerns have raised increasing interest in biocontrol strategies to limit disease spread in vineyards, especially by using some promising beneficial bacteria. Here, we first examined in planta the biocontrol capacity of Bacillus subtilis PTA-271 against N. parvum Np-Bt67 strain producing both (-)-terremutin and (R)-mellein. We then focused on the direct effects of PTA-271 on pathogen growth and the fate of pure phytotoxins, and explored the capacity of PTA-271 to induce or prime grapevine immunity upon pathogen infection or phytotoxin exposure. Results provided evidence that PTA-271 significantly protects grapevine cuttings against N. parvum and significantly primes the expression of PR2 (encoding a β-1,3-glucanase) and NCED2 (9-cis-epoxycarotenoid dioxygenase involved in abscisic acid biosynthesis) genes upon pathogen challenge. Using in vitro plantlets, we also showed that PTA-271 triggers the expression of salicylic acid- and jasmonic acid-responsive genes, including GST1 (encoding a glutathione-S-transferase) involved in detoxification process. However, in PTA-271-pretreated plantlets, exogenous (-)-terremutin strongly lowered the expression of most of upregulated genes, except GST1. Data also indicated that PTA-271 can detoxify both (-)-terremutin and (R)-mellein and antagonize N. parvum under in vitro conditions. Our findings highlight (-)-terremutin and (R)-mellein as key aggressive molecules produced by N. parvum that may weaken grapevine immunity to promote Botryosphaeria dieback symptoms. However, PTA-271 can efficiently attenuate Botryosphaeria dieback by enhancing some host immune responses and detoxifying both phytotoxins produced by N. parvum

Bacillus subtilis PTA-271 counteracts Botryosphaeria dieback in grapevine, triggering immune responses and detoxification of fungal phytotoxins

Plant pathogens have evolved various strategies to enter hosts and cause diseases. Particularly Neofusicoccum parvum, a member of Botryosphaeria dieback consortium, can secrete the phytotoxins (-)-terremutin and (R)-mellein during grapevine colonization. The contribution of phytotoxins to Botryosphaeria dieback symptoms still remains unknown. Moreover, there are currently no efficient control strategies of this disease, and agro-environmental concerns have raised increasing interest in biocontrol strategies to limit disease spread in vineyards, especially by using some promising beneficial bacteria. Here, we first examined in planta the biocontrol capacity of Bacillus subtilis PTA-271 against N. parvum Np-Bt67 strain producing both (-)- terremutin and (R)-mellein. We then focused on the direct effects of PTA-271 on pathogen growth and the fate of pure phytotoxins, and explored the capacity of PTA- 271 to induce or prime grapevine immunity upon pathogen infection or phytotoxin exposure. Results provided evidence that PTA-271 significantly protects grapevine cuttings against N. parvum and significantly primes the expression of PR2 (encoding a β-1,3-glucanase) and NCED2 (9-cis-epoxycarotenoid dioxygenase involved in abscisic acid biosynthesis) genes upon pathogen challenge. Using in vitro plantlets, we also showed that PTA-271 triggers the expression of salicylic acid- and jasmonic acid-responsive genes, including GST1 (encoding a glutathione-S-transferase) involved in detoxification process. However, in PTA-271-pretreated plantlets, exogenous (-)-terremutin strongly lowered the expression of most of upregulated genes, except GST1. Data also indicated that PTA-271 can detoxify both (-)- terremutin and (R)-mellein and antagonize N. parvum under in vitro conditions. Our findings highlight (-)-terremutin and (R)-mellein as key aggressive molecules produced by N. parvum that may weaken grapevine immunity to promote Botryosphaeria dieback symptoms. However, PTA-271 can efficiently attenuate Botryosphaeria dieback by enhancing some host immune responses and detoxifying both phytotoxins produced by N. parvum

Cultivar Contributes to the Beneficial Effects of Bacillus subtilis PTA-271 and Trichoderma atroviride SC1 to Protect Grapevine Against Neofusicoccum parvum

[EN] Grapevine trunk diseases (GTDs) are a big threat for global viticulture. Without effective chemicals, biocontrol strategies are developed as alternatives to better cope with environmental concerns. A combination of biological control agents (BCAs) could even improve sustainable disease management through complementary ways of protection. In this study, we evaluated the combination of Bacillus subtilis (Bs) PTA-271 and Trichoderma atroviride (Ta) SC1 for the protection of Chardonnay and Tempranillo rootlings against Neofusicoccum parvum Bt67, an aggressive pathogen associated to Botryosphaeria dieback (BD). Indirect benefits offered by each BCA and their combination were then characterized in planta, as well as their direct benefits in vitro. Results provide evidence that (1) the cultivar contributes to the beneficial effects of Bs PTA-271 and Ta SC1 against N. parvum, and that (2) the in vitro BCA mutual antagonism switches to the strongest fungistatic effect toward Np-Bt67 in a three-way confrontation test. We also report for the first time the beneficial potential of a combination of BCA against Np-Bt67 especially in Tempranillo. Our findings highlight a common feature for both cultivars: salicylic acid (SA)-dependent defenses were strongly decreased in plants protected by the BCA, in contrast with symptomatic ones. We thus suggest that (1) the high basal expression of SA-dependent defenses in Tempranillo explains its highest susceptibility to N. parvum, and that (2) the cultivar-specific responses to the beneficial Bs PTA-271 and Ta SC1 remain to be further investigated.Funding This work was supported by a French Grant from the Region GRAND-EST France and the City of GRAND-REIMS France through the BIOVIGNE Ph.D. program, whose functioning is supported by BELCHIM Crop Protection France. DG was supported by the Ramon y Cajal program, Spanish Government (RyC-2017-23098).Leal, C.; Richet, N.; Guise, J.; Gramaje, D.; Armengol Fortí, J.; Fontaine, F.; Trotel-Aziz, P. (2021). Cultivar Contributes to the Beneficial Effects of Bacillus subtilis PTA-271 and Trichoderma atroviride SC1 to Protect Grapevine Against Neofusicoccum parvum. Frontiers in Microbiology. 12:1-17. https://doi.org/10.3389/fmicb.2021.726132S1171

Vitis methods to understand and develop strategies for diagnosis and sustainable control of grapevine trunk diseases

Vitis vinifera is affected by many diseases every year, depending on causal agents, susceptibility of cultivars, and climate region. Some are caused by a single agent, such as gray mold caused by Botrytis cinerea or powdery mildew caused by Erysiphe necator. Others result from the actions of a complex of pathogens such as grapevine trunk diseases (GTDs). GTDs are presently among the most devastating diseases in viticulture worldwide because both the economic losses and the long-term sustainability of vineyards are strongly affected. The complexity of GTDs results from the diversity of associated fungi, the undetermined period of latency within the vine (asymptomatic status), the erratic foliar symptom expression from one year to the next, and, probably correlated with all of these points, the lack of efficient strategies to control them. Distinct methods can be beneficial to improve our knowledge of GTDs. In vitro bioassays with cell suspensions, calli, foliar discs, full leaves, or plantlets, and in vivo natural bioassays with cuttings, grafted plants in the greenhouse, or artificially infected ones in the vineyard, can be applied by using progressive integrative levels of in vitro and in vivo, depending on the information searched. In this review, the methods available to understand GTDs are described in terms of experimental procedures, main obtained results, and deliverable prospects. The advantages and disadvantages of each model are also discussed

Use of beneficial bacteria and their secondary metabolites to control grapevine pathogen diseases

Grapevine is one of the most important economic crops yielding berries, wine products as well as derivates. However, due to the large array of pathogens inducing diseases on this plant, considerable amounts of pesticides—with possible negative impact on the environment and health—have been used and are currently used in viticulture. To avoid negative impacts of such products and to ensure product quality, a substantial fraction of pesticides needs to be replaced in the near future. One solution can be related to the use of beneficial bacteria inhabiting the rhizo- and/or the endosphere of plants. These biocontrol bacteria and their secondary metabolites can reduce directly or indirectly pathogen diseases by affecting pathogen performance by antibiosis, competition for niches and nutrients, interference with pathogen signaling or by stimulation of host plant defenses. Due to the large demand for biocontrol of grapevine diseases, such biopesticides, their modes of actions and putative consequences of their uses need to be described. Moreover, the current knowledge on new strains from the rhizo- and endosphere and their metabolites that can be used on grapevine plants to counteract pathogen attack needs to be discussed. This is in particular with regard to the control of root rot, grey mould, trunk diseases, powdery and downy mildews, pierce’s disease, grapevine yellows as well as crown gall. Future prospects on specific beneficial microbes and their secondary metabolites that can be used as elicitors of plant defenses and/or as biocontrol agents with potential use in a more sustainable viticulture will be further discussed

Pseudomonas spp.-induced systemic resistance to Botrytis cinerea is associated with induction and priming of defence responses in grapevine

Non-pathogenic rhizobacteria Pseudomonas spp. can reduce disease in plant tissues through induction of a defence state known as induced systemic resistance (ISR). This resistance is based on multiple bacterial determinants, but nothing is known about the mechanisms underlying rhizobacteria-induced resistance in grapevine. In this study, the ability of Pseudomonas fluorescens CHA0 and Pseudomonas aeruginosa 7NSK2 to induce resistance in grapevine against Botrytis cinerea is demonstrated. Both strains also triggered an oxidative burst and phytoalexin (i.e. resveratrol and viniferin) accumulation in grape cells and primed leaves for accelerated phytoalexin production upon challenge with B. cinerea. Treatment of cell cultures with crude cell extracts of bacteria strongly enhanced oxidative burst, but resulted in comparable amounts of phytoalexins and resistance to B. cinerea to those induced by living bacteria. This suggests the production of bacterial compounds serving as inducers of disease resistance. Using other strains with different characteristics, it is shown that P. fluorescens WCS417 (Pch-deficient), P. putida WCS358 (Pch- and SA-deficient) and P. fluorescens Q2-87 (a DAPG producer) were all capable of inducing resistance to an extent similar to that induced by CHA0. However, in response to WCS417 (Pch-negative) the amount of H2O2 induced is less than for the CHA0. WCS417 induced low phytoalexin levels in cells and lost the capacity to prime for phytoalexins in the leaves. This suggests that, depending on the strain, SA, pyochelin, and DAPG are potentially effective in inducing or priming defence responses. The 7NSK2 mutants, KMPCH (Pch- and Pvd-negative) and KMPCH-567 (Pch-, Pvd-, and SA-negative) induced only partial resistance to B. cinerea. However, the amount of H2O2 triggered by KMPCH and KMPCH-567 was similar to that induced by 7NSK2. Both mutants also led to a low level of phytoalexins in grapevine cells, while KMPCH slightly primed grapevine leaves for enhanced phytoalexins. This highlights the importance of SA, pyochelin, and/or pyoverdin in priming phytoalexin responses and induced grapevine resistance by 7NSK2 against B. cinerea

The control of proline consumption by abscisic acid during osmotic stress recovery of canola leaf discs

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