Gray mold caused by Botrytis cinerea limits grape production in Chile. Bernardo A. Latorre, Karina Elfar, and Enrique E. Corresponding author: blatorre uc.
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In vitro inhibition of Botrytis cinerea with extracts of wild grapevine Vitis spp. RVS presented the second higher inhibition rate. This dose contained phenolic compounds Botrytis cinerea Teleomorph: Botryotinia fuckeliana , the causal agent of gray rot, damages a wide range of plants worldwide Piesik et al.
This fungus has developed resistance to some conventional fungicides, particularly benzimidazole and dicarboximide fungicides Panebianco et al. Multiple applications of the different chemical control treatments to fruit and vegetables may cause health problems to field workers and consumers, export restrictions due to pesticide residues, damage to the environment and harmful effects on organisms beneficial to agriculture Yadav et al.
An alternative for integral plant disease management is to use natural compounds produced by some plants that have antifungal properties Compean and Ynalvez, Among those compounds are isoflavonoids, diterpenoids, alcaloids, essential oils, stilbenes and polypeptides Soylu et al. In particular, Vitis vinifera has been reported to produce several compounds with antimicrobial and antifungal activity, such as myricetin, ellagic acid, kaempferol, quercitin, and gallic acid, among others Schnee et al.
Within the phenolic compounds present in fruit peel extracts of a mix of three Chilean cultivars of V. Also, resveratrol RVS , a compound that was isolated for the first time from white hellebores Veratrum album and belongs to the stilbene group, has been reported as being active against different pathogenic fungi in grapevine crops, i.
Erysiphe necator , Plasmopara viticola and B. On the other hand, micelial growth of B. Over 16 wild species of the genus Vitis have been reported to be present in Mexico Cruz, These plants are being systematically studied to enhance their use. In particular, the leaves of wild grapevines Vitis spp.
Tobar-Reyes et al. Given that the presence of phenolic compounds in wild grapevine indicates their potential as a natural control agent for plant diseases, in this study we evaluated their in vitro antifungal activity against B.
Later, using the hyphal tip technique, a portion of mycelial tissue was transferred to another Petri dish containing MCAvA to obtain a pure isolate of the fungus. Festival strawberries that had been previously disinfected as described earlier. Once the typical symptoms of gray rot appeared, the fungus was identified by comparing the morphological structures observed mycelium type and color, septation, conidiophores, conidiophore branching, conidia length and width using the Barnett and Hunter keys The specialized descriptions by Ellis and Crous et al.
Samples of the fungus were also sent to the Laboratory of Phytosanitary Diagnosis of Colegio de Postgraduados, Montecillo, Mexico, where they were molecularly analyzed. Once the species was confirmed, the fungus was grown to perform the tests of this experiment. Healthy leaves of uniform size and color were obtained Franco et al.
Afterwards, the extract was filtered and concentrated in a rotary evaporator until 50 mL of stock solution free of the extraction solvent was obtained.
Using gallic acid as a standard, the total amount of phenols contained in extracts of wild grapevine leaves EHVS was determined using the method of Folin-Ciocalteu Mora et al. The result was expressed in gallic acid equivalents EAG.
For the mobile phase we used water: acetonitrile: acetic acid The flow level was 1 mL min -1 and we allowed a five minute stabilization interval between each sample. The signal was monitored at nm Lorrain et al. The samples were injected three times in the chromatographic system and the compounds detected using the standard internal method were quantified.
Table 1. Concentration of phenolic compounds in oat-agar adding extracts of leaves of wild grapevine Vitis spp. EAG: Equivalents of gallic acid. Each solution was poured into properly labeled Petri dishes and allowed to solidify.
Each of the 19 treatments were repeated four times using three Petri dishes per replication; each of the Petri dishes was an experimental unit. The experiment was performed twice, and because of the similarity of both results, the statistical analysis included in this report corresponds to only one experiment. Measurement of each replication ended when the surface of MCAvA in all the TA dishes was completely covered by mycelium. The Petri dishes within the incubator were arranged in a completely randomized design.
At the end of the experiment, MCAvA spores were collected from each treatment using a glass rod and sterile distilled water. For each previously described experimental unit, an aliquot of the suspension containing conidia was taken and transferred to a Neubauer chamber to count the conidia following the methodology of Moo-Koh et al. Two hundred microliters of the suspension were poured in Petri dishes with MCAvA supplemented with the different doses of each of the 19 treatments.
The conidia were considered germinated when the length of the germ tube was equal to or greater than the diameter of the conidium. The percentage of spore germination inhibition IGC was determined with the formula used by Soylu et al.
The experimental design and number of replications per treatment were the same ones used for mycelium growth. Four days after inoculation with Botrytis sp. On the other hand, in MCAvA, colonies showed white concentric mycelium growth of velvety consistency; seven days later, the mycelium turned gray Figure 1B.
Under a light microscope, we observed long, septate, pigmented conidia with smooth walls, apically branched and with bunches of conidia Figure 1C. All the features observed corresponded to B. Figure 1. Isolate of Botrytis cinerea from Capsicum annuum fruits. A Symptoms of gray rot in strawberry fruits. Four-day old colony cultivated in oat-agar. C Branched tip of a conidiophore with conidia bunches; D Botrytis cinerea conidia. Table 2. Figure 2. On the other hand, all the treatments showed IGC.
This study suggests that the difference among the accessions in the total content of phenolic compounds, and especially of the three phenols identified, is mainly the result of genetics, as the plant tissues of the three accessions came from the same germplasm bank. The inhibitory effect of the development of B. Vio-Michaelis et al. In particular, the action mode of polyphenols against B. All this inhibits the development of mycelium and prevents sporulation Adrian y Jeandet, ; Minova et al.
It has been suggested that polyphenols of the type 3b-hydroxy-kaurenoic acid are involved in the reduction of B. These results may be due to results reported by Guerrero et al. In spite of the lower control in vitro of B. The widespread presence of wild grapevine in Mexico and its low cost are other factors that also contribute to the interest in fully identifying the phenols present in EHVS and their particular effect on the studied fungus.
The methanolic extracts of leaves of three wild grapevine Vitis spp. Reactive grade resveratrol showed better control of the fungus than the extracts of wild grapevine leaves.
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Proceedings of the Latvian Academy of Sciences
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