Management of gray mold in vine crops

In all wine-growing areas of the world, pests and diseases constitute one of the biggest obstacles to the expansion of vine cultivation, as they affect both the quantity and quality of the final product (Kuhn & Nickel, 1998). In Serra Gaúcha, gray rot is a disease that stands out for the problems it causes and for leading to losses in the quality of fruits in the field and in wine (Garrido & Sônego, 2004; Sônego et al.

Gray rot, gray rot botrytis or gray mold, is caused by Botryotinia fuckeliana (from Bary) Whetzel, conidial form of Botrytis cinerea Pers.Fr. The fungus has more than 200 known hosts, which contributes to its spread, and is present in practically all vineyards in the world, causing damage to both the productivity and quality of the grape (Sônego et al, 2005). It survives in the soil, in cultural remains, in the bark of branches and also in mummified fruits from the previous harvest. In spring, spores are produced and infect new leaves and clusters before flowering. This fungus develops well at high relative humidity values, above 90%, and temperatures between 18ºC and 23ºC (Jackson, 2000; Magalhães et al, 2008; Sonego et al, 2005). Infection of bunches can occur through the parts of the flowers that are susceptible, with the pathogen remaining latent until the berries ripen, when gray rot appears. The calyptra (set of vine flower petals) and the flower receptacle are the predominant sites of infection. B. cinerea (Elmer & Michailides, 2004; Viret et al.

Os Damage caused by this pathogen is a result of peduncular necrosis that occurs at the beginning of cluster formation and gray rot itself in ripe berries at the end of the cycle.

Control of this disease, in most cases, is based on the use of chemical fungicides recommended for the crop. However, it is necessary to highlight that other measures must be used, such as: avoiding the use of rootstocks and excess fertilization that provides too much vigor to the plants; alternate chemical groups; avoid the use of fungicides derived from dithiocarbamic acid; remove leaves and branches that excessively cover the bunches and provide conditions favorable to the disease; remove infected cultural remains; carry out chemical treatment during winter (for example, with lime sulfur mixture) to reduce inoculum sources; and avoid cultivars with very compact clusters (Sônego et al, 2005). In Figure 2B it is possible to see how the compaction of bunches can provide a favorable condition for the occurrence of the disease. In work carried out in New Zealand, the authors observed that the sources of inoculum of B. cinerea they can be in different parts of the plant, such as rachis, tendrils, petioles and branches, which are left in the field after pruning as crop residue or in the crown of the plant – the rachis being the part of the plant with the greatest amount of inoculum. B. cinerea, both in the canopy and in the rest of the culture. It was also possible to observe that the largest amount of inoculum was present during flowering and in the rest of the crop present in the planting line. This shows the importance of taking precautions during flowering time, as it is a potential way for the pathogen to enter the plant (Jaspers et al.

Work has been carried out by researchers around the world in the hope of increasing the options for strategies for managing gray rot in grapes, without the need to necessarily use chemical fungicides. The use of chitosan in grapevines can induce an increase in the production of resistance substances in the plant. B. cinerea, known as phytoalexins – resveratrol and viniferin, for example – providing a reduction in the disease in grape leaves (Aziz et al, 2006). Other authors observed a direct effect of chitosan on the B. cinerea paralyzing the growth of the fungus (without causing its death, however) and indirectly with the increase in the activity of enzymes responsible for making the plant cell wall stronger – peroxidase and phenylalanine ammonia-lyase (Camili et al, 2007; Reglinski et al, 2010). The action of biocontrol agents such as Trichoderma harzianum e Bacillus subtilis in the control of gray rot has also been observed, including making it possible to reduce the use of chemical fungicides to half the dose (Esterio et al, 2000; Harman et al, 1996). Several other reports of the effect of chitosan, Trichoderma spp. And B. subtilis are also cited by Elmer & Reglinski (2006).

Currently, work is being carried out at the IFRS/Campus Bento Gonçalves Plant Health Laboratory to adapt technologies already researched elsewhere in Brazil and around the world. In the 2011/2012 harvest, tests were carried out with chitosan (bioactive substance, 1,5L/ha, spray volume 300L/ha, commercial product at a concentration of 3%), potassium phosphite 40-20 (foliar fertilizer, 0,6L/ ha, spray volume 300L/ha), Trichoderma sp. (biocontrol fungus, 2L/ha, spray volume 300L/ha, commercial product with a concentration of 10⁸ CFU/ml) and Bacillus amyloliquefaciens (biocontrol bacteria, dose 2L/ha, spray volume 300L/ha, concentration of 3,99 x 10⁷ cells/ml). These alternatives were tested on plants of the Chardonnay and Tempranillo cultivars, grown in an espalier system. The applications were made to replace the recommended chemical treatments (control), in the pre-flowering, flowering and bunch maturation periods. As a control, in the area with Chardonnay, six applications were made using the fungicides pyrimethanil (concentration 300g a.i./L) and mancozeb (concentration 800g a.i./kg) and in the area with Tempranillo, four applications were made using the fungicides methyl thiophanate (concentration 700g a.i./L). kg), mancozeb (concentration 800g a.i./kg) and pyrimethanil (concentration 300g a.i./L), according to the manufacturers' recommendations.

In the area with the Chardonnay cultivar, it can be observed that treatments with chitosan, Trichoderma sp. and phosphite provided greater productivity per bunch than the treatment with B. amyloliquefaciens and the control, not differing from each other, with emphasis on the better performance of the portion treated with chitosan (Table 1). In relation to the incidence of gray rot (percentage of diseased plants or parts of diseased plants in a group) and severity of the disease (percentage of the area or volume of tissue covered by symptoms of the disease) it is clear that the evaluated treatments were as effective in the control of gray rot as chemical fungicides applied in the control plot, with the exception of the plot treated with phosphite, in which the worst performance was observed in both aspects. In the area with the Tempranillo cultivar (Table 2), there was a greater occurrence of gray rot in general in all treatments. However, unlike the area with Chardonnay, treatments with B. amyloliquefaciens on Trichoderma sp. showed a lower incidence of gray mold than the control treated with chemical fungicides, and when assessing severity, treatment with B. amyloliquefaciens showed better performance than the control.

It is important to highlight that so far the results obtained are partial and from just one harvest, however, based on the results obtained, it is considered that alternative products based on Trichoderma sp., chitosan and Bacillus amyloliquefaciens have potential to control gray rot in the field, as they showed efficiency equal to or greater than the chemical control used. More studies should be carried out in other harvests with different climatic conditions and to verify the effect of these alternatives on the quality of the wines produced. In no way is it recommended to use these products to control B. cinerea on vine.

Just for information and not as a recommendation for producers, it is interesting to know that in Brazil there are already organic products based on Bacillus subtilis e Bacillus spumilus registered for control of B. cinerea in strawberries and apples (Agrofit, 2012).

Figure 1 - Symptom of B. cinerea in Cabernet Sauvignon cultivar leaf. Bento Gonçlaves (RS) 2012

Figure 2 - Symptom of B. cinerea in the initial phase of bunch formation in Moscato cultivar. (A) Symptom on the complete side of the bunch and (B) symptom only on the lower tip of the bunch. Bento Gonçalves (RS), 2012

Figure 3 - Signs of B. cinerea in (A) bunch of grapes from the cultivar Itália in the final stage of ripening and (B) bunch of grapes from the cultivar Cabernet Sauvignon with the appearance of the disease in the place of greatest compaction of berries. Bento Gonçalves (RS), 2012.

Table 1 - Assessment of productivity per bunch, incidence and severity of gray rot in Chardonnay grapes. Bento Gonçalves (RS), 2012

Treatment	Productivity (Kg/bunch)	Incidence	Severity
chitosan	0,382 a*	29,18b*	7,65b*
Trichoderma sp.	0,304 ab	40,95 ab	11,44 ab
Phosphite 40-20	0,333 ab	56,58 to	16,92 to
Bacillus amyloliquefaciens	0,259 b	38,6 ab	10,7 ab
Witness	0,274 b	27,61 b	7,47 b
CV (%)	22,93	40,13	41,66

Treatment

Productivity (Kg/bunch)

Incidence

Severity

chitosan

0,382 a*

29,18b*

7,65b*

Trichoderma sp.

0,304 ab

40,95 ab

11,44 ab

Phosphite 40-20

0,333 ab

56,58 to

16,92 to

Bacillus amyloliquefaciens

0,259 b

38,6 ab

10,7 ab

Witness

0,274 b

27,61 b

7,47 b

CV (%)

22,93

40,13

41,66

*Averages followed by the same letter in the column do not differ statistically from each other at the 5% level of significance using the Duncan test.

Tabela 2 - Assessment of incidence and severity of gray rot in grapes of the Tempranillo cultivar. Bento Gonçalves (RS), 2012

Treatment	Incidence	Severity
chitosan	95,53 a*	28,79 ab*
Trichoderma sp.	82,35 b	23,04 bc
Phosphite 40-20	92,48 ab	31,81 to
Bacillus amyloliquefaciens	65,58 c	17,86 c
Witness	96,25 to	27,90 ab
CV (%)	9,24	18,59

Treatment

Incidence

Severity

chitosan

95,53 a*

28,79 ab*

Trichoderma sp.

82,35 b

23,04 bc

Phosphite 40-20

92,48 ab

31,81 to

Bacillus amyloliquefaciens

65,58 c

17,86 c

Witness

96,25 to

27,90 ab

CV (%)

9,24

18,59

*Averages followed by the same letter in the column do not differ statistically from each other at the 5% level of significance using the Duncan test.