- Email: [email protected]
Increase in Cladosporium spp. populations and rot of wine grapes associated with leaf removal
Crop Protection 30 (2011) 52e56
Contents lists available at ScienceDirect
Crop Protection journal homepage: www.elsevier.com/locate/cropro
Increase in Cladosporium spp. populations and rot of wine grapes associated with leaf removal B.A. Latorre*, E.X. Briceño, R. Torres Facultad de Agronomía e Ingeniería Forestal, Pontificia Universidad Católica de Chile, Casilla 306-22, Santiago, Chile
a r t i c l e i n f o
a b s t r a c t
Article history: Received 2 March 2010 Received in revised form 25 August 2010 Accepted 27 August 2010
Leaf removal reduces the epiphytic populations of several filamentous fungi found on grapevine (Vitis vinifera). Consequently this practice is used to prevent foliar diseases of grapevines and rots of grapes. In this study, the effects of leaf removal on Cladosporium rot (Cladosporium cladosporioides and Cladosporium herbarum), which often affects ‘Cabernet Sauvignon’ in Chile, were characterized. The effects of leaf removal on epiphytic populations of Cladosporium spp. on grape berry surfaces and on Cladosporium rot development were investigated. Three leaf removal treatments were compared: (i) severe leaf removal, where leaves from two to three nodes above, opposite and from all nodes below clusters were removed; (ii) mild leaf removal, where leaves opposite each cluster were removed; and (iii) no leaf removal. Regardless of the leaf removal treatment, low population levels of Cladosporium spp. were detected early in the ontogenic development of grape berries which increased as the berries matured, reaching maximum populations on overripe berries. Based on our results, severe leaf removal favors the growth of Cladosporium spp. on grape berries and increases the prevalence of Cladosporium rot at harvest. This increase in Cladosporium spp. was correlated with an increase in lenticel damage in ‘Cabernet Sauvignon’ and ‘Sauvignon blanc’ vines subjected to severe leaf removal. Considering that Cladosporium rot significantly reduces yield and wine quality, farmers should avoid continuous exposure of grape clusters to sunlight in order to prevent severe outbreaks of Cladosporium rot. Ó 2010 Elsevier Ltd. All rights reserved.
Keywords: Cladosporium Diseases Grapevine Pest management Vitis vinifera
1. Introduction Cladosporium rot, caused by Cladosporium cladosporioides and Cladosporium herbarum, is a common disease of grapevines (Vitis vinifera L.) in Chile, particularly in ‘Cabernet Sauvignon’ vines that are commonly harvested very late in the season, when grapes are partially senescent (Briceño and Latorre, 2007, 2008). This delay in harvest appears to be needed to obtain a complete phenolic ripeness of the berries to ensure aroma and flavor development for optimal wine quality (Saint-Cricq et al., 1998). However, a delay in harvest favors Cladosporium rot, which reduces yield and affects the quality of wines (Briceño et al., 2009; Pszczólkowski et al., 2001). Leaf removal has been demonstrated to be an effective canopy management strategy to reduce the incidence and severity of foliar diseases and rots of grapevines (Chellemi and Marois, 1992; Duncan et al., 1995; Gubler et al., 1987; Stapleton and Grant, 1992; Stapleton et al., 1995). Consequently, farmers normally remove leaves from the basal portion of the shoots, leaving clusters exposed to air flow and sunlight after bloom, a practice that affects the microclimate of
* Corresponding author. Tel.: þ562 585 4159; fax: þ562 5534130. E-mail address: [email protected] (B.A. Latorre). 0261-2194/$ e see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.cropro.2010.08.022
grapevines by increasing temperature and reducing relative humidity (English et al., 1989). Numerous fungal genera have been recovered from the surface of grape berries, including species of Aspergillus, Botrytis, Cladosporium, Penicillium, and Alternaria (Díaz et al., 2009; Donoso and Latorre, 2006; Thompson and Latorre, 1999). The populations of these species increase as the berry mature. This was demonstrated for a population of Cladosporium spp. that developed on the surface of apparently healthy grape berries (Briceño and Latorre, 2008). This study was conducted to determine the effect of leaf removal on the epiphytic populations of Cladosporium spp. and on the development of Cladosporium rot on wine grape clusters.
2. Material and methods 2.1. Plant material Leaf removal experiments were conducted in commercial ‘Cabernet Sauvignon’ and ‘Sauvignon blanc’ vineyards, in Alto Jahuel (33 430 6000 S, 70 420 000 W) and ‘Cabernet Sauvignon’ in Alhué (33 2100000 S, 71080 0000 W). Both localities are characterized by a Mediterranean climate, with rains concentrated in winter months
B.A. Latorre et al. / Crop Protection 30 (2011) 52e56
53
Fig. 1. Grapevine (Vitis vinifera) ‘Cabernet Sauvignon’ subjected to a non-defoliated vine treatment (A) and a severe leaf removal treatment (B). Cladosporium rot (C) and lenticel damage (D) developed on ‘Sauvignon blanc’ grape berries in vines subjected to a severe leaf removal treatment.
and warm summer months. The ‘Cabernet Sauvignon’ and ‘Sauvignon blanc’ vines were 7 and 15 year-old, respectively, and planted on their own roots at 2.5 1.2 m, with rows oriented north to south and trained in a bi-lateral cordon trellis system with three wires at 0.9, 1.2 and 1.5 cm above ground. In all experiments, grapevines were managed as is customary for wine grapes in Chile (Gil and Pszczólkowski, 2007), except that foliar fungicides were not applied.
leaves were removed manually on the east and west side of the vines. However, evaluations were made separately on each vine side, considering that light interception on the east side of vines oriented north to south varies between 8 and 10 h, in contrast to 12e19 h on the west side (Gil and Pszczólkowski, 2007; Smart, 1973).
2.2. Leaf removal
The effect of leaf removal on the epiphytic populations of Cladosporium spp. was monitored from the date of leaf removal (fruit set) through harvest on grapevines ‘Cabernet Sauvignon’, subjected to a severe leaf removal treatment in Alto Jahuel in 2007 and 2008 and Alhué in 2008. For each sample, 50 berries were randomly selected, suspended and shaken for 5 min in 50 ml of sterile 0.05% Tween-80 for 5 min. An aliquot (100 mL) of each suspension was sprayed in a 90 mm diameter Petri dish containing acidified potato dextrose agar (2% dehydrated mashed potatoes, 2% glucose and 20 g L1 agar plus 0.5 ml L1 92% lactic acid added after autoclaving)
The effect of three leaf removal treatments on Cladosporium rot and lenticel damage were studied: (i) severe leaf removal, consisting of removing leaves from two to three nodes above, opposite each cluster, and leaves from all nodes below clusters, leaving them exposed to sunlight continuously from fruit set to harvest. (ii) Mild leaf removal, consisting of removing leaves opposite each cluster about 4e5 weeks after fruit set, and (iii) no leaf removal, where nondefoliated plants were left as controls (Fig. 1). In all experiments,
2.3. Effect of leaf removal on the dynamic of Cladosporium ssp.
54
B.A. Latorre et al. / Crop Protection 30 (2011) 52e56
Fig. 2. Effect of a severe leaf removal on the epiphytic populations of Cladosporium spp. on grape berries of V. vinifera ‘Cabernet Sauvignon’. (A and B) Alto Jahuel 2007 and 2008, respectively. (C) Alhué, 2008. Veraison (berry beginning to lose green color) was approximately 63 and 70 days after full bloom (DAFB) in Alto Jahuel in 2007 and 2008, respectively, and 73 DAFB in Alhué in 2008.
that was supplemented (MPDA) per liter with 0.05 g tetracycline (SigmaeAldrich), and 0.1 g streptomycin (SigmaeAldrich). One milliliter of Igepal CO-630 (SigmaeAldrich, Atlanta, USA) was added as a colony growth restrictor. Plates were incubated at 20 C for 6 days to determine the number of colonies of Cladosporium spp. on each disk.
Cladosporium rot (y) obtained in ‘Cabernet Sauvignon’ clusters was studied by regression analysis.
2.4. Effect of leaf removal on Cladosporium rot and lenticel damage
Independent of location, year and side of the vine, the population dynamics of Cladosporium spp. on the surface of apparently healthy grapes exhibited a similar trend that was characterized by a relatively low density early in the season, increasing rapidly after veraison (when a berry begins to lose its green color), with maximum values at harvest. For instance, in grape clusters subjected to a severe leaf removal treatment, population densities increased 11- to 34-fold in Alto Jahuel and 132- to 346-fold in Alhué. In contrast, population densities increased 3- to 10-fold and 2.6- to 86-fold on berries from vines that maintained their canopies unaltered in Alto Jahuel and Alhué, respectively. An exponential relationship best described the relation between population densities and DAFB (Fig. 2). The mean population densities of Cladosporium spp. were significantly (P < 0.012) higher on berries exposed to sunlight on the west side relative to those exposed on the east side of the vines in Alto Jahuel. However, the side of the vine had no effect on the population densities of Cladosporium spp. obtained in Alhué (Table 1). Leaf removal treatments had a significant (P < 0.001) effect on populations of Cladosporium spp. in Alto Jahuel and Alhué in 2008, but the effect was non-significant in Alto Jahuel in 2007. The interaction between leaf removal treatments and side of the vines were significant in Alto Jahuel (P ¼ 0.002) in 2008 and Alhué 2008 (P ¼ 0.001) (Table 1).
At harvest, the prevalence of Cladosporium rot was evaluated in 25 berry samples collected arbitrarily from five grape clusters from each replicate in each trial. At the same time, the severity of the lenticel damage was evaluated in each berry by counting the necrotic lenticels under a stereoscopic microscope. 2.5. Experimental design and statistical analysis Treatments were arranged as randomized complete blocks (blocked within rows) with a 3 2 (leaf removal treatments grapevine sides) factorial arrangement of treatments, with four replicates of at least 10 vines each. Data were subjected to a twoway analysis of variance, and means were analyzed separately using Tukey’s multiple comparison test (P < 0.05), using SigmaStat 3.1 (Systat Software Inc., San José, CA, USA). If needed, prior to analysis, data were normalized by log10(x þ 1) transformation; however, non-transformed data are presented. The effect of leaf removal on the epiphytic populations of Cladosporium spp. which developed on ‘Cabernet Sauvignon’ clusters was described by regression analysis between x ¼ days after full bloom (DAFB) and y ¼ Cladosporium spp. populations. Similarly, the relationship between lenticel damage (x) and incidence of
3. Results 3.1. Effect of leaf removal on the dynamic of Cladosporium ssp.
B.A. Latorre et al. / Crop Protection 30 (2011) 52e56 Table 1 Effect of leaf removal on epiphytic populations of Cladosporium spp. on grape (Vitis vinifera) berries ‘Cabernet Sauvignon’. Treatments
Population of Cladosporium spp. on the surface of apparently healthy berries, cfu cm2 Alto Jahuel
Alhué
2007
2008
2008
Leaf removal levels (LRLs)a Severe 2758.8 Mild 2503.1 None 1976.7 df 2 F 1.27 P 0.304 SED 1.151
5337.3 5431.8 2202.4 3 10.23 0.001 0.349
6149.2 3317.1 3229.9 3 40.37 <0.001 0.228
Grapevine side (GS)a East West df F P SED
1588.1a 3237.5b 1 18.80 <0.001 0.329
2985.2 5662.3 1 7.71 0.012 0.315
4334.5 4128.3 1 2.22 0.154 0.206
LRL GS interaction df F P SED
2 0.35 0.706 0.433
2 8.62 0.002 0.415
2 12.59 0.001 0.037
55
a moldy appearance to the berries that only affected the berries superficially (Fig. 1C). Leaf removal treatments had a significant effect on the incidence of Cladosporium rot in ‘Sauvignon blanc’ (P < 0.001), but the effect was not significant on ‘Cabernet Sauvignon’ (P ¼ 0.301). However, the grapevine side of the vines significantly (P < 0.043) affected the level of Cladosporium rot in both grapevine cultivars. The interaction between leaf removal treatment and grapevine side was only significant (P ¼ 0.035) in ‘Cabernet Sauvignon’ (Table 2). Leaf removal significantly increased Cladosporium rot in ‘Sauvignon blanc’, with a mean of 61.6 diseased berries per cluster; while only four and six diseased berries per cluster were obtained in vines subjected to mild leaf removal and no leaf removal, respectively (Table 2). In ‘Cabernet Sauvignon’, only mean differences in leaf removal treatments on the west side of the plants were significant (P ¼ 0.05). 3.3. Effect of leaf removal on lenticel damage
a Means followed by the same letter in each column did not differ significantly according to Tukey’s test (P ¼ 0.05). Data were log10(x þ 1) transformed before analysis, but non-transformed data are presented.
Leaf removal treatments increased the population density of Cladosporium spp. by 40e175% in severely defoliated vines relative to non-defoliated vines. However, differences between populations found in berries from unaltered canopies and the populations in berries from severe removal treatments were significant (P ¼ 0.05) only for Alto Jahuel and Alhué in 2008. 3.2. Effect of leaf removal on Cladosporium rot Cladosporium rot appeared on mature grapes (total soluble solids [TSS] > 22%), and was characterized by the presence of superficial olive-green colonies of Cladosporium spp. that confer
Lenticel damage was characterized by the development of necrotic spherical dots that were brown and about 1.0e2.0 mm in diameter, often surrounded by a reddish halo, which were apparent before veraison (Fig. 1D). Independent of the grapevine cultivar, leaf removal had a significant (P < 0.012) effect on the severity of lenticel damage, significantly increasing in vines subjected to a severe leaf removal (Table 2). The severity of lenticel damage was not affected by the side of the vines where grape cluster were hanging, and the interaction between leaf removal and the side of the vines was not significant (Table 2). An exponential model, y ¼ 0.0022e0.2303x, R2 ¼ 0.77, best explained the relation between x ¼ lenticel damage (x) and y ¼ Cladosporium rot (Fig. 3). 4. Discussion This study demonstrates that a high level of leaf removal increases epiphytic populations of Cladosporium spp. on the surface of wine grape berries and increases the prevalence of Cladosporium rot at harvest. Based on the negative effect of leaf removal on epiphytic populations of Botrytis cinerea and other grape pathogens (Duncan et al., 1995), mainly caused by increasing air movement
Table 2 Analysis of variance showing the effect of leaf removal on the development of Cladosporium rot and lenticel damage of grapes (V. vinifera). Treatments
Cabernet Sauvignon
Sauvignon blanc Cladosporium rot n cluster1
Lenticel damage n berry1
12.0b 9.9a 10.1a 2 5.71 0.012 4.953
61.6b 4.0a 6.0a 2 15.59 <0.001 0.587
24.2b 16.8a 18.3a 2 43.01 <0.001 5.473
0.9 2.5 1 4.72 0.043 0.397
10.5 10.8 1 0.18 0.681 4.475
15.1a 32.6b 1 5.62 0.029 0.529
19.9 19.6 1 0.21 0.649 4.946
2 4.03 0.035 0.523
2 0.12 0.222 5.890
2 3.82 0.894 0.696
2 0.13 0.378 2.089
Cladosporium rot n cluster
1
Lenticel damage n berry
1
a
Leaf removal levels (LRLs) Severe 2.2 Mild 1.5 None 1.4 df 2 F 1.28 P 0.301 SED 0.440 Grapevine side (GS)a East West df F P SED LRL GS interaction df F P SED
a Means followed by the same letter in each column were not significantly different according to Tukey’s test (P ¼ 0.05). Data were log10(x þ 1) transformed before analysis, but non-transformed data are presented.
56
B.A. Latorre et al. / Crop Protection 30 (2011) 52e56
Cladosporium spp. were 66.1 and 233.6 times higher than Penicillium spp. in unaltered ‘Cabernet Sauvignon’, 70 and 152 DAFB in Alto Jahuel, respectively. The environmental conditions, characteristic of a Mediterranean climate, with low relative humidity and an absence of summer rains, appear to highly favor the survival and development of Cladosporium spp. in Chilean vineyards. In conclusion, leaf removal increases the epiphytic populations of Cladosporium spp. on grape berry surfaces and it increases Cladosporium rot on ‘Cabernet Sauvignon’ and ‘Sauvignon blanc’. Therefore, leaf removal should be used cautiously avoiding over exposure of grape clusters to sun.
Cladosporium rot, no. cluster
-1
8
6
4
y = 0.022e0.2303x R2 = 0.77
2
Acknowledgments
0 8
12
16
20
24
Lenticel damage,no.berry -1
We thank Viña Santa Rita and Viña Ventisquero for allowing us to use their plantations. This research project was supported by Consorcio Empresarial para la Vid y el Vino, Vinnova project 05CTE01-09.
Fig. 3. Relationship between lenticel damage and Cladosporium rot obtained in grapevine (V. vinifera) ‘Cabernet Sauvignon’ subjected to a severe leaf removal.
References
around grape berries (English et al., 1989), canopy management, including leaf removal, shoot topping and shoot positioning, has been recommended to preventively control Botrytis bunch rot and other grapevine diseases (Chellemi and Marois, 1992; Gubler et al., 1987; Stapleton and Grant, 1992; Stapleton et al., 1995). However, considering that Cladosporium rot significantly reduces yield and wine quality (Briceño et al., 2009; Pszczólkowski et al., 2001), farmers should avoid continuous exposure of grape clusters to sunlight so as to prevent severe Cladosporium rot outbreaks. The incidence of Cladosporium rot was at least partially associated with an increase in lenticel damage observed in ‘Cabernet Sauvignon’ and ‘Sauvignon blanc’ vines subjected to a severe leaf removal. It is possible that the necrotic lesions that develop around lenticels might provide a substrate for the establishment and colonization of grape berries by Cladosporium spp. Additionally, Cladosporium spp. produce melanin-like pigments in the cell wall of conidiophores and conidia, which confer protection against solar radiation (Ulevicius et al., 2000; Valero et al., 2007). Therefore, increasing solar radiation of the grape clusters through leaf removal may not affect their survival, which may explain the high densities of Cladosporium spp. obtained on grape clusters subjected to severe leaf removal in this study. Regardless of the leaf removal treatment, our results confirm the presence of epiphytic populations of Cladosporium spp. on apparently healthy grape clusters. These populations were detected very early in the ontogenic development of grape berries and increased as the berries matured, reaching maximum numbers on overripe berries (Briceño and Latorre, 2008). As has been suggested for other pathogens affecting grape berries (Commenil et al., 1997; Jeandet et al., 1991; Mlikota Gabler et al., 2003), it is postulated that changes in cuticle porosity, cuticle fractures, lower capacity for developing defense reactions, and the decrease of epicuticular wax content in ripe to overripe grapes may promote the presence of sugary and watery exudates that stimulate the superficial development of Cladosporium spp. in overripe berries. However, further studies are needed to better characterize the dynamics of the epiphytic populations of Cladosporium on grape berries. The populations of Cladosporium spp. were always higher than the populations of other filamentous fungi, such as species of Alternaria, Aspergillus, Botrytis, Penicillium and Rhizopus (results not reported in this article). For instance, population densities of
Briceño, E.X., Latorre, B.A., 2007. Outbreaks of Cladosporium rot associated with delayed harvest wine grapes in Chile. Plant Dis. 91, 1060. Briceño, E.X., Latorre, B.A., 2008. Characterization of Cladosporium rot in grapevines, a problem of growing importance in Chile. Plant Dis. 92, 1635e1642. Briceño, E.X., Latorre, B.A., Bordeu, E., 2009. Effect of Cladosporium rot on the composition and aromatic compounds of red wine. Span. J. Agric. Res. 7, 119e128. Chellemi, D.O., Marois, J.J., 1992. Influence of leaf removal, fungicide applications, and fruit maturity on incidence and severity of grape powdery mildew. Am. J. Enol. Vitic. 43, 53e57. Commenil, P., Brunet, L., Audran, J.C., 1997. The development of the grape berry cuticle in relation to susceptibility to bunch rot disease. J. Exp. Bot. 48, 1599e1607. Díaz, G.A., Torres, R., Vega, M., Latorre, B.A., 2009. Ochratoxigenic Aspergillus species on grapes from Chilean vineyards and Aspergillus threshold levels on grapes. Int. J. Food Microbiol. 133, 195e199. Donoso, A., Latorre, B.A., 2006. Characterization of blue mold caused by Penicillium spp. in cold stored table grapes. Cien. Inv. Agric. 33, 119e130. Duncan, R.A., Stapleton, J.J., Leavitt, G.M., 1995. Population dynamics of epiphytic mycoflora and occurrence of bunch rots of wine grapes as influenced by leaf removal. Plant Pathol. 44, 956e965. English, J.T., Thomas, C.S., Marois, J.J., Gubler, W.D., 1989. Microclimates of grapevine canopies associated with leaf removal and control of Botrytis bunch rot. Phytopathology 79, 395e401. Gil, G.F., Pszczólkowski, P., 2007. Viticultura, Fundamentos para Optimizar Producción y Calidad. Ediciones Universidad Católica, Santiago, Chile, 535 pp. Gubler, W.D., Marois, J.J., Bledsoe, A.M., Bettiga, U., 1987. Control of Botrytis bunch rot of grape with canopy management. Plant Dis. 71, 599e601. Jeandet, P., Bessis, R., Gautheron, B., 1991. The production of resveratrol (3,5,4-trihydroxystilbene) by grape berries in different developmental stages. Am. J. Enol. Vitic. 42, 41e46. Mlikota Gabler, F., Smilanick, J.L., Mansour, M., Ramming, D.W., Mackey, B.E., 2003. Correlations of morphological, anatomical, and chemical features of grape berries with resistance to Botrytis cinerea. Phytopathology 93, 1263e1273. Pszczólkowski, P., Latorre, B.A., Ceppi di Lecco, C., 2001. Efecto de los mohos presentes en uvas cosechadas tardíamente sobre la calidad de los mostos y vinos Cabernet Sauvignon. Cien. Inv. Agric 28, 157e163. Saint-Cricq, N., Vivas, N., Glories, Y., 1998. Maturité phénolique: définition et contrôle. Rev. Fr. OEnol. 98, 22e25. Smart, R.E., 1973. Sunlight interception by vineyards. Am. J. Enol. Vitic. 24, 141e147. Stapleton, J.J., Grant, R.S., 1992. Leaf removal for nonchemical control of the summer bunch rot complex of wine grapes in the San Joaquin Valley. Plant Dis. 76, 205e208. Stapleton, J.J., Leavitt, G.M., Verdegaal, P.S., 1995. Integration of leaf removal and reduced fungicide applications for management of grape powdery mildew in the San Joaquin Valley. Calif. Agric. 49, 33e36. Thompson, J.R., Latorre, B.A., 1999. Characterization of Botrytis cinerea from table grapes in Chile using RAPD-PCR. Plant Dis. 83, 1090e1094. Ulevicius, V., Peciulyte, D., Mordas, G., Lugauskas, A., 2000. Field study on changes in viability of airborne fungal propagules exposed to solar radiation. J. Aerosol Sci. 31, S961eS962. Valero, A., Begum, M., Leong, S.L., Hocking, A.D., Ramos, A.J., Sanchis, V., Maırín, S., 2007. Effect of germicidal UVC light on fungi isolated from grapes and raisins. Lett. Appl. Microbiol. 45, 238e243.
Contents lists available at ScienceDirect
Crop Protection journal homepage: www.elsevier.com/locate/cropro
Increase in Cladosporium spp. populations and rot of wine grapes associated with leaf removal B.A. Latorre*, E.X. Briceño, R. Torres Facultad de Agronomía e Ingeniería Forestal, Pontificia Universidad Católica de Chile, Casilla 306-22, Santiago, Chile
a r t i c l e i n f o
a b s t r a c t
Article history: Received 2 March 2010 Received in revised form 25 August 2010 Accepted 27 August 2010
Leaf removal reduces the epiphytic populations of several filamentous fungi found on grapevine (Vitis vinifera). Consequently this practice is used to prevent foliar diseases of grapevines and rots of grapes. In this study, the effects of leaf removal on Cladosporium rot (Cladosporium cladosporioides and Cladosporium herbarum), which often affects ‘Cabernet Sauvignon’ in Chile, were characterized. The effects of leaf removal on epiphytic populations of Cladosporium spp. on grape berry surfaces and on Cladosporium rot development were investigated. Three leaf removal treatments were compared: (i) severe leaf removal, where leaves from two to three nodes above, opposite and from all nodes below clusters were removed; (ii) mild leaf removal, where leaves opposite each cluster were removed; and (iii) no leaf removal. Regardless of the leaf removal treatment, low population levels of Cladosporium spp. were detected early in the ontogenic development of grape berries which increased as the berries matured, reaching maximum populations on overripe berries. Based on our results, severe leaf removal favors the growth of Cladosporium spp. on grape berries and increases the prevalence of Cladosporium rot at harvest. This increase in Cladosporium spp. was correlated with an increase in lenticel damage in ‘Cabernet Sauvignon’ and ‘Sauvignon blanc’ vines subjected to severe leaf removal. Considering that Cladosporium rot significantly reduces yield and wine quality, farmers should avoid continuous exposure of grape clusters to sunlight in order to prevent severe outbreaks of Cladosporium rot. Ó 2010 Elsevier Ltd. All rights reserved.
Keywords: Cladosporium Diseases Grapevine Pest management Vitis vinifera
1. Introduction Cladosporium rot, caused by Cladosporium cladosporioides and Cladosporium herbarum, is a common disease of grapevines (Vitis vinifera L.) in Chile, particularly in ‘Cabernet Sauvignon’ vines that are commonly harvested very late in the season, when grapes are partially senescent (Briceño and Latorre, 2007, 2008). This delay in harvest appears to be needed to obtain a complete phenolic ripeness of the berries to ensure aroma and flavor development for optimal wine quality (Saint-Cricq et al., 1998). However, a delay in harvest favors Cladosporium rot, which reduces yield and affects the quality of wines (Briceño et al., 2009; Pszczólkowski et al., 2001). Leaf removal has been demonstrated to be an effective canopy management strategy to reduce the incidence and severity of foliar diseases and rots of grapevines (Chellemi and Marois, 1992; Duncan et al., 1995; Gubler et al., 1987; Stapleton and Grant, 1992; Stapleton et al., 1995). Consequently, farmers normally remove leaves from the basal portion of the shoots, leaving clusters exposed to air flow and sunlight after bloom, a practice that affects the microclimate of
* Corresponding author. Tel.: þ562 585 4159; fax: þ562 5534130. E-mail address: [email protected] (B.A. Latorre). 0261-2194/$ e see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.cropro.2010.08.022
grapevines by increasing temperature and reducing relative humidity (English et al., 1989). Numerous fungal genera have been recovered from the surface of grape berries, including species of Aspergillus, Botrytis, Cladosporium, Penicillium, and Alternaria (Díaz et al., 2009; Donoso and Latorre, 2006; Thompson and Latorre, 1999). The populations of these species increase as the berry mature. This was demonstrated for a population of Cladosporium spp. that developed on the surface of apparently healthy grape berries (Briceño and Latorre, 2008). This study was conducted to determine the effect of leaf removal on the epiphytic populations of Cladosporium spp. and on the development of Cladosporium rot on wine grape clusters.
2. Material and methods 2.1. Plant material Leaf removal experiments were conducted in commercial ‘Cabernet Sauvignon’ and ‘Sauvignon blanc’ vineyards, in Alto Jahuel (33 430 6000 S, 70 420 000 W) and ‘Cabernet Sauvignon’ in Alhué (33 2100000 S, 71080 0000 W). Both localities are characterized by a Mediterranean climate, with rains concentrated in winter months
B.A. Latorre et al. / Crop Protection 30 (2011) 52e56
53
Fig. 1. Grapevine (Vitis vinifera) ‘Cabernet Sauvignon’ subjected to a non-defoliated vine treatment (A) and a severe leaf removal treatment (B). Cladosporium rot (C) and lenticel damage (D) developed on ‘Sauvignon blanc’ grape berries in vines subjected to a severe leaf removal treatment.
and warm summer months. The ‘Cabernet Sauvignon’ and ‘Sauvignon blanc’ vines were 7 and 15 year-old, respectively, and planted on their own roots at 2.5 1.2 m, with rows oriented north to south and trained in a bi-lateral cordon trellis system with three wires at 0.9, 1.2 and 1.5 cm above ground. In all experiments, grapevines were managed as is customary for wine grapes in Chile (Gil and Pszczólkowski, 2007), except that foliar fungicides were not applied.
leaves were removed manually on the east and west side of the vines. However, evaluations were made separately on each vine side, considering that light interception on the east side of vines oriented north to south varies between 8 and 10 h, in contrast to 12e19 h on the west side (Gil and Pszczólkowski, 2007; Smart, 1973).
2.2. Leaf removal
The effect of leaf removal on the epiphytic populations of Cladosporium spp. was monitored from the date of leaf removal (fruit set) through harvest on grapevines ‘Cabernet Sauvignon’, subjected to a severe leaf removal treatment in Alto Jahuel in 2007 and 2008 and Alhué in 2008. For each sample, 50 berries were randomly selected, suspended and shaken for 5 min in 50 ml of sterile 0.05% Tween-80 for 5 min. An aliquot (100 mL) of each suspension was sprayed in a 90 mm diameter Petri dish containing acidified potato dextrose agar (2% dehydrated mashed potatoes, 2% glucose and 20 g L1 agar plus 0.5 ml L1 92% lactic acid added after autoclaving)
The effect of three leaf removal treatments on Cladosporium rot and lenticel damage were studied: (i) severe leaf removal, consisting of removing leaves from two to three nodes above, opposite each cluster, and leaves from all nodes below clusters, leaving them exposed to sunlight continuously from fruit set to harvest. (ii) Mild leaf removal, consisting of removing leaves opposite each cluster about 4e5 weeks after fruit set, and (iii) no leaf removal, where nondefoliated plants were left as controls (Fig. 1). In all experiments,
2.3. Effect of leaf removal on the dynamic of Cladosporium ssp.
54
B.A. Latorre et al. / Crop Protection 30 (2011) 52e56
Fig. 2. Effect of a severe leaf removal on the epiphytic populations of Cladosporium spp. on grape berries of V. vinifera ‘Cabernet Sauvignon’. (A and B) Alto Jahuel 2007 and 2008, respectively. (C) Alhué, 2008. Veraison (berry beginning to lose green color) was approximately 63 and 70 days after full bloom (DAFB) in Alto Jahuel in 2007 and 2008, respectively, and 73 DAFB in Alhué in 2008.
that was supplemented (MPDA) per liter with 0.05 g tetracycline (SigmaeAldrich), and 0.1 g streptomycin (SigmaeAldrich). One milliliter of Igepal CO-630 (SigmaeAldrich, Atlanta, USA) was added as a colony growth restrictor. Plates were incubated at 20 C for 6 days to determine the number of colonies of Cladosporium spp. on each disk.
Cladosporium rot (y) obtained in ‘Cabernet Sauvignon’ clusters was studied by regression analysis.
2.4. Effect of leaf removal on Cladosporium rot and lenticel damage
Independent of location, year and side of the vine, the population dynamics of Cladosporium spp. on the surface of apparently healthy grapes exhibited a similar trend that was characterized by a relatively low density early in the season, increasing rapidly after veraison (when a berry begins to lose its green color), with maximum values at harvest. For instance, in grape clusters subjected to a severe leaf removal treatment, population densities increased 11- to 34-fold in Alto Jahuel and 132- to 346-fold in Alhué. In contrast, population densities increased 3- to 10-fold and 2.6- to 86-fold on berries from vines that maintained their canopies unaltered in Alto Jahuel and Alhué, respectively. An exponential relationship best described the relation between population densities and DAFB (Fig. 2). The mean population densities of Cladosporium spp. were significantly (P < 0.012) higher on berries exposed to sunlight on the west side relative to those exposed on the east side of the vines in Alto Jahuel. However, the side of the vine had no effect on the population densities of Cladosporium spp. obtained in Alhué (Table 1). Leaf removal treatments had a significant (P < 0.001) effect on populations of Cladosporium spp. in Alto Jahuel and Alhué in 2008, but the effect was non-significant in Alto Jahuel in 2007. The interaction between leaf removal treatments and side of the vines were significant in Alto Jahuel (P ¼ 0.002) in 2008 and Alhué 2008 (P ¼ 0.001) (Table 1).
At harvest, the prevalence of Cladosporium rot was evaluated in 25 berry samples collected arbitrarily from five grape clusters from each replicate in each trial. At the same time, the severity of the lenticel damage was evaluated in each berry by counting the necrotic lenticels under a stereoscopic microscope. 2.5. Experimental design and statistical analysis Treatments were arranged as randomized complete blocks (blocked within rows) with a 3 2 (leaf removal treatments grapevine sides) factorial arrangement of treatments, with four replicates of at least 10 vines each. Data were subjected to a twoway analysis of variance, and means were analyzed separately using Tukey’s multiple comparison test (P < 0.05), using SigmaStat 3.1 (Systat Software Inc., San José, CA, USA). If needed, prior to analysis, data were normalized by log10(x þ 1) transformation; however, non-transformed data are presented. The effect of leaf removal on the epiphytic populations of Cladosporium spp. which developed on ‘Cabernet Sauvignon’ clusters was described by regression analysis between x ¼ days after full bloom (DAFB) and y ¼ Cladosporium spp. populations. Similarly, the relationship between lenticel damage (x) and incidence of
3. Results 3.1. Effect of leaf removal on the dynamic of Cladosporium ssp.
B.A. Latorre et al. / Crop Protection 30 (2011) 52e56 Table 1 Effect of leaf removal on epiphytic populations of Cladosporium spp. on grape (Vitis vinifera) berries ‘Cabernet Sauvignon’. Treatments
Population of Cladosporium spp. on the surface of apparently healthy berries, cfu cm2 Alto Jahuel
Alhué
2007
2008
2008
Leaf removal levels (LRLs)a Severe 2758.8 Mild 2503.1 None 1976.7 df 2 F 1.27 P 0.304 SED 1.151
5337.3 5431.8 2202.4 3 10.23 0.001 0.349
6149.2 3317.1 3229.9 3 40.37 <0.001 0.228
Grapevine side (GS)a East West df F P SED
1588.1a 3237.5b 1 18.80 <0.001 0.329
2985.2 5662.3 1 7.71 0.012 0.315
4334.5 4128.3 1 2.22 0.154 0.206
LRL GS interaction df F P SED
2 0.35 0.706 0.433
2 8.62 0.002 0.415
2 12.59 0.001 0.037
55
a moldy appearance to the berries that only affected the berries superficially (Fig. 1C). Leaf removal treatments had a significant effect on the incidence of Cladosporium rot in ‘Sauvignon blanc’ (P < 0.001), but the effect was not significant on ‘Cabernet Sauvignon’ (P ¼ 0.301). However, the grapevine side of the vines significantly (P < 0.043) affected the level of Cladosporium rot in both grapevine cultivars. The interaction between leaf removal treatment and grapevine side was only significant (P ¼ 0.035) in ‘Cabernet Sauvignon’ (Table 2). Leaf removal significantly increased Cladosporium rot in ‘Sauvignon blanc’, with a mean of 61.6 diseased berries per cluster; while only four and six diseased berries per cluster were obtained in vines subjected to mild leaf removal and no leaf removal, respectively (Table 2). In ‘Cabernet Sauvignon’, only mean differences in leaf removal treatments on the west side of the plants were significant (P ¼ 0.05). 3.3. Effect of leaf removal on lenticel damage
a Means followed by the same letter in each column did not differ significantly according to Tukey’s test (P ¼ 0.05). Data were log10(x þ 1) transformed before analysis, but non-transformed data are presented.
Leaf removal treatments increased the population density of Cladosporium spp. by 40e175% in severely defoliated vines relative to non-defoliated vines. However, differences between populations found in berries from unaltered canopies and the populations in berries from severe removal treatments were significant (P ¼ 0.05) only for Alto Jahuel and Alhué in 2008. 3.2. Effect of leaf removal on Cladosporium rot Cladosporium rot appeared on mature grapes (total soluble solids [TSS] > 22%), and was characterized by the presence of superficial olive-green colonies of Cladosporium spp. that confer
Lenticel damage was characterized by the development of necrotic spherical dots that were brown and about 1.0e2.0 mm in diameter, often surrounded by a reddish halo, which were apparent before veraison (Fig. 1D). Independent of the grapevine cultivar, leaf removal had a significant (P < 0.012) effect on the severity of lenticel damage, significantly increasing in vines subjected to a severe leaf removal (Table 2). The severity of lenticel damage was not affected by the side of the vines where grape cluster were hanging, and the interaction between leaf removal and the side of the vines was not significant (Table 2). An exponential model, y ¼ 0.0022e0.2303x, R2 ¼ 0.77, best explained the relation between x ¼ lenticel damage (x) and y ¼ Cladosporium rot (Fig. 3). 4. Discussion This study demonstrates that a high level of leaf removal increases epiphytic populations of Cladosporium spp. on the surface of wine grape berries and increases the prevalence of Cladosporium rot at harvest. Based on the negative effect of leaf removal on epiphytic populations of Botrytis cinerea and other grape pathogens (Duncan et al., 1995), mainly caused by increasing air movement
Table 2 Analysis of variance showing the effect of leaf removal on the development of Cladosporium rot and lenticel damage of grapes (V. vinifera). Treatments
Cabernet Sauvignon
Sauvignon blanc Cladosporium rot n cluster1
Lenticel damage n berry1
12.0b 9.9a 10.1a 2 5.71 0.012 4.953
61.6b 4.0a 6.0a 2 15.59 <0.001 0.587
24.2b 16.8a 18.3a 2 43.01 <0.001 5.473
0.9 2.5 1 4.72 0.043 0.397
10.5 10.8 1 0.18 0.681 4.475
15.1a 32.6b 1 5.62 0.029 0.529
19.9 19.6 1 0.21 0.649 4.946
2 4.03 0.035 0.523
2 0.12 0.222 5.890
2 3.82 0.894 0.696
2 0.13 0.378 2.089
Cladosporium rot n cluster
1
Lenticel damage n berry
1
a
Leaf removal levels (LRLs) Severe 2.2 Mild 1.5 None 1.4 df 2 F 1.28 P 0.301 SED 0.440 Grapevine side (GS)a East West df F P SED LRL GS interaction df F P SED
a Means followed by the same letter in each column were not significantly different according to Tukey’s test (P ¼ 0.05). Data were log10(x þ 1) transformed before analysis, but non-transformed data are presented.
56
B.A. Latorre et al. / Crop Protection 30 (2011) 52e56
Cladosporium spp. were 66.1 and 233.6 times higher than Penicillium spp. in unaltered ‘Cabernet Sauvignon’, 70 and 152 DAFB in Alto Jahuel, respectively. The environmental conditions, characteristic of a Mediterranean climate, with low relative humidity and an absence of summer rains, appear to highly favor the survival and development of Cladosporium spp. in Chilean vineyards. In conclusion, leaf removal increases the epiphytic populations of Cladosporium spp. on grape berry surfaces and it increases Cladosporium rot on ‘Cabernet Sauvignon’ and ‘Sauvignon blanc’. Therefore, leaf removal should be used cautiously avoiding over exposure of grape clusters to sun.
Cladosporium rot, no. cluster
-1
8
6
4
y = 0.022e0.2303x R2 = 0.77
2
Acknowledgments
0 8
12
16
20
24
Lenticel damage,no.berry -1
We thank Viña Santa Rita and Viña Ventisquero for allowing us to use their plantations. This research project was supported by Consorcio Empresarial para la Vid y el Vino, Vinnova project 05CTE01-09.
Fig. 3. Relationship between lenticel damage and Cladosporium rot obtained in grapevine (V. vinifera) ‘Cabernet Sauvignon’ subjected to a severe leaf removal.
References
around grape berries (English et al., 1989), canopy management, including leaf removal, shoot topping and shoot positioning, has been recommended to preventively control Botrytis bunch rot and other grapevine diseases (Chellemi and Marois, 1992; Gubler et al., 1987; Stapleton and Grant, 1992; Stapleton et al., 1995). However, considering that Cladosporium rot significantly reduces yield and wine quality (Briceño et al., 2009; Pszczólkowski et al., 2001), farmers should avoid continuous exposure of grape clusters to sunlight so as to prevent severe Cladosporium rot outbreaks. The incidence of Cladosporium rot was at least partially associated with an increase in lenticel damage observed in ‘Cabernet Sauvignon’ and ‘Sauvignon blanc’ vines subjected to a severe leaf removal. It is possible that the necrotic lesions that develop around lenticels might provide a substrate for the establishment and colonization of grape berries by Cladosporium spp. Additionally, Cladosporium spp. produce melanin-like pigments in the cell wall of conidiophores and conidia, which confer protection against solar radiation (Ulevicius et al., 2000; Valero et al., 2007). Therefore, increasing solar radiation of the grape clusters through leaf removal may not affect their survival, which may explain the high densities of Cladosporium spp. obtained on grape clusters subjected to severe leaf removal in this study. Regardless of the leaf removal treatment, our results confirm the presence of epiphytic populations of Cladosporium spp. on apparently healthy grape clusters. These populations were detected very early in the ontogenic development of grape berries and increased as the berries matured, reaching maximum numbers on overripe berries (Briceño and Latorre, 2008). As has been suggested for other pathogens affecting grape berries (Commenil et al., 1997; Jeandet et al., 1991; Mlikota Gabler et al., 2003), it is postulated that changes in cuticle porosity, cuticle fractures, lower capacity for developing defense reactions, and the decrease of epicuticular wax content in ripe to overripe grapes may promote the presence of sugary and watery exudates that stimulate the superficial development of Cladosporium spp. in overripe berries. However, further studies are needed to better characterize the dynamics of the epiphytic populations of Cladosporium on grape berries. The populations of Cladosporium spp. were always higher than the populations of other filamentous fungi, such as species of Alternaria, Aspergillus, Botrytis, Penicillium and Rhizopus (results not reported in this article). For instance, population densities of
Briceño, E.X., Latorre, B.A., 2007. Outbreaks of Cladosporium rot associated with delayed harvest wine grapes in Chile. Plant Dis. 91, 1060. Briceño, E.X., Latorre, B.A., 2008. Characterization of Cladosporium rot in grapevines, a problem of growing importance in Chile. Plant Dis. 92, 1635e1642. Briceño, E.X., Latorre, B.A., Bordeu, E., 2009. Effect of Cladosporium rot on the composition and aromatic compounds of red wine. Span. J. Agric. Res. 7, 119e128. Chellemi, D.O., Marois, J.J., 1992. Influence of leaf removal, fungicide applications, and fruit maturity on incidence and severity of grape powdery mildew. Am. J. Enol. Vitic. 43, 53e57. Commenil, P., Brunet, L., Audran, J.C., 1997. The development of the grape berry cuticle in relation to susceptibility to bunch rot disease. J. Exp. Bot. 48, 1599e1607. Díaz, G.A., Torres, R., Vega, M., Latorre, B.A., 2009. Ochratoxigenic Aspergillus species on grapes from Chilean vineyards and Aspergillus threshold levels on grapes. Int. J. Food Microbiol. 133, 195e199. Donoso, A., Latorre, B.A., 2006. Characterization of blue mold caused by Penicillium spp. in cold stored table grapes. Cien. Inv. Agric. 33, 119e130. Duncan, R.A., Stapleton, J.J., Leavitt, G.M., 1995. Population dynamics of epiphytic mycoflora and occurrence of bunch rots of wine grapes as influenced by leaf removal. Plant Pathol. 44, 956e965. English, J.T., Thomas, C.S., Marois, J.J., Gubler, W.D., 1989. Microclimates of grapevine canopies associated with leaf removal and control of Botrytis bunch rot. Phytopathology 79, 395e401. Gil, G.F., Pszczólkowski, P., 2007. Viticultura, Fundamentos para Optimizar Producción y Calidad. Ediciones Universidad Católica, Santiago, Chile, 535 pp. Gubler, W.D., Marois, J.J., Bledsoe, A.M., Bettiga, U., 1987. Control of Botrytis bunch rot of grape with canopy management. Plant Dis. 71, 599e601. Jeandet, P., Bessis, R., Gautheron, B., 1991. The production of resveratrol (3,5,4-trihydroxystilbene) by grape berries in different developmental stages. Am. J. Enol. Vitic. 42, 41e46. Mlikota Gabler, F., Smilanick, J.L., Mansour, M., Ramming, D.W., Mackey, B.E., 2003. Correlations of morphological, anatomical, and chemical features of grape berries with resistance to Botrytis cinerea. Phytopathology 93, 1263e1273. Pszczólkowski, P., Latorre, B.A., Ceppi di Lecco, C., 2001. Efecto de los mohos presentes en uvas cosechadas tardíamente sobre la calidad de los mostos y vinos Cabernet Sauvignon. Cien. Inv. Agric 28, 157e163. Saint-Cricq, N., Vivas, N., Glories, Y., 1998. Maturité phénolique: définition et contrôle. Rev. Fr. OEnol. 98, 22e25. Smart, R.E., 1973. Sunlight interception by vineyards. Am. J. Enol. Vitic. 24, 141e147. Stapleton, J.J., Grant, R.S., 1992. Leaf removal for nonchemical control of the summer bunch rot complex of wine grapes in the San Joaquin Valley. Plant Dis. 76, 205e208. Stapleton, J.J., Leavitt, G.M., Verdegaal, P.S., 1995. Integration of leaf removal and reduced fungicide applications for management of grape powdery mildew in the San Joaquin Valley. Calif. Agric. 49, 33e36. Thompson, J.R., Latorre, B.A., 1999. Characterization of Botrytis cinerea from table grapes in Chile using RAPD-PCR. Plant Dis. 83, 1090e1094. Ulevicius, V., Peciulyte, D., Mordas, G., Lugauskas, A., 2000. Field study on changes in viability of airborne fungal propagules exposed to solar radiation. J. Aerosol Sci. 31, S961eS962. Valero, A., Begum, M., Leong, S.L., Hocking, A.D., Ramos, A.J., Sanchis, V., Maırín, S., 2007. Effect of germicidal UVC light on fungi isolated from grapes and raisins. Lett. Appl. Microbiol. 45, 238e243.