Isidre Llorente

Evaluation of BSPcast Disease Warning System in Reduced Fungicide Use Programs for Management of Brown Spot of Pear

A forecasting model (BSPcast) developed for prediction of brown spot (Stemphylium vesicarium) of pear was evaluated as an advisory system for reduced fungicide use in disease management programs. Eleven trials were performed during 1995, 1996, and 1997 in five orchards in two different climatic areas in Catalunya (Spain) and Emilia-Romagna (Italy). Values of 3-day cumulative daily infection risk (CR) provided by the model were used to determine risk periods during the growing season of pear and were taken as thresholds to schedule fungicide sprays. The fungicide application programs tested using the model consisted of guided schedules with CR action thresholds of 0.4, 0.5, or 0.6, and fixed sprays following a standard commercial schedule. In nine out of 10 trials, no significant differences were observed in disease incidence on fruit at harvest between the fixed-spray commercial schedule and guided sprays using thresholds of 0.4 or 0.5. The average savings in number of fungicide sprays applied using BSPcast compared with the fixed-spray schedule were 20 to 70% when using fungicides with a 15-day protection period (kresoxim-methyl or procymidone) and ranged from 20 to 50% when using a fungicide with a 7-day protection period (thiram).

Susceptibility of selected European pear cultivars to infection by Stemphylium vesicarium and influence of leaf and fruit age.

Nonwounded fruit of 16 European pear cultivars (Pyrus communis) and potted Conference pear plant were inoculated with conidia of Stemphylium vesicarium. Cultivars Passe Crassane, Alexandrine, Conference, Doyenne du Comice, Duc de Bordeaux, Abate Fetel, and General Leclerc were highly susceptible. Cultivars Williams, Blanquilla, Beurre Hardy, Louis Bonne, Grand Champion, and Highland were slightly or not susceptible. The susceptibility of fruit decreased logarithmically from fruit set to harvest in very susceptible cultivars. Younger leaves developed 1.8-3 times more disease than older leave

Brown Spot of Pear: An Emerging Disease of Economic Importance in Europe

Brown spot of pear (BSP), a fungal disease caused by Stemphylium vesicarium (Wallr.) E. Simmons, is economically important in pear-production areas in Europe (Fig. 1) (4,14,18,31,42). The first outbreaks were reported in 1975 in the Emilia-Romagna region in Italy on Abate Fetel pear (31). In 1984, the disease was detected in Spain in the Catalunya region on Passe Crassane and Conference pears (42). Subsequently, it has been reported from France (1987), Portugal (1996), The Netherlands (1997), and Belgium (2002) (4,14,18,35). The relative importance of BSP has increased significantly over the past 10 years in Europe, and it is now considered of similar or even higher incidence than apple scab. Epidemics may be severe, and depending on the year and pear growing area, global losses may be between 1 and 10% of total production, in spite of the control measures applied. Levels of disease (5 to 10%) one year may be followed by up to 90% infected fruit in the next year (28,42).

Effect of relative humidity and interrupted wetness periods on brown spot severity of pear caused by Stemphylium vesicarium

ABSTRACT Field observations in four pear orchards during 5 years from April to October indicated that days with uninterrupted wetness of variable length represented 83.9% of the total days studied. However, days with surface wetness interruptions and with high relative humidity (RH) (>/=90%) without wetness occurred with a frequency of 7.1 and 6.2%, respectively. Accordingly, the effect of interruption of 24-h wetness periods by dry periods of high or low RH on infections caused by Stemphylium vesicarium on pear was determined. Pear plants inoculated with conidia of S. vesicarium were exposed to a 12-h wet period followed by a dry period of variable length (0, 3, 6, 12, 18, or 24 h) and a second wet period of 12 h. The dry period consisted either of low (60%) or high (96%) RH. The infection process was irreversibly stopped under low RH during dry periods between wetness, but continued at high RH. The effect of high RH on disease severity in the absence of wetness was also determined. Pear plants inoculated with S. vesicarium were exposed to periods of variable length (3 to 24 h) either at high RH (96%) in the presence of wetness or at high RH (96%) without wetness. No infections were observed on plants incubated under high RH without wetness, indicating that conidia of S. vesicarium require the presence of a water film in the plant surface to develop infections on pear.

Infection Potential of Pleospora allii and Evaluation of Methods for Reduction of the Overwintering Inoculum of Brown Spot of Pear

The capacity for germination and pathogenicity to pear leaves of ascospores of Pleospora allii, the teleomorph of Stemphylium vesicarium, causal agent of brown spot of pear, were studied in vitro. Most ascospores germinated within 1 h at temperatures between 15 and 20°C, and the optimum temperature for germination was 18.9°C. Infections developed on wounded and non-wounded detached pear leaves, but were more frequent on wounded leaves. The minimum infective dose was one ascospore per wound. Biological, chemical, and mechanical methods for decreasing overwintering inoculum of P. allii were evaluated. Ascospores were discharged from March to May, depending on the orchard and year. Leaf shredding or removal were the most effective methods of reducing overwintering inoculum. Biological control methods based on application of Thichodermasp. formulations were partially effective. Chemical methods based on copper and urea treatments were ineffective.

Control of brown spot of pear by reducing the overwintering inoculum through sanitation

Stemphylium vesicarium, the causal agent of brown spot of pear, overwinters in the leaf residues of pear and herbaceous plants of the orchard floor. Pseudothecia of the teleomorph, Pleospora allii, are formed on these residues where they produce ascospores. New methods were tested aimed at reducing this overwintering inoculum and increasing the efficacy of control of brown spot of pear. Sanitation methods were evaluated in nine trials in Girona (Spain) and Ferrara (Italy) over a 4-year period. The sanitation methods were leaf litter removal in December to February, and application of biological control agents (commercial formulates of Trichoderma spp.) to the orchard ground cover from February to May. Fungicides were also applied to the trees during the pear-growing season, scheduled according to the BSPcast model. The different methods were tested as stand-alone applications or in combination. All methods consistently reduced the disease incidence at harvest on fruit with an efficacy between 30 to 60% for leaf litter removal and more than 60% for the combination of leaf litter removal and biological control. Efficacy of sanitation alone (leaf litter removal and biological control) in reducing the brown spot level on fruit was similar in most of the trials to the efficacy obtained when fungicides were applied alone. However, integration of sanitation methods and fungicides did not improve the efficacy of disease control over the level provided by fungicides alone.

An update on control of brown spot of pear

Brown spot of pear is a fungal disease producing high economical losses in several pear-growing areas in Europe. Fungicide applications during the growing period either at fixed schedule or delivered according to the BSPcast forecasting system are not enough to control the disease under favorable conditions. New strategies have been introduced to control the inoculum production using sanitation methods. These methods are based on combinations of leaf litter removal during winter and biological control agent applications during late winter, spring and summer. These practices reduce both the inoculum pressure and disease levels. Therefore, the resulting optimized disease management consists of a combination of sanitation methods applied during the whole year with chemical fungicides scheduled according to the BSPcast forecasting model during the vegetative period. It is expected that the control of brown spot could be further refined upon availability of rapid methods for inoculum potential analysis. However, this analysis is difficult due to the variability in pathogenicity within the pathogen population.

Postinfection Activity of Synthetic Antimicrobial Peptides Against Stemphylium vesicarium in Pear

Brown spot of pear is a fungal disease of economic importance caused by Stemphylium vesicarium that affects the pear crops in Europe. Due to the characteristics of this disease and the moderate efficacy of available fungicides, the effectiveness of control measures is very limited; however, synthetic antimicrobial peptides (AMPs) may be a complement to these fungicides. In the present study, 12 AMPs of the CECMEL11 library were screened for fungicidal activity against S. vesicarium. In vitro experiments showed that eight AMPs significantly reduced the germination of conidia. The most effective peptides, BP15, BP22, and BP25, reduced fungal growth and sporulation at concentrations below 50 μM. Leaf assays showed that preventive application of BP15 and BP22 did not reduce infection; however, when the peptides were applied curatively, infection was significantly reduced. The use of a BP15 fluorescein 5-isothiocyanate conjugate revealed that the peptide binds to hyphae and germ tubes and produces malformations that irreversibly stop their development.

Interaction of antifungal peptide BP15 with Stemphylium vesicarium, the causal agent of brown spot of pear.

Peptide BP15 has shown antifungal activity against several plant pathogenic fungi, including Stemphylium vesicarium, the causal agent of brown spot of pear. BP15 inhibits the germination, growth and sporulation of S. vesicarium and displays post-infection activity by stopping fungal infection in pear leaves. In this work, live-cell imaging was undertaken to understand the antifungal mechanism of BP15. A double-staining method based on the combination of calcofluor white and SYTOX green coupled with epifluorescence microscopy was used to investigate fungal cell permeabilization and alterations in fungal growth induced by BP15. GFP-transformants of S. vesicarium were obtained and exposed to rhodamine-labelled BP15. Confocal laser microscopy provided evidence of peptide internalization by hyphae, resulting in fungal cell disorganization and death. S. vesicarium membrane permeabilization by BP15 was found to be peptide-concentration dependent. BP15 at MIC and sub-MIC concentrations (10 and 5 μM, respectively) inhibited S. vesicarium growth and produced morphological alterations to germ tubes, with slow and discontinuous compromise of fungal cell membranes. Fungal cell membrane disruption was immediately induced by BP15 at 100 μM, and this was accompanied by rapid peptide internalization by S. vesicarium hyphae. Peptide BP15 interacted with germ tubes and hyphae of S. vesicarium but not with conidial cells.

A model for predicting Xanthomonas arboricola pv. pruni growth as a function of temperature

A two-step modeling approach was used for predicting the effect of temperature on the growth of Xanthomonas arboricola pv. pruni, causal agent of bacterial spot disease of stone fruit. The in vitro growth of seven strains was monitored at temperatures from 5 to 35°C with a Bioscreen C system, and a calibrating equation was generated for converting optical densities to viable counts. In primary modeling, Baranyi, Buchanan, and modified Gompertz equations were fitted to viable count growth curves over the entire temperature range. The modified Gompertz model showed the best fit to the data, and it was selected to estimate the bacterial growth parameters at each temperature. Secondary modeling of maximum specific growth rate as a function of temperature was performed by using the Ratkowsky model and its variations. The modified Ratkowsky model showed the best goodness of fit to maximum specific growth rate estimates, and it was validated successfully for the seven strains at four additional temperatures. The model generated in this work will be used for predicting temperature-based Xanthomonas arboricola pv. pruni growth rate and derived potential daily doublings, and included as the inoculum potential component of a bacterial spot of stone fruit disease forecaster.