Use of Phosphite Salts in Laboratory and Semicommercial Tests to Control Citrus Postharvest Decay.

Potassium phosphite (KP) concentrations that inhibited the germination of 50% of Penicillium digitatum conidia were 229, 334, 360, 469, 498, or 580 mg/liter at pH 3, 4, 5, 6, 7, or 8, respectively. Increasing phosphate content in media reduced phosphite toxicity. To control green or blue mold, fruit were inoculated with P. digitatum or P. italicum, then immersed 24 h later in KP, calcium phosphite (CaP), sodium carbonate, sodium bicarbonate, or potassium sorbate for 1 min at 20 g/liter for each at 25 or 50°C. Mold incidence was lowest after potassium sorbate, CaP, or KP treatments at 50°C. CaP was often more effective than KP but left a white residue on fruit. KP was significantly more effective when fruit were stored at 10 or 15°C after treatment compared with 20°C. Acceptable levels of control were achieved only when KP was used in heated solutions or with fungicides. KP was compatible with imazalil (IMZ) and other fungicides and improved their effectiveness. KP increased thiabendazole or IMZ residues slightly. Phosphite residues did not change during storage for 3 weeks, except they declined when KP was applied with IMZ. KP caused no visible injuries or alteration in the rate of color change of citrus fruit in air or ethylene at 5 µl/liter.

Preharvest Fungicide, Potassium Sorbate, or Chitosan Use on Quality and Storage Decay of Table Grapes.

Potassium sorbate, a program of four fungicides, or one of three chitosan formulations were applied to clusters of 'Thompson Seedless' grape berries at berry set, pre-bunch closure, veraison, and 2 or 3 weeks before harvest. After storage at 2°C for 6 weeks, the natural incidence of postharvest gray mold was reduced by potassium sorbate, the fungicide program, or both together in a tank mixture, in 2009 and 2010. In 2011, the experiment was repeated with three chitosan products (OII-YS, Chito Plant, and Armour-Zen) added. Chitosan or fungicide treatments significantly reduced the natural incidence of postharvest gray mold among grape berries. Berries harvested from vines treated by two of the chitosan treatments or the fungicide program had fewer infections after inoculation with Botrytis cinerea conidia. None harmed berry quality and all increased endochitinase activity. Chitosan decreased berry hydrogen peroxide content. One of the chitosan formulations increased quercetin, myricetin, and resveratrol content of the berry skin. In another experiment, 'Princess Seedless' grape treated with one of several fungicides before 4 or 6 weeks of cold storage had less decay than the control. Fenhexamid was markedly superior to the other fungicides for control of both the incidence and spread of gray mold during storage.

Control of Postharvest Gray Mold of Table Grapes in the San Joaquin Valley of California by Fungicides Applied During the Growing Season.

Fungicides applied before harvest were evaluated to control postharvest gray mold of table grapes, caused by Botrytis cinerea. The concentrations of thiophanate methyl (THM), iprodione (IPR), cyprodinil (CYP), pyraclostrobin + boscalid (PS+BO), pyrimethanil (PYR), or fenhexamid (FEN) that inhibited the growth of four isolates sensitive to these fungicides by 50% (EC50) were 12.4, 2.5, 0.61, 0.29/0.57, 0.26, or 0.17 mg liter-1, respectively. THM, IPR, CYP, PS+BO, PYR, or FEN were applied to detached 'Thompson Seedless' berries at the equivalent of the maximum approved rates of 600, 500, 270, 59/116, 370, or 290 mg liter-1, respectively, except PS+BO, which were used at 54.2% of their current registered maximum rates. The berries were inoculated with B. cinerea 48 or 24 h before treatment or 24 or 48 h after treatment. Gray mold 2 weeks after treatment and storage at 15°C was lowest after FEN application, followed by PYR, CYP, IPR, PS+BO, and THM. In commercial vineyards, one application of FEN, PYR, CYP, or PS+BO, all at their current maximum approved rates, 2 weeks before harvest reduced postharvest gray mold by approximately 50%. When fungicides were applied repeatedly after berry set either in mixtures or alternated with fungicides of different mode of action classes, postharvest gray mold was reduced by about 50% using a commercial air-blast sprayer and by 70 to 87% using a hand-held sprayer that was directed into the clusters. The fungicide sensitivity of isolates collected in numerous vineyards indicated those with reduced sensitivity to all of the tested fungicides, except FEN, were common. The efficacy of preharvest fungicide regimes was not sufficient to replace postharvest sulfur dioxide fumigation.

Influence of Temperature and Humidity on Survival of Penicillium digitatum and Geotrichum citri-aurantii.

Longevity of conidia of Penicillium digitatum (cause of citrus green mold) and arthroconidia of Geotrichum citri-aurantii (cause of sour rot of citrus) was determined under controlled temperature and relative humidity (RH) or ambient summer conditions in central California. Longevity at low RH was longer than at high RH. Hours to kill 99% of the conidia (LT99) of nine P. digitatum isolates were determined at 50°C and 75 or 95% RH. At 75 and 95% RH, the LT99 was 24.9 and 4.9 h, respectively. The LT99 was 30 and 42 days, respectively, for conidia of P. digitatum under ambient conditions at two outdoor locations. The LT99 of arthroconidia of G. citriaurantii from colonies cultured on potato dextrose agar was briefer than that of conidia of P. digitatum. At 45°C and 75 or 95% RH, the LT99 was about 4 and 2 h, respectively, whereas at 50°C, none was viable after 1 h at either humidity. Sanitation is an important practice for managing these diseases. Since there was little or no survival of either fungus after 1 day at 50°C and 75% RH or higher, we conclude that commercial sanitation could employ a similar regime.

Influence of Temperature, Inoculation Interval, and Dosage on Biofumigation with Muscodor albus to Control Postharvest Gray Mold on Grapes.

Control of postharvest gray mold, caused by Botrytis cinerea, on Thompson Seedless grape by biofumigation with a rye grain formulation of Muscodor albus, a fungus that produces volatiles lethal to many microorganisms, was evaluated. The influences of temperature, biofumigant dosage, and interval between inoculation and treatment on disease incidence and severity on detached single berries were assessed. When biofumigation began within 24 h after inoculation, higher M. albus dosages (≥50 g of the M. albus grain formulation per kilogram of grapes at 20°C or 100 g/kg at 5°C) stopped infections and control persisted after M. albus removal. Biofumigation was more effective at 20 than 5°C. Among inoculated clusters inside clamshell boxes incubated for 7 days at 15°C, gray mold incidence was reduced from 20.2% among untreated grape fruit to less than 1%, when ≥5 g of the formulation per kilogram of grapes was added. Among grape berries commercially packaged in ventilated polyethylene cluster bags incubated for 7 days at 15°C, gray mold incidence was 40.5% among untreated fruit and 11.1 or 6.7% when the formulation at 5 or 20 g/kg, respectively, had been added. In the same packaging, among grape berries incubated for 28 days at 0.5°C, gray mold incidence was 42.8% among untreated fruit and 4.8 or 4.0% when the formulation at 5 or 10 g/kg, respectively, had been added. Lower dosages (≤20 g/kg) suppressed disease development while M. albus was present; however, after their removal, B. cinerea resumed growth and gray mold incidence increased. Placement of M. albus inside grape packages significantly controlled gray mold and may be a feasible approach to manage postharvest decay of table grape.

Pre- and Postharvest Treatments to Control Green Mold of Citrus Fruit During Ethylene Degreening.

Two approaches, fungicide applications to trees before harvest and drenching fruit after harvest, were evaluated to minimize postharvest green mold, caused by Penicillium digitatum, particularly among fruit subjected to ethylene gas after harvest, a practice termed "degreening" that eliminates green rind color. Preharvest applications of thiophanate methyl (TM) controlled postharvest green mold consistently. In five tests, green mold among degreened orange fruit was 16% when TM was applied 1 week before harvest; whereas, among fruit not treated, the incidence was 89.5%. Thiabendazole (TBZ) applied to harvested fruit in bins before degreening also was very effective. TBZ effectiveness was enhanced by mild heating (41°C), adding sodium bicarbonate, and immersing fruit, rather than drenching them, with the solution. With these measures, an isolate of P. digitatum with a high level of TBZ resistance was significantly controlled. In semicommercial tests with naturally inoculated fruit, TBZ and sodium bicarbonate treatment reduced green mold incidence from 11% among untreated orange fruit to 2%. TBZ residues in lemon fruit at 41°C were about twice those treated at 24°C. Neither TM before harvest nor TBZ and sodium bicarbonate applied after harvest influenced green color removal during degreening of orange fruit. Sodium bicarbonate slightly reduced the rate of lemon color change.

Preharvest Chitosan and Postharvest UV Irradiation Treatments Suppress Gray Mold of Table Grapes.

The effectiveness of chitosan treatment of table grapes, alone or in combination with ultraviolet-C (UV-C) radiation, to control postharvest gray mold caused by Botrytis cinerea, was determined in California, United States. The influence of these treatments on catechin and resveratrol contents and chitinase activity in grape berry skins also was assessed. Clusters of cvs. Thompson Seedless, Autumn Black, and Emperor were sprayed in the vineyard with 1% chitosan, then harvested daily for 5 days. Promptly after harvest, they were inoculated with B. cinerea. Decay incidence and disease severity were significantly reduced by chitosan, which was most effective on berries harvested 1 or 2 days after treatment. In another experiment, grape berries were sprayed in the vineyard with chitosan, harvested 2 days later, irradiated for 5 min with UV-C (0.36 J/cm2), and inoculated with B. cinerea 2 days later. Combined chitosan and UV-C treatments applied to cv. Autumn Black or selection B36-55 were synergistic in reducing gray mold incidence and severity compared with either treatment alone. Preharvest chitosan treatment increased neither concentration of catechin or resveratrol nor activity of chitinase in berry skin. Conversely, UV-C irradiation, alone or combined with chitosan treatment, induced catechin in cv. Autumn Black berries and trans-resveratrol in both cv. Autumn Black and selection B36-55.

Impact of Postharvest Hot Water or Ethanol Treatment of Table Grapes on Gray Mold Incidence, Quality, and Ethanol Content.

The influence of brief immersion of grape berries in water or ethanol at ambient or higher temperatures on the postharvest incidence of gray mold (caused by Botrytis cinerea) was evaluated. The incidence of gray mold among grape berries that were untreated, or immersed for 1 min in ethanol (35% vol/vol) at 25 or 50°C, was 78.7, 26.2, and 3.4 berries/kg, respectively, after 1 month of storage at 0.5°C and 2 days at 25°C. Heated ethanol was effective up to 24 h after inoculation, but less effective when berry pedicels were removed before inoculation. Rachis appearance, epicuticular wax content and appearance, and berry shatter were unchanged by heated ethanol treatments, whereas berry color changed slightly and treated grape berries were more susceptible to subsequent infection. Ethanol and acetaldehyde contents of grape berries were determined 1, 7, and 14 days after storage at 0.5°C following treatment for 30 or 90 s at 30, 40, or 50°C with water, or 35% ethanol. Highest residues (377 µg/g of ethanol and 13.3 µg/g of acetaldehyde) were in berries immersed for 90 s at 50°C in ethanol. Among ethanol-treated grape berries, the ethanol content declined during storage, whereas acetaldehyde content was unchanged or increased. Untreated grape berries initially contained ethanol at 62 µg/g, which then declined. Acetaldehyde content was 0.6 µg/g initially and changed little during storage.

Influence of pH and NaHCO3 on Effectiveness of Imazalil to Inhibit Germination of Penicillium digitatum and to Control Postharvest Green Mold on Citrus Fruit.

In vitro, spores of Penicillium digitatum germinated without inhibition between pH 4 and 7, but were inhibited at higher pH. Estimated concentrations of imazalil (IMZ) in potato-dextrose broth-Tris that caused 50% reduction in the germination of spores (ED50) of an IMZ-sensitive isolate M6R at pH 4, 5, 6, and 7 were 0.16, 0.11, 0.015, and 0.006 µg/ml, respectively. ED50 IMZ concentrations of an IMZ-resistant isolate D201 at pH 4, 5, 6, and 7 were 5.9, 1.4, 0.26, and 0.07 µg/ml, respectively. The natural pH within 2-mm-deep wounds on lemon was 5.6 to 5.1 and decreased with fruit age. IMZ effectiveness to control green mold and its residues increased with pH. The pH in wounds on lemon fruit 24 h after immersion in 1, 2, or 3% NaHCO3 increased from pH 5.3 to 6.0, 6.3, and 6.7, respectively. NaHCO3 dramatically improved IMZ performance. Green mold incidence among lemon fruit inoculated with M6R and treated 24 h later with IMZ at 10 µg/ml, 1% NaHCO3, or their combination was 92, 55, and 22%, respectively. Green mold among lemon fruit inoculated with D201 and treated 24 h later with water, IMZ at 500 µg/ml, 3% NaHCO3, or their combination was 96.3, 63.0, 44.4, and 6.5%, respectively. NaHCO3 did not influence IMZ fruit residue levels.

Survival of spores of Rhizopus stolonifer, Aspergillus niger, Botrytis cinerea and Alternaria alternata after exposure to ethanol solutions at various temperatures.

AIMS: To quantify and model the toxicity of brief exposures of spores of Rhizopus stolonifer, Aspergillus niger, Botrytis cinerea and Alternaria alternata to heated, aqueous ethanol solutions. These fungi are common postharvest decay pathogens of fresh grapes and other produce. Sanitation of produce reduces postharvest losses caused by these and other pathogens. METHODS AND RESULTS: Spores of the fungi were exposed to solutions containing up to 30% (v/v) ethanol at 25-50 degrees C for 30 s, then their survival was determined by germination on semisolid media. Logistical, second-order surface-response models were prepared for each fungus. Subinhibitory ethanol concentrations at ambient temperatures became inhibitory when heated at temperatures much lower than those that cause thermal destruction of the spores by water alone. At 40 degrees C, the estimated ethanol concentrations that inhibited the germination of 50% (LD(50)) of the spores of B. cinerea, A. alternata, A. niger and R. stolonifer were 9.7, 13.5, 19.6 and 20.6%, respectively. CONCLUSIONS: Ethanol and heat combinations were synergistic. Control of spores of these fungi could be accomplished with much lower temperatures and ethanol concentrations when combined compared with either used alone. Botrytis cinerea and A. alternata were less resistant to the combination than A. niger or R. stolonifer.

Quantification of the Toxicity of Aqueous Chlorine to Spores of Penicillium digitatum and Geotrichum citri-aurantii.

Chlorine toxicity to Penicillium digitatum and Geotrichum citri-aurantii, causes of green mold and sour rot of citrus, respectively, was quantified. In 3% wt/vol NaHCO3 containing 200 µg free chlorine per ml at pH 8.3, 95% of P. digitatum spores died (LT95) by 180 s at 5°C, while only 32 s were required at 24°C. The LT95 of G. citri-aurantii arthrospores was 108 and 31 s at 5 and 24°C, respectively. Mortality slowed 2- to 4-fold for each unit of increase from pH 7 to 10. The LT95 of P. digitatum spores in 200 µg free chlorine per ml at 24°C at pH 7, 8, 9, and 10 was 13.2, 19.1, 29.4, and 88.4 s, respectively. The LT95 of G. citri-aurantii at pH 7, 8, 9, and 10 was 3.0, 12.6, 56.6, and 114 s, respectively. Models were prepared describing mortality. Brief immersion in 200 µg free chlorine per ml reduced viable spores of P. digitatum and G. citri-aurantii from 106 to 103 spores per lemon, and naturally occurring yeast and molds from 106 to 104 CFU. In fruit wound-inoculated and immersed 24 h later in water, 4,000 µg free chlorine per ml, or 3% wt/vol NaHCO3, green mold occurrence after storage was 98.5, 68.3 and 7.5%, respectively.

Comparison of Effects of Acidic Electrolyzed Water and NaOCl on Tilletia indica Teliospore Germination.

Definitive identification of free teliospores of Tilletia indica, causal agent of Karnal bunt of wheat, requires polymerase chain reaction (PCR)-based diagnostic tests. Since direct PCR amplification from teliospores has not been reliable, teliospores first must be germinated in order to obtain adequate DNA. We have routinely surface-sterilized teliospores for 2 min with 0.4% (vol/vol) sodium hypochlorite (NaOCl) to stimulate germination and produce axenic cultures. However, we observed that some spores were killed even with a 2-min NaOCl treatment, the shortest feasible duration. Decreasing the NaOCl concentration in our study from 0.4% to 0.3 and 0.2%, respectively, increased teliospore germination, but treatment times longer than 2 min still progressively reduced the germination percentages. In testing alternative methods, we found "acidic electrolyzed water" (AEW), generated by electrolysis of a weak solution of sodium chloride, also surface-sterilized and increased the rate of T. indica teliospore germination. In a representative experiment comparing the two methods, NaOCl (0.4%) for 2 min and AEW for 30 min increased germination from 19% (control) to 41 and 54%, respectively, by 7 days after treatment. Because teliospores can be treated with AEW for up to 2 h with little, if any, loss of viability, compared with 1 to 2 min for NaOCl, treatment with AEW has certain advantages over NaOCl for surface sterilizing and increasing germination of teliospores of suspect T. indica.

Improved Control of Green Mold of Citrus with Imazalil in Warm Water Compared with Its Use in Wax.

The effectiveness of imazalil for the control of citrus green mold (caused by Penicillium digitatum) improved significantly when fruit were treated with heated aqueous solutions of the fungicide as compared with the current commercial practice of spraying wax containing imazalil on fruit. When applied at less than 500 µg·ml-1 in solutions heated to 37.8°C, control of postharvest green mold of citrus was significantly superior to applications of 4,200 µg·ml-1 imazalil in wax sprayed on fruit at ambient temperatures. The improvement in imazalil efficacy was obtained with a decrease in fungicide residues on the fruit. Residues of about 3.5 µg·g-1 imazalil deposited by the application of imazalil in wax reduced the incidence of green mold on lemons from 94.4% among untreated controls to 15.1%, whereas an equal residue deposited by passing fruit through heated aqueous imazalil reduced green mold incidence to 1.3%. Similar differences were found in tests with oranges. Residues of 2 and 3.5 µg·g-1 imazalil were needed to control the sporulation of P. digitatum on oranges and lemons, respectively. The mode of application of imazalil did not influence control of sporulation. The influence of immersion time, imazalil concentration, and solution temperature on imazalil residues on oranges and lemons was determined in tests using commercial packing equipment, and a model that describes residue deposition was developed. Residues after a 30- or 60-s treatment in heated aqueous imazalil were sufficient to control sporulation, but residues after 15-s treatments were too low and required an additional application of 1,070 µg·ml-1 imazalil in wax to deposit an amount of imazalil sufficient to control sporulation. An imazalil-resistant isolate of P. digitatum was significantly controlled by heated aqueous imazalil. The incidence of green mold of navel oranges was reduced from 98.8 to 17.4% by treatment in 410 µg·ml-1 imazalil at 40.6°C for 90 s. However, control of the resistant isolate required imazalil residues on the fruit of 7.9 µg·g-1, which is within the U.S. tolerance of 10 µg·g-1 but above the 5 µg·g-1 tolerance of some countries that import citrus fruit from the United States.

Influence of Concentration of Soda Ash, Temperature, and Immersion Period on the Control of Postharvest Green Mold of Oranges.

Oranges were inoculated with spores of Penicillium digitatum, the citrus green mold pathogen, and immersed 24 h later in heated soda ash (Na2CO3, sodium carbonate) solutions to control postharvest citrus green mold. Oranges were immersed for 1 or 2 min in solutions containing 0, 2, 4, or 6% (wt/vol) soda ash heated to 35.0, 40.6, 43.3, or 46.1°C. After 3 weeks of storage at 10°C, the number of decayed oranges was determined. Soda ash significantly controlled green mold in every test. The most effective control of green mold was obtained at 40.6 or 43.3°C with 4 or 6% soda ash. The concentration of soda ash greatly influenced efficacy, whereas the influences of temperature or immersion period on soda ash efficacy were small. Solutions of 4 and 6% soda ash were similar in efficacy and provided superior control of green mold compared with 2% soda ash. The control of green mold by soda ash solutions heated to 40.6 or 43.3°C was slightly superior to control by solutions heated to 35.0 or 46.1°C. The control of green mold by 1-min immersion of inoculated oranges in heated soda ash solutions was inferior to immersion for 2 min, but the magnitude of the difference, particularly with 6% soda ash, was small. A second-order response surface model without interactions was developed that closely described the influence of soda ash concentration, temperature, and immersion period on efficacy. The efficacy of soda ash under commercial conditions was better than that predicted by the model, probably because under commercial conditions the fruit were rinsed less thoroughly with water after treatment than in laboratory tests. The primary finding of this work was that soda ash controlled 24-h-old green mold infections at commercially useful levels using shorter immersion periods and lower temperatures than those recommended by other workers for the use of soda ash on lemons. The oranges were not visibly injured in any test.

Germinability of Teliospores of Tilletia indica after Hot Water and Sodium Hypochlorite Treatments.

Hot water and sodium hypochlorite (NaOCl) were evaluated to eradicate teliospores of the Karnal bunt fungus, Tilletia indica, for the purpose of decontaminating grain storage and handling equipment. The germinability of free teliospores and teliospores within the sori of infected wheat was assessed. Temperatures of 25, 60, and 80°C, NaOCl concentrations (wt/vol, pH 11.5) of 0, 0.53, and 1.60%, and immersion periods of 1, 5, 15, and 30 min were evaluated. In other tests, the influence of pH on NaOCl potency and of a delay between treatment and water rinsing were evaluated. Immersion at 80°C in water alone or with NaOCl killed both free teliospores and those within the sori of infected seeds within 1 min. NaOCl at 1.60% at 25°C killed teliospores suspended in water within 15 min, but some teliospores inside sori survived 30 min of this treatment. NaOCl adjusted to pH 8 before use was superior to NaOCl at pH 11.5. An application of 1.60% NaOCl at 25°C for 5 min followed by a 10-min delay before the seeds were rinsed in fresh water killed free teliospores but not all teliospores within sori. This treatment was more effective than the 5-min treatment alone but inferior to the 15-min treatment with NaOCl at a concentration of 1.60%. Because teliospores within the sori of infected seeds are partially protected and much more resistant to NaOCl, we recommend the removal and disposal of seeds from equipment before the treatments are applied. NaOCl radically altered the appearance of the teliospores, leaving a persistent visual indication that they had been treated, while hot water treatment alone did not. Therefore, it is beneficial to add NaOCl to hot water, although the improvement in the sporicidal efficacy was often small.