Botrytis californica, a new cryptic species in the B. cinerea species complex causing gray mold in blueberries and table grapes.

The Botrytis cinerea species complex comprises two cryptic species, originally referred to Group I and Group II based on Bc-hch gene RFLP haplotyping. Group I was described as a new cryptic species B. pseudocinerea During a survey of Botrytis spp. causing gray mold in blueberries and table grapes in the Central Valley of California, six isolates, three from blueberries and three from table grapes, were placed in Group I but had a distinct morphological character with conidiophores significantly longer than those of B. cinerea and B. pseudocinerea We compared these with B. cinerea and B. pseudocinerea by examining morphological and physiological characters, sensitivity to fenhexamid and phylogenetic analysis inferred from sequences of three nuclear genes. Phylogenetic analysis with the three partial gene sequences encoding glyceraldehyde-3-phosate dehydrogenase (G3PDH), heat-shock protein 60 (HSP60) and DNA-dependent RNA polymerase subunit II (RPB2) supported the proposal of a new Botrytis species, B. californica, which is closely related genetically to B. cinerea, B. pseudocinerea and B. sinoviticola, all known as causal agents of gray mold of grapes. Botrytis californica caused decay on blueberry and table grape fruit inoculated with the fungus. This study suggests that B. californica is a cryptic species sympatric with B. cinerea on blueberries and table grapes in California.

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.

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.

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.