Production of patulin and citrinin by Penicillium expansum from British Columbia (Canada) apples.

Twenty-four isolates of Penicillium expansum Link from British Columbia (Canada) apples were cultured in yeast-extract sucrose (YES) at 25°C for 28 days to investigate production of patulin and citrinin. These isolates proved to be potent producers of citrinin, patulin, or in most cases, both mycotoxins. In every isolate, citrinin, patulin, or both compounds were produced at levels as high as 565 µg/mL (mean 269 µg/mL) and 100 µg/mL (mean 31 µg/mL), respectively. Of the 24 isolates, 4 produced citrinin only, and 2 produced patulin only. Overall, 83% of the isolates formed patulin and 91% formed citrinin. YES broth proved to be an effective medium for patulin and citrinin production. Other workers have noted that production of these mycotoxins in culture often presages production in fruits, so these results might help Canadian fruit processors evaluate and minimize mycotoxin levels in their products.

First Report of a New Postharvest Disease of Pear Fruit Caused by Sphaeropsis pyriputrescens in Canada.

A survey of stored d'Anjou pears was conducted in British Columbia (BC), Canada in January 2006 to determine if Sphaeropsis rot was present in BC as had been reported previously for apples and pears in Washington (1,2). Sphaeropsis pyriputrescens Xiao & J.D. Rogers produces decay similar to Botrytis cinerea that originates from the stem or calyx end. Of 3,614 pears sampled, 55 (1.5%) had symptoms similar to those described for Sphaeropsis rot. Isolations were made from each infected pear onto acidified potato dextrose agar (APDA) dishes and incubated at 20°C for 5 to 7 days. Twenty-seven cultures resembling S. pyriputrescens were induced to produce pycnidia by exposing them to 12-h cycles of alternating light and dark periods at 20°C (1). Conidia extracted from pycnidia were then streaked onto PDA dishes and incubated at 20°C for 12 to 24 h from which single-spore cultures were made. These isolates developed a dense, white-to-cream mycelium that turned yellow over time; black pycnidia were formed on the culture dishes after 4 weeks. Conidia were brown, clavate to subglobose to irregular, and similar in size (16 × 10 µm) to previous descriptions (1). Identification of S. pyriputrescens was confirmed by using DNA sequence data from the ß-tubulin and ribosomal genes. Sequences from S. pyriputrescens from Washington (1) were compared with those from BC, Canada. Isolates from Canada shared 99 to 100% sequence homology with those from Washington. Two of the BC isolates (DAOM 238917 and 238918) were deposited in the Canadian Culture Collection, Ottawa, ON and their corresponding sequences were placed in the GenBank database (NCBI, Bethesda, MD) with accession nos. EU156037 and EU156040 (ribosomal gene) and EU156048 and EU 156050 (ß-tubulin gene), respectively. Five isolates from different locations in BC and two isolates from Washington were tested for pathogenicity on d'Anjou pears and four apple cultivars (Ambrosia, Fuji, Gala, and Granny Smith). Plugs (3 mm in diameter) removed from 2-week-old cultures were placed into two corresponding wounds on each of five fruit per cultivar. The fruit were then placed at 1 or 20°C for 22 or 7 days, respectively, when the diameters of the decay areas were recorded. All isolates were pathogenic on pears (P = <0.05). Decay lesion diameter was greater at 1°C, ranging from 46.8 to 57.9 mm, than at 20°C, ranging from 32.6 to 44.2 mm. All BC isolates were also pathogenic on the fruit of each apple cultivar (P = <0.05), although at 20°C, decay areas were smaller than on pears, and at 1°C, very little rot developed. Koch's postulates were completed by reisolating S. pyriputrescens from the inoculated pears and apples and identifying the isolates as above. Although S. pyriputrescens was only observed on pears in BC, research in Washington indicates that it is a more serious problem on apples (2). To our knowledge, this is the first documented report of the occurrence of S. pyriputrescens in Canada. References: (1) C. L. Xiao and J. D. Rogers. Plant Dis. 88:114, 2004. (2) C. L. Xiao et al. Plant Dis. 88:223, 2004.

First Report of Brown Rot in Wine Grapes Caused by Monilinia fructicola in Canada.

A survey was conducted in 2001 and 2002 to determine incidence of fruit pathogens in wine grapes (Vitis vinifera), an important crop in the southern interior of British Columbia (BC), Canada. Grape clusters were sampled every 2 weeks from June to October at eight vineyard sites located from Osoyoos in the south to Kelowna, approximately 100 km to the north. In the laboratory, the berry clusters were surface disinfested for 0.5 min in 70% ethanol, followed by 1 min in 0.5% sodium hypochlorite, and rinsed twice in sterile distilled water. The berries were placed on potato dextrose agar (PDA) amended with 15 ml/liter of 85% lactic acid and incubated at 20°C for 1 week. During the 2002 survey, a fungus resembling Monilinia fructicola (G. Wint.) Honey was observed sporulating on immature 'Pinot noir' grapes from Kelowna that were sampled on 14 August. Later in the growing season, a similar fungus was detected on 'Riesling' grapes from Summerland sampled on 11 September. There was no evidence of brown rot near the vineyard in Kelowna, but diseased stonefruit were present near the vineyard in Summerland. Subsequent identification of the fungus from 'Riesling' as M. fructicola was based on morphological characters and DNA sequence data for the internal transcribed spacer (ITS) regions of the nuclear ribosomal rRNA genes. The sequenced isolate was deposited in the Canadian Collection of Fungus Cultures as DAOM 231119, and the ITS sequence was accessioned in GenBank as AY289185. Colony growth on PDA was rapid and in concentric rings with the colony margin complete, microconidia abundant, and macroconidia 12 to 13 µm long. Macroconidia germinated with a long germ tube before branching. These characteristics distinguished this fungus from M. laxa, a closely related species that is slow growing with lobed colony margins, produces few microconidia, and germ tubes that branch close to the conidium (1). The complete ITS sequence for DAOM 231119 was a 100% match to other sequences deposited for M. fructicola (Z73777, AF010500, and U21815). On the basis of comparisons of available data, ITS sequences for M. fructicola (three complete ITS, seven partial ITS) and M. laxa (8 complete ITS, 10 partial ITS) differed consistently at four nucleotide positions. The fungus identified as M. fructicola was tested for pathogenicity on mature surface-sterilized 'Pinot noir' and 'Riesling' grapes. Under humid conditions, buff-colored sporodochia bearing conidia developed over the surface of the infected berries. This indicates that M. fructicola can cause decay of wine grapes and could be confused with bunch rot caused by Botrytis cinerea. Previously, M. fructicola was reported on grapes in Oklahoma, but likely these grapes were not Vitis vinifera (2). To our knowledge, this is the first report of brown rot caused by M. fructicola on wine grapes in North America. References: (1) L. R. Batra. World Species of Monilinia (Fungi): Their Ecology, Biosystematics and Control. Mycologia Memoir No. 16. Gerbrüder Borntraeger, Berlin/Stuttgart, 1991. (2) D. A. Preston. Host Index of Oklahoma Plant Diseases, Tech. Bull. No. 21. Oklahoma Agricultural and Mechanical College, Agricultural Experiment Station, Stillwater, 1945.

Disinfection of mung bean seed with gaseous acetic acid.

Mung bean seed inoculated with Salmonella Typhimurium, Escherichia coli O157:H7, and Listeria monocytogenes (3 to 5 log CFU/g) was exposed to gaseous acetic acid in an aluminum fumigation chamber. Salmonella Typhimurium and E. coli O157:H7 were not detected by enrichment of seeds treated with 242 microl of acetic acid per liter of air for 12 h at 45 degrees C. L. monocytogenes was recovered by enrichment from two of 10 25-g seed samples treated in this manner. Fumigation with gaseous acetic acid was also lethal to indigenous bacteria and fungi on mung bean seed. The treatment did not significantly reduce seed germination rates, and no differences in surface microstructure were observed between treated and untreated seed viewed by scanning electron microscopy.

First Report of Powdery Mildew, Caused by Erysiphe cichoracearum, on Coneflowers.

Purple coneflowers (Echinacea purpurea) are grown in North America and Europe for their medicinal properties and as ornamental plants. In September 1997 and again in 1998, a previously undescribed disease was noticed on fully grown coneflower plants in Summerland and Oliver, British Columbia. Mycelia were observed on stems, foliage, and flowers, and distinct dark red to black, round (approximately 5 mm in diameter) lesions were observed on the flower petals. The disease appeared similar to powdery mildews that have been reported on numerous genera of the Asteraceae. Samples of the diseased tissue were examined and the salient features of the fungus on two specimens were determined: cleistothecia infrequent, subglobose or flattened on the side next to the leaf surface, 121 to 209 µm in diameter; epidermal (surface) cells 20 µm in diameter; appendages hyphoid, 5 µm in diameter, up to 200 µm long; asci, 10 to 19 in each cleistothecium, broadly ellipsoid, 47 to 85 × 28 to 37 µm with a short stalk, about 8 to 13 µm long and 8 µm in diameter; ascospores, immature, two per ascus, ellipsoid to broadly ellipsoid, 17 to 25 × 11 to 13 µm, thin walled, hyaline, and smooth; conidia oblong with sides slightly convex and apices truncate, 27 to 40 × 14 to 20 µm, walls hyaline, thin, smooth. Based on the occurrence of asci that contained two ascospores and the hyphoid appendages on the cleistothecia we concluded that the fungus was Erysiphe cichoracearum DC. Damage due to this disease was minimal in 1997 and 1998 because it developed very late in the growing season and occurred sporadically within the plantings. In order to complete Koch's postulates, Echinacea purpurea plants grown in the greenhouse were inoculated with a conidial suspension (105 to 106 conidia per ml) from field-infected plants. Powdery mildew first appeared 3 months later, eventually infecting leaves and stems of 12 of 49 inoculated plants. It was distinctly white and in discrete patches on leaves, compared with coalescing dark brown areas on the stems. Microscopic examination of the conidia confirmed that they were E. cichoracearum. Although powdery mildew caused by E. cichoracearum has been widely reported on lettuce, safflower, and other cultivated and wild Compositae, we found no reference to it on Echinacea spp. in Canada (1,2), the U.S. (3), or elsewhere in the world (4). The specimens have been deposited in the National Mycological Herbarium of Canada (DAOM) with accession numbers 225933 and 225934 for Oliver and Summerland, B.C., respectively. References: (1) U. Braun. Beih. Nova Hedwigia 89:1, 1987. (2) I. L. Conners. 1967. An annotated index of plant diseases in Canada and fungi recorded on plants in Alaska, Canada, and Greenland. Canada Dept. of Agric. Pub. 1251. (3) D. F. Farr et al. 1989. Fungi on Plants and Plant Products in the United States. American Phytopathological Society, St. Paul, MN. (4) J. Ginns. 1986. Compendium of plant disease and decay fungi in Canada, 1960-1980. Agriculture Canada Pub. 1813.

Fumigation of Fruit with Short-Chain Organic Acids to Reduce the Potential of Postharvest Decay.

Vapors of acetic (1.9 or 2.5 µl/liter), formic (1.2 µl/liter), and propionic (2.5 µl/liter) acids were tested for postharvest decay control on 8 cherry, 14 pome, and 3 citrus fruit cultivars. Surfacesterilized fruit were inoculated with known fungal pathogens by drying 20-µl drops of spore suspension on marked locations on each fruit, placing at 10°C to equilibrate for approximately 24 h, and fumigating by evaporating the above acids in 12.7-liter airtight fumigation chambers for 30 min. Immediately after fumigation, the fruit were removed, aerated, aseptically injured, and placed at 20°C until decay occurred. All three fumigants controlled Monilinia fructicola, Penicillium expansum, and Rhizopus stolonifer on cherry. Formic acid increased fruit pitting on six of eight cultivars and was the only organic acid to increase blackening of cherry stems when compared to the control. Decay of pome fruit caused by P. expansum was reduced from 98% to 16, 4, or 8% by acetic, formic, and propionic acids, respectively, without injury to the fruit. Decay of citrus fruit by P. digitatum was reduced from 86 to 11% by all three acids, although browning of the fruit peel was observed on grapefruit and oranges fumigated with formic acid.

First Report of Mucor Rot in Commercially Sold Cherries Caused by Mucor piriformis.

In August 1996, a commercial retailer of high quality cherries from the Okanagan Valley of British Columbia, Canada, reported severe losses in at least 150 boxes (11.3 kg per box) of packed cherries cv. Stella. Upon examination of a representative box of these cherries, it was found that approximately 80% of the fruit was affected by what appeared to be Mucor spp. The decayed cherries were concentrated below the surface layer and small pockets of cherries had prominent salt-and-pepper-colored whiskers emanating from individual cherries. Counting the exact number of rotten cherries was difficult because they disintegrated on handling. Several infected cherries were surface sterilized by immersing in a 0.5% NaClO solution for 1 min and rinsing with sterile, distilled water. Pieces of the surface-sterilized cherries were plated onto dishes containing potato dextrose agar and incubated at 6°C for 10 days. Ten typical isolates derived from single spores from these dishes were all identified as Mucor piriformis A. Fischer (1). Samples of 200 g of cherries cv. Sweetheart (20 ± 1 cherries) were sprayed with a water suspension containing 1 × 106 sporangiospores per ml with one of the isolates identified as M. piriformis and incubated at 1, 6, 10, 15, and 20°C for up to 25 days. This same procedure was repeated for two other isolates of M. piriformis isolated from infected cherries. As a control, cherries were misted with sterile water and incubated at 20°C for 6 days. At 6°C, a temperature at which cherries are often stored, the three isolates decayed, respectively, 100, 60, and 60% of the cherries in 18 days. In general, all three isolates caused greater than 50% decay at 20, 15, 10, and 1°C in 6, 6, 11, and 25 days, respectively. Decay did not occur in cherries misted only with sterile water. Careful visual examination of decayed cherries revealed similar whiskerlike growth of Mucor as seen in the commercial cherries. Microscopic examination of sporangiophores infecting the fruit found them to be the same as the original M. piriformis isolate, successfully completing Koch's postulates. Previous to this report, M. piriformis, M. hiemalis, and M. plumbeus were isolated from Lambert cherries stored for 4 weeks at 4.5°C in Oregon (2); however, it is unclear if all or any of these isolates were pathogenic because pathogenicity tests were not conducted. References: (1) T. J. Michailides and R. A. Spotts. Plant Dis. 74:537, 1990. (2) P. G. Sanderson and R. A. Spotts. Fungic. Nematic. Tests 47:43, 1992.