First Report of Aspergillus niger Causing Postharvest Fruit Rot of Cherry in the Prefectures of Imathia and Pella, Northern Greece

In June 2011, symptoms of postharvest rot were observed on approximately 3% of all cherries collected from commercial orchards of cultivars Lapen and Ferrovia in the prefectures of Imathia and Pella (northern Greece). Fruit were harvested in a timely manner to avoid overripeness. No wounds or other predisposing injuries were observed on the infected fruits. Lesions enlarged rapidly and separated easily from healthy tissue when pressure was applied. Infected tissues were pale and water soaked and the associated fungal spores were dark and powdery and easily liberated when mature. The fungus grew rapidly and produced black colonies on acidified potato dextrose agar (2.5 ml of 85% lactic acid per liter of nutrient medium) after 5 days at 24°C. Identification of the pathogen was based on morphological characteristics (1). The conidial head was radiate, vesicles were nearly spherical and covered with metulae and phialides (biseriate). Conidia were globose (3 to 5 μm in diameter) and usually very rough with irregular ridges, bars, and verrucae. Koch's postulates were completed in the laboratory by inoculating mature cherry fruits (cv. Lapen). The fruits were surface sterilized by dipping in 10% chloride bleach solution, allowed to dry in a laminar flow hood, and wounded with a sharp glass rod that was 2 mm in diameter. A 40-μl drop of a suspension containing 20,000 conidia per ml of water was placed on each wound. There were 20 inoculated and 20 control fruits (similarly wounded and inoculated with a 40-μl drop of sterile distilled water) in a randomized design and incubated at 24 to 26°C for 6 days. Koch's postulates were satisfied after reisolating the fungus from inoculated fruit that developed symptoms similar to those observed on fruit collected from orchards. Control fruits did not show any symptom of the disease. To our knowledge, this is the first report of the occurrence of Aspergillus niger as the causal agent of postharvest rots of cherries in Greece. Postharvest fruit rots caused by A. niger have been reported in cherry orchards of other countries around the world (2). Because this disease causes postharvest rots of cherry fruits, measures may need to be implemented to manage the pathogen. References: (1) M. A. Klich. Page 12 in: Identification of Common Aspergillus Species. Centraalbureau Voor Schimmelcultures, Utrecht, the Netherlands, 2002. (2) A. Valiuskaite et al. Phytopathol. Pol. 35:197, 2005.

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First Report of Stemphylium botryosum Causing Leaf Blight of Kiwi in the Province Imathia, Northern Greece

Plant Disease ◽

10.1094/pdis-92-4-0650b ◽

2008 ◽

Vol 92 (4) ◽

pp. 650-650 ◽

Cited By ~ 1

Author(s):

T. Thomidis ◽

T. J. Michailides

Keyword(s):

Apical Cell ◽

Morphological Characteristics ◽

Leaf Blight ◽

Width Ratio ◽

Northern Greece ◽

Cell Width ◽

Koch’S Postulates ◽

First Report ◽

Stemphylium Botryosum ◽

Koch's Postulates

In Greece, kiwi (Actinidia deliciosa) is mostly found in the northern part of the country where approximately 440,000 ha are grown. In the summer of 2006, a Stemphylium sp. was frequently isolated from leaves of kiwi (cv. Hayward) grown in the province of Imathia. Symptomatic leaves were covered with irregular, necrotic, brown areas. Lesions had a distinct margin that, in some cases, covered a wide part of the diseased leaves. Intense symptoms were frequently observed and associated with defoliation. This Stemphylium sp. was consistently isolated from diseased leaves onto potato dextrose agar (PDA) after surface sterilization with 0.1% chlorine solution. On the basis of morphological characteristics of mycelia, dimensions (length 20 to 29 μm and width 14 to 21 μm) and mean length/width ratio (1.42 μm) of conidia, and width and apical cell width of condiophores, the fungus was identified as Stemphylium botryosum (Wallr.) (2,3) Koch's postulates were completed in the laboratory by inoculating leaves of kiwi (cv. Hayward) with an isolate of S. botryosum originated from a symptomatic leaf of a Hayward kiwi. Twenty leaves were surface sterilized by dipping them into 0.1% chlorine solution for 2 to 3 min, washing in sterile distilled water, and allowing them to dry in a laminar flow hood. A leaf was then placed into a petri plate containing a wet, sterilized paper towel. Inoculation was made by transferring a 5-mm-diameter mycelial disc from the margins of a 7-day-old culture onto the center of each leaf surface. Petri plates were closed and incubated at 25°C with 12 h of light for 6 days. Koch's postulates were satisfied when the same S. botryosum was reisolated from 100% of inoculated leaves that developed symptoms similar to those observed in the vineyards. Leaves inoculated with a PDA plug alone (with no S. botryosum) did not develop any symptoms. Previously, Alternaria alternata was reported as the causal agent of a leaf spot pathogen of kiwi (1,4). To our knowledge, this is the first report of the occurrence of S. botryosum causing leaf blight of kiwi in Greece and worldwide. This pathogen can cause a high level of defoliation in diseased plants. References: (1) L. Corazza et al. Plant Dis. 83:487, 1999. (2) M. B. Ellis. Dematiaceous Hyphomycetes. Mycology Institute. London, England, 1971. (3) E. G. Simmons. Mycologia 61:1, 1969. (4) C. Tsahouridou and C. C. Thanassoulopoulos. Plant Dis. 84:371, 2000

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First Report of Phomopsis amygdali Causing Fruit Rot on Peaches in Greece

Plant Disease ◽

10.1094/pd-90-1551c ◽

2006 ◽

Vol 90 (12) ◽

pp. 1551-1551 ◽

Cited By ~ 3

Author(s):

T. J. Michailides ◽

T. Thomidis

Keyword(s):

Prunus Persica ◽

Morphological Characteristics ◽

Fruit Rot ◽

Peach Fruit ◽

Koch’S Postulates ◽

Canker Disease ◽

First Report ◽

Immature Fruit ◽

Diseased Fruit ◽

Koch's Postulates

In the summer of 2005, the fungus Phomopsis amygdali (Del.) Tuset & Portilla was frequently isolated from decayed peaches (Prunus persica cv. Andross) grown in the province of Imathia, Greece. Fruit infected by P. amygdali developed gray-to-brown decay lesions with white mycelium forming on the surface of lesions. Identification of the pathogen was based on morphological characteristics. Dark-pigmented pycnidia (flask-shaped, conidia-bearing fruiting bodies) were produced over the surface of potato dextrose agar. The pycnidia exuded conidia in white tendrils 7 days later. Koch's postulates were completed in the laboratory by inoculating mature and immature cv. Andross peach fruits with an isolate of P. amygdali isolated from decayed cv. Andross peaches. Thirty peach fruit were surface sterilized by dipping them into 0.1% chlorine solution and allowing them to dry in a laminar flow hood. The peach fruit were wounded with a 2-mm diameter glass rod and a 40-μl drop of 5 × 105 conidia of P. amygdali per milliliter suspension was applied to the wound. Thirty control fruits were similarly wounded and inoculated with a 40-μl drop of sterile water. All inoculated and noninoculated fruit were incubated at 24 to 26°C for 7 days. Koch's postulates were satisfied when the same fungus was reisolated from 100% of inoculated mature and immature fruit that developed symptoms similar to diseased fruit collected from orchards. Although P. amygdali has been previously reported as a causal agent of canker disease (2) and fruit rots of peaches (1) in other countries, to our knowledge, this is the first report of the occurrence of P. amygdali causing a fruit rot of peaches in Greece. References: (1) Y. Ko and S. Sun. Plant Pathol. Bull. 12:212, 2003. (2) E. I. Zehr, Constriction canker. Page 31 in: Compendium of Stone Fruit Diseases. J. M. Ogawa et al., eds. The American Phytopathological Society, St. Paul, MN, 1995.

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First Report of Aspergillus niger Causing Root Rot of Peanut in China

Plant Disease ◽

10.1094/pdis-05-14-0530-pdn ◽

2015 ◽

Vol 99 (2) ◽

pp. 284-284 ◽

Cited By ~ 3

Author(s):

M. L. Xu ◽

J. G. Yang ◽

J. X. Wu ◽

Y. C. Chi ◽

L. H. Xie

Keyword(s):

Aspergillus Niger ◽

Root Rot ◽

Genomic Dna ◽

Dense Layer ◽

Causal Organism ◽

Centraalbureau Voor ◽

Koch’S Postulates ◽

First Report ◽

Initial Symptoms ◽

Koch's Postulates

Peanut (Arachis hypogaea) is one of the most important oil crops and food legumes worldwide. China sows approximately 3.5 million hectares each year and produces 40% of the world's peanuts. Fungal diseases are among the main biotic stresses affecting peanut production. Root rot is a serious disease caused by several fungi. Pythium spp., Fusarium spp., and Rhizopus spp. are some of the root rot fungi that have been reported in China. In 2012 and 2013, root rot symptoms were observed in several fields in Laixi District, Qingdao City, Shandong Province, China. The first symptoms appeared in July. Initial symptoms of the disease were brown spots on the stem base and root. Affected plants were stunted, with leaf chlorosis, reduced growth, or sudden wilting. As disease progressed, the infected tissues showed brown discoloration and rot, and abundant dark brown and black powdery spores were visible on the surfaces of affected parts. Eventually, affected plants collapsed and died. To isolate the causal organism, roots and stems were cut into sections, which were surface-disinfected with 70% ethanol solution (v/v) for 20 s, soaked in 0.1% mercuric chloride solution for 50 s, rinsed with sterilized water three times, dried, placed on Czapek's Dox agar supplemented with chloramphenicol (100 μg/ml), and incubated at 28°C for 7 days. Fungal colonies were white initially and then covered with a dense layer of dark brown or black conidial heads. The conidial head was radiate; vesicles were nearly spherical and covered with irregular metulae and phialides. Conidia were globose or subglobose (3.0 to 5.5 μm in diameter), dark brown to black, with rough cell walls. Total genomic DNA was extracted from mycelia using the EasyPure Genomic DNA Kit (TransGEN, Beijing, China). The rDNA-ITS region was amplified using PCR with the universal fungal primers ITS1 and ITS4 (2). The purified products were separately sequenced in both directions using the same primer pair. The sequences (GenBank Accession No. KJ848716) obtained were 99% similar to the ITS sequence of isolates of Aspergillus niger. This, together with the morphological characters (1) described above, suggested that the microorganism we had isolated was A. niger. Koch's postulates were completed in the laboratory by inoculating peanut. Thirty Huayu20 peanut seeds were placed in a 500-ml sterile pot with 300 g of autoclaved soil. Twenty days after seedling emergence, 15 peanut plants were wounded with a needle and inoculated with 5 ml of conidia suspension (106 ml−1). The same number of peanuts were similarly wounded and inoculated with 5 ml of sterile distilled water to serve as controls in the same pot. All peanuts were kept in a randomized complete block design at 30°C under a 12-h photoperiod. After 7 days, disease symptoms similar to those observed in the field appeared in all inoculated but not in non-inoculated peanuts. The tests were repeated three times in the greenhouse. Koch's postulates were satisfied after re-isolating the A. niger from inoculated peanuts using the method described above. To our knowledge, this is the first report of A. niger causing root rot in peanut in China. References: (1) M. A. Klich. Page 12 in: Identification of Common Aspergillus Species. Centraalbureau voor Schimmelcultures, Utrecht, The Netherlands, 2002. (2) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990.

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First Report of Leaf Spot Disease on Walnut Caused by Colletotrichum fioriniae in China

Plant Disease ◽

10.1094/pdis-09-14-0938-pdn ◽

2015 ◽

Vol 99 (2) ◽

pp. 289-289 ◽

Cited By ~ 9

Author(s):

Y. Z. Zhu ◽

W. J. Liao ◽

D. X. Zou ◽

Y. J. Wu ◽

Y. Zhou

Keyword(s):

Dna Sequences ◽

Leaf Spot ◽

Juglans Regia ◽

Morphological Characteristics ◽

Leaf Spot Disease ◽

Koch’S Postulates ◽

First Report ◽

Spot Disease ◽

Leaf Spots ◽

Koch's Postulates

In May 2014, a severe leaf spot disease was observed on walnut tree (Juglans regia L.) in Hechi, Guangxi, China. Leaf spots were circular to semicircular in shape, water-soaked, later becoming grayish white in the center with a dark brown margin and bordered by a tan halo. Necrotic lesions were approximately 3 to 4 mm in diameter. Diseased leaves were collected from 10 trees in each of five commercial orchards. The diseased leaves were cut into 5 × 5 mm slices, dipped in 75% ethanol for 30 s, washed three times in sterilized water, sterilized with 0.1% (w/v) HgCl2 for 3 min, and then rinsed five times with sterile distilled water. These slices were placed on potato dextrose agar (PDA), followed by incubating at 28°C for about 3 to 4 days. Fungal isolates were obtained from these diseased tissues, transferred onto PDA plates, and incubated at 28°C. These isolates produced gray aerial mycelium and then became pinkish gray with age. Moreover, the reverse of the colony was pink. The growth rate was 8.21 to 8.41 mm per day (average = 8.29 ± 0.11, n = 3) at 28°C. The colonies produced pale orange conidial masses and were fusiform with acute ends, hyaline, sometimes guttulate, 4.02 to 5.25 × 13.71 to 15.72 μm (average = 4.56 ± 0.31 × 14.87 ± 1.14 μm, n = 25). The morphological characteristics and measurements of this fungal isolate matched the previous descriptions of Colletotrichum fioriniae (Marcelino & Gouli) R.G. Shivas & Y.P. Tan (2). Meanwhile, these characterizations were further confirmed by analysis of the partial sequence of five genes: the internal transcribed spacer (ITS) of the ribosomal DNA, beta-tubulin (β-tub) gene, glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene, chitin synthase 3(CHS-1) gene, and actin (ACT) gene, with universal primers ITS4/ITS5, T1/βt2b, GDF1/GDR1, CHS1-79F/CHS1-354R, and ACT-512F/ACT-783R, respectively (1). BLAST of these DNA sequences using the nucleotide database of GenBank showed a high identify (ITS, 99%; β-tub, 99%; GAPDH, 99%; CHS-1, 99%; and ACT, 100%) with the previously deposited sequences of C. fioriniae (ITS, KF278459.1, NR111747.1; β-tub, AB744079.1, AB690809.1; GAPDH, KF944355.1, KF944354.1; CHS-1, JQ948987.1, JQ949005.1; and ACT, JQ949625.1, JQ949626.1). Koch's postulates were fulfilled by inoculating six healthy 1-year-old walnut trees in July 2014 with maximum and minimum temperatures of 33 and 26°C. The 6-mm mycelial plug, which was cut from the margin of a 5-day-old colony of the fungus on PDA, was placed onto each pin-wounded leaf, ensuring good contact between the mycelium and the wound. Non-colonized PDA plugs were placed onto pin-wounds as negative controls. Following inoculation, both inoculated and control plants were covered with plastic bags. Leaf spots, similar to those on naturally infected plants, were observed on the leaves inoculated with C. fioriniae within 5 days. No symptoms were observed on the negative control leaves. Finally, C. fioriniae was re-isolated from symptomatic leaves; in contrast, no fungus was isolated from the control, which confirmed Koch's postulates. To our knowledge, this is the first report of leaf disease on walnut caused by C. fioriniae. References: (1) L. Cai et al. Fungal Divers. 39:183, 2009. (2) R. G. Shivas and Y. P. Tan. Fungal Divers. 39:111, 2009.

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First Report of Cylindrocarpon pauciseptatum Associated with Root Rot and Decline of Peach in Southern Italy (Apulia Region)

Plant Disease ◽

10.1094/pdis-10-11-0858 ◽

2012 ◽

Vol 96 (5) ◽

pp. 764-764 ◽

Cited By ~ 4

Author(s):

T. Yaseen ◽

Y. Ahmed ◽

A. M. D'Onghia ◽

M. Digiaro

Keyword(s):

Prunus Persica ◽

Southern Italy ◽

Vascular Lesions ◽

Universal Primers ◽

Stone Fruits ◽

Centraalbureau Voor ◽

Koch’S Postulates ◽

First Report ◽

Apulia Region ◽

Koch's Postulates

During a survey for the sanitary status of stone fruits in southern Italy (Apulia region), symptoms of low vigor, sparse foliage, and chlorosis of leaves, frequently leading to decline or death of the plants, were observed on 3- to 5-year-old peach trees (Prunus persica) cvs. Tardi Belle, Zee Lady, and O'Henry grafted on GF677. Brown-to-black discolorations of the wood were observed in cross-sections of the trunks just below the graft union. Samples were collected from May to June 2010 from two symptomatic orchards in Brindisi and Foggia provinces. Small pieces of brownish, vascular wood and necrotic root tissues were surface disinfested, placed onto potato dextrose agar (PDA), and incubated for 7 days at 25°C in the dark. Single-conidial isolates were subsequently grown on PDA at 25°C for 10 days. Fungal colonies were presumptively identified as members of the genus Cylindrocarpon on the basis of their morphological and conidial characteristics. On PDA, the isolates developed abundant mycelium, which gradually became yellowish or partially brownish. Macroconidia were predominantly three septate, straight and cylindrical with both ends broadly rounded. Chlamydospores and ovoidal microconidia were observed on synthetic nutrient-poor agar (1). Sequence of the ribosomal internal transcribed spacer (ITS) region was obtained using universal primers (ITS6-ITS4) and deposited in GenBank (Accession No. HE577846). This sequence revealed 100% genetic identity with a sequence from Cylindrocarpon pauciseptatum Schroers & Crous (Accession No. EF607090), a recently described species (3). In nature, several species of the genus Cylindrocarpon affect a large number of woody plants, mainly grapevine, olive, and stone fruits, in which they attack the root surface (2). To verify Koch's postulates, the roots of 20 3-month-old peach seedlings (GF305) were dipped for 30 min in a spore suspension of the fungus (1 × 108 conidia ml–1). Seedlings were then transplanted in an artificial soil mix and held under controlled conditions in a greenhouse at 24°C. Typical black-foot symptoms developed on 92% of the inoculated plants within 3 months, whereas the control plants, whose roots had been dipped in distilled water, remained healthy. C. pauciseptatum was reisolated from infected tissues and internal vascular lesions of 45% of the inoculated plants, but none of the plants used as controls, fulfilling Koch's postulates. To our knowledge, this is the first report of this pathogen on peach in the Apulia Region of Italy. Currently, C. pauciseptatum is limited to a few orchards where presumably it was introduced with infected propagating material from extra-regional nurseries. C. pauciseptatum has the potential to negatively affect the stone fruit industry in Italy including reducing nursery production and productivity and vigor of trees in orchards, or even rapid death of young trees. References: (1) W. Gams et al. CBS Course of Mycology. 4th ed. Centraalbureau voor Schimmelcultures, Baarn, the Netherlands, 1998. (2) M. E. S. Hernandez et al. Eur. J. Plant Pathol. 104:347, 1998. (3) H. J. Schroers et al. Mycol. Res. 112:82, 2008.

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First report of Stemphylium eturmiunum causing leaf spot and black spot on apple in Zhaotong, Yunnan, China

Plant Disease ◽

10.1094/pdis-08-21-1834-pdn ◽

2021 ◽

Author(s):

Cong Zhou ◽

Xin Long Pan ◽

Bao Hua Kong ◽

Yueqiu He ◽

Jun Ma ◽

...

Keyword(s):

Leaf Spot ◽

Yunnan Province ◽

Management Practices ◽

Black Spot ◽

Parsimony Method ◽

Koch’S Postulates ◽

First Report ◽

The World ◽

Postharvest Rot ◽

Koch's Postulates

Apple is the largest fruit tree crop in the world, and China is the largest apple-producing County in the world. Zhaotong, Yunnan Province is a typical cold and mountainous apple-producing area in China. However, apple production is threatened by diseases during the entire growing season, and among them, apple leaf spot and fruit black spot are severe. In previous reports, the main pathogen causing apple leaf spot and fruit black spot was Alternaria sp. (Lior, et al, 2017), while different pathogens were identified. In the current study, seven red Fuji apple fruit with typical black spot samples were collected randomly in Dongda company orchard, Sujiayuan town, Zhaotong, Yunnan on March 25, 2021. The spots on the surface of these apples appear rounded, the diseased parts turn brown or black in colour and the flesh became soften and rotten. The tissues of fruit epidermis at the edge between diseased and healthy parts were cut, soaked in 75% alcohol for 30 s, washed with sterile water three times, and air-dried. Five pieces of tissue were placed on PDA medium amended with rifampicin (50 mg/ml) and incubated in the dark at 25 ℃ for 3-5 days. After colonies grew, mycelial clumps were picked out from the edges of the colonies, transferred to new PDA plates, and incubated at 25 ℃ for 6 days. The diameter of the colonies reached up to 5.7 cm. A representative isolate was retained for further work and was named P6-3-1. The hyphae were white and dense at an early stage, the culture medium on the underside became yellow and the middle parts of the colonies were darker. With maturity, hyphae were clumped, became red with other colors interspersed, and the medium became dark red. Light brown spores were produced, with more vertical septa and fewer transverse septa. Two to three transverse septa were generally observed with obvious constriction at the transverse septa. Average spore size was 22.83 µm ± 2.04 µm × 14.58 µm ± 1.97 µm. DNA was extracted from mycelium, purified and amplified with two pairs of primers, ITS1/ITS4 (White et al. 1990) and gpdF/gpdR (Marcos P. S. Câmara, et al. 2002). The PCR products were sequenced and deposited in GenBank (accession NO.OK560128 and OK627661 ). The similarity of ITS sequences between the isolate and MH843733 (Stemphylium eturmiunum strain ST14) was 100%, and that of gpd sequences between the isolate and MH843728 (Stemphylium eturmiunum strain ST20) was 100%. The maximum parsimony method of Mega7.0 was used and demonstrated that the studied isolate converged to the same branch as Stemphylium eturmiunum. Koch's postulates was applied to identify the pathogenicity of this isolate. A disc of P6-3-1-culture on PDA (5 mm in diameter) was placed on apple leaves and fruit wounds. Sterile PDA was used as a control. All plants were kept in a growth chamber at 25-30 ℃. Four days after inoculation, the disease spot was observed on the inoculated sites and fruit, and with the extension of incubation time, the diseased spots continue to grow, and the leaf spots were not limited by the veins. The pathogen was re-isolated from the inoculated leaves and fruit, satisfying Koch’s postulates. This pathogen can also cause postharvest rot of sweet cherry (Alice Spadoni, et al, 2020), postharvest rot on tomato (Prencipe Simona, et al, 2021), etc. This is the first report that Stemphylium eturmiunum can cause apple leaf spot and fruit black spot in Yunnan province, China. The apple black spot caused by Stemphylium eturmiunum was accurately identified. By distinguishing between the two similar diseases mentioned above, resistance to the host and management practices can be accrued based on the characteristics of the pathogen, its epidemiological pattern and the choice of an effective chemical fungicide.

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First report of Fusarium petroliphilum causing fruit rot of spaghetti squash in California

Plant Disease ◽

10.1094/pdis-08-21-1683-pdn ◽

2021 ◽

Author(s):

Albre Brown ◽

Marinell C Soriano ◽

Suzanne Rooney-Latham ◽

Cheryl L. Blomquist

Keyword(s):

Cucurbita Pepo ◽

Fruit Size ◽

Elongation Factor ◽

Fruit Rot ◽

Post Inoculation ◽

Koch’S Postulates ◽

First Report ◽

Light Yellow ◽

Fusarium Petroliphilum ◽

Koch's Postulates

Spaghetti squash (Cucurbita pepo L. subsp. pepo) is a yellow-skinned squash that forms translucent spaghetti-like strands when cooked. California leads the nation in total squash production, the majority of which is grown in the San Joaquin Valley. In October of 2019, severe fruit rot of C. pepo L. subsp. Pepo (C. pepo) was observed in fruit harvested from seven cultivated fields in San Joaquin County, California. Infected fields incurred up to 30% postharvest losses. At harvest, fruit appeared healthy. After ten days in a shaded storage shed, scattered buff to tan ringed lesions extending into the flesh of infected fruits were observed. Lesions had visible sporodochia at the center that were variable in size and continued to expand in storage. Tissue (∼1 mm3) from the lesion margins of symptomatic fruit (n=8) was surface sterilized in 75 % ethanol for 1 min then 0.6% sodium hypochlorite for a minute, and aseptically transferred to half strength acidified potato dextrose agar (0.5 APDA) and incubated at 22–25 °C. Fungal colonies which grew from the pieces were light yellow, with mycelium that was flat and mucoid. Sporodochial conidia were falcate and robust with 3 to 5 septa and measured from 44.2 to 51.6 × 4.6 to 5.9 μm (average 46.3 × 5.2 μm). Aerial conidia were profuse, borne on short monophialides, ovoid to reniform, and measured 5.1 to 12.6 μm × 3.2 to 5.6 μm (average 4.2 × 6.1 μm). DNA extracted from two isolates, was amplified with primers ITS1/ITS4, and EF1-728F/EF1-986R using PCR, to obtain sequences from the internal transcribed spacer (ITS) (White 1990), and elongation factor 1α (EF1α) (Carbone et al. 1999) genetic regions. Sequences from both isolates were identical. Sequences from isolate MVAP50001827, GenBank nos. MZ081401 (ITS) and MZ102267 (EF-1α) matched 100% to sequences of representative isolates of Fusarium petroliphilum (Q.T. Chen & X.H. Fu; Short et al., 2013, MB 802539) from Cucurbita species, MF535516 (ITS) and MF580776 (EF-1α) respectively (González, V. et al. 2018). To fulfill Koch’s postulates, conidia were harvested from a culture of isolate MVAP50001827 and grown for 7 days on 0.5 APDA at room temperature (22–25 °C). A 3-cc syringe with a 25-gauge needle was used to wound and inject 200 μl of 1 × 106 conidia ml–1 into three equally spaced points 1 mm deep into the rind of C. pepo fruit (n=4). C. pepo fruit (n=4) serving as negative controls were treated similarly with 200 μl of sterile deionized water. Fruit was incubated in a growth chamber at 27 °C under 12-h diurnal cycle lighting conditions. Ten days post inoculation, lesions densely covered with white sporodochia had expanded to 7 cm diameter and 5 cm deep on average (average fruit size 31×11 cm). Twenty days post inoculation, severe fruit rot was observed. F. petroliphilum did not grow from the controls, and was successfully reisolated from the symptomatic inoculated fruits, completing Koch’s postulates. Seeds inside the inoculated fruits were completely colonized and covered in conidia. Twenty-five seeds from the source seed lot was tested for F. petroliphilum by surface sterilizing and plating onto 0.5 APDA. No F. petroliphilum grew from tested seed. Postharvest fungal diseases can affect profitability of winter squash, which is often held in storage, and sold when market prices are optimal. To our knowledge, this is the first report of Fusarium petroliphilum infecting spaghetti squash (Cucurbita pepo L. subsp. pepo) in California.

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First Report of Botryosphaeria dothidea Causing Shoot Blight of Kiwifruit (Actinidia deliciosa) in Greece

Plant Disease ◽

10.1094/pdis-07-10-0528 ◽

2010 ◽

Vol 94 (12) ◽

pp. 1503-1503 ◽

Cited By ~ 4

Author(s):

T. Thomidis ◽

E. Exadaktylou

Keyword(s):

Random Amplified Polymorphic Dna ◽

Morphological Characteristics ◽

Fungal Mycelium ◽

Adhesive Tape ◽

Petroleum Jelly ◽

Koch’S Postulates ◽

First Report ◽

Agar Plug ◽

Dna Primers ◽

Koch's Postulates

In the spring of 2010, in commercial orchards located in the Prefecture of Pieria in northern Greece, wilted shoots of kiwifruit cv. Hayward were observed. Blighted shoots took on a distinct dark color. Isolations from the lower margins of the cankers were made by plating sodium-hypochlorite-treated shoot tissue sections of approximately 3 mm on acidified (2.5 ml of 85% lactic acid per liter of nutrient medium to create a pH = 3.5 after autoclaving) potato dextrose agar. Plates were incubated at 23°C for 5 days, and a fast-growing, mouse-gray colored fungus was consistently isolated from diseased stems. Identification of the pathogen was based on morphological characteristics and confirmed by using the four random amplified polymorphic DNA primers (K19 [CAC AGG CGG A], K20 [GTG TCG CGA G], R13 [GGA CGA CAA G], and R15 [GGA CAA CGA G], suggested by Ma et al. (2). This fungus formed darkly pigmented pycnidia (170 × 155 μm), while the conidia observed in these bodies were one-celled, hyaline, ellipsoidal to fusoid with distinctly truncate bases, and measured 10.9 to 21.55 × 3.25 to 10.10 μm. The pycnidia exuded conidia in white tendrils. Koch's postulates were completed in the laboratory by inoculating 20 segments (6 cm long and 1.5 to 2 cm in diameter) of 1-year-old woody shoots of kiwifruit cv. Hayward. Using a cork borer, a 7-mm-diameter wound was created in the middle of each shoot segment by removing the bark and a 6-mm-diameter agar plug bearing mycelia from a 15-day-old culture of B. dothidea was inserted into the wound. The wound was covered with petroleum jelly and wrapped with adhesive tape to prevent desiccation. Ten control segments were similarly wounded and inoculated with an agar disk without fungal mycelium. All inoculated and noninoculated shoot segments were incubated at 25°C in moist chambers, after which the resulting necrosis was recorded. Koch's postulates were satisfied after reisolating the fungus from inoculated shoots that developed symptoms similar to those observed on shoots collected from orchards. Although B. dothidea has been previously reported to cause dieback on kiwifruit in Japan (1), to our knowledge, this is the first report of the occurrence of B. dothidea on kiwifruit in Greece. This pathogen can cause a high level of shoot blights in diseased plants and presents a significant threat to the commercial kiwifruit production in Greece. References: (1) M. Kinugawa and T. Sato. Ann. Phytopathol. Soc. Jpn. 69:373, 2003. (2) Z. Ma et al. Phytopathology 91:665, 2001.

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First Report of Anthracnose Fruit Rot of Strawberry Caused by Colletotrichum acutatum in Montenegro

Plant Disease ◽

10.1094/pdis-02-12-0108-pdn ◽

2012 ◽

Vol 96 (7) ◽

pp. 1066-1066 ◽

Cited By ~ 1

Author(s):

J. Latinovic ◽

N. Latinovic ◽

J. Tiodorovic ◽

A. Odalovic

Keyword(s):

Morphological Characteristics ◽

Its Region ◽

Fruit Rot ◽

Colletotrichum Acutatum ◽

Pathogenicity Test ◽

Centraalbureau Voor ◽

First Report ◽

Strawberry Anthracnose ◽

Production Area ◽

Ascigerous Stage

Strawberries (Fragaria × ananassa) in Montenegro have become an increasingly important economic crop in recent years. During May 2011, severe fruit damage in strawberry cv. Clery was observed in two fields in the Podgorica region. Fruit symptoms were typical for strawberry anthracnose: sunken, dark brown to black circular lesions appeared on maturing fruits. However, no stem, crown, or foliar symptoms were observed. Under wet conditions, orange masses of conidia were produced in acervuli in the center of lesions. Conidia were hyaline, aseptate, cylindrical, with pointed ends, measuring 9.8 to 17.2 (mean 14.3) × 2.5 to 6.1 (mean 4.4) μm. Colonies on potato dextrose agar (PDA) were initially white, then turned gray as conidia formed in orange to salmon pink masses around the center of the culture. Setae or an ascigerous stage were never observed in culture or on the host. Koch's postulates were fulfilled by inoculating ripe and unripe asymptomatic fruits (20 of each, removed from strawberry plants cv. Clery) with the isolated fungus. Fruits were sprayinoculated (106 conidia/ml). An equal number of noninoculated fruits were used as a control. After incubation time of 2 to 3 days at 25°C in a moist chamber, symptoms appeared on inoculated ripe fruits. On unripe fruits, the lesions developed only 3 to 4 days after the inoculation. No symptoms were found on control fruits. The fungus was reisolated from fruits, after which typical morphological characteristics developed in culture as described above. On the basis of the symptoms, the morphological and cultural characteristics of the fungus, and the pathogenicity test, the disease was identified as strawberry anthracnose caused by Colletotrichum acutatum, which is in accordance with previous reports (1,2,3,4). The isolate was submitted to the Centraalbureau voor Schimmelcultures in the Netherlands (CBS 131813). The internal transcribed spacer (ITS) region of the fungal DNA was amplified with ITS1F and ITS4 primers, sequenced, and submitted to NCBI GenBank (Accession No. JQ424934). BLASTn searches of GenBank using the ITS sequence revealed 99% similarity with database sequences of C. acutatum. Since the pathogen was found in the main Montenegrin strawberry production area, it poses a threat to strawberry production in Montenegro. To our knowledge, this is the first report of anthracnose fruit rot of strawberry in Montenegro. References: (1) S. G. Bobev et al. Plant Dis. 86:1178, 2002. (2) F. M. Dai et al. Plant Dis. 90:1460, 2006. (3) U. Nilsson et al. Plant Dis. 89:1242, 2005. (4) A. Stensvand et al. Plant Dis. 85:558, 2001.

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First Report of Magnaporthe poae, Cause of Summer Patch Disease on Annual Bluegrass, in Canada

Plant Disease ◽

10.1094/pdis-06-12-0530-pdn ◽

2012 ◽

Vol 96 (11) ◽

pp. 1698-1698 ◽

Cited By ~ 3

Author(s):

M. M. I. Bassoriello ◽

K. S. Jordan

Keyword(s):

Morphological Characteristics ◽

Root Tissue ◽

Golf Course ◽

Koch’S Postulates ◽

First Report ◽

Annual Bluegrass ◽

Streptomycin Sulfate ◽

Magnaporthe Poae ◽

Patch Disease ◽

Koch's Postulates

The ectotrophic, root-infecting fungus Magnaporthe poae Landschoot & Jackson, the causal agent of summer patch disease in the U.S. (2), is implicated in the damage and loss of annual bluegrass (Poa annua L.) on golf course greens. This pathogenic fungus, one of the important root pathogens of turfgrass, attacks and colonizes susceptible turfgrass roots suffering from environmental or cultural stresses. Over 100 turf samples that exhibited symptoms (chlorotic circular or irregular patches of ≥15 cm in diameter with necrotic crowns and discolored roots) reminiscent of summer patch were collected from 77 southwestern Ontario golf courses from July to August of 2009 and 2010. Roots and crowns were often covered with dark, ectotrophic runner hyphae, lobed hyphopodia, and growth cessation structures, characteristic of M. poae. Sections of root tissue were surface sterilized in 0.6% sodium hypochlorite (NaOCl) for 5 min. Sterilized root tissue was plated on potato dextrose agar (PDA) containing 50 mg L–1 streptomycin sulfate and incubated at 28°C for 7 to 10 days. A fungus with morphological characteristics (hyaline mycelium that appears gray or olive-brown when mature) similar to those of M. poae (1) was consistently isolated (≥100 isolates were obtained) and used to identify M. poae through molecular techniques and Koch's postulates. DNA was extracted from the fungal mycelium of the collected isolates using the PowerPlant DNA isolation kit (MO BIO Laboratories, Inc., Carlsbad, CA). The rDNA internal transcribed spacer (ITS) regions of the isolates (≥100 isolates) were amplified by PCR using universal fungal rDNA primers ITS 4 (5′-TCCTCCGCTTATTGATATGC-3′) and ITS 5 (5′- GGAAGTAAAAGTCGTAACAAGG-3′) (3). The purified PCR products were sequenced (GenBank Accession No. JX134588 through JX134601) and a BLAST search exhibited seven isolates with 99% (MAG3, MAG6, MAG13, MAG16, and MAG17) and 100% (MAG1 and MAG14) similarity to M. poae in the GenBank database. Pathogenicity of four isolates (MAG1, MAG3, MAG6, and MAG14) was confirmed with Koch's postulates. Sixteen healthy P. annua core samples (four replicates of each treatment/isolate) collected from an Ontario golf course were inoculated with 25 mg M. poae-infested Kentucky bluegrass seed (Poa pratensis L.; 12.5 mg inoculum applied at the surface of the potting medium and 12.5 mg inoculum applied on the foliar surface) and were placed in a growth chamber with 12-h day/night cycles at 30/25°C and approximate relative humidity. After 2 to 3 weeks, inoculated plants exhibited chlorotic foliage and necrotic roots covered with dark ectotrophic runner hyphae and lobed hyphopodia. Infected root sections from each replication were surface sterilized and placed on PDA containing 50 mg L–1 streptomycin sulfate. The fungal cultures exhibited morphological characteristics consistent with M. poae (1). To our knowledge, this is the first report of summer patch caused by M. poae in Canada. References: (1) B. B. Clarke and A. B. Gould, eds. Turfgrass Patch Diseases Caused by Ectotrophic Root-Infecting Fungi. The American Phytopathological Society, St. Paul, MN, 1993. (2) P. J. Landschoot and N. Jackson. Mycol. Res. 93:59, 1989. (3) T. J. White et al. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. Pages 315-322 in: PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al. eds. Academic Press, San Diego, CA, 1990.

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