First Report of Myrothecium Leaf Spot of Hemionitis arifolia Caused by Myrothecium roridum in China

Hemionitis arifolia (Burm.) Moore. was grown commercially as an ornamental plant in China. In January 2010, a new foliar disease with typical leaf spot symptoms was observed on H. arifolia in Dongguan City, Guangdong Province. Approximately 10% of the plants in the Dongguan nursery were affected. Leaf spots were circular to subcircular, dark brown, with distinct concentric zones, and ranged from 10 to 20 mm in diameter. Lesions developed mostly on the lower leaves and black sporodochia with white mycelial tufts were produced mostly in older lesions under high humidity. Single-spore isolates from lesions plated on potato dextrose agar (PDA) produced white, floccose colonies and dark green-to-black sporodochia. Colonies reached 60 mm on PDA at 25°C after 14 days. Conidiophores branched repeatedly. Conidiogenous cells in whorls of two to six on ultimate branches were hyaline, cylindrical, and 13 to 20 × 1.2 to 1.8 μm. Conidia were hyaline, cylindrical, mostly with both rounded ends, occasionally one blunt end, and mean size was 6.1 (4.5 to 7.0) × 2.3 (1.8 to 3.0) μm. These characteristics were consistent with the descriptions of Myrothecium roridum Tode ex Fr. (2–4). The internal transcribed spacer regions including ITS1, ITS2, and the 5.8S rRNA of one isolate were PCR amplified and sequenced. A BLAST search in GenBank revealed highest similarity (99%) to M. roridum sequences from isolates collected from Germany (Accession Nos. AJ302001.1 and AJ301995.1). The M. roridum sequence from the Chinese isolate was submitted to GenBank (Accession No. JF343832). To confirm pathogenicity, five leaves on five H. arifolia plants were inoculated with 5-mm mycelial plugs from the periphery of 7-day-old cultures; sterile PDA plugs were used as the control treatment. Plants were covered with plastic bags and incubated in a growth chamber at 28°C. Necrotic lesions appeared 2 to 3 days after inoculation and the symptoms described above were observed after 7 days on all inoculated leaves, whereas sterile PDA plugs did not produce any leaf lesion. The pathogen was reisolated from the inoculated leaves and confirmed to be M. roridum on the basis of morphological characteristics. There are approximately 271 hosts of M. roridum (1), including many ornamental plants such as salvia (2) and garden hydrangea (3). To our knowledge, this is the first report of Myrothecium leaf spot on H. arifolia. Because the disease caused damage to the foliage and reduced the ornamental value of H. arifolia plants, control measures may need to be implemented for production of this species in ornamental nurseries. References: (1) D. F. Farr and A. Y. Rossman. Fungal Databases. Systematic Mycology and Microbiology Laboratory, ARS, USDA. Retrived from http://nt.ars-grin.gov/fungaldatabases/ , 6 February 2011. (2) J. A. Mangandi et al. Plant Dis. 91:772, 2007. (3) M. T. Mmbaga et al. Plant Dis. 94:1266, 2010. (4) M. Tulloch. Mycol. Pap. No. 130. CMI, Wallingford, UK, 1972.

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First Report of Alternaria Leaf Spot of Banana Caused by Alternaria alternata in the United States

Plant Disease ◽

10.1094/pdis-01-13-0007-pdn ◽

2013 ◽

Vol 97 (8) ◽

pp. 1116-1116 ◽

Cited By ~ 3

Author(s):

V. Parkunan ◽

S. Li ◽

E. G. Fonsah ◽

P. Ji

Keyword(s):

United States ◽

Alternaria Alternata ◽

Leaf Spot ◽

Morphological Characteristics ◽

The United States ◽

Its Sequencing ◽

Single Spore ◽

First Report ◽

Alternaria Leaf Spot ◽

Leaf Spots

Research efforts were initiated in 2003 to identify and introduce banana (Musa spp.) cultivars suitable for production in Georgia (1). Selected cultivars have been evaluated since 2009 in Tifton Banana Garden, Tifton, GA, comprising of cold hardy, short cycle, and ornamental types. In spring and summer of 2012, 7 out of 13 cultivars (African Red, Blue Torres Island, Cacambou, Chinese Cavendish, Novaria, Raja Puri, and Veinte Cohol) showed tiny, oval (0.5 to 1.0 mm long and 0.3 to 0.9 mm wide), light to dark brown spots on the adaxial surface of the leaves. Spots were more concentrated along the midrib than the rest of the leaf and occurred on all except the newly emerged leaves. Leaf spots did not expand much in size, but the numbers approximately doubled during the season. Disease incidences on the seven cultivars ranged from 10 to 63% (10% on Blue Torres Island and 63% on Novaria), with an average of 35% when a total of 52 plants were evaluated. Six cultivars including Belle, Ice Cream, Dwarf Namwah, Kandarian, Praying Hands, and Saba did not show any spots. Tissue from infected leaves of the seven cultivars were surface sterilized with 0.5% NaOCl, plated onto potato dextrose agar (PDA) media and incubated at 25°C in the dark for 5 days. The plates were then incubated at room temperature (23 ± 2°C) under a 12-hour photoperiod for 3 days. Grayish black colonies developed from all the samples, which were further identified as Alternaria spp. based on the dark, brown, obclavate to obpyriform catenulate conidia with longitudinal and transverse septa tapering to a prominent beak attached in chains on a simple and short conidiophore (2). Conidia were 23 to 73 μm long and 15 to 35 μm wide, with a beak length of 5 to 10 μm, and had 3 to 6 transverse and 0 to 5 longitudinal septa. Single spore cultures of four isolates from four different cultivars were obtained and genomic DNA was extracted and the internal transcribed spacer (ITS1-5.8S-ITS2) regions of rDNA (562 bp) were amplified and sequenced with primers ITS1 and ITS4. MegaBLAST analysis of the four sequences showed that they were 100% identical to two Alternaria alternata isolates (GQ916545 and GQ169766). ITS sequence of a representative isolate VCT1FT1 from cv. Veinte Cohol was submitted to GenBank (JX985742). Pathogenicity assay was conducted using 1-month-old banana plants (cv. Veinte Cohol) grown in pots under greenhouse conditions (25 to 27°C). Three plants were spray inoculated with the isolate VCT1FT1 (100 ml suspension per plant containing 105 spores per ml) and incubated under 100% humidity for 2 days and then kept in the greenhouse. Three plants sprayed with water were used as a control. Leaf spots identical to those observed in the field were developed in a week on the inoculated plants but not on the non-inoculated control. The fungus was reisolated from the inoculated plants and the identity was confirmed by morphological characteristics and ITS sequencing. To our knowledge, this is the first report of Alternaria leaf spot caused by A. alternata on banana in the United States. Occurrence of the disease on some banana cultivars in Georgia provides useful information to potential producers, and the cultivars that were observed to be resistant to the disease may be more suitable for production. References: (1) E. G. Fonsah et al. J. Food Distrib. Res. 37:2, 2006. (2) E. G. Simmons. Alternaria: An identification manual. CBS Fungal Biodiversity Center, Utrecht, Netherlands, 2007.

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First Report of Myrothecium roridum Causing Leaf Spot on Garden Hydrangea in the United States

Plant Disease ◽

10.1094/pdis-04-10-0264 ◽

2010 ◽

Vol 94 (10) ◽

pp. 1266-1266 ◽

Cited By ~ 7

Author(s):

M. T. Mmbaga ◽

Y. Li ◽

M.-S. Kim

Keyword(s):

United States ◽

Leaf Spot ◽

Incubation Temperature ◽

Morphological Characteristics ◽

The United States ◽

Control Treatment ◽

Medium Size ◽

First Report ◽

Detached Leaves ◽

Myrothecium Roridum

Garden hydrangea (Hydrangea macrophylla) is a popular flowering shrub that grows well in Tennessee but foliar diseases impact their appearance, health, and market value. Leaves of garden hydrangea showed necrotic lesions with concentric rings of brown and dark brown at the Tennessee State University Research Center in McMinnville. A fungus was recovered from June and July leaf samples with 20% frequency of isolation from approximately 40 leaf pieces that were surface sterilized and plated in potato dextrose agar (PDA). Isolates developed white colonies and dark gray-to-black, spore-bearing mycelial cushions (sporodochia) that formed on older colonies (30 to 45 days old) at 25 ± 2°C. Conidia were hyaline to slightly dark, one-celled, ovoid to elongate with rounded ends, and 2.0 to 2.5 × 5.5 to 6.5 μm. These morphological characteristics were consistent with those described for Myrothecium roridum Tode ex Fr. (1). DNA sequence for three isolates of this fungus showed identical internal transcribed spacer (ITS) region sequences (GenBank Accession No. HM215150) with 99% maximum sequence identity to M. roridum isolates (GenBank Accession Nos. AJ301994.1 and AJ608978). Another close match (97%) was with M. gramineum (GenBank Accession No. FJ235084) and M. tongaense (GenBank Accession No. AY254157). Pathogenicity of M. roridum was evaluated on detached leaves from three hydrangea cultivars, Nikko Blue, All Summer Beauty, and Blue bird. Four, medium-size, detached leaves were placed in moist chambers and inoculated with 5-mm mycelial plugs from 14-day-old cultures; sterile PDA was used as the control treatment. A randomized, complete-block experimental design was used with a replication of four leaves per cultivar. Incubation temperature was 26 ± 2°C. Necrotic lesions started 4 to 5 days after inoculation in all inoculated leaves; lesions expanded to cover 10 to 25% of the leaf surface and formed concentric rings; sterile PDA plugs did not produce leaf lesions. This experiment was repeated twice and similar symptoms were produced; M. roridum was reisolated from all inoculated leaves. Spray inoculation of detached leaves of hydrangea cv. Pretty Maiden with 5 × 104 spores/ml produced similar symptoms; leaves sprayed with water remained symptom free. M. roridum has a wide host range and similar symptoms have been reported on other ornamentals including salvia (2), begonia ( http://mrec.ifas.ufl.edu/foliage/folnotes/begonias.htm ), gardenia ( http://cfextension.ifas.ufl.edu/agriculture/ nursery_production/ documents/Gardenia.pdf ), and cotton (3). To our knowledge, this is the first report of M. roridum causing leaf spot on H. macrophylla in the United States. References: (1) M. B. Ellis. Page 465 in: More Damatacous Hyphomycetes. CABI, Wallingford, UK. 1993. (2) J. A. Mangandi et al. Plant Dis. 91:772, 2007. (3) R. L. Munjal. Indian Phytopathol. New Delhi, 13:150, 1960.

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First Report of Leaf Spot on Gerbera jamesonii Caused by Corynespora cassiicola in China

Plant Disease ◽

10.1094/pdis-12-11-1081-pdn ◽

2012 ◽

Vol 96 (6) ◽

pp. 915-915

Author(s):

Z. R. Shi ◽

M. M. Xiang ◽

Y. X. Zhang ◽

J. H. Huang

Keyword(s):

Leaf Spot ◽

Control Measures ◽

Control Treatment ◽

Leaf Spot Disease ◽

Corynespora Cassiicola ◽

Gerbera Jamesonii ◽

Cut Flower ◽

Single Spore ◽

Internal Transcribed Spacer Region ◽

First Report

Gerbera (Gerbera jamesonii Bolus ex. Hook f.) is a popular cut flower and flowering potted plant. In August 2011, a new leaf spot disease was observed on double-type Gerbera growing in outdoor ground beds in Guangzhou, Guangdong Province, China. Approximately 30% of about 20,000 Gerbera plants in the Guangzhou ground beds were affected. Leaf spots were round or irregular with grayish centers surrounded by dark brown borders and ranged from 5 to 15 mm in diameter. Leaves with multiple lesions became blighted. A fungus was isolated from the lesions and single-spore isolates plated on potato dextrose agar (PDA) produced gray, floccose colonies, which reached 65 mm on PDA after 7 days at 28°C. Conidiophores were brown or olivaceous, cylindrical, straight and unbranched, two to seven septations, and 25 to 83 × 4 to 7 μm. Conidiogenous cells were olivaceous or brown, cylindrical, and 11 to 21 × 4 to 6 μm. Conidia were borne singly or in chains of two to five, brown, cylindrical, straight to slightly curved, two to eight pseudosepta, and 30 to 90 × 5.5 to 11.5 μm (mean 70.4 × 7.3 μm), with a conspicuous hilum. These characteristics were consistent with the description of Corynespora cassiicola (Berk. & M.A. Curtis.) C.T. Wei (1). The internal transcribed spacer region (ITS) of one isolate (GenBank Accession No. JN853778) was amplified using primers ITS4 and ITS5 (3) and sequenced. A BLAST search in GenBank revealed highest similarity (99%) to sequences of C. cassiicola (AY238606.1 and FJ852715.1). Pathogenicity tests were conducted on 10 potted double-type Gerbera plants. Five wounded and five unwounded leaves on each plant were inoculated with 5-mm mycelial plugs from the periphery of 5-day-old cultures of the isolated fungus. The plugs were put on the leaf surface and secured with sterile wet cotton. Sterile PDA plugs were used as the control treatment on different leaves of the same plants that were inoculated. Plants were covered with plastic bags and incubated in a growth chamber with 12 h of light at 28°C. Necrotic lesions appeared on wounded leaves after 2 to 3 days of incubation and on unwounded leaves 5 to 7 days after incubation. Symptoms on wounded and unwounded leaves were similar to those observed in the field, whereas control leaves inoculated with sterile PDA plugs remained symptomless. C. cassiicola was consistently reisolated from these lesions. Although there are approximately 644 reported hosts of C. cassiicola (2), to our knowledge, this is the first report of C. cassiicola leaf spot on G. jamesonii. Because the disease caused damage to the foliage and affected the flowering of the plants, control measures may need to be implemented for the production of Gerbera in cut flower nurseries. References: (1) M. B. Ellis. CMI Mycol. Pap. 65:15, 1957. (2) D. F. Farr and A. Y. Rossman. Fungal Databases, Systematic Mycology and Microbiology Laboratory, ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/ , 21 November 2011. (3) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, San Diego, 1990.

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First Report of Corynespora cassiicola Causing Leaf Spot of Cassava in China

Plant Disease ◽

10.1094/pdis-94-7-0916a ◽

2010 ◽

Vol 94 (7) ◽

pp. 916-916 ◽

Cited By ~ 1

Author(s):

X.-B. Liu ◽

T. Shi ◽

C.-P. Li ◽

J.-M. Cai ◽

G.-X. Huang

Keyword(s):

Leaf Spot ◽

Morphological Characteristics ◽

Its Region ◽

Pathogenic Fungi ◽

Commonwealth Mycological Institute ◽

Corynespora Cassiicola ◽

Single Spore ◽

First Report ◽

Leaf Spots ◽

Main Vein

Cassava (Manihot esculenta) is an important economic crop in the tropical area of China. During a survey of diseases in July and September of 2009, leaf spots were observed on cassava plants at three separate plantations in Guangxi (Yunfu and Wuming) and Hainan (Baisha) provinces. Circular or irregular-shaped leaf spots were present on more than one-third of the plants. Spots were dark brown or had white papery centers delimited by dark brown rims and surrounded by a yellow halo. Usually, the main vein or small veinlets adjacent to the spots were dark. Some defoliation of plants was evident at the Wuming location. A fungus was isolated from symptomatic leaves from each of the three locations and designated CCCGX01, CCCGX02, and CCCHN01. Single-spore cultures of these isolates were incubated on potato dextrose agar (PDA) for 7 days with a 12-h light/dark cycle at a temperature of 28 ± 1°C. Conidiophores were straight to slightly curved, unbranched, and pale to light brown. Conidia were formed singly or in chains, obclavate to cylindrical, straight or curved, subhyaline-to-pale olivaceous brown, 19.6 to 150.3 μm long and 5.5 to 10.7 μm wide at the base, with 4 to 13 pseudosepta. Morphological characteristics of the specimen and their conidia were similar to the descriptions for Corynespora cassiicola (2). The isolate CCCGX01 was selected as a representative for molecular identification. Genomic DNA was extracted by the cetyltrimethylammoniumbromide protocol (3) from mycelia and used as a template for amplification of the internal transcribed spacer (ITS) region of rDNA with primer pair ITS1/ITS4. The sequence (GenBank Accession No. GU138988) exactly matched several sequences (e.g., GenBank Accession Nos. FJ852715, EF198117, and AY238606) of C. cassiicola (1). Young, healthy, and fully expanded green leaves of cassava cv. SC205 were surface sterilized. Ten leaves were inoculated with 10-μl drops of 104 ml suspension of conidia and five leaves were inoculated with the same volume of sterile water to serve as controls. After inoculation, leaves were placed in a dew and dark chamber for 36 h at 25°C and subsequently transferred to the light for 5 days. All inoculated leaves with isolates showed symptoms similar to those observed in natural conditions, whereas the controls remained symptom free. The morphological characteristics of reisolated conidia that formed on the diseased parts were identical with the nature isolates. To our knowledge, this is the first report of leaf spot caused by C. cassiicola on cassava in China. References: (1) L. J. Dixon et al. Phytopathology 99:1015, 2009. (2) M. B. Ellis et al. Corynespora cassiicola. No. 303 in: CMI Description of Pathogenic Fungi and Bacteria. Commonwealth Mycological Institute, Kew, UK 1971. (3) J. R. Xu et al. Genetics 143:175, 1996.

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First Report of Leaf Spot of Soybean Caused by Aristastoma guttulosum in China

Plant Disease ◽

10.1094/pdis-05-12-0454-pdn ◽

2012 ◽

Vol 96 (11) ◽

pp. 1694-1694

Author(s):

X. F. Zhu ◽

Y. Pan ◽

L. J. Chen ◽

Y. X. Duan ◽

Y. Y. Wang

Keyword(s):

Leaf Spot ◽

Chinese Isolate ◽

Its Sequence ◽

Light Cycle ◽

Single Spore ◽

Internal Transcribed Spacer Region ◽

Dark Light ◽

First Report ◽

Leaf Spots ◽

Soybean Leaves

In fall, 2008, leaf spots were observed during the flowering stage of the Zhong Huang 13 cultivar of soybean in the fields of Anhui Province, China. The leaf spots were irregularly shaped, necrotic, brown-black, and surrounded by yellow halos. Often, on a given leaf, several spots joined one another to form a large blighted area. Finally, those leaves turned yellow followed by defoliation. Damaged leaves showed scattered black spots (i.e., numerous pycnidia) on the lower side. Fresh material was collected from infected plants and a single spore of the putative causal pathogen was isolated on potato dextrose agar (PDA) and incubated at 25°C during a 12-h dark/light cycle. The isolate produced a white fungal colony and black pycnidia after 30 days. The pycnidia are characterized as globose, dark brown-black, and distinctly papillate, with ostiolar setae, and are more or less straight, unbranched, and tapered at the apex. The conidia are clavate, hyaline, mostly with three transverse septa per cell; conidia are either straight or slightly bent, obviously guttulate, and 16 to 29 × 2.5 to 3.5 μm. This pathogen is similar to other Aristastoma guttulosum Sutton (1964), but with the following differences: (a) it has more than 10 versus 4 to 9 setae; (b) conidia are 16 to 29 × 2.5 to 3.5 μm versus 32 to 42 × 3.9 to 4.6 μm as reported for A. guttulosum (1). Conidia of the Chinese isolate were used to inoculate leaves of soybean. Five soybean leaves from potted plants, 1 month old, were sprayed with a suspension of conidia in water. Conidia were harvested from PDA cultures and the suspension was adjusted to 3 × 105 conidia/ml with a hemocytometer. Five leaves were sprayed with sterile distilled water as controls. Inoculated plants were kept in the greenhouse. All five of the inoculated leaves displayed the same symptoms observed in the fields. The symptoms developed initially as brown pinhead spots on the upper side of the leaves, gradually increasing to large brown spots. These spots were irregularly shaped, brown and necrotic in the center and surrounded by a yellow halo. Black pycnidia appeared after 1 week whereas the controls remained asymptomatic. The pathogen was reisolated from the inoculated soybean leaves according to standard Koch's postulates. Primers ITS1 and ITS4 were used in PCR reactions to amplify the internal transcribed spacer region (ITS) (3). Sequencing was performed using the same primers. The ITS sequence (GenBank Accession No. JF825548.1) for this pathogen (587 bp) was submitted to a BLAST search in GenBank. Since the ITS sequence of the genus Aristastoma has never been previously submitted, results did not show high similarity with any extant GenBank sequences. The genus Aristastoma Tehon (1933) was described by Tehon (2). Five of the species in this genus were described by Sutton (1). The number of septate conidium and lack of obvious guttulate within the conidium are the morphological basis to separate these five species. Morphological features of the pathogen from soybean leaves in China were slightly different from those of A. guttulosum. To our knowledge, this is the first report of leaf spot caused by A. guttulosum on soybean in China. References: (1) B. C. Sutton. Mycological Papers. 97:10, 1964. (2) L. R. Tehon. Mycologia XXV. 25:249, 1933. (3) T. J. White et al. Academic Press, San Diego, 1990.

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First Report of Pestalotiopsis microspora Causing Leaf Spot of Hidcote (Hypericum patulum) in Japan

Plant Disease ◽

10.1094/pdis-94-8-1064b ◽

2010 ◽

Vol 94 (8) ◽

pp. 1064-1064 ◽

Cited By ~ 3

Author(s):

M. Zhang ◽

H. Y. Wu ◽

T. Tsukiboshi ◽

I. Okabe

Keyword(s):

Leaf Spot ◽

Morphological Characteristics ◽

Its Region ◽

Basal Cells ◽

Single Spore ◽

Management Options ◽

First Report ◽

Leaf Spots ◽

Plastic Bags ◽

Pestalotiopsis Microspora

Hidcote, Hypericum patulum Thunb. ex Murray, is a deciduous shrub that is cultivated as an ornamental in landscape gardens and courtyards in Japan. In early August 2008, severe leaf spotting was observed on plants growing in a courtyard in Nasushiobara, Tochigi, Japan. More than 30% of the leaves on five shrubs exhibited leaf spot symptoms. Small, round, pale brown lesions were initially observed. Later, they expanded to 5 to 12 mm in diameter, round to irregular-shaped with pale brown centers and dark brown margins. Under continuously wet or humid conditions, black acervuli developed on the leaf lesions. Conidia were straight or slightly curved, fusiform to clavate, and five-celled with constrictions at the septa. Conidia ranged from 17 to 21 × 5 to 8 μm with hyaline apical and basal cells. Fifteen percent of apical cells had two and the rest had three appendages (setulae) ranging from 10 to 21 μm long. The basal hyaline cell tapered into a 2 to 4 μm pedicel. The three median cells ranged from light or dark brown to olive green. These morphological characteristics matched those of Pestalotiopsis microspora (Speg.) G.C. Zhao & N. Li (1,2). The identity of the fungus was confirmed by DNA sequencing of the internal transcribed spacer (ITS) region (GenBank Accession No. GU908473) from single-spore isolates, which revealed 100% homology with those of other P. microspora isolates (e.g., GenBank Accession Nos. FJ459950 and DQ456865). Koch's postulates were confirmed using leaves of three detached branches of a field-grown asymptomatic plant of H. patulum. Thirty leaves of each branch were inoculated by placing mycelial plugs obtained from the periphery of 7-day-old single-spore cultures on the leaf surface. Potato dextrose agar plugs without mycelium served as controls. Leaves on branches were covered with plastic bags for 24 h to maintain high relative humidity in a greenhouse (approximately 24 to 28°C). After 5 days, all inoculated leaves showed symptoms identical to those described above, whereas control leaves remained symptom free. Reisolation of the fungus from lesions on inoculated leaves confirmed that the causal agent was P. microspora. To our knowledge, this is the first report of leaf spots on H. patulum caused by P. microspora in Japan. Management options may have to be developed and implemented to protect Hidcote plants in areas where leaf spot cannot be tolerated. References: (1) P. A. Saccardo. Sylloge Fungorum III:789, 1884. (2) G. C. Zhao and N. Li. J. Northeast For. Univ. 23(4):21, 1995.

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First Report of a Leaf Spot on Hazel Leaves Caused by Phyllosticta coryli in Liaoning Province of China

Plant Disease ◽

10.1094/pdis-02-13-0213-pdn ◽

2013 ◽

Vol 97 (9) ◽

pp. 1254-1254 ◽

Cited By ~ 10

Author(s):

J. Sun ◽

D.-M. Wang ◽

X.-Y. Huang ◽

Z.-H. Liu

Keyword(s):

Leaf Spot ◽

Control Strategies ◽

Morphological Characteristics ◽

Leaf Tissue ◽

Its Region ◽

Liaoning Province ◽

Single Spore ◽

First Report ◽

Leaf Spots ◽

Microscopic Morphology

Hazel (Corylus heterophylla Fischl) is an important nut tree grown in China, especially in Liaoning Province, and is rich in nutritional and medicinal values. In August 2011, leaf spotting was observed on hybrid hazel (Dawei) leaves in Paotai Town, Wafangdian County of Liaoning Province. By August 2012, the disease had spread to Zhangdang Town, Fushun County. Symptoms initially appeared on both sides of leaves as pinpoint brown spots, which enlarged and developed into regular, dark brown lesions, 3 to 9 mm in diameter. The lesions were lighter in color in the center compared to the margin. To identify the pathogen, leaf pieces (3 to 5 mm) taken from the margins, including both symptomatic and healthy portions of leaf tissue, were surface-disinfected first in 75% ethanol for 5 s, next in 0.1% aqueous mercuric chloride for 50 s, and then rinsed with sterilized water three times. Leaf pieces were incubated on potato dextrose agar (PDA) at 25°C for 14 days in darkness. Single spore isolates were obtained from individual conidia. For studies of microscopic morphology, isolates were grown on synthetic nutrient agar (SNA) in slide cultures. Colonies grew up to 45 to 48 mm in diameter on PDA after 14 days. Pycnidia appeared on the colonies after 12 days. Conidiophores were short. Pycnidia were dark brown, subglobose, and 150 to 205 μm in diameter. Conidia were unicellular, colorless, ovoid to oval, and from 2.4 to 4.5 × 1.6 to 2.4 μm. On the basis of these morphological characteristics, the isolates were tentatively identified as Phyllosticta coryli Westend (2). The rDNA internal transcribed spacer (ITS) region was amplified using primers ITS1 and ITS4 and sequenced (GenBank Accession No. KC196068). The 490-bp amplicons had 100% identity to an undescribed Phyllosticta species isolated from Cornus macrophylla in Gansu, Tianshui, China (AB470897). On the basis of morphological characteristics and nucleotide homology, the isolate was tentatively identified as P. coryli. Koch's postulates were fulfilled in the growth chamber on hazelnut leaves inoculated with P. coryli conidial suspensions (107 conidia ml–1). Eight inoculated 1-year-old seedlings (Dawei) were incubated under moist conditions for 8 to 10 days at 25°C. All leaf spots that developed on inoculated leaves were similar in appearance to those observed on diseased hazel leaves in the field. P. coryli was recovered from lesions and its identity was confirmed by morphological characteristics. P. coryli was first reported as a pathogen of hazel leaves in Bull of Belgium (2). In China, P. coryli was first reported on Corylus heterophylla Fisch. in Jilin Province (1). To our knowledge, this is the first report of P. coryli causing leaf spot on hybrid hazel in Liaoning Province of China. The outbreak and spread of this disease may decrease the yield of hazelnut in northern regions of China. More studies are needed on control strategies, including the possible resistance of hazel cultivars to P. coryli. References: (1) Y. Li et al. J. Shenyang Agric. Univ. 25:153, 1994. (2) P. A. Saccardo. Sylloge Fungorum Vol. III, page 31, 1884.

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First Report of Leaf Spot on Tibouchina semidecandra Caused by Beltrania rhombica in China

Plant Disease ◽

10.1094/pdis-04-12-0363-pdn ◽

2012 ◽

Vol 96 (9) ◽

pp. 1380-1380 ◽

Cited By ~ 1

Author(s):

Z. R. Shi ◽

M. M. Xiang ◽

Y. X. Zhang ◽

J. H. Huang

Keyword(s):

Leaf Spot ◽

Its Region ◽

Control Measures ◽

Control Treatment ◽

Sterile Water ◽

Single Spore ◽

First Report ◽

Potted Plants ◽

Detached Leaves

Tibouchina semidecandra Cogn. is a popular ornamental plant in tropical and subtropical areas (1). In August 2011, a leaf spot was observed on approximately 70% of 5,000 potted plants of T. semidecandra in a nursery in Zhongshan, Guangdong Province, China. Each leaf spot was round with a brown center surrounded by a reddish brown border, and ranged from 8 to 10 mm in diameter. A fungus was isolated consistently from the lesions by surface-sterilizing symptomatic leaf sections (each 3 cm2) with 75% alcohol for 8 s, washing the sections with sterile water, soaking the sections in 3% NaOCl for 15 s, rinsing the sections with sterile water three times, and then placing the sections on potato dextrose agar (PDA) at 28°C. Each of three single-spore isolates on PDA produced gray, floccose colonies that reached 70 mm in diameter after 5 days at 28°C. Setae were dark brown, straight, erect, distantly and inconspicuously septate, and 125 to 193 × 3.0 to 4.5 μm. Conidiophores were light brown, cylindrical, simple or sometimes branched at the base, and 105 to 202 × 3 to 5 μm. Separating cells were hyaline, oval, and 12 to 13 × 4 to 5 μm. Conidia were unequally biconic, unicellular, dark brown with a pale brown or subhyaline band just above the widest part, and 26 to 31 × 8.5 to 12 μm (mean 27.3 × 10.6 μm) with a conspicuous appendage at the apex that was 6 to 14 × 1 to 1.8 μm. These characteristics were consistent with the description of Beltrania rhombica Penz. (3). The internal transcribed spacer (ITS) region of the ribosomal DNA (rDNA) of one isolate (GenBank Accession No. JN853777) was amplified using primers ITS4 and ITS5 (4) and sequenced. A BLAST search in GenBank revealed 97% similarity to the ITS sequence of an isolate of B. rhombica (GU797390.1). To confirm pathogenicity of the isolate, ten detached leaves from 3-month-old plants of T. semidecandra ‘Purple Glorybush’ were inoculated in vitro with 5-mm diameter, colonized mycelial plugs from the periphery of 5-day-old cultures of the isolated fungus. The agar plugs were put on the leaf surface and secured with sterile, moist cotton. Sterile PDA plugs were similarly used as the control treatment on ten detached leaves. Leaves were placed in petri dishes and incubated in a growth chamber with 12 h of light/day at 28°C. Necrotic lesions appeared on leaves after 2 to 3 days of incubation, whereas control leaves inoculated with sterile PDA plugs remained asymptomatic. B. rhombica was consistently reisolated from the lesions using the same method described above, but was not reisolated from the control leaves. Although there are approximately 77 reported hosts of B. rhombica (2), to our knowledge, this is the first report of B. rhombica causing a leaf spot on T. semidecandra. Because the disease caused foliar damage and reduced the ornamental value of the nursery plants, control measures may need to be implemented for this species in nurseries. References: (1) M. Faravani and B. H. Bakar. J. Food Agric. Env. Pap. 5:234, 2007. (4) D. F. Farr and A. Y. Rossman. Fungal Databases. Systematic Microbiology Laboratory, ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/ , 30 Mar. 2012. (2) K. A. Pirozyski and S. D. Patil. Can. J. Bot. Pap. 48:567, 1970. (3) T. J. White et al. Page 315 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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First Report of Leaf Spot Disease Caused by Cercosporella pfaffiae on Brazilian Ginseng (Pfaffia glomerata) in Brazil

Plant Disease ◽

10.1094/pdis-06-12-0614-pdn ◽

2012 ◽

Vol 96 (11) ◽

pp. 1702-1702 ◽

Cited By ~ 2

Author(s):

A. R. Machado ◽

D. B. Pinho ◽

M. Silva ◽

O. L. Pereira

Keyword(s):

Leaf Spot ◽

Morphological Characteristics ◽

Leaf Spot Disease ◽

Single Spore ◽

First Report ◽

Pfaffia Glomerata ◽

Leaf Spots ◽

Agar Plug ◽

Brazilian Ginseng ◽

Two Samples

Pfaffia glomerata (Spreng) Pedersen (Amaranthaceae) and other species in this genus, popularly known as “Brazilian ginseng,” have been marketed and used for many years in folk medicine for the treatment of various diseases (1). In January 2012, samples of P. glomerata with leaf spots were collected in the city of Viçosa, state of Minas Gerais, Brazil. Two samples were deposited in the herbarium at the Universidade Federal de Viçosa (VIC31849 and VIC31851). The diseased leaves were examined using a stereomicroscope (75×). The fungal structures were scraped with a scalpel from the plant surface and mounted in lactophenol. Thirty measurements of all of the relevant morphological characters were obtained using light microscopy for the identification of the species. To confirm the identification, fungal DNA from single-spore pure culture was isolated from the diseased leaves on PDA, and the DNA was amplified using primers ITS1 and ITS4 for the ITS region (GenBank Accession No. JQ990331) and LR0R and LR5 for partial 28S rDNA (Accession No. JQ990330). Sequencing was performed by Macrogen, Korea. The symptoms observed were leaf spots, subcircular, usually up to 6 mm diameter, initially yellowish becoming brown to reddish, margin indefinite, with the formation of fungal structures, hypophyllous, white, scattered, or grouped. Conidiophores were very numerous in dense subsynnematal fascicles, moderately brown at the base but for most of the length subhyaline, 42.5 to 350 × 2.5 to 3.5 μm, showing conidial scars. Conidia formed singly, 22.5 to 77.5 × 5 to 6 μm, hyaline, hilum slightly thickened, and refractive. These characteristics show that the fungus found on P. glomerata matched well with the description of Cercosporella pfaffiae (2). Koch's postulates were fulfilled by inoculation of 6-mm-diameter PDA plugs with the isolate mycelia on leaves of P. glomerata. Six plants were inoculated with the isolate and six plants were inoculated with an isolate-free agar plug. Inoculated plants were maintained in a moist chamber for 24 hours and subsequently in a greenhouse at 26°C. Leaf spot was observed in inoculated plants 15 days after inoculation, and symptoms were similar to those in the field. All non-inoculated plants remained healthy. A Megablast search of the NCBI GenBank nucleotide sequence database using the ITS sequence retrieved C. virgaureae as the closest match [GenBank GU214658; Identity = 458/476 (96%), Gaps = 2/476 (0%)]. To confirm the identification, Bayesian inference analyses were employed, and the tree was deposited in TreeBASE (Study S12680). The analysis placed our isolate in the same clade with the type species of Cercosporella. Molecular studies and morphological characteristics confirm our identification. C. pfaffiae has been previously reported in P. iresinoides (H.B.K.) Spreng. in Trinidad and Gomphrena glomerata L. in Argentina (2). To our knowledge, this is the first report of C. pfaffiae causing disease in P. glomerata in Brazil and it may become a serious problem for some medicinal plant growers, due to the severity of the disease and the lack of chemical products for this pathogen. References: (1) Neto et al. J. Ethnopharmacol. 96:87, 2005. (2) U. Braun. A Monograph of Cercosporella, Ramularia and Allied Genera (Phytopathogenic Hyphomycetes). Eching bei Müchen, IHW-Verlage. Vol. 1, p. 68, 1995.

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First Report of Leaf Spot of Sweet Basil Caused by Cercospora guatemalensis in Korea

Plant Disease ◽

10.1094/pdis-05-12-0436-pdn ◽

2012 ◽

Vol 96 (10) ◽

pp. 1580-1580

Author(s):

J. H. Park ◽

K. S. Han ◽

J. Y. Kim ◽

H. D. Shin

Keyword(s):

Leaf Spot ◽

Morphological Characteristics ◽

Its Region ◽

Culture Collection ◽

Distilled Water ◽

First Report ◽

Sweet Basil ◽

Organic Farm ◽

Leaf Spots ◽

Close Phylogenetic Relationship

Sweet basil, Ocimum basilicum L., is a fragrant herb belonging to the family Lamiaceae. Originated in India 5,000 years ago, sweet basil plays a significant role in diverse cuisines across the world, especially in Asian and Italian cooking. In October 2008, hundreds of plants showing symptoms of leaf spot with nearly 100% incidence were found in polyethylene tunnels at an organic farm in Icheon, Korea. Leaf spots were circular to subcircular, water-soaked, dark brown with grayish center, and reached 10 mm or more in diameter. Diseased leaves defoliated prematurely. The damage purportedly due to this disease has reappeared every year with confirmation of the causal agent made again in 2011. A cercosporoid fungus was consistently associated with disease symptoms. Stromata were brown, consisting of brown cells, and 10 to 40 μm in width. Conidiophores were fasciculate (n = 2 to 10), olivaceous brown, paler upwards, straight to mildly curved, not geniculate in shorter ones or one to two times geniculate in longer ones, 40 to 200 μm long, occasionally reaching up to 350 μm long, 3.5 to 6 μm wide, and two- to six-septate. Conidia were hyaline, acicular to cylindric, straight in shorter ones, flexuous to curved in longer ones, truncate to obconically truncate at the base, three- to 16-septate, and 50 to 300 × 3.5 to 4.5 μm. Morphological characteristics of the fungus were consistent with the previous reports of Cercospora guatemalensis A.S. Mull. & Chupp (1,3). Voucher specimens were housed at Korea University herbarium (KUS). An isolate from KUS-F23757 was deposited in the Korean Agricultural Culture Collection (Accession No. KACC43980). Fungal DNA was extracted with DNeasy Plant Mini DNA Extraction Kits (Qiagen Inc., Valencia, CA). The complete internal transcribed spacer (ITS) region of rDNA was amplified with the primers ITS1/ITS4 and sequenced. The resulting sequence of 548 bp was deposited in GenBank (Accession No. JQ995781). This showed >99% similarity with sequences of many Cercospora species, indicating their close phylogenetic relationship. Isolate of KACC43980 was used in the pathogenicity tests. Hyphal suspensions were prepared by grinding 3-week-old colonies grown on PDA with distilled water using a mortar and pestle. Five plants were inoculated with hyphal suspensions and five plants were sprayed with sterile distilled water. The plants were covered with plastic bags to maintain a relative humidity of 100% for 24 h and then transferred to a 25 ± 2°C greenhouse with a 12-h photoperiod. Typical symptoms of necrotic spots appeared on the inoculated leaves 6 days after inoculation, and were identical to the ones observed in the field. C. guatemalensis was reisolated from symptomatic leaf tissues, confirming Koch's postulates. No symptoms were observed on control plants. Previously, the disease was reported in Malawi, India, China, and Japan (2,3), but not in Korea. To our knowledge, this is the first report of C. guatemalensis on sweet basil in Korea. Since farming of sweet basil has recently started on a commercial scale in Korea, the disease poses a serious threat to safe production of this herb, especially in organic farming. References: (1) C. Chupp. A Monograph of the Fungus Genus Cercospora. Ithaca, NY, 1953. (2) D. F. Farr and A. Y. Rossman. Fungal Databases. Systematic Mycology & Microbiology Laboratory, ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/ , May 5, 2012. (3) J. Nishikawa et al. J. Gen. Plant Pathol. 68:46, 2002.

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