First Report of Alternaria alternata Causing Black Leaf Spot of Rubber Tree in China

We first reported Alternaria heveae (E.G. Simmons ) to be the pathogen that caused black leaf spot of rubber tree (Hevea brasiliensis Muell. Arg) in Heikou county in July 2014 (1). Black leaf spots that resembled the symptoms caused by A. heveae were observed on the leaves of rubber trees of the whole propagule collection nursery in Jingping County (22°68′ N and 103°05′ E) of Yunnan Province. Black foliar spots (0.1 to 2 mm in diameter) surrounded by a yellow halo with lesions slightly sunken on the leaf surface were observed. To confirm whether the disease was caused by the same pathogen, 5-mm2 sections were removed from the leading edge of the lesion and were surface-sterilized in 75% ethanol, air-dried, plated on potato carrot agar (PCA), and incubated at 28°C in the dark. Colonies of the fungus on PCA had round margins and little aerial mycelia with gray-black coloration after 6 days of growth on PCA (2). Medium brown conidia were found to be in short chains of two to eight spores, ovoid, obclavate, and obpyriform, with or without a short conical or cylindrical-shaped apical beak. Conidia ranged from 22.5 to 67.5 μm long (mean 39.9 μm) × 10 to 15 μm wide (mean 12.5 μm; 100 colodia were measured), with three to six transverse septa and zero to three longitudinal or oblique septa. Morphological characteristics matched the descriptions of A. alternata [(Fries) Keissler] (4).The ITS1-5.8S-ITS2 region of one single-spore isolate, Ah02JP1, was amplified with primers ITS1 and ITS4. The PCR product was sequenced directly and deposited in GenBank (Accession No. KM111289). A BLAST search of the GenBank database revealed 100% similarity with A. alternata isolates KJ829535.1, KJ677246.1, and KF813070.1. Therefore, the pathogen was identified as A. alternata on the basis of its morphological characteristics and ITS sequence. Pathogenicity of a representative isolate, Ah02JP1 was confirmed using a field rubber tree inoculation method. Three rubber plants (the clone of rubber tree Yunyan77-4) were grown to the copper-colored leaf stage. Leaves were spray-inoculated (104 conidia per milliliter spore suspension) until drops were equally distributed using a manual pressure sprayer. Three rubber plants sprayed with sterile distilled water were used as controls. After inoculation, the plants were covered with plastic bags to maintain high relative humidity. The plastic bags were removed 2 days post-inoculation (dpi), and the plants were monitored daily for symptom development. Five days post-inoculation, spots similar to the original ones seen on the field trees developed on all inoculated leaves, while control leaves remained symptomless. A. alternata was re-isolated from spray-inoculated leaves, confirming Koch's postulates. A. alternata has been reported as the causal agent of leaf blight of rubber tree in India, which initially appeared as minute spots on leaves and enlarged with the growth of the leaves (3). However, in the present study, the symptoms (black leaf spots) remained small over time after inoculation. To our knowledge, this is the first report of A. alternata on rubber tree in China. Correct identification of pathogens is essential for disease management strategies. This report will establish a foundation for the further study of Alternaria alternata to address the disease effectively. References: (1) Z. Y. Cai et al. Plant Dis. 98:1011, 2014. (2) E. Mirkova. J. Phytopathol. 151:323, 2003. (3) C. B. Roy et al. J. Plantation Crops 34:499, 2006. (4) T. Y. Zhang. Page 32 in: Flora Fungorum Sinicorum, Vol. 16: Alternaria. Science Press, Beijing, 2003.

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First Report of Alternaria heveae Causing Black Leaf Spot of Rubber Tree in China

Plant Disease ◽

10.1094/pdis-01-14-0065-pdn ◽

2014 ◽

Vol 98 (7) ◽

pp. 1011-1011 ◽

Cited By ~ 1

Author(s):

Z. Y. Cai ◽

Y. X. Liu ◽

G. X. Huang ◽

M. Zhou ◽

G. Z. Jiang ◽

...

Keyword(s):

Rubber Tree ◽

Morphological Characteristics ◽

Black Spot ◽

Its Region ◽

Post Inoculation ◽

First Report ◽

Leaf Spots ◽

Plastic Bags ◽

Manual Pressure ◽

Tree Leaf

Rubber tree (Hevea brasiliensis Muell. Arg.) is an important industrial crop of tropical areas for natural rubber production. In October 2013, foliar spots (0.1 to 0.4 mm in diameter), black surrounded by a yellow halo, and with lesions slightly sunken were observed on the rubber tree leaf in a growing area in Heikou County of Yunnan Province. Lesion tissues removed from the border between symptomatic and healthy tissue were surface sterilized in 75% ethanol and air-dried, plated on PDA plates, and incubated at 28°C with alternating day/night cycles of light. The pathogen was observed growing out of many of the leaf pieces, and produced abundant conidia. Colonies 6.1 cm in diameter developed on potato carrot agar (PCA) after 7 days, with well-defined concentric rings of growth. Colonies on PCA were composed of fine, dark, radiating, surface and subsurface hyphae. Conidia produced in PCA culture were mostly solitary or in short chains of 2 to 5 spores, long ovoid to clavate, and light brown, 40 to 81.25 × 8 to 20 μm (200 colonies were measured), with 3 to 6 transverse septa and 0 to 2 longitudinal or oblique septa. Morphological characteristics were similar to those described for Alternaria heveae (3,4). A disease of rubber tree caused by Alternaria sp. had been reported in Mexico in 1947 (2). DNA of Ah01HK13 isolate was extracted for PCR and sequencing of the ITS region with ITS1 and ITS4 primers was completed. From the BLAST analysis, the sequence of Ah01HK13 (GenBank Accession No. KF953884), had 97% similarity to A. dauci, 96% identical to A. macrospora (AY154701.1 and DQ156342.1, respectively), indicating the pathogen belonged to Alternaria genus. According to morphological characteristics, this pathogen was identified as A. heveae. Pathogenicity of representative isolate, Ah01HK13 was confirmed using a field rubber tree inoculation method. Three rubber plants (the clone of rubber tree Yunyan77-4) were grown to the copper-colored leaf stage and inoculated by spraying spore suspension (concentration = 104 conidia/ml) to the copper-colored leaves until drops were equally distributed on it using manual pressure sprayer. Three rubber plants sprayed with sterile distilled water were used as controls. After inoculation, the plants were covered with plastic bags. The plastic bags were removed after 2 days post-inoculation (dpi) and monitored daily for symptom development (1). The experiment was repeated three times. The typical 0.1 to 0.4 mm black leaf spots were observed 7 dpi. No symptoms were observed on control plants. A fungus with the same colony and conidial morphology as A. heveae were re-isolated from leaf lesions on inoculated rubber plants, but not from asymptomatic leaves of control plants, fulfilling Koch's postulates. Based on these results, the disease was identified as black spot of rubber tree caused by A. heveae. To our knowledge, this is the first report of A. heveae on rubber tree in China. References: (1) Z. Y. Cai et al. Microbiol Res. 168:340, 2013. (2) W. J. Martin. Plant Dis. Rep. 31:155, 1947. (3) E. G. Simmons. Mycotaxon 50:262, 1994. (4) T. Y. Zhang. Page 111 in: Flora Fungorum Sinicorum: Alternaria, Science Press, Beijing, 2003.

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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 Leaf Spot Caused by Alternaria alternata on Kiwifruit in Italy

Plant Disease ◽

10.1094/pdis.1999.83.5.487d ◽

1999 ◽

Vol 83 (5) ◽

pp. 487-487 ◽

Cited By ~ 3

Author(s):

L. Corazza ◽

L. Luongo ◽

M. Parisi

Keyword(s):

New Zealand ◽

Alternaria Alternata ◽

Leaf Spot ◽

Southern Italy ◽

Morphological Characteristics ◽

Potato Dextrose Agar ◽

First Report ◽

Leaf Spots ◽

Detached Leaves ◽

Pathogenicity Tests

A leaf spot of kiwifruit (Actinidia deliciosa (A. Chev.) C. F. Liang & A. R. Ferg.) leaves was recently observed on plants of the cultivar Hayward in an orchard near Salerno, in southern Italy. The affected plants showed early severe defoliation. The fungus isolated from the infected leaves was identified as Alternaria alternata (Fr.:Fr.) Keissl., based on conidial morphological characteristics. Pathogenicity tests were made by inoculating detached leaves of male pollinator cultivar Tomuri and the female cultivars Hayward and Bruno with a 7-mm disk taken from actively growing cultures of the fungus on potato dextrose agar (PDA). After 14 days, necrotic leaf spots developed and A. alternata was consistently isolated from the inoculated leaves. A. alternata has been observed as a pathogen on leaves and fruits in New Zealand. In the Mediterranean, it has been reported in Israel (2) and in the island of Crete (1). This is the first report of Alternaria leaf spot on kiwifruit in Italy. References: (1) V. A. Bourbos and M. T. Skoudridakis. Petria 7:111, 1997. (2) A. Sive and D. Resnizky. Alon Hanotea 41:409, 1987.

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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 Bipolaris peregianensis Causing Leaf Spot of Cynodon dactylon in China

Plant Disease ◽

10.1094/pdis-12-11-1066-pdn ◽

2012 ◽

Vol 96 (6) ◽

pp. 917-917 ◽

Cited By ~ 2

Author(s):

Z. Y. Wang ◽

S. N. Xie ◽

Y. Wang ◽

H. Y. Wu ◽

M. Zhang

Keyword(s):

Leaf Spot ◽

Cynodon Dactylon ◽

Control Strategies ◽

Tap Water ◽

Morphological Characteristics ◽

Its Region ◽

Basal Cells ◽

First Report ◽

Leaf Spots ◽

Plastic Bags

Cynodon dactylon is widely cultivated as a sod crop in warm climates worldwide. In early September 2009, heavy leaf spot infection was observed on C. dactylon from Zhengzhou, Henan, China. Early symptoms appeared as small, elliptical, pale brown lesions on the leaves. Lesions later expanded to 5 to 10 mm long and 1 to 2 mm wide, becoming brown to dark brown. A fungus was consistently isolated from leaf spots on potato dextrose agar (PDA), but with poor sporulation. Morphological characteristics were observed from single-conidium cultures on tap water agar + wheat straw (TWA+W) after 5 to 7 days. Conidiophores were light to medium brown, cylindrical, solitary or clustered, unbranched, usually with basal cells enlarged, and 94.5 to 147.0 × 4.0 to 9.0 μm. Conidia were fusoid, strongly curved, end cells broadly hemiellipsoidal, brown, 58.5 to 84.5 × 13.5 to 18.5 μm, with 6 to 10 septa. These morphological characteristics are similar to those of Bipolaris peregianensis (2). The identity of our fungus was confirmed to be B. peregianensis by DNA sequencing of the internal transcribed spacer (ITS) region (GenBank Accession No. JQ316121), which was 99% homologous to those of other B. peregianensis isolates (= Cochliobolus peregianensis; Accession Nos AF071328 and AF158111) (1). Koch's postulates were performed with the leaves of three pots of C. dactylon. Leaves were sprayed with 1 × 105 conidia/ml of B. peregianensis; an equal number of leaves in the pots of the same plant sprayed with sterile distilled water served as the control. All test plants were covered with plastic bags for 24 h to maintain high relative humidity at 23 to 25°C. After 7 days, more than 50% of inoculated leaves showed symptoms identical to those observed in natural condition, whereas controls remained symptom free. Reisolation of the fungus from lesions on inoculated leaves confirmed that the causal agent was B. peregianensis. To our knowledge, this is the first report of leaf spots caused by B. peregianensis on C. dactylon in China. The disease cycle and the control strategies in the regions are being further studied. References: (1) M. L. Berbee et al. Mycologia 91:964, 1999. (2) A. Sivanesan. Mycol. Pap. 158:1, 1987.

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First Report of Leaf Spot Caused by Cercospora fukushiana on New Guinea Impatiens in Korea

Plant Disease ◽

10.1094/pdis-04-14-0363-pdn ◽

2014 ◽

Vol 98 (9) ◽

pp. 1280-1280

Author(s):

B. S. Kim ◽

K. S. Baek ◽

C. H. Pak ◽

J. H. Park ◽

H. D. Shin

Keyword(s):

Leaf Spot ◽

Morphological Characteristics ◽

Its Region ◽

New Guinea ◽

Culture Collection ◽

Distilled Water ◽

First Report ◽

Leaf Spots ◽

Plastic Bags ◽

New Guinea Impatiens

New Guinea impatiens, Impatiens hawkeri W. Bull, is widely cultivated as a potted plant and garden plant. In July 2013, hundreds of young plants (cv. Fanfare) showing symptoms of leaf spot with approximately 50% incidence were found in polyethylene tunnels in Yongin City, Korea. Leaf spots were circular to oblong, reaching 6 mm or more in diameter. The spots were initially uniformly brown to reddish brown, turning gray with reddish brown margin. Diseased plants defoliated prematurely and were abandoned without marketing due to signs of discoloration and yellowing on leaves. A cercosporoid fungus was consistently observed in association with disease symptoms. Stromata were brown, small, and composed of a few swollen hyphal cells. Conidiophores were emerging through the cuticle, fasciculate (n = 2 to 20), olivaceous to brown, paler toward the apex, straight to mildly curved, geniculate, 30 to 260 μm long, 3.5 to 5 μm wide, 1- to 6-septate, and with conspicuous conidial scars. Conidia were hyaline and acicular. Smaller conidia were straight and longer conidia were mildly curved. Conidia were subacute to obtuse at the apex, truncate to obconically truncate at the base, 2- to 18-septate, 30 to 320 × 3.5 to 5.5 μm, and with thickened, darkened hila at the base. Morphological characteristics of the fungus were consistent with the previous reports of Cercospora fukushiana (Matsuura) W. Yamam. (1). Voucher specimens were housed in the Korea University herbarium (KUS). An isolate from KUS-F27438 was deposited in the Korean Agricultural Culture Collection (Accession No. KACC47640). Fungal DNA was extracted with DNeasy Plant Mini Kits (Qiagen Inc., Valencia, CA). The complete internal transcribed spacer (ITS) region of rDNA was amplified with the primers ITS1/ITS4 (4) and sequenced. The resulting sequence of 497 bp was deposited in GenBank (Accession No. KJ620981). This showed >99% similarity with sequence of C. fukushiana (EF600954) on I. balsamina from Korea. Isolate of KACC47640 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 10 days after inoculation, and were identical to the symptoms observed in the field. C. fukushiana was re-isolated from symptomatic leaf tissues, confirming Koch's postulates. No symptoms were observed on water-inoculated control plants. Previously, leaf spots of Impatiens spp. associated with C. apii, C. balsaminae, and C. fukushiana have been reported (1,2,3). To our knowledge, this is the first report of C. fukushiana on I. hawkeri in Korea. Our observations in the nurseries of I. hawkeri suggest that low humidity with good ventilation as well as plant hygiene in greenhouses might be main strategies for preventing this disease. References: (1) C. Chupp. A Monograph of the Fungus Genus Cercospora. Ithaca, NY, 1953. (2) D. F. Farr and A. Y. Rossman. Fungal Databases. Syst. Mycol. Microbiol. Lab., online publication, ARS, USDA, retrieved March 25, 2014. (3) J. M. Soares et al. Plant Dis. 93:1214, 2009. (4) 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 Zonate Leaf Spot of Artocarpus altilis Caused by Cristulariella moricola in Taiwan

Plant Disease ◽

10.1094/pdis.2002.86.10.1179a ◽

2002 ◽

Vol 86 (10) ◽

pp. 1179-1179 ◽

Cited By ~ 1

Author(s):

B. Y. Hu ◽

W. W. Hsiao ◽

C. H. Fu

Keyword(s):

Leaf Spot ◽

Morphological Characteristics ◽

Eastern United States ◽

First Report ◽

Artocarpus Altilis ◽

Leaf Spots ◽

Plastic Bags ◽

Basal Septum ◽

Bull's Eye ◽

Northern Taiwan

Breadfruit (Artocarpus altilis (Parkinson) Fosberg) is an important landscape and garden tree in Taiwan. During the spring of 2002, zonate leaf spots of breadfruit were observed at a Taipei nursery in northern Taiwan. Initially, several small, brown, zonate lesions developed on leaves. As lesions enlarged, they coalesced, leading to blighting of leaves and premature defoliation. Sporophores on the host were generally hypophyllous but sometimes amphigenous, solitary, erect, easily detachable, and as much as 850 μm long. The upper portion of the sporophore is considered an individual conidium and consisted of a pyramidal head that was fusiform to ventricose and cristulate, 495 to 534 μm long and 210 to 290 μm wide at the broadest point. Branches within the pyramidal head were short and compact, and dichotomously or trichotomously branched. The conidia were hyaline, broad, septate, tapering toward an acute apex, and sometimes constricted at the basal septum. Conidiophores were 400 to 680 × 20 to 100 μm. The fungus was isolated from infected tissue and maintained on potato dextrose agar (PDA). Sclerotia were produced on PDA after 4 to 5 weeks at 20°C without light, but conidia were not observed in culture. The fungus was identified as Cristulariella moricola (Hino) Redhead based on morphological characteristics (1,2). To complete Koch's postulates, three sporophores from infected leaves or three sclerotia from cultures were placed individually on each of 10 breadfruit leaves. The plants were placed in plastic bags and incubated at 16 to 20°C. Symptoms were observed after 2 to 3 days on 100% of plants inoculated with sporophores and after 6 days on 50% of plants inoculated with sclerotia. The pathogen was reisolated from lesions on plants inoculated with sporophores and sclerotia. No symptoms were observed on the control plants. C. moricola has been known to cause a bull's eye or zonate leaf spot and defoliation on woody and annual plants, including at least 51 host species and 36 families distributed in the central and eastern United States (1). To our knowledge, this is the first report of zonate leaf spot and defoliation of breadfruit caused by C. moricola. References: (1) T. T. Chang. Bull. Taiwan For. Res. Inst. New Ser. 10 (2):235, 1995. (2) S. A. Redhead. Can. J. Bot. 53:700, 1975.

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First report of Alternaria tenuissima causing leaf black spot on pecan in China

Plant Disease ◽

10.1094/pdis-04-21-0757-pdn ◽

2021 ◽

Author(s):

Lichun Yan ◽

Xin Yang ◽

Zhuangwei Wang ◽

Yaming Qian ◽

Haijun Zhu ◽

...

Keyword(s):

Dna Sequences ◽

Leaf Spot ◽

Morphological Characteristics ◽

Black Spot ◽

Effective Control ◽

Post Inoculation ◽

Alternaria Tenuissima ◽

First Report ◽

Histone 3 ◽

Black Leaf Spot

China is one of the largest markets for pecan (Carya illinoinensis) consumption in the world, and also, pecan production in China has been increased by years since 2008 (Zhang et al. 2015). From July to September in the year 2019 and 2020, leaf black spot was observed on several pecan cultivars including Pawnee, Burkett, Kiowa and Western schley in the germplasm in Liuhe county, Nanjing, Jiangsu Province, China. Disease incidence was approximately 40% in 2019 and 50% in 2020 respectively. Small, dark brown to black spots on leaves were observed initially, and spots expanded quickly into circular or irregular when spots coalesced. In severe cases, the disease can obviously weaken the tree vigor, ultimately leading to losses in yield. Disease symptoms were not observed on the fruits. To determine the causal agent of black leaf spot, symptomatic leaves were collected and cut into pieces (approximately 3 × 3 cm2), surface sterilized with 1% sodium hypochlorite for 2 min and 75% ethanol for 30 s and rinsed twice with sterile distilled ddH2O. Dried tissues were placed on potato dextrose agar (PDA) amended with rifampin (Solarbio, Beijing, China) at a final concentration of 100 µg/mL and incubated at 25°C for 7 days in darkness. Five colonies were obtained and purified by single spore culture for morphological characterization. Colonies were initially white, turned to dark olivaceous with white margin and moderate to abundant gray aerial hyphae. Conidiophores were linear, light brown in color and appeared as individuals or in clusters. Conidia were pale brown to brown, typically obclavate or obpyriform (8 – 49 µm× 3 – 18 µm), with one to five transverse septa and zero to three longitudinal septa (n=50). Conidia of all isolates produced no beaks or a short beak. Based on conidial morphological characteristics, isolates were tentatively identified as Alternaria tenuissima (Simmons 2007). To further confirm the five isolates, the internal transcribed spacer (ITS) region of ribosomal DNA (rDNA), partial region of the histone 3 (H3) gene, translation elongation factor 1-α gene (TEF) and the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were amplified with PCR primer sets ITS1/ITS4 (White et al. 1990), H3-1a/H3-1b (Glass and Donaldson 1995), EF1-728F/EF1-986R (Carbone and Kohn 1999), and GDF1/GDR1 (Berbee et al. 1999) respectively. The sequences were deposited in GenBank (ITS, MN822659 to MN822661 and MZ182355 to MZ182356; histone 3, MN840997 to MN840999 and MZ202355 to MZ202356; TEF, MZ246595 to MZ246599; GAPDH, MZ246590 to MZ246594). BLAST analysis of the resulting sequences showed 99% to 100 % nucleotide identical to those of A. tenuissima isolates (KP278184 [ITS]; MH824352 [H3]; MN046379 [TEF]; MK683840 [GAPDH]). Therefore, based on morphological characteristics and DNA sequences data, the five isolates were identified as A. tenuissima. To determine the pathogenicity of the five isolates, 10 µL of 105 conidia /mL suspension from each isolate was placed to three intact young leaves (Pawnee) respectively. Leaves inoculated with ddH2O in the same manner served as the controls. Inoculated leaves were placed in a growth chamber at 28°C with 95% relatively humidity (RH). Black spot symptoms appeared on all inoculated leaves by 7 days post inoculation, A. tenuissima were re-isolated from the inoculated leaves with isolates and reconfirmed by morphological characteristics, thus fulfilling the Koch’s postulates. The controls remained symptomless. Pathogenicity tests were conducted twice. To our knowledge, this is the first report of A. tenuissima causing black leaf spot on pecan in China. Although we have not found any fruit infection, we think the disease is also a potential threat to pecan fruits and production. If more reports of this pathogen are found on pecans, then it is necessary to study and develop effective control strategies.

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First Report of Leaf Spot caused by Bipolaris oryzae on Switchgrass in Tennessee

Plant Disease ◽

10.1094/pdis-01-13-0070-pdn ◽

2013 ◽

Vol 97 (12) ◽

pp. 1654-1654 ◽

Cited By ~ 3

Author(s):

A. L. Vu ◽

M. M. Dee ◽

J. Zale ◽

K. D. Gwinn ◽

B. H. Ownley

Keyword(s):

Leaf Spot ◽

Panicum Virgatum ◽

Morphological Characteristics ◽

Its Region ◽

Spore Production ◽

Post Inoculation ◽

Sterile Water ◽

Bipolaris Oryzae ◽

First Report ◽

Symptomatic Leaf

Knowledge of pathogens in switchgrass, a potential biofuels crop, is limited. In December 2007, dark brown to black irregularly shaped foliar spots were observed on ‘Alamo’ switchgrass (Panicum virgatum L.) on the campus of the University of Tennessee. Symptomatic leaf samples were surface-sterilized (95% ethanol, 1 min; 20% commercial bleach, 3 min; 95% ethanol, 1 min), rinsed in sterile water, air-dried, and plated on 2% water agar amended with 3.45 mg fenpropathrin/liter (Danitol 2.4 EC, Valent Chemical, Walnut Creek, CA) and 10 mg/liter rifampicin (Sigma-Aldrich, St. Louis, MO). A sparsely sporulating, dematiaceous mitosporic fungus was observed. Fungal plugs were transferred to surface-sterilized detached ‘Alamo’ leaves on sterile filter paper in a moist chamber to increase spore production. Conidia were ovate, oblong, mostly straight to slightly curved, and light to olive-brown with 3 to 10 septa. Conidial dimensions were 12.5 to 17 × 27.5 to 95 (average 14.5 × 72) μm. Conidiophores were light brown, single, multiseptate, and geniculate. Conidial production was polytretic. Morphological characteristics and disease symptoms were similar to those described for Bipolaris oryzae (Breda de Haan) Shoemaker (2). Disease assays were done with 6-week-old ‘Alamo’ switchgrass grown from seed scarified with 60% sulfuric acid and surface-sterilized in 50% bleach. Nine 9 × 9-cm square pots with approximately 20 plants per pot were inoculated with a mycelial slurry (due to low spore production) prepared from cultures grown on potato dextrose agar for 7 days. Cultures were flooded with sterile water and rubbed gently to loosen mycelium. Two additional pots were inoculated with sterile water and subjected to the same conditions to serve as controls. Plants were exposed to high humidity by enclosure in a plastic bag for 72 h. Bags were removed, and plants were incubated at 25/20°C with 50 to 60% relative humidity. During the disease assay, plants were kept in a growth chamber with a 12-h photoperiod of fluorescent and incandescent lighting. Foliar leaf spot symptoms appeared 5 to 14 days post-inoculation for eight of nine replicates. Control plants had no symptoms. Symptomatic leaf tissue was processed and plated as described above. The original fungal isolate and the pathogen recovered in the disease assay were identified using internal transcribed spacer (ITS) region sequences. The ITS region of rDNA was amplified with PCR and primer pairs ITS4 and ITS5 (4). PCR amplicons of 553 bp were sequenced, and sequences from the original isolate and the reisolated pathogen were identical (GenBank Accession No. JQ237248). The sequence had 100% nucleotide identity to B. oryzae from switchgrass in Mississippi (GU222690, GU222691, GU222692, and GU222693) and New York (JF693908). Leaf spot caused by B. oryzae on switchgrass has also been described in North Dakota (1) and was seedborne in Mississippi (3). To our knowledge, this is the first report of B. oryzae from switchgrass in Tennessee. References: (1) D. F. Farr and A. Y. Rossman. Fungal Databases. Systematic Mycology and Microbiology Laboratory, ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/, 28 June 2012. (2) J. M. Krupinsky et al. Can. J. Plant Pathol. 26:371, 2004. (3) M. Tomaso-Peterson and C. J. Balbalian. Plant Dis. 94:643, 2010. (4) T. J. White et al. Pages 315-322 in: PCR Protocols: a Guide to Methods and Applications. M. A. Innis et al. (eds), Acad. Press, San Diego, 1990.

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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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