First report of banana (Musa acuminate L. AAA Cavendish, cv. Formosana) leaf spot disease caused by Curvularia geniculata in China

Banana (Musa acuminate L.) is an important tropical fruit in China. During 2019-2020, a new leaf spot disease was observed on banana (M. acuminate L. AAA Cavendish, cv. Formosana) at two orchards of Chengmai county (19°48ʹ41.79″ N, 109°58ʹ44.95″ E), Hainan province, China. In total, the disease incidence was about 5% of banana trees (6 000 trees). The leaf spots occurred sporadically and were mostly confined to the leaf margin, and the percentage of the leaf area covered by lesions was less than 1%. Symptoms on the leaves were initially reddish brown spots that gradually expanded to ovoid-shaped lesions and eventually become necrotic, dry, and gray with a yellow halo. The conidia obtained from leaf lesions were brown, erect or curved, fusiform or elliptical, 3 to 4 septa with dimensions of 13.75 to 31.39 µm × 5.91 to 13.35 µm (avg. 22.39 × 8.83 µm). The cells of both ends were small and hyaline while the middle cells were larger and darker (Zhang et al. 2010). Morphological characteristics of the conidia matched the description of Curvularia geniculata (Tracy & Earle) Boedijn. To acquire the pathogen, tissue pieces (15 mm2) of symptomatic leaves were surface disinfected in 70% ethanol (10 s) and 0.8% NaClO (2 min), rinsed in sterile water three times, and transferred to potato dextrose agar (PDA) for three days at 28°C. Grayish green fungal colonies appeared, and then turned fluffy with grey and white aerial mycelium with age. Two representative isolates (CATAS-CG01 and CATAS-CG92) of single-spore cultures were selected for molecular identification. Genomic DNA was extracted from the two isolates, the internal transcribed spacer (ITS), large subunit ribosomal DNA (LSU rDNA), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), translation elongation factor 1-alpha (TEF1-α) and RNA polymerase II second largest subunit (RPB2) were amplified and sequenced with universal primers ITS1/ITS4, LROR/LR5, GPD1/GPD2, EF1-983F/EF1-2218R and 5F2/7cR, respectively (Huang et al. 2017; Raza et al. 2019). The sequences were deposited in GenBank (MW186196, MW186197, OK091651, OK721009 and OK491081 for CATAS-CG01; MZ734453, MZ734465, OK091652, OK721100 and OK642748 for CATAS-CG92, respectively). For phylogenetic analysis, MEGA7.0 (Kumar et al. 2016) was used to construct a Maximum Likelihood (ML) tree with 1 000 bootstrap replicates, based on a concatenation alignment of five gene sequences of the two isolates in this study as well as sequences of other Curvularia species obtained from GenBank. The cluster analysis revealed that isolates CATAS-CG01 and CATAS-CG92 were C. geniculata. Pathogenicity assays were conducted on 7-leaf-old banana seedlings. Two leaves from potted plants were stab inoculated by puncturing into 1-mm using a sterilized needle and placing 10 μl conidial suspension (2×106 conidia/ml) on the surface of wounded leaves and equal number of leaves were inoculated with sterile distilled water serving as control (three replicates). Inoculated plants were grown in the greenhouse (12 h/12 h light/dark, 28°C, 90% relative humidity). Necrotic lesions on inoculated leaves appeared seven days after inoculation, whereas control leaves remained healthy. The fungus was recovered from inoculated leaves, and its taxonomy was confirmed morphologically and molecularly, fulfilling Koch’s postulates. C. geniculata has been reported to cause leaf spot on banana in Jamaica (Meredith, 1963). To our knowledge, this is the first report of C. geniculata on banana in China.

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First Report of a Leaf Spot Disease of Bells-of-Ireland (Moluccella laevis) Caused by Cercospora apii in California

Plant Disease ◽

10.1094/pdis.2003.87.2.203a ◽

2003 ◽

Vol 87 (2) ◽

pp. 203-203

Author(s):

S. T. Koike ◽

S. A. Tjosvold ◽

J. Z. Groenewald ◽

P. W. Crous

Keyword(s):

Leaf Spot ◽

Sequence Data ◽

Disease Incidence ◽

Leaf Spot Disease ◽

Conidial Suspension ◽

Internal Transcribed Spacer Region ◽

First Report ◽

Spot Disease ◽

Leaf Spots ◽

Initial Symptoms

Bells-of-Ireland (Moluccella laevis) (Lamiaceae) is an annual plant that is field planted in coastal California (Santa Cruz County) for commercial cutflower production. In 2001, a new leaf spot disease was found in these commercially grown cutflowers. The disease was most serious in the winter-grown crops in 2001 and 2002, with a few plantings having as much as 100% disease incidence. All other plantings that were surveyed during this time had at least 50% disease. Initial symptoms consisted of gray-green leaf spots. Spots were generally oval in shape, often delimited by the major leaf veins, and later turned tan. Lesions were apparent on both adaxial and abaxial sides of the leaves. A cercosporoid fungus having fasciculate conidiophores, which formed primarily on the abaxial leaf surface, was consistently associated with the spots. Based on morphology and its host, this fungus was initially considered to be Cercospora molucellae Bremer & Petr., which was previously reported on leaves of M. laevis in Turkey (1). However, sequence data obtained from the internal transcribed spacer region (ITS1, ITS2) and the 5.8S gene (STE-U 5110, 5111; GenBank Accession Nos. AY156918 and AY156919) indicated there were no base pair differences between the bells-of-Ireland isolates from California, our own reference isolates of C. apii, as well as GenBank sequences deposited as C. apii. Based on these data, the fungus was subsequently identified as C. apii sensu lato. Pathogenicity was confirmed by spraying a conidial suspension (1.0 × 105 conidia/ml) on leaves of potted bells-of-Ireland plants, incubating the plants in a dew chamber for 24 h, and maintaining them in a greenhouse (23 to 25°C). After 2 weeks, all inoculated plants developed leaf spots that were identical to those observed in the field. C. apii was again associated with all leaf spots. Control plants, which were treated with water, did not develop any symptoms. The test was repeated and the results were similar. To our knowledge this is the first report of C. apii as a pathogen of bells-of-Ireland in California. Reference: (1) C. Chupp. A Monograph of the Fungus Genus Cercospora. Cornell University Press, Ithaca, New York, 1954.

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First Report of Leaf Spot Disease Caused by Colletotrichum gloeosporioides on Chinese Bean Tree in China

Plant Disease ◽

10.1094/pdis-03-12-0261-pdn ◽

2013 ◽

Vol 97 (1) ◽

pp. 138-138 ◽

Cited By ~ 1

Author(s):

B. Z. Fu ◽

M. Yang ◽

G. Y. Li ◽

J. R. Wu ◽

J. Z. Zhang ◽

...

Keyword(s):

Leaf Spot ◽

Colletotrichum Gloeosporioides ◽

Disease Incidence ◽

Phylogenetic Analyses ◽

Morphological Characteristics ◽

Leaf Spot Disease ◽

First Report ◽

Spot Disease ◽

Rdna Its ◽

Its Sequence Analysis

Chinese bean tree, Catalpa fargesii f. duciouxii (Dode) Gilmour, is an ornamental arbor plant. Its roots, leaves, and flowers have long been used for medicinal purposes in China. During July 2010, severe outbreaks of leaf spot disease on this plant occurred in Kunming, Yunnan Province. The disease incidence was greater than 90%. The symptoms on leaves began as dark brown lesions surrounded by chlorotic halos, and later became larger, round or irregular spots with gray to off-white centers surrounded by dark brown margins. Leaf tissues (3 × 3 mm), cut from the margins of lesions, were surface disinfected in 0.1% HgCl2 solution for 3 min, rinsed three times in sterile water, plated on potato dextrose agar (PDA), and incubated at 28°C. The same fungus was consistently isolated from the diseased leaves. Colonies of white-to-dark gray mycelia formed on PDA, and were slightly brown on the underside of the colony. The hyphae were achromatic, branching, septate, and 4.59 (±1.38) μm in diameter on average. Perithecia were brown to black, globose in shape, and 275.9 to 379.3 × 245.3 to 344.8 μm. Asci that formed after 3 to 4 weeks in culture were eight-spored, clavate to cylindrical. The ascospores were fusiform, slightly curved, unicellular and hyaline, and 13.05 to 24.03 × 10.68 to 16.02 μm. PCR amplification was carried out by utilizing universal rDNA-ITS primer pair ITS4/ITS5 (2). Sequencing of the PCR products of DQ1 (GenBank Accession No. JN165746) revealed 99% similarity (100% coverage) with Colletotrichum gloeosporioides isolates (GenBank Accession No. FJ456938.1, No. EU326190.1, No. DQ682572.1, and No. AY423474.1). Phylogenetic analyses (MEGA 4.1) using the neighbor-joining (NJ) algorithm placed the isolate in a well-supported cluster (>90% bootstrap value based on 1,000 replicates) with other C. gloeosporioides isolates. The pathogen was identified as C. gloeosporioides (Penz.) Penz. & Sacc. (teleomorph Glomerella cingulata (Stoneman) Spauld & H. Schrenk) based on the morphological characteristics and rDNA-ITS sequence analysis (1). To confirm pathogenicity, Koch's postulates were performed on detached leaves of C. fargesii f. duciouxii, inoculated with a solution of 1.0 × 106 conidia per ml. Symptoms similar to the original ones started to appear after 10 days, while untreated leaves remained healthy. The inoculation assay used three leaves for untreated and six leaves for treated. The experiments were repeated once. C. gloeosporioides was consistently reisolated from the diseased tissue. C. gloeosporioides is distributed worldwide causing anthracnose on a wide variety of plants (3). To the best of our knowledge, this is the first report of C. gloeosporioides causing leaf spots on C. fargesii f. duciouxii in China. References: (1) B. C. Sutton. Page 1 in: Colletotrichum: Biology, Pathology and Control. CAB International. Wallingford, UK, 1992. (2) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990. (3) J. Yan et al. Plant Dis. 95:880, 2011.

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First Report of Leaf Spot Disease Caused by Epicoccum nigrum on Lablab purpureus in India

Plant Disease ◽

10.1094/pdis-07-13-0798-pdn ◽

2014 ◽

Vol 98 (2) ◽

pp. 284-284 ◽

Cited By ~ 12

Author(s):

S. Mahadevakumar ◽

K. M. Jayaramaiah ◽

G. R. Janardhana

Keyword(s):

Leaf Spot ◽

Leaf Surface ◽

Disease Incidence ◽

Leaf Spot Disease ◽

Post Inoculation ◽

Conidial Suspension ◽

Lablab Purpureus ◽

First Report ◽

Spot Disease ◽

Epicoccum Nigrum

Lablab purpureus (L.) Sweet (Indian bean) is an important pulse crop grown in arid and semi-arid regions of India. It is one of the most widely cultivated legume species and has multiple uses. During a September 2010 survey, we recorded a new leaf spot disease on L. purpureus in and around Mysore district (Karnataka state) with 40 to 80% disease incidence in 130 ha of field crop studied, which accounted for 20 to 35% estimated yield loss. The symptoms appeared as small necrotic spots on the upper leaf surface. The leaf spots were persistent under mild infection throughout the season with production of conidia in clusters on abaxial leaf surface. A Dueteromyceteous fungus was isolated from affected leaf tissues that were surface sterilized with 2% NaOCl2 solution then washed thrice, dried, inoculated on potato dextrose agar (PDA) medium, and incubated at 28 ± 2°C at 12 h alternate light and dark period for 7 days. The fungal colony with aerial mycelia interspersed with dark cushion-shaped sporodochia consists of short, compact conidiophores bearing large isodiametric, solitary, muricate, brown, globular to pear shaped conidia (29.43 to 23.92 μm). Fungal isolate was identified as Epicoccum sp. based on micro-morphological and cultural features (1). Further authenticity of the fungus was confirmed by PCR amplification of the internal transcribed spacer (ITS) region using ITS1/ITS4 universal primer. The amplified PCR product was purified, sequenced directly, and BLASTn search revealed 100% homology to Epicoccum nigrum Link. (DQ093668.1 and JX914480.1). A representative sequence of E. nigrum was deposited in GenBank (Accession No. KC568289.1). The isolated fungus was further tested for its pathogenicity on 30-day-old healthy L. purpureus plants under greenhouse conditions. A conidial suspension (106 conidia/ml) was applied as foliar spray (three replicates of 15 plants each) along with suitable controls. The plants were kept under high humidity (80%) for 5 days and at ambient temperature (28 ± 2°C). The appearance of leaf spot symptoms were observed after 25 days post inoculation. Further, the pathogen was re-isolated and confirmed by micro-morphological characteristics. E. nigrum has been reported to cause post-harvest decay of cantaloupe in Oklahoma (2). It has also been reported as an endophyte (3). Occurrence as a pathogen on lablab bean has not been previously reported. To our knowledge, this is the first report of the occurrence of E. nigrum on L. purpureus in India causing leaf spot disease. References: (1) H. L. Barnet and B. B. Hunter. Page 150 in: Illustrated Genera of Imperfect Fungi, 1972. (2) B. D. Bruten et al. Plant Dis. 77:1060, 1993. (3) L. C. Fávaro et al. PLoS One 7(6):e36826, 2012.

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First Report of Tomato Gray Leaf Spot Disease Caused by Stemphylium solani in Malaysia

Plant Disease ◽

10.1094/pdis-03-12-0223-pdn ◽

2012 ◽

Vol 96 (8) ◽

pp. 1226-1226

Author(s):

A. Nasehi ◽

J. B. Kadir ◽

M. A. Zainal Abidin ◽

M. Y. Wong ◽

F. Mahmodi

Keyword(s):

Leaf Spot ◽

Disease Incidence ◽

Gray Leaf Spot ◽

Leaf Spot Disease ◽

Conidial Suspension ◽

Internal Transcribed Spacer Region ◽

First Report ◽

Spot Disease ◽

Pcr Product ◽

Morphological Criteria

In June 2011, tomatoes (Solanum lycopersicum) in major growing areas of the Cameron Highlands and the Johor state in Malaysia were affected by a leaf spot disease. Disease incidence exceeded 80% in some severely infected regions. Symptoms on 50 observed plants initially appeared on leaves as small, brownish black specks, which later became grayish brown, angular lesions surrounded by a yellow border. As the lesions matured, the affected leaves dried up and became brittle and later developed cracks in the center of the lesions. A survey was performed in these growing areas and 27 isolates of the pathogen were isolated from the tomato leaves on potato carrot agar (PCA). The isolates were purified by the single spore technique and were transferred onto PCA and V8 agar media for conidiophore and conidia production under alternating light (8 hours per day) and darkness (16 hours per day) (4). Colonies on PCA and V8 agar exhibited grey mycelium and numerous conidia were formed at the terminal end of conidiophores. The conidiophores were up to 240 μm long. Conidia were oblong with 2 to 11 transverse and 1 to 6 longitudinal septa and were 24 to 69.6 μm long × 9.6 to 14.4 μm wide. The pathogen was identified as Stemphylium solani on the basis of morphological criteria (2). In addition, DNA was extracted and the internal transcribed spacer region (ITS) was amplified by universal primers ITS5 and ITS4 (1). The PCR product was purified by the commercial PCR purification kit and the purified PCR product sequenced. The resulting sequences were 100% identical to published S. solani sequences (GenBank Accestion Nos. AF203451 and HQ840713). The amplified ITS region was deposited with NCBI GenBank under Accession No. JQ657726. A representative isolate of the pathogen was inoculated on detached 45-day-old tomato leaves of Malaysian cultivar 152177-A for pathogenicity testing. One wounded and two nonwounded leaflets per leaf were used in this experiment. The leaves were wounded by applying pressure to leaf blades with the serrated edge of a forceps. A 20-μl drop of conidial suspension containing 105 conidia/ml was used to inoculate these leaves (3). The inoculated leaves were placed on moist filter paper in petri dishes and incubated for 48 h at 25°C. Control leaves were inoculated with sterilized distilled water. After 7 days, typical symptoms for S. solani similar to those observed in the farmers' fields developed on both wounded and nonwounded inoculated leaves, but not on noninoculated controls, and S. solani was consistently reisolated. To our knowledge, this is the first report of S. solani causing gray leaf spot of tomato in Malaysia. References: (1) M. P. S. Camara et al. Mycologia 94:660, 2002. (2) B. S. Kim et al. Plant Pathol. J. 15:348, 1999. (3) B. M. Pryor and T. J. Michailides. Phytopathology 92:406, 2002. (4) E. G. Simmons. CBS Biodiversity Series 6:775, 2007.

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First report of leaf spot disease caused by Stemphylium eturmiunum on American sweetgum (Liquidambar styraciflua L.) in China

Plant Disease ◽

10.1094/pdis-09-20-1877-pdn ◽

2021 ◽

Author(s):

Yun-fei Mao ◽

Li Jin ◽

Huiyue Chen ◽

Xiang-rong Zheng ◽

Minjia Wang ◽

...

Keyword(s):

Leaf Spot ◽

Disease Incidence ◽

Morphological Characteristics ◽

Liquidambar Styraciflua ◽

Leaf Spot Disease ◽

Distilled Water ◽

Conidial Suspension ◽

First Report ◽

Wood Processing ◽

Light Yellow

American sweetgum (Liquidambar styraciflua L.) is an important tree for landscaping and wood processing. In recent years, leaf spots on American sweetgum with disease incidence of about 53% were observed in about 1200 full grown plants in a field (about 8 ha) located in Pizhou, Jiangsu Province, China. Initially, dense reddish-brown spots appeared on both old and new leaves. Later, the spots expanded into dark brown lesions with yellow halos. Symptomatic leaf samples from different trees were collected and processed in the laboratory. For pathogen isolation, leaf sections (4×4mm) removed from the lesion margin were surface sterilized with 75% ethanol for 20s and then sterilized in 2% NaOCl for 30s, rinsed three times in sterile distilled water, incubated on potato dextrose agar (PDA) at 25 °C in the darkness. After 5 days of cultivation, the pure culture was obtained by single spore separation. 6 isolate samples from different leaves named FXA1 to FXA6 shared nearly identical morphological features. The isolate FXA1 (codes CFCC 54675) was deposited in the China Center for Type Culture Collection. On the PDA, the colonies were light yellow with dense mycelium, rough margin, and reverse brownish yellow. Conidiophores (23–35 × 6–10 µm) (n=60) were solitary, straight to flexuous. Conidia (19–34 × 10–21 µm) (n=60) were single, muriform, oblong, mid to deep brown, with 1 to 6 transverse septa. These morphological characteristics resemble Stemphylium eturmiunum (Simmons 2001). Genomic DNA was extracted from mycelium following the CTAB method. The ITS region, gapdh, and cmdA genes were amplified and sequenced with the primers ITS5/ITS4 (Woudenberg et al. 2017), gpd1/gpd2 (Berbee et al. 1999), and CALDF1/CALDR2 (Lawrence et al. 2013), respectively. A maximum likelihood phylogenetic analysis based on ITS, gapdh and cmdA (accession nos. MT898502-MT898507, MT902342-MT902347, MT902336-MT902341) sequences using MEGA 7.0 revealed that the isolates were placed in the same clade as S. eturmiunum with 98% bootstrap support. All seedlings for pathogenicity tests were enclosed in plastic transparent incubators to maintain high relative humidity (90%-100%) and incubated in a greenhouse at 25°C with a 12-h photoperiod. For pathogenicity, the conidial suspension (105 spores/ml) of each isolate was sprayed respectively onto healthy leaves of L. styraciflua potted seedlings (2-year-old, 3 replicate plants per isolate). As a control, 3 seedlings were sprayed with sterile distilled water. After 7 days, dense reddish-brown spots were observed on all inoculated leaves. In another set of tests, healthy plants (3 leaves per plant, 3 replicate plants per isolate) were wound-inoculated with mycelial plugs (4×4mm) and inoculated with sterile PDA plugs as a control. After 7 days, brown lesions with light yellow halo were observed on all inoculation sites with the mycelial plugs. Controls remained asymptomatic in the entire experiment. The pathogen was reisolated from symptomatic tissues and identified as S. eturmiunum but was not recovered from the control. The experiment was repeated twice with the similar results, fulfilling Koch’s postulates. S. eturmiunum had been reported on tomato (Andersen et al. 2004), wheat (Poursafar et al. 2016), garlic (L. Fu et al. 2019) but not on woody plant leaves. To our knowledge, this is the first report of S. eturmiunum causing leaf spot on L. styraciflua in the world. This disease poses a potential threat to American sweetgum and wheat in Pizhou.

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First Report of Gray Leaf Spot of Mango (Mangifera indica) Caused by Pestalotiopsis mangiferae in Taiwan

Plant Disease ◽

10.1094/pdis-91-12-1684b ◽

2007 ◽

Vol 91 (12) ◽

pp. 1684-1684 ◽

Cited By ~ 6

Author(s):

Y. Ko ◽

K. S. Yao ◽

C. Y. Chen ◽

C. H. Lin

Keyword(s):

Leaf Spot ◽

Mangifera Indica ◽

Gray Leaf Spot ◽

Leaf Spot Disease ◽

Continuous Exposure ◽

Conidial Suspension ◽

Tropical Fruit ◽

First Report ◽

Spot Disease ◽

Initial Symptoms

Mango (Mangifera indica L.; family Anacardiaceae) is one of the world's most important fruit crops and is widely grown in tropical and subtropical regions. Since 2001, a leaf spot disease was found in mango orchards of Taiwan. Now, the disease was observed throughout (approximately 21,000 ha) Taiwan in moderate to severe form, thus affecting the general health of mango trees and orchards. Initial symptoms were small, yellow-to-brown spots on leaves. Later, the irregularly shaped spots, ranging from a few millimeters to a few centimeters in diameter, turned white to gray and coalesced to form larger gray patches. Lesions had slightly raised dark margins. On mature lesions, numerous black acervuli, measuring 290 to 328 μm in diameter, developed on the gray necrotic areas. Single conidial isolates of the fungus were identified morphologically as Pestalotiopsis mangiferae (Henn.) Steyaert (2,3) and were consistently isolated from the diseased mango leaves on acidified (0.06% lactic acid) potato dextrose agar (PDA) medium incubated at 25 ± 1°C. Initially, the fungus grew (3 mm per day) on PDA as a white, chalky colony that subsequently turned gray after 2 weeks. Acervuli developed in culture after continuous exposure to light for 9 to 12 days at 20 to 30°C. Abundant conidia oozed from the acervulus as a creamy mass. The conidia (17.6 to 25.4 μm long and 4.8 to 7.1 μm wide) were fusiform and usually straight to slightly curved with four septa. Three median cells were olivaceous and larger than the hyaline apical and basal cells. The apical cells bore three (rarely four) cylindrical appendages. Pathogenicity tests were conducted with either 3-day-old mycelial discs or conidial suspension (105 conidia per ml) obtained from 8- to 10-day-old cultures. Four leaves on each of 10 trees were inoculated. Before inoculation, the leaves were washed with a mild detergent, rinsed with tap water, and then surface sterilized with 70% ethanol. Leaves were wounded with a needle and exposed to either a 5-mm mycelial disc or 0.2 ml of the spore suspension. The inoculated areas were wrapped with cotton pads saturated with sterile water and the leaves were covered with polyethylene bags for 3 days to maintain high relative humidity. Wounded leaves inoculated with PDA discs alone served as controls. The symptoms described above were observed on all inoculated leaves, whereas uninoculated leaves remained completely free from symptoms. Reisolation from the inoculated leaves consistently yielded P. mangiferae, thus fulfilling Koch's postulates. Gray leaf spot is a common disease of mangos in the tropics and is widely distributed in Africa and Asia (1–3); however, to our knowledge, this is the first report of gray leaf spot disease affecting mango in Taiwan. References: (1) T. K. Lim and K. C. Khoo. Diseases and Disorders of Mango in Malaysia. Tropical Press. Malaysia, 1985. (2) J. E. M. Mordue. No. 676 in: CMI Descriptions of Pathogenic Fungi and Bacteria. Surrey, England, 1980. (3) R. C. Ploetz et al. Compendium of Tropical Fruit Diseases. The American Phytopathological Society. St. Paul, MN, 1994.

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First report of leaf spot caused by Colletotrichum fructicola on Dalbergia hupeana in China

Plant Disease ◽

10.1094/pdis-08-21-1842-pdn ◽

2021 ◽

Author(s):

Yang Zhou ◽

Rou Ye ◽

Qin Ying ◽

Yang Zhang ◽

Linping Zhang

Keyword(s):

Phylogenetic Tree ◽

Leaf Spot ◽

Disease Incidence ◽

Morphological Characteristics ◽

Jiangxi Province ◽

Leaf Spot Disease ◽

Sterile Water ◽

First Report ◽

Spot Disease ◽

Colletotrichum Fructicola

Dalbergia hupeana is a kind of wood and medicinal tree widely distributed in southern China. Since 2019, a leaf spot disease was observed on the leaves of D. hupeana in Gangxia village, Luoting town in Jiangxi Province, China (28°52′53″N, 115°44′58″E). The disease incidence was estimated to be above 50%. The symptoms began as small spots that gradually expanded, developing a brown central and dark brown to black margin. The spots ranged from 4 to 6 mm in diameter. Leaf pieces (5 × 5 mm) from lesion margins were surface sterilized in 70% ethanol for 30 s followed by 2% NaOCl for 1 min and then rinsed three times with sterile water. Tissues were placed on potato dextrose agar (PDA) and incubated at 25°C. Pure cultures were obtained by monosporic isolation. Fifteen strains with similar morphological characterizations were isolated, and three representative isolates (JHT-1, JHT-2, and JHT-3) were chosen and used for further study. Colonies on PDA of three isolates were grayish-green with white edges and dark green on the reverse side. Conidia were transparent, cylindrical with rounded ends, and measured 3.6-5.3 µm × 9.5-15.2 µm (3.7 ± 0.2 × 13.6 ± 1.1 µm, n = 100). Appressoria were dark brown, globose or subcylindrical, and ranged from 6.2-9.2 µm× 5.1-6.8 µm (7.9 ± 0.4 × 5.9 ± 0.3 µm, n=100). The morphological characteristics of the three strains were consistent with the description of species in the Colletotrichum gloeosporioides complex (Weir et al. 2012). The internal transcribed spacer (ITS) regions, actin (ACT), calmodulin (CAL), chitin synthase (CHS-1) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and beta-tubulin 2 (TUB2) were amplified from genomic DNA for the three isolates using primers ITS1/ITS4, ACT-512F/ACT-783R, CL1/CL2, CHS-79F/CHS-345R, GDF/GDR and T1/Bt2b (Weir et al. 2012), respectively. The sequences were deposited in GenBank (Accession Nos. MZ482016 - MZ482018 for ITS; MZ463636 - MZ463638 for ACT; MZ463648- MZ463650 for CAL; MZ463639 - MZ463641 for CHS-1; MZ463642 - MZ463644 for GAPDH; MZ463645 - MZ463647 for TUB2). A neighbor-joining phylogenetic tree was constructed with MEGA 7.0 using the concatenation of multiple sequences (ITS, ACT, GAPDH, TUB2, CHS-1, CAL) (Kumar et al. 2016). According to the phylogenetic tree, three isolates fall within the Colletotrichum fructicola clade (boot support 99%). Based on morphological characteristics and phylogenetic analysis, three isolates were identified as C. fructicola. The pathogenicity of three isolates was conducted on two-yr-old seedlings (30 cm tall) of D. hupeana. Healthy leaves were wounded with a sterile needle and then inoculated with 10 μL spore suspension (106 conidia per mL). Controls were treated with sterile water. All plants were covered with transparent plastic bags and incubated in a greenhouse at 28°C with a 12 h photoperiod (relative humidity > 80%). Within five days, the inoculated leaves developed lesions similar to those observed in the field, whereas controls were asymptomatic. The experiments repeated three times showed similar results. The infection rate was 100%. C. fructicola was re-isolated from the lesions, whereas no fungus was isolated from control leaves. C. fructicola can cause leaf diseases in a variety of hosts, including Aesculus chinensis (Sun et al. 2020), Peucedanum praeruptorum (Ma et al. 2020), and Mandevilla × amabilis (Sun et al. 2020). C. brevisporum and C. gigasporum were also reported to infect Dalbergia odorifera (Chen et al. 2021; Wan et al. 2018). However, This is the first report of C. fructicola associated with leaf spot disease on D. hupeana in China. These results will help to develop effective strategies for appropriately managing this newly emerging disease.

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First report of Colletotrichum fructicola and Colletotrichum nymphaeae causing leaf spot on Rubus corchorifolius in Zhejiang province, China

Plant Disease ◽

10.1094/pdis-01-21-0198-pdn ◽

2021 ◽

Author(s):

Ju Wu ◽

Hanrong Wang ◽

Li Fang ◽

Yunye Xie ◽

Lianping Wang

Keyword(s):

Leaf Spot ◽

Type Strain ◽

Disease Incidence ◽

Morphological Characteristics ◽

Pathogenic Fungi ◽

Zhejiang Province ◽

Colletotrichum Acutatum ◽

Leaf Spot Disease ◽

Conidial Suspension ◽

First Report

Rubus corchorifolius is one of the most economically important fruit trees, (Tian et al. 2021). A severe leaf spot disease on leaves of R. corchorifolius was observed in Longquan county, Zhejiang province (118°42’E, 27°42’N) in 2019, with disease incidence of more than 20% on affected plants. The symptoms on leaves of the naturally affected plants were early necrotic lesion with white centers, surrounded by yellow halos (< 5 mm). Later, lesions were expanded with yellowish-brown centers, surrounded by yellow halos (< 5 mm). Putative pathogenic fungi were isolated as described by Fang (1998) and two pure single-colony fungal strains (FPZ1 and FPZ2) were selected for further analysis. The fungi was cultured on potato dextrose agar (PDA) medium for 6 days, at 25°C. The colonies had gray-green centers, white aerial mycelium and gelatinous orange conidial masses. The conidia were unicellular, smooth-walled, hyaline, cylindrical with obtuse to rounded ends, the size 10.15 to 14.09 µm (mean = 12.95 µm, n = 50) × 4.36 to 6.19 µm (mean = 5.19 µm, n = 50) were single, brown to dark brown, ovoid or irregular in shape, and 5.59 to 12.99 µm (mean = 8.77 µm, n = 50) × 4.68 to 10.36 µm (mean = 6.50 µm, n = 50). The characteristics of FPZ1 were consistent with the description of species in the Colletotrichum gloeosporioides complex (Weir et al. 2012). The conidia of FPZ2 were hyaline, smooth-walled, one-celled, fusiform, the size 9.34 to 14.09 µm (mean = 11.92 µm, n = 50) × 3.26 to 6.15 µm (mean = 4.89 µm, n = 50). Appressoria were single, darker brown, elliptical or irregular in outline, and 4.49 to 15.06 µm (mean = 9.88 µm, n = 50) × 3.23 to 7.42 µm (mean = 5.72 µm, n = 50) in size. The characteristics of FPZ2 were consistent with species of the Colletotrichum acutatum complex (Damn et al. 2012). For molecular identification of strains, the internal transcribed spacer (ITS), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), beta-tubulin (TUB), chitin synthase (CHS-1), and actin (ACT) genes were sequenced (Weir et al. 2012). For the strain FPZ1, the five sequences obtain were deposited in GenBank (MT846907, MT849313, MT849317, MT849315 and MT849319, respectively). A BLAST search of FPZ1 sequences showed 99% identity with the five loci sequences of type strain C. fructicola ICMP 18581 (JX010165, JX010033, JX010405, JX009866 and FJ907426) (Jayawardena et al. 2016). Similarly, for the strain FPZ2, the five sequences (MT846885, MT849314, MT849318, MT849316 and MT849320, respectively) had 99% identity with the five loci sequences of type strain C. nymphaeae CBS 515.78 (JQ948197, JQ948527, JQ949848, JQ948858 and JQ949518, respectively) (Jayawardena et al. 2016). Based on morphological characteristics and phylogenetic analysis, FPZ1 was identified as C. fructicola and FPZ2 as C. nymphaeae, respestively. For pathogenicity tests, 10 μL conidial suspension (1 × 106 conidia per ml) of FPZ1 was used to inoculate five healthy, non-wounded detached leaves, while five leaves inoculated with sterilized water served as control. The experiment was repeated three times, and all leaves were kept on a mist bench at 27°C and relative humidity 80% for 6 days. The inoculation sites of both FPZ1 and FPZ2 became brown and necrotic, while control leaves developed no symptoms. C. fructicola and C. nymphaeae were re-isolated from the lesions of inoculated leaves, fulfilling Koch’s postulates. To our knowledge, this is the first report of C. fructicola and C. nymphaeae causing leaf spot on Rubus corchorifolius in China, and reports on the prevalence of C. gloeosporioides and C. acutatum species complexes will be beneficial to management of anthracnose in R. corchorifolius.

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First report of Colletotrichum fioriniae causing anthracnose on mu oil tree in China

Plant Disease ◽

10.1094/pdis-03-21-0502-pdn ◽

2021 ◽

Author(s):

Yang Zhang ◽

Guangqiang Li ◽

Dou Yang ◽

Ruoling Zhang ◽

Songze Wan

Keyword(s):

Leaf Spot ◽

Disease Incidence ◽

Juglans Regia ◽

Morphological Characteristics ◽

Leaf Spot Disease ◽

Pathogenicity Test ◽

First Report ◽

Spot Disease ◽

Initial Symptoms ◽

Oil Plant

Mu oil tree (Vernicia montana) is an economically important woody oil plant, which is widely distributed in southern China. In mid-May 2020, a leaf spot disease was observed on the leaves of mu oil tree in Taihe County in Jiangxi Province, China (26°55′25.55″N, 114°49′5.85″E). The disease incidence was estimated to be above 40%. Initial symptoms were circular red-brown spots which were 1-2 mm in diameter, then enlarged with red-brown center. In later stages, the spots coalesced and formed large patches, and subsequently red-brown centers of lesions gradually dried and fell out, forming a “shot hole” appearance. To identify the pathogen, diseased leaves were collected from Taihe County. Leaf tissues (5 × 5 mm) were cut from the margins of typical symptomatic lesions, surface- sterilized in 75% ethanol for 30 seconds and 3% sodium hypochlorite for 60 seconds, then rinsed with sterile distilled water three times. Leaf pieces were placed on potato dextrose agar (PDA; 1.5%, Difco-BD Diagnostics) and incubated at 25 °C in the dark. Pure cultures were obtained from individual conidia by recovering single spores. On PDA, colonies were initially white and cottony. The mycelia then became pinkish to deep-pink with time at the center on the front side and pink on the reverse side. Colonies produced pale orange conidial masses after 9 days. Conidia were fusiform with acute ends, smooth-walled, hyaline, and measured 3.6–5.5 × 8.1–14.5 µm (4.5 ± 0.5 × 10.6 ± 1.0 µm, n = 100). The morphological characteristics of the isolate matched the descriptions of Colletotrichum acutatum complex (Damm et al. 2012). For molecular identification, the internal transcribed spacer (ITS) region, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), chitin synthase (CHS-1), beta-tubulin 2 (TUB2), and actin (ACT) were sequenced using the primers ITS1/ITS4, GDF/GDR, CHS-79F/CHS-345R, T1/Bt2b, ACT-512F/ACT-783R, respectively (Weir et al. 2012). The obtained sequences were deposited into the GenBank [accession nos. MW584317 (ITS); MW656269 (GAPDH); MW656270 (TUB2); MW656268 (CHS-1); MW656267 (ACT)]. All the sequences showed 94 to 100% similarity with those of C. fioriniae. A neighbor-joining phylogenetic tree was generated by combining all the sequenced loci using MEGA7.0 (Kumar et al. 2016). The isolate TH-M4 clustered with C. fioriniae, having 99% bootstrap support. Base on the morphology and multi-gene phylogeny, isolate TH-M4 was identified as C. fioriniae (Damm et al. 2012). To confirm pathogenicity, 20 healthy leaves of 10 mu oil trees (3-year-old) grown outdoors were inoculated with a drop of spore suspension (106 conidia per mL) of the isolate TH-M4 in September 2020. Another 10 plants were inoculated with sterile water as the control. The leaves were wounded with a sterile toothpick. All the inoculated leaves were covered with black plastic bags to maintain humidity for 2 days. The pathogenicity test was repeated twice. The resulting symptoms were similar to those on the original infected plants, whereas the control leaves remained asymptomatic. The same fungus was re-isolated from the lesions on the inoculated plant, fulfilling Koch’s postulates. C. fioriniae has been recorded as anthracnose pathogen on Mahonia aquifolium (Garibaldi et al. 2020), Paeonia lactiflora (Park et al. 2020), Solanum melongena (Xu et al. 2020), and Juglans regia (Varjas et al. 2020). To our knowledge, this is the first report of C. fioriniae associated with leaf spot disease on mu oil tree in China. This study provided crucial information for epidemiologic studies and appropriate control strategies for this oil plant disease.

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First Report of Leaf Spot in Farfugium japonicum Caused by Alternaria tenuissima in Korea

Plant Disease ◽

10.1094/pdis-03-13-0253-pdn ◽

2013 ◽

Vol 97 (10) ◽

pp. 1382-1382 ◽

Cited By ~ 1

Author(s):

J. H. Lee ◽

D. S. Kim ◽

H. J. Cho ◽

G.-H. Gang ◽

Y.-S. Kwak

Keyword(s):

Leaf Spot ◽

Morphological Characteristics ◽

Spot Size ◽

Plant Diseases ◽

Universal Primers ◽

Leaf Spot Disease ◽

Alternaria Tenuissima ◽

First Report ◽

Spot Disease ◽

Farfugium Japonicum

Farfugium japonicum (L.) Kitam (common name: Leopard plant) is known as a medical herb and belongs to family Asteraceae (1). In June 2012, a leaf spot disease was observed on the leaf surface of F. japonicum at a forest research plot Jinju, Gyeongnam province, Korea. More than 95% of F. japonicum plants were infected and leaf spot symptom appeared in the regions under our investigation. Light brownish symptoms initially developed and the spot size gradually increased and turned dark brown with an irregular shape as the disease progressed (spot size 1 to 10 mm in diameter). At the late stage of disease, spots became hollow and completely dehydrated. The infected leaves were easily crumbled, possibly due to dryness. To isolate the causal agent, the infected leaves were surface disinfected and pieces of leaves were placed on water agar (WA). Nine isolates were isolated from 10 pieces of the infected leaves. Fungi mycelia from the WA were transferred on potato dextrose agar (PDA) and incubated at 28°C for 7 days. The colonies were purple navy to black and conidia spores developed on the media. The morphological characteristics of spores were multi-septate, dark brown, pyriform, and 6.7 to 12.8 × 22.2 to 38.4 μm. The spores had 1 to 4 transverse and 0 to 3 longitudinal septa. The morphological characteristics of the isolates showed considerably similar to well-known Alternaria tenuissima (2). The leaf spot disease caused by A. cinerariae of F. japonicum was reported from Japan (3). Spores of A. cinerariae are golden brown to brown with 3 to 9 transverse and 0 to 6 longitudinal septa and are 87.5 × 28.7 μm (avg.) (3). To verify pathogenicity of the isolate, the pure cultured fungi on the PDA medium was taken (4 mm in diameter) and placed on healthy leaves of Leopard plant. The artificially inoculated leaves were placed on wet filter paper in Petri dishes and incubated at 25°C and 80% humidity. At 7 days after inoculation, similar disease symptoms developed on 8 out of 10 infected Leopard plant leaves. The pathogen was reisolated from artificially infected leaves. To identify in molecular biology level, genomic DNA was extracted and the ITS-rDNA region was amplified using universal primers ITS1 and ITS4. The amplified PCR product was purified and sequenced (528 bp) with ITS1 and ITS4 primers for both directions and then deposited in GenBank (Accession No. KC415611.1). The BLAST search showed that it matched previously reported A. tenuissima with 100% identity. To the best of our knowledge, this is the first report of Leopard plant leaf spot disease in Korea. References: (1) E. Y. Kim et al. J. Ethnopharmacol. 146:40, 2013. (2) E. G. Simmons. Page 1 in: Alternaria Biology, Plant Diseases and Metabolites. J. Chelchowski and A. Visconti, eds. Elsevier, Amsterdam, 1992. (3) T. Sakoda et al. Res. Bull. Pl. Prot. Japan 46:73, 2010.

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