First Report of Zucchini Collapse by Fusarium solani f. sp. cucurbitae Race 1 and Plectosporium tabacinum in Italy

Zucchini plant collapse has been often associated with Fusarium solani f. sp. cucurbitae race 1, which is the causal agent of Fusarium crown and foot rot disease of cucurbits. In Italy, F. solani f. sp. cucurbitae race 1 has been reported on zucchini (Cucurbita pepo) in a greenhouse in the Tuscany Region (4). In spring 2005, a severe outbreak was observed on zucchini in a vast area of cultivation in the province of Venice. Isolations from necrotic vessels gave more than 20 single-spore cultures. On the basis of morphological characteristics, they were identified as F. solani (2) and Plectosporium tabacinum (3). The internal transcribed spacer (ITS) region of rDNA was amplified and sequenced. A fragment of 454 and 531 bp was 99% homologous with sequence PSU66732 and AF150472 of F. solani f. sp. cucurbitae race 1 and P. tabacinum, respectively, in the NCBI database. The nucleotide sequences have been assigned Accession No. AM408782 for F. solani f. sp. cucurbitae race 1 and AM408781 for P. tabacinum. Pathogenicity tests were conducted with four isolates of each species on 15-day-old zucchini plants and on fruit. Plants were inoculated by dipping the roots in a conidial suspension of 106 spores ml-1 for 10 min. Control plants were dipped in sterile water. Five replicates for the inoculated and control plants were used. All plants were maintained in a greenhouse at approximately 24°C. After 14 days, inoculations with F. solani f. sp. cucurbitae race 1 gave symptoms of a cortical rot at the base of the stem with a progressive yellows and wilting of leaves, while plants inoculated with P. tabacinum displayed a moderate wilting. Fruit were washed under running water, disinfected with a solution of 3% sodium hypochlorite and 5% ethanol for 1 min, and inoculated with 6-mm-diameter mycelial plugs cut from the margin of 10-day-old cultures grown on PDA. Plugs were inserted into holes (approximately 2 mm deep) made with a sterile 7-mm-diameter cork borer. Five replicates per isolate were used. Fruit were kept at room temperature (22 to 24°C) in a moist chamber. All isolates induced symptoms of fruit rotting 10 days after inoculation. All controls remained healthy. The colonies reisolated from the inoculated plants and fruit were morphologically identical to the original isolates. The results obtained proved that F. solani f. sp. cucurbitae race 1 can be considered the major pathogen in zucchini collapse, at the same time P. tabacinum may play a role in this syndrome as reported for other cucurbits (1). To our knowledge, this is the first report of zucchini plant collapse caused by F. solani f. sp. cucurbitae race 1 and P. tabacinum, and the first report of P. tabacinum on zucchini in Italy. References: (1) V. J. Garcia-Jimenez et al. EPPO Bull. 30:169, 2000. (2) P. E. Nelson et al. Fusarium Species: An Illustrated Manual for Identification. Pennsylvania State University, University Park, 1983. (3) M. E. Palm et al. Mycologia 87:397, 1995. (4) G. Vannacci and P. Gambogi. Phytopathol. Mediterr. 19:103, 1980.

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First Report of Tuber Rot Disease of Kala Zeera Caused by a Member of the Fusarium solani Species Complex in India

Plant Disease ◽

10.1094/pdis-02-12-0148-pdn ◽

2012 ◽

Vol 96 (7) ◽

pp. 1067-1067 ◽

Cited By ~ 1

Author(s):

V. Gupta ◽

D. John ◽

V. K. Razdan ◽

S. K. Gupta

Keyword(s):

Fusarium Solani ◽

Species Complex ◽

Morphological Characteristics ◽

Distilled Water ◽

Conidial Suspension ◽

Potato Dextrose Broth ◽

First Report ◽

Fusarium Solani Species Complex ◽

Rot Disease ◽

Tuber Rot

Bunium persicum (Kala zeera, also black cumin) is an economically important culinary crop that is cultivated for its seed pods and its tuberlike roots. In India, high-altitude regions of Himachal Pradesh, including the Padder valley and the Gurez area of Jammu and Kashmir, are areas of kalazeera production (3). In 2008 to 2009, tuber rot disease of kala zeera was observed during the late spring season in the Padder valley. Symptomatic plants were distributed in localized areas in the field and the symptoms included drying of foliage and rotting of tubers. White mycelia were found on the tubers at the late stages of disease development. Incidence of infection in the surveyed area was 80 to 90%. Yield losses were 50 to 60%. To isolate the causal pathogen, we cultured tissues from symptomatic tubers. Small bits of the infected tissue were surface disinfested in 0.1% mercuric chloride, followed by rinsing three times in sterile distilled water. The surface disinfested tissues were plated on potato dextrose agar (PDA) and incubated at 27°C for 4 days. Pure cultures of the mycelium from the diseased tissues were transferred to a second set of PDA for species identification. The fungus produced three types of spores: small, one-celled, oval microconidia; large, slightly curved, septate macroconidia; and rounded, thick-walled chlamydospores. Microconidia were mostly non-septate and 8.91 to 15.73 × 2.3 to 3.5 μm, whereas macroconidia were three- to five-septate and were 35.55 to 54.74 × 3.91 to 6.5 μm. On the basis of morphological characteristics (1), the fungus was identified and deposited as a member of the Fusarium solani species complex in the Indian Type Culture Collection, New Delhi (ID No. 8422.11). To confirm pathogenicity, healthy tubers were submerged for 20 min in a conidial suspension of the isolated fungus (1 × 105 cfu/ml), which was prepared in potato dextrose broth, incubated for 10 days at 27°C, and centrifuged at 140 rpm. Noninoculated controls were submerged in distilled water. Inoculated and control tubers were then planted in separate pots filled with sterilized soil and kept in a shade house. Symptoms appeared on inoculated tubers 9 to 10 days after planting. Signs of the pathogen in the form of mycelia were present. The tubers rotted and died 12 to 15 days after inoculation. Control tubers did not display any symptoms. F. solani species complex was reisolated from inoculated tubers, fulfilling Koch's postulates. F. solani has been reported to cause corm rot on gladiolus and saffron (2). To our knowledge, this is the first report of the F. solani species complex as pathogenic to tubers of kalazeera in India. References: (1) C. Booth. The Genus Fusarium. 47, 1971. (2) L. Z. Chen et al. J. Shanghai Agric. College 12:240, 1994. (3) K. S. Panwar et al. Agriculture Situation in India. 48:151, 1993.

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First report of Fusarium incarnatum causing fruit rot of litchi in China

Plant Disease ◽

10.1094/pdis-11-20-2393-pdn ◽

2021 ◽

Author(s):

Zhaoyin Gao ◽

Jiaobao Wang ◽

Zhengke Zhang ◽

Min Li ◽

Deqiang Gong ◽

...

Keyword(s):

Southern China ◽

Fusarium Species ◽

Morphological Characteristics ◽

Fruit Rot ◽

Conidial Suspension ◽

Single Spore ◽

Internal Transcribed Spacer Region ◽

First Report ◽

Litchi Fruit ◽

His3 Gene

Litchi (Litchi chinensis Sonn.) is an indigenous tropical and subtropical fruit in Southern China with an attractive appearance, delicious taste, and good nutritional value (Jiang et al. 2003). In March 2020, brown rots were observed on nearly ripe litchi fruits (cv. Guihuaxiang) in an orchard of Lingshui county, Hainan province of China (18.615877° N, 109.948871° E). About 5% fruits were symptomatic in the field, and the disease caused postharvest losses during storage. The initial infected fruits had no obvious symptoms on the outer pericarp surfaces, but appeared irregular, brown to black-brown lesions in the inner pericarps around the pedicels. Then lesions expanded and became brown rots. Small tissues (4 mm × 4 mm) of fruit pericarps were cut from symptomatic fruits, surface-sterilized in 1% sodium hypochlorite for 3 min, rinsed in sterilized water three times, plated on potato dextrose agar (PDA) and incubated at 28℃ in the darkness. Morphologically similar colonies were isolated from 85% of 20 samples after 4 days of incubation. Ten isolates were purified using a single-spore isolation method. The isolates grown on PDA had abundant, fluffy, whitish to yellowish aerial mycelia, and the reverse side of the Petri dish was pale brown. Morphological characteristics of conidia were further determined on carnation leaf-piece agar (CLA) (Leslie et al. 2006). Macroconidia were straight to slightly curved, 3- to 5-septates with a foot-shaped basal cell, tapered at the apex, 2.70 to 4.43 µm × 18.63 to 37.58 µm (3.56 ± 0.36 × 28.68 ± 4.34 µm) (n = 100). Microconidia were fusoid to ovoid, 0- to 1-septate, 2.10 to 3.57 µm × 8.18 to 18.20 µm (2.88 ± 0.34 × 11.71 ± 1.97 µm) (n = 100). Chlamydospores on hyphae singly or in chains were globose, subglobose, or ellipsoidal. Based on cultural features and morphological characteristics, the fungus was identified as a Fusarium species (Leslie et al. 2006). To further confirm the pathogen, DNA was extracted from the 7-day-old aerial mycelia of three isolates (LZ-1, LZ-3, and LZ-5) following Chohan et al. (2019). The sequences of the internal transcribed spacer region of rDNA (ITS), translation elongation factor-1 alpha (tef1) gene, and histone H3 (his3) gene were partially amplified using primers ITS1/ITS4, EF1-728F/EF1-986R, and CYLH3F/CYLH3R, respectively (Funnell-Harris et al. 2017). The nucleotide sequences were deposited in GenBank (ITS: 515 bp, MW029882, 533 bp, MW092186, and 465 bp, MW092187; tef1: 292 bp, MW034437, 262 bp, MW159143, and 292 bp, MW159141; his3: 489 bp, MW034438, 477 bp, MW159142, and 474 bp, MW159140). The ITS, tef1, and his3 genes showed 99-100% similarity with the ITS (MH979697), tef1 (MH979698), and his3 (MH979696) genes, respectively of Fusarium incarnatum (TG0520) from muskmelon fruit. The phylogenetic analysis of the tef1 and his3 gene sequences showed that the three isolates clustered with F. incarnatum. Pathogenicity tests were conducted by spraying conidial suspension (1×106 conidia/ml) on wounded young fruits in the orchid. Negative controls were sprayed with sterilized water. Fruits were bagged with polythene bags for 24 hours and then unbagged for 10 days. Each treatment had 30 fruits. The inoculated fruits developed symptoms similar to those observed in the orchard and showed light brown lesions on the outer pericarp surfaces and irregular, brown to black-brown lesions in the inner pericarps, while the fruits of negative control remained symptomless. The same fungus was successfully recovered from symptomatic fruits, and thus, the test for the Koch’s postulates was completed. F. semitectum (synonym: F. incarnatum) (Saha et al. 2005), F. oxysporum (Bashar et al. 2012), and F. moniliforme (Rashid et al. 2015) have been previously reported as pathogens causing litchi fruit rots in India and Bangladesh. To our knowledge, this is the first report of Fusarium incarnatum causing litchi fruit rot in China.

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First report of Fusarium falciforme (FSSC 3+4) causing root rot on chickpea in Mexico

Plant Disease ◽

10.1094/pdis-05-21-1123-pdn ◽

2021 ◽

Author(s):

Sixto Velarde Felix ◽

Victor Valenzuela ◽

Pedro Ortega ◽

Gustavo Fierros ◽

Pedro Rojas ◽

...

Keyword(s):

Fusarium Solani ◽

Root Rot ◽

Species Complex ◽

Elongation Factor ◽

Morphological Characteristics ◽

Aerial Mycelium ◽

Pathogenicity Test ◽

Conidial Suspension ◽

Single Spore ◽

First Report

Chickpea (Cicer aretinium L.) is a legume crop of great importance worldwide. In January 2019, wilting symptoms on chickpea (stunted grow, withered leaves, root rot and wilted plants) were observed in three fields of Culiacan Sinaloa Mexico, with an incidence of 3 to 5%. To identify the cause, eighty symptomatic chickpea plants were sampled. Tissue from roots was plated on potato dextrose agar (PDA) medium. Typical Fusarium spp. colonies were obtained from all root samples. Ten pure cultures were obtained by single-spore culturing (Ff01 to Ff10). On PDA the colonies were abundant with white aerial mycelium, hyphae were branched and septae and light purple pigmentation was observed in the center of old cultures (Leslie and Summerell 2006). From 10-day-old cultures grown on carnation leaf agar medium, macroconidias were falciform, hyaline, with slightly curved apexes, three to five septate, with well-developed foot cells and blunt apical cells, and measured 26.6 to 45.8 × 2.2 to 7.0 μm (n = 40). The microconidia (n = 40) were hyaline, one to two celled, produced in false heads that measured 7.4 to 20.1 (average 13.7) μm × 2.4 to 8.9 (average 5.3) μm (n = 40) at the tips of long monophialides, and were oval or reniform, with apexes rounded, 8.3 to 12.1 × 1.6 to 4.7 μm; chlamydospores were not evident. These characteristics fit those of the Fusarium solani (Mart.) Sacc. species complex, FSSC (Summerell et al. 2003). The internal transcribed spacer and the translation elongation factor 1 alpha (EF1-α) genes (O’Donnell et al. 1998) were amplified by polymerase chain reaction and sequenced from the isolate Ff02 and Ff08 (GenBank accession nos. KJ501093 and MN082369). Maximum likelihood analysis was carried out using the EF1-α sequences (KJ501093 and MN082369) from the Ff02 and Ff08 isolates and other species from the Fusarium solani species complex (FSSC). Phylogenetic analysis revealed the isolate most closely related with F. falciforme (100% bootstrap). For pathogenicity testing, a conidial suspension (1x106 conidia/ml) was prepared by harvesting spores from 10-days-old cultures on PDA. Twenty 2-week-old chickpea seedlings from two cultivars (P-2245 and WR-315) were inoculated by dipping roots into the conidial suspension for 20 min. The inoculated plants were transplanted into a 50-hole plastic tray containing sterilized soil and maintained in a growth chamber at 25°C, with a relative humidity of >80% and a 12-h/12-h light/dark cycle. After 8 days, the first root rot symptoms were observed on inoculating seedlings and the infected plants eventually died within 3 to 4 weeks after inoculation. No symptoms were observed plants inoculated with sterilized distilled water. The fungus was reisolated from symptomatic tissues of inoculated plants and was identified by sequencing the partial EF1-α gene again and was identified as F. falciforme (FSSC 3 + 4) (O’Donnell et al. 2008) based on its morphological characteristics, genetic analysis, and pathogenicity test, fulfilling Koch’s postulates. The molecular identification was confirmed via BLAST on the FusariumID and Fusarium MLST databases. Although FSSC has been previously reported causing root rot in chickpea in USA, Chile, Spain, Cuba, Iran, Poland, Israel, Pakistan and Brazil, to our knowledge this is the first report of root rot in chickpea caused by F. falciforme in Mexico. This is important for chickpea producers and chickpea breeding programs.

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First Report of Root Rot on Pulsatilla koreana Caused by Fusarium oxysporum in China

Plant Disease ◽

10.1094/pdis-09-12-0849-pdn ◽

2013 ◽

Vol 97 (3) ◽

pp. 425-425 ◽

Cited By ~ 1

Author(s):

D. Su ◽

J. F. Fu

Keyword(s):

Fusarium Oxysporum ◽

Root Rot ◽

Its Region ◽

Liaoning Province ◽

Conidial Suspension ◽

Single Spore ◽

Laboratory Manual ◽

First Report ◽

Medicinal Value ◽

Rot Disease

Windflowers (Pulsatilla spp.) are perennial medicinal plants in the family Ranunculaceae with high economic as well as medicinal value in China. It is commonly used as traditional Chinese medicine (1). In May 2012, a root rot disease was observed on windflower (Pulsatilla koreana Nakai) at flowering stages in fields of Liaoning Province, China. The diseased area was estimated to be over 500 ha in the province and the yield was reduced by 30% on average with up to 45% yield losses in some fields. As the disease progressed, brown lesion production extended onto lateral and main roots, and aboveground tissues shriveled and decayed; in severe cases, white mycelium was clearly visible on diseased root tissue. Isolations from symptomatic roots were made on potato dextrose agar (PDA) and single-spore cultures were obtained. Colonies were initially white, but became pale violet with age, and purple pigments were produced in the agar. Microconidia were abundant, unicellular, oval to reniform, and ranged from 5.6 to 13.1 (9.3) × 2.8 to 4.2 (3.2) μm. Macroconidia were sparse, three-septate, slightly curved, and ranged from 21.9 to 39.4 (31.2) × 3.4 to 4.5 (3.9) μm. The isolated fungus was morphologically similar to Fusarium oxysporum (2). Two isolates were selected for molecular identification, and the internal transcribed spacer (ITS) region of rDNA was amplified using the primers ITS1/ITS4 (3) and sequenced. The obtained sequences (GenBank Accession Nos. JX669525 and JX669526) showed 99% homology with the sequences of F. oxysporum in GenBank (GQ121303). For pathogenicity tests, the isolate was cultured on PDA for 10 days at 25°C. Inoculations were performed on 10 healthy P. koreana plants by spraying a conidial suspension (2.0 × 105 microconidia ml–1) on roots previously wounded with a metal needle. Ten non-treated plants used as controls were sprayed with distilled water. The inoculated plants were incubated at 25°C under conditions of 12/12 h (light and dark). After 2 weeks, root rot symptoms were similar to the original symptoms observed under field conditions. No disease was observed on water-inoculated control plants. The same fungus was reisolated from the roots of infected plants, satisfying Koch's postulates. To our knowledge, this is the first report of F. oxysporum on P. koreana in China. The disease was hitherto scarcely reported in any other countries, and may deserve more attention in the future. References: (1) S. C. Bang et al. J. Nat. Prod. 68:268, 2005. (2) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Professional, Ames, IA, 2006. (3) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990.

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First Report of Maize Stalk Rot Caused by Fusarium nelsonii in China

Plant Disease ◽

10.1094/pdis-01-21-0089-pdn ◽

2021 ◽

Author(s):

Xiaojie Zhang ◽

Cheng Guo ◽

Chunming Wang ◽

Tianwang Zhou

Keyword(s):

Fusarium Species ◽

Morphological Characteristics ◽

Molecular Characteristics ◽

Conidial Suspension ◽

Basal Cells ◽

Sterile Water ◽

Single Spore ◽

First Report ◽

Stalk Rot ◽

Maize Plants

Maize (Zea Mays L.) is one of the main crops in Ningxia Province, China, and stalk rot has become a serious disease of maize in this area. Infected plants showed softening of the stalks at lower internodes, which lodged easily and died prematurely during grain filling, and the pith tissue internally appeared to be disintegrating and slightly brown to reddish. In September 2018, symptomatic tissue was collected from seventeen locations in Ningxia. The incidence ranged from 5% to 40% in surveyed fields, reaching as high as 86% in certain plots. The discolored stalk pith tissues from the lesion region were cut into small pieces (approximately 0.5 × 0.2 cm), superficially disinfected with 75% ethanol for 1 min and rinsed three times with sterile water before plating on potato dextrose agar (PDA) medium with chloromycetin. The purified strains were obtained by single-spore separation and transferred to PDA and carnation leaf agar (CLA) medium. Morphological and molecular characteristics confirmed the presence of nine Fusarium species in these samples, including Fusarium graminearum species complex and Fusarium verticillioides. Four isolates of Fusarium nelsonii were recovered from samples collected in Shizuishan and Wuzhong. On PDA plates, the floccose to powdery, white to rose-colored aerial mycelia were produced and covered plates after 8 days of incubation, producing abundant mesoconidia and chlamydospores. Mesoconidia were fusiform or lanceolate until slightly curved with 0-3 septa, and chlamydospores were initially smooth and transparent, and became verrucous and light brown. Macroconidia produced in CLA were straight or curved and falcate, usually having 3-5 septa, with beak-shaped strongly curved apical cells and foot-shaped basal cells. Two isolates (SS-1-7 and ZY-2-2) were selected for molecular identification, and the total DNA was extracted using a fungal genomic DNA separation kit (Sangon Biotechnology, Shanghai, China). Sequence comparison of EF-1α (GenBank accession numbers MW294197 and MW294198) and RPB2 (Accession MW294176 and MW294177) genes showed 97% homology with the sequences of F. nelsonii reported in GenBank (accession MN120760 for TEF and accession MN120740 for RPB2). Pathogenicity tests with two isolates (SS-1-7 and ZY-2-2) were performed by individually inoculating five 10-leaf stage maize plants at between the 2nd and 3rd stem nodes from the soil level with 20 μl conidial suspension at a concentration of 106 conidia/ml as described by Zhang et al. (2016). Five maize plants inoculated with sterile water were used as controls. The inoculated plants were kept at 25 ± 0.5°C in the greenhouse with a photoperiod of 12 h. After 30 days, all plants inoculated with the conidial suspension formed an internal dark brown necrotic area around the inoculation site, whereas the control plants showed no symptoms. The pathogen was re-isolated from the necrotic tissue of the inoculated plants and identified by morphological characteristics as F. nelsonii. This species was first described by Marasas et al. (1998), and it is expanding its host range and has been isolated from sorghum, Medicago, wheat, and cucumber (Ahmad et al. 2020). The pathogen should be paid more attention owing to a serious risk of trichothecene and aflatoxin contamination (Astoreca et al. 2019; Lincy et al. 2011). To our knowledge, this is the first report of maize stalk rot caused by F. nelsonii in China. References: Ahmad, A., et al. 2020. Plant disease.1542 https://doi.org/10.1094/PDIS-11-19-2511-PDN Astoreca, A. L., et al. 2019. Eur. J. Plant Pathol. 155:381. Lincy, S. V., et al. 2011. World J. Microbiol. Biotechnol. 27:981. Marasas, W. F. O., et al. 1998. Mycologia 90:505. Zhang, Y., et al. 2016. PLoS Pathog. 12:e1005485. Funding: This research was financially supported by National R & D Plan of China (No.2019QZKK0303); Ningxia Agriculture and Forestry Academy Science and Technology Cooperation Project (DW-X-2018019)

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First Report of Ramularia coleosporii causing Leaf Spot on Perilla frutescens in Korea

Plant Disease ◽

10.1094/pdis-01-21-0223-pdn ◽

2021 ◽

Author(s):

Md Aktaruzzaman ◽

Tania Afroz ◽

Hyo-Won Choi ◽

Byung Sup Kim

Keyword(s):

Leaf Spot ◽

Ipomoea Batatas ◽

Rust Fungus ◽

Morphological Characteristics ◽

Perilla Frutescens ◽

Herbaceous Plant ◽

Pathogenicity Test ◽

Conidial Suspension ◽

Single Spore ◽

First Report

Perilla (Perilla frutescens var. japonica), a member of the family Labiatae, is an annual herbaceous plant native to Asia. Its fresh leaves are directly consumed and its seeds are used for cooking oil. In July 2018, leaf spots symptoms were observed in an experimental field at Gangneung-Wonju National University, Gangneung, Gangwon province, Korea. Approximately 30% of the perilla plants growing in an area of about 0.1 ha were affected. Small, circular to oval, necrotic spots with yellow borders were scattered across upper leaves. Masses of white spores were observed on the leaf underside. Ten small pieces of tissue were removed from the lesion margins of the lesions, surface disinfected with NaOCl (1% v/v) for 30 s, and then rinsed three times with distilled water for 60 s. The tissue pieces were then placed on potato dextrose agar (PDA) and incubated at 25°C for 7 days. Five single spore isolates were obtained and cultured on PDA. The fungus was slow-growing and produced 30-50 mm diameter, whitish colonies on PDA when incubated at 25ºC for 15 days. Conidia (n= 50) ranged from 5.5 to 21.3 × 3.5 to 5.8 μm, were catenate, in simple or branched chains, ellipsoid-ovoid, fusiform, and old conidia sometimes had 1 to 3 conspicuous hila. Conidiophores (n= 10) were 21.3 to 125.8 × 1.3 to 3.6 μm in size, unbranched, straight or flexuous, and hyaline. The morphological characteristics of five isolates were similar. Morphological characteristics were consistent with those described for Ramularia coleosporii (Braun, 1998). Two representative isolates (PLS 001 & PLS003) were deposited in the Korean Agricultural Culture Collection (KACC48670 & KACC 48671). For molecular identification, a multi-locus sequence analysis was conducted. The internal transcribed spacer (ITS) regions of the rDNA, partial actin (ACT) gene and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene were amplified using primer sets ITS1/4, ACT-512F/ACT-783R and gpd1/gpd2, respectively (Videira et al. 2016). Sequences obtained from each of the three loci for isolate PLS001 and PLS003 were deposited in GenBank with accession numbers MH974744, MW470869 (ITS); MW470867, MW470870 (ACT); and MW470868, MW470871 (GAPDH), respectively. Sequences for all three genes exhibited 100% identity with R. coleosporii, GenBank accession nos. GU214692 (ITS), KX287643 (ACT), and 288200 (GAPDH) for both isolates. A multi-locus phylogenetic tree, constructed by the neighbor-joining method with closely related reference sequences downloaded from the GenBank database and these two isolates demonstrated alignment with R. coleosporii. To confirm pathogenicity, 150 mL of a conidial suspension (2 × 105 spores per mL) was sprayed on five, 45 days old perilla plants. An additional five plants, to serve as controls, were sprayed with sterile water. All plants were placed in a humidity chamber (>90% relative humidity) at 25°C for 48 h after inoculation and then placed in a greenhouse at 22/28°C (night/day). After 15 days leaf spot symptoms, similar to the original symptoms, developed on the leaves of the inoculated plants, whereas the control plants remained symptomless. The pathogenicity test was repeated twice with similar results. A fungus was re-isolated from the leaf lesions on the inoculated plants which exhibited the same morphological characteristics as the original isolates, fulfilling Koch’s postulates. R. coleosporii has been reported as a hyperparasite on the rust fungus Coleosporium plumeriae in India & Thailand and also as a pathogen infecting leaves of Campanula rapunculoides in Armenia, Clematis gouriana in Taiwan, Ipomoea batatas in Puerto Rico, and Perilla frutescens var. acuta in China (Baiswar et al. 2015; Farr and Rossman 2021). To the best of our knowledge, this is the first report of R. coleosporii causing leaf spot on P. frutescens var. japonica in Korea. This disease poses a threat to production and management strategies to minimize leaf spot should be developed.

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First Report of Leaf Spot Caused by Botrytis cinerea on Cardamine hupingshanensis in China

Plant Disease ◽

10.1094/pdis-04-21-0698-pdn ◽

2021 ◽

Author(s):

Jun Guo ◽

Jin Chen ◽

Zhao Hu ◽

Jie Zhong ◽

Jun Zi Zhu

Keyword(s):

Botrytis Cinerea ◽

Morphological Characteristics ◽

Gray Mold ◽

Hunan Province ◽

Conidial Suspension ◽

Heat Shock Protein 60 ◽

Basal Cells ◽

Single Spore ◽

Internal Transcribed Spacer Region ◽

First Report

Cardamine hupingshanensis is a selenium (Se) and cadmium (Cd) hyperaccumulator plant distributed in wetlands along the Wuling Mountains of China (Zhou et al. 2018). In March of 2020, a disease with symptoms similar to gray mold was observed on leaves of C. hupingshanensis in a nursery located in Changsha, Hunan Province, China. Almost 40% of the C. hupingshanensis (200 plants) were infected. Initially, small spots were scattered across the leaf surface or margin. As disease progressed, small spots enlarged to dark brown lesions, with green-gray, conidia containing mold layer under humid conditions. Small leaf pieces were cut from the lesion margins and were sterilized with 70% ethanol for 10 s, 2% NaOCl for 2 min, rinsed with sterilized distilled water for three times, and then placed on potato dextrose agar (PDA) medium at 22°C in the dark. Seven similar colonies were consistently isolated from seven samples and further purified by single-spore isolation. Strains cultured on PDA were initially white, forming gray-white aerial mycelia, then turned gray and produced sclerotia after incubation for 2 weeks, which were brown to blackish, irregular, 0.8 to 3.0 × 1.2 to 3.5 mm (n=50). Conidia were unicellular, globose or oval, colourless, 7.5 to 12.0 × 5.5 to 8.3 μm (n=50). Conidiophores arose singly or in group, straight or flexuous, septate, brownish to light brown, with enlarged basal cells, 12.5 to 22.1 × 120.7 to 310.3 μm. Based on their morphological characteristics in culture, the isolates were putatively identified as Botrytis cinerea (Ellis 1971). Genomic DNA of four representative isolates, HNSMJ-1 to HNSMJ-4, were extracted by CTAB method. The internal transcribed spacer region (ITS), glyceraldehyde-3-phosphate dehydrogenase gene (G3PDH), heat-shock protein 60 gene (HSP60), ATP-dependent RNA helicaseDBP7 gene (MS547) and DNA-dependent RNA polymerase subunit II gene (RPB2) were amplified and sequenced using the primers described previously (Aktaruzzaman et al. 2018) (MW820311, MW831620, MW831628, MW831623 and MW831629 for HNSMJ-1; MW314722, MW316616, MW316617, MW316618 and MW316619 for HNSMJ-2; MW820519, MW831621, MW831627, MW831624 and MW831631 for HNSMJ-3; MW820601, MW831622, MW831626, MW831625 and MW831630 for HNSMJ-4). BLAST searches showed 99.43 to 99.90% identity to the corresponding sequences of B. cinerea strains, such as HJ-5 (MF426032.1, MN448500.1, MK791187.1, MH727700.1 and KX867998.1). A combined phylogenetic tree using the ITS, G3PDH, HSP60 and RPB2 sequences was constructed by neighbor-joining method in MEGA 6. It revealed that HNSMJ-1 to HNSMJ-4 clustered in the B. cinerea clade. Pathogenicity tests were performed on healthy pot-grown C. hupingshanensis plants. Leaves were surface-sterilized and sprayed with conidial suspension (106 conidia/ mL), with sterile water served as controls. All plants were kept in growth chamber with 85% humidity at 25℃ following a 16 h day-8 h night cycle. The experiment was repeated twice, with each three replications. After 4 to 7 days, symptoms similar to those observed in the field developed on the inoculated leaves, whereas controls remained healthy. The pathogen was reisolated from symptomatic tissues and identified using molecular methods, confirming Koch’s postulates. B. cinerea has already been reported from China on C. lyrate (Zhang 2006), a different species of C. hupingshanensis. To the best of our knowledge, this is the first report of B. cinerea causing gray mold on C. hupingshanensis in China and worldwide. Based on the widespread damage in the nursery, appropriate control strategies should be adopted. This study provides a basis for studying the epidemic and management of the disease.

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First Report of Leaf Blight Caused by Septoria allii on Garlic Chives in Korea

Plant Disease ◽

10.1094/pdis-08-12-0809-pdn ◽

2013 ◽

Vol 97 (1) ◽

pp. 147-147

Author(s):

J. H. Park ◽

S. E. Cho ◽

K. S. Han ◽

H. D. Shin

Keyword(s):

Disease Incidence ◽

Morphological Characteristics ◽

Its Region ◽

Leaf Blight ◽

Plant Diseases ◽

Cultivated Plants ◽

Sequence Information ◽

Conidial Suspension ◽

Korean Society ◽

First Report

Garlic chives, Allium tuberosum Roth., are widely cultivated in Asia and are the fourth most important Allium crop in Korea. In June 2011, a leaf blight of garlic chives associated with a Septoria spp. was observed on an organic farm in Hongcheon County, Korea. Similar symptoms were also found in fields within Samcheok City and Yangku County of Korea during the 2011 and 2012 seasons. Disease incidence (percentage of plants affected) was 5 to 10% in organic farms surveyed. Diseased voucher specimens (n = 5) were deposited at the Korea University Herbarium (KUS). The disease first appeared as yellowish specks on leaves, expanding to cause a leaf tip dieback. Half of the leaves may be diseased within a week, especially during wet weather. Pycnidia were directly observed in leaf lesions. Pycnidia were amphigenous, but mostly epigenous, scattered, dark brown to rusty brown, globose, embedded in host tissue or partly erumpent, separate, unilocular, 50 to 150 μm in diameter, with ostioles of 20 to 40 μm in diameter. Conidia were acicular, straight to sub-straight, truncate at the base, obtuse at the apex, hyaline, aguttulate, 22 to 44 × 1.8 to 3 μm, mostly 3-septate, occasionally 1- or 2-septate. These morphological characteristics matched those of Septoria allii Moesz, which is differentiated from S. alliacea on conidial dimensions (50 to 60 μm long) (1,2). A monoconidial isolate was cultured on potato dextrose agar (PDA). Two isolates have been deposited in the Korean Agricultural Culture Collection (Accession Nos. KACC46119 and 46688). Genomic DNA was extracted using the DNeasy Plant Mini DNA Extraction Kit (Qiagen Inc., Valencia, CA). The internal transcribed spacer (ITS) region of rDNA was amplified using the ITS1/ITS4 primers and sequenced. The resulting sequence of 482-bp was deposited in GenBank (JX531648 and JX531649). ITS sequence information was at least 99% similar to those of many Septoria species, however no information was available for S. allii. Pathogenicity was tested by spraying leaves of three potted young plants with a conidial suspension (2 × 105 conidia/ml), which was harvested from a 4-week-old culture on PDA. Control leaves were sprayed with sterile water. The plants were placed in humid chambers (relative humidity 100%) for the first 48 h. After 7 days, typical leaf blight symptoms started to develop on the leaves of inoculated plants. S. allii was reisolated from the lesions of inoculated plants, confirming Koch's postulates. No symptoms were observed on control plants. The host-parasite association of A. tuberosum and S. allii has been known only from China (1). S. alliacea has been recorded on several species of Allium, e.g. A. cepa, A. chinense, A. fistulosum, and A. tuberosum from Japan (4) and A. cepa from Korea (3). To the best of our knowledge, this is the first report of S. allii on garlic chives. No diseased plants were observed in commercial fields of garlic chives which involved regular application of fungicides. The disease therefore seems to be limited to organic garlic chive production. References: (1) P. K. Chi et al. Fungous Diseases on Cultivated Plants of Jilin Province, Science Press, Beijing, China, 1966. (2) P. A. Saccardo. Sylloge Fungorum Omnium Hucusque Congnitorum. XXV. Berlin, 1931. (3) The Korean Society of Plant Pathology. List of Plant Diseases in Korea, Suwon, Korea, 2009. (4) The Phytopathological Society of Japan. Common Names of Plant Diseases in Japan, Tokyo, Japan, 2000.

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First Report of Curvularia Leaf Blight on Curcuma wenyujin Caused by Curvularia clavata in China

Plant Disease ◽

10.1094/pdis-04-12-0392-pdn ◽

2013 ◽

Vol 97 (1) ◽

pp. 138-138 ◽

Cited By ~ 4

Author(s):

X. Y. Chen ◽

J. D. Feng ◽

Z. Su ◽

C. Sui ◽

X. Huang

Keyword(s):

Morphological Characteristics ◽

Its Region ◽

Leaf Blight ◽

Hainan Province ◽

Continuous Cropping ◽

Single Spore ◽

Chinese Medicinal Herb ◽

First Report ◽

Causal Pathogen ◽

Blight Disease

Curcuma wenyujin Y.H. Chen & C. Ling is a traditional Chinese medicinal herb in the Zingiberaceae family. Commonly known as Wen yujin, the root is widely used for alleviating pain and protecting the liver. A severe leaf blight disease was observed in three C. wenyujin farms in Hainan Province of China in October 2010. The obvious symptoms of leaf blight, yellow to brown irregular lesions (1 to 20 cm) on C. wenyujin, usually began at the tips of leaves and the main veins. This disease, especially severe from August to October, caused heavy damage and 100% of mature plants (10 months old) in farms were infected. The disease was most severe when continuous cropping was performed and showed slight improvement when rotation was adopted. Farmers usually sprayed carbendazim (50% WP) and thiophanate-methyl (70% WP) to control this disease, but these treatments were not effective. To isolate the causal pathogen, diseased plants were collected in October 2010 from a field of the Hainan Branch Institute of Medicinal Plant Development in Hainan Province. Lesion tissue was removed from the border between symptomatic and healthy tissue, surface sterilized in 75% ethanol for 1 min, washed in three changes of sterile distilled water, transferred to potato dextrose agar (PDA) plates, and incubated at 28°C for 7 days. Single spore cultures of five isolates were obtained and identified as Curvularia clavata based on morphological characteristics (1). Conidia measured 20 to 29 × 7.5 to 10.5 μm (n = 100), were curved, 3-septate, and the third cell from the base was larger and darker than the others. Mycelia of single spore cultures growing on PDA for 5 days were used for DNA extraction using a plant genomic DNA kit (TIANGEN, Beijing). The internal transcribed spacer (ITS) region of the rDNA was amplified using primers ITS1 and ITS4. The amplicons were 562 bp in length (GenBank Accession No. JQ730852) and had 99% nucleotide identity with the GenBank Accession No. JN021115 and AF071336 of C. clavata. Pathogenicity tests were conducted using fresh and healthy detached Curcuma wenyujin leaves. Mycelial discs (10 mm) removed from a 5-day-old colony on PDA were used for inoculation. Each isolate was inoculated on three distinct leaves (two distinct inoculations per leaf). Three additional leaves inoculated with sterile PDA discs were used as control. Inoculated leaves were covered with a polythene film to maintain high humidity. Leaves in trays were kept in a growth chamber at 28°C and observed for symptom appearance every day. Five days after inoculation, inoculated leaves developed blight symptoms similar to those observed on naturally infected leaves. No symptoms were observed on non-inoculated leaves. C. clavata was reisolated from the inoculated leaves, thus fulfilling Koch's postulates. C. clavata has been previously reported to be economically important on a number of other hosts (2). To our knowledge, this is the first report of Curvularia leaf blight on Curcuma wenyujin caused by C. clavata in China. References: (1) A. M Mandokhot et al. Eur. J. Plant Pathol.78:65, 1972. (2) T. Y. Zhang et al. Flora fungorum sinicorum: Beijing, China, 2010.

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First Report on Ovate-leaf Atractylodes Damping-off Caused by Fusarium oxysporum species Complex in South Korea

Plant Disease ◽

10.1094/pdis-05-21-1107-pdn ◽

2021 ◽

Author(s):

Oliul Hassan ◽

Taehyun Chang

Keyword(s):

South Korea ◽

Fusarium Oxysporum ◽

Species Complex ◽

Disease Incidence ◽

Fusarium Species ◽

Morphological Characteristics ◽

Conidial Suspension ◽

Sterile Water ◽

Damping Off ◽

First Report

In South Korea, ovate-leaf atractylodes (OLA) (Atractylodes ovata) is cultivated for herbal medicine. During May to June 2019, a disease with damping off symptoms on OLA seedlings were observed at three farmer fields in Mungyeong, South Korea. Disease incidence was estimated as approximately 20% based on calculating the proportion of symptomatic seedlings in three randomly selected fields. Six randomly selected seedlings (two from each field) showing damping off symptoms were collected. Small pieces (1 cm2) were cut from infected roots, surface-sterilized (1 minute in 0.5% sodium hypochlorite), rinsed twice with sterile water, air-dried and then plated on potato dextrose agar (PDA, Difco, and Becton Dickinson). Hyphal tips were excised and transferred to fresh PDA. Six morphologically similar isolates were obtained from six samples. Seven-day-old colonies, incubated at 25 °C in the dark on PDA, were whitish with light purple mycelia on the upper side and white with light purple at the center on the reverse side. Macroconidia were 3–5 septate, curved, both ends were pointed, and were 19.8–36.62 × 3.3–4.7 µm (n= 30). Microconidia were cylindrical or ellipsoid and 5.5–11.6 × 2.5–3.8 µm (n=30). Chlamydospores were globose and 9.6 –16.3 × 9.4 – 15.0 µm (n=30). The morphological characteristics of present isolates were comparable with that of Fusarium species (Maryani et al. 2019). Genomic DNA was extracted from 4 days old cultures of each isolate of SRRM 4.2, SRRH3, and SRRH5, EF-1α and rpb2 region were amplified using EF792 + EF829, and RPB2-5f2 + RPB2-7cr primer sets, respectively (Carbone and Kohn, 1999; O'Donnell et al. 2010) and sequenced (GenBank accession number: LC569791- LC569793 and LC600806- LC600808). BLAST query against Fusarium loci sampled and multilocus sequence typing database revealed that 99–100% identity to corresponding sequences of the F. oxysporum species complex (strain NRRL 28395 and 26379). Maximum likelihood phylogenetic analysis with MEGA v. 6.0 using the concatenated sequencing data for EF-1α and rpb2 showed that the isolates belonged to F. oxysporum species complex. Each three healthy seedlings with similar sized (big flower sabju) were grown for 20 days in a plastic pot containing autoclaved peat soil was used for pathogenicity tests. Conidial suspensions (106 conidia mL−1) of 20 days old colonies per isolate (two isolates) were prepared in sterile water. Three pots per strain were inoculated either by pouring 50 ml of the conidial suspension or by the same quantity of sterile distilled water as control. After inoculation, all pots were incubated at 25 °C with a 16-hour light/8-hour dark cycle in a growth chamber. This experiment repeated twice. Inoculated seedlings were watered twice a week. Approximately 60% of the inoculated seedlings per strain wilted after 15 days of inoculation and control seedlings remained asymptomatic. Fusarium oxysporum was successfully isolated from infected seedling and identified based on morphology and EF-1α sequences data to confirm Koch’s postulates. Fusarium oxysporum is responsible for damping-off of many plant species, including larch, tomato, melon, bean, banana, cotton, chickpea, and Arabidopsis thaliana (Fourie et al. 2011; Hassan et al.2019). To the best of our knowledge, this is the first report on damping-off of ovate-leaf atractylodes caused by F. oxysporum in South Korea. This finding provides a basis for studying the epidemic and management of the disease.

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