First Report of Pyrus pyrifolia ‘Cuiguan’ Fruit Rot Caused by Monilinia fructicola in Southern China

The early-ripening pear (i.e., Asian pear; Pyrus pyrifolia (Burm. f.) Nak.) cultivar ‘Cuiguan’ is an important fruit crop in southern China. In July 2020, an unknown fruit rot was observed on pear, which were harvested from an orchard in Xiajiang County of Jiangxi Province (27.62° N, 115.33° E), during storage in postharvest lab of Jiangxi Agricultural University. The incidence of this disease was 15% of 1000 post-harvest pear fruit (P. pyrifolia cv ‘Cuiguan’) after 10 d in storage room (20°C, 90% relative humidity). Initial symptoms were small brown nearly circular (diameter 10-20 mm) lesions with water-soaked edges on the fruit surface. The lesions expanded as concentric circles, and a gray-white mold developed in the center of the lesions. Ten fruits with typical symptoms were collected and surface-sterilized with 75% ethanol for 30 s. Small fragments (5×5 mm) at the junction of diseased and healthy tissues were disinfected with 1% sodium hypochlorite for 2 min, washed with sterile water 3 times, transferred to potato dextrose agar (PDA), and incubated at 28 ± 1℃ for 3 days. Five fungal colonies that looked similar were single-spored. The resultant fungal colonies produced irregular concentric rings with abundant gray-white aerial mycelia and gradually turned gray-brown. Conidia were unicellular, hyaline, lemon-shaped, 11.1-24.8 × 10.3-17.4 μm (n=50) in size and were produced in branched monilioid chains. These morphological characteristics were consistent with Monilinia fructicola (G. Winter) Honey (Hilber-Bodmer et al. 2010; Peter et al. 2015). To further confirm the species identification, genomic DNA of a representative isolate was obtained using an extraction kit (Biocolor, Shanghai, China), and primers ITS1/ITS4 (Munda et al. 2010), IMf F/IMf R (Ma et al. 2003), MO368-5 F/MO368-10 R (Côté et al. 2004), Mon-TubF1/Mon-TubR1, and Mon-G3pdhF/Mon-G3pdhR (Hu et al. 2011) were used to amplify the internal transcribed spacer (ITS), microsatellite, and polymorphic regions, as well as the glyceraldehyde-3-phosphate dehydrogenase (g3pdh) and beta-tubulin (tub2) genes, respectively. A BLAST search with obtained DNA sequences (GenBank Accession No. MW740236, MW788382, MW788383, MZ243141, and MZ243142, respectively) indicated 100% identity with M. fructicola (GenBank Accession No. LC312668.1 (513/513 bp), AY237426.1 (438/438 bp), FM994904.1 (490/500 bp), MN709392.1 (744/744 bp), and HQ908768.1 (1534/1534 bp), respectively). To confirm pathogenicity, 20 μl of a spore suspension (1.0 × 106 spores/ml) prepared from 7-day old PDA colonies of each of the five isolates was applied to the surface of 10 needle-wounded and 10 nonwounded, surface-disinfected ‘Cuiguan’ pear fruit. Ten wounded and nonwounded pears were inoculated with sterile water as controls. The experiment was repeated three times. All fruit were incubated at 25℃, 90% relative humidity. After 5 days, all wounded and nonwounded pears inoculated with M. fructicola showed symptoms similar to those observed in the storage room. Symptoms of nonwounded pears were milder than the wounded inoculated pears, while the control fruit remained healthy. A fungus with similar morphology to M. fructicola was re-isolated from the inoculated fruits, and thus, Koch’s postulates were fulfilled. To our knowledge, M. fructicola has been reported to cause brown rot of pear fruit in northern China (Zhu et al. 2016), but this is the first report of M. fructicola causing rot on P. pyrifolia in southern China. As an emerging rot disease in this region, and based on its economic importance in other pear growing regions, its presence is of concern the ‘Cuiguan’ pear fruit industry.

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First Report of Alternaria alternata causing fruit Rot on Tetradium ruticarpum in China

Plant Disease ◽

10.1094/pdis-08-20-1853-pdn ◽

2020 ◽

Author(s):

Miaolian Xiang ◽

Shucheng Li ◽

Fan Wu ◽

Xianyang Zhao ◽

Yinbao Wang ◽

...

Keyword(s):

Relative Humidity ◽

Alternaria Alternata ◽

Sequence Data ◽

Elongation Factor ◽

Fruit Rot ◽

Effective Control ◽

Molecular Characteristics ◽

Sterile Water ◽

First Report ◽

Initial Symptoms

Tetradium ruticarpum, previously and commonly known as Evodia rutaecarpa, is a tree that produces a fruit which is one of the most important traditional Chinese medicine herbs in China (Zhao et al. 2015). In July 2019, an investigation of diseases of T. ruticarpum was conducted in the farmland of Ruichang County (29.68° N, 115.65° E), Jiujiang City, China. An unknown fruit rot disease was observed and the incidence rate was estimated to be 60% to 70% within a 5,000 m2 area. The early symptoms appeared as small circular to irregular dark brown or black spots on the fruit, which gradually coalesced to a light brown-to-black discoloration and caused fruit rot. To identify the causal agent of the disease, 10 diseased fruits were collected and surface disinfected with 2% sodium hypochlorite for 2 min, 70% ethanol for 30 s, rinsed in sterile water and dried on filter paper. Tissues from non-symptomatic tissue as well as from the margin between healthy and affected edge were incubated on potato dextrose agar (PDA) at 25±1°C (12 h light/dark) with 90% relative humidity for 5 days. The colonies were brown to black with abundant whitish margins. Conidiophores were brown and measured 20.40 – 43.10×1.30 – 4.20 μm (25.47 × 2.35 µm on average, n=50). Conidia produced in single or branched chains, were obclavate or ovoid, approximately 9.90 – 32.80×6.50 – 14.50 μm (28.75×12.57 µm on average, n=50) with 2 to 5 transverse septa and 0 to 3 longitudinal septa. The colonies were consistent with Alternaria alternata (Simmons 2007). For molecular identification, the f partial internal transcribed spacer (ITS) regions, Glyceraldehyde-3-phosphate dehydrogenase (gapdh) genes, translation elongation factor 1-alpha (TEF) and Alternaria major allergen (Alt a1) gene of the isolate were amplified using primers ITS1/ITS4 (White et al. 1990), GDF/GDR (Templeton et al. 1992), EF1-728F/EF1-986R (Carbone and Kohn 1999) and Alt-for/Alt-rev (Hong et al. 2005). Sequence data showed 100% homology to A. alternata (GenBank accessions No.MN625176.1 (570/570 bp), MK683866.1 (618/618 bp), MK637432.1 (281/281 bp), KT315515.1 (488/488 bp)), respectively and the sequence data were deposited into GenBank with accession numbers MN897753 (ITS), MT041998 (gapdh), MT041999 (TEF), and MT042000 (Alt a1). Based on both morphological and molecular characteristics, the pathogen was identified as A. alternata. To confirm pathogenicity, 10 μl of a spore suspension (1.0 × 106 conidia/ml) obtained from 5-day-old PDA cultures of the strain were inoculated on 20 wounded (using sterile needle) and 20 nonwounded healthy T. ruticarpum fruits previously disinfected in 75% ethanol. Control fruits including 20 wounded fruits and 20 nonwounded fruits were inoculated with sterilized water. All fruits were incubated at 25±1°C (12 h light/dark) with 90% relative humidity. Four days later, all the wounded and non-wounded fruits showed the initial symptoms of black rot which was similar to that observed in the field, while the wounded and nonwounded fruits treated with sterile water remained healthy. The same pathogen was again isolated from the inoculated fruits. The pathogenicity experiment was repeated three times with the same results. As far as we know, this is the first report of A. alternata causing fruits rot on T. ruticarpum in China, and the identification of the pathogen will provide useful information for developing effective control strategies.

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First Report of Sclerotinia Stem Rot and Watery Soft Rot Caused by Sclerotinia sclerotiorum on Sand Rocket (Diplotaxis tenuifolia) in Italy

Plant Disease ◽

10.1094/pd-89-1241c ◽

2005 ◽

Vol 89 (11) ◽

pp. 1241-1241 ◽

Cited By ~ 4

Author(s):

A. Garibaldi ◽

A. Minuto ◽

M. L. Gullino

Keyword(s):

Relative Humidity ◽

Sclerotinia Sclerotiorum ◽

Severe Disease ◽

Soil Surface ◽

Soft Rot ◽

Sclerotinia Stem Rot ◽

First Report ◽

Initial Symptoms ◽

Stem Necrosis ◽

White Mycelium

Several species of Diplotaxis (D. tenuifolia, D. erucoides, and D. muralis), known as wild or sand rocket, are widely cultivated in Italy. Rocket is used in Mediterranean cuisine as salad, a component of packaged salad products, and as a garnish for food. In winter 2003, a severe disease was observed on D. tenuifolia grown in unheated glasshouses on commercial farms near Albenga in northern Italy. Initial symptoms included stem necrosis at the soil level and darkening of leaves. As stem necrosis progressed, infected plants wilted and died. Wilt, characterized by the presence of soft and watery tissues, occurred within a few days on young plants. The disease was extremely severe in the presence of high relative humidity and mild temperature (15°C). Necrotic tissues became covered with white mycelium that produced dark sclerotia. Diseased stem tissue was disinfested for 1 min in 1% NaOCl and plated on potato dextrose agar (PDA) amended with 100 ppm streptomycin sulfate. Sclerotinia sclerotiorum (1) was consistently recovered from infected stem pieces. Sclerotia observed on infected plants measured 1.23 to 3.00 × 1.40 to 5.38 mm (average 2.10 × 2.85 mm). Sclerotia produced on PDA measured 1.00 to 4.28 × 1.00 to 6.01 mm (average 2.38 × 3.23 mm). Pathogenicity of three isolates obtained from infected plants was confirmed by inoculating 30-day-old plants of D. tenuifolia grown in 18-cm-diameter pots in a glasshouse. Inoculum, 2 g per pot of wheat kernels infested with mycelium and sclerotia of each isolate, was placed on the soil surface around the base of each plant. Three replicates of five pots each were used per isolate. Noninoculated plants served as controls. The inoculation trial was repeated once. All plants were kept at temperatures ranging between 10 and 26°C (average 15°C) with an average relative humidity of 80% and were watered as needed. Inoculated plants developed symptoms of leaf yellowing within 12 days, soon followed by the appearance of white mycelium and sclerotia, and eventually wilted. Control plants remained symptomless. S. sclerotiorum was reisolated from inoculated plants. To our knowledge, this is the first report of infection of D. tenuifolia by S. sclerotiorum in Italy as well as worldwide. The disease currently has been observed in the Liguria Region but not yet in other areas where sand rocket is cultivated. The economic importance of this disease for the crop can be considered medium at the moment, but is expected to increase in the future. Reference: (1) N. F. Buchwald. Den. Kgl. Veterin.er-og Landbohojskoles Aarsskrift, 75, 1949.

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First Report of Brown Rot Caused by Monilinia fructicola Affecting Peach Orchards in Slovenia

Plant Disease ◽

10.1094/pdis-94-9-1166a ◽

2010 ◽

Vol 94 (9) ◽

pp. 1166-1166 ◽

Cited By ~ 15

Author(s):

A. Munda ◽

M. Viršček Marn

Keyword(s):

Prunus Persica ◽

Brown Rot ◽

Fruit Rot ◽

Morphological Identification ◽

Monilinia Fructicola ◽

Conidial Suspension ◽

Stone Fruits ◽

The European Union ◽

First Report ◽

Representative Isolate

Monilinia fructicola, the causal agent of brown rot, is a destructive fungal pathogen that affects mainly stone fruits (Prunoideae). It causes fruit rot, blossom wilt, twig blight, and canker formation and is common in North and South America, Australia, and New Zealand. M. fructicola is listed as a quarantine pathogen in the European Union and was absent from this region until 2001 when it was detected in France. In August 2009, mature peaches (Prunus persica cv. Royal Glory) with brown rot were found in a 5-year-old orchard in Goriška, western Slovenia. Symptoms included fruit lesions and mummified fruits. Lesions were brown, round, rapidly extending, and covered with abundant gray-to-buff conidial tufts. The pathogen was isolated in pure culture and identified based on morphological and molecular characters. Colonies on potato dextrose agar (PDA) incubated at 25°C in darkness had an average daily growth rate of 7.7 mm. They were initially colorless and later they were light gray with black stromatal plates and dense, hazel sporogenous mycelium. Colony margins were even. Sporulation was abundant and usually developed in distinct concentric zones. Limoniform conidia, produced in branched chains, measured 10.1 to 17.7 μm (mean = 12.1 μm) × 6.2 to 8.6 μm (mean = 7.3 μm) on PDA. Germinating conidia produced single germ tubes whose mean length ranged from 251 to 415 μm. Microconidia were abundant, globose, and 3 μm in diameter. Morphological characters resembled those described for M. fructicola (1). Morphological identification was confirmed by amplifying genomic DNA of isolates with M. fructicola species-specific primers (2–4). Sequence of the internal transcribed spacer (ITS) region (spanning ITS1 and ITS 2 plus 5.8 rDNA) of a representative isolate was generated using primers ITS1 and ITS4 and deposited in GenBank (Accession No. GU967379). BLAST analysis of the 516-bp PCR product revealed 100% identity with several sequences deposited for M. fructicola in NCBI GenBank. Pathogenicity was tested by inoculating five mature surface-sterilized peaches with 10 μl of a conidial suspension (104 conidia ml–1) obtained from one representative isolate. Sterile distilled water was used as a control. Peaches were wounded prior to inoculation. After 5 days of incubation at room temperature and 100% relative humidity, typical brown rot symptoms developed around the inoculation point, while controls showed no symptoms. M. fructicola was reisolated from lesion margins. Peach and nectarine orchards in a 5-km radius from the outbreak site were surveyed in September 2009 and M. fructicola was confirmed on mummified fruits from seven orchards. The pathogen was not detected in orchards from other regions of the country, where only the two endemic species M. laxa and M. fructigena were present. To our knowledge, this is the first report of M. fructicola associated with brown rot of stone fruits in Slovenia. References: (1) L. R. Batra. Page 106 in: World Species of Monilinia (Fungi): Their Ecology, Biosystematics and Control. J. Cramer, Berlin, 1991. (2) M.-J. Côté et al. Plant Dis. 88:1219, 2004. (3) K. J. D. Hughes et al. EPPO Bull. 30:507, 2000. (4) R. Ioos and P. Frey. Eur. J. Plant Pathol. 106:373, 2000.

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First Report of Nigrospora osmanthi Causing Leaf Spot on Tartary Buckwheat in China

Plant Disease ◽

10.1094/pdis-08-20-1773-pdn ◽

2020 ◽

Author(s):

Quan Shen ◽

Xixu Peng ◽

Feng He ◽

Shaoqing Li ◽

Zuyin Xiao ◽

...

Keyword(s):

Relative Humidity ◽

Leaf Spot ◽

Plant Pathogens ◽

Hunan Province ◽

Tartary Buckwheat ◽

Tween 20 ◽

Pathogenicity Test ◽

Conidial Suspension ◽

Sterile Water ◽

First Report

Buckwheat (Fagopyrum tataricum) is a traditional short-season pseudocereal crop originating in southwest China and is cultivated around the world. Antioxidative substances in buckwheat have been shown to provide many potential cardiovascular health benefits. Between August and November in 2019, a leaf spot was found in several Tartary buckwheat cv. Pinku1 fields in Xiangxiang County, Hunan Province, China. The disease occurred throughout the growth cycle of buckwheat after leaves emerged, and disease incidence was approximately 50 to 60%. Initially infected leaves developed a few round lesions, light yellow to light brown spots. Several days later, lesions began to enlarge with reddish brown borders, and eventually withered and fell off. Thirty lesions (2×2 mm) collected from three locations with ten leaves in each location were sterilized in 70% ethanol for 10 sec, in 2% sodium hypochlorite for 30 sec, rinsed in sterile water for three times, dried on sterilized filter paper, and placed on a potato dextrose PDA with lactic acid (3 ml/L), and incubated at 28°C in the dark for 3 to 5 days. Fungal colonies were initially white and later turned black with the onset ofsporulation. Conidia were single-celled, black, smooth, spherical to subspherical, and measured 9.2 to 15.6 µm long, and 7.1 to 11.6 µm wide (n=30). Each conidium was terminal and borne on a hyaline vesicle at the tip of conidiophores. Morphologically, the fungus was identified as Nigrospora osmanthi (Wang et al. 2017). Identification was confirmed by amplifying and sequencing the ITS region, and translation elongation factor 1-alpha (TEF1-α) and partial beta-tublin (TUB2) genes using primers ITS1/ITS4 (Mills et al. 1992), EF1-728F/EF-2 (Carbone and Kohn 1999; O’Donnell et al. 1998) and Bt-2a/Bt-2b (Glass et al. 1995), respectively. BLAST searches in GenBank indicated the ITS (MT860338), TUB2 (MT882054) and TEF1-α (MT882055) sequences had 99.80%, 99% and 100% similarity to sequences KX986010.1, KY019461.1 and KY019421.1 of Nigrospora osmanthi ex-type strain CGMCC 3.18126, respectively. A neighbor-joining phylogenetic tree constructed using MEGA7.0 with 1,000 bootstraps based on the concatenated nucleotide sequences of the three genes indicated that our isolate was closely related to N. osmanthi. Pathogenicity test was performed using leaves of healthy F. tataricum plants. The conidial suspension (1 × 106 conidia/ml) collected from PDA cultures with 0.05% Tween 20 buffer was used for inoculation by spraying leaves of potted 20-day-old Tartary buckwheat cv. Pinku1. Five leaves of each plant were inoculated with spore suspensions (1 ml per leaf). An equal number of control leaves were sprayed with sterile water to serve as a control. The treated plants were kept in a greenhouse at 28°C and 80% relative humidity for 24 h, and then transferred to natural conditions with temperature ranging from 22 to 30°C and relative humidity ranging from 50 to 60%. Five days later, all N. osmanthi-inoculated leaves developed leaf spot symptoms similar to those observed in the field, whereas control leaves remained healthy. N. osmanthi was re-isolated from twelve infected leaves with frequency of 100%, fulfilling Koch’s postulates. The genus Nigrospora has been regarded by many scholars as plant pathogens (Fukushima et al. 1998) and N. osmanthi is a known leaf blight pathogen for Stenotaphrum secundatum (Mei et al. 2019) and Ficus pandurata (Liu et al. 2019) but has not been reported on F. tataricum. Nigrospora sphaerica was also detected in vegetative buds of healthy Fagopyrum esculentum Moench (Jain et al. 2012). To our knowledge, this is the first report of N. osmanthi causing leaf spot on F. tataricum in China and worldwide. Appropriate strategies should be developed to manage this disease.

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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 Leaf Spot Caused by Colletotrichum fructicola on Myrica rubra in China

Plant Disease ◽

10.1094/pdis-10-21-2138-pdn ◽

2021 ◽

Author(s):

Shucheng Li ◽

Liuhua Xiao ◽

Fan Wu ◽

Yinbao Wang ◽

Mingshu Jia ◽

...

Keyword(s):

Leaf Spot ◽

Juglans Regia ◽

Morphological Characteristics ◽

Effective Control ◽

Bootstrap Support ◽

Sterile Water ◽

First Report ◽

Myrica Rubra ◽

Initial Symptoms ◽

Colletotrichum Fructicola

Myrica rubra is an important fruit tree with high nutritional and economic value, which is widely cultivated in multiple regions of China. In January 2021, an unknown disease which caused leaf spot with approximately 20% (n=100 investigated plants) of incidence was discovered on the leaves of M.rubra in Jiujiang City of Jiangxi Province (29.71° N, 115.97° E). The initial symptoms were small pale brown spots (1 to 2 mm diameter) on the leaves, which gradually expanded into round or irregular dark brown spots with the occurrence of the disease, and the lesion developed necrotic tissues in the center at later stages, eventually leading the leaves to chlorotic and wilted. Ten diseased leaves with typical symptoms were collected and the leaf tissue (5 × 5 mm) at junction of diseased and healthy portion were cut. The surfaces were disinfected with 75% ethanol for 45 s, 1% sodium hypochlorite for 1 min, and rinsed in sterile water for 3 times then transferred to potato dextrose agar (PDA) at 28 ± 1 ℃ for 3 days. Five fungal single isolates with similar morphology were purified from single spores. On PDA medium, the colonies initially appeared white with numerous aerial hyphae, and the center of the colony turned gray at later stages, less sporulation. While on modified czapek-dox medium (Peptone 3g, K2HPO4 1g, MgSO4·7H2O 0.5g, KCl 0.5g, FeSO4 0.01g, Maltose 30g, Agar 15g, Distilled water 1000 mL, pH=7.0), the mycelia of the colony were sparse and produced a large number of small bright orange particles (conidial masses). Conidia were single-celled, transparent, smooth-walled, 1-2 oil globule, cylindrical with slightly blunt rounded ends, 14.45-18.44 × 5.54-6.98 μm (av=16.27 μm × 6.19 μm, n=50) in size. These morphological characteristics of the pathogen were similar to the descriptions of Colletotrichum fructicola (Ruan et al, 2017; Yang et al, 2021). To further confirm the identity of the pathogen, genomic DNA from a representative isolate was extracted with DNA Extraction Kit (Yeasen, Shanghai, China), and the internal transcribed spacer (ITS), glyceraldehyde-3-phosphatedehydrogenase (GAPDH), calmodulin gene (CAL), actin (ACT) and chitin synthase 1 (CHS 1) were amplified by using the primers ITS1/ITS4 (Gardes et al, 1993), GDF/GDR (Templeton et al, 1992), CL1C/CL2C (Weir et al, 2012), ACT-512F/ACT-783R and CHS-79F/CHS-345R (Carbone et al, 1999), respectively. The PCR amplified sequences were submitted to GenBank (GenBank Accession No. ITS, MW740334; GAPDH, MW759805; CAL, MW759804; ACT, MW812384; CHS-1, MW759803) and aligned with GenBank showed 100% identity with C. fructicola (GenBank Accession No. ITS, MT355821.1 (546/546 bp); GAPDH, MT374664.1 (255/255 bp); CAL, MK681354.1 (741/741 bp); ACT, MT364655.1 (262/262 bp); CHS, MT374618.1 (271/271 bp)). Phylogenetic tree using the maximum likelihood methods with Kimura 2-parameter model and combined ITS-ACT-GAPDH-CHS-CAL concatenated sequences, bootstrap nodal support for 1000 replicates in MEGA7.0, revealed that the isolate was assigned to C. fructicola strain (ICMP 18581 and CBS 125397) (Yang et al. 2021) with 98% bootstrap support. Pathogenicities of were tested on fifteen healthy M. rubra plants (five for wounded inoculation, five for nonwounded inoculation, and five for controls) in the orchard. Twenty leaves were marked from each plant, and disinfected the surface with 75% ethanol. Ten μL spore suspension (1.0 × 106 conidia/ml) of each isolate from 7-day-old culture were inoculated on the surface of 20 needle-wounded and 20 nonwounded leaves, respectively. Healthy leaves were inoculated with sterile water as controls by the same method. All inoculated leaves were sprayed with sterile water and covered with plastic film to remained humidification. After 5 days, all the wounded leaves which were inoculated with C. fructicola showed similar symptoms to those observed on the original leaves. Symptoms of nonwounded leaves were milder than the wounded inoculated leaves, while control leaves remained healthy. Finally, the C. fructicola was re-isolated from the inoculated leaves. C. fructicola has been reported on Juglans regia, Peucedanum praeruptorum, Paris polyphylla var. Chinensis in China (Wang et al, 2017; Ma et al, 2020; Zhou et al, 2020). As far as we know, this is the first report of C. fructicola causing leaf spot on M.rubra in China. This result contributes to better understand the pathogens causing diseases of M.rubra in this region of China and develop effective control strategies.

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First Report and Epidemics of Buffelgrass Blight Caused by Pyricularia grisea in South Texas

Plant Disease ◽

10.1094/pdis.1999.83.4.398d ◽

1999 ◽

Vol 83 (4) ◽

pp. 398-398 ◽

Cited By ~ 4

Author(s):

O. Rodriguez ◽

J. Gonzalez-Dominguez ◽

J. P. Krausz ◽

G. N. Odvody ◽

J. P. Wilson ◽

...

Keyword(s):

Drought Stress ◽

Relative Humidity ◽

The United States ◽

South Texas ◽

Pyricularia Grisea ◽

Severe Drought ◽

Commonwealth Mycological Institute ◽

Unique Type ◽

First Report ◽

Initial Symptoms

A blight on buffelgrass, Cenchrus ciliaris L., has been observed for several years in south Texas and Mexico. The disease did not reach epidemic proportions until 1996. The causal agent, identified as Pyricularia grisea (Cooke) Sacc., is a common pathogen of grasses and other cultivated crops. Several Pennisetum spp. have been reported as hosts of Pyricularia spp.; this is the first report of buffelgrass as a host of this pathogen (1,2). Pathogenicity of P. grisea on buffelgrass was confirmed by greenhouse inoculations of 2-month-old buffelgrass plants with conidia washed with distilled water from monoconidial isolations of the pathogen, grown on potato dextrose agar, from infected leaves collected in several locations in south Texas and Mexico. Plants were placed for 8 h every night inside a plastic enclosure with a humidifier, simulating the high relative humidity conditions prevalent during the epidemic. Typical lesions developed after 7 days. The pathogen was re-isolated from the lesions after 10 days, fulfilling Koch's postulates. Conidia harvested from the sporulating samples were hyaline, transversely septate, with one to three septa, most of them having two. Conidia were obpyriform, with hylum often protuberant, measuring 20.6 to 26.3 μm in length and 8.5 to 10.1 μm wide. These measurements are consistent with those given for Pyricularia spp. by Ellis (1). Conidiophores were hyaline, single, slender, and unbranched. Initial symptoms were dark, discolored spots on the leaf that developed into tan, round to elliptical, necrotic lesions with a dark red border and a yellow, chlorotic halo. With increasing severity, lesions can coalesce, killing the entire leaf blade. Under heat and moisture stress, leaves with few lesions and yellow discoloration will wilt completely. Except for the presence of distinct lesions, wilted plants appear to be suffering from severe drought stress or herbicide injury. Losses vary from a few lesions to wilted whole plants and entire pastures. The pathogen also reduces the quantity and quality of seed by infecting involucres of the head. In the absence of the disease, even under severe moisture or drought stress, buffelgrass is able to thrive. Common T-4464 buffelgrass, which is highly susceptible to P. grisea, was introduced into south Texas in the late 1940s and is currently grown on 8 to 10 million acres in south Texas and Mexico. Buffelgrass reproduces by obligate apomixis, in which seeds are formed without sexual fertilization. Consequently, the progeny are genetically identical to the maternal parent. The monoculture of this grass with its unique type of reproduction encompasses millions of acres with genetically identical plants. Interaction of inoculum with weather conditions (nights with 8 to 10 h of more than 75% relative humidity) in 1996, 1997, and the late summer of 1998 produced epidemics of buffelgrass blight throughout south Texas and northern Mexico. P. grisea was also isolated from lesions on grassburr Cenchrus incertus M. A. Curtis collected throughout the area. References: (1) M. B. Ellis 1971. Dematiaceous hyphomycetes. Commonwealth Mycological Institute, Kew, Surrey, England. (2) D. F. Farr et al. 1989. Fungi of Plants and Plant Products in the United States. American Phytopathological Society, St. Paul, MN.

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First Report of Colletotrichum nymphaeae Causing Blossom Blight, Peduncle Rot, and Fruit Rot on Pyrus pyrifolia in Brazil

Plant Disease ◽

10.1094/pdis-12-18-2263-pdn ◽

2019 ◽

Vol 103 (8) ◽

pp. 2133-2133

Author(s):

R. R. Moreira ◽

D. P. Vandresen ◽

C. Glienke ◽

L. L. May-De-Mio

Keyword(s):

Fruit Rot ◽

Pyrus Pyrifolia ◽

First Report

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First Report of Alternaria tenuissimain Causing Leaf Spot of Celery (Apium graveolens) in China

Plant Disease ◽

10.1094/pdis-09-20-2083-pdn ◽

2020 ◽

Author(s):

Jianqiang Zhang ◽

Kangli Wu ◽

Xiaomeng Zhang ◽

Jiajia Li ◽

Abdramane salah zene ◽

...

Keyword(s):

Dna Sequences ◽

Leaf Spot ◽

Leaf Age ◽

Apium Graveolens ◽

Control Group ◽

Sterile Water ◽

Internal Transcribed Spacer Region ◽

Data Set ◽

First Report ◽

Initial Symptoms

Celery (Apium graveolens) is one of the most widely grown vegetables in the world. A survey in Anding District of Gansu Province in 2019 showed that the incidence of celery leaf spot was 25%-45%. The disease mainly occurs in late June and July. The leaf spot is conducive to the onset at high temperature and humidity environment. The initial symptoms were many small light brown, irregular-shaped on the leaves. The lesions gradually enlarged in the later stage of the disease, and multiple lesions coalesced to form large irregular brown spots, eventually the whole leaves died. A 3~4mm leaf tissue was cut from the junction of the diseased leaf and the healthy area, the leaf tisse was surface-sterilized in 1.5% NaClO for 1 min and washed with sterile water. Then, it was incubated on potato dextrose agar (PDA) and obtained the pure culture (Q1). After 5 days of cultivation at 25°C, the fungal colonies were olivaceous to dark olive with white margins and abundant aerial mycelia. The conidia were obclavate or ellipsoid, pale brown, with 3~4 longitudinal septa and 2~7 transverse septa, and measured 20.0 to 50.0 × 3.5 to 14.0μm (n=50). Conidiophores were septate, arising singly, and measured 3.5 to 40.0 × 2.5 to 4.5 μm (n=50). Based on morphological characteristics, the fungus was preliminarily identified as A.tenuissima (Simmons 2007). To further confirm the identification, the internal transcribed spacer region (ITS), translation elongation factor 1-α gene (TEF), RNA polymerase II second largest subunit (RPB2), major allergen Alt a 1 gene (Alt a 1), endopolygalacturonase gene (endoPG), anonymous gene region (OPA10-2) and glyceraldehyde 3-phos-phatedehydrogenase (GAPDH) were amplified and sequenced using primers ITS1/ITS4 (Peever et al. 2004), EF1-728F/EF1-986R (Carbone et al. 1999), RPB2-5F2/RPB2-5R (Sung et al. 2007), Alt-for/Alt-rev (Hong et al. 2005), EPG-specific/EPG-3b (Peever et al. 2004), OPA10-2R/OPA10-2L (Peever et al. 2004) and Gpd1/Gpd2 (Berbee et al. 1999) (GenBank accession no.MN046364, MW016001, MW016002, MW016003, MW016004, MW016005, MW016006). DNA sequences of TEF, RPB2, endoPG, OPA10-2 and GAPDH were 100% identical to those of A. tenuissima (MN256108, MK605866, KP789503, JQ859829 and MK683802), but ITS and Alt a 1 were 100% similarity with A. tenuissima (MN615420, JQ282277) and A. alternate (MT626589, KP123847). The ITS and Alt a 1 sequence did not distinguish A. tenuissima from the A. alternate complex. Maximum likelihood phylogenetic analyses were performed for the combined data set with TEF, RPB2, and endoPG using MEGA6 under the Tamura-Nei model (Kumar et al. 2016). The isolate Q1 clustered with type strain A. tenuissima CBS 918.96. The 20 celery plants of 4-7 leaf age were used test the pathogenicity of Q1, the ten plants were sprayed with 20ml of spore suspension (1×105 spores/ml), the control was sprayed with 20mL sterile water, which were placed in a growth chamber (25℃, a 14h light and 10h dark period, RH > 80%). Eight days after inoculation, 40% of the leaves formed lesions, which were consistent with the field observation，the control group was asymptomatic. The pathogen was reisolated from infected leaves to fulfill Koch’s postulates. To our knowledge, this is the first report of A. tenuissima causing leaf spot on celery in China.

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First Report of Dickeya fangzhongdai Causing Peduncle Soft Rot of Banana in China

Plant Disease ◽

10.1094/pdis-11-21-2513-pdn ◽

2021 ◽

Author(s):

Di Yang ◽

Chan Juan Du ◽

Yunfeng Ye ◽

Lian Fu Pan ◽

Jin Zhang ◽

...

Keyword(s):

16S Rdna ◽

Dna Sequences ◽

Southern China ◽

Intergenic Spacer ◽

Soft Rot ◽

Bacterial Suspension ◽

Sterile Water ◽

Internal Tissue ◽

Flower Bud ◽

First Report

Banana (Musa spp.) is a popular fruit all over the world, and it’s also an important cash crop with a planting area of 358,924 ha in southern China. In July 2020, a peduncle soft rot disease occurred on dwarf banana (Musa sp. cv. Guangfen) in Guigang city (N22°50'29″, E109° 43'34″), Guangxi province, China. More than 20% plants were infected in the banana plantation. The first external sign of the disease appeared on the incisional wound after the flower bud was cut off from the peduncle. The symptom initially appeared as a black lesion on the wound, then extended into the internal tissue of the whole peduncle. In the later stages, the internal tissue became soft and rot, occasionally formed a necrotic cavity, and eventually led to the black rot of the whole peduncle with a foul smell. To isolate the pathogen, the internal lesion tissues of 5 mm × 5 mm were collected between the border of symptomatic and healthy tissue, treated with 75% ethanol for 10 s, and 0.1% HgCl2 for 3 min, then rinsed with sterile water for three times. Sterilized tissue fragments were cut to pieces with sterilized surgical shears and soaked in 5 mL sterile water, then shaken for 10 min in a vortex oscillator. The suspension was diluted 1000 times with sterilized water，then plated on nutrient-agar medium and incubated at 28℃ in darkness for 24 h. Among the 32 isolates, 23 pure bacterial cultures with similar morphology were predominantly obtained from the samples. These bacteria were gram-negative, and their colonies were initially yellowish white with irregular edges and smooth surfaces, then turned to grayish blue after 72 h incubated at 28℃. The representative isolates GZF2-2 and GZF1-8 were selected for further identification. Genomic DNA was isolated from the bacteria and the 16S rDNA was amplified with primers 27F/1492R (Weisburg et al. 1991) and sequenced. The obtained sequences (GenBank Accession No. MZ768922 and OK668082) showed >99% identities to several records of Dickeya fangzhongdai deposited in NCBI GenBank (1400/1404 bps for GZF2-2 to KT992690, 1409/1417 bps for GZF1-8 to MT613398) based on BLAST analysis. In addition, the recA, fusA, gapA, purA, rplB, dnaX genes and the 16S-23S intergenic spacer (IGS) regions of the two isolates were also amplified and sequenced (GenBank Accession Nos. OK634381-OK634382, OK634369- OK634370, OK634373-OK634374, OK634377-OK634378, OK634385-OK634386, OK634365- OK634366 and OK631722-OK631723) as described by Tian et al. (2016). All the DNA sequences matched that of D. fangzhongdai strains JS5T (percent identities>99.06%), PA1 and ECM-1 in GenBank. Neighbor-joining phylogenetic analysis by software MegaX (Kumar et al. 2018) based on the 16S rDNA sequences revealed that the two isolates were in the same clade with reported D. fangzhongdai strains. Multilocus sequence analysis of the other seven regions also showed the two representative isolates were belong to D. fangzhongdai. Therefore, the isolates were identified as D. fangzhongdai. Pathogenicity of isolate GZF2-2 was investigated to demonstrate Koch’s postulate. The end of the banana peduncles of 6 healthy plants were cut off, and 10 mL bacterial suspension (108 CFU/mL) was inoculated to the fresh wound on the plants using sterile brushes. Six control plants were inoculated with sterilized water. All the inoculated peduncles were covered with plastic bags to maintain high humidity. After 28 days, all the peduncles inoculated with strain GZF2-2 showed soft rot symptoms similar to those observed in the field, while the controls remained symptomless. The same bacteria were re-isolated from the symptomatic peduncles and confirmed by sequencing the 16S rDNA. D. fangzhongdai has been reported to cause soft rot on onion (Ma et al. 2020) and bleeding cankers on pear trees (Chen et al. 2020). To the best of our knowledge, this is the first report of D. fangzhongdai causing peduncle soft rot on banana in China.

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