First Report of Pepper Fruit Rot Caused by Fusarium concentricum in China

Pepper (Capsicum annuum L.) is an important vegetable crop worldwide. Some Fusarium species can cause pepper fruit rot, leading to significant yield losses of pepper production and, for some Fusarium species, potential risk of mycotoxin contamination. A total of 106 diseased pepper fruit samples were collected from various pepper cultivars from seven provinces (Gansu, Hainan, Heilongjiang, Hunan, Shandong, Shanghai, and Zhejiang) in China during the 2012 growing season, where pepper production occurs on approximately 25,000 ha. Pepper fruit rot symptom incidence ranged from 5 to 20% in individual fields. Symptomatic fruit tissue was surface-sterilized in 0.1% HgCl2 for 1 min, dipped in 70% ethanol for 30 s, then rinsed in sterilized distilled water three times, dried, and plated in 90 mm diameter petri dishes containing potato dextrose agar (PDA). After incubation for 5 days at 28°C in the dark, putative Fusarium colonies were purified by single-sporing. Forty-three Fusarium strains were isolated and identified to species as described previously (1,2). Morphological characteristics of one strain were identical to those of F. concentricum. Aerial mycelium was reddish-white with an average growth rate of 4.2 to 4.3 mm/day at 25°C in the dark on PDA. Pigments in the agar were formed in alternating red and orange concentric rings. Microconidia were 0- to 1-septate, mostly 0-septate, and oval, obovoid to allantoid. Macroconidia were relatively slender with no significant curvature, 3- to 5-septate, with a beaked apical cell and a foot-shaped basal cell. To confirm the species identity, the partial TEF gene sequence (646 bp) was amplified and sequenced (GenBank Accession No. KC816735). A BLASTn search with TEF gene sequences in NCBI and the Fusarium ID databases revealed 99.7 and 100% sequence identity, respectively, to known TEF sequences of F. concentricum. Thus, both morphological and molecular criteria supported identification of the strain as F. concentricum. This strain was deposited as Accession MUCL 54697 (http://bccm.belspo.be/about/mucl.php). Pathogenicity of the strain was confirmed by inoculating 10 wounded, mature pepper fruits that had been harvested 70 days after planting the cultivar Zhongjiao-5 with a conidial suspension (1 × 106 spores/ml), as described previously (3). A control treatment consisted of inoculating 10 pepper fruits of the same cultivar with sterilized distilled water. The fruit were incubated at 25°C in a moist chamber, and the experiment was repeated independently in triplicate. Initially, green to dark brown lesions were observed on the outer surface of inoculated fruit. Typical soft-rot symptoms and lesions were observed on the inner wall when the fruit were cut open 10 days post-inoculation. Some infected seeds in the fruits were grayish-black and covered by mycelium, similar to the original fruit symptoms observed at the sampling sites. The control fruit remained healthy after 10 days of incubation. The same fungus was isolated from the inoculated infected fruit using the method described above, but no fungal growth was observed from the control fruit. To our knowledge, this is the first report of F. concentricum causing a pepper fruit rot. References: (1) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Ames, IA, 2006. (2) K. O'Donnell et al. Proc. Nat. Acad. Sci. USA 95:2044, 1998. (3) Y. Yang et al. 2011. Int. J. Food Microbiol. 151:150, 2011.

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First Report of Diaporthe hongkongensis Causing Fruit rot on Peach (Prunus persica) in China

Plant Disease ◽

10.1094/pdis-07-20-1505-pdn ◽

2021 ◽

Author(s):

Zhou Zhang ◽

Zheng Bing Zhang ◽

Yuan Tai Huang ◽

FeiXiang Wang ◽

Wei Hua Hu ◽

...

Keyword(s):

Prunus Persica ◽

Fruit Rot ◽

Hunan Province ◽

Post Inoculation ◽

Distilled Water ◽

Conidial Suspension ◽

Internal Transcribed Spacer Region ◽

First Report ◽

Representative Isolate ◽

Rot Disease

Peach [Prunus persica (L.) Batsch] is an important deciduous fruit tree in the family Rosaceae and is a widely grown fruit in China (Verde et al., 2013). In July and August 2018, a fruit rot disease was observed in a few peach orchards in Zhuzhou city, the Hunan Province of China. Approximately 30% of the fruit in more than 400 trees was affected. Symptoms displayed were brown necrotic spots that expanded, coalesced, and lead to fruit being rotten. Symptomatic tissues excised from the margins of lesions were surface sterilized in 70% ethanol for 10 s, 0.1% HgCl2 for 2 min, rinsed with sterile distilled water three times, and incubated on potato dextrose agar (PDA) at 26°C in the dark. Fungal colonies with similar morphology developed, and eight fungal colonies were isolated for further identification. Colonies grown on PDA were grayish-white with white aerial mycelium. After an incubation period of approximately 3 weeks, pycnidia developed and produced α-conidia and β-conidia. The α-conidia were one-celled, hyaline, fusiform, and ranged in size from 6.0 to 8.4 × 2.1 to 3.1 μm, whereas the β-conidia were filiform, hamate, and 15.0 to 27.0 × 0.8 to 1.6 μm. For molecular identification, total genomic DNA was extracted from the mycelium of a representative isolate HT-1 and the internal transcribed spacer region (ITS), β-tubulin gene (TUB), translation elongation factor 1-α gene (TEF1), calmodulin (CAL), and histone H3 gene (HIS) were amplified and sequenced (Meng et al. 2018). The ITS, TUB, TEF1, CAL and HIS sequences (GenBank accession nos. MT740484, MT749776, MT749778, MT749777, and MT749779, respectively) were obtained and in analysis by BLAST against sequences in NCBI GenBank, showed 99.37 to 100% identity with D. hongkongensis or D. lithocarpus (the synonym of D. hongkongensis) (Gao et al., 2016) (GenBank accession nos. MG832540.1 for ITS, LT601561.1 for TUB, KJ490551.1 for HIS, KY433566.1 for TEF1, and MK442962.1 for CAL). Pathogenicity tests were performed on peach fruits by inoculation of mycelial plugs and conidial suspensions. In one set, 0.5 mm diameter mycelial discs, which were obtained from an actively growing representative isolate of the fungus on PDA, were placed individually on the surface of each fruit. Sterile agar plugs were used as controls. In another set, each of the fruits was inoculated by application of 1 ml conidial suspension (105 conidia/ml) by a spray bottle. Control assays were carried out with sterile distilled water. All treatments were maintained in humid chambers at 26°C with a 12-h photoperiod. The inoculation tests were conducted twice, with each one having three fruits as replications. Six days post-inoculation, symptoms of fruit rot were observed on inoculated fruits, whereas no symptoms developed on fruits treated with agar plugs and sterile water. The fungus was re-isolated and identified to be D. hongkongensis by morphological and molecular methods, thus fulfilling Koch’s Postulates. This fungus has been reported to cause fruit rot on kiwifruit (Li et al. 2016) and is also known to cause peach tree dieback in China (Dissanayake et al. 2017). However, to our knowledge, this is the first report of D. hongkongensis causing peach fruit rot disease in China. The identification of the pathogen will provide important information for growers to manage this disease.

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First report of chili pepper fruit rot caused by Fusarium incarnatum in China

Plant Disease ◽

10.1094/pdis-09-19-1996-pdn ◽

2021 ◽

Author(s):

Xiao Qin Zhu ◽

Dongmei Liu ◽

Quanchun Hong ◽

Yifang Lu ◽

Dongli Pei

Keyword(s):

Chili Pepper ◽

Fruit Rot ◽

Effective Control ◽

Economic Losses ◽

Hainan Province ◽

Pathogenicity Test ◽

Distilled Water ◽

Conidial Suspension ◽

First Report ◽

Pepper Fruit

Pepper (Capsicum annuum L.), with annual production over 1 million tons, is ranked the first vegetable crop in Hainan Province, China. In December 2018, fruit rot of chili pepper , with yield loss of up to 15%, was found in 10 fields (12 hm2) in Yacheng (18°N, 109°E), Hainan Province, China. Water-soaked and soft lesions developed on fruit, with white to light gray fungal mycelium present inside. The diseased fruit turned soft and decayed at the later stages. Diseased tissue was cut into 12 pieces of 0.5×0.5 cm, surface-disinfected with 2% sodium hypochlorite for 2 min, followed by 70% ethanol for 30 s, rinsed with sterile distilled water five times, and plated onto potato dextrose agar (PDA). After growing on PDA for 2 to 3 days at 28°C in an incubator without light, 10 pure culture isolates were obtained. All isolates had abundant dense white aerial mycelia that became beige with age. The macroconidia were slightly curved with four to seven septa, 29.51 to 42.15 × 4.29 to 6.22 μm. Spindle-shaped mesoconidia with three to four septa were abundantly produced, 20.34 to 24.54 × 4.58 to to 11.70 × 2.35 to 3.20 μm. Chlamydospores were absent. Based on the morphological characteristics, the fungus was tentatively identified as Fusarium incarnatum (Leslie and Summerell 2006). An isolate SQHP-01 was chosen for molecular identification and pathogenicity test. Two DNA fragments of the isolate, the internal transcribed spacer (ITS) and translation elongation factor genes (EF-1α) were amplified for sequencing. BLAST analysis showed that sequences of ITS (GenBank acc. no. MN317371) and EF-1α (acc. No. MN928788) had 99.61 to 100% identity with those of known F. incarnatum (MN480497 and KF993969). Phylogenetic analysis was conducted using neighbor-joining algorithm based on ITS and EF-1a genes separately, and the isolate was well clustered with F. incarnatum both with 100% bootstrap support. Pathogenicity test of the isolate were carried out twice on five healthy chili pepper fruit. After surface-disinfection, fruit were wounded at three different points and 20 μl of conidial suspension (106 conidia/ml) were deposited on each wound. Unwounded inoculation was conducted by spreading 100 μl of the suspension on each fruit surface including the pedicel and calyx. The fruit spread with sterile distilled water represented the negative control. All fruit treatments were placed on the moist sterile cotton in moist chambers at 25°C with 16 h light and 8 h darkness. After 4 to 6 days, water-soaked necrotic lesions appeared on the wounded fruit, the symptoms identical to those observed in the field. Water-soaked necrotic lesions developed on the pedicel and calyx of unwounded fruit. No symptoms were observed on the control fruit. The morphology and sequences of re-isolated fungal isolates from the tested peppers were the same as the original isolate. To our knowledge, this is the first report of F. incarnatum (synonym of F. semitectum) causing fruit rot on chili pepper in China. F. incarnatum has been reported to cause root rot of greenhouse pepper in China (Li et al. 2018), fruit rot of bell pepper in Trinidad (Ramdial et al. 2016) and Pakistan (Tariq et al. 2018). Effective control strategies need to be developed to prevent the economic losses caused by the disease in chili pepper.

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First Report of Fusarium Maize Ear Rot Caused by Fusarium kyushuense in China

Plant Disease ◽

10.1094/pdis-05-13-0558-pdn ◽

2014 ◽

Vol 98 (2) ◽

pp. 279-279 ◽

Cited By ~ 2

Author(s):

J.-H. Wang ◽

H.-P. Li ◽

J.-B. Zhang ◽

B.-T. Wang ◽

Y.-C. Liao

Keyword(s):

Fusarium Species ◽

Morphological Characteristics ◽

Distilled Water ◽

Ear Rot ◽

Tubulin Gene ◽

Average Growth ◽

First Report ◽

Maize Ear Rot ◽

Maize Ear ◽

Β Tubulin

From September 2009 to October 2012, surveys to determine population structure of Fusarium species on maize were conducted in 22 provinces in China, where the disease incidence ranged from 5 to 20% in individual fields. Maize ears with clear symptoms of Fusarium ear rot (with a white to pink- or salmon-colored mold at the ear tip) were collected from fields. Symptomatic kernels were surface-sterilized (1 min in 0.1% HgCl2, and 30 s in 70% ethanol, followed by three rinses with sterile distilled water), dried, and placed on PDA. After incubation for 3 to 5 days at 28°C in the dark, fungal colonies displaying morphological characteristics of Fusarium spp. (2) were purified by transferring single spores and identified to species level by morphological characteristics (2), and DNA sequence analysis of translation elongation factor-1α (TEF) and β-tubulin genes. A large number of Fusarium species (mainly F. graminearum species complex, F. verticillioides, and F. proliferatum) were identified. These Fusarium species are the main causal agents of maize ear rot (2). Morphological characteristics of six strains from Anhui, Hubei, and Yunnan provinces were found to be identical to those of F. kyushuense (1), which was mixed with other Fusarium species in the natural infection in the field. Colonies grew fast on PDA with reddish-white and floccose mycelia. The average growth rate was 7 to 9 mm per day at 25°C in the dark. Reverse pigmentation was deep red. Microconidia were obovate, ellipsoidal to clavate, and 5.4 to 13.6 (average 8.8) μm in length. Macroconidia were straight or slightly curved, 3- to 5-septate, with a curved and acute apical cell, and 26.0 to 50.3 (average 38.7) μm in length. No chlamydospores were observed. Identity of the fungus was further investigated by sequence comparison of the partial TEF gene (primers EF1/2) and β-tubulin gene (primers T1/22) of one isolate (3). BLASTn analysis of the TEF amplicon (KC964133) and β-tubulin gene (KC964152) obtained with cognate sequences available in GenBank database revealed 99.3 and 99.8% sequence identity, respectively, to F. kyushuense. Pathogenicity tests were conducted twice by injecting 2 ml of a prepared spore suspension (5 × 105 spores/ml) into maize ears (10 per isolate of cv. Zhengdan958) through silk channel 4 days post-silk emergence under field conditions in Wuhan, China. Control plants were inoculated with sterile distilled water. The ears were harvested and evaluated 30 days post-inoculation. Reddish-white mold was observed on inoculated ears and the infected kernels were brown. No symptoms were observed on water controls. Koch's postulates were fulfilled by re-isolating the pathogen from infected kernels. F. kyushuense, first described on wheat in Japan (1), has also been isolated from rice seeds in China (4). It was reported to produce both Type A and Type B trichothecene mycotoxins (1), which cause toxicosis in animals. To our knowledge, this is the first report of F. kyushuense causing maize ear rot in China and this disease could represent a serious risk of yield losses and mycotoxin contamination in maize and other crops. The disease must be considered in existing disease management practices. References: (1) T. Aoki and K. O'Donnell. Mycoscience 39:1, 1998. (2) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Ames, IA, 2006. (3) F. Van Hove et al. Mycologia 103:570, 2011. (4) Z. H. Zhao and G. Z. Lu. Mycotaxon 102:119, 2007.

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First report of Fusarium incarnatum causing spikelet rot on rice in China

Plant Disease ◽

10.1094/pdis-12-20-2660-pdn ◽

2021 ◽

Author(s):

Ling Wang ◽

S. L. Ge ◽

Kehan Zhao ◽

huang Shiwen

Keyword(s):

Natural Science ◽

Disease Incidence ◽

Science And Technology ◽

Agricultural Science ◽

Zhejiang Province ◽

Maturity Stage ◽

Post Inoculation ◽

Distilled Water ◽

Conidial Suspension ◽

First Report

Rice (Oryza sativa L.) is the most important and widely grown crop, covering about 29.9 million ha of total cultivation area in China. In the last decade, spikelet rot disease on rice became much more frequent in the middle and lower reaches of the Yangtze River, China. Fusarium proliferatum (Matsush.) Nirenberg ex Gerlach & Nirenberg was reported to be a causal agent of spikelet rot on rice in Hangzhou, Zhejiang province (Huang et al. 2012). In September 2019, a survey was conducted to understand the etiology of the disease in the main rice growing regions of Jinshan District of Shanghai. Symptomatic panicles exhibiting reddish or brown discoloration on the glumes were collected from different rice fields, where disease incidence was estimated to be between 20 to 80%. Diseased glumes were cut into small sections (5 × 5 mm) from the boundary of necrotic and healthy tissues, surface-sterilized with 75% ethanol for 30 s and 3% sodium hypochlorite for 90 s, rinsed twice with sterile distilled water, then placed onto 1/5 strength potato dextrose agar (PDA). After 3 to 5 days of incubation at 28°C in the dark, fungal growth with Fusarium-like colonies were transferred to PDA and purified by the single-spore isolation method. A total of 12 isolates were obtained and colonies showed loosely floccose, white mycelium and pale-yellow pigmentation on PDA. Microconidia were ovoid mostly with 0 to 1 septum, and measured 4.2 to 16.6 × 2.5 to 4.1 μm (n = 50). After 5-7 days of inoculation on carnation leaf agar (CLA), macroconidia produced usually had 3 to 5 septa, slightly curved at the apex, ranging from 15.7 to 39.1 × 3.3 to 5.0 μm (n = 50). Chlamydospores were produced in hyphae, most often solitary in short chains or in clumps, ellipsoidal or subglobose with thick and roughened walls. Molecular identification was performed on the representative isolates (JS3, JS9, and JS21). The rDNA internal transcribed spacer (ITS), translation elongation factor (TEF-1α) and β-tubulin (β-TUB) genes were amplified and sequenced using the paired primers ITS1/ITS4 (White et al. 1990), EF1/EF2 (O’Donnell et al. 1998) and T1/T22 (O’Donnell and Cigelnik 1997), respectively. The obtained sequences were deposited in GenBank under accession numbers MT889972 to MT889974 (ITS), MT895844 to MT895846 (TEF-1α), and MT895841 to MT895843 (β-TUB), respectively. BLASTn search of the sequences revealed 99 to 100% identity with ITS (MF356578), TEF-1α (HM770725) and β-TUB (GQ915444) of Fusarium incarnatum isolates. FUSARIUM-ID (Geiser et al. 2004) analysis showed 99 to 100% similarity with sequences of the F. incarnatum-equiseti species complex (FIESC) (FD_01651 and FD_01628). In addition, a phylogenetic analysis based on the concatenated nucleotide sequences placed the isolates in the F. incarnatum clade at 100% bootstrap support. Thus, both morphological observations and molecular criteria supported identification of the isolates as F. incarnatum (Desm.) Sacc (synonym: Fusarium semitectum) (Leslie and Summerell 2006, Nirenberg 1990). Pathogenicity tests were performed on susceptible rice cultivar ‘Xiushui134’. At pollen cell maturity stage, a 2-ml conidial suspension (5 × 105 macroconidia/ml) of each isolate was injected into 10 rice panicles. Control plants were inoculated with sterile distilled water. Then, the pots were kept in a growth chamber at 28°C, 80% relative humidity, and 12 h/12 h light (10,000 lux)/dark. The experiment was repeated two times for each isolate. Two weeks post-inoculation, all inoculated panicles showed similar symptoms with the original samples, whereas no symptoms were observed on the control. The pathogen was re-isolated from inoculated panicles and identified by the method described above to fulfill Koch's postulates. Previous studies reported that F. incarnatum reproduced perithecia to overwinter on rice stubble as the inoculum of Fusarium head blight of wheat in southern China (Yang et al. 2018). To our knowledge, this is the first report of spikelet rot on rice caused by F. incarnatum in China. Further investigation is needed to gain a better understanding its potential geographic distribution of this new pathogen on rice crop. References: (1) Huang, S. W., et al. 2011. Crop Prot. 30: 10. (2) White, T. J., et al. 1990. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA. (3) O’Donnell, K., et al. 1998. Proc. Natl. Acad. Sci. U.S.A. 95: 2044. (4) O'Donnell, K., Cigelnik, E. 1997. Mol. Phylogenet. Evol. 7: 103. (5) Geiser, D. M., et al. 2004. Eur. J. Plant Pathol. 110: 473. (6) Leslie, J. F., and Summerell, B. A. 2006. The Fusarium Laboratory Manual. Blackwell, Ames, IA. (7) Nirenberg, H. I. 1990. Stud. Mycol. 32: 91. (8) Yang, M. X., et al. 2018. Toxins. 10: 115. The author(s) declare no conflict of interest. Funding: Funding was provided by National Natural Science Foundation of China (grant no. 31800133), Zhejiang Provincial Natural Science Foundation of China (grant no. LQ18C140005), Key Research and Development Program of Zhejiang Province (grant no. 2019C02018), Shanghai Science and Technology for Agriculture Promotion Project (2019-02-08-00-08-F01127), and the Agricultural Science and Technology Innovation Program of China Academy of Agricultural Science (CAAS-ASTIP-2013- CNRRI).

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First Report of Walnut Canker Caused by Fusarium incarnatum from India

Plant Disease ◽

10.1094/pdis-04-11-0352 ◽

2011 ◽

Vol 95 (12) ◽

pp. 1587-1587

Author(s):

B. Singh ◽

C. S. Kalha ◽

V. K. Razdan ◽

V. S. Verma

Keyword(s):

Fusarium Species ◽

Juglans Regia ◽

Infected Plant ◽

Distilled Water ◽

Conidial Suspension ◽

State University ◽

Single Spore ◽

Jammu And Kashmir ◽

First Report ◽

Branch Dieback

While screening newly introduced cultivars of walnut (Juglans regia) at Bhaderwah (Mini Kashmir), Jammu and Kashmir, India in September 2008, 60% of grafted plants were found to be dying because of a cankerous growth observed on seedling stems. Later, these symptoms extended to lateral branches. In the surveyed nurseries, cvs. SKU 0002 and Opex Dachaubaria were severely affected by the disease. Cankers were also observed in all walnut nurseries in the area with several wild seedlings also being observed to be exhibiting similar cankerous symptoms on stem and branches. Necrotic lesions from cankerous tissues on seedling stems were surface disinfested with 0.4% NaOCl for 1 min and these disinfected cankerous tissues were grown on potato dextrose agar (potato-250 g, dextrose-15 g, agar-15 g, distilled water-1 liter). A Fusarium sp. was isolated consistently from these cankerous tissues, which was purified using single-spore culture. Carnation leaf agar was used for further culture identification (2,3). The fungal colony was floccose, powdery white to rosy in appearance when kept for 7 days at 25 ± 2°C. Macroconidia were straight to slightly curved, four to eight septate and 30 to 35 × 3.5 to 5.7 μm. These are characteristics consistent with Fusarium incarnatum (3). Pathogenicity was confirmed by spraying a conidial suspension (1 × 106 conidia/ml) onto bruised branches of 1-year-old walnut plants (cv. Opex Dachaubaria) while sterile distilled water sprays were used for the controls. Inoculated plants were incubated at 20 ± 2°C and 85% relative humidity for 48 h. Fifty days following inoculation, branch dieback followed by canker symptoms developed on inoculated plants. Control plants remained healthy with no symptoms of canker. F. incarnatum (Roberge) Sacc. was repeatedly isolated from inoculated walnut plants, thus satisfying Koch's postulates. Infected plant material has been deposited at Herbarium Crytogamae Indiae Orientalis (ITCC-6874-07), New Delhi. To our knowledge, this is the first report of walnut canker caused by F. incarnatum (Roberge) Sacc. from India. This fungus was previously reported to be affecting walnut in Italy (1) and Argentina (4). References: (1) A. Belisario et al. Informatore Agrario 21:51, 1999. (2) J. C. Gilman. A Manual of Soil Fungi. The Iowa State University Press, Ames, 1959. (3) P. E. Nelson et al. Fusarium Species. An Illustrated Manual for Identification. The Pennsylvania State University Press, University Park, 1983. (4) S. Seta et al. Plant Pathol. 53:248, 2004.

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First Report of Fusarium proliferatum Causing Fruit Rot of Grapes (Vitis vinifera) in Pakistan

International Journal of Phytopathology ◽

10.33687/phytopath.007.02.2589 ◽

2018 ◽

Vol 7 (2) ◽

pp. 85-88 ◽

Cited By ~ 1

Author(s):

Salman Ghuffar ◽

Gulshan Irshad ◽

Fengyan Zhai ◽

Asif Aziz ◽

Hafiz M. Asadullah M. Asadullah ◽

...

Keyword(s):

Vitis Vinifera ◽

Negative Control ◽

Fusarium Proliferatum ◽

Fruit Rot ◽

Pathogenicity Test ◽

Distilled Water ◽

Conidial Suspension ◽

Fruit Crop ◽

First Report ◽

Thin Walled

Grapes (Vitis vinifera) are the important fruit crop in Pakistan, mostly cultivated for edible purpose. In September 2016, unusual fruit rot symptoms were observed 3-5 days after harvesting on grapes cv. Kishmishi in post-harvest packing houses in Jehlum district (32°56'22.3"N 73°43'31.4"E) of Punjab province. To determine the disease incidence, a total of 10 boxes of grapes from 5 different locations were selected randomly. Each box contained average 12 bunches and 30 bunches out of 120 inspected bunches displayed typical symptoms of the disease. The initial Symptoms were small, round, water-soaked lesions that rapidly developed into soft, white to light pink mycelium near the centre of infected fruits (Figure 1). A total of 186 symptomatic berries were surface sterilized with 1% sodium hypochlorite, rinsed three times with sterile distilled water and dried by placing on filter paper for 45 sec. Sterilized tissues (approximately 4 mm3) were excised and incubated on potato dextrose agar (PDA) medium at 25 ± 4°C. One week after incubation, colonies with abundant aerial mycelium were initially white, cottony and turned to violet and dark purple with age (Figure 2). A total of 25 isolates were examined morphologically. Macroconidia were slender, thin-walled, 3 to 5 septate, curved apical cell, with 20.9 to 45.2 × 3.2 to 7.1 μm and Microconidia were thin-walled, aseptate, club-shaped with 4.5 to 11.2 × 2.3 to 4.1 μm (Figure 3). These characteristics best fit for the description of Fusarium proliferatum (Leslie and Summerell, 2006). Portions of the internal transcribed spacer (ITS) region were sequenced (White et al., 1990). Sequences of two isolates Fus 07 and Fus 09 (GenBank Accessions; MH444366 and MH464139) showed 100% identity to the corresponding gene sequences of Fusarium proliferatum (GenBank Accessions; MH368119, MF033172 and KU939071) (Figure 4). Pathogenicity test was performed by inoculation with 50-μl conidial suspension (1 × 106conidia/ml) of two isolates onto three non-wounded and four wounded asymptomatic grapes berries. Sterile distilled water was used for a negative control (Figure 5). The experiment was conducted twice and berries were incubated at 25 ± 2°C in sterile moisture chambers (Ghuffar et al., 2018). White to light pink mycelium in appearance with the original symptoms were observed on both wounded and non-wounded inoculated berries after 3 days, whereas no symptoms were observed on the negative control. The morphology of the fungus that was re-isolated from each of the inoculated berries was identical to that of the original cultures. Fusarium proliferatum, one of the destructive species, causes diseases like foot-rot of corn (Farr et al., 1990), root rot of soybean (Díaz Arias et al., 2011), bakanae of rice (Zainudin et al., 2008), wilt of date palm (Khudhair et al., 2014), tomato wilt (Chehri, 2016) and tomato fruit rot (Murad et al., 2016). To our knowledge, this is the first report of Fusarium proliferatum causing fruit rot of grapes in Pakistan, where the disease poses a significant threat to the sustainability of this major fruit crop.

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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 incarnatum-equiseti Species Complex Causing Leaf Spot on Rockmelon (Cucumis melo L.) in Malaysia

Plant Disease ◽

10.1094/pdis-06-20-1380-pdn ◽

2020 ◽

Author(s):

Siti Izera Ismail ◽

Nur Ainina Noor Asha ◽

Dzarifah Zulperi

Keyword(s):

Leaf Spot ◽

Cucumis Melo ◽

Species Complex ◽

Fruit Rot ◽

Peat Moss ◽

Distilled Water ◽

Conidial Suspension ◽

Isolation Technique ◽

First Report ◽

Cucumis Melo L

Rockmelon, (Cucumis melo L.) is an economically important crop cultivated in Malaysia. In October 2019, severe leaf spot symptoms with a disease incidence of 40% were observed on the leaves of rockmelon cv. Golden Champion at Faculty of Agriculture, Universiti Putra Malaysia (UPM). Symptoms appeared as brown necrotic spots, 10 to 30 mm in diameter, with spots surrounded by chlorotic halos. Pieces (5 x 5 mm) of diseased tissue were sterilized with 0.5% NaOCl for 1 min, rinsed three times with sterile distilled water, plated onto potato dextrose agar (PDA) and incubated at 25°C for 7 days with a 12-h photoperiod. Nine morphologically similar isolates were obtained by using single spore isolation technique and a representative isolate B was characterized further. Colonies were abundant, whitish aerial mycelium with orange pigmentation. The isolates produced macroconidia with 5 to 6 septa, a tapered with pronounced dorsiventral curvature and measured 25 to 30 μm long x 3 to 5 μm wide. Microconidia produced after 12 days of incubation were single-celled, hyaline, ovoid, nonseptate, and 1.0 to 3.0 × 4.0 to 10.0 µm. Morphological characteristics of the isolates were similar to the taxonomic description of Fusarium equiseti (Leslie and Summerell 2006). Genomic DNA was extracted from fresh mycelium using DNeasy Plant Mini kit (Qiagen, USA). To confirm the identity of the fungus, two sets of primers, ITS4/ITS5 (White et al. 1990) and TEF1-α, EF1-728F/EF1-986R (Carbone and Kohn 1999) were used to amplify complete internal transcribed spacer (ITS) and partial translation elongation factor 1-alpha (TEF1-α) genes, respectively. BLASTn search in the NCBI database using ITS and TEF-1α sequences revealed 99 to 100% similarities with species of both F. incarnatum and F. equiseti. BLAST analysis of these in FUSARIUM-ID database showed 100% and 99% similarity with Fusarium incarnatum-F. equiseti species complex (FIESC) (NRRL34059 [EF-1α] and NRRL43619 [ITS]) respectively (Geiser et al. 2004). The ITS and TEF1-α sequences were deposited in GenBank (MT515832 and MT550682). The isolate was identified as F. equiseti, which belongs to the FIESC based on morphological and molecular characteristics. Pathogenicity was conducted on five healthy leaves of 1-month-old rockmelon cv. Golden Champion grown in 5 plastic pots filled with sterile peat moss. The leaves were surface-sterilized with 70% ethanol and rinsed twice with sterile-distilled water. Then, the leaves were wounded using 34-mm-diameter florist pin frog and inoculated by pipetting 20-μl conidial suspension (1 × 106 conidia/ml) of 7-day-old culture of isolate B onto the wound sites. Control leaves were inoculated with sterile-distilled water only. The inoculated plants were covered with plastic bags for 5 days and maintained in a greenhouse at 25 °C, 90% relative humidity with a photoperiod of 12-h. After 7 days, inoculated leaves developed necrotic lesions similar to the symptoms observed in the field while the control treatment remained asymptomatic. The fungus was reisolated from the infected leaves and was morphologically identical to the original isolate. F. equiseti was previously reported causing fruit rot of watermelon in Georgia (Li and Ji 2015) and China (Li et al. 2018). This pathogen could cause serious damage to established rockmelon as it can spread rapidly in the field. To our knowledge, this is the first report of a member of the Fusarium incarnatum-F.equiseti species complex causing leaf spot on Cucumis melo in Malaysia.

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First Report of Fusarium Head Blight of Wheat Caused by Fusarium sacchari in China

Plant Disease ◽

10.1094/pdis-08-14-0829-pdn ◽

2015 ◽

Vol 99 (1) ◽

pp. 160-160 ◽

Cited By ~ 6

Author(s):

J.-H. Wang ◽

X.-D. Peng ◽

S.-H. Lin ◽

A.-B. Wu ◽

S.-L. Huang

Keyword(s):

Fusarium Head Blight ◽

Yangtze River ◽

Fusarium Species ◽

Gene Sequencing ◽

Morphological Characteristics ◽

Distilled Water ◽

The Yangtze River ◽

Head Blight ◽

Average Growth ◽

First Report

Fusarium head blight (FHB), or scab, caused by Fusarium species, is an economically devastating disease of wheat and other cereal crops worldwide. FHB epidemics in wheat occur frequently in China, especially along the middle and lower reaches of the Yangtze River, including Jiangsu and Shanghai. In 2013, wheat spikes showing clear FHB symptoms were collected from fields in Jiangsu and Shanghai. Symptomatic seeds were surface-sterilized for 1 min with a 5% sodium hypochlorite solution and dipping in 70% ethanol for 30 s, then rinsed three times in sterile distilled water and dried. They were placed onto potato dextrose agar (PDA) and incubated for 3 to 5 days at 28°C in the dark. Fungal colonies displaying morphological characteristics of Fusarium spp. (1,2) were purified by the single-spore technique and characterized at the species level by morphological observations (1,2) and translation elongation factor 1-α (TEF) gene sequencing. The results indicated that members of the Fusarium graminearum clade were predominant on wheat, while the morphological characteristics of 16 isolates were found to be identical to those of F. sacchari (1,2). Colonies on PDA were densely cottony, initially pale but becoming violet with age. The average growth rate was 6 to 8 mm per day at 25°C in the dark. Reverse pigmentation was brownish red to violet-brown. Microconidia, abundant in the aerial mycelium and formed in false heads, were oval to ellipsoidal in shape, primarily zero-septate, measuring 5.7 to 18.8 (average 10.6) μm in length. Macroconidia were slender, three- to five-septate, with a curved apical cell and a poorly developed basal cell, 26.3 to 68.9 (average 44.0) μm in length. No chlamydospores were observed. Two F. sacchari strains (Y37 and S43), isolated from Jiangsu and Shanghai, respectively, were investigated by sequence comparison of their partial TEF gene sequences (Accession Nos. KM233195 and KM233196). BLASTn analysis of the TEF sequences obtained with sequences available in the GenBank database revealed 99.8 and 99.5% sequence identity to F. sacchari (GenBank Accession Nos. JF740708 and JF740709). Pathogenicity tests were conducted by injecting 10 μl of a spore suspension (5 × 105 spores/ml) into wheat florets (20 per isolate of cv. Yangmai16), which were then grown under field conditions in Shanghai. Control plants were inoculated with sterile distilled water. Spikes were harvested and evaluated 14 days post-inoculation. Reddish white mold was observed on inoculated wheat spikes; in addition, spikelets adjacent to the inoculation point and the infected florets were brown. No symptoms were observed on water controls. Koch's postulates were fulfilled by reisolating the pathogen from infected florets and identifying them by TEF gene sequencing. F. sacchari is the cause of an important disease of sugar cane, pokkah boeng (1), and has been reported to produce the mycotoxin beauvericin, which causes toxicosis in human and other animals (3). To our knowledge, this is the first report of F. sacchari causing wheat head blight in China. The report contributes to an improved understanding of the composition of Fusarium species on wheat in the lower reaches of the Yangtze River in China, which will be useful for exploring appropriate disease management strategies in this region. References: (1) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Ames, IA, 2006. (2) J. F. Leslie et al. Mycologia 97:718, 2005. (3) A. Moretti et al. Int. J. Food Microbiol. 118:158, 2007.

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First Report of a New Postharvest Rot in Sweet Cherries Caused by Aureobasidium pullulans

Plant Disease ◽

10.1094/pdis-07-13-0740-pdn ◽

2014 ◽

Vol 98 (3) ◽

pp. 424-424 ◽

Cited By ~ 7

Author(s):

Y. K. Kim

Keyword(s):

Aureobasidium Pullulans ◽

Deionized Water ◽

Causal Agent ◽

Fruit Rot ◽

Conidial Suspension ◽

Centraalbureau Voor ◽

First Report ◽

Control Fruit ◽

Initial Symptoms ◽

Sweet Cherries

During August to October 2012, several cherry packers in central Washington State reported that a significant volume of sweet cherries (Prunus avium) (cvs. Staccato, Sweetheart, and Lapin) were rotten by an unknown fungal pathogen after packing. Of 14 boxes (9 kg per box) of commercially packed cherries rejected by a retailer, the average incidence of the decay was 68%. Initial symptoms on infected fruit appeared as soft, slippery skin with tan discoloration and later skin cracking, epidermal breakdown, and severe pitting were observed. To isolate the causal agent, decayed fruit were rinsed with water, sprayed with 70% ethanol, and air-dried in a laminar hood. After removing the fruit skin with a sterile scalpel, small fragments of fruit flesh between decayed and healthy tissue were cut and placed on potato dextrose agar (PDA) acidified with 0.1% lactic acid. The plates were incubated at 20°C for 7 days and sub-cultured on PDA to obtain pure cultures. The colonies initially appeared white to cream, yeast-like, and later turned to light yellow to pink or brown with age. Conidia were hyaline, smooth-walled, single-celled, and ellipsoidal with variable shape and size. The fungus was identified as Aureobasidium pullulans (de Bary) G. Arnaud based on its morphology (1). The identity of three representative isolates were further confirmed by analysis of nucleotide sequences of the internal transcribed spacer (ITS) regions amplified using the primers ITS1/ITS4. A BLAST search showed that the sequences had 99% homology (E-value = 0.0) with that of A. pullulans deposited at GenBank (Accession No. JF440584.1). The nucleotide sequence of the isolate, A625, has been assigned GenBank Accession No. KF569512. To test pathogenicity, three single-spore isolates were grown on PDA at 20°C. Cultures grown on 10-day-old PDA were flooded with 20 ml of sterile deionized water, and the resulting conidial suspensions were filtered through two layers of cheesecloth and adjusted to 5 × 105 conidia/ml with a hemacytometer. Organic cherry fruit (cv. Bing for isolate A625 and cv. Sweetheart for isolates A755 and A757) were surface-disinfested in 0.6% sodium hypochlorite solution for 5 min, rinsed twice with deionized water, and air-dried. Ten fruit per replicate, four replications per treatment were inoculated with the conidial suspension using a hand sprayer and placed on sterilized wet paper towel in a plastic container. Control fruit were sprayed with sterile water. All fruit were incubated at 22 ± 1°C for 5 days. The experiments were conducted twice. The same symptoms of skin cracking and epidermal breakdown developed on 73% of the inoculated fruit, while no such symptoms appeared on the control fruit. Koch's postulates were fulfilled by re-isolating the fungus from the symptomatic fruit. A. pullulans, a ubiquitous saprophytic fungus on many fruits, has been reported as a causal agent of melting decay in grapes (2). To the best of our knowledge, this is the first report of postharvest fruit rot in sweet cherries caused by A. pullulans. References: (1) E. J. Hermanides-Nijhof. Aureobasidium and related genera. Pages 141-181 in: The Black Yeasts and Allied Hyphomycetes. Stud. Mycol. No. 15. Centraalbureau voor Schimmelcultures, Baarn, The Netherlands, 1977. (2) D. P. Morgan and T. J. Michailides. Plant Dis. 88:1047, 2004.

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