First Report of Phyllosticta capitalensis Causing Black Freckle Disease on Rubus chingii in China

Rubus chingii is used as an important traditional Chinese medicine, and belongs to the family Rosaceae. The fruit has multiple pharmacological activities, including antioxidant, anti-inflammatory, and improving cognitive impairment (Na Han et al. 2012). In June 2019, a new fungal infection was observed on the leaves of R. chingii in Qiandongnan Miao and Dong Autonomous Prefecture, Guizhou Province, China, forming small lesions with reddish-brown edges along leaf veins. Over 500 plants were surveyed, and nearly 20% of the plants were symptomatic. The diseased plants grew poorly and appeared stunted, and severely affected plants died. Five symptomatic leaves were randomly collected from the field and washed with tap water and distilled water successively. The edges of infected leaf tissue were cut into small pieces (4 to 5 mm2), surface sterilized with 70% ethanol for 30 s and 0.1% HgCl2 for 1 minute, and then rinsed three times in sterile distilled water (Chen et al. 2016). The same fungus was isolated from 41 pieces. The hyphae of a representative isolate were gray, the colony surface was granular, the edges were uneven and white, and the culture turned black over time with black spherical conidia. Conidia were nearly elliptical, unicellular, and each with a hyaline, unstable apical appendage, 3 to 10 µm long. The size of conidia was 10 to 18 μm in length and 4 to 8 μm in width. These morphological characteristics are consistent with those described for the fungus Phyllosticta capitalensis. (Wikee et al. 2013). For an accurate identification, genomic DNA of a representative isolate of the pathogen was extracted to amplify the internal transcribed spacer (ITS) region, the transcription elongation factor (tefa-1), and actin (ACT) genes with the ITS1/ITS4, EF1-728F/EF1-986R, and ACT-512F/ACT-783R (Cheng, L. L. ,et al. 2019), respectively. The ITS, tefa-1 and actin gene sequences were deposited in GenBank and assigned accession numbers MW308365, MW714380 and MW714381, respectively. BLAST search analysis of GenBank (NCBI) showed that the sequences had 100% similarity with those of Phyllosticta capitalensis (GenBank accession no. ITS, MN548091; tefa-1, MN958711; and ACT, MN565575). The pathogenicity of Phyllosticta capitalensis was verified using six healthy detached leaves from healthy R. chingii plants around 40 cm tall. A total of nine plants were used, and three leaves from each plant were artificially inoculated. Each wound was inoculated with conidial suspension (106 mL-1), while the control leaves were coated by sterile water. All the treated plants were covered with plastic bags for 2 days, incubated at 28ºC and 85% relative humidity, with a 12-hour photoperiod. After 15 days following inoculation, the injured leaves showed similar symptoms to the above-mentioned lesions, while the control and uninjured leaves were still healthy. P. capitalensis were reisolated from inoculated leaves, fulfilling Koch’s postulates. P. capitalensis is an endophyte, widely distributed in various host plants in China. (Lu, J. M, et al. 2016). To the best of our known, this is the first report of black freckle disease caused by P. capitalensis on Rubus chingii in China. P. capitalensis is a destructive plant pathogen with an unusually broad host range and our findings will be useful for its management and for further research. The author(s) declare no conflict of interest.

Download Full-text

First Report of Fusarium Wilt Caused by Fusarium equiseti on Cabbage (Brassica oleracea var. capitate L.) in Korea

Plant Disease ◽

10.1094/pdis-06-20-1278-pdn ◽

2020 ◽

Author(s):

Tania Afroz ◽

Samnyu JEE ◽

Hyo-Won Choi ◽

Ji Hyeon Kim ◽

Awraris Derbie Assefa ◽

...

Keyword(s):

Brassica Oleracea ◽

Fusarium Wilt ◽

Apical Cell ◽

Morphological Characteristics ◽

Distilled Water ◽

Conidial Suspension ◽

Mlst Database ◽

Fusarium Equiseti ◽

First Report ◽

Representative Isolate

Cabbage (Brassica oleracea var. capitate L.) is an important vegetable crop that is widely cultivated throughout the world. In August 2019, wilting symptoms on cabbage (stunted growth, withered leaves, and wilted plants) were observed in a cabbage field of Pyeongchang, Gangwon Province, with an incidence of 5 to 10%. To identify the cause, symptomatic root tissue was excised, surface-sterilized with 70% ethanol, and rinsed thrice with sterile distilled water. The samples were dried on blotter paper, placed onto potato dextrose agar (PDA), and incubated at 25°C for 1 week. Five morphologically similar fungal isolates were sub-cultured and purified using the single spore isolation method (Choi et al. 1999). The fungus produced colonies with abundant, loosely floccose, whitish-brown aerial mycelia and pale-orange pigmentation on PDA. Macroconidia had four 4 to six 6 septa, a foot-shaped basal cell, an elongated apical cell, and a size of 20.2 to 31.8 × 2.2 to 4.1 μm (n = 30). No microconidia were observed. Chlamydospores were produced from hyphae and were most often intercalary, in pairs or solitary, globose, and frequently formed chains (6.2? to 11.7 μm, n = 10). Based on these morphological characteristics, the fungus was identified as Fusarium equiseti (Leslie and Summerell 2006). A representative isolate was deposited in the Korean Agricultural Culture Collection (KACC48935). For molecular characterization, portions of the translation elongation factor 1-alpha (TEF-1α) and second largest subunit of RNA polymerase II (RPB2) genes were amplified from the representative isolate using the primers pair of TEF-1α (O’Donnell et al. 2000) and GQ505815 (Fusarium MLST database), and sequenced. Searched BLASTn of the RPB2 sequence (MT576587) to the Fusarium MLST database showed 99.94% similarity to the F. incarnatum-equiseti species complex (GQ505850) and 98.85 % identity to both F. equiseti (GQ505599) and F. equiseti (GQ505772). Further, the TEF-1α sequence (MT084815) showed 100% identity to F. equiseti (KT224215) and 99.85% identity to F. equiseti (GQ505599), respectively. Therefore, the fungus was identified as F. equiseti based on morphological and molecular identification. For pathogenicity testing, a conidial suspension (1 × 106 conidia/ml) was prepared by harvesting macroconidia from 2-week-old cultures on PDA. Fifteen 4-week-old cabbage seedlings (cv. 12-Aadrika) were inoculated by dipping roots into the conidial suspension for 30 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 4 days, the first wilt symptoms were observed on inoculated seedlings, and the infected plants eventually died within 1 to 2 weeks after inoculation. No symptoms were observed in plants inoculated with sterilized distilled water. The fungus was re-isolated from symptomatic tissues of inoculated plants and its colony and spore morphology were identical to those of the original isolate, thus confirming Koch's postulates. Fusarium wilt caused by F. equiseti has been reported in various crops, such as cauliflower in China, cumin in India, and Vitis vinifera in Spain (Farr and Rossman 2020). To our knowledge, this is the first report of F. equiseti causing Fusarium wilt on cabbage in Korea. It This disease poses a threat to cabbage production in Korea, and effective disease management strategies need to be developed.

Download Full-text

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.

Download Full-text

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.

Download Full-text

First Report of Nigrospora Leaf Blight on Tea Caused by Nigrospora sphaerica in India

Plant Disease ◽

10.1094/pdis-05-14-0545-pdn ◽

2015 ◽

Vol 99 (3) ◽

pp. 417-417 ◽

Cited By ~ 11

Author(s):

J. Dutta ◽

S. Gupta ◽

D. Thakur ◽

P. J. Handique

Keyword(s):

Camellia Sinensis ◽

West Bengal ◽

Morphological Characteristics ◽

Its Region ◽

Causative Organism ◽

Distilled Water ◽

Conidial Suspension ◽

Tea Leaves ◽

First Report ◽

Initial Symptoms

Tea [Camellia sinensis (L.) O. Kuntze] is an economically important non-alcoholic caffeine-containing beverage crop widely cultivated for leaves in India, especially in the Darjeeling district of West Bengal. In May 2012, distinct blight symptoms were observed on leaves of popular tea cultivars AV-2, Tukdah 78, Rungli Rungliot 17/144, and Bannockburn 157 in commercial tea estates of the Darjeeling district. This disease reduces yield and quality of the leaves. The initial symptoms were frequently observed on the young leaf margins and apices. Foliar symptoms are characterized by grayish to brown, semicircular or irregular shaped lesions, often surrounded by pale yellow zones up to 9 mm in diameter. The lesions later expand and the affected leaves turn grayish to dark brown and eventually the dried tissue falls, leading to complete defoliation of the plant. The disease causes damage to leaves of all ages and is severe in young leaves. A portion of the symptomatic leaf tissues were surface sterilized in 70% ethanol for 30 s, then in 2% NaClO for 3 min, rinsed three times in sterile distilled water, and plated onto potato dextrose agar (PDA). The fungal colonies were initially white and then became grayish to brown with sporulation. Conidia were spherical to sub spherical, single-celled, black, 19 to 21 μm in diameter, and were borne on a hyaline vesicle at the tip of each conidiophore. Morphological characteristics of the isolates were concurring to those of Nigrospora sphaerica (1). Moreover, the internal transcribed spacer (ITS) region of the ribosomal RNA was amplified by using primers ITS1 and ITS4 and sequenced (GenBank Accession No. KJ767520). The sequence was compared to the GenBank database through nucleotide BLAST search and the isolate showed 100% similarity to N. sphaerica (KC519729.1). On the basis of morphological characteristics and nucleotide homology, the isolate was identified as N. sphaerica. Koch's postulates were fulfilled in the laboratory on tea leaves inoculated with N. sphaerica conidial suspension (106 conidia ml−1) collected from a 7-day-old culture on PDA. Six inoculated 8-month-old seedlings of tea cultivars AV-2 and S.3/3 were incubated in a controlled environment chamber at 25°C and 80 to 85% humidity with a 12-h photoperiod. In addition, three plants of each cultivar were sprayed with sterile distilled water to serve as controls. Twelve to 14 days after inoculation, inoculated leaves developed blight symptoms similar to those observed on naturally infected tea leaves in the field. No symptoms were observed on the control leaves. The pathogen was re-isolated from lesions and its identity was confirmed by morphological characteristics. It was reported that N. sphaerica is frequently encountered as a secondary invader or as a saprophyte on many plant species and also as a causative organism of foliar disease on several hosts worldwide (2,3). To our knowledge, this is first report of N. sphaerica as a foliar pathogen of Camellia sinensis in Darjeeling, West Bengal, India, or worldwide. References: (1) M. B. Ellis. Dematiaceous Hyphomycetes. CMI, Kew, Surrey, UK, 1971. (2) D. F. Farr and A. Y. Rossman. Fungal Databases, Syst. Mycol. Microbiol. Lab., ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/ July 01, 2013. (3) E. R. Wright et al. Plant Dis. 92:171, 2008.

Download Full-text

First report of fruit blight caused by Alternaria alternata on sesame in Northeast China

Plant Disease ◽

10.1094/pdis-01-21-0047-pdn ◽

2021 ◽

Author(s):

Hongsen Cheng ◽

De Xue Gao ◽

Huijie Sun ◽

Yanbin Na ◽

Jing Xu

Keyword(s):

Alternaria Alternata ◽

Morphological Characteristics ◽

Its Region ◽

Liaoning Province ◽

Oilseed Crop ◽

Distilled Water ◽

Conidial Suspension ◽

First Report ◽

Health Products ◽

Blight Disease

Sesame (Sesamum indicum L.) is an important oilseed crop in China and it is also used in food and health products. In August of 2019, a blight sesame fruit was observed in a field of Liaoyang city, Liaoning province of China. Initial disease symptoms consisted of brown or dark brown spots on fruit. With time, lesions coalesced and the whole fruit turned dark brown or black. Most of the diseased fruit had thin and small, deformed, necrotic, hardened cracked epidermal lesions. Lesions were also produced on stem and petioles leading to leaf abscission. The disease results in premature fruit death, and in turn, considerable yield losses. To determine the causal agent, symptomatic fruit with developing lesions were collected, and surface sterilized in 2% NaClO for 3 min, rinsed three times in distilled water, and plated onto PDA medium. After incubation at 25°C for 5 days, a dark olivaceous fungus with abundant, branched, brown to black, and septate hyphae was consistently isolated. Twenty single spores were separated with an inoculation needle under stereomicroscope. The conidia were in chains, brown, obclavate, ovoid or ellipsoid, with 1-6 transverse septa and 0-4 longitudinal or oblique septa 12.5 to 45 × 6.5 to 14.5 μm in size. Conidiophores were septate, light brown to olive brown, measuring 22-60 μm × 2-4 μm. The morphological characteristics of the 20 isolates all matched the description of Alternaria alternata (Simmons, 2007). The internal transcribed spacer (ITS) region of rDNA of 15 isolates was amplified using primers ITS1/ITS4 (White et al. 1990) and EF1-728F/EF1-986R (Carbone et al. 1999) and sequenced. Identical sequences were obtained and the sequence of the isolate ZMHG12 was submitted to GenBank (Accession no. MW418181 and MW700316). BLAST analysis of the sequences of the isolates of ZMHG12 showed 100% to A. alternata (KP739875 and LC132712). In pathogenicity tests, a conidial suspension (2.5 × 105 conidia per ml) was prepared from 7 days-old cultures of isolate ZMHG12 grown on PDA at 25°C. Fruit of 10 two-month-old potted sesame plants (Variety “Liaozhi 8”) were sprayed with the conidia suspension until runoff. Another 10 plants sprayed with distilled water to served as non-inoculated controls. All plants were maintained for 48 h in a humid chamber with a temperature of 25°C to 26°C, and then moved to a greenhouse. Ten days after inoculation, all fruit of inoculated plants exhibited symptoms similar to those observed in the field and non-inoculated control plants remained symptomless. The experiment was repeated twice with similar results. A. alternata has been reported as a pathogen caused leaf blight disease of sesame in Pakistan (Nayyar et al. 2017). To our knowledge, this is the first report of A.alternata causing fruit blight of sesame in China. To date, we have observed the disease on sesames in fields of Fuxin, Chaoyang and Tieling city in Liaoning Province, and Tongliao city in Inner Mongolia of China, and it has become an important disease in sesame production of China. References : Simmons E. G. 2007. Alternaria: An identification manual. CBS Fungal Biodiversity Center, Utrecht, Netherlands. White T. J., et al. 1990. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego. Carbone I., et al. 1999. Mycologia, 91: 553-556. Nayyar, B. G., et al. 2017. Plant Pathology Journal, 33 (6): 543-553.

Download Full-text

First Report of Fusarium ipomoeae Causing Peanut leaf Spot in China

Plant Disease ◽

10.1094/pdis-01-21-0226-pdn ◽

2021 ◽

Author(s):

Manlin Xu ◽

Xia Zhang ◽

Jing Yu ◽

zhiqing Guo ◽

Ying Li ◽

...

Keyword(s):

Leaf Spot ◽

Genomic Dna ◽

Block Design ◽

Management Strategies ◽

Morphological Characteristics ◽

Ethanol Solution ◽

Distilled Water ◽

Conidial Suspension ◽

Internal Transcribed Spacer Region ◽

First Report

Peanut (Arachis hypogaea L.) is one of the most economically important crops as an important source of edible oil and protein. In August 2020, circular to oval-shaped brown leaf spots (2-6 mm in diameter) with well-defined borders surrounded by a yellow margin were observed on peanut plant leaves in Laixi City, Shandong Province, China. Symptomatic plants randomly distributed in the field, the incidence was approximately 5%. Leave samples were collected consisted of diseased tissue and the adjacent healthy tissue. The samples were dipped in a 70% (v/v) ethanol solution for 30 s and then soaked in a 0.1% (w/v) mercuric chloride solution for 60 s. The surface-sterilized tissues were then rinsed three times with sterile distilled water, dried and placed on Czapek Dox agar supplemented with 100 μg/ml of chloramphenicol. The cultures were incubated in darkness at 25 °C for 3–5 days. Fungal colonies were initially white and radial, turning to orange-brown in color, with abundant aerial mycelia. Macroconidia were abundant, 4 to 7 septate, with a dorsiventral curvature, and were 3.3–4.5 × 18.5–38.1 μm (n=100) in size; microconidia were absent; chlamydospores were produced in chains or clumps, ellipsoidal to subglobose, and thick walled. The morphological characteristics of the conidia were consistent with those of Fusarium spp. To identify the fungus, an EasyPure Genomic DNA Kit (TransGEN, Beijing, China) was used to extract the total genomic DNA from mycelia. The internal transcribed spacer region (ITS rDNA) and the translation elongation factor 1-α gene (TEF1) were amplified with primers ITS1/ITS4 (White et al. 1990) and EF1/EF2 (O’Donnell et al. 1998), respectively. Based on BLAST analysis, sequences of ITS (MT928727) and TEF1 (MT952337) showed 99.64% and 100% similarity to the ITS (MT939248.1), TEF1 (GQ505636.1) of F. ipomoeae isolates. Sequence analysis confirmed that the fungus isolated from the infected peanut was F. ipomoeae (Xia et al. 2019). The pathogenicity of the fungus was tested in the greenhouse. Twenty two-week-old peanut seedlings (cv. Huayu20) grown in 20-cm pots (containing autoclaved soil) were sprayed with a conidial suspension (105 ml−1) from a 15-day-old culture. Control plants were sprayed with distilled water. The experiment was conducted as a randomized complete block design, and placed at 25 °C under a 12-h photoperiod with 90% humidity. Symptoms similar to those in the field were observed on leaves treated with the conidial suspension ten days after inoculation, but not on control plants. F. ipomoeae was re-isolated from symptomatic leaves but not from the control plants. Reisolation of F. ipomoeae from inoculated plants fulfilled Koch's postulates. To our knowledge, this is the first report of F. ipomoeae causing peanut leaf spot in China. Our report indicates the potential spread of this pathogen in China and a systematic survey is required to develop effective disease management strategies.

Download Full-text

First report of Fusarium commune causing root and crown rot on maize in Italy

Plant Disease ◽

10.1094/pdis-01-21-0075-pdn ◽

2021 ◽

Author(s):

Monica Mezzalama ◽

Vladimiro Guarnaccia ◽

Ilaria Martino ◽

Giulia Tabome ◽

Maria Lodovica GULLINO

Keyword(s):

Plant Disease ◽

Tap Water ◽

Morphological Characteristics ◽

Plant Diseases ◽

Crown Rot ◽

Conidial Suspension ◽

Fusarium Spp ◽

First Report ◽

Root And Crown Rot ◽

Pathogenicity Tests

Maize (Zea mays L.) is a cereal crop of great economic importance in Italy; production is currently of 62,587,469 t, with an area that covers 628,801 ha, concentrated in northern Italy (ISTAT 2020). Fusarium species are associated with root and crown rot causing failures in crop establishment under high soil moisture. In 2019 maize seedlings collected in a farm located in San Zenone degli Ezzelini (VI, Italy) showed root and crown rot symptoms with browning of the stem tissues, wilting of the seedling, and collapsing due to the rotting tissues at the base of the stem. The incidence of diseased plants was approximately 15%. Seedlings were cleaned thoroughly from soil residues under tap water. Portions (about 3-5 mm) of tissue from roots and crowns of the diseased plants were cut and surface disinfected with a water solution of NaClO at 0.5% for 2 minutes and rinsed in sterile H20. The tissue fragments were plated on Potato Dextrose Agar (PDA) amended with 50 mg/l of streptomycin sulfate and incubated for 48-72 hours at 25oC. Over the 80 tissue fragments plated, 5% were identified as Fusarium verticillioides, 60% as Fusarium spp., 35% developed saprophytes. Fusarium spp. isolates that showed morphological characteristics not belonging to known pathogenic species on maize were selected and used for further investigation while species belonging to F. oxysporum were discarded. Single conidia of the Fusarium spp. colonies were cultured on PDA and Carnation Leaf Agar (CLA) for pathogenicity tests, morphological and molecular identification. The colonies showed white to pink, abundant, densely floccose to fluffy aerial mycelium. Colony reverse showed light violet pigmentation, in rings on PDA. On CLA the isolates produced slightly curved macronidia with 3 septa 28.1 - 65.5 µm long and 2.8-6.3 µm wide (n=50). Microconidia were cylindrical, aseptate, 4.5 -14.0 µm long and 1.5-3.9 µm wide (n=50). Spherical clamydospores were 8.8 ± 2.5 µm size (n=30), produced singly or in pairs on the mycelium, according to the description by Skovgaard et al. (2003) for F. commune. The identity of two single-conidia strains was confirmed by sequence comparison of the translation elongation factor-1α (tef-1α), and RNA polymerase II subunit (rpb2) gene fragments (O’Donnell et al. 2010). BLASTn searches of GenBank, and Fusarium-ID database, using the partial tef-1α (MW419921, MW419922) and rpb2 (MW419923, MW419924) sequences of representative isolate DB19lug07 and DB19lug20, revealed 99% identity for tef-1α and 100% identity to F. commune NRRL 28387(AF246832, AF250560). Pathogenicity tests were carried out by suspending conidia from a 10-days old culture on PDA in sterile H2O to 5×104 CFU/ml. Fifty seeds were immersed in 50 ml of the conidial suspension of each isolate for 24 hours and in sterile water (Koch et al. 2020). The seeds were drained, dried at room temperature, and sown in trays filled with a steamed mix of white peat and perlite, 80:20 v/v, and maintained at 25°C and RH of 80-85% for 14 days with 12 hours photoperiod. Seedlings were extracted from the substrate, washed under tap water, and observed for the presence of root and crown rots like the symptoms observed on the seedlings collected in the field. Control seedlings were healthy and F. commune was reisolated from the symptomatic ones and identified by resequencing of tef-1α gene. F. commune has been already reported on maize (Xi et al. 2019) and other plant species, like soybean (Ellis et al. 2013), sugarcane (Wang et al. 2018), potato (Osawa et al. 2020), indicating that some attention must be paid in crop rotation and residue management strategies. To our knowledge this is the first report of F. commune as a pathogen of maize in Italy. References Ellis M L et al. 2013. Plant Disease, 97, doi: 10.1094/PDIS-07-12-0644-PDN. ISTAT. 2020. http://dati.istat.it/Index.aspx?QueryId=33702. Accessed December 28, 2020. Koch, E. et al. 2020. Journal of Plant Diseases and Protection. 127, 883–893 doi: 10.1007/s41348-020-00350-w O’Donnell K et al. 2010. J. Clin. Microbiol. 48:3708. https://doi.org/10.1128/JCM.00989-10 Osawa H et al. 2020. Journal of General Plant Pathology, doi.org/10.1007/s10327-020-00969-5. Skovgaard K 2003. Mycologia, 95:4, 630-636, DOI: 10.1080/15572536.2004.11833067. Wang J et al. 2018. Plant Disease, 102, doi/10.1094/PDIS-07-17-1011-PDN Xi K et al. 2019. Plant Disease, 103, doi/10.1094/PDIS-09-18-1674-PDN

Download Full-text

First Report of Seedling Stem Rot on Jinxianlian (Anoectochilus roxburghii) Caused by Fusarium oxysporum in China

Plant Disease ◽

10.1094/pdis-09-21-2066-pdn ◽

2021 ◽

Author(s):

Chuan-Qing Zhang ◽

X. Y. Chen ◽

Ya-hui Liu ◽

Dejiang Dai

Keyword(s):

Fusarium Oxysporum ◽

Morphological Characteristics ◽

Aerial Mycelium ◽

Morphological Features ◽

Stem Rot ◽

Broad Host Range ◽

Distilled Water ◽

Internal Transcribed Spacer Region ◽

First Report ◽

Anoectochilus Roxburghii

Anoectochilus roxburghii is an important Chinese herbal medicine plant belonging to Orchidaceae and known as Jinxianlian. This orchid is cultivated and mostly adopted to treat diabetes and hepatitis. About 2 billion artificially cultivated seedlings of Jinxianlian are required each year and approximately $600 million in fresh A. roxburghii seedlings is produced in China. From 2011, sporadic occurrence of stem rot on Jinxianlian have been observed in greenhouses in Jinhua City (N29°05′, E119°38′), Zhejiang Province. In 2018, nearly 30% of seedlings of Jinxianlian grown in greenhouse conditions were affected by stem rot in Jinhua City. Symptoms initially occurred in the stem at the soil line causing dark discoloration lesions, rotted tissues, wilting, and eventually leading to the death of the plants. A total of 23 diseased seedlings collected from seven different greenhouses were surface sterilized with 1.5% sodium hypochlorite for 3 min, then rinsed in water. Pieces of tissues disinfected from each sample were plated on 2% potato dextrose agar (PDA), and incubated at 25°C in the dark for 5 days (Kirk et al. 2008). A total of 19 isolates were recovered. They developed colonies with purple mycelia and beige or orange colors after 7 days of incubation under 25°C on PDA and carnation leaf agar (CLA) media (Kirk et al. 2008; Zhang et al. 2016). Colonies on PDA had an average radial growth rate of 3.1 to 4.0 mm /d at 25°C. Colony surface was pale vinaceous, floccose with abundant aerial mycelium. On CLA, aerial mycelium was sparse with abundant bright orange sporodochia forming on the carnation leaves. Microconidia were hyaline and oval-ellipsoid to cylindrical (3.7 to 9.3 × 1.3 to 2.9 μm) (n=19). Macroconidia were 3 to 5 septate and fusoid-subulate with a pedicellate base (27.4 to 35.6 × 3.2 to 4.2 μm) (n=19). These morphological features were consistent with Fusarium oxysporum (Sun et al. 2008; Lombard et al., 2019). To confirm the identification based on these morphological features, the internal transcribed spacer region (ITS) and translation elongation factor1 (TEF) were amplified from the DNA of 3 out of 19 isolates chosen at random respectively using the set primer ITS1/ITS4 and EF1/ EF2 (Sun, S., et al. 2018; Lombard et al., 2019). BLAST analysis revealed that the ITS sequences (OK147619, OK147620, OK147621) had 99% identity to that of F. oxysporum isolate JJF2 (GenBank MN626452) and TEF sequence (OK155999, OK156000, OK156001) had 100% identity to that of F. oxysporum isolate gss100 (GenBank MH341210). A multilocus phylogenetic analysis by Bayesian inference (BI) and maximum likelihood (ML) trees based on ITS and TEF indicated that the pathogen grouped consistently with F. oxysporum. Three out of 19 isolates chosen at random were selected to evaluate pathogenicity. Uninfected healthy A. roxburghii seedlings about 40 day-old planted in sterilized substrates were sprayed with distilled water containing 2 x 106 conidia per ml suspensions as inoculums, and plants sprayed with distilled water alone served as controls. Plants were then incubated at 25°C and 85% relative humidity. Ten plants were inoculated for each isolate. After 10 days, all plants inoculated developed stem rot symptoms, while control plants remained healthy. Cultures of Fusarium spp. were re-isolated only from inoculated plants with the frequency of 100% and re-identified by morphological characteristics as F. oxysporum, fulfilling Koch’s postulates. To the best of our knowledge, this is the first report of F. oxysporum causing stem rot on A. roxburghii seedlings. As F. oxysporum is a devastating pathogenic fungus with a broad host range, measures should be taken in advance to manage stem rot of A. roxburghii.

Download Full-text

First Report of Septoria Blight of Parsley Caused by Septoria petroselini in the Mediterranean Region of Turkey

Plant Disease ◽

10.1094/pdis.2003.87.1.99a ◽

2003 ◽

Vol 87 (1) ◽

pp. 99-99 ◽

Cited By ~ 3

Author(s):

S. Kurt

Keyword(s):

Mediterranean Region ◽

Morphological Characteristics ◽

Potato Dextrose Agar ◽

Distilled Water ◽

Conidial Suspension ◽

Sterile Water ◽

First Report ◽

Foliar Symptoms ◽

Polyethylene Bags ◽

The Mediterranean

During December 2001 to March 2002, Septoria blight of parsley was observed in approximately 500 ha of commercial parsley crops in Arsuz County, Hatay, in the Mediterranean Region of Turkey. Incidence of disease ranged from 42 to 80%. Symptoms included irregularly shaped, grayish brown spots (average 3 to 8 mm diameter) with a slightly darker brown margin of necrotic tissue that developed into tan-to-brown lesions surrounded by chlorotic halo on the leaves. Oval-shaped lesions were observed occasionally on petioles. Lesions contained erumpent, dark brown, flask-shaped pycnidia with the ostiole on the upper surface of the foliage. Thirty samples, consisting of diseased leaves and petioles of parsley, were collected from each field. Infected tissues were surface-sterilized in 1% NaOCl for 2 min, rinsed in sterile distilled water, placed on petri dishes containing potato dextrose agar (PDA), and incubated for 10 to 14 days at 25°C. The fungus formed long, multiseptate (0 to 4), hyaline, filiform conidia (14 to 29 μm × 0.5 to 1.9 μm), and short conidiophores within the pycnidia. Based on the morphological characteristics of the fungus, the pathogen was identified as Septoria petroselini Desm. (1). Monoconidial cultures of 18 isolates were prepared. Pathogenicity was confirmed by brush-inoculating slightly wounded foliage of 5- to 7- week-old parsley plants (cv. Kereviz yapragi) with a conidial suspension (106 conidia per ml of sterile water) of each isolate of S. petroselini. Control plants that were brush-inoculated with distilled water and inoculated plants were placed in clear polyethylene bags that were closed and incubated at 20°C for 48 h. The bags were removed, and plants were maintained in a dew chamber for 21 days at 65 to 70% relative humidity. Foliar symptoms developed 15 days after inoculation and appeared similar to lesions observed in the field. Yellowing and necrosis of leaves was also observed on >60% of inoculated plants. No lesions developed on the control plants. The pathogen was readily reisolated on PDA from inoculated plants. To our knowledge, this is the first report of Septoria blight of parsley in the Mediterranean Region of Turkey. Reference: (1) R. F. Cerkauskas and J. Uyenaka. Plant Dis. 74:1037, 1990.

Download Full-text

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

Plant Disease ◽

10.1094/pdis-09-20-1877-pdn ◽

2021 ◽

Author(s):

Yun-fei Mao ◽

Li Jin ◽

Huiyue Chen ◽

Xiang-rong Zheng ◽

Minjia Wang ◽

...

Keyword(s):

Leaf Spot ◽

Disease Incidence ◽

Morphological Characteristics ◽

Liquidambar Styraciflua ◽

Leaf Spot Disease ◽

Distilled Water ◽

Conidial Suspension ◽

First Report ◽

Wood Processing ◽

Light Yellow

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

Download Full-text