scholarly journals Morphology, Molecular Phylogeny, and Pathogenicity of Neofusicoccum parvum, Associated with Leaf Spot Disease of a New Host, the Japanese Bay Tree (Machilus thunbergii)

Forests ◽  
2021 ◽  
Vol 12 (4) ◽  
pp. 440
Author(s):  
Sungyu Choi ◽  
Narayan Chandra Paul ◽  
Kye-Han Lee ◽  
Hyun-Jun Kim ◽  
Hyunkyu Sang
Keyword(s):  
Leaf Spot ◽  
Morphological Characteristics ◽  
Aerial Mycelium ◽  
Its Region ◽  
Leaf Spot Disease ◽  
New Host ◽  
Neofusicoccum Parvum ◽  
Spot Disease ◽  
Leaf Spots ◽  
Machilus Thunbergii

During a survey of diseased plants on Wando Island, Korea from May to June 2020, a severe leaf spot disease was observed in the upper leaves of Japanese bay tree (Machilus thunbergii). Early symptoms were light blackish spots on the leaf surface and enlargement of older spots. Dry leaf spots surrounded with deep black margins were common throughout the plants. Symptomatic leaf samples were collected, and the causal pathogen was isolated on potato dextrose agar (PDA). Three fungal isolates (CMML20-1, CMML20-3, and CMML20-4) were cultured on PDA for morphological characterization at 25 °C in the darkness. Fungal colonies were circular, fast-growing, olivaceous to dark grey, and with abundant aerial mycelium. Sporulation was induced in 14 h-10 h light-dark conditions, and the conidia were single-celled, thin-walled with a smooth surface, ellipsoid with round apices, and measuring 17.5–20.5 (avg. 17.5) μm × 7.5–10.0 (7.9) μm. The morphological characteristics resembled those typical for Neofusicoccum parvum. Molecular identification was confirmed by partially sequencing the internal transcribed spacer (ITS) region and the translation elongation factor 1-α (EF1-α) genes. Pathogenicity tests were conducted on detached leaves and whole plants of M. thunbergii. High disease prevalence was observed, and Koch postulates were fulfilled. This is the first worldwide report of N. parvum causing leaf spots on Machilus thunbergii.

scholarly journals First Report of Leaf Spot Disease in Coconut Seedling Caused by Bipolaris setariae in China

Plant Disease ◽  
2014 ◽  
Vol 98 (12) ◽  
pp. 1742-1742 ◽  
Author(s):  
X.-Q. Niu ◽  
F.-Y. Yu ◽  
H. Zhu ◽  
W.-Q. Qin
Keyword(s):  
Leaf Spot ◽  
Sequence Similarity ◽  
Cocos Nucifera ◽  
Morphological Characteristics ◽  
Aerial Mycelium ◽  
Its Region ◽  
Leaf Spot Disease ◽  
First Report ◽  
Spot Disease ◽  
Cocos Nucifera L

Coconut (Cocos nucifera L.), an important oilseed as well as a multipurpose perennial plantation crop, is distributed and planted in humid tropical areas. In October 2012, a new leaf spot disease was observed on 3-year-old coconut seedlings in Wenchang, Hainan Province, China. The symptom first appeared as spindly or elliptical and brown flecks with water-soaked lesions that became yellow with the progress of the disease. In the later stage of the disease, the lesions merged together, gradually expanding to the leaf apex. In recent years, the disease has been prevalent in all the nursery gardens surveyed. Once young leaves got infected and nearly all the leaves of the tree showed diseased symptoms, the coconut eventually became defoliated. The pathogen was isolated from the lesion margin, surface sterilized with 75% ethanol and 0.1% mercury bichloride, washed by sterile distilled water, and then placed excising pieces of leaves from the leision margin onto potato dextrose agar (PDA). Plates were incubated at 25°C for 4 days. After 7 days, the colony was grayish black and produced black pigment in the medium. Aerial mycelium was fluffy, septate, and branched, the conidiophores were slightly flexuous or straight, 5 to 11 μm thick, and produced curved, spindle-shaped, or fusiform, septate conidia with 4 to 10 septa, measuring 39 to 86 × 9 to 16 μm, with a slightly protuberant hilum, truncated. Based on the symptoms and mycelial and conidial characters above, the fungus was identified as Bipolaris setariae (1). The pathogenicity was established and repeated for six times by following Koch's postulates. Two 1-year-old coconut seedlings were washed with sterilized water and six leaves were wounded with a sterile needle and then inoculated by spraying them with a suspension of conidia of the isolate. The seedlings were kept in two incubators at 25°C for 12 days. Inoculated leaves showed typical symptoms similar to those described above. The pathogen was re-isolated from inoculated leaves. Morphological characteristics were identical to the original isolated fungus. In contrast, the control leaves did not show any symptoms. The genomic DNA of this fungus was extracted, amplification of the internal transcribed spacer (ITS) region was performed with primer ITS1 and ITS4, and the purified PCR product was sequenced (GenBank Accession No. KJ605157). BLASTn analysis revealed 99% sequence similarity with four B. setariae isolates (HE792936.1, JX462256, GU073108.1, and FJ606786.1). Morphologic characters and sequence analysis of the ITS rDNA confirmed that the pathogen was B. setariae. Bipolaris incurvata has been reported causing disease on coconut (2), but B. setariae was not previously reported on coconut. So far, this is the first report of B. setariae caused coconut seedling leaf spot disease in Hainan, China. References: (1) K. C. da Cunha et al. J. Clin. Microbiol. 50:4061, 2012. (2) A. Kamalakannan et al. New Dis. Rep. 12:18, 2005.

scholarly journals First Report of Leaf Spot Disease on Walnut Caused by Colletotrichum fioriniae in China

Plant Disease ◽  
2015 ◽  
Vol 99 (2) ◽  
pp. 289-289 ◽  
Author(s):  
Y. Z. Zhu ◽  
W. J. Liao ◽  
D. X. Zou ◽  
Y. J. Wu ◽  
Y. Zhou
Keyword(s):  
Dna Sequences ◽  
Leaf Spot ◽  
Juglans Regia ◽  
Morphological Characteristics ◽  
Leaf Spot Disease ◽  
Koch’S Postulates ◽  
First Report ◽  
Spot Disease ◽  
Leaf Spots ◽  
Koch's Postulates

In May 2014, a severe leaf spot disease was observed on walnut tree (Juglans regia L.) in Hechi, Guangxi, China. Leaf spots were circular to semicircular in shape, water-soaked, later becoming grayish white in the center with a dark brown margin and bordered by a tan halo. Necrotic lesions were approximately 3 to 4 mm in diameter. Diseased leaves were collected from 10 trees in each of five commercial orchards. The diseased leaves were cut into 5 × 5 mm slices, dipped in 75% ethanol for 30 s, washed three times in sterilized water, sterilized with 0.1% (w/v) HgCl2 for 3 min, and then rinsed five times with sterile distilled water. These slices were placed on potato dextrose agar (PDA), followed by incubating at 28°C for about 3 to 4 days. Fungal isolates were obtained from these diseased tissues, transferred onto PDA plates, and incubated at 28°C. These isolates produced gray aerial mycelium and then became pinkish gray with age. Moreover, the reverse of the colony was pink. The growth rate was 8.21 to 8.41 mm per day (average = 8.29 ± 0.11, n = 3) at 28°C. The colonies produced pale orange conidial masses and were fusiform with acute ends, hyaline, sometimes guttulate, 4.02 to 5.25 × 13.71 to 15.72 μm (average = 4.56 ± 0.31 × 14.87 ± 1.14 μm, n = 25). The morphological characteristics and measurements of this fungal isolate matched the previous descriptions of Colletotrichum fioriniae (Marcelino & Gouli) R.G. Shivas & Y.P. Tan (2). Meanwhile, these characterizations were further confirmed by analysis of the partial sequence of five genes: the internal transcribed spacer (ITS) of the ribosomal DNA, beta-tubulin (β-tub) gene, glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene, chitin synthase 3(CHS-1) gene, and actin (ACT) gene, with universal primers ITS4/ITS5, T1/βt2b, GDF1/GDR1, CHS1-79F/CHS1-354R, and ACT-512F/ACT-783R, respectively (1). BLAST of these DNA sequences using the nucleotide database of GenBank showed a high identify (ITS, 99%; β-tub, 99%; GAPDH, 99%; CHS-1, 99%; and ACT, 100%) with the previously deposited sequences of C. fioriniae (ITS, KF278459.1, NR111747.1; β-tub, AB744079.1, AB690809.1; GAPDH, KF944355.1, KF944354.1; CHS-1, JQ948987.1, JQ949005.1; and ACT, JQ949625.1, JQ949626.1). Koch's postulates were fulfilled by inoculating six healthy 1-year-old walnut trees in July 2014 with maximum and minimum temperatures of 33 and 26°C. The 6-mm mycelial plug, which was cut from the margin of a 5-day-old colony of the fungus on PDA, was placed onto each pin-wounded leaf, ensuring good contact between the mycelium and the wound. Non-colonized PDA plugs were placed onto pin-wounds as negative controls. Following inoculation, both inoculated and control plants were covered with plastic bags. Leaf spots, similar to those on naturally infected plants, were observed on the leaves inoculated with C. fioriniae within 5 days. No symptoms were observed on the negative control leaves. Finally, C. fioriniae was re-isolated from symptomatic leaves; in contrast, no fungus was isolated from the control, which confirmed Koch's postulates. To our knowledge, this is the first report of leaf disease on walnut caused by C. fioriniae. References: (1) L. Cai et al. Fungal Divers. 39:183, 2009. (2) R. G. Shivas and Y. P. Tan. Fungal Divers. 39:111, 2009.

scholarly journals Characterization of Neopestalotiopsis Species Associated with Mango Grey Leaf Spot Disease in Sinaloa, Mexico

Pathogens ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 788
Author(s):  
Saida S. Gerardo-Lugo ◽  
Juan M. Tovar-Pedraza ◽  
Sajeewa S. N. Maharachchikumbura ◽  
Miguel A. Apodaca-Sánchez ◽  
Kamila C. Correia ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Leaf Spot ◽  
Elongation Factor ◽  
Its Region ◽  
Morphological Characterization ◽  
Leaf Spot Disease ◽  
Grey Leaf Spot ◽  
Spot Disease ◽  
Leaf Spots ◽  
Mango Leaves

Mango is one of the most popular and nutritious fruits in the world and Mexico is the world’s largest exporter. There are many diseases that directly affect fruit yield and quality. During the period 2016–2017, leaves with grey leaf spots were collected from 28 commercial mango orchards distributed in two main production areas in Sinaloa State of Mexico, and 50 Neopestalotiopsis isolates were obtained. Fungal identification of 20 representative isolates was performed using morphological characterization and phylogenetic analysis based on the internal transcribed spacer (ITS) region of ribosomal DNA, part of the translation elongation factor 1-alpha (TEF) and the β-tubulin (TUB) genes. Phylogenetic analysis indicated that the 20 isolates from this study formed four consistent groups, however, overall tree topologies do not consistently provide a stable and sufficient resolution. Therefore, even though morphological and phylogenetic separation is evident, these isolates were not assigned to any new taxa and were tentatively placed into four clades (clades A–D). Pathogenicity tests on detached mango leaves of cv. Kent showed that the 20 isolates that belong to the four Neopestalotiopsis clades from this study and induce lesions on mango leaves. This is the first report of species of Neopestalotiopsis causing mango grey leaf spot disease in Mexico.

scholarly journals First Report of Neofusicoccum parvum Causing Panicle Blight and Leaf Spot on Vitis heyneana in China

Plant Disease ◽  
2015 ◽  
Vol 99 (3) ◽  
pp. 417-417 ◽  
Author(s):  
D. D. Wu ◽  
G. Fu ◽  
Y. F. Ye ◽  
F. Y. Hu ◽  
H. F. Mou ◽  
...  
Keyword(s):  
Leaf Spot ◽  
Severe Disease ◽  
Morphological Characteristics ◽  
Aerial Mycelium ◽  
Its Region ◽  
Average Diameter ◽  
Wine Production ◽  
Neofusicoccum Parvum ◽  
First Report ◽  
Panicle Blight

The climbing vine, Vitis heyneana Roem. & Schult, is a member of the grape family endemic to Asia. Its fruits are used in wine production, and its roots, stems, and leaves can be used in medicinal materials. This plant is grown in Southwest China, as well as in India, Bhutan, and Nepal. Mulao Autonomous County in Guangxi Province is the only artificial cultivation area in China. During the summer of 2013, a panicle blight and leaf spot were detected on V. heyneana on four farms in Mulao Autonomous County. The symptoms were observed from the onset of florescence through fruit harvest. Brown lesions initially appeared at the base of a panicle and then extended to the whole panicle, finally causing the panicle to die and fruit to drop. When the disease developed on leaves, the symptom initially appeared as small dark brown circular spots, later enlarging into irregular spots (average diameter 6 mm) with a light brown center and dark brown rim. With severe disease, some individual leaves were affected by numerous spots, leading to premature senescence. Small sections of diseased tissue excised from 10 panicle and 10 leaf samples were plated on potato dextrose agar (PDA) and incubated at 28°C. Fungal colonies developed, initially with abundant white aerial mycelium, which turned olivaceous gray after 5 days and formed black pycnidia after 25 days. The conidia were hyaline, ellipsoidal to fusiform, externally smooth, thin-walled, and nonseptate. Thirty conidia were measured; the dimensions were 12.0 to 17.5 × 4.0 to 6.0 μm. Morphological characteristics of the isolates were similar to the descriptions of Neofusicoccum parvum (3). The isolate MPT-1 was selected as a representative for molecular identification. Genomic DNA was extracted and used for PCR to amplify the internal transcribed spacer (ITS) region and partial translation elongation factor 1-alpha (EF1-α) gene, using primers ITS1/ITS4 and EF1-728F/EF1-986R, respectively. The obtained ITS sequence (GenBank Accession No. KJ599627) and EF1-α sequence (KM921768) showed >99% homology with several GenBank sequences of N. parvum. Morphological and molecular results confirmed the isolate as N. parvum. For pathogenicity tests, detached, young healthy panicles and leaves of V. heyneana were surface-sterilized, wounded by sterile needle, and inoculated with mycelial plugs (3 mm in diameter) of four N. parvum isolates. Ten panicles and 10 leaves were used for every isolate. Control panicles and leaves were treated with sterile PDA plugs. All the samples were placed in a humid chamber (RH 90%, 28°C, 12 h of light) for 3 days. Symptoms similar to those observed in the field developed on all panicles and leaves inoculated with N. parvum isolates. N. parvum was reisolated from all inoculated, symptomatic tissues. The controls remained symptomless. N. parvum has been reported to cause trunk canker on V. vinifera (2), dieback on Cupressus funebris (3), and a leaf spot on Myristica fragrans (1). To our knowledge, this is the first report of N. parvum causing panicle blight and leaf spot on V. heyneana in China. Panicle blight caused a large number of fruits to drop and reduced the yield seriously. Some effective measures should be taken to control this disease. References: (1) V. Jayakumar et al. New Dis. Rep. 23:19, 2011. (2) J. Kaliternam et al. Plant Dis. 97:1656, 2013. (3) S. B. Li et al. Plant Dis. 94:641, 2010.

scholarly journals Alternaria tenuissima Causing Leaf Spot Disease on Amygdalus triloba in Xinjiang of China

Plant Disease ◽  
2020 ◽  
Author(s):  
Xiao Fei Chen ◽  
Yanqiao He ◽  
Aomen He ◽  
Juxia He ◽  
Qiongqiong Li ◽  
...  
Keyword(s):  
Leaf Spot ◽  
Fungal Pathogen ◽  
Morphological Characteristics ◽  
Its Region ◽  
Leaf Spot Disease ◽  
Diseased Plant ◽  
Distilled Water ◽  
Single Spore ◽  
Alternaria Tenuissima ◽  
Spot Disease

Amygdalus triloba (Rosaceae; previously Prunus triloba) is a deciduous, flowering shrub that is widely used in the greening and beautification of lawns, parks and courtyards in China. In late May 2019, a leaf spot disease of A. triloba was observed on approximately 35% of plants in the Xinjiang Alaer city (40˚33′20′′N, 81˚17′19′′E). The disease symptoms began as small, suborbicular, brown spots on the leaves. As the disease progressed, the spots enlarged and coalesced into large necrotic areas and resulted in premature defoliation. Leaf sections (5 x 5 mm) from infected leaves were surface - sterilized with 75% ethanol for 30 s and 0.1% HgCl2 for 1 min, rinsed three times in sterile distilled water and then incubated on potato dextrose agar (PDA). Fifteen fungal isolates showing similar morphological characteristics were obtained by single-spore isolation. On the PDA plates, all fungal colonies had a dark olive color with loose, cottony mycelium. On the potato carrot agar, the fungus formed unbranched spore chains, but occasionally formed one or two lateral branches. Conidiophores were short, hazel-colored, septae, arising singly, and measuring 15.1 to 61.8 × 1.8 to 4.2 µm (35.2 ± 1.4 × 2.3 ± 0.1 µm, n = 50). Mature conidia were ellipsoidal to ovoid with a short conical beak at the tip, light brown with zero to three longitudinal septa and one to five transverse septa, and measuring 19.3 to 30.8 × 7.2 to 12.5 µm (21.8 ± 0.3 × 9.5 ± 0.2 µm, n = 50). Based on the cultural and morphological traits, the pathogen was preliminary identified as Alternaria tenuissima (Simmons 2007). Genomic DNA was extracted from the representative isolate YALAR-1, and the internal transcribed spacer (ITS) region, the partial coding sequence of endopolygalacturonase (endoPG), the glyceradehyde -3- phosphate dehydrogenase (GAPDA), the partial region of the histone 3 (H3) genes were amplified using primers ITS1/ITS4 (White et al. 1990), PG2b/PG3a (Andrew et al. 2009), GDF1/GDR1 (Berbee et al. 1999) and H3-1a/H3-1b (Glass and Donaldson 1995), respectively. The amplicons were sequenced and deposited in GenBank [MT459807 (ITS), MT459808 (endoPG), MT459805 (GAPDA), MT459806 (H3)]. MegaBLAST analyses revealed that our ITS, endoPG, GAPDA, and H3 sequences were 99-100% identical to those of A. tenuissima isolates in GenBank [AF347032 (ITS), KP124026 (endoPG), AY278809 (GAPDA), KF997086 (H3)], confirming the identity of the pathogen as A. tenuissima. Pathogenicity tests were performed by inoculating the fungus onto healthy, mature leaves of A. triloba in the field. Twenty five leaves (five leaves/plant) were sprayed with spore suspensions (1 × 106 spores/ml) of each fungal pathogen, and the same number of leaves were sprayed with distilled water as controls. Inoculated and control leaves were covered with clear plastic bags for 3 days. The experiment was repeated three times. Twelve days after inoculation, the observed symptoms were similar to the original symptoms and the same fungal pathogen was reisolated from the inoculated leaves and identified as A. tenuissima based on morphological features and sequence analysis. The control leaves remained asymptomatic and no fungus was isolated from these leaves. Previously, a leaf spot of A. triloba caused by Alternaria brassicae was reported in Dalian, China (Xie et al. 2017). In order to control this disease effectively, further studies are needed on the biology and ecology of A. tenuissima and A. brassicae respectively. To our knowledge, this is the first report of A. tenuissima associated with leaf spot disease on A. triloba in China. In late September 2020, the diseased plant rate increased to 38% in Alaer city. If the disease control and prevention is neglected, the landscape of Alaer city will be affected seriously. So, in order to effectively control the spread of the disease, it is urgent now to study the sensitivity of pathogen to fungicide and carry out the field efficacy trials. References: Andrew, M., et al. 2009. Mycologia. 101:95. Berbee, M. L., et al. 1999. Mycologia. 91:964. Glass, N. L., and Donaldson, G. C. 1995. Appl. Environ. Microbiol. 61:1323. Simmons, E. G. 2007. Alternaria: An Identification Manual. CBS Fungal Biodiversity Centre, Utrecht, The Netherlands. White, T. J., et al. 1990. PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA. Xie, Y., et al. 2017. Liaoning Agricultural Sciences. 6: 73.

scholarly journals First Report of Leaf Spot Caused by Alternaria alternata on Chinese Dwarf Banana in China

Plant Disease ◽  
2014 ◽  
Vol 98 (5) ◽  
pp. 691-691 ◽  
Author(s):  
B. Z. Fu ◽  
Z. H. Zhang ◽  
L. H. Wang ◽  
G. Y. Li ◽  
J. Z. Zhang ◽  
...  
Keyword(s):  
Alternaria Alternata ◽  
Leaf Spot ◽  
Morphological Characteristics ◽  
Pcr Amplification ◽  
Its Region ◽  
Leaf Spot Disease ◽  
First Report ◽  
Spot Disease ◽  
Rdna Its ◽  
Its Sequence Analysis

The Chinese dwarf banana (Ensete lasiocarpum) is one of the ornamental bananas that belongs to Musaceae family. The plant is native to the southwestern China, where it grows semi-wild in the mountains between 1,500 and 2,500 m above sea level. During July 2011, a leaf spot disease on this plant was observed in the campus and parks in Kunming, Yunnan Province. The incidence level was about 22%, mainly on the old leaves. The leaf symptoms were irregular spots with gray to off-white centers surrounded by dark brown margins, and usually also surrounded by chlorotic halos. Leaf tissues (3 × 5 mm), cut from the margins of lesions, were surface-disinfected (95% ethanol for 3 min, 0.1% HgCl2 for 2 min, rinsed three times with sterile water), plated on potato sucrose agar (PSA), and incubated at 26°C under natural lights. The same fungus was consistently isolated from the diseased leaves. Colonies of white-to-dark gray mycelia formed on PSA that were black on the underside. The colonies were further identified as Alternaria sp. based on the dark brown, obclavate to obpyriform catenulate conidia with longitudinal and transverse septa tapering to a prominent beak attached in chains on a simple and short conidiophore (2). Conidia were 5.26 to 30.26 μm long and 3.95 to 15.79 μm wide, averaging 10.21 (±3.17) × 20.02 (±5.75) μm (n = 50), with a beak length of 0 to 7.89 μm, and had 3 to 8 transverse and 0 to 3 longitudinal septa. PCR amplification was carried out by utilizing universal rDNA-ITS primer pair ITS4/ITS5 (1). The ITS region of isolate DY1 (GenBank Accession No. KF516556) was 572 bp in length. BLAST search revealed 99% identity with two Alternaria alternata isolates (JF440581.1 and GQ121322.2). Phylogenetic analysis (MEGA 5.1) using the neighbor-joining algorithm placed the isolate in a well-supported cluster with other A. alternata isolates. The pathogen was identified as A. alternate (Fr.:Fr.) Keissler based on the morphological characteristics and rDNA-ITS sequence analysis. To confirm pathogenicity, Koch's postulates were performed on detached leaves of E. lasiocarpum inoculated with mycelial plugs with ddH2O and agar plugs as a control. Leaf spots identical to those observed in the field developed in 9 days on the inoculated leaves but not on the control. The inoculation assay used three leaves, totaling 72 spots for control and 36 spots for inoculation. The experiments were repeated once. A. alternata was consistently re-isolated from the inoculated leaves. The symptom developed easier with wounds. To our knowledge, this is the first report of E. lasiocarpum leaf spot disease caused by A. alternata in China and the world. References: (1) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990. (2) T. Y. Zhang. Flora Fungorum Sinicorum, Vol. 16: Alternaria. Science Press, Beijing, China, 2003.

scholarly journals First Report of leaf spot disease caused by Colletotrichum gloeosporioides on Chaenomeles sinensis in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Hang Ni ◽  
Wei-Liang Kong ◽  
Qiao-qiao Zhang ◽  
Xiao-Qin Wu
Keyword(s):  
Leaf Spot ◽  
Economic Value ◽  
Morphological Characteristics ◽  
Bootstrap Support ◽  
Leaf Spot Disease ◽  
Accurate Identification ◽  
First Report ◽  
Spot Disease ◽  
Leaf Spots ◽  
Chaenomeles Sinensis

Chaenomeles sinensis is a shrub or small arbor of the genus Chaenomeles in Rosaceae, which is widely planted in China. It is a kind of garden ornamental plant and has high economic value. Since 2020, a leaf disease occurred on the foliage of C. sinensis at the campus of Nanjing Forestry University, Nanjing, China. After investigating, C. sinensis was found with leaf spot disease at a 100% infection rate, which causing gigantic ornamental loss. Leaf spots are round to irregular distributing on the leaves, in addition, the color of spots is brown. There are yellow halos on the edge of the lesion. Small leaf tissues (3 to 4 mm2) from lesion margins were surface sterilized with 75% ethanol for 30s and then rinsed with sterile dH2O for three times. Afterwards, placed on potato dextrose agar (PDA) at 25°C. Pure cultures were obtained by monosporic isolation, and a representative isolate (NJTJ.1) was obtained. When cultured on PDA, the colony of NJTJ.1 was white and cottony. On the reverse side, the color of colony nearly light yellow. The colony were placed in the liquid Carboxymethyl cellulose (CMC) medium. After culturing for 24h in a shaker at 25℃ and 150rmp/min, the spore liquid was taken by us. The conidia were one-celled, straight, hyaline, subcylindrical with rounded ends and measured 15.1 to 23.6× 5.4 to 7.9 µm (n =30). Appressoria were one-celled, brown, thick-walled, ellipsoidal, and measured 7.7 to 13.8 × 6.4 to 10.3 µm (n =30). The morphological characteristics of NJTJ.1 fitted with the description of the Colletotrichhum gloeosporioides complex (Weir et al., 2012). For accurate identification, the internal transcribed spacer (ITS), and the genes encoding glyceraldehyde-3-phosphate dehydrogenase (GAPDH), actin (ACT) and chitin synthase (CHS-1) were amplified with primers ITS1/ITS4, GDF/GDR, ACT-512F/ACT-783R, and CHS-79F/CHS-345R (Zhu et al, 2019), respectively. The sequences were deposited in GenBank [Accession Nos.MT984264, MW030495 and MW030496 to MW030497 for NJTJ.1]. A Blast search of GenBank showed that ITS, GAPDH, ACT and CHS-1 sequences of NJTJ.1 were 99%, 99%, 100% and 100% identical to those of C. gloeosporioides (MH571757.1 ,KY995355.1 , MN058143.1 and MN313581.1). A neighbor-joining phylogenetic tree was generated by combining all sequenced loci in MEGA7. The isolate NJTJ.1 clustered in the C. gloeosporioides clade with 99% bootstrap support. The pathogenicity of the NJTJ.1 was verified both on detached and living leaves. The detached leaves were inoculated with 5-mm mycelial plugs cut from the edge of 6-day old cultures on PDA and 20 μL of spore suspension (106 conidia/mL) and each treatment had 5 replicates. Controls were treated with sterile dH2O. The inocula were placed at a distance of 2 to 3 cm on the leaves which were wounded with a sterile needle. All of them were placed in 20-cm dishes on wet filter paper at 25°C. After 5 days, all the inoculated points showed lesions which were similar to those outdoor observed. Whereas, controls were asymptomatic.At the same time, the plugs of C. gloeosporioides were inoculated on living leaves.After 7 days, the leaves which were inoculated also appeared lesions. This is the first report of C. gloeosporioides causing leaf blotch on Chaenomeles sinensis in China. These data will help develop effective strategies for managing this disease.

scholarly journals First Report of Leaf Spot Caused by Pseudocercospora subsessilis on Melia azedarach in Korea

Plant Disease ◽  
2013 ◽  
Vol 97 (7) ◽  
pp. 993-993
Author(s):  
S. T. Seo ◽  
C. H. Shin ◽  
J. H. Park ◽  
H. D. Shin
Keyword(s):  
Leaf Spot ◽  
Morphological Characteristics ◽  
Its Region ◽  
The United States ◽  
Melia Azedarach ◽  
Leaf Spot Disease ◽  
Limiting Factor ◽  
First Report ◽  
Leaf Spots ◽  
White Leaf

Melia azedarach L., called chinaberry, is native to Southeast Asia and Australia. The trees are commonly planted as ornamentals in the southern part of Korea. In October 2010, a leaf spot disease was observed on trees for the first time in Wando, Korea. Further surveys conducted from 2010 to 2012 showed that the disease occurs on trees in Jeju, Seogwipo, and Tongyeong cities as well as Wando county with nearly 100% incidence. Leaf spots were circular to semicircular, later becoming angular, small, pale brown in the center with a dark brown margin, and later becoming milky white. Leaf spots sometimes coalesced to blight the entire leaf and were capable of rapidly defoliating whole trees in late September. Fruiting was amphigenous, but mostly hypogenous. Stromata were substomatal, globular, dark brown, and 25 to 70 μm in diameter. Conidiophores were densely fasciculate, pale olivaceous to pale brown, substraight to mildly curved, not geniculate, 10 to 30 μm long, 2.5 to 4.5 μm wide, and aseptate or uniseptate. Conidia were pale olivaceous, generally darker than conidiophores, cylindric to obclavate, substraight in shorter ones, curved to mildly sinuous in longer ones, obconically truncate at the base, obtuse at the apex, 2- to 14-septate, 16 to 120 × 3 to 5 μm, guttulate, and had inconspicuous hila. Morphological characteristics of the fungus were consistent with the previous descriptions of Pseudocercospora subsessilis (Syd. & P. Syd.) Deighton (2). Voucher specimens (n = 6) were deposited in the Korea University Herbarium (KUS). An isolate from KUS-F25395 was deposited in the Korean Agricultural Culture Collection (KACC45688). The complete internal transcribed spacer (ITS) region of rDNA was amplified with the primers ITS1/ITS4 (3) and sequenced. The resulting sequence of 517 bp was deposited in GenBank (Accession No. JX993904). A BLAST search in GenBank revealed that the sequence shows >99% similarity (1 bp substitution) with a sequence of P. subsessilis ex M. azedarach from Cuba (GU269815). For pathogenicity tests, hyphal suspensions were prepared by grinding 3-week-old colonies grown on potato dextrose agar with distilled water using a mortar and pestle. Five 3-year-old chinaberry trees were inoculated with hyphal suspensions using a fine haired paint brush. Three healthy trees of the same age, serving as controls, were sprayed with sterile water. The plants were covered with plastic bags to maintain 100% relative humidity for 24 h and then transferred to a greenhouse. Typical symptoms of necrotic spots that appeared on the inoculated leaves 10 days after inoculation were identical to the ones observed in the field. P. subsessilis was reisolated from symptomatic leaf tissues, confirming Koch's postulates. No symptoms were observed on control plants. The disease has been reported in several Asian countries as well as in Cuba and the United States (1). To our knowledge, this is the first report of leaf spot on chinaberry caused by P. subsessilis in Korea. The observed high incidence and severity suggest that this disease can be a limiting factor in utilizing this tree species as ornamentals in public areas. References: (1) D. F. Farr and A. Y. Rossman. Fungal Databases. Syst. Mycol. Microbiol. Lab., Online publication, ARS, USDA, Retrieved October 22, 2012. (2) Y. L. Guo and W. H. Hsieh. The genus Pseudocercospora in China. International Academic Publishers, Beijing, China, 1995. (3) T. J. White et al. PCR Protocols. Academic Press, San Diego, CA, 1990.

scholarly journals COMBRETUM INDICUM – A NEW HOST RECORD OF ALTERNARIA BRASSICAE LEAF SPOT DISEASE FROM PAKISTAN

2017 ◽  
Vol 29 (1) ◽  
pp. 171
Author(s):  
Naureen Akhtar ◽  
Rahila Hafeez ◽  
Waheed Anwar
Keyword(s):  
Leaf Spot ◽  
Host Record ◽  
Its Region ◽  
First Record ◽  
Nucleotide Sequence Analysis ◽  
Leaf Spot Disease ◽  
Alternaria Brassicae ◽  
New Host ◽  
Causal Organism ◽  
Spot Disease

Combretum indicum leaf spot disease was observed during a survey of horticultural plants in Lahore. Isolation of causal organism was carried out from the disease leaves. The pathogen, Alternaria brassicae, was first identified considering morphological characters. The morphology based identification was confirmed by nucleotide sequence analysis of ITS region of rDNA and phylogentic analysis of this fungus with closely related other fungal species. Pathogenicity of A. brassicae was verified following the Koch’s pathogenicity postulates. This report represents the first record of leaf spot of Combretum indicum by Alternaria brassicae from Pakistan.

scholarly journals The First Report of Calonectria Pteridis Causing a Leaf Spot Disease on Serenoa repens in China

Plant Disease ◽  
2014 ◽  
Vol 98 (6) ◽  
pp. 854-854
Author(s):  
W. Yang ◽  
L. Zheng ◽  
C. Wang ◽  
C.-P. Xie
Keyword(s):  
Leaf Spot ◽  
Morphological Characteristics ◽  
Its Region ◽  
Leaf Spot Disease ◽  
Fluorescent Light ◽  
Brown Pigment ◽  
Morphological Identification ◽  
Serenoa Repens ◽  
First Report ◽  
Spot Disease

Serenoa repens [(Bartr) J. K. Small] is an important medicinal plant with their extracts is one of the three most effective drugs to cure benign prostatic hyperplasia (BPH). Also it can be used as an ornamental plant for garden. In November 2010, a new leaf spot disease was found on S. repens in Danzhou, Hainan Province, China. Disease occurred very seriously, with the incidence close or up to 100%, even leading to plant drying and death. Initially, the leaves had circular water-soaked dots, and had an obvious yellow halo on the edge, then expanded into oval, circular, or irregular shaped spots. Eventually the spot was beige and gray in the center and dark brown and slightly concave on the edge. The pathogen was isolated following the method reported by Fang (3) and prepared for further characterization. On potato dextrose agar (PDA) medium, the pathogen formed round and red-brown colonies with neat edges of a sandy beige color. A white powdery substance was formed on the surface of the colony, and it produced reddish-brown pigment on the back. On carnation leaf agar (CLA), only large macroconidium was observed. Macroconidiophores containing a stipe bearing penicillate suites of fertile branches, terminating in a clavate vesicle (5.9-) 6.4 (-6.9) × (33.8-) 39.6 (-46.7) μm. Conidiogenous apparatus had primary branches aseptate or rarely 1-septate and were (21.8-) 28.7 (-38.6) μm long, secondary branches were aseptate and (18.8-) 29.9 (-39.9) μm long, and tertiary branches were aseptate and (14.2-) 17.4 (-19.9) μm long. Macroconidium and microconidium were observed on water agar (WA) at 30 days. Macroconidium was colorless, cylindrical, rounded at both ends, 1 to 3 hyaline septate, but mainly one, and (4.5-) 5.2 (-6.2) × (71.3-) 84.1 (-98.0) μm; microconidium was colorless, cylindrical, both ends obtuse, curved or straight, 1-septate, and (24.8-) 33.2 (-45.2) × (2.5-) 3.5 (-5.0) μm. It could produce microsclerotia on PDA, CLA, and WA media. Morphological characteristics of the specimen examined were similar to Calonectria pteridis. In the genus of Calonectria, only C. pteridis could produce bending microconidium on WA medium (2). To confirm the morphological identification, primer pair ITS1/ITS4 were used for amplification of the ITS region of rDNA. Its sequence (GenBank Accession No. KF994926) showed 99% identity with C. pteridis Crous, M.J. Wingf. & Alfenas. (GQ280617.1). In addition, the translation elongation factor 1-alpha gene sequence was amplified (KF994927) and it showed 100% identify with C. pteridis (FJ918564.1) (1). Thus, the pathogen was identified as C. pteridis. To confirm pathogenicity, conidial suspensions (105 conidia ml−1) of the pathogen were inoculated with healthy leaves of 10 plants by pinprick inoculation method. Control plants were inoculated with water. Plants were maintained at 28°C in a greenhouse with constant humidity (RH 90%) and a 12-h photoperiod of fluorescent light. Symptoms similar to the original ones appeared after 7 days, while the control plants remained healthy. The tests were repeated three times and the pathogen was re-isolated from the leaves of inoculated plants and confirmed to be C. pteridis by both morphology and molecular characterization. To our knowledge, this is the first report of leaf spot caused by C. pteridis on S. repens in China. References: (1) I. Carbone and L. M. Kohn. Mycologia 91:553, 1999. (2) P. W. Crous and M. J. Wingfield. Mycotaxon 51:341, 1994. (3) Z. D. Fang. Plant Disease Research Methods, 3rd edition. China Agriculture Press, Beijing, 1998.

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