scholarly journals First Report of Leaf Spot Disease in Withania coagulans Caused by Alternaria alternata in India

Plant Disease ◽  
2013 ◽  
Vol 97 (3) ◽  
pp. 420-420
Author(s):  
A. Sharma ◽  
V. Singh ◽  
G. Singh ◽  
P. K. Pati
Keyword(s):  
Medicinal Plant ◽  
Alternaria Alternata ◽  
Leaf Spot ◽  
Management Program ◽  
Leaf Spot Disease ◽  
Post Inoculation ◽  
Isolation Technique ◽  
Leaf Spots ◽  
Withania Coagulans ◽  
Initial Symptoms

Withania coagulans (Paneer doddi) is a medicinal plant in the Solanaceae (1) that grows in northwestern India. An unknown disease appeared in the Amritsar district of Punjab, India on W. coagulans starting in 2009, and was noticed annually in the post-monsoon seasons through 2011. The plants were grown in a net house (74.82323 to 74.82332°E, 31.63678 to 31.63688°N, 221 m elevation), Guru Nanak Dev University, Amritsar. Symptoms first appeared as brown to black spots (2 to 10 mm in diameter) that were sometimes surrounded by a yellow halo, starting on the lower leaves and gradually spreading to the upper leaves. Spots were apparent on both the dorsal and ventral sides of the leaves. In the most severe outbreak of the disease, more than 90% of the leaf area was covered with the leaf spot symptoms. The pathogen was isolated by placing pieces of surface-sterilized leaf spots on potato dextrose agar (PDA), and the hyphae that emerged from the leaf pieces were sub-cultured to PDA. Initially, the fungal colony was white, and gradually turned olive-brown as it matured. Conidiophores were mostly simple, usually becoming geniculate by sympodial elongation. Conidiophores were mostly branched and conidia were arranged in acropetal chains of 6 to 10. Conidia were 15 to 25 μm long and 5 to 10 μm wide, with 3 to 4 transverse septa and 2 to 3 longitudinal septa, muriform, ovoid to ellipsoid, with a broadly rounded base and an apical beak. Based on these morphological characters, the pathogen was identified as Alternaria alternata (Fr.) Keissler (4). The identification of the pathogen was also confirmed by MTCC, Chandigarh, India and deposited in their culture collection with Accession No. MTCC-10939 ( http://mtcc.imtech.res.in ). During preliminary studies, the optimum temperature for its growth was 25 ± 2°C. Pathogenicity of the fungus was demonstrated on three potted W. coagulans plants after spray-inoculating leaves with a spore suspension of 3 × 105 spores per ml. A batch of three plants sprayed with sterile distilled water served as controls. Both inoculated and control plants were incubated at 100% relative humidity for 5 days and transferred to a glasshouse at 25 ± 2°C. Initial symptoms developed on a few plants at 5 to 7 days post-inoculation, and the same leaf symptoms described above from field plants became apparent on all inoculated plants by 10 to 14 days. Control plants did not develop symptoms. The pathogen was recovered from leaf spots using the isolation technique described above, and was identified as A. alternata, thus fulfilling Koch's postulates. Although the leaf spot diseases caused by A. alternata and A. dianthicola were reported on W. somnifera (2,3), to the best of our knowledge, there is no report of either pathogen infecting W. coagulans. Thus, the identification of pathogen will facilitate the disease management program and for enhancing the commercial value of this important endangered medicinal plant. References: (1) S. A. Gilani et al. Afr. J. Biotechnol. 8:2948, 2009. (2) C. K. Maiti et al. Plant Dis. 91:467, 2007. (3) P. K. Pati et al. Indian J Microbiol. 48:432, 2008. (4) E. G. Simmons. Alternaria: An identification manual. American Phytopathological Society, St. Paul, MN, 2007.

scholarly journals First Report of a Leaf Spot Disease of Bells-of-Ireland (Moluccella laevis) Caused by Cercospora apii in California

Plant Disease ◽  
2003 ◽  
Vol 87 (2) ◽  
pp. 203-203
Author(s):  
S. T. Koike ◽  
S. A. Tjosvold ◽  
J. Z. Groenewald ◽  
P. W. Crous
Keyword(s):  
Leaf Spot ◽  
Sequence Data ◽  
Disease Incidence ◽  
Leaf Spot Disease ◽  
Conidial Suspension ◽  
Internal Transcribed Spacer Region ◽  
First Report ◽  
Spot Disease ◽  
Leaf Spots ◽  
Initial Symptoms

Bells-of-Ireland (Moluccella laevis) (Lamiaceae) is an annual plant that is field planted in coastal California (Santa Cruz County) for commercial cutflower production. In 2001, a new leaf spot disease was found in these commercially grown cutflowers. The disease was most serious in the winter-grown crops in 2001 and 2002, with a few plantings having as much as 100% disease incidence. All other plantings that were surveyed during this time had at least 50% disease. Initial symptoms consisted of gray-green leaf spots. Spots were generally oval in shape, often delimited by the major leaf veins, and later turned tan. Lesions were apparent on both adaxial and abaxial sides of the leaves. A cercosporoid fungus having fasciculate conidiophores, which formed primarily on the abaxial leaf surface, was consistently associated with the spots. Based on morphology and its host, this fungus was initially considered to be Cercospora molucellae Bremer & Petr., which was previously reported on leaves of M. laevis in Turkey (1). However, sequence data obtained from the internal transcribed spacer region (ITS1, ITS2) and the 5.8S gene (STE-U 5110, 5111; GenBank Accession Nos. AY156918 and AY156919) indicated there were no base pair differences between the bells-of-Ireland isolates from California, our own reference isolates of C. apii, as well as GenBank sequences deposited as C. apii. Based on these data, the fungus was subsequently identified as C. apii sensu lato. Pathogenicity was confirmed by spraying a conidial suspension (1.0 × 105 conidia/ml) on leaves of potted bells-of-Ireland plants, incubating the plants in a dew chamber for 24 h, and maintaining them in a greenhouse (23 to 25°C). After 2 weeks, all inoculated plants developed leaf spots that were identical to those observed in the field. C. apii was again associated with all leaf spots. Control plants, which were treated with water, did not develop any symptoms. The test was repeated and the results were similar. To our knowledge this is the first report of C. apii as a pathogen of bells-of-Ireland in California. Reference: (1) C. Chupp. A Monograph of the Fungus Genus Cercospora. Cornell University Press, Ithaca, New York, 1954.

scholarly journals First Report of Leaf Spot Caused by Alternaria alternata on Marigold (Tagetes erecta) in Beijing, China

Plant Disease ◽  
2014 ◽  
Vol 98 (8) ◽  
pp. 1153-1153 ◽  
Author(s):  
Y. Li ◽  
J. Shen ◽  
B. H. Pan ◽  
M. X. Guo ◽  
Q. X. Wang ◽  
...  
Keyword(s):  
Alternaria Alternata ◽  
Leaf Spot ◽  
Leaf Spot Disease ◽  
Tagetes Erecta ◽  
Pathogenicity Test ◽  
First Report ◽  
Leaf Spots ◽  
Commercial Crop ◽  
Necrotic Spots ◽  
Beijing China

Marigold (Tagetes erecta) is an important commercial crop and 200 ha are planted every year in the Beijing district of China. A leaf spot disease of T. erecta was observed during 2012 and 2013 in the Beijing district. The disease was widespread, with 60 to 75% of the fields affected. Leaves of the affected plants had small, brown, necrotic spots on most of the foliage. Yield losses of flowers of up to 20 to 30% were reported. The spots gradually enlarged, becoming irregular in shape, or remained circular, and with concentric rings or zones. In the later stages of infection, the spots coalesced, and the leaves withered, dried, and fell from the plants (4). A fungus was consistently isolated on potato dextrose agar (PDA) from the infected leaves of T. erecta. After 6 days of incubation at 26°C and a 12-h photoperiod, the fungus produced colonies that were flat, with a rough upper surface (2). The conidiophores were short. Conidia varied from 18 × 6 to 47 × 15 μm and were medium to dark brown or olive-brown in color, short beaked, borne in long chains, oval and bean shaped, with 1 to 5 transverse septa and 0 to 2 longitudinal septa. The rDNA of the internal transcribed spacer regions 1 and 2 and the 5.8S gene in seven isolates were amplified using primers ITS1 (5′-TCCGTAGGTGAACCTGCGG-3′) and ITS4 (5′-TCCTCCGCTTATTGATATGC-3′). The nucleotide sequence was the same as isolate No. 7, which was deposited in GenBank (Accession No. KF307207). A BLAST search showed 97% identity with the strain Alternaria alternata GNU-F10 (KC752593). Seven isolates were also confirmed as A. alternata by PCR identification performed by specific primers (C_for/C_rev) of A. alternata (1). Seven isolates were grown on PDA for 2 weeks and the conidia harvested. A 5-μl drop of spore suspension (1 × 105 spores/ml) was placed on each leaflet of 140 detached, surface-sterilized T. erecta leaves. Twenty leaves were inoculated with sterile distilled water as a control. The leaves were incubated in a growth chamber at 80 to 90% relative humidity, 50 to 60 klx/m2 light intensity, and a 12-h photoperiod. After 6 days, leaf spots similar to the original developed at inoculation sites for all isolates and A. alternata was consistently re-isolated. The control leaves remained symptomless. The pathogenicity test was performed three times. Leaf spot of T. erecta caused by Alternaria spp. is well known in Asian countries such as Japan (3). To our knowledge, this is the first report of A. alternata on T. erecta in the Beijing district of China. References: (1) T. Gat. Plant Dis. 96:1513, 2012. (2) E. Mirkova. J. Phytopathol. 151:323, 2003. (3) K. Tomioka. J. Gen. Plant Pathol. 66:294, 2000. (4) T. Y. Zhang. Page 284 in: Flora Fungorum Sinicorum, Volume 16: Alternaria. Science Press, Beijing, 2003.

scholarly journals First Report of Alternaria alternata Causing Leaf Spots of Tea (Camellia sinensis) in China

Plant Disease ◽  
2014 ◽  
Vol 98 (5) ◽  
pp. 697-697 ◽  
Author(s):  
L. X. Zhou ◽  
W. X. Xu
Keyword(s):  
Camellia Sinensis ◽  
Alternaria Alternata ◽  
Plant Diseases ◽  
Unknown Etiology ◽  
Leaf Spot Disease ◽  
Post Inoculation ◽  
Tea Leaves ◽  
First Report ◽  
Leaf Spots ◽  
Young Leaves

Tea is the most popular non-alcoholic beverage crop in the world, which originated in China and has been cultivated in over 45 countries. In recent years, a leaf spot disease of unknown etiology has been observed on young leaves of tea trees (Camellia sinensis) grown in Luotian county, Hubei Province, China. Observed symptoms display grayish brown to white spots (about 1 cm in diameter) surrounded by brown edges. Over 20% of the young leaves were affected on surveyed trees. To identify the pathogen, six symptomatic tea leaves were collected from six individual tea trees of unknown variety in August 2012. A thin section (3 to 5 mm) of symptomatic tissue was sterilized in a bleach solution of 3% hypochlorite and placed on potato dextrose agar (PDA) medium at 25°C in darkness for isolation. Six fungal colonies displaying gray-brown and gray-white aerial mycelia were consistently recovered from lesions of the six leaves, termed as T1 to T6, respectively. Conidia produced on the colonies were olive brown, obpyriform, short conical beak at the tip, 0 to 3 vertical and 1 to 6 transverse septa, and length × width of 7.1 to 31.7 (avg. 20.1) × 2.9 to 12.7 (avg. 7.2) μm. T1 to T6 were identified as Alternaria alternata on the basis of morphological characterization, respectively (2). Confirmation of the species identification was obtained by molecular characterization of their internal transcribed spacer (ITS) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) regions amplified from the genomic DNAs using the universal primers (1). The results revealed identical sequences of ITS (GenBank Accession No. KF699530) and GAPDH among the six isolates. BLAST searches showed that they had the highest similarity with A. alternata strains, with 98.3% for ITS (AJ276055) and 96.2% for GAPDH (EF513205), deposited in fungus database ( http://www.mycobank.org/ ). Pathogenicity tests were conducted on the detached leaves expanding for 10 to 20 days of two tea varieties (cvs. Fudingdabai and Taicha No. 12) in triplicate by placing 4 mm diameter discs from 5-day-old PDA plates of T3 and T6, which were incubated in an incubator at 25°C with a 12-h photoperiod for 7 days. All inoculated leaves with or without wound treatment developed brown spots similar to the original ones at 7 days post inoculation (dpi) while the control leaves inoculated with non-colonized PDA plugs remained asymptomatic. Isolates recovered from diseased samples were of the same morphology and ITS sequence as the inoculated ones. Alternaria alternata had been described on C. sinensis in China (3), but it was only reported as a severe foliar fungal pathogen of tea in North Bengal, India (1), and to our knowledge, this is the first report of A. alternata causing leaf spots on tea leaves (C. sinensis) in China. In addition to quantity loss, the species may result in a decrease of quality of tea crop considering that it can produce Alternaria toxins related to animal and public health. The etiologic identification of the disease is expected to provide useful information for its control. References: (1) B. N. Chakraborty et al. Plant Pathol. 55:303, 2006. (2) E. G. Simmons. Page 1 in: Alternaria Biology, Plant Diseases and Metabolites. J. Chelchowski and A. Visconti, eds. Elsevier, Amsterdam, 1992. (3) F. L. Tai. Page 1527 in: Sylloge Fungorum Sinicorum. eds. Sci. Press Acad. Sin. Beijing, 1979. (4) B. S. Weir et al. Stud. Mycol. 73:115, 2012.

scholarly journals Identification of Fusarium ipomoeae as the causative agent of leaf spot disease in Bletilla striata in China

Plant Disease ◽  
2020 ◽  
Author(s):  
Lv-Yin Zhou ◽  
Shuang-Feng Yang ◽  
Shao-Mei Wang ◽  
Jing-Wen Lv ◽  
Wei Qiang Wan ◽  
...  
Keyword(s):  
Leaf Spot ◽  
Morphological Characteristics ◽  
Leaf Spot Disease ◽  
Molecular Characteristics ◽  
Post Inoculation ◽  
Sodium Hypochlorite Solution ◽  
Transparent Plastic ◽  
Bletilla Striata ◽  
Spot Disease ◽  
Initial Symptoms

Bletilla striata (Thunb.) Rchb. f. (Orchidaceae) is traditionally used for hemostasis and detumescence in China. In April 2019, a leaf spot disease on B. striata was observed in plant nurseries in Guilin, Guangxi Province, China, with an estimated incidence of ~30%. Initial symptoms include the appearance of circular or irregular brown spots on leaf surfaces, which progressively expand into large, dark brown, necrotic areas. As lesions coalesce, large areas of the leaf die, ultimately resulting in abscission. To isolate the pathogen, representative samples exhibiting symptoms were collected, leaf tissues (5 × 5 mm) were cut from the junction of diseased and healthy tissue, surface-disinfected in 1% sodium hypochlorite solution for 2 min, rinsed three times in sterile water, plated on potato dextrose agar (PDA) medium, and incubated at 28°C (12-h light-dark cycle) for 3 days. Hyphal tips from recently germinated spores were transferred to PDA to obtain pure cultures. Nine fungal isolates with similar morphological characteristics were obtained. Colonies on PDA were villose, had a dense growth of aerial mycelia and appeared pinkish white from above and greyish orange at the center and pinkish-white at the margin on the underside. Macroconidia were smooth, and hyaline, with a dorsiventral curvature, hooked to tapering apical cells, and 3- to 5-septate. Three-septate macroconidia were 21.2 to 32.1 × 2.4 to 3.9 μm (mean ± SD: 26.9 ± 2.5 × 3.2 ± 0.4 μm, n = 30); 4-septate macroconidia were 29.5 to 38.9 × 3.0 to 4.3 μm (mean ± SD: 33.5 ± 2.6 × 3.6 ± 0.3 μm, n = 40); and 5-septate macroconidia were 39.3 to 55.6 × 4.0 to 5.4 μm (mean ± SD: 48.0 ± 3.9 × 4.5 ± 0.3 μm, n = 50). These morphological characteristics were consistent with F. ipomoeae, a member of the Fusarium incarnatum-equiseti species complex (FIESC) (Wang et al. 2019). To confirm the fungal isolate’s identification, the genomic DNA of the single-spore isolate BJ-22.3 was extracted using the CTAB method (Guo et al. 2000). The internal transcribed space (ITS) region of rDNA, translation elongation factor-1 alpha (TEF-1α), and partial RNA polymerase second largest subunit (RPB2) were amplified using primer pairs [ITS1/ITS4 (White et al. 1990), EF-1/EF-2 (O’Donnell et al. 1998), and 5f2/11ar (Liu, Whelen et al. 1999, Reeb, Lutzoni et al. 2004), respectively]. The ITS (MT939248), TEF-1α (MT946880), and RPB2 (MT946881) sequences of the BJ-22.3 isolate were deposited in GenBank. BLASTN analysis of these sequences showed over 99% nucleotide sequence identity with members of the FIESC: the ITS sequence showed 99.6% identity (544/546 bp) to F. lacertarum strain NRRL 20423 (GQ505682); the TEF-1α sequence showed 99.4% similarity (673/677 bp) to F. ipomoeae strain NRRL 43637 (GQ505664); and the RPB2 sequence showed 99.6% identity (1883/1901 bp) to F. equiseti strain GZUA.1657 (MG839492). Phylogenetic analysis using concatenated sequences of ITS, TEF-1α, and RPB2 showed that BJ-22.3 clustered monophyletically with strains of F. ipomoeae. Therefore, based on morphological and molecular characteristics, the isolate BJ-22.3 was identified as F. ipomoeae. To verify the F. ipomoeae isolate’s pathogenicity, nine 1.5-year-old B. striata plants were inoculated with three 5 × 5 mm mycelial discs of strain BJ-22.3 from 4-day-old PDA cultures. Additionally, three control plants were inoculated with sterile PDA discs. The experiments were replicated three times. All plants were enclosed in transparent plastic bags and incubated in a greenhouse at 26°C for 14 days. Four days post-inoculation, leaf spot symptoms appeared on the inoculated leaves, while no symptoms were observed in control plants. Finally, F. ipomoeae was consistently re-isolated from leaf lesions from the infected plants. To our knowledge, this is the first report of F. ipomoeae causing leaf spot disease on B. striata in China. The spread of this disease might pose a serious threat to the production of B. striata. Growers should implement disease management to minimize the risks posed by this pathogen.

scholarly journals First Report of Leaf Spot Disease on Cinnamomum camphora (Camphor tree) Caused by Epicoccum poaceicola in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Da Li ◽  
Tianning Zhang ◽  
Qingni Song ◽  
Jun Liu ◽  
Haiyan Zhang ◽  
...  
Keyword(s):  
Leaf Spot ◽  
Leaf Growth ◽  
Leaf Spot Disease ◽  
Molecular Characteristics ◽  
Post Inoculation ◽  
Cinnamomum Camphora ◽  
First Report ◽  
Spot Disease ◽  
Leaf Spots ◽  
Camphor Tree

As an important industrial, pharmaceutical and evergreen shade tree (Singh and Jawaid 2012), the camphor tree (Cinnamomum camphora) has been coppiced in Jiangxi Province, China. From 2017 to 2020, we noticed many camphor trees with leaf spots, with an incidence estimated at 50 to 75%, which could severely inhibit leaf growth and reduce their biomass. A dark-green circle with a watery spot appeared on the infected leaves at the initial stage, and necrosis with forming shot-spots surrounded by yellow halos occurred (Figure 1 A). Five leaves with typical symptoms were sampled and washed with tap water for ca. 15 min. Isolation and morphological analysis were performed following the method of Bao et al. (2019). Among 61 fungal isolates, 49 showed the same culture characters. Colonies on PDA were villose and regular, the reverse was scarlet at the edge of the colony, which was ca. 8.75 cm after 7 days of inoculation (Figure 1 I). Chlamydospores were aseptate, dark brown, smooth, in chains or solitary, ellipsoidal to ovoid, 4.8–9.6 × 4.8–11.1 μm (Figure 1 J). The pycnidia were produced on PDA and varied from 47.4 to 85.8 µm (mean 60.2 µm) × 38.6 to 66.8 μm (mean 49.7 μm) (n = 17) (Figure 1 K). Conidia were hyaline, unicellular, elliptical to ovoid, 4.3-6.4 µm (mean 5.1 µm) × 2.3-3.3 µm (mean 2.8 µm) (n = 52) (Figure 1 L). Pathogenicity tests of isolate XW-9 was carried out in the field. Ten leaves were wounded with a sterilized insect needle and inoculated with mycelium plugs (7-mm diameter). Non-colonized PDA plugs served as the negative controlIn addition, conidial suspensions (105 conidia/mL) of isolate XW-9 were sprayed on surface-sterilized leaves with a further ten leaves being sprayed with sterile water as the control. Symptoms described in this study appeared in 100% of the mycelium-inoculated leaves and more than 80% of the conidium-inoculated leaves after 7 days post-inoculation (Figure 1 B-E). No symptoms were seen in the controls (Figure 1 C). Three days after inoculation, brown spots resembling those observed in the field developed on the inoculated leaves, and some lesions turned into shot holes on the infected leaves (Figure 1 G & H). However, no symptoms were observed on the controls (Figure 1 F). The fungus was re-isolated from the margins of the leaf spots and labelled P-XW-9A. The gene regions for ITS, LSU, tub2, RPB2 and ACT of isolates XW-9 and P-XW-9A were amplified and sequenced. The sequences of rDNA-ITS, LSU, tub2, RPB2 and ACT of XW-9 were GenBank MW142397, MW130844, MW165322, MW446945 and MW165324, respectively and those of P-XW-9A were GenBank MW142398, MW130845, MW165323, MW446946 and MW165325, respectively (Lumbsch, et al. 2000; Aveskamp, et al. 2009; Hou et al. 2020). Phylogenetic analysis using concatenated sequences of ITS, LSU, RPB2, and tub2 showed that isolates XW-9 and P-XW-9A formed a single clade with the reference strain of E. poaceicola CBS 987.95 (Figure 2). Thus, XW-9 was identified as E. poaceicola based on its morphological and molecular characteristics. Significantly, the recovered isolate P-XW-9A also aligned with E. poaceicola fulfilling the criteria for Koch's Postulates. E. poaceicola was only reported as a fungal pathogen of Phyllostachys viridis in China (Liu et al. 2020). To our knowledge, this is the first report of leaf spot disease on camphor trees caused by E. poaceicola in China and our findings will be useful for its management.

scholarly journals First Report of Zonate Leaf Spot of Vigna vexillata var. tsusimensis Caused by Cristulariella moricola

Plant Disease ◽  
2002 ◽  
Vol 86 (4) ◽  
pp. 440-440 ◽  
Author(s):  
H. B. Lee ◽  
C.-J. Kim
Keyword(s):  
Leaf Spot ◽  
Morphological Characteristics ◽  
Leaf Spot Disease ◽  
Eastern United States ◽  
Leaf Spots ◽  
Vigna Vexillata ◽  
Initial Symptoms ◽  
Pathogenicity Tests ◽  
Basal Septum ◽  
Bull's Eye

A zonate leaf spot disease on a wild bean variety, Vigna vexillata L. var. tsusimensis Mat., occurred in the patch fields and foothills of Chungnam and Kyonggi districts in Korea during late September, October, and early November of 1999 to 2001. The zonate lesions were particularly prevalent in October following periods of heavy dew accumulation. Initial symptoms were small, circular lesions with darkbrown marginal rings that later developed into large spots with characteristic target-shaped rings. The spots were gray to bright or blackish brown, progressed rapidly, and sometimes fused together to form lesions of up to 20 mm in diameter. Sporophores on the natural host were generally hypophyllous but sometimes amphigenous, abundant on large spots, fewer on small spots, solitary, erect, easily detachable, and up to 864 μm long. The upper portion of the sporophore is considered an individual conidium and consisted of a pyramidal head that was fusiform to ventricose and cristulate, 495 to 534 μm long, and 210 to 290 μm wide at the broadest point. Branches within the pyramidal head were short and compact, and dichotomously or trichotomously branched. The central axis within the conidium was hyaline, broad, septate, tapering toward an acute apex, and sometimes constricted at the basal septum. Conidiophores were 272 to 330 μm long and up to 24 μm wide. The fungus was identified as Cristulariella moricola (Hino) Redhead based on morphological characteristics (1,2). The fungus was isolated from Vigna leaf spots, placed on 2% water agar or potato dextrose agar (PDA), and maintained on PDA amended with 2% Vigna leaf extract. For pathogenicity tests, 4- to 5-week-old leaves of V. vexillata var. tsusimensis were surface-sterilized in 1% NaOCl. Agar disks (approximately 10 mm diameter) containing mycelia of the fungus were placed on the upper leaf surface. The inoculated plants (two leaflets per plant × 2) were then sprayed with distilled water, covered with premoistened polyethylene bags, and incubated at 15 to 25°C. Within 5 days, small leaf spots appeared that were similar to those originally observed on all inoculated leaflets. Uninoculated control leaves exposed to the same environmental conditions remained healthy. C. moricola was consistently reisolated from the infected leaves. The hyphomycete fungus C. moricola has been known to cause a bull's eye or zonate leaf spot and defoliation on woody and annual plants, including at least 73 host species and 36 families distributed in the central and eastern United States and Japan (1). In Asia, the occurrence of Cristulariella spp. on several hosts has been reported only in Taiwan and Japan (3,4). No species in the genus has ever been reported from Korea. To our knowledge, V. vexillata var. tsusimensis represents a previously unreported host for C. moricola. References: (1) M. C. Niedbalski et al. Mycologia 75:988, 1983. (2) S. A. Redhead. Mycologia 71:1248, 1979. (3). H. J. Su and S. C. Leu. Plant Dis. 67:915, 1983. (4) T. Yokoyama and K. Tubaki. Trans. Mycol. Soc. Jpn. 15:189, 1974.

scholarly journals First Report of Leaf Spot Disease Caused by Stemphylium vesicarium on Fagopyrum esculentum in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Taixiang Chen ◽  
Yangju Qi ◽  
Lihua Wang ◽  
James Francis White ◽  
Chunjie Li
Keyword(s):  
Leaf Spot ◽  
Apical Cell ◽  
Morphological Features ◽  
Fagopyrum Esculentum ◽  
Leaf Spot Disease ◽  
Width Ratio ◽  
Back Side ◽  
Stemphylium Vesicarium ◽  
Leaf Spots ◽  
Initial Symptoms

Common buckwheat (Fagopyrum esculentum) is a widely cultivated non-grass cereal. It has a considerable market value with nutritional qualities and ability to treat or prevent cancers, hypertension and diabetes (Cawoy et al., 2006). In July-August 2018 and 2019, leaves exhibiting unfamiliar symptoms were observed in Huan County and Huachi County, Gansu, China, with 15% incidence and moderate to severe severity across the field. Initial symptoms consisted of small chlorotic, circular to oval leaf spots. As disease progressed, the spots enlarged and turned light brown to brown with chlorotic margins. When the disease was severe, the leaf spots coalesced and the leaves became prematurely chlorotic and senescent. Spots occurred mostly but not exclusively on older foliage. Diseased tissues were surface sterilized with 75% ethanol for 20 s and 0.1% NaClO for 2 min, placed on PDA medium and incubated at 20 °C for 48-72 h. A total 24 isolates were obtained and purified through single-spore cultures, 19 of which were characterized. Colonies on PDA of all 19 isolates were identical, exhibiting a light gray color, with whitish aerial mycelium that later turned light brown on the reverse of the culture plates, and sporulated sparsely. Conidia were brown colored, cylindrical, and borne singly, often had three main transverse septa, at which points there were conspicuous constrictions, and measured (17-37) μm long × (13-21) μm wide (n=30) in V8 juice agar, (22-38) μm long × (11-19) μm wide (n=30) in PCA, (21-41) μm long × (13-20) μm wide (n=30) in SNA, the mean length/width ratio was 1.6 to 2.0 in V8 juice agar, 1.6 to 2.5 in PCA, 1.3 to 2.2 in SNA. Conidiophores were unbranched, 5.1 to 7.9 μm wide. Dictyospores were produced on well-differentiated conidiophores, the apical cell of conidiophore was slightly to distinctly swollen. Based on morphological features, the isolates were tentatively identified as a member of the Stemphylium vesicarium species complex (Koike et al., 2013; Simmons, 1967). Genomic DNA of representative isolate B1 was extracted, and the internal transcribed spacer (ITS) region and calmodulin gene (cmdA) were amplified using ITS1/4, V9G/ITS4, and CALDF1/CALDR1, respectively. The resulting sequences were deposited in GenBank (acc. nos. MT629829, MW406903, MW417122). Nucleotide BLAST similarity analysis of the sequence fragment of ITS and cmdA from isolate B1 resulted in higher than 99% (99.32% for ITS1/4, 100% for V9G/ITS4, and 100% for CALDF1/CALDR1) identity with S. vesicarium strains (GenBank acc. nos. LC512757, MH863402 and MH206181). Based on morphological features and molecular data, the buckwheat isolates were identified as S. vesicarium. To verify pathogenicity, the back side of leaves from six asymptomic plants were inoculated by spraying the spore suspension (104 spores/ml) harvested from isolate B1 grown on V8 juice agar for 20 days. Control plants were sprayed with sterile water. Each plant was covered with a black plastic bag for 48 h and then was kept in a greenhouse. Stemphylium spot symptoms were observed on all inoculated leaves after 14 days, whereas control leaves were symptomless. The pathogen was reisolated from symptomatic leaf spots, micromorphological features and colony characters of the reisolated fungi were identical to the original isolate. To our knowledge, this is the first documentation of leaf spot of buckwheat caused by S. vesicarium in China, and the first characterization of a Stemphylium foliar pathogen on this crop.

scholarly journals First Report of Leaf Spot Caused by Corynespora cassiicola on Viburnum odoratissimum Ker-Gawl. var. awabuki (K. Koch) Zabel ex Rumpl. (sweet viburnum) in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Tianning Zhang ◽  
Huanhuan Liu ◽  
Qingni Song ◽  
Jun Liu ◽  
Qingpei Yang ◽  
...  
Keyword(s):  
Leaf Spot ◽  
Leaf Spot Disease ◽  
Conidial Suspension ◽  
Puncture Wound ◽  
First Report ◽  
Leaf Spots ◽  
Fungal Isolates ◽  
Initial Symptoms ◽  
Viburnum Odoratissimum ◽  
Sweet Viburnum

Sweet viburnum [Viburnum odoratissimum Ker-Gawl. var. awabuki (K. Koch) Zabel ex Rumpl.] belonging to the family Adoxaceae, is a medical and landscape plant, native to Korea (Jeju Island), Taiwan, and Japan (Edita 1988). In June and September 2019, leaf spots were observed on approximately 65% to 80% of sweet viburnum plants in a hedgerow located in Fenghe Xincheng District (28°41'52.9"N 115°52'14.3"E) in Nanchang, China. Initial symptoms of disease appeared as dark brown spots surrounded by red halos (Figure 1 A), which expanded irregularly. Finally, the center of the lesions desiccated and became light-brown, surrounded by a deep-red halos (Figure 1 B). Ten leaf samples with typical symptoms were collected and washed with tap water for about 15 min. The tissue between the healthy and necrotic area (ca. 4 mm × 4 mm) was cut with a sterile scalpel and surface sterilized with 70% alcohol for 45 s, 2% NaClO for 2 min, washed in sterile deionized water three times, dried on sterilized filter paper, then placed in Petri dishes and incubated at 25℃ in the dark. After 3 to 5 days, the hyphal tips from the edges of growing colonies were transferred to fresh PDA dishes. Eventually, 54 fungal isolates were obtained and, of these, 39 isolates were identical in their morphological characteristics. Morphological analysis was performed according with Ellis (1971). The isolate S18, chosen as representative, formed a gray to grayish brown colony with concentric circleson PDA, and a diameter of 8.5 to 9 cm after 7 days incubation at 25℃ (Figure 1 G). Conidia were hyaline, straight or slightly curved, needle shaped, truncate at the base, and acuminate at the tip, with 2 to 6 pseudosepta, 18.90 to 38.38 µm (avg. = 27.51 µm) × 1.64 to 4.50 µm (avg. = 2.60 µm) (n = 36) (Figure 1 H). The genes of fungal isolates (i.e., ITS, tub2 and ACT) were amplified with ITS4/ITS5 for ITS (White, Bruns et al. 1990), Bt2a/Bt2b for tub2 (Glass and Donaldson 1995) and ACT783R/ACT512F for ACT (Carbone and Kohn 1999) and sequenced. The sequences were deposited in GenBank (MW165772 for ITS, MW175900 for ACT and MW168659 for tub2), which showing greater than 99.1% similarity to multiple C. cassiicola accessions, respectively. Pathogenicity tests were performed on healthy leaves in field by inoculating surface-sterilized mature leaves with puncture wound (Figure C) and non-wounded young leaves with 20 µL of a conidial suspension (105 conidia ml-1) (Figure F and G) at 26℃. After 4 to 7 days, all inoculated leaves reproduced similar symptoms as observed initially in the field (Figure 1 C, E and F). To fulfill Koch’s postulates, the fungus was isolated on PDA from the margins of leaf spots on inoculated leaves and confirmed as C. cassiicola by morphological characters and ITS gene sequencing. Previously, C. cassiicola was reported as an endophyte on Viburnum spp. and Viburnum odoratissimum (Alfieri et al. 1994). More recently, C. cassiicola has been reported as a pathogen of many plant species in China, such as kiwifruit (Cui, Gong et al. 2015), American sweetgum (Mao, Zheng et al. 2021), castor bean (Tang, Liu et al. 2020), and holly mangrove (Xie, He et al. 2020). To our knowledge, this is the first report of leaf spot disease on sweet viburnum caused by C. cassiicola in China and the precise identification of the causal agent will be useful for its management.

scholarly journals First Report of Alternaria alternata Causing Leaf Spot on Menisper-mum dauricum DC. in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Hai feng Sun ◽  
Ming yu Wei ◽  
Na Li ◽  
Yu Yan
Keyword(s):  
Alternaria Alternata ◽  
Leaf Spot ◽  
Aerial Mycelium ◽  
Leaf Spot Disease ◽  
Small Lesion ◽  
Leaf Margin ◽  
Pathogenicity Test ◽  
First Report ◽  
Spot Disease ◽  
Initial Symptoms

Menispermum dauricum DC. is an ornamental plant used in traditional Chinese medicine. (Tang et al. 1992). In September 2019, a leaf spot on M. dauricum DC. was first found in a medicinal plant plantation in Harbin city (45.80°N, 126.53°E), Heilongjiang Province, China. The incidence was 76-90% on the 0.02 ha plantation. The initial symptoms were irregular black and brown spots on the leaves. The lesions expanded and coalesced, eventually leading to blight. Fresh leaf samples from ten M. dauricum plants with typical symptoms were collected. The areas of leaf between symptomatic and healthy tissue (5㎜×5㎜) were cut and surface disinfeated in 75% ethanol for 2 min, and with 1% HgCl2 for 1 min, and then rinsed three times with sterile water. Small lesion pieces were incubated on potato dextrose agar (PDA) for 7 days at 25℃, in the dark. Ten fungal isolates were obtained and transferred onto new PDA and potato carrot agar (PCA) plates to establish pure cultures. After 8 days, the colonies on PDA were 75-86㎜ in diameter, circular, with distinct concentric rings and a whitish aerial-mycelium margin, cottony, light gray to dark bluish brown. The colonies on the PCA were olive-green and bordered by white aerial hyphae. A total of 150 conidia were single or in short chains, obclavate, oval or inverted pear, light brown to brown, smooth or slightly spiny, with 1 to 6 transverse septa, 0 to 4 longitudinal or oblique septa, not narrow or slightly narrowed at the separation, 22.5-42.5×7.5-15.5㎛, and rostrate. Conidiophores were simple, erect, or ascending, dark brown, geniculate, septate, and with one or several conidial scars, 32.5-77.5×3.0-5.0㎛. Beaks were columnar or conical, 7.5-22.5×2.5-3.5㎛. Morphologically, all ten isolates were most similar to Alternaria alternata (Simmons 2007). For further identification of the fungus at the molecular level, internal transcribed spacer rDNA regions (ITS), RNA polymerase second largest subunit gene (RPB2) and Alternaria major allergen (Alt a 1) were amplified and sequenced using the primers ITS1 and ITS4, RPB2-5F2 and RPB2-7CR, Alt-for and Alt-rev (Woudenberg et al. 2015). The resulting sequences were deposited in GenBank (ITS: MT995193, MZ150794, RPB2: MT999483, MZ170963, Alt a 1: MT802122, MZ170962). BLAST search of these sequences showed 99%-100% homology with the ITS (FJ196306), RPB2 (KC584375) and Alt a 1 (KT315515) of the type strain CBS 916.96 of A. alternata, respectively. Thus, the fungus was identified as A. alternata based on the morphology and molecular analysis. For the pathogenicity test, spore suspensions (1×106 spores/mL) of the representative isolates BFG001 and BFG002 were sprayed onto the leaves of six healthy plants, separately. As a control, six plants were treated with sterile distilled water. The plants used in the experiment were covered with plastic bags and incubated at 25℃ for 10 days. Eight days after inoculation, irregular, slightly sunken black leaf spots appeared at the leaf margin. The experiment was repeated three times with the same method. The same fungus was successfully re-isolated from the leaves of the inoculated plants, fulfilling Koch’s postulates. No symptoms were observed on control plants. To our knowledge, this is the first report of leaf spot disease on M. dauricum DC. caused by A. alternata in the world. The appearance of leaf spot disease reduces the yield and quality of Chinese medicinal materials. This report has laid the foundation for the further research and control of leaf spot disease.

scholarly journals First Report of Fusarium commune Causing Leaf Spot Disease on Bletilla striata in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Wang Hanyi ◽  
Hou Xiuming ◽  
Xueming Huang ◽  
Meng Gao ◽  
Tingting Chen ◽  
...  
Keyword(s):  
Leaf Spot ◽  
Morphological Characteristics ◽  
Leaf Spot Disease ◽  
Molecular Characteristics ◽  
Post Inoculation ◽  
Bletilla Striata ◽  
First Report ◽  
Spot Disease ◽  
Severe Stage ◽  
Initial Symptoms

Bletilla striata (Thunb.) Rchb. f. (Orchidaceae family, known as Baiji in Chinese) is a perennial herb and has been traditionally used for hemostasis and detumescence in China. In April of 2020, a leaf spot disease on B. striata was observed in plant nurseries (∼0.2 h) in Guilin, Guangxi Province, China. Approximately 20% of the plants were symptomatic, of which 150 plants were randomly selected for investigation. Initial symptoms include the appearance of small, circular or irregular light brown spots, randomly scattered on the edges and surfaces of the leaves, which progressively expand into large, suborbicular or irregular-shaped dark brown, necrotic areas. At the severe stage, the lesions coalesced into large necrotic areas and ultimately resulted in leaf abscission. To isolate the pathogen, three representative plants exhibiting symptoms were collected from the nurseries. Leaf tissues (5 × 5 mm) were cut from the margin of necrotic lesions (n = 18), surface-disinfected in 1% sodium hypochlorite (NaOCl) solution for 2 min, then rinsed three times in sterile water before isolation. The tissues were plated on potato dextrose agar (PDA) medium, and incubated at 28°C (12-h photoperiod) for 3 days. Hyphal tips from recently germinated spores were transferred to PDA to obtain pure cultures. Nine fungal isolates with similar morphological characteristics were obtained. Three single-spore isolates, BJ23.1, BJ55.1, and BJ91.3, were subjected to further morphological and molecular characterisation. Colonies on PDA plates were villose, had a dense growth of aerial mycelia and appeared white (1A1) to yellowish white (3A2). Macroconidia were smooth, hyaline, straight to slightly curved, usually contained three or five septa, and measuring 23.3 to 42.1 × 3.0 to 6.2 μm (mean ± SD: 31.2 ± 5.1 × 4.2 ± 0.6 μm, n = 50). Microconidia were generally cylindrical, straight to slightly curved, aseptate, and measuring 7.2 to 18.8 × 2.5 to 4.3 μm (mean ± SD: 12.1 ± 2.8 × 3.3 ± 0.5 μm, n = 62). Morphological characteristics are similar to those of F. commune (Skovgaard et al. 2003). For molecular identification, the genomic DNA of the isolates BJ23.1, BJ55.1, and BJ91.3 were extracted using the CTAB method (Guo et al. 2000). The internal transcribed spacer (ITS) region of rDNA, partial translation elongation factor-1 alpha (TEF-1α), RNA polymerase second largest subunit (RPB2), and the mitochondrial small subunit rDNA (mtSSU) genes were amplified using primer pairs [ITS1/ITS4 (White et al. 1990), EF-1/EF-2 (O’Donnell et al. 1998), and 5f2/11ar (Liu et al. 1999, Reeb et al. 2004), MS1/MS2 (Li et al. 1994), respectively]. The obtained sequences were deposited in NCBI GenBank under the following accession numbers: ITS (MZ424697 to MZ424699), TEF-1α (MZ513467 to MZ513469), RPB2 (MZ513473 to MZ513475), and mtSSU (MZ513470 to MZ513472). BLAST® analysis of the deposited sequences showed 99 to 100% identity with those of F. commune present in GenBank (Accession numbers: DQ016205, MH582348, MH582181, AF077383). In addition, a phylogenetic analysis using concatenated sequences of ITS, TEF-1α, mtSSU genes showed that BJ23.1, BJ55.1, and BJ91.3 located on the same clade with strains of F. commune. Therefore, based on morphological and molecular characteristics, the isolates were identified as F. commune (Skovgaard et al. 2003, Stewart et al. 2006). Pathogenicity was tested using 1.5-year-old B. striata plants. Healthy leaves on plants were inoculated with 5 × 5 mm mycelial discs of strains BJ23.1, BJ55.1, and BJ91.3 from 3-day-old PDA cultures, each isolate was inoculated onto three plants; three other plants inoculated with sterile PDA discs served as controls. All plants were enclosed in transparent plastic bags and incubated in a greenhouse at 28°C for 14 days (12-h photoperiod). Three days post-inoculation, leaf spot symptoms appeared on the inoculated leaves. No symptoms were detected on control plants. Experiments were replicated three times with similar results. To fulfill Koch’s postulates, F. commune was consistently re-isolated from symptomatic tissue and confirmed by morphology and sequencing, whereas no fungus was isolated from the control plants. F. commune has been reported to cause diseases on some plants, including sugarcane (Wang et al. 2018), maize (Xi et al. 2019) and Wax Gourd (Zeng et al. 2020). To our knowledge, this is the first report of F. commune causing leaf spot disease on B. striata in China. Identification of this pathogen provides the information for further studies to develop management strategies to control the disease.

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