First Report of Leaf Spot Caused by Alternaria alternata on Marigold (Tagetes erecta) in 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.

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First Report of Leaf Spot on Industrial Hemp (Cannabis sativa L.) Caused by Alternaria alternata in China

Plant Disease ◽

10.1094/pdis-01-21-0130-pdn ◽

2021 ◽

Author(s):

Lili Tang ◽

Xixia Song ◽

Liguo Zhang ◽

Jing Wang ◽

Shuquan Zhang

Keyword(s):

Leaf Spot ◽

Cannabis Sativa ◽

Leaf Spot Disease ◽

Molecular Characteristics ◽

Pathogenicity Test ◽

Conidial Suspension ◽

Sterile Water ◽

First Report ◽

Leaf Spots ◽

Industrial Hemp

Industrial hemp is an economically important plant with traditional uses for textiles, paper, building materials, food and medicine (Li 1974; Russo et al. 2008; Zlas et al. 1993). In August 2020, an estimated 80% of the industrial hemp plants with leaf spots were observed in greenhouse in Minzhu town, Harbin City, Heilongjiang Province, China (45.8554°N, 126.8167°E), resulting in yield losses of 20%. Leaf symptoms began as small spots on the upper surface of leaves and gradually developed into brown spots with light yellow halos. These irregular spots expanded gradually and eventually covered the entire leaf; the center of the spots was easily perforated. To identify the pathogen, 20 diseased leaves were collected, and small sections of (3 to 5 mm) were taken from the margins of lesions of infected leaves. The pieces were sterilized with 75% alcohol for 30 s, a 0.1% mercuric chloride solution for 1 min, and then rinsed three times with sterile water. Samples were then cultured on potato dextrose agar at 28℃ in darkness for 4 days. A single-spore culture was obtained by monosporic isolation. Conidiophores were simple or branched, straight or flexuous, brown, and measured 22 to 61 μm long × 4 to 5 μm wide (n = 50). Conidia were solitary or in chains, brown or dark brown, obclavate, obpyriform or ellipsoid. Conidia ranged from 23 to 55 μm long × 10 to 15 μm wide (n = 50) with one to eight transverse and several longitudinal septa. For molecular identification (Jayawardena et al. 2019), genomic DNA of pathogenic isolate (MZ1287) was extracted by a cetyltrimethylammonium bromide protocol. Four gene regions including the rDNA internal transcribed spacer (ITS), glyceraldehyde-3-phosplate dehydrogenase (GAPDH), translation elongation factor 1-alpha (TEF1) and RNA polymerase II beta subunit (RPB2) were amplified with primers ITS1/ITS4, GDF1/GDR1, EF1-728F/EF1-986R and RPB2-5F/RPB2-7cR, respectively (White et al. 1990). Resulting sequences were deposited in GenBank with accession numbers of MW272539.1, MW303956.1, MW415414.1 and MW415413.1, respectively. A BLASTn analysis showed 100% homology with A. alternata (GenBank accession nos. MN615420.1, MH926018.1, MN615423.1 and KP124770.1), respectively. A neighbor-joining phylogenetic tree was constructed by combining all sequenced loci in MEGA7. The isolate MZ1287 clustered in the A. alternata clade with 100% bootstrap support. Thus, based on morphological (Simmons 2007) and molecular characteristics, the pathogen was identified as A. alternata. To test pathogenicity, leaves of ten healthy, 2-month-old potted industrial hemp plants were sprayed using a conidial suspension (1×106 spores/ml). Control plants were sprayed with sterile water. All plants were incubated in a greenhouse at 25℃ for a 16 h light and 8 h dark period at 90% relative humidity. The experiment was repeated three times. After two weeks, leaf spots of industrial hemp developed on the inoculated leaves while the control plants remained asymptomatic. The A. alternata pathogen was re-isolated from the diseased leaves on inoculated plants, fulfilling Koch's postulates. Based on morphology, sequencing, and pathogenicity test, the pathogen was identified as A. alternata. To our knowledge, this is the first report of A. alternata causing leaf spot disease of industrial hemp (Cannabis sativa L.) in China and is worthy of our attention for the harm it may cause to industrial hemp production.

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

Plant Disease ◽

10.1094/pdis-07-21-1447-pdn ◽

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.

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

Plant Disease ◽

10.1094/pdis-08-20-1842-pdn ◽

2021 ◽

Author(s):

Yun-fei Mao ◽

Xiang-rong Zheng ◽

Fengmao Chen

Keyword(s):

Leaf Spot ◽

Morphological Characteristics ◽

Liquidambar Styraciflua ◽

Bootstrap Support ◽

Leaf Spot Disease ◽

Pathogenicity Test ◽

Conidial Suspension ◽

Corynespora Cassiicola ◽

First Report ◽

Leaf Spots

American sweetgum (Liquidambar styraciflua L.) is a forest plant native to North America, which has been introduced into other countries due to its ornamental and medicinal values. In June 2019, symptoms of leaf spots on sweetgum were observed in a field (5 ha) located in Xuzhou, Jiangsu Province, China. On this field, approximately 45% of 1,000 trees showed the same symptoms. Symptoms were observed showing irregular or circular dark brown necrotic lesions approximately 5 to 15 mm in diameter with a yellowish margin on the leaves. To isolate the pathogen, diseased leaf sections (4×4mm) were excised from the margin of the lesion, surface-sterilized with 0.1% NaOCl for 90 s, rinsed 4 times in sterile distilled water, air dried and then transferred on potato dextrose agar (PDA) medium at 25°C in the dark. Pure cultures were obtained by monospore isolation after subculture. Ten purified isolates, named FXI to FXR, were transferred to fresh PDA and incubated as above to allow for morphological and molecular identification. After 7 days, the aerial mycelium was abundant, fluffy and exhibited white to greyish-green coloration. The conidia were dark brown or olive, solitary or produced in chains, obclavate, with 1 to 15 pseudosepta, and measured 45 to 200µm  10 to 18µm. Based on morphological features, these 10 isolates were identified as Corynespora cassiicola (Ellis et al. 1971). Genomic DNA of each isolate was extracted from mycelia using the cetyltrimethylammonium bromide (CTAB) method. The EF-1α gene and ITS region were amplified and sequenced with the primer pairs rDNA ITS primers (ITS4/ITS5) (White et al. 1990) and EF1-728F/EF-986R (Carbone et al.1999) respectively. The sequences were deposited in GenBank. BLAST analysis revealed that the ITS sequence had 99.66% similarity to C. cassiicola MH255527 and that the EF-1α sequence had 100% similarity to C. cassiicola KX429668A. maximum likelihood phylogenetic analysis based on EF-1α and ITS sequences using MEGA 7 revealed that ten isolates were placed in the same clade as C. cassiicola (Isolate: XQ3-1; accession numbers: MH572687 and MH569606, respectively) at 98% bootstrap support. Based on the morphological characteristics and phylogenetic analyses, all isolates were identified as C. cassiicola. For the pathogenicity test, a 10 µl conidial suspension (1×105 spores/ml) of each isolate was dripped onto healthy leaves of 2-year-old sweetgum potted seedlings respectively. Leaves inoculated with sterile water served as controls. Three plants (3 leaves per plant) were conducted for each treatment. The experiment was repeat twice. All seedlings were enclosed in plastic transparent incubators to maintain high relative humidity (90% to 100%) and incubated in a greenhouse at 25°C with a 12-h photoperiod. After 10 days, leaves inoculated with conidial suspension of each isolate showed symptoms of leaf spots, similar to those observed in the field. Control plants were remained healthy. In order to reisolate the pathogen, surface-sterilized and monosporic isolation was conducted as described above. The same fungus was reisolated from the lesions of symptomatic leaves, and its identity was confirmed by molecular and morphological approaches, thus fulfilling Koch’s postulates. Chlorothalonil and Boscalid can be used to effectively control Corynespora leaf spot (Chairin T et al.2017). To our knowledge, this is the first report of leaf spot caused by C. cassiicola on L. styraciflua in China.

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First Report of Leaf Spot in Farfugium japonicum Caused by Alternaria alternata in China

Plant Disease ◽

10.1094/pdis-02-21-0395-pdn ◽

2021 ◽

Author(s):

Hui Wang ◽

Hong Liu ◽

Xun Lu ◽

Qian Zhou

Keyword(s):

Alternaria Alternata ◽

Leaf Spot ◽

Economic Losses ◽

Leaf Spot Disease ◽

Pathogenicity Test ◽

Distilled Water ◽

Conidial Suspension ◽

Isolation Technique ◽

First Report ◽

Farfugium Japonicum

Farfugium japonicum (L.) Kitam (with the common name leopard plant) is known as a garden and medical herb, and belongs to the family Asteraceae. In May 2019, a leaf spot disease was observed on the upper leaf surface of F. japonicum in Changsha city, Hunan province, China. More than 98% of the F. japonicum plants were infected in a garden of Donghu district (28°13′ N; 112°56′ E). Leaf symptoms included small (1 to 10 mm in diameter), brown spots that were circular, tan to gray in the center and distinct brownish-yellow margins. Severely affected leaves were blighted and plants were dying. For isolation, symptomatic leaf tissue was surface sterilized, rinsed in sterile distilled water, and plated on potato dextrose agar (PDA) amended with a 50 μg/ml streptomycin sulfate followed by incubation at 25°C in darkness. By a single-spore isolation technique, pure fungal cultures were obtained and displayed gray-brown and gray-white aerial mycelia after five days of incubation. One representative isolate (HnAa-1) was selected for further studies. Conidia of HnAa-1 were olive brown, obpyriform, either branched or unbranched with a short beak, 1 to 5 transverse septa, and 0 to 3 longitudinal or oblique septa. The conidia were 10 to 35 μm long and 2 to 12 μm wide. HnAa-1 was identified as an Alternaria sp. on the basis on morphological characterization by Simmons (1). Further identification to species level was made by molecular analyses. DNA of HnAa-1 was extracted from the regions internal transcribed spacer (ITS), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and partial Alt a 1 major allergen (ALT) gene. Amplification and sequencing was carried out with the method described by Woudenberg et al.(2) . BLASTn searches showed that the ITS, GAPDH and ALT sequences had the highest similarity with A. alternata strains, with 100% (548/548) identities for ITS (GQ169728), 100% (567/567) identities for GAPDH (MK903028) and 99.36% (466/469) identities for ALT (MN184998). Moreover, the ITS, GAPDH and ALT sequences had more than 99% identities with the epitype CBS 916.96 of A. alternata (ITS: AF347031; GAPDH: AY278808; ALT: AY563301). The ITS, GAPDH and ALT sequences of HnAa-1 were submitted to GenBank (Accession No. MT767170, No. MW115639 and No. MW316727). Pathogenicity tests were conducted by spraying a 10 ml conidial suspension (1.0 ×105 conidia /mL) on surfaces of leaves of three healthy plants (8-week-old). Leaves of three healthy plants were sprayed with sterile distilled water as a control treatment. All inoculated plants were maintained in growth chamber at 25°C with a 12-h photoperiod. The pathogenicity test was repeated twice. After five days inoculation, typical brown spots and necrotic lesions similar to those observed in the field, had developed on all inoculated plants but not on water-treated control plants. Alternaria alternata was re-isolated from the symptomatic tissue of inoculated plants but not from the control plants, and re-identified with morphological and molecular methods, which fulfilled Koch's postulates. This host-pathogen association has been reported in Korea (3), but it is the first report of A. alternata causing leaf spots on F. japonicum in China. Since A. alternata is a ubiquitous and very important plant pathogen causing leaf spot diseases in over 100 species plant, the occurrence of this disease is a serious threat to F.japonicum and might lead to economic losses. Therefore, appropriate prevention strategies to F.japonicum should be adopted.

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First Report of Alternaria alternata Causing Leaf Spot on English Walnut

Plant Disease ◽

10.1094/pdis.1999.83.7.696a ◽

1999 ◽

Vol 83 (7) ◽

pp. 696-696 ◽

Cited By ~ 5

Author(s):

A. Belisario ◽

E. Forti ◽

L. Corazza ◽

H. A. van Kesteren

Keyword(s):

Alternaria Alternata ◽

Leaf Spot ◽

Juglans Regia ◽

Deciduous Trees ◽

Sterile Water ◽

First Report ◽

Leaf Spots ◽

Pathogenicity Tests ◽

Necrotic Spots ◽

English Walnut

A foliar disease of English walnut (Juglans regia L.) was observed in 1998 in an orchard in northern Italy. Symptoms consisted of circular, necrotic spots bordered with concentric zones of darker tissue. Average lesion diameter was 20 mm, although lesions could extend to half of the leaflet lamina. A fungus identified as Alternaria alternata (Fr.:Fr.) Keissl. (1) was observed on the necrotic tissue and was consistently isolated from the margins of the necrosis. Conidia from leaves were brown, ellipsoid to ovoid (primary conidia 28 to 60 × 8 to 15 μm; secondary conidia 10 to 30 × 7 to 13 μm), with walls often ornamented, 1 to 6 transversely septate and 0 to 3 longitudinally septate. For pathogenicity tests, three isolates were grown on potato dextrose agar for 2 weeks. Inoculations were performed on detached, surface-sterilized, healthy J. regia leaflets. Four drops (5 μl each) of a sterile water suspension of 1 × 105 conidia per ml were placed on each leaflet; three leaves per isolate were used. Leaves were incubated in a moist chamber. After 10 days, leaf spots similar to the original symptoms developed on all the inoculated points for all three isolates, and the pathogen was reisolated. Control leaflets inoculated with sterile, distilled water remained symptomless. The experiment was performed three times and the results were similar. Alternaria alternata is a well-known pathogen on many crops but a few records report this fungus as a causal agent of leaf spot on deciduous trees. This is the first report of A. alternata on English walnut. Reference: (1) E. G. Simmons. Mycotaxon 37:79, 1990.

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First Report of a Leaf Spot on Conyza sumatrensis Caused by Phoma macrostoma in China

Plant Disease ◽

10.1094/pdis-03-11-0228 ◽

2012 ◽

Vol 96 (1) ◽

pp. 148-148 ◽

Cited By ~ 1

Author(s):

J. Liu ◽

H. D. Luo ◽

W. Z. Tan ◽

L. Hu

Keyword(s):

Leaf Spot ◽

Fungal Pathogens ◽

Biological Diversity ◽

Biocontrol Agent ◽

Continuous Light ◽

The United States ◽

Leaf Spot Disease ◽

Pathogenicity Test ◽

Dry Land ◽

First Report

Conyza sumatrensis (Asteraceae), an annual or biennial plant, is native to North and South America. It is an invasive, noxious weed that is widespread in southern and southeastern China. It invades farm land and causes great losses to dry land crops, including wheat, corn, and beans. It also reduces biological diversity by crowding out native plants in the infested areas (3,4). During a search for fungal pathogens that could serve as potential biological control agents of C. sumatrensis, a leaf spot disease was observed in 2010 in Chongqing, China. An isolate (SMBC22) of a highly virulent fungus was obtained from diseased leaves. Pathogenicity tests were performed by placing 6-mm-diameter mycelial disks of 7-day-old potato dextrose agar (PDA) cultures of SMBC22 on leaves of 15 healthy greenhouse-grown plants of C. sumatrensis; the same number of control plants was treated with sterile PDA disks. Treated plants were covered with plastic bags for 24 h and maintained in a growth chamber with daily average temperatures of 24 to 26°C, continuous light (3,100 lux), and high relative humidity (>90%). Lesions similar to those observed in the field were first obvious on the SMBC22-inoculated leaves 3 days after inoculation. Symptoms became severe 7 to 9 days after inoculation. Control plants remained healthy. The fungus was reisolated from inoculated and diseased leaves and it was morphologically the same as SMBC22. The pathogenicity test was conducted three times. A survey of 10 southern and southeastern Chinese provinces revealed that the disease was widespread and it attacked leaves and stems of seedlings and mature plants of C. sumatrensis. Lesions on leaves were initially small, circular, and water soaked. The typical lesion was ovoid or fusiform, dark brown, and surrounded by a yellow halo. The spots coalesced to form large lesions and plants were often completely blighted. Fungal colonies of SMBC22 on PDA plates were initially white and turned dark gray. Colonies were circular with smooth edges with obvious rings of pycnidia on the surface. Aerial hyphae were short and dense. Pycnidia, black and immersed or semi-immersed in the medium, were visible after 12 days of incubation. Pycnidia were 72 to 140 μm in diameter. Conidia were produced in the pycnidia and were hyaline, unicellular, ellipsoidal, and 4.4 to 6.1 × 1.6 to 2.2 μm. To confirm identification of the fungus, genomic DNA was extracted from mycelia of a 7-day-old culture on PDA at 25°C (2). The internal transcribed spacer (ITS) gene of rDNA was amplified using primers ITS4/ITS5. The gene sequence was 524 bp long and registered in NCBI GenBank (No. HQ645974). BLAST analysis showed that the current sequence had 99% homology to an isolate of Phoma macrostoma (DQ 404792) from Cirsium arvense (Canada thistle) in Canada and reported to cause chlorotic symptoms on that host plant (1). To our knowledge, this is the first report of P. macrostoma causing disease on C. sumatrensis in China. P. macrostoma, thought of as a biocontrol agent of broadleaf weeds in Canada, has been patented in the United States. The current isolate of P. macrostoma is considered as a potential biocontrol agent of C. sumatrensis. References: (1) P. R. Graupner et al. J. Nat. Prod. 66:1558, 2004. (2) S. Takamatsu et al. Mycoscience 42:135, 2001. (3) W. Z. Tan et al. Page 177 in: Manual of Emergency Control Technology Invasive Pests in China. G. L. Zhang, ed. Science Press, Beijing, 2010. (4) C. Wang et al. J. Wuhan Bot. Res. 28:90, 2010.

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First Report of Alternaria Leaf Spot of Banana Caused by Alternaria alternata in the United States

Plant Disease ◽

10.1094/pdis-01-13-0007-pdn ◽

2013 ◽

Vol 97 (8) ◽

pp. 1116-1116 ◽

Cited By ~ 3

Author(s):

V. Parkunan ◽

S. Li ◽

E. G. Fonsah ◽

P. Ji

Keyword(s):

United States ◽

Alternaria Alternata ◽

Leaf Spot ◽

Morphological Characteristics ◽

The United States ◽

Its Sequencing ◽

Single Spore ◽

First Report ◽

Alternaria Leaf Spot ◽

Leaf Spots

Research efforts were initiated in 2003 to identify and introduce banana (Musa spp.) cultivars suitable for production in Georgia (1). Selected cultivars have been evaluated since 2009 in Tifton Banana Garden, Tifton, GA, comprising of cold hardy, short cycle, and ornamental types. In spring and summer of 2012, 7 out of 13 cultivars (African Red, Blue Torres Island, Cacambou, Chinese Cavendish, Novaria, Raja Puri, and Veinte Cohol) showed tiny, oval (0.5 to 1.0 mm long and 0.3 to 0.9 mm wide), light to dark brown spots on the adaxial surface of the leaves. Spots were more concentrated along the midrib than the rest of the leaf and occurred on all except the newly emerged leaves. Leaf spots did not expand much in size, but the numbers approximately doubled during the season. Disease incidences on the seven cultivars ranged from 10 to 63% (10% on Blue Torres Island and 63% on Novaria), with an average of 35% when a total of 52 plants were evaluated. Six cultivars including Belle, Ice Cream, Dwarf Namwah, Kandarian, Praying Hands, and Saba did not show any spots. Tissue from infected leaves of the seven cultivars were surface sterilized with 0.5% NaOCl, plated onto potato dextrose agar (PDA) media and incubated at 25°C in the dark for 5 days. The plates were then incubated at room temperature (23 ± 2°C) under a 12-hour photoperiod for 3 days. Grayish black colonies developed from all the samples, which were further identified as Alternaria spp. 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 23 to 73 μm long and 15 to 35 μm wide, with a beak length of 5 to 10 μm, and had 3 to 6 transverse and 0 to 5 longitudinal septa. Single spore cultures of four isolates from four different cultivars were obtained and genomic DNA was extracted and the internal transcribed spacer (ITS1-5.8S-ITS2) regions of rDNA (562 bp) were amplified and sequenced with primers ITS1 and ITS4. MegaBLAST analysis of the four sequences showed that they were 100% identical to two Alternaria alternata isolates (GQ916545 and GQ169766). ITS sequence of a representative isolate VCT1FT1 from cv. Veinte Cohol was submitted to GenBank (JX985742). Pathogenicity assay was conducted using 1-month-old banana plants (cv. Veinte Cohol) grown in pots under greenhouse conditions (25 to 27°C). Three plants were spray inoculated with the isolate VCT1FT1 (100 ml suspension per plant containing 105 spores per ml) and incubated under 100% humidity for 2 days and then kept in the greenhouse. Three plants sprayed with water were used as a control. Leaf spots identical to those observed in the field were developed in a week on the inoculated plants but not on the non-inoculated control. The fungus was reisolated from the inoculated plants and the identity was confirmed by morphological characteristics and ITS sequencing. To our knowledge, this is the first report of Alternaria leaf spot caused by A. alternata on banana in the United States. Occurrence of the disease on some banana cultivars in Georgia provides useful information to potential producers, and the cultivars that were observed to be resistant to the disease may be more suitable for production. References: (1) E. G. Fonsah et al. J. Food Distrib. Res. 37:2, 2006. (2) E. G. Simmons. Alternaria: An identification manual. CBS Fungal Biodiversity Center, Utrecht, Netherlands, 2007.

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

Plant Disease ◽

10.1094/pdis.2003.87.2.203a ◽

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.

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

Plant Disease ◽

10.1094/pdis-09-14-0938-pdn ◽

2015 ◽

Vol 99 (2) ◽

pp. 289-289 ◽

Cited By ~ 9

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.

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First Report of Alternaria Alternata Causing Brown Leaf Spot on Apricot (Prunus Armeniaca) in Karbala Province of Iraq

IOP Conference Series Earth and Environmental Science ◽

10.1088/1755-1315/910/1/012080 ◽

2021 ◽

Vol 910 (1) ◽

pp. 012080

Author(s):

Zeinab L. Hameed ◽

Adnan A. Lahuf ◽

Mundher T. Jasim ◽

Hassan M. Mohsen ◽

Bahaa J. Kadim ◽

...

Keyword(s):

Alternaria Alternata ◽

Leaf Spot ◽

Causative Factor ◽

Prunus Armeniaca ◽

Leaf Spot Disease ◽

First Report ◽

Spot Disease ◽

Brown Leaf Spot ◽

Rdna Sequencing ◽

Brown Leaf

Abstract During a survey in season 018, leaf spot symptoms were commonly observed on apricot (Prunus armeniaca) trees in the orchards of Al-Hussainiya district in Karbala Province of Iraq. The symptomatic leaves were gathered, the associated fungus was isolated and characterized relied on its morphological features and ITS-rDNA sequencing. The causative factor was found to be the fungus Alternaria alternata that caused distinguishable leaf spot symptoms on the inoculated leaves of apricot. Based on a review of previous references relatedof this disease in Iraq, this is the first report of the brown leaf spot disease caused by A. alternata in Karbala province of Iraq.

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