First Report of Rhexocercosporidium panacis Causing Rusty Root of Panax ginseng in Northeastern China

In northeastern China, Asian ginseng (Panax ginseng) roots exhibited reddish brown lesions of various sizes, irregular shapes, and diffuse margins, typical of rusty root disease. The lesions remain superficial, smooth, and limited to the epidermal and peridermal tissues. In September 2013, 10 symptomatic roots were collected from each of three fields in Jilin and Heilongjiang provinces. One piece of symptomatic skin tissue from each root was excised, surface-disinfested in 1% NaClO for 3 min, rinsed three times with sterile water, and then placed on tetracycline-amended (50 μg/ml) potato dextrose agar. After incubation at 22 ± 1°C in the dark for a week, small olivaceous black colonies developed from the symptomatic tissue from five of the 30 samples. No spores were observed. A single hyphal tip of each colony was transferred to a fresh V8 agar plate to purify the culture. Two-week-old colonies on V8 agar were olivaceous gray, and 42 to 46 mm in diameter with an outer white margin (3 to 5 mm wide). Conidia produced in V8 broth after 3 weeks with a 12-h photoperiod were straight and hyaline, cylindrical or subcylindrical with no or one septum. Mature conidia were 12.8 to 21.8 × 2.2 to 4.5 μm (mean 18.2 × 3.0 μm, n = 100 conidia for each of three isolates). Three isolates selected randomly were further identified by analyzing the partial sequences of the ITS region of rDNA with primers ITS4 and ITS5 (5), and partial sequences of β-tubulin with the primers tub2F and tub2R (1). Sequences of the three isolates (GenBank Accession Nos. KJ149287, KJ149288, and KJ149290 to 93) showed 99% to 100% homology with previously identified and deposited Rhexocercosporidium panacis isolates (DQ2499992 and DQ457119) for both loci (3). Therefore, the three isolates were identified as R. panacis and deposited in China General Microbiological Culture Collection Center (CGMCC3.17259 to 61). Pathogenicity of R. panacis in Asian ginseng was investigated using these three isolates as described previously with slight modifications (4). Bare roots of 3-year-old Asian ginseng were surface-disinfested as described above, and inoculated with mycelial plugs (4 mm diameter) cut from the margin of actively growing colonies of the isolates on V8 agar. Three mycelial plugs were placed on each root at 3-cm intervals and four roots (replicates) were inoculated for each isolate. Four additional roots were inoculated with non-colonized agar plugs as control. The treated roots were placed on moist filter paper in an enamel tray. The plates were sealed with plastic wrap to prevent desiccation and incubated in the dark at 18 ± 1°C. Four weeks post inoculation, all the inoculated ginseng roots showed red-brown lesions, which turned to dark red or black over time. R. panacis was recovered from symptomatic roots for all isolates and confirmed by ITS sequence analysis. The mock-inoculated control roots remained symptomless and no R. panacis was isolated. The inoculation experiment was repeated and showed the same results. R. panacis was reported in 2006 to infect roots of Panax quinquefolius (2,3,4). To our knowledge, this is the first report of R. panacis causing rusty root of P. ginseng. References: (1) P. R. Hirsch et al. Mycol. Res. 104:435, 2000. (2) Z. K. Punja et al. Can. J. Plant Pathol. 35:503, 2013. (3) R. D. Reeleder. Mycologia. 99:91, 2007. (4) R. D. Reeleder et al. Phytopathology 96:1243, 2006. (5) T. J. White et al. PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990.

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First Report of Dollar Spot of Sandbur Caused by Sclerotinia homoeocarpa in Oklahoma

Plant Disease ◽

10.1094/pdis-03-14-0244-pdn ◽

2014 ◽

Vol 98 (8) ◽

pp. 1160-1160

Author(s):

F. Flores ◽

N. R. Walker

Keyword(s):

Warm Season ◽

Its Region ◽

Small Subunit ◽

Type I ◽

Academic Press ◽

Sclerotinia Homoeocarpa ◽

Dollar Spot ◽

First Report ◽

Small Subunit Rdna ◽

Disturbed Soils

Sandbur (Cenchrus incertus Curtis) is a warm-season, annual, noxious, grassy weed native to southern North America. It is common in sandy, disturbed soils and can also be found in home lawns and sport fields where low turf density facilitates its establishment. In July 2013, after a period of frequent rainfall and heavy dew, symptoms of dollar spot-like lesions (1) were observed on sandbur plants growing in a mixed stand of turf-type and native warm-season grasses in Logan County, Oklahoma. Lesions, frequently associated with leaf sheaths, were tan and surrounded by a dark margin. Symptomatic leaves were surface sterilized and plated on potato dextrose agar amended with 10 ppm rifampicin, 250 ppm ampicillin, and 5 ppm fenpropathrin. After incubation, a fungus morphologically identical to Sclerotinia homoeocarpa Bennett was consistently isolated. The nuclear ribosomal internal transcribed spacer (ITS) region of two different isolates, SCL2 and SCL3, were amplified using primers ITS4 and ITS5 (2). The DNA products were sequenced and BLAST analyses were used to compare sequences with those in GenBank. The sequence for isolate SLC2 was 869 bp, contained a type I intron in the 18S small subunit rDNA, and was identical to accession EU123803. The ITS sequence for isolate SLC3 was 535 bp and identical to accession EU123802. Twenty-five-day-old seedlings of C. incertus were inoculated by placing 5-mm-diameter agar plugs, colonized by mycelia of each S. homoeocarpa isolate, onto two of the plants' leaves. Plugs were held in place with Parafilm. Two plants were inoculated with each isolate and sterile agar plugs were placed on two leaves of another seedling as control. Plants were incubated in a dew chamber at 20°C and a 12-h photoperiod. After 3 days of incubation, water-soaked lesions surrounded by a dark margin appeared on inoculated plants only. Fungi that were later identified as S. homoeocarpa isolates SLC2 and SLC3 by sequencing of the ITS region were re-isolated from symptomatic leaves, fulfilling Koch's postulates. To our knowledge, this is the first report of dollar spot on sandbur. References: (1) R. W. Smiley et al. Page 22 in: Compendium of Turfgrass Diseases. 3rd ed. The American Phytopathological Society, St. Paul, MN, 2005. (2) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990.

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First Report of Leaf Spot caused by Bipolaris oryzae on Switchgrass in Tennessee

Plant Disease ◽

10.1094/pdis-01-13-0070-pdn ◽

2013 ◽

Vol 97 (12) ◽

pp. 1654-1654 ◽

Cited By ~ 3

Author(s):

A. L. Vu ◽

M. M. Dee ◽

J. Zale ◽

K. D. Gwinn ◽

B. H. Ownley

Keyword(s):

Leaf Spot ◽

Panicum Virgatum ◽

Morphological Characteristics ◽

Its Region ◽

Spore Production ◽

Post Inoculation ◽

Sterile Water ◽

Bipolaris Oryzae ◽

First Report ◽

Symptomatic Leaf

Knowledge of pathogens in switchgrass, a potential biofuels crop, is limited. In December 2007, dark brown to black irregularly shaped foliar spots were observed on ‘Alamo’ switchgrass (Panicum virgatum L.) on the campus of the University of Tennessee. Symptomatic leaf samples were surface-sterilized (95% ethanol, 1 min; 20% commercial bleach, 3 min; 95% ethanol, 1 min), rinsed in sterile water, air-dried, and plated on 2% water agar amended with 3.45 mg fenpropathrin/liter (Danitol 2.4 EC, Valent Chemical, Walnut Creek, CA) and 10 mg/liter rifampicin (Sigma-Aldrich, St. Louis, MO). A sparsely sporulating, dematiaceous mitosporic fungus was observed. Fungal plugs were transferred to surface-sterilized detached ‘Alamo’ leaves on sterile filter paper in a moist chamber to increase spore production. Conidia were ovate, oblong, mostly straight to slightly curved, and light to olive-brown with 3 to 10 septa. Conidial dimensions were 12.5 to 17 × 27.5 to 95 (average 14.5 × 72) μm. Conidiophores were light brown, single, multiseptate, and geniculate. Conidial production was polytretic. Morphological characteristics and disease symptoms were similar to those described for Bipolaris oryzae (Breda de Haan) Shoemaker (2). Disease assays were done with 6-week-old ‘Alamo’ switchgrass grown from seed scarified with 60% sulfuric acid and surface-sterilized in 50% bleach. Nine 9 × 9-cm square pots with approximately 20 plants per pot were inoculated with a mycelial slurry (due to low spore production) prepared from cultures grown on potato dextrose agar for 7 days. Cultures were flooded with sterile water and rubbed gently to loosen mycelium. Two additional pots were inoculated with sterile water and subjected to the same conditions to serve as controls. Plants were exposed to high humidity by enclosure in a plastic bag for 72 h. Bags were removed, and plants were incubated at 25/20°C with 50 to 60% relative humidity. During the disease assay, plants were kept in a growth chamber with a 12-h photoperiod of fluorescent and incandescent lighting. Foliar leaf spot symptoms appeared 5 to 14 days post-inoculation for eight of nine replicates. Control plants had no symptoms. Symptomatic leaf tissue was processed and plated as described above. The original fungal isolate and the pathogen recovered in the disease assay were identified using internal transcribed spacer (ITS) region sequences. The ITS region of rDNA was amplified with PCR and primer pairs ITS4 and ITS5 (4). PCR amplicons of 553 bp were sequenced, and sequences from the original isolate and the reisolated pathogen were identical (GenBank Accession No. JQ237248). The sequence had 100% nucleotide identity to B. oryzae from switchgrass in Mississippi (GU222690, GU222691, GU222692, and GU222693) and New York (JF693908). Leaf spot caused by B. oryzae on switchgrass has also been described in North Dakota (1) and was seedborne in Mississippi (3). To our knowledge, this is the first report of B. oryzae from switchgrass in Tennessee. References: (1) D. F. Farr and A. Y. Rossman. Fungal Databases. Systematic Mycology and Microbiology Laboratory, ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/, 28 June 2012. (2) J. M. Krupinsky et al. Can. J. Plant Pathol. 26:371, 2004. (3) M. Tomaso-Peterson and C. J. Balbalian. Plant Dis. 94:643, 2010. (4) T. J. White et al. Pages 315-322 in: PCR Protocols: a Guide to Methods and Applications. M. A. Innis et al. (eds), Acad. Press, San Diego, 1990.

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First Report of Leaf Blight and Stem Canker of Pachysandra terminalis Caused by Pseudonectria pachysandricola in Korea

Plant Disease ◽

10.1094/pdis-09-11-0813 ◽

2012 ◽

Vol 96 (2) ◽

pp. 287-287

Author(s):

K. S. Han ◽

J. H. Park ◽

S. E. Cho ◽

H. D. Shin

Keyword(s):

United States ◽

Its Region ◽

Stem Canker ◽

The United States ◽

Culture Collection ◽

Leaf Blight ◽

First Report ◽

Cornell University ◽

Plastic Bags ◽

Young Leaves

Pachysandra terminalis Siebold & Zucc., known as Japanese pachysandra, is a creeping evergreen perennial belonging to the family Buxaceae. In April 2011, hundreds of plants showing symptoms of leaf blight and stem canker with nearly 100% incidence were found in a private garden in Suwon, Korea. Plants with the same symptoms were found in Seoul in May and Hongcheon in August. Affected leaves contained tan-to-yellow brown blotches. Stem and stolon cankers first appeared as water soaked and developed into necrotic lesions. Sporodochia were solitary, erumpent, circular, 50 to 150 μm in diameter, salmon-colored, pink-orange when wet, and with or without setae. Setae were hyaline, acicular, 60 to 100 μm long, and had a base that was 4 to 6 μm wide. Conidiophores were in a dense fascicle, not branched, hyaline, aseptate or uniseptate, and 8 to 20 × 2 to 3.5 μm. Conidia were long, ellipsoid to cylindric, fusiform, rounded at the apex, subtruncate at the base, straight to slightly bent, guttulate, hyaline, aseptate, 11 to 26 × 2.5 to 4.0 μm. A single-conidial isolate formed cream-colored colonies that turned into salmon-colored colonies on potato dextrose agar (PDA). Morphological and cultural characteristics of the fungus were consistent with previous reports of Pseudonectria pachysandricola B.O. Dodge (1,3,4). Voucher specimens were housed at Korea University (KUS). Two isolates, KACC46110 (ex KUS-F25663) and KACC46111 (ex KUS-F25683), were accessioned in the Korean Agricultural Culture Collection. Fungal DNA was extracted with DNeasy Plant Mini DNA Extraction Kits (Qiagen Inc., Valencia, CA). The complete internal transcribed spacer (ITS) region of rDNA was amplified with the primers ITS1/ITS4 and sequenced using ABI Prism 337 automatic DNA sequencer (Applied Biosystems, Foster, CA). The resulting sequence of 487 bp was deposited in GenBank (Accession No. JN797821). This showed 100% similarity with a sequence of P. pachysandricola from the United States (HQ897807). Isolate KACC46110 was used in pathogenicity tests. Inoculum was prepared by harvesting conidia from 2-week-old cultures on PDA. Ten young leaves wounded with needles were sprayed with conidial suspensions (~1 × 106 conidia/ml). Ten young leaves that served as the control were treated with sterile distilled water. Plants were covered with plastic bags to maintain a relative humidity of 100% at 25 ± 2°C for 24 h. Typical symptoms of brown spots appeared on the inoculated leaves 4 days after inoculation and were identical to the ones observed in the field. P. pachysandricola was reisolated from 10 symptomatic leaf tissues, confirming Koch's postulates. No symptoms were observed on control plants. Previously, the disease was reported in the United States, Britain, Japan, and the Czech Republic (2,3), but not in Korea. To our knowledge, this is the first report of P. pachysandricola on Pachysandra terminalis in Korea. Since this plant is popular and widely planted in Korea, this disease could cause significant damage to nurseries and the landscape. References: (1) B. O. Dodge. Mycologia 36:532, 1944. (2) D. F. Farr and A. Y. Rossman. Fungal Databases. Systematic Mycology and Microbiology Laboratory, ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/ , September 24, 2011. (3) I. Safrankova. Plant Prot. Sci. 43:10, 2007. (4) W. A. Sinclair and H. H. Lyon. Disease of Trees and Shrubs. 2nd ed. Cornell University Press, Ithaca, NY, 2005.

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First Report of Leaf Spot of Sweet Basil Caused by Cercospora guatemalensis in Korea

Plant Disease ◽

10.1094/pdis-05-12-0436-pdn ◽

2012 ◽

Vol 96 (10) ◽

pp. 1580-1580

Author(s):

J. H. Park ◽

K. S. Han ◽

J. Y. Kim ◽

H. D. Shin

Keyword(s):

Leaf Spot ◽

Morphological Characteristics ◽

Its Region ◽

Culture Collection ◽

Distilled Water ◽

First Report ◽

Sweet Basil ◽

Organic Farm ◽

Leaf Spots ◽

Close Phylogenetic Relationship

Sweet basil, Ocimum basilicum L., is a fragrant herb belonging to the family Lamiaceae. Originated in India 5,000 years ago, sweet basil plays a significant role in diverse cuisines across the world, especially in Asian and Italian cooking. In October 2008, hundreds of plants showing symptoms of leaf spot with nearly 100% incidence were found in polyethylene tunnels at an organic farm in Icheon, Korea. Leaf spots were circular to subcircular, water-soaked, dark brown with grayish center, and reached 10 mm or more in diameter. Diseased leaves defoliated prematurely. The damage purportedly due to this disease has reappeared every year with confirmation of the causal agent made again in 2011. A cercosporoid fungus was consistently associated with disease symptoms. Stromata were brown, consisting of brown cells, and 10 to 40 μm in width. Conidiophores were fasciculate (n = 2 to 10), olivaceous brown, paler upwards, straight to mildly curved, not geniculate in shorter ones or one to two times geniculate in longer ones, 40 to 200 μm long, occasionally reaching up to 350 μm long, 3.5 to 6 μm wide, and two- to six-septate. Conidia were hyaline, acicular to cylindric, straight in shorter ones, flexuous to curved in longer ones, truncate to obconically truncate at the base, three- to 16-septate, and 50 to 300 × 3.5 to 4.5 μm. Morphological characteristics of the fungus were consistent with the previous reports of Cercospora guatemalensis A.S. Mull. & Chupp (1,3). Voucher specimens were housed at Korea University herbarium (KUS). An isolate from KUS-F23757 was deposited in the Korean Agricultural Culture Collection (Accession No. KACC43980). Fungal DNA was extracted with DNeasy Plant Mini DNA Extraction Kits (Qiagen Inc., Valencia, CA). The complete internal transcribed spacer (ITS) region of rDNA was amplified with the primers ITS1/ITS4 and sequenced. The resulting sequence of 548 bp was deposited in GenBank (Accession No. JQ995781). This showed >99% similarity with sequences of many Cercospora species, indicating their close phylogenetic relationship. Isolate of KACC43980 was used in the pathogenicity tests. Hyphal suspensions were prepared by grinding 3-week-old colonies grown on PDA with distilled water using a mortar and pestle. Five plants were inoculated with hyphal suspensions and five plants were sprayed with sterile distilled water. The plants were covered with plastic bags to maintain a relative humidity of 100% for 24 h and then transferred to a 25 ± 2°C greenhouse with a 12-h photoperiod. Typical symptoms of necrotic spots appeared on the inoculated leaves 6 days after inoculation, and were identical to the ones observed in the field. C. guatemalensis was reisolated from symptomatic leaf tissues, confirming Koch's postulates. No symptoms were observed on control plants. Previously, the disease was reported in Malawi, India, China, and Japan (2,3), but not in Korea. To our knowledge, this is the first report of C. guatemalensis on sweet basil in Korea. Since farming of sweet basil has recently started on a commercial scale in Korea, the disease poses a serious threat to safe production of this herb, especially in organic farming. References: (1) C. Chupp. A Monograph of the Fungus Genus Cercospora. Ithaca, NY, 1953. (2) D. F. Farr and A. Y. Rossman. Fungal Databases. Systematic Mycology & Microbiology Laboratory, ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/ , May 5, 2012. (3) J. Nishikawa et al. J. Gen. Plant Pathol. 68:46, 2002.

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First Report of Anthracnose of Onion Caused by Colletotrichum coccodes in Ohio

Plant Disease ◽

10.1094/pdis-01-14-0032-pdn ◽

2014 ◽

Vol 98 (9) ◽

pp. 1271-1271 ◽

Cited By ~ 2

Author(s):

F. Baysal-Gurel ◽

N. Subedi ◽

D. P. Mamiro ◽

S. A. Miller

Keyword(s):

Leaf Tissue ◽

Its Region ◽

The United States ◽

Tween 20 ◽

Colletotrichum Coccodes ◽

Academic Press ◽

Conidial Suspension ◽

First Report ◽

Allium Cepa L ◽

Total Dna

Dry bulb onion (Allium cepa L. cvs. Pulsar, Bradley, and Livingston) plants with symptoms of anthracnose were observed in three commercial fields totaling 76.5 ha in Huron Co., Ohio, in July 2013. Symptoms were oval leaf lesions and yellowing, curling, twisting, chlorosis, and death of leaves. Nearly half of the plants in a 32.8-ha field of the cv. Pulsar were symptomatic. Concentric rings of acervuli with salmon-colored conidial masses were observed in the lesions. Conidia were straight with tapered ends and 16 to 23 × 3 to 6 μm (2). Colletotrichum coccodes (Wallr.) S. Hughes was regularly isolated from infected plants (2). Culturing diseased leaf tissue on potato dextrose agar (PDA) amended with 30 ppm rifampicin and 100 ppm ampicillin at room temperature yielded white aerial mycelia and salmon-colored conidial masses in acervuli. Numerous spherical, black microsclerotia were produced on the surface of colonies after 10 to 14 days. To confirm pathogen identity, total DNA was extracted directly from a 7-day-old culture of isolate SAM30-13 grown on PDA, using the Wizard SV Genomic DNA Purification System (Promega, Madison, WI) following the manufacturer's instructions. The ribosomal DNA internal transcribed spacer (ITS) region was amplified by PCR using the primer pair ITS1 and ITS4 (2), and sequenced. The sequence, deposited in GenBank (KF894404), was 99% identical to that of a C. coccodes isolate from Michigan (JQ682644) (1). Ten onion seedlings cv. Ebenezer White at the two- to three-leaf stage of growth were spray-inoculated with a conidial suspension (1 × 105 conidia/ml containing 0.01% Tween 20, with 10 ml applied/plant). Plants were maintained in a greenhouse (21 to 23°C) until symptoms appeared. Control plants were sprayed with sterilized water containing 0.01% Tween 20, and maintained in the same environment. After 30 days, sunken, oval lesions each with a salmon-colored center developed on the inoculated plants, and microscopic examination revealed the same pathogen morphology as the original isolates. C. coccodes was re-isolated consistently from leaf lesions. All non-inoculated control plants remained disease-free, and C. coccodes was not re-isolated from leaves of control plants. C. coccodes was reported infecting onions in the United States for the first time in Michigan in 2012 (1). This is the first report of anthracnose of onion caused by C. coccodes in Ohio. Unusually wet, warm conditions in Ohio in 2013 likely contributed to the outbreak of this disease. Timely fungicide applications will be necessary to manage this disease in affected areas. References: (1) A. K. Lees and A. J. Hilton. Plant Pathol. 52:3. 2003. (2) L. M. Rodriguez-Salamanca et al. Plant Dis. 96:769. 2012. (3) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990.

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First Report of Fruit Rot of Melon Caused by Fusarium asiaticum in China

Plant Disease ◽

10.1094/pdis-08-20-1857-pdn ◽

2020 ◽

Author(s):

Fangmin Hao ◽

Quanyu Zang ◽

Weihong Ding ◽

Erlei Ma ◽

Yunping Huang ◽

...

Keyword(s):

Disease Incidence ◽

Elongation Factor ◽

Morphological Characteristics ◽

Its Region ◽

Fruit Rot ◽

Post Inoculation ◽

Basal Cells ◽

First Report ◽

Fusarium Asiaticum ◽

Sterile Needle

Melon (Cucumis melo L.) is a member of the Cucurbitaceae family, an important economical and horticultural crop, which is widely grown in China. In May 2020, fruit rot disease with water-soaked lesions and pink molds on cantaloupe melons was observed in several greenhouses with 50% disease incidence in Ningbo, Zhejiang Province in China. In order to know the causal agent, diseased fruits were cut into pieces, surface sterilized for 1 min with 1% sodium hypochlorite (NaClO), 2 min with 75% ethyl alcohol, rinsed in sterile distilled water three times (Zhou et al. 2018), and then placed on potato dextrose agar (PDA) medium amended with streptomycin sulfate (100 μg/ml) plates at 25°C for 4 days. The growing hyphae were transferred to new PDA plates using the hyphal tip method, putative Fusarium colonies were purified by single-sporing. Twenty-five fungal isolates were obtained and formed red colonies with white aerial mycelia at 25°C for 7 days, which were identified as Fusarium isolates based on the morphological characteristics and microscopic examination. The average radial mycelial growth rate of Fusarium isolate Fa-25 was 11.44 mm/day at 25°C in the dark on PDA. Macroconidia were stout with curved apical and basal cells, usually with 4 to 6 septa, and 29.5 to 44.2 × 3.7 to 5.2 μm on Spezieller Nährstoffarmer agar (SNA) medium at 25°C for 10 days (Leslie and Summerell 2006). To identify the species, the internal transcribed spacer (ITS) region and translational elongation factor 1-alpha (TEF1-α) gene of the isolates were amplified and cloned. ITS and TEF1-α was amplified using primers ITS1/ITS4 and EF1/EF2 (O’Donnell et al. 1998), respectively. Sequences of ITS (545 bp, GenBank Accession No. MT811812) and TEF1-α (707 bp, GenBank Acc. No. MT856659) for isolate Fa-25 were 100% and 99.72% identical to those of F. asiaticum strains MSBL-4 (ITS, GenBank Acc. MT322117.1) and Daya350-3 (TEF1-α, GenBank Acc. KT380124.1) in GenBank, respectively. A phylogenetic tree was established based on the TEF1-α sequences of Fa-25 and other Fusarium spp., and Fa-25 was clustered with F. asiaticum. Thus, both morphological and molecular characterizations supported the isolate as F. asiaticum. To confirm the pathogenicity, mycelium agar plugs (6 mm in diameter) removed from the colony margin of a 2-day-old culture of strain Fa-25 were used to inoculate melon fruits. Before inoculation, healthy melon fruits were selected, soaked in 2% NaClO solution for 2 min, and washed in sterile water. After wounding the melon fruits with a sterile needle, the fruits were inoculated by placing mycelium agar plugs on the wounds, and mock inoculation with mycelium-free PDA plugs was used as control. Five fruits were used in each treatment. The inoculated and mock-inoculated fruits were incubated at 25°C with high relative humidity. Symptoms were observed on all inoculated melon fruits 10 days post inoculation, which were similar to those naturally infected fruits, whereas the mock-inoculated fruits remained symptomless. The fungus re-isolated from the diseased fruits resembled colony morphology of the original isolate. The experiment was conducted three times and produced the same results. To our knowledge, this is the first report of fruit rot of melon caused by F. asiaticum in China.

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First Report of Leaf Blight Caused by Nigrospora sphaerica on Curcuma in China

Plant Disease ◽

10.1094/pdis-04-11-0284 ◽

2011 ◽

Vol 95 (9) ◽

pp. 1190-1190

Author(s):

L. X. Zhang ◽

J. H. Song ◽

G. J. Tan ◽

S. S. Li

Keyword(s):

Disease Incidence ◽

Its Region ◽

Leaf Blight ◽

Morphological Data ◽

Future Research ◽

Leaf Margin ◽

Academic Press ◽

First Report ◽

Pathogenicity Tests ◽

Leaf Pathogen

Curcuma (family Zingiberaceae) is commonly cultivated for the use of rhizomes within traditional Chinese medicines. In October 2009 and 2010, severe leaf blight was observed on Curcuma wenyujin Y.H. Chen & C. Ling (4) in fields located in Ruian, China. The area of cultivation in Ruian encompasses 90% of the production in Zhejiang Province. Disease incidence was approximately 90% of plants observed in affected fields. Early symptoms were yellow-to-brown, irregular-shaped lesions on the leaf margin or tip. After several days, lesions expanded along the mid-vein until the entire leaf was destroyed. Blighted leaves turned grayish to dark brown and withered, and severely affected plants died. Eight fungal isolates were recovered from symptomatic C. wenyujin leaves, collected from eight different fields, on potato dextrose agar (PDA). These fungal colonies were initially white, becoming light to dark gray and produced black, spherical to subspherical, single-celled conidia (14 to 17 × 12 to 15 μm), which were borne on a hyaline vesicle at the tip of the conidiophores. On the basis of these morphological features, the isolates appeared to be similar to Nigrospora sphaerica (2). Strain ZJW-1 was selected as a representative for molecular identification. Genomic DNA was extracted from the isolate, and the internal transcribed spacer (ITS) region of the ribosomal DNA (ITS1-5.8S-ITS2) was amplified using ITS1 (5′-TCCGTAGGTGAACCTGCGG-3′) and ITS4 (5′-TCCTCCGCTTATTGATATGC-3′) primers (3). The ITS region was further cloned and sequenced (GenBank Accession No. JF738028) and was 99% identical to N. sphaerica (GenBank Accession No. FJ478134.1). On the basis of morphological data and the ITS rDNA sequence, the isolate was determined to be N. sphaerica. Pathogenicity tests were conducted on four leaves of four C. wenyujin plants by placing agar pieces (5 mm in diameter) from 8-day-old cultures on pushpin-wounded leaves. An equal number of control plants were wounded and inoculated with noncolonized PDA agar pieces. Plants were placed in moist chambers at 25°C with a 12-h photoperiod. Brown-to-black lesions were observed on wounded leaves after 3 days and expanded to an average of 56 × 40 mm 15 days after inoculation. No symptoms developed on the control leaves. The pathogen was reisolated from the margins of necrotic tissues but not from the controls. The pathogen has been reported as a leaf pathogen on several hosts worldwide (1). To our knowledge, this is the first report of N. sphaerica as a leaf pathogen of C. wenyujin in China. Future research will focus primarily on management of this disease. References: (1) D. F. Farr and A. Y. Rossman. Fungal Databases, Systematic Mycology and Microbiology Laboratory, USDA-ARS, Retrieved from http://nt.ars-grin.gov/fungaldatabases/ , March 31, 2011. (2) E. W. Mason. Trans. Brit. Mycol. Soc. 12:152, 1927. (3) T. J. White et al. PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990. (4) J. Zhao et al. Molecules 15:7547, 2010.

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First Report of Alternaria heveae Causing Black Leaf Spot of Rubber Tree in China

Plant Disease ◽

10.1094/pdis-01-14-0065-pdn ◽

2014 ◽

Vol 98 (7) ◽

pp. 1011-1011 ◽

Cited By ~ 1

Author(s):

Z. Y. Cai ◽

Y. X. Liu ◽

G. X. Huang ◽

M. Zhou ◽

G. Z. Jiang ◽

...

Keyword(s):

Rubber Tree ◽

Morphological Characteristics ◽

Black Spot ◽

Its Region ◽

Post Inoculation ◽

First Report ◽

Leaf Spots ◽

Plastic Bags ◽

Manual Pressure ◽

Tree Leaf

Rubber tree (Hevea brasiliensis Muell. Arg.) is an important industrial crop of tropical areas for natural rubber production. In October 2013, foliar spots (0.1 to 0.4 mm in diameter), black surrounded by a yellow halo, and with lesions slightly sunken were observed on the rubber tree leaf in a growing area in Heikou County of Yunnan Province. Lesion tissues removed from the border between symptomatic and healthy tissue were surface sterilized in 75% ethanol and air-dried, plated on PDA plates, and incubated at 28°C with alternating day/night cycles of light. The pathogen was observed growing out of many of the leaf pieces, and produced abundant conidia. Colonies 6.1 cm in diameter developed on potato carrot agar (PCA) after 7 days, with well-defined concentric rings of growth. Colonies on PCA were composed of fine, dark, radiating, surface and subsurface hyphae. Conidia produced in PCA culture were mostly solitary or in short chains of 2 to 5 spores, long ovoid to clavate, and light brown, 40 to 81.25 × 8 to 20 μm (200 colonies were measured), with 3 to 6 transverse septa and 0 to 2 longitudinal or oblique septa. Morphological characteristics were similar to those described for Alternaria heveae (3,4). A disease of rubber tree caused by Alternaria sp. had been reported in Mexico in 1947 (2). DNA of Ah01HK13 isolate was extracted for PCR and sequencing of the ITS region with ITS1 and ITS4 primers was completed. From the BLAST analysis, the sequence of Ah01HK13 (GenBank Accession No. KF953884), had 97% similarity to A. dauci, 96% identical to A. macrospora (AY154701.1 and DQ156342.1, respectively), indicating the pathogen belonged to Alternaria genus. According to morphological characteristics, this pathogen was identified as A. heveae. Pathogenicity of representative isolate, Ah01HK13 was confirmed using a field rubber tree inoculation method. Three rubber plants (the clone of rubber tree Yunyan77-4) were grown to the copper-colored leaf stage and inoculated by spraying spore suspension (concentration = 104 conidia/ml) to the copper-colored leaves until drops were equally distributed on it using manual pressure sprayer. Three rubber plants sprayed with sterile distilled water were used as controls. After inoculation, the plants were covered with plastic bags. The plastic bags were removed after 2 days post-inoculation (dpi) and monitored daily for symptom development (1). The experiment was repeated three times. The typical 0.1 to 0.4 mm black leaf spots were observed 7 dpi. No symptoms were observed on control plants. A fungus with the same colony and conidial morphology as A. heveae were re-isolated from leaf lesions on inoculated rubber plants, but not from asymptomatic leaves of control plants, fulfilling Koch's postulates. Based on these results, the disease was identified as black spot of rubber tree caused by A. heveae. To our knowledge, this is the first report of A. heveae on rubber tree in China. References: (1) Z. Y. Cai et al. Microbiol Res. 168:340, 2013. (2) W. J. Martin. Plant Dis. Rep. 31:155, 1947. (3) E. G. Simmons. Mycotaxon 50:262, 1994. (4) T. Y. Zhang. Page 111 in: Flora Fungorum Sinicorum: Alternaria, Science Press, Beijing, 2003.

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First Report of Internal Black Rot Caused byNeoscytalidium dimidiatumonHylocereus undatus(Pitahaya) Fruit in Israel

Plant Disease ◽

10.1094/pdis-05-13-0535-pdn ◽

2013 ◽

Vol 97 (11) ◽

pp. 1513-1513 ◽

Cited By ~ 9

Author(s):

D. Ezra ◽

O. Liarzi ◽

T. Gat ◽

M. Hershcovich ◽

M. Dudai

Keyword(s):

Its Region ◽

Stem Canker ◽

Brown Spot ◽

Black Rot ◽

Post Inoculation ◽

Single Spore ◽

Flower Opening ◽

Disease Symptoms ◽

First Report ◽

Β Tubulin

Pitahaya (Hylocereus undatus [Haw.] Britton & Rose) was introduced to Israel in 1994, and is grown throughout the country. In the summer of 2009, fruit with internal black rot was collected from a field in central Israel. Symptomatic tissue from the black rot was placed on potato dextrose agar (PDA) plates amended with 12 μg/ml tetracycline and incubated at 25°C for 3 days. A dark, gray to black, fast-growing fungus was isolated from all samples (10 fruits). For identification, single-spore cultures were grown on PDA at 25°C for 5 days, and colonies with gray to black, wooly mycelium were formed. The mycelia were branched and septate (4 to 8 μm wide). The arthroconidia were dark brown, thick-walled, and one-celled, 6.3 to 14.2 × 2.0 to 4.5 μm (n = 5), and ovate to rectangular. Based on these characteristics, the fungus was identified as Neoscytalidium dimidiatum (Penz.) Crous & Slippers (1). The internal transcribed spacer (ITS) region of rDNA and β-tubulin gene were amplified using ITS1 and ITS4, T121 (2), and Bt1b (3) primers, respectively, and then sequenced (GenBank Accessions KF000372 and KF020895, respectively). Both sequences were identical to sequences previously deposited in GenBank. The ITS (561 bp) and β-tubulin (488 bp) sequences exhibited 99% and 100% identity, and 100% and 84% coverage, respectively, to N. dimidiatum (JX524168 and FM211185, respectively). Thus, the results of the molecular identifications confirmed the morphological characterization. To establish fungal pathogenicity and the mechanism of infection, 60 flowers in a disease-free orchard were marked to form three different treatments (15 flowers per treatment): inoculations of the flower tube by inserting PDA plugs (0.5 × 0.5 cm) from a 5-day-old culture to the base of the flower, inoculations of the flower stigma by placing the fungus plug on intact, or pre-wounded flower stigma. The wounds were made by scratching the stigma with a sterile scalpel. For each treatment, five additional flowers were used as negative controls in which the PDA plugs did not contain any fungus. All flowers were hand-pollinated and left to grow for a month until the fruit had ripened. Only flowers inoculated by insertion of the fungus into the flower tube developed black rot in the fruit (8 of 15 fruit) 3 to 4 weeks post inoculation, suggesting involvement of the flower tube in the mechanism of infection. All other treatments and controls failed to develop any detectable disease symptoms. N. dimidiatum was reisolated from the rot, fulfilling Koch's postulates. Flowers with wounded stigma developed significantly smaller fruit. Interestingly, diseased fruit changed color about a week before ripening from the flower opening downwards, whereas healthy fruit changed color from the attachment point to the stem upwards. These results indicate that N. dimidiatum is the pathogen of pitahaya internal black rot disease. Recently, this pathogen was reported to cause brown spot disease and stem canker disease of pitahaya in China (4) and Taiwan (5), respectively. To date, the disease can be detected in all orchards in Israel, with up to 50% of the fruit being infected. Since the disease symptoms of the Israeli isolate are located in the fruit, the commercial loss due to pathogen attack is significant. To our knowledge, this is the first report of internal black rot caused by N. dimidiatum on pitahaya fruit in Israel.References: (1) P. W. Crous et al. Stud. Mycol. 55:235, 2006. (2) K. O'Donnell and E. Cigelnik. Mol. Phylo, Evol. 7:103, 1997. (3) N. L. Glass and G. C. Donaldson. Appl. Environ. Microiol. 61:1323, 1995. (4) G. B. Lan and Z. F. He. Plant Dis. 96:1702, 2012. (5) M. F. Chuang et al. Plant Dis. 96:906, 2012.

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First Report of Diaporthe australafricana Associated with Stem Canker on Blueberry in Chile

Plant Disease ◽

10.1094/pdis-12-11-1025-pdn ◽

2012 ◽

Vol 96 (5) ◽

pp. 768-768 ◽

Cited By ~ 3

Author(s):

B. A. Latorre ◽

K. Elfar ◽

J. G. Espinoza ◽

R. Torres ◽

G. A. Díaz

Keyword(s):

Vitis Vinifera ◽

Vascular Tissue ◽

Its Region ◽

Pathogenic Fungi ◽

Stem Canker ◽

Vaccinium Corymbosum ◽

Academic Press ◽

First Report ◽

Potted Plants ◽

Blastn Analysis

Stem cankers of blueberry (Vaccinium corymbosum L.) have been observed on as much as 15% of the plants in plantations in central and southern Chile since 2006. Symptoms consisted of apical necrosis of the shoots and brown-to-reddish necrotic lesions on the stems. Internally, a brown-to-reddish discoloration of the vascular tissue can be observed. Twenty, single-plant samples were collected in 12 blueberry plantings (approximately 33°27′ to 40°53′S). Isolations from the margins of the necrotic lesions on the stems were made by plating small pieces (5 mm) on potato dextrose agar acidified with 0.5 μl/ml of 92% lactic acid (APDA). The plates were incubated at 20°C for 5 to 7 days, and hyphal tips of white colonies with septate and hyaline mycelium were transferred to APDA. Colonies were then transferred to autoclaved Pinus radiata needles on 2% water agar and incubated for 20 days at 20°C. Twelve isolates producing black pycnidia and alpha conidia were tentatively identified as a Phomopsis sp. (teleomoph Diaporthe Nitschke). Other fungi, including Botryosphaeriaceae spp. and Pestalotiopsis spp., were also isolated. Alpha conidia were smooth, unicellular, hyaline, fusoid, biguttulate, and 6.4 to 7.9 × 2.3 to 3.3 μm (n = 20). Beta conidia were not observed. The internal transcribed spacer (ITS) region of the rDNA was amplified using primers ITS1 and ITS2 (4) and sequenced. BLASTn analysis of the 473-bp fragment (GenBank Accession No. JQ045712) showed 100% identity to Diaporthe australafricana Crous & J.M. van Niekerk from Vitis vinifera (3). The pathogenicity of D. australafricana was studied on blueberry cv. O'Neal using detached stems (n = 4) in the laboratory, on 2-year-old potted plants (n = 4) in a greenhouse, and on attached stems of mature plants (n = 4) established in the ground. Inoculations were done by placing mycelial plugs taken from 7-day-old APDA cultures in a 7-mm long incision made on the stems. Inoculations with sterile mycelium plugs served as negative controls. Inoculation sites were wrapped with Parafilm to avoid rapid dehydration. Dark brown, necrotic lesions on the internal tissues were obtained on all inoculated stems 15 days after inoculation. Mean lesion lengths were 18.0 ± 7.4 mm on detached stems, 7.8 ± 6.9 mm on stems of 2-year-old plants, and 7.3 ± 2.5 mm on mature plants in the field. No symptoms developed on control stems. Reisolations were successful in 100% of the inoculated stems and D. australafricana was confirmed by the presence of pycnidia and alpha conidia. To our knowledge, this is the first report of D. australafricana causing stem canker in V. corymbosum. Previously, this pathogen has been reported to be affecting Vitis vinifera in Australia and South Africa (3). These results do not exclude that other plant-pathogenic fungi may be involved in this syndrome (1,2). References: (1) J. G. Espinoza et al. Plant Dis 92:1407, 2008. (2) J. G. Espinoza et al. Plant Dis. 93:1187, 2009. (3) J. M. van Niekerk et al. Australas. Plant Pathol. 34:27, 2005. (4) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, NY, 1990.

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