First Report of Botryosphaeria rhodina Causing Shoot Blight of Pistachio in California

In the summers of 2000 and 2001, shoot blight was observed in pistachios (Pistacia vera L.) grown in Kern County, California. Black, necrotic lesions developed at the base of shoots originating from contaminated or partially infected buds. Infection moved upward resulting in a progressive wilting and blighting of leaves. Leaf blades on infected shoots withered, and petioles became necrotic. Symptoms have been considered characteristic of infection by Botryosphaeria dothidea (Moug.:Fr.) Ces. & de Not., but this pathogen causes panicle and shoot blight of pistachio (1). However, there were no symptoms of any fruit panicle infections on trees we observed. Isolations on acidified potato dextrose agar from the base of blighted shoots in both years revealed a fast-growing fungus producing pycnidia which was identified as the anamorph Lasiodiplodia theobromae (Pat.) Griffon & Maubl. of B. rhodina Berk. & Curt. Arx. Identification of the pathogen was based on characteristic dark brown, oval pycnidiospores with striations on the surface of the spore along the long axis. Pathogenicity tests were performed on 12 Kerman pistachio trees grown at Kearney Agricultural Center, in Parlier, CA, using three isolates recovered from pistachios grown in two locations. Six to 16 current season shoots of pistachio trees (1 to 2 shoots per tree) were wounded with a 5-mm-diameter cork borer, and a mycelial plug of 5-day-old cultures of B. rhodina was inserted in each wound. Shoots were wrapped with Parafilm to prevent desiccation of inoculum. Six other shoots (one per tree) were inoculated similarly with mycelial agar plugs of a pistachio isolate of B. dothidea and served as positive controls, while six similar shoots were inoculated with only agar plugs and served as negative controls. Wilting of lower leaves in the majority of inoculated shoots started within 4 days for B. rhodina and 7 days for B. dothidea. Depending on the isolate of B. rhodina, 1 to 5 shoots and 50 to 80% of leaves were blighted within 7 days after inoculation. All inoculated shoots were left on the trees until 3 to 4 months after inoculation, pruned and assessed again. For inoculations done in September 2001, 33 to 71% of shoots were blighted, and the rest had cankers ranging from 22.5 to 28 mm long and 13.5 to 23.5 mm wide. A majority (67 to 100%) of shoots had pycnidia of the pathogen present. For inoculations done in October 2001, none of the shoots was blighted, but cankers ranged from 5 to 55.4 mm long and 6 to 22 mm wide and 33.3 to 100% developed pycnidia. B. rhodina was isolated from all inoculated shoots but not from negative controls or those inoculated with B. dothidea. Inoculations of shoots with B. dothidea produced similar symptoms as those of B. rhodina. Shoots that served as negative controls did not develop symptoms. Because panicle and shoot blight of pistachio caused by B. dothidea has developed to epidemic levels in commercial pistachio orchards and is of concern to the pistachio industry in California, it would be of interest to monitor how much shoot blight caused by B. rhodina would eventually develop over the years in commercial pistachio orchards. A survey was initiated in 2002 to determine how widespread B. rhodina is in California pistachios. To our knowledge, this is the first report worldwide of B. rhodina causing shoot blight of pistachio. Reference: (1) T. Michailides. Panicle and shoot blight. Page 68 in: Compendium of Nut Crop Diseases in Temperate Zones. B. L. Teviotdale, T. J. Michailides, and J. W. Pscheidt, eds. American Phytopathological Society, St. Paul, MN 2002.

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First Report of Aspergillus Vine Canker of Table Grapes Caused by Aspergillus niger

Plant Disease ◽

10.1094/pdis.2002.86.1.75a ◽

2002 ◽

Vol 86 (1) ◽

pp. 75-75 ◽

Cited By ~ 6

Author(s):

T. J. Michailides ◽

W. Peacock ◽

P. Christensen ◽

D. P. Morgan ◽

D. Felts

Keyword(s):

Aspergillus Niger ◽

Potato Dextrose Agar ◽

American Phytopathological Society ◽

Current Season ◽

First Report ◽

Table Grapes ◽

Lateral Shoots ◽

Mycelial Plug ◽

Cork Borer ◽

Bunch Rot

A vine canker was first observed in the San Joaquin Valley, CA, in fall 1989, on exceptionally vigorous 1-year-old cv. Redglobe vines (Vitis vinifera) when vines were trained up the stakes. Since 1989, the same canker symptoms have been observed in Tulare, Kern, Fresno, and Riverside (Coachella Valley, CA) counties on cv. Redglobe, Crimson Seedless, Chardonnay, and Grenache vines. In affected vineyards, the disease resulted in the retraining of 2 to 6.1% of vines the following spring, using a shoot originating from below the canker. In a sample of 54 infected vines collected in 1997, 65% of cankers were found at the branching (crotch) of the vine, 24% along the shoot, or both (11%). All infections started through wounds caused by removing lateral shoots or leaves when the vine was topped to form cordons or possibly through growth cracks that occur on rapidly growing 1-year-old shoots. The first symptoms usually appear in August as red pinhead-size drops of sap on the surface of discolored tissue. By October to November, the canopies of vines girdled by the canker prematurely display fall colors and are very distinct from healthy vines. The trunk is slightly swollen and spongy where the canker occurs. Internal canker tissue is discolored and dead. Black spores are abundant within the canker, on the surface of the canker, or both. Callous tissue is often associated with the canker as the vine attempts to repair the damage with new tissue. Canker length can range from 3.5 to 26.5 cm (average 7.0 cm) and can affect the shoot's cross section from 0.4 cm to completely girdling the shoot (up to 9.0 cm in circumference). Isolations from cankers or black sporulation inside the canker on acidified potato dextrose agar (APDA) consistently yielded Aspergillus niger van Tiegh. Six well-matured current-season canes of cv. Redglobe in an experimental vineyard at Kearney Agricultural Center were inoculated by inserting a 7-mm plug of mycelium from actively growing cultures on APDA in a cut made with a 7-mm cork borer or by brushing spores of the culture over the surface of six canes wounded with a sterile razor. Six canes were inoculated with a 7-mm plug of APDA and used as noninoculated controls. Inoculated sites were sealed with Parafilm to avoid dehydration. Inoculation of grapevines with A. niger resulted in cankers similar to those observed in commercial vineyards 5 months after inoculation. Cankers ranged from 2.4 to 4.2 cm for mycelial-plug inoculation (100% of canes infected) and 2.3 to 7.3 cm for spore-brushing inoculation (67% infected). Noninoculated control canes were not infected. In another experiment, inoculation of 10 canes each with A. niger on 17 May, 10 June, 2 July, 21 July, and 16 August resulted in 50, 60, 90, 90, and 100% canker formation, respectively, 5 to 8 months after inoculation, suggesting summer inoculations were more effective than spring inoculations. Reisolation from infected canes on APDA revealed A. niger. Aspergillus species in section Nigri have been reported to be among the pathogens involved in the bunch rot complex (1,2), but to our knowledge, this is the first report of A. niger causing a serious canker of vigorously growing grape vines. References: (1) W. B. Hewitt. Berry rots and raisin molds. Pages 26–28 in: Compendium of Grape Diseases. R. C. Pearson and A. C. Gohen, eds. The American Phytopathological Society, St. Paul, MN, 1994. (2) W. R. Jarvis and J. A. Traquair. Plant Dis. 68:718, 1984.

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A Severe Outbreak of Charcoal Rot in Cantaloupe Melon Caused by Macrophomina phaseolina in Chile

Plant Disease ◽

10.1094/pdis-06-12-0588-pdn ◽

2013 ◽

Vol 97 (1) ◽

pp. 141-141 ◽

Cited By ~ 7

Author(s):

C. J. Jacob ◽

C. Krarup ◽

G. A. Díaz ◽

B. A. Latorre

Keyword(s):

Macrophomina Phaseolina ◽

Morphological Identification ◽

Internal Transcribed Spacer Region ◽

Charcoal Rot ◽

Agar Plug ◽

Fruit Decay ◽

Mycelial Plug ◽

Pathogenicity Tests ◽

Cork Borer ◽

Cucumis Melo L

A severe outbreak of charcoal rot was observed in cantaloupe melon (Cucumis melo L.) in the summer of 2011 to 2012 in Curacaví Valley, Chile. Prior to harvest, of 72 plants per cultivar, charcoal rot prevalence varied from 32% to 82% in cvs. Colima, Charantias, Navigator, Origami, Otero, and Samoa. Symptoms were wilting and leaf browning associated with water-soaked lesions at the base of the crown with amber to dark brown exudates. Lesions dried out progressively, turned tan, and cracked. Affected plants declined and died before harvest. Reddish fruit decay was observed. Symptomatic stem and root samples (n = 97) were collected, surface disinfected (96% ethanol, 30 s), plated on PDA acidified with 0.5 ml/liter of 92% lactic acid (APDA), and incubated at 20 ± 1°C. A white, fast-growing mycelium was obtained that turned gray to black after 7 days due to the presence of spherical to oblong black microsclerotia 136 ± 52 μm (n = 80) in diameter. On the basis of colony morphology and microsclerotia, 57 isolates (59%), obtained from 97 melon samples, were tentatively identified as Macrophomina phaseolina (Tassi) Goid. (2,3). The morphological identification of four isolates M1HB-B, M2CO-B, M3CH-R, and M4OT-B (GenBank Accession Nos. JX203630, JX203631, JX203632, and JX203633) was confirmed by sequencing of the internal transcribed spacer region (ITS1-5.8S-ITS2) of rDNA, using primers ITS4 and ITS5, with >99% similarity with the sequences of M. phaseolina (GenBank Accession No. HQ660592) (4). Pathogenicity tests were conducted with isolates M1HB-B, M2CO-B, M3CH-R, and M4OT-B on melon fruits cvs. Colima, Origami, Charantias, and Diva. Four mature melon fruits per cultivar per isolate were surface disinfected with 0.5% sodium hypochlorite for 2 min before inserting a mycelium plug (19 mm2) in a 6 mm diameter hole made with a sterile cork borer. An equal number of perforated fruits in which a sterile agar plug was inserted were left as non-inoculated controls. After 8 days of incubation at 20°C, inoculated fruits developed a spherical, reddish, soft necrotic lesion of 15 to 20 mm in diameter in the pulp. Non-inoculated fruits remained symptomless. The pathogenicity of the four isolates was also studied in 3-month-old melon plants (n = 4) cvs. Colima and Navigator. Plants were inoculated by inserting a mycelial plug (9 mm2) underneath the epidermis of the crown, 5 cm above the soil level. The inoculation site was immediately wrapped with Parafilm to avoid dehydration. An equal number of non-inoculated, but injured plants, treated with a sterile agar plug, were left as controls. After 21 days of incubation under greenhouse conditions (17 ± 5.5°C), all inoculated plants developed water-soaked to dry necrotic lesions, 20 to 70 mm long, yellow to tan in color. No symptoms were obtained in non-inoculated controls. M. phaseolina was reisolated in 84% and 100% of the inoculated plants and fruits, respectively. To our knowledge, this study is the first report of charcoal rot in cantaloupe melon in Chile, previously found on watermelon and melon group inodorus (1). Charcoal rot appears as an emerging disease that aggressively affects current cantaloupe melon cultivars in central Chile. References: (1) G. Apablaza. Cien. Inv. Agr. 20:101, 1993. (2) B. D. Bruton and E. V. Wann. Charcoal rot. Page 9 in: Compendium of Cucurbit Diseases. T. A. Zitter, D. L. Hopkins, and C. E. Thomas, eds. APS, St. Paul, MN, 1996. (3) S. Kaur et al. Crit. Rev. Microbiol. 38:136, 2012. (4) J. Q. Zhang et al. Plant Dis. 95:872, 2011.

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First Report of Fusarium solani Fruit Rot of Pumpkin (Cucurbita pepo) in Trinidad

Plant Disease ◽

10.1094/pdis-93-5-0547b ◽

2009 ◽

Vol 93 (5) ◽

pp. 547-547

Author(s):

S. N. Rampersad

Keyword(s):

Fusarium Solani ◽

Cucurbita Pepo ◽

Morphological Characteristics ◽

Soft Rot ◽

The United States ◽

Fruit Rot ◽

Laboratory Manual ◽

First Report ◽

Pathogenicity Tests ◽

Negative Controls

Trinidad is a major exporter of pumpkins (Cucurbita pepo L.) to other Caribbean countries, Canada, and the United States. Producers and exporters have reported 50 to 80% yield losses because of soft rot and overnight collapse of fruit at the pre- and postharvest stages. Severe fruit rot occurred in fields in Victoria County in South Trinidad between April and May 2006 (mid-to-late dry season) with an increase in the severity and number of affected fruit in the rainy season (July to December). Symptoms began as water-soaked lesions on the fruit of any age at the point of contact with the soil. The disease progressed to a soft rot with leakage and whole fruit collapse. A dark brown, soft decay also developed at the base of the main vines. Fusarium solani was isolated on selective fusarium agar and potato dextrose agar (PDA) (1) after 7 to 10 days of incubation at 25°C. The pathogen was identified by morphological characteristics and pathogenicity tests. Colonies were fast growing with white aerial mycelia and a cream color on the reverse side; hyphae were septate and hyaline, conidiophores were unbranched, and microconidia were abundant, thin walled, hyaline, fusiform to ovoid, generally one to two celled, and 8 to 10 × 2 to 4 μm. Macroconidia were hyaline, two to three celled, moderately curved, thick walled, and 25 to 30 × 4 to 6 μm. Pathogenicity tests for 10 isolates were conducted on 2-week-old pumpkin seedlings (cv. Jamaican squash; seven plants per isolate) and mature pumpkin fruit (2). Briefly, seedlings were inoculated by dipping their roots in a spore suspension (1 × 104 spores per ml) for 20 min. The plants were repotted in sterile potting soil. For negative controls, plant roots were dipped in sterile water. After the rind of fruit was swabbed with 70% ethanol followed by three rinses with sterile distilled water, 0.4-cm-diameter agar plugs of the isolates were inserted into wounds made with a sterile 1-cm-diameter borer. Sterile PDA plugs served as negative controls. Fruit were placed in sealed, clear, plastic bags. Inoculated plants and fruit were placed on greenhouse benches (30 to 32°C day and 25 to 27°C night temperatures) and monitored over a 30-day period. Tests were repeated once. Inoculated fruit developed a brown, spongy lesion that expanded from the initial wound site over a period of approximately 17 days after inoculation. White mycelia grew diffusely over the lesion. Inoculated plants developed yellow and finally necrotic leaves and lesions developed on stems at the soil line approximately 21 days after inoculation. No symptoms developed on the control plants. The fungus was reisolated from symptomatic tissue, fulfilling Koch's postulates. To my knowledge, this is the first report of Fusarium fruit rot of pumpkin in Trinidad. References: (1) J. Leslie and B. Summerell. Page 1 in: The Fusarium Laboratory Manual. Blackwell Publishing, Oxford, 2006. (2) W. H. Elmer. Plant Dis. 80:131, 1996.

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First Report of Lasidiplodia theobromae and Cryptovalsa ampelina Associated with Grapevine Decline from Castilla y León, Spain

Plant Disease ◽

10.1094/pdis-93-5-0545c ◽

2009 ◽

Vol 93 (5) ◽

pp. 545-545 ◽

Cited By ~ 4

Author(s):

M. T. Martin ◽

L. Martin ◽

M. T. de-Francisco ◽

R. Cobos

Keyword(s):

Dna Sequences ◽

Morphological Characteristics ◽

Tubulin Gene ◽

Current Season ◽

First Report ◽

Grapevine Decline ◽

Eutypa Dieback ◽

Castilla Y Leon ◽

Pathogenicity Tests ◽

Β Tubulin

Symptoms of grapevine decline were surveyed. Samples from mature vines exhibiting external symptoms of Eutypa dieback and Esca were collected, as were young plants with and without external symptoms, and fungal isolations were performed. In 2007, 3-year-old grapevines (cv. Tempranillo grafted onto 110R rootstock) with low vigor, reduced foliage, and vascular streaking in the wood were observed. Small pieces of discolored wood were placed onto malt extract agar supplemented with 0.25 g/liter of chloramphenicol, incubated at 25°C, and resulting colonies were transferred to potato dextrose agar (PDA). Isolates were characterized by abundant aerial and fast-growing mycelium covering the plate surface after 3 days, mycelium became dark green. Pycnidia contained thick-walled, aseptate conidia 15 to 35 × 10 to 15 μm. Lasidiplodia theobromae was identified based on morphological characteristics (3) and confirmed by banding patterns obtained after the digestion of the 1,200-bp amplicon generated with ITS1 and NL4 primers (2) using restriction endonucleases (2). Single-spore cultures were generated and DNA sequences of the rDNA internal transcribed spacer region, partial sequence of the 5′ end of the β-tubulin gene, and a fragment of the elongation factor further confirmed the identification and revealed genetic similarity with other isolates of L. theobromae. A sequence of each fragment was deposited in GenBank with Accession Nos. EU600925, EU597297, and EU597298, respectively. Pathogenicity tests were conducted on four replicate rootstocks (110R) and 15 canes of current-season growth (cv. Tempranillo). Plants were inoculated with an agar plug containing L. theobromae; controls were treated with agar only. Grapevines were maintained in a greenhouse at 20 to 25°C. After 3 months, L. theobromae was reisolated from internal vascular lesions in 100 and 66% of inoculated rootstocks and canes, respectively. Control plants were asymptomatic and L. theobromae was not recovered. Using the same methodology, a fungus identified based on morphological characteristics in culture as Cryptovalsa ampelina (1) was isolated from grapevines (cv. Tempranillo) planted in 1987. Cultures in PDA were white to creamy white and cottony with diffuse margins. Colonies covered the 90-mm-diameter petri dish surface in 5 days. Conidia were 20 to 23 × 1 to 1.5 μm, unicellular, hyaline, and filiform. PCR amplifications of the DNA extracts of C. ampelina with Camp-1 and Camp-2R primers gave a characteristic DNA fragment of 300 bp (3) and DNA sequences of the ITS4-ITS5 amplicons (GenBank Accession No. EU597296) confirmed the identification. For the first time, the 5′ end of the β-tubulin gene was sequenced and deposited in GenBank (Accession No. EU600926). Pathogenicity tests were conducted as described above for L. theobromae. Both pathogens were examined in the same experiment. C. ampelina was reisolated from internal brown streaking lesions in 25% of the rootstocks and 33% of the canes. Control plants exhibited no symptoms. L. theobromae appeared to be a more aggressive pathogen than C. ampelina on grapevine with more internal brown streaking and greater recovery of pathogen from inoculated samples. To our knowledge, this is the first report of L. theobromae and C. ampelina causing grapevine decline in Castilla y León. References: (1) J. Luque et al. Phytopathol. Mediterr. 45:S101, 2006. (2) M. T. Martin and R. Cobos. Phytopathol. Mediterr. 46:18, 2007. (3) D. Pavlic et al. Stud. Mycol. 50:313, 2004.

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First Report of 16S rDNA II Group Phytoplasma on Polygala mascatense, a Weed in Oman

Plant Disease ◽

10.1094/pd-90-0248c ◽

2006 ◽

Vol 90 (2) ◽

pp. 248-248 ◽

Cited By ~ 1

Author(s):

S. Livingston ◽

M. O. Al-Azri ◽

N. A. Al-Saady ◽

A. M. Al-Subhi ◽

A. J. Khan

Keyword(s):

16S Rdna ◽

Nested Pcr ◽

Pcr Amplification ◽

Perennial Herb ◽

Direct Pcr ◽

First Report ◽

Palm Trees ◽

National Botanic Garden ◽

Negative Controls ◽

Positive Controls

Polygala mascatense Boiss. (family Polygalaceae) is a common weed found in neglected farms, under date palm trees, and in stony locations throughout the Sultanate of Oman (1). It is a perennial herb approximately 30 to 40 cm tall, has slender branches, is woody at the base, and has linear leaves with purple flowers. Recently (November 2004), in the interior region of Oman (210 km south of Muscat), some polygala plants were found stunted with small leaves, bushy growth, and the floral parts were showing phyllody symptoms. Total genomic DNA extracted from asymptomatic and symptomatic plants with modified cetyltrimethylammoniumbromide (CTAB) buffer method (4) was used as a template for direct polymerase chain reaction (PCR) amplification of phytoplasma 16S rDNA with P1/P7 primers. Direct PCR product was used as template DNA for nested PCR with primers R16F2n/R16R2. DNA from plants infected with alfalfa and lime witches'-broom phytoplasma was used as positive controls, and DNA from healthy plants and water was used as negative controls. Products from nested PCR (1.2 kb) were analyzed by using single endonuclease enzyme digestion (restriction fragment length polymorphism [RFLP]) with Tru9I, HaeIII, HhaI, TaqI, AluI, and RsaI (3). The results showed the presence of a 1.8-kb product amplified with direct PCR and a 1.2-kb product of the nested PCR from infected polygala and the positive controls, whereas no PCR products were observed in the negative controls. The PCR assay confirmed the presence of phytoplasma causing witches'-broom disease in polygala. The RFLP results showed the polygala phyto-plasma to be most similar to the alfalfa phytoplasma, a member of 16SrII group (2). Infected polygala weeds may serve as a reservoir for alfalfa witches'-broom phytoplasma that causes annual losses over $25 million to alfalfa cultivation in Oman (2). A detailed investigation needs to be carried out to establish transmission of phytoplasma from polygala to alfalfa. To our knowledge, this is the first report of phytoplasma infecting polygala weeds in Oman. References: (1) S. A. Ghazanfar. Pages 95–96 in: An Annotated Catalogue of the Vascular Plants in Oman. Scripta Botanica Belgica Meise, National Botanic Garden of Belgium, 1992. (2) A. J. Khan et al. Phytopathology 92:1038, 2002. (3) I. M. Lee et al. Int. J. Syst. Bacteriol. 1153, 1998. (4) M. A. Saghai-Maroof et al. Proc. Natl. Acad. Sci. USA 81:8014, 1984.

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Etiology of Botryosphaeria Panicle and Shoot Blight of Pistachio (Pistacia vera) caused by Botryosphaeriaceae in Italy

Plant Disease ◽

10.1094/pdis-08-21-1672-re ◽

2021 ◽

Author(s):

Giorgio Gusella ◽

Daniel P. Lawrence ◽

Dalia Aiello ◽

Yong Luo ◽

Giancarlo Polizzi ◽

...

Keyword(s):

Complex Disease ◽

Phylogenetic Analyses ◽

Morphological Characterization ◽

Fungal Species ◽

Lesion Length ◽

Pistacia Vera ◽

Mycelial Plug ◽

Shoot Blight ◽

Pathogenicity Tests ◽

A Minor

Pistachio (Pistacia vera) is an important crop in Italy, traditionally cultivated in Sicily (southern Italy) for several decades now. In recent years, new orchards have been planted in new areas of the island. Field surveys conducted in 2019 revealed the presence of symptomatic trees showing shoot dieback, cankers, fruit spots, and leaf lesions. Isolations from symptomatic samples consistently yielded fungal species in the Botryosphaeriaceae family. Identification of collected isolates was conducted using morphological and molecular analyses. Morphological characterization was based on conidia measurements of representative isolates and also effects of temperatures on mycelial growth was evaluated. DNA data derived from sequencing the ITS, tef1-α and tub2 gene regions were analyzed via phylogenetic analyses (Maximum Parsimony and Maximum Likelihood). Results of the analyses confirmed the identity of Botryosphaeria dothidea, Neofusicoccum hellenicum and Neofusicoccum mediterraneum. Pathogenicity tests were conducted on detached twigs and in the fields both on shoots as well as on fruit clusters using the mycelial plug technique. The inoculation experiments revealed that among the Botryosphaeriaceae species identified in this study N. hellenicum (occasionally detected) and N. mediterraneum were the most aggressive based on lesion length on shoots and fruits. N. mediterraneum was the most widespread among the orchards while B. dothidea can be considered a minor pathogen involved in this complex disease of pistachio. Moreover, to our knowledge, this is the first report of N. hellenicum in Italy.

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First Report of Forsythia Shoot Blight Caused by Phytophthora nicotianae in Connecticut

Plant Disease ◽

10.1094/pdis-03-14-0297-pdn ◽

2014 ◽

Vol 98 (9) ◽

pp. 1278-1278

Author(s):

J. A. LaMondia ◽

D. W. Li ◽

A. M. Madeiras ◽

R. L. Wick

Keyword(s):

Disease Incidence ◽

Morphological Characteristics ◽

Aerial Mycelium ◽

Phytophthora Nicotianae ◽

First Report ◽

Sterile Needle ◽

Shoot Blight ◽

Pathogenicity Tests ◽

Half Strength ◽

Internal Necrosis

Blighting of Forsythia × intermedia ‘Showoff’ was first observed affecting several hundred plants in a commercial nursery in Connecticut in September 2012. Symptoms included wilting, leaf and stem blight, and dieback progressing to plant death. A Phytophthora sp. was isolated from symptomatic tissues on half-strength potato dextrose agar (½PDA). Colonies were white and cottony on ½PDA, reaching 9 mm in 15 days at 25°C, but colorless and inconspicuous on pimaricin, ampicillin, rifampicin, pentachloronitrobenzene agar (PARP) with sparse and limited aerial mycelium, reaching 60 mm in 15 days at 25°C. The characteristics of the pathogen were observed and measured from a 3-month-old colony on ½PDA. Sporangia were abundant, various in shape, ovoid, ellipsoid to pyriform or limoniform, occasionally gourd shaped or irregular; (17.9) 27.2 to 41.4 (47.3) × (12.6) 19.1 to 30.5 (36.7) μm (n = 30), length/breadth ratio 1.4 ± 0.2, papillate and noncaducous. Papillae measured 2.9 ± 0.8 × 3.4 ± 0.8 μm (n = 10). Chlamydospores were present, 23.4 ± 3.1 × 22 ± 3.3 μm (n = 10). Oogonia and oospores were not observed. Arachnoid mycelia were present. These morphological characteristics are consistent with Phytophthora nicotianae Breda de Haan (1). The identity of the pathogen was confirmed as P. nicotianae by BLAST analysis of ITS, Cox II, and beta tubulin gene sequences (99% match for the three sequences, E value = 0). Pathogenicity tests were conducted four times on healthy liners of Forsythia × intermedia ‘Showoff’ grown in 10-cm-diameter pots. Leaves and stems were wounded by pricking with a sterile needle and six plants were inoculated with 0.25 cm2 plugs of the pathogen growing on ½PDA placed on three leaves and in three stem nodes and covered with Parafilm. Controls consisted of an equal number of plants wounded and inoculated with ½PDA alone. All plants were placed inside high humidity chambers for 24 h and then transferred to a greenhouse for up to 1 month. Typical symptoms developed within 1 week of inoculation and the pathogen was re-isolated from diseased tissue. Disease incidence was nearly 100% of inoculated leaves and stems and not observed in control plants without the pathogen. Three replicate 6-week-old broadleaf tobacco ‘C9’ plants were each inoculated with tobacco or forsythia isolates of P. nicotianae or sterile media alone, by wounding stems and placing colonized 0.25 cm2 ½PDA plugs into wounds and covering with Parafilm. After 1 week, stems were split and the length of internal necrosis in the stem measured. Disease resulted from inoculation with both the tobacco and forsythia isolates and stem necrosis averaged 43 and 23 mm for tobacco or forsythia isolates, respectively. No necrosis was observed in the pathogen-free controls. P. nicotianae has been reported from the basal stem and roots of F. viridissima in Italy (2) and from shoots of Forsythia × intermedia in Virginia (3). To our knowledge, this is the first report of P. nicotianae causing shoot blight on Forsythia in the northeastern United States. References: (1) J. van. Breda de Haan. Mededeelingenuit's Lands Plantentuin Batavia. 15:57, 1896. (2) S. O. Cacciola et al. Plant Dis. 78:525, 1994. (3) C. X. Hong et al. Plant Dis. 89:430, 2005.

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First Report of Stem and Leaf Blight Caused by Sclerotinia sclerotiorum on Eustoma in Taiwan

Plant Disease ◽

10.1094/pdis-02-12-0168-pdn ◽

2012 ◽

Vol 96 (6) ◽

pp. 910-910

Author(s):

Y. M. Shen ◽

C. H. Chao ◽

F. C. Wang ◽

H. L. Liu ◽

T. C. Huang

Keyword(s):

Sclerotinia Sclerotiorum ◽

Leaf Blight ◽

Pathogenicity Test ◽

First Report ◽

Average Temperature ◽

Specific Pcr ◽

Sclerotinia Blight ◽

Pathogenicity Tests ◽

Rice Water ◽

Negative Controls

Eustoma (Eustoma russellianum) is an economically important cut flower in Taiwan. Each year more than 1.7 million dozen flowers, mainly exported to Japan in the winter, are produced in greenhouses. In January 2011, eustoma plants with stem and leaf blight symptoms were observed in some greenhouses in Changhua County, Taiwan, at an incidence of 2%. Brown and rotten lesions were presented on the stem and nearby leaves, with white mycelia growing on the surface and black sclerotia (up to 7 mm long) produced inside the stem. Infected plants were completely blighted and eventually died. Diseased stem tissues collected from the field were surface sterilized for 3 min in 0.6% NaOCl, rinsed with sterilized distilled water, and plated on potato dextrose agar. White fungal colonies were consistently isolated. The cultures produced large sclerotia at the peripheries of the plates. Internal transcribed spacer (ITS) sequences of two voucher isolates were determined and deposited in GenBank (Accession Nos. JQ653934 and JQ653935). The sequences were 100% identical to that of Sclerotinia sclerotiorum strain ATCC MYA-4521 (Accession No. FJ810516). In addition, PCR amplified DNA fragments (approximately 630 bp) were obtained by the S. sclerotiorum specific primer pair MP_SsF and MP_UniR (1). On the basis of morphology, ITS sequence homology, and the specific PCR detection, the fungus was identified as S. sclerotiorum. The two fungal isolates (BCRC34830 and BCRC34831) were deposited in Bioresource Collection and Research Center, Hsinchu, Taiwan. Pathogenicity tests were conducted on 1-month-old, second flush eustoma cultivars Ex Rosa Pink Flash and Rosina Blue Ver. 2 after primary flowers had been harvested in the greenhouse. Fungal inoculum consisting of Tref horticultural substrate and wet sterilized rice colonized by S. sclerotiorum BCRC34830 (substrate-rice-water ratio of 2:1:1) was placed near the base of the plants. Ten plants of each cultivar were inoculated with about 800 g of the mixture. Sterile mixture applied to an equal number of plants served as negative controls. Eight plants of each cultivar showed blight symptoms after 1 month of incubation at an average temperature of 26°C. All control plants remained healthy. The pathogen reisolated from the inoculated stems produced sclerotia identical to those isolated in the field, fulfilling Koch's postulates. The pathogenicity test was repeated with similar results. S. sclerotiorum has been reported on eustoma in Argentina (2). To our knowledge, this is the first report of Sclerotinia blight on eustoma in Taiwan. Although the disease was not prevalent on eustoma, the inoculum could be dormant in the greenhouse soil. Awareness of the potential perennial problem could increase the quality of the flowers exported and benefit the flower industry. References: (1) S. Hirschhäuser and J. Fröhlich. Int. J. Food Microbiol. 118:151, 2007. (2) S. Wolcan et al. Plant Dis. 80:223, 1996.

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First Report of Branch Rot of Lonicera japonica Caused by Sclerotium delphinii in China

Plant Disease ◽

10.1094/pdis-01-14-0053-pdn ◽

2014 ◽

Vol 98 (8) ◽

pp. 1155-1155

Author(s):

M. Zhang ◽

X. J. Wang ◽

Y. Li ◽

Y. H. Geng ◽

H. Y. Wu

Keyword(s):

Disease Incidence ◽

Its Region ◽

Lonicera Japonica ◽

First Report ◽

Plastic Bags ◽

Mycelial Plug ◽

Pathogenicity Tests ◽

White Mycelium ◽

Sclerotium Delphinii ◽

Stem Lesions

Honeysuckle flower (Lonicera japonica Thunb.) is a perennial, traditional Chinese medicine plant, widely cultivated in China. In early June 2013, heavy branch rot infection was observed on L. japonica in an approximately 10,000-m2 field in Linyi, Shandong, China. The disease incidence was 30 to 40%. Early symptoms appeared as small, elliptoid, pale brown lesions on the branches. Lesions expanded into 50 to 100 mm long and 3 to 7 mm wide, brown, elongated spots. The upper branches wilted after the lesions expanded around the stems. A fungus was consistently isolated from stem lesions on potato dextrose agar (PDA) that was morphologically similar to S. delphinii, with white mycelium, round to irregularly shaped reddish-brown sclerotia that were 2 to 4 mm diameter (2). The identity of the fungus was confirmed by DNA sequencing of the internal transcribed spacer (ITS) region (GenBank Accession No. KJ145328), which was 99% homologous to those of other S. delphinii isolates (JN241578 and AB075314) (1). Pathogenicity tests were conducted with three 2-year-old seedlings grown in 20-cm-diameter pots at 25 to 30°C during experiments in greenhouse. Ten branches from the three plants pricked by needle were inoculated with a mycelial plug (0.4 cm diameter) harvested from the periphery of a 4-day-old colony. An equal number of branches pricked by needle serving as controls were mock-inoculated with plugs of PDA medium. Inoculated branches were covered with plastic bags for 24 h to maintain high relative humidity and incubated at about 25°C. Plugs were removed 48 h after inoculation. After 3 days, nine inoculated branches showed symptoms identical to those observed in the field under natural conditions, whereas controls remained symptom-free. Re-isolation of the fungus from lesions on inoculated branches confirmed that the causal agent was S. delphinii. Pathogenicity tests were repeated three times by the same methods with the same results. To our knowledge, this is the first report of S. delphinii infecting Lonicera japonica in China. References: (1) I. Okabe and N. Matsumoto. Mycol. Res. 107:164, 2003. (2) Z. K. Punja and A. Damiani. Mycologia. 88:694, 1996.

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First Report of Seimatosporium botan Associated with Trunk Disease of Grapevine (Vitis vinifera) in Chile

Plant Disease ◽

10.1094/pdis-05-12-0478-pdn ◽

2012 ◽

Vol 96 (11) ◽

pp. 1696-1696 ◽

Cited By ~ 4

Author(s):

G. A. Díaz ◽

K. Elfar ◽

B. A. Latorre

Keyword(s):

Cross Sections ◽

Lesion Length ◽

Woody Tissue ◽

First Report ◽

Trunk Disease ◽

Agar Plug ◽

Significant Difference ◽

Pathogenicity Tests ◽

Cork Borer ◽

Dark Green

Grapevines are planted on 180,000 ha in Chile. In 2010 and 2011, necrotic lesions and hard texture were observed on woody tissue on 10-year-old vines of cvs. Cabernet Sauvignon, Carménère, Moscatel de Alejandría, and Pedro Jimenez in Ovalle (lat. 30°58′ S) and Cauquenes (lat. 35°58′ S). Symptoms were on 10 to 25% of the arm cross sections, resembling symptoms caused by Botryosphaeriaceae (4). Prevalence of 5% was estimated visually in Ovalle (n = 920 grapevines) and Cauquenes (n = 350 grapevines). Small pieces (3 mm) of necrotic tissues from the margins of lesions in cordons (n = 32) were surface sterilized (96% ethanol, 15 s), and plated on acidified PDA plus 0.5 ml/liter of 92% lactic acid, 0.005% tetracycline, 0.01% streptomycin, and 0.1% Igepal CO-630 (Sigma-Aldrich, St. Louis, MO) (APDA). The plates were incubated at 20°C for 14 days. Isolates (n = 12) were obtained from the yellow to dark green slimy colonies with white irregular margins, staining brown the underside of APDA plates. Black acervuli and ellipsoid to fusiform conidia were obtained. Conidia were triple septated, with hyaline upper and bottom cells and brown middle cells (n = 30) of 17.7 ± 1.2 × 5.8 ± 0.8 μm. A basal conidial appendage (6.2 ± 1.0 μm) was always obtained, but conidia having appendages at both ends also were observed. Morphologically, these isolates were identified as Seimatosporium botan Sat. Hatak. & Y. Harada (2). The identification of isolates sei-302 and sei-316 was confirmed by amplifying and sequencing the region ITS1-5.8S-ITS2 of rDNA using ITS4 and ITS5 primers (GenBank Accession Nos. JN088482 and JN088483). BLAST analyses showed 100% similarity with S. botan (Accession No. HM067840) (2). Pathogenicity tests were conducted with isolates sei-302 and sei-316 on detached green shoots (GS) and on rooted 2-year-old vines ‘Carménère.’ Rooted vines were inoculated at the base of canes and trunks. Inoculations were performed by placing a mycelial agar plug taken from APDA on a wound aseptically made with a cork borer. Wounds were sealed with Parafilm to avoid a rapid dehydration. The inoculated GS were incubated for 2 weeks in a moisture chamber (relative humidity >80%) at 20°C. Inoculated 2-year-old vines were placed in a lath-house for 7 and 15 months for canes and trunk inoculation, respectively. An equal number of GS and vines were inoculated with sterile agar plugs and left as controls. Necrotic lesions with mean of 23.7 ± 2.5 mm on GS, 50.5 ± 3.4 mm on canes, and 41.9 ± 2.3 mm on trunks developed. No significant difference (P < 0.05) was obtained in lesion length between S. botan isolates. After 7 months, 40% of inoculated canes had died. No symptoms were observed in GS controls and rooted control vines treated with sterile agar plugs. S. botan was reisolated from 93 to 100% of the inoculated samples. Previously, S. botan was reported as pathogenic in Paeonia suffruticosa (1), and Seimatosporium sp. was isolated from V. vinifera in California, but their pathogenicity was not demonstrated (3). To our knowledge, this is the first report of pathogenic isolates of S. botan associated with trunk disease of grapevines. These results contribute to the knowledge of the trunk disease of grapevines worldwide. References: (1) Y. Duan et al. Plant Dis. 95:226, 2011. (2) S. Hatakeyama et al. Mycoscience 45:106, 2004. (3) Z. Morales et al. Phytopathol. Mediterr. 49:109, 2010. (4) J. R. Úrbez-Torres. Phytopathol. Mediterr. 50:S5, 2011.

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