First Report of Umbel Browning and Stem Necrosis Caused by Diaporthe angelicae on Carrot in France

In 2011, carrot (Daucus carota L.) seed production occurred on 2,900 ha, which accounts for approximately 25% of the area devoted to the production of vegetable fine seeds. Since 2007, symptoms of umbel browning have been regularly observed in carrot production areas located in the central region. Initially, triangular necrotic lesions appeared on carrot umbels that later spread to the entire umbels and often progressed to the stems. Diseased umbels became dried prematurely, compromising seed development. The loss in seed production was estimated at approximately 8% of the harvested carrot umbels during the cropping seasons of spring and summer 2007 and 2008 in France. In collaboration with seed companies, diseased carrot stems were collected from seven fields of seed production (eight plants per field) and a fungus was isolated from the tissue. The cultures were grown on malt (2%) agar (1.5%) medium and incubated for 2 weeks at 22°C in darkness. Young fungal colonies were white and a brownish green pigmentation developed when the colonies became older. The same color was observed from the top and on the reverse of the colonies. To induce sporulation, isolates were grown on water agar (1.5%) medium in the presence of carrot stem fragments for 1 week at 22°C in darkness, followed by 1 week at 22°C in white light under a 16-h photoperiod. Pycnidia were produced on stem fragments and contained alpha and beta conidia typical of the genus Diaporthe (2). Alternatively, pycnidia were also obtained on malt agar medium after 2 weeks of culture at 25°C in white light under a 12-h photoperiod. The size of alpha and beta conidia was 6.3 ± 0.5 × 2.3 ± 0.4 μm and 23.3 ± 1.8 × 0.9 ± 0.2 μm, respectively (n = 170). In order to confirm the identification at the genus level and determine the species, DNA was extracted from the mycelium of three representative isolates and the ITS regions of the ribosomal DNA were amplified using universal primers (1). The sequences of the amplified products (GenBank Accession Nos. KF240772 to KF240774) were 100% identical with the ITS sequence of a Diaporthe angelicae isolate deposited in the NCBI database (CBS 111592 isolate, KC343027). To confirm pathogenicity, the three isolates of D. angelicae were inoculated on carrot umbels in the greenhouse. A total of nine plants were inoculated (three plants per isolate). Using a micropipette, 10 μl of a conidial suspension containing alpha and beta conidia (105 conidia mL–1) were deposited at the base of the primary umbel and two secondary umbels, which were wounded before inoculation using a scalpel blade. Seven inoculated plants developed triangular, necrotic lesions that were typical umbel browning. D. angelicae was re-isolated on malt agar medium from the inoculated diseased carrot umbels. To our knowledge, this is the first report of D. angelicae in carrot cultivated for seed production in France. The disease resembles the lesions described in the Netherlands in 1951 on carrot inflorescence caused by Phomopsis dauci (3). In future experiments, it would be crucial to precisely determine if D. angelicae could be transmitted to the seeds. References: (1) M. A. Innis et al. PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990. (2) J. M. Santos and A. J. L. Philips. Fungal Divers. 34:111, 2009. (3) J. A. von Arx. Eur. J. Plant Pathol. 57:44, 1951.

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First Report of Fusarium oxysporum f. sp. lycopersici Race 3 and F. oxysporum f. sp. radicis-lycopersici in Tomatoes in the Azapa Valley of Chile

Plant Disease ◽

10.1094/pdis-03-14-0303-pdn ◽

2014 ◽

Vol 98 (10) ◽

pp. 1432-1432 ◽

Cited By ~ 4

Author(s):

G. Sepúlveda-Chavera ◽

W. Huanca ◽

R. Salvatierra-Martínez ◽

B. A. Latorre

Keyword(s):

Fusarium Oxysporum ◽

Tap Water ◽

Universal Primers ◽

Conidial Suspension ◽

Laboratory Manual ◽

First Report ◽

Solanum Lycopersicum L ◽

Vascular Discoloration ◽

Seedling Roots ◽

Pcr Assays

Tomato (Solanum lycopersicum L.) is an important crop in the Azapa Valley (18°35′ S, 69°30′ W) in northern Chile, with approximately 600 ha of fresh tomatoes under greenhouses. Cultivars resistant to Fusarium oxysporum f. sp. lycopersici (FOL) races 1 and 2 are mainly used. However, in 2012 and 2013, Fusarium wilt incidence was 2 to 3%. Symptoms appeared unilaterally and consisted of yellowing, leaf wilting of lower leaves, dark brown vascular discoloration, and plant death. The aim of this study was to determine the causal agent of tomato wilt in seven tomato greenhouses in the Azapa Valley. Stem samples (5 × 5 mm) were obtained 10 cm of the stem base from wilted tomatoes ‘Naomi’ (BIOAMERICA S.A., Chile) or from Maxifort tomato rootstock (De Ruiter Seed, USA), both FOL resistant to races 1 and 2. Samples were washed with tap water, surface sterilized with 1% NaClO for 3 min, and incubated on sterile moist paper towels in petri plates for 5 days at 22°C. Mycelial fragments from white colonies, emerging from diseased tissues, were transferred to PDA. Six Fusarium isolates were characterized by the presence of hyaline macroconidia, mostly 3 to 5 septate, slightly curved (19.2 to 32.1 × 2.9 to 4.5 μm) and single-celled, oval to elongated microconidia (3.1 to 8.9 × 2.0 to 4.0 μm). Chlamydospores were single or in pairs. These isolates were identified as F. oxysporum (3). The identity of F. oxysporum was confirmed by PCR assays using genomic DNA of each isolated and the universal primers Uni F and Uni R that generate a 672-bp PCR product. The pathogenic form and races were determined by PCR assays using the specific primers uni, sp13, sp23, and sprl that were able to discriminate all the three FOL races as well as F. oxysporum f. sp. radicis-lycopersici (FORL) isolates (2). The sp13 and sp23 primers amplified DNA bands of 445 and 518 bp, confirming the identity of FOL race 3. However, sprl amplified a fragment of 947 bp corresponding to FORL (2). Pathogenicity tests were conducted on 25-day-old seedlings (10 seedlings per isolate) of tomato ‘Poncho Negro,’ which is susceptible to FOL and FORL. Seedling roots were cut, submerged for 5 min in conidial suspension of 2 × 106 conidia/ml, and transplanted to 250-ml plastic containers with sterile substrate (sand/peat, 1:1). Equally treated non-inoculated seedlings were left as controls. The first symptoms induced by each of the five FOL isolates appeared 8 days after incubation under greenhouse and were characterized by yellowing of older leaves, sometimes affecting one side of the plant, vascular discoloration of the stem, and eventually plant death. In contrast, all seedlings inoculated with a FORL isolate developed a necrotic lesion and vascular discoloration at the base of the stems near the soil line, followed by wilting and plant death. Control plants remained asymptomatic. F. oxysporum was re-isolated only from inoculated plants, completing Koch's postulates. FOL and FORL were reported earlier in other tomato growing areas of Chile (1), located over 1,000 km south of the Azapa Valley. However, this is the first report of FOL race 3 and FORL in the Azapa Valley and FOL race 3 is reported for the first time in Chile. References: (1) S. Acuña. Compendio de Fitopatógenos de Cultivos Agrícolas. Servicio Agrícola y Ganadero. Gobierno de Chile, 2008. (2) Y. Hirano and T. Arie. J. Gen. Plant Pathol. 72:273, 2006. (3) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Ames, IA, 2006.

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First Report of Alternaria Leaf Spot Caused by Alternaria sp. on Highbush Blueberry (Vaccinium corymbosum) in South Korea

Plant Disease ◽

10.1094/pdis-04-14-0344-pdn ◽

2014 ◽

Vol 98 (10) ◽

pp. 1434-1434

Author(s):

J.-H. Kwon ◽

D.-W. Kang ◽

M.-G. Cheon ◽

J. Kim

Keyword(s):

South Korea ◽

Leaf Spot ◽

Disease Incidence ◽

Vaccinium Corymbosum ◽

Its Rdna ◽

Universal Primers ◽

Conidial Suspension ◽

First Report ◽

Representative Isolate ◽

Alternaria Sp

In South Korea, the culture, production, and consumption of blueberry (Vaccinium corymbosum) have increased rapidly over the past 10 years. In June and July 2012, blueberry plants with leaf spots (~10% of disease incidence) were sampled from a blueberry orchard in Jinju, South Korea. Leaf symptoms included small (1 to 5 mm in diameter) brown spots that were circular to irregular in shape. The spots expanded and fused into irregularly shaped, large lesions with distinct dark, brownish-red borders. The leaves with severe infection dropped early. A fungus was recovered consistently from sections of surface-disinfested (1% NaOCl) symptomatic leaf tissue after transfer onto water agar and sub-culture on PDA at 25°C. Fungal colonies were dark olive and produced loose, aerial hyphae on the culture surfaces. Conidia, which had 3 to 6 transverse septa, 1 to 2 longitudinal septa, and sometimes also a few oblique septa, were pale brown to golden brown, ellipsoid to ovoid, obclavate to obpyriform, and 16 to 42 × 7 to 16 μm (n = 50). Conidiophores were pale to mid-brown, solitary or fasciculate, and 28 to 116 × 3 to 5 μm (n = 50). The species was placed in the Alternaria alternata group (1). To confirm the identity of the fungus, the complete internal transcribed spacer (ITS) rDNA region of a representative isolate, AAVC-01, was amplified using ITS1 and ITS4 primers (2). The DNA products were cloned into the pGEM-T Easy vector (Promega, Madison, WI) and the resulting pOR13 plasmid was sequenced using universal primers. The resulting 570-bp sequence was deposited in GenBank (Accession No. KJ636460). Comparison of ITS rDNA sequences with other Alternaria spp. using ClustalX showed ≥99% similarity with the sequences of A. alternata causing blight on Jatropha curcas (JQ660842) from Mexico and Cajannus cajan (JQ074093) from India, citrus black rot (AF404664) from South Africa, and other Alternaria species, including A. tenuissima (WAC13639) (3), A. lini (Y17071), and A. longipes (AF267137). Two base substitutions, C to T at positions 345 and 426, were found in the 570-bp amplicon. Phylogenetic analysis revealed that the present Alternaria sp. infecting blueberry grouped separately from A. tenuissima and A. alternata reported from other hosts. A representative isolate of the pathogen was used to inoculate V. corymbosum Northland leaves for pathogenicity testing. A conidial suspension (2 × 104 conidia/ml) from a single spore culture and 0.025% Tween was spot inoculated onto 30 leaves, ranging from recently emerged to oldest, of 2-year-old V. corymbosum Northland plants. Ten leaves were treated with sterilized distilled water and 0.025% Tween as a control. The plants were kept in a moist chamber with >90% relative humidity at 25°C for 48 h and then moved to a greenhouse. After 15 days, leaf spot symptoms similar to those observed in the field developed on the inoculated leaves, whereas the control plants remained asymptomatic. The causal fungus was re-isolated from the lesions of the inoculated plants to fulfill Koch's postulates. To our knowledge, this is the first report of Alternaria sp. on V. corymbosum in South Korea. References: (1) E. G. Simmons. Page 1797 in: Alternaria: An Identification Manual. CBS Fungal Biodiversity Centre, Utrecht, The Netherlands, 2007. (2) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990. (3) M. P. You et al. Plant Dis. 98:423, 2014.

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First report of banana (Musa acuminate L. AAA Cavendish, cv. Formosana) leaf spot disease caused by Curvularia geniculata in China

Plant Disease ◽

10.1094/pdis-09-21-2001-pdn ◽

2021 ◽

Author(s):

Yanxiang Qi ◽

Yanping Fu ◽

Jun Peng ◽

Fanyun Zeng ◽

Yanwei Wang ◽

...

Keyword(s):

Leaf Spot ◽

Disease Incidence ◽

Morphological Characteristics ◽

Universal Primers ◽

Leaf Spot Disease ◽

Leaf Margin ◽

Conidial Suspension ◽

Tropical Fruit ◽

First Report ◽

Spot Disease

Banana (Musa acuminate L.) is an important tropical fruit in China. During 2019-2020, a new leaf spot disease was observed on banana (M. acuminate L. AAA Cavendish, cv. Formosana) at two orchards of Chengmai county (19°48ʹ41.79″ N, 109°58ʹ44.95″ E), Hainan province, China. In total, the disease incidence was about 5% of banana trees (6 000 trees). The leaf spots occurred sporadically and were mostly confined to the leaf margin, and the percentage of the leaf area covered by lesions was less than 1%. Symptoms on the leaves were initially reddish brown spots that gradually expanded to ovoid-shaped lesions and eventually become necrotic, dry, and gray with a yellow halo. The conidia obtained from leaf lesions were brown, erect or curved, fusiform or elliptical, 3 to 4 septa with dimensions of 13.75 to 31.39 µm × 5.91 to 13.35 µm (avg. 22.39 × 8.83 µm). The cells of both ends were small and hyaline while the middle cells were larger and darker (Zhang et al. 2010). Morphological characteristics of the conidia matched the description of Curvularia geniculata (Tracy & Earle) Boedijn. To acquire the pathogen, tissue pieces (15 mm2) of symptomatic leaves were surface disinfected in 70% ethanol (10 s) and 0.8% NaClO (2 min), rinsed in sterile water three times, and transferred to potato dextrose agar (PDA) for three days at 28°C. Grayish green fungal colonies appeared, and then turned fluffy with grey and white aerial mycelium with age. Two representative isolates (CATAS-CG01 and CATAS-CG92) of single-spore cultures were selected for molecular identification. Genomic DNA was extracted from the two isolates, the internal transcribed spacer (ITS), large subunit ribosomal DNA (LSU rDNA), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), translation elongation factor 1-alpha (TEF1-α) and RNA polymerase II second largest subunit (RPB2) were amplified and sequenced with universal primers ITS1/ITS4, LROR/LR5, GPD1/GPD2, EF1-983F/EF1-2218R and 5F2/7cR, respectively (Huang et al. 2017; Raza et al. 2019). The sequences were deposited in GenBank (MW186196, MW186197, OK091651, OK721009 and OK491081 for CATAS-CG01; MZ734453, MZ734465, OK091652, OK721100 and OK642748 for CATAS-CG92, respectively). For phylogenetic analysis, MEGA7.0 (Kumar et al. 2016) was used to construct a Maximum Likelihood (ML) tree with 1 000 bootstrap replicates, based on a concatenation alignment of five gene sequences of the two isolates in this study as well as sequences of other Curvularia species obtained from GenBank. The cluster analysis revealed that isolates CATAS-CG01 and CATAS-CG92 were C. geniculata. Pathogenicity assays were conducted on 7-leaf-old banana seedlings. Two leaves from potted plants were stab inoculated by puncturing into 1-mm using a sterilized needle and placing 10 μl conidial suspension (2×106 conidia/ml) on the surface of wounded leaves and equal number of leaves were inoculated with sterile distilled water serving as control (three replicates). Inoculated plants were grown in the greenhouse (12 h/12 h light/dark, 28°C, 90% relative humidity). Necrotic lesions on inoculated leaves appeared seven days after inoculation, whereas control leaves remained healthy. The fungus was recovered from inoculated leaves, and its taxonomy was confirmed morphologically and molecularly, fulfilling Koch’s postulates. C. geniculata has been reported to cause leaf spot on banana in Jamaica (Meredith, 1963). To our knowledge, this is the first report of C. geniculata on banana in China.

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First Report of Colletotrichum boninense Causing Anthracnose on Pepper in China

Plant Disease ◽

10.1094/pdis-04-12-0403-pdn ◽

2013 ◽

Vol 97 (1) ◽

pp. 138-138 ◽

Cited By ~ 7

Author(s):

Y. Z. Diao ◽

J. R. Fan ◽

Z. W. Wang ◽

X. L. Liu

Keyword(s):

Internal Transcribed Spacer ◽

Severe Disease ◽

Causal Agent ◽

Its Sequences ◽

Universal Primers ◽

Pathogenicity Test ◽

Conidial Suspension ◽

Single Spore ◽

First Report ◽

Species Specific

Anthracnose, caused by Colletotrichum spp., is a severe disease and results in large losses in pepper (Capsicum frutescens) production in China (4). Colletotrichum boninense is one of the Colletotrichum species in pepper in China. In August 2011, anthracnose symptoms (circular, sunken lesions with orange to black spore masses) were observed on pepper fruits in De-Yang, Sichuan Province, China. Three single-spore isolates (SC-6-1, SC-6-2, SC-6-3) were obtained from the infected fruits. A 5-mm diameter plug was transferred to potato dextrose agar (PDA); the isolates formed colonies with white margins and circular, dull orange centers. The conidia were cylindrical, obtuse at both ends, and 10.5 to 12.6 × 4.1 to 5.0 μm. The colonies grew rapidly at 25 to 28°C, and the average colony diameter was 51 to 52 mm after 5 days on PDA at 25°C. Based upon these characters, the causal agent was identified as C. boninense. To confirm the identity of the isolates, the internal transcribed spacer (ITS) regions were amplified with the ITS1/ITS4 universal primers (1). The internal transcribed spacer (ITS) sequences (Accession No. JQ926743) of the causal fungus shared 99 to 100% homology with ITS sequences of C. boninense in GenBank (Accession Nos. FN566865 and EU822801). The identity of the causal agent as C. boninense was also confirmed by species-specific primers (Col1/ITS4) (2). In a pathogenicity test, five detached ripe pepper fruits were inoculated with 1 μl of a conidial suspension (106 conidia/mL) or five fruits with 1 μl of sterile water were kept as control. After 7 days in a moist chamber at 25°C, typical anthracnose symptoms had developed on the five inoculated fruits but not on control fruits. C. boninense was reisolated from the lesions, and which was confirmed by morphology and molecular methods as before. There have reports of C. boninense infecting many species of plants, including pepper (3). To our knowledge, this is the first report of C. boninense causing anthracnose on pepper in China. References: (1) A. K. Lucia et al. Phytopathology 93:581, 2002. (2) S. A. Pileggi et al. Can. J. Microbiol. 55:1081, 2009. (3) H. J. Tozze et al. Plant Dis. 93:106, 2009. (4) M. L. Zhang. J. Anhui Agri. Sci. 2:21, 2000.

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First Report of Anthracnose Caused by Colletotrichum gloeosporioides in Pumpkin in Trinidad

Plant Disease ◽

10.1094/pdis-94-8-1062a ◽

2010 ◽

Vol 94 (8) ◽

pp. 1062-1062

Author(s):

S. N. Rampersad

Keyword(s):

Colletotrichum Gloeosporioides ◽

Phylogenetic Analyses ◽

Pcr Amplification ◽

Positive Control ◽

Universal Primers ◽

Conidial Suspension ◽

Specific Primers ◽

First Report ◽

Main Production ◽

Species Specific

In Trinidad, pumpkin (Cucurbita pepo L. and C. moschata L.) is extensively grown for local and international export markets. In November 2008, symptoms of foliar chlorosis and necrosis were observed in 15 commercial pumpkin fields located in the main production areas of St. George East, Caroni, Victoria, and St. Patrick counties. Severely infected plants were unable to support fruit maturation, which resulted in yield loss. The pathogen was isolated from surface-sterilized tissues of symptomatic plants. Colonies on potato dextrose agar (PDA) were white to cream with gray spore masses in the center. Conidia were hyaline, cylindrical with rounded ends, aseptate, and measured 12.5 to 16.5 μm × 3.5 to 5.0 μm. PCR amplification was carried out with ITS4/5 universal primers (4) and species-specific primers, CgInt/ITS4 (1), using a positive control of Colletotrichum gloeosporioides (courtesy of D. Perez-Brito). Species-specific primers generated a single amplicon, ~450 bp long, which corresponded with the positive control. The ITS1 region (1) of pumpkin isolates (GenBank No. GU320190) was 100% identical to cognate sequences of C. gloeosporioides isolates (GenBank Nos. AY841136 and FJ624257). Phylogenetic analyses (MEGA 4 – Molecular Evolutionary Genetic Analysis Software version 4 for Windows) using the neighbor-joining (NJ) algorithm placed the pumpkin isolates in a well-supported cluster (>90% bootstrap value based on 1,000 replicates) with other C. gloeosporioides isolates. The tree was rooted with C. crassipes (GenBank No. AJ536230). The pathogen was similar to C. gloeosporioides (Penz.) Penz. & Sacc. (3). In pathogenicity tests, six plants (cv. Jamaican squash) for each of five isolates were spray inoculated to runoff with a conidial suspension (1.0 × 106 conidia/ml). Negative controls were sprayed with sterile distilled water. In repeated tests, plants were symptomatic of infection 7 days postinoculation. There were no symptoms on control plants. Koch's postulates were fulfilled with the reisolation of the pathogen from symptomatic leaf tissues. Anthracnose is a serious threat to cucurbit production; however, infection is not common in pumpkin and squash (2). To my knowledge, this is the first report of C. gloeosporioides causing widespread anthracnose infection in pumpkin in Trinidad. References: (1) A. E. Brown et al. Phytopathology 86:523, 1996. (2) G. Kelly. Acta Hortic. (ISHS) 731:479, 2007. (3) B. C. Sutton. Page 1 in: Colletotrichum: Biology, Pathology and Control. CAB International. Wallingford, UK, 1992. (4) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990.

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First Report of Phomopsis longicolla Causing Stem Canker of Eggplant in Guangdong, China

Plant Disease ◽

10.1094/pdis-07-13-0790-pdn ◽

2014 ◽

Vol 98 (3) ◽

pp. 426-426 ◽

Cited By ~ 4

Author(s):

C. Shu ◽

J. Chen ◽

H. Huang ◽

Y. He ◽

E. Zhou

Keyword(s):

Disease Incidence ◽

Guangdong Province ◽

Stem Canker ◽

Universal Primers ◽

Distilled Water ◽

Conidial Suspension ◽

Phomopsis Longicolla ◽

Fungal Isolation ◽

First Report ◽

Absorbent Cotton

Eggplant (Solanum melongena L.) is an economically important vegetable crop worldwide. In August 2012, severe stem cankers were observed on eggplant at the early stage of maturation in several fields in Guangdong Province, China. Diseased plants raised cankers on the stems and branches, which resulted in wilting and stunting. No symptoms developed on eggplant fruit. Disease incidence was as high as 40% within affected fields. By using routine fungal-isolation methods and single-spore purification technique, five single-conidial isolates were obtained from each diseased stem. Colonies were grayish-white, circular, and got yellow pigmentation when placed in acidified potato dextrose agar (PDA) in an incubator at pH 4.5 and 25°C with a 12-h photoperiod. Stromata were black, large, and spreading in a concentric pattern. Conidiomata were pycnidial, and the pycnidia were round, oblate, triangular or irregular, and unilocular. Conidiophores were colorless, separated, dichotomous, and 10.0 to 18.0 × 1.5 to 2.0 μm. Alpha conidia were single-celled, ellipsoidal to fusiform, guttulate, and 6.0 to 8.0 × 2.0 to 2.5 μm. Beta conidia, produced on oat meal agar in 2 weeks at 25°C in the dark, were filiform, hamate, and 16.0 to 28.0 × 0.7 to 1.0 μm. Based on these morphological characters, the fungus was identified as Phomopsis longicolla Hobbs (1). The ITS-rDNA sequence (GenBank Accession No. KC886605) of the isolate EPPL1 of this fungus (P. longicolla EPPL1) was obtained by using universal primers ITS5/ITS4 (1). BLAST searches showed a 98% homology with the sequence of the ITS region of rDNA of P. longicolla. Phylogenetic analysis showed that P. longicolla EPPL1 clustered with P. longicolla SYJM15 and formed a distinct clade distantly related to P. vexans PV3 (GU373630), a well-known pathogen of eggplant. Digestion of PCR-amplified DNA with Alu I yielded two restriction fragments of sizes consistent with those reported for P. longicolla (2). Pathogenicity tests were performed on 30-day-old plants of cv. Yuefengzihongqie grown in a plastic pot (1 liter) in a greenhouse by using mycelial plugs and conidial suspensions of isolate EPPL1 as inocula. A mycelial plug (4 mm in diameter) from a 7-day-old PDA culture was placed on stems of both wounded and non-wounded plants and covered with sterile absorbent cotton moistened with sterile distilled water. Both wounded and non-wounded plants were inoculated with 0.5 ml of conidial suspension (1 × 106 conidia ml–1) dropped onto sterile absorbent cotton covering the stems. Control assays were performed with agar plugs and sterile distilled water only. Inoculated plants were placed in a greenhouse with a 12-h photoperiod at 28°C. Each treatment was replicated on five plants, and the test was repeated. Twenty-five days after inoculation, both wounded and non-wounded plants inoculated with either method showed raised cankers at the points of inoculation and canker lesions similar to those observed in the field expanded up and down the stems to reach lengths of 15 to 30 mm. Later, sparse, small, black pycnidia formed on the surface of the lesions. The inoculated plants exhibited stunting and premature senescence compared to controls. P. longicolla was re-isolated from the infected stems of inoculated plants. Control plants were asymptomatic. To our knowledge, this is the first report of P. longicolla causing stem canker in eggplant in Guangdong, China. Considering the economic importance of eggplant in Guangdong Province and throughout the world, further study of phomopsis stem canker of eggplant is warranted. References: (1) T. W. Hobbs et al. Mycologia 77:535, 1985. (2) A. W. Zhang et al. Plant Dis. 81:1143, 1997.

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First Report of Mango Malformation Disease Caused by Fusarium mangiferae in Spain

Plant Disease ◽

10.1094/pdis-07-11-0599 ◽

2012 ◽

Vol 96 (2) ◽

pp. 286-286 ◽

Cited By ~ 21

Author(s):

M. Crespo ◽

F. M. Cazorla ◽

J. M. Hermoso ◽

E. Guirado ◽

M. Maymon ◽

...

Keyword(s):

Agar Medium ◽

Economic Losses ◽

Conidial Suspension ◽

Control Strain ◽

Potato Dextrose Agar Medium ◽

Floral Tissue ◽

First Report ◽

Specific Pcr ◽

Mango Malformation ◽

Fusarium Mangiferae

Mango (Mangifera indica L.) malformation disease (MMD) is one of the most important diseases affecting this crop worldwide, which causes severe economic losses because of the reduction of productivity. Symptoms of MMD in Spain were observed for the first time in April of 2006 in three mango orchards in the Axarquia Region (southern Spain). Symptoms included an abnormal development of vegetative shoots with shortened internodes and dwarfed leaves and hypertrophied short and thickened panicles. In the years of 2006, 2009, and 2010, isolates of Fusarium were obtained from vegetative shoots and floral tissue of symptomatic mango trees from 21 different orchards of cvs. Keitt, Kent, Osteen, Tommy Atkins, and a variety of minor commercial cultivars, all showing typical symptoms of MMD. Different Fusarium-like strains were isolated from infected tissues. Colonies from single-spored isolates possessed dark purple-to-salmon-colored mycelium when grown on potato dextrose agar medium. On fresh carnation leaf agar medium, mycelium contained aerial conidiophores possessing three- to five-celled macroconidia and abundant microconidia in false heads from mono- and polyphialides; while cream-orange-colored sporodochia were produced on the surface of the medium, typical for Fusarium mangiferae. The identification of 37 isolates was confirmed as F. mangiferae by species-specific PCR analysis with the primer pair 1-3 F/R that amplified a 608-bp DNA fragment from all Spanish isolates as well as a representative Israeli control strain, Fus 34, also designated as MRC7560 (2). Pathogenicity using four representative isolates, UMAF F02, UMAF F10, UMAF F17, and UMAF F38 of F. mangiferae from Spain as well as isolate MRC7560, was tested on 2-year-old healthy mango seedlings cv. Keitt by inoculating 15 buds from three different trees with a 20-μl conidial suspension (5 × 107 conidia per ml) per isolate (1). This experiment was conducted twice with two independent sets of plants and at different times (March and November 2010). Typical mango malformation symptoms were detected after bud break in March 2011, 5 and 12 months after inoculation. Symptoms were observed for 60% of the inoculated buds with the four F. mangiferae Spanish isolates and 75% with the MRC7560 control strain, but not with water-inoculated control plants. Recovered isolates from the infected floral and vegetative malformed buds were identical morphologically to those inoculated, and the specific 608-bp fragment described for F. mangiferae was amplified with specific-PCR, thus fulfilling Koch's postulates. To our knowledge, this is the first report of mango malformation disease caused by F. mangiferae in Spain and Europe. References: (1) S. Freeman et al. Phytopathology 89:456, 1999. (2) Q. I. Zheng and R. C. Ploetz. Plant Pathol. 51:208, 2002.

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First Report of Gray Leaf Spot on Pepper Caused by Stemphylium solani in Malaysia

Plant Disease ◽

10.1094/pdis-03-12-0262-pdn ◽

2012 ◽

Vol 96 (8) ◽

pp. 1227-1227 ◽

Cited By ~ 4

Author(s):

A. Nasehi ◽

J. B. Kadir ◽

M. A. Zainal Abidin ◽

M. Y. Wong ◽

F. Abed Ashtiani

Keyword(s):

Leaf Spot ◽

Disease Incidence ◽

Leaf Tissue ◽

Gray Leaf Spot ◽

Universal Primers ◽

Conidial Suspension ◽

Internal Transcribed Spacer Region ◽

First Report ◽

Morphological Criteria ◽

Pepper Leaves

Symptoms of gray leaf spot were first observed in June 2011 on pepper (Capsicum annuum) plants cultivated in the Cameron Highlands and Johor State, the two main regions of pepper production in Malaysia (about 1,000 ha). Disease incidence exceeded 70% in severely infected fields and greenhouses. Symptoms initially appeared as tiny (average 1.3 mm in diameter), round, orange-brown spots on the leaves, with the center of each spot turning gray to white as the disease developed, and the margin of each spot remaining dark brown. A fungus was isolated consistently from the lesions using sections of symptomatic leaf tissue surface-sterilized in 1% NaOCl for 2 min, rinsed in sterile water, dried, and plated onto PDA and V8 agar media (3). After 7 days, the fungal colonies were gray, dematiaceous conidia had formed at the end of long conidiophores (19.2 to 33.6 × 12.0 to 21.6 μm), and the conidia typically had two to six transverse and one to four longitudinal septa. Fifteen isolates were identified as Stemphylium solani on the basis of morphological criteria described by Kim et al. (3). The universal primers ITS5 and ITS4 were used to amplify the internal transcribed spacer region (ITS1, 5.8, and ITS2) of ribosomal DNA (rDNA) of a representative isolate (2). A 570 bp fragment was amplified, purified, sequenced, and identified as S. solani using a BLAST search with 100% identity to the published ITS sequence of an S. solani isolate in GenBank (1). The sequence was deposited in GenBank (Accession No. JQ736024). Pathogenicity of the fungal isolate was tested by inoculating healthy pepper leaves of cv. 152177-A. A 20-μl drop of conidial suspension (105 spores/ml) was used to inoculate each of four detached, 45-day-old pepper leaves placed on moist filter papers in petri dishes (4). Four control leaves were inoculated similarly with sterilized, distilled water. The leaves were incubated at 25°C at 95% relative humidity for 7 days. Gray leaf spot symptoms similar to those observed on the original pepper plants began to develop on leaves inoculated with the fungus after 3 days, and S. solani was consistently reisolated from the leaves. Control leaves did not develop symptoms and the fungus was not reisolated from these leaves. Pathogenicity testing was repeated with the same results. To our knowledge, this is the first report of S. solani causing gray leaf spot on pepper in Malaysia. References: (1) S. F. Altschul et al. Nucleic Acids Res. 25:3389, 1997. (2) M. P. S. Camara et al. Mycologia 94:660, 2002. (3) B. S. Kim et al. Plant Pathol. J. 15:348, 1999. (4) B. M. Pryor and T. J. Michailides. Phytopathology 92:406, 2002.

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First Report of Nectria haematococca Causing Stem Rot of Soybean in China

Plant Disease ◽

10.1094/pdis-07-11-0626 ◽

2012 ◽

Vol 96 (3) ◽

pp. 457-457 ◽

Cited By ~ 1

Author(s):

Y. Gai ◽

R. Pan ◽

D. Xu ◽

M. Deng ◽

W. Chen ◽

...

Keyword(s):

Elongation Factor ◽

Morphological Characteristics ◽

Its Region ◽

Stem Rot ◽

Universal Primers ◽

Conidial Suspension ◽

Fruiting Bodies ◽

Nectria Haematococca ◽

First Report ◽

Soybean Seedlings

In October 2010, soybean (Glycine max) plants growing in commercial soybean fields in Zengcheng City, Guangdong Province developed symptoms consisting of stem and root rot, yellowing, and defoliation of leaves. Reddish, spherical fruiting bodies appeared in lesions that developed on stems. Plants with symptoms were sampled from fields. Fruiting bodies were excised from diseased tissues. Microscopic examination revealed that they were perithecia, globose to pyriform, and measured 197 to 260 μm in diameter and 226 to 358 μm long. When squeezed gently, cylindrical to clavate asci, 7.2 to 9.6 μm in diameter and 75.4 to 92.0 μm long, containing eight ascospores were exuded from the perithecia. Ascospores were ellipsoid to obovate, two celled, slightly constricted at the septum, had longitudinal striations, and measured 4.9 to 6.0 μm in diameter and 10.6 to 15.0 μm long. The fungus was isolated from the basal stem tissues of diseased soybean plants and cultured on potato dextrose agar (PDA) medium amended with streptomycin sulfate. On PDA, the culture developed into blue-pigmented colonies with whitish mycelium that produced oval to cylindrical microconidia. Microconidia had 0 to 1 septum, ranged from 2.5 to 5.2 × 7.6 to 29.4 μm, and were produced on monophialides. Macroconidia were cylindrical to falcate, thick walled, 2 to 5 septa, and 3.5 to 6.0 × 25.4 to 66.8 μm. Chlamydospores were present and ranged from 6.8 to 13.6 × 5.5 to 9.5 μm. Orange-to-reddish perithecia were readily formed in old culture. These morphological characteristics were consistent with descriptions of Nectria haematococca (anamorph Fusarium solani) (1). The rDNA internal transcribed spacer (ITS) region and the fragment of translation elongation factor 1-alpha (EF1-α) genes of the fungus were amplified, respectively, with universal primers ITS1/ITS4 and ef1/ef2 primers and sequenced. BLAST searches showed that the ITS sequences of three isolates (GenBank Accession Nos. JN015069, JN190942, and JN190943) had 99% similarity with those of N. haematococca(GenBank Accession Nos. DQ535186, DQ535185, and DQ535183) and the EF1-α sequences of three isolates (GenBank Accession Nos. JN874641, JN874642, and JN874643) had 100% similarity with those of F. solani (GenBank Accession Nos. DQ247265 and DQ247327). Completion of Koch's postulates confirmed the pathogenicity of the isolates in a replicated experiment. Thirty-day-old soybean seedlings of cultivar Huaxia No. 3 were inoculated by soaking their root systems in a conidial suspension (106 conidia per ml) for 30 min and then transplanted in plastic pots (20 cm in diameter) and incubated at 25 ± 2°C in a greenhouse. Control plants were treated with sterile water in the same way. There were four plants per pot and there were six replicates for each treatment. Within 3 weeks, more than 70% of the inoculated plants exhibited symptoms of leaf yellowing, stem rot, and root rots while control plants were symptomless. N. haematococca was reisolated from the diseased plants. To our knowledge, this is the first report of N. haematococca causing stem rot of soybean in China and the first description of sexual reproduction of F. solani causing soybean stem rot in nature. This pathogen may pose a serious threat to soybean production in China where soybean is a main crop. Reference: (1) C. Booth. The Genus Fusarium. CAB International, Wallingford, UK, 1971.

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First report of strains within the Pythium spinosum species complex causing carrot cavity spot in California

Plant Disease ◽

10.1094/pdis-08-21-1731-pdn ◽

2021 ◽

Author(s):

Misbah J. Chaudhry ◽

Jaspreet K. Sidhu ◽

Joe J Nuñez ◽

Jeroen T. F. Gillard ◽

Isolde M. Francis

Keyword(s):

Species Complex ◽

Tap Water ◽

Negative Control ◽

Coi Gene ◽

Universal Primers ◽

Peat Moss ◽

Tree Seedling ◽

First Report ◽

Daucus Carota L ◽

Cavity Spot

Carrots (Daucus carota L. subsp. sativus (Hoffm.)) with typical symptoms of cavity spot, i.e., sunken, round to elliptical lesions of 2-5 mm long (Hiltunen and White 2002), were collected from two locations in California, Bakersfield and Riverside, in January and July 2019, respectively. Carrots were rinsed in tap water, 4-mm2 lesion fragments were pressed into selective corn meal agar (CMA-PARP; Schrandt et al. 1994) and incubated at 23ºC in the dark for four days. Identification of pure cultures was performed via amplification and sequence analysis of two genomic regions, the Internal Transcribed Spacer 1-5.8S-ITS2 (ITS) region and the cytochrome C oxidase subunit 1 (COI) gene, using the universal primers UN-UP18S42/UN-LO28S576B (Schroeder et al. 2006) and OomCOXI-Levup/OomCOXI-Levlo (Robideau et al. 2011), respectively. Via BLAST, two isolates from organically grown carrots in Bakersfield (MCIF19) and Riverside (JSCS19), with identical ITS sequences (GenBank Acc. Nos. MZ799354 and MZ799355, respectively), showed 99.61% similarity (1021/1025 bp) to that of Pythium spinosum (AY598701.2). Yet, the COI of MCIF19 (MZ803207) showed 98.72% similarity (692/701 bp) to that of Pythium paroecandrum (GU071818.1), while the COI of JSCS19 (MZ803208) was identical (701/701 bp) to that of Pythium kunmingense (GU071820.1), a rarely isolated species considered within the species complex of P. spinosum (Robideau et al. 2011). According to these results, the isolates were identified as belonging to the P. spinosum species complex, part of Pythium Clade F (Lévesque and De Cock 2004; Robideau et al. 2011). Further research is needed to clarify the exact taxonomic status of both isolates. Koch’s postulates were completed using two different assays. Each assay was done twice and with carrots of the cultivar Maverick. Surface-sterilized, freshly harvested, mature carrots, in a plastic box lined with moistened sterile paper towels, were inoculated each with four CMA plugs (5-mm diameter) with actively growing mycelium of each isolate. CMA plugs, non-inoculated or colonized by a known pathogenic P. violae strain, were used as the negative and positive control, respectively. Boxes were closed to maintain humidity and incubated at 23ºC in the dark. Lesions similar to the ones caused by P. violae were observed at day 3 for all plugs of both isolates. No symptoms were observed for the negative control, even after extending the incubation to 7 days. In a more natural assay, four non-treated carrot seeds were planted in tree seedling pots (25 x 6.5 cm) containing sterilized 50/50 peat moss/sand combined with 15-ml V8 broth (Schrandt et al. 1994) with densely grown mycelium. The same inoculation treatments were used as for the carrot disk assay. Plants (one plant/pot, four plants/treatment) were maintained at 23ºC under a 16 h photoperiod with daily watering (20 ml). At 14 weeks, the carrots inoculated with P. violae and the two test isolates showed cavity spot lesions while no symptoms were observed on carrots growing in non-inoculated medium. For both assays, pathogens were re-isolated from rinsed symptomatic tissue and their identity was confirmed using the molecular analysis described above. No oomycetes were recovered from the non-inoculated carrots. Although several Pythium species have been associated with cavity spot before, this is, to our knowledge, the first report of strains within the P. spinosum species complex causing carrot cavity spot in California and elsewhere. Funding: This research was made possible by the California Fresh Carrot Advisory Board (FRA-21). References: Hiltunen, L.H., and White, J. G. 2002. Ann. Appl. Biol. 141:201. Lévesque, C. A., and De Cock, W. A. M. 2004. Mycol. Res. 108:1363. Robideau, G.P., et al. 2011. Mol. Ecol. Resour. 11:1002. Schrandt K. K., et al. 1994. Plant Dis. 78:335. Schroeder, K.L., et al. 2006. Phytopathology 96:637. Supplementary material: Supplementary figure S1 Supplementary figure S2

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