First Report of a New Postharvest Rot in Sweet Cherries Caused by Aureobasidium pullulans

During August to October 2012, several cherry packers in central Washington State reported that a significant volume of sweet cherries (Prunus avium) (cvs. Staccato, Sweetheart, and Lapin) were rotten by an unknown fungal pathogen after packing. Of 14 boxes (9 kg per box) of commercially packed cherries rejected by a retailer, the average incidence of the decay was 68%. Initial symptoms on infected fruit appeared as soft, slippery skin with tan discoloration and later skin cracking, epidermal breakdown, and severe pitting were observed. To isolate the causal agent, decayed fruit were rinsed with water, sprayed with 70% ethanol, and air-dried in a laminar hood. After removing the fruit skin with a sterile scalpel, small fragments of fruit flesh between decayed and healthy tissue were cut and placed on potato dextrose agar (PDA) acidified with 0.1% lactic acid. The plates were incubated at 20°C for 7 days and sub-cultured on PDA to obtain pure cultures. The colonies initially appeared white to cream, yeast-like, and later turned to light yellow to pink or brown with age. Conidia were hyaline, smooth-walled, single-celled, and ellipsoidal with variable shape and size. The fungus was identified as Aureobasidium pullulans (de Bary) G. Arnaud based on its morphology (1). The identity of three representative isolates were further confirmed by analysis of nucleotide sequences of the internal transcribed spacer (ITS) regions amplified using the primers ITS1/ITS4. A BLAST search showed that the sequences had 99% homology (E-value = 0.0) with that of A. pullulans deposited at GenBank (Accession No. JF440584.1). The nucleotide sequence of the isolate, A625, has been assigned GenBank Accession No. KF569512. To test pathogenicity, three single-spore isolates were grown on PDA at 20°C. Cultures grown on 10-day-old PDA were flooded with 20 ml of sterile deionized water, and the resulting conidial suspensions were filtered through two layers of cheesecloth and adjusted to 5 × 105 conidia/ml with a hemacytometer. Organic cherry fruit (cv. Bing for isolate A625 and cv. Sweetheart for isolates A755 and A757) were surface-disinfested in 0.6% sodium hypochlorite solution for 5 min, rinsed twice with deionized water, and air-dried. Ten fruit per replicate, four replications per treatment were inoculated with the conidial suspension using a hand sprayer and placed on sterilized wet paper towel in a plastic container. Control fruit were sprayed with sterile water. All fruit were incubated at 22 ± 1°C for 5 days. The experiments were conducted twice. The same symptoms of skin cracking and epidermal breakdown developed on 73% of the inoculated fruit, while no such symptoms appeared on the control fruit. Koch's postulates were fulfilled by re-isolating the fungus from the symptomatic fruit. A. pullulans, a ubiquitous saprophytic fungus on many fruits, has been reported as a causal agent of melting decay in grapes (2). To the best of our knowledge, this is the first report of postharvest fruit rot in sweet cherries caused by A. pullulans. References: (1) E. J. Hermanides-Nijhof. Aureobasidium and related genera. Pages 141-181 in: The Black Yeasts and Allied Hyphomycetes. Stud. Mycol. No. 15. Centraalbureau voor Schimmelcultures, Baarn, The Netherlands, 1977. (2) D. P. Morgan and T. J. Michailides. Plant Dis. 88:1047, 2004.

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First Report of Pepper Fruit Rot Caused by Fusarium concentricum in China

Plant Disease ◽

10.1094/pdis-03-13-0325-pdn ◽

2013 ◽

Vol 97 (12) ◽

pp. 1657-1657 ◽

Cited By ~ 6

Author(s):

J. H. Wang ◽

Z. H. Feng ◽

Z. Han ◽

S. Q. Song ◽

S. H. Lin ◽

...

Keyword(s):

Fusarium Species ◽

Fruit Rot ◽

Post Inoculation ◽

Distilled Water ◽

Conidial Suspension ◽

Average Growth ◽

First Report ◽

Control Fruit ◽

Pepper Fruit ◽

Fruit Samples

Pepper (Capsicum annuum L.) is an important vegetable crop worldwide. Some Fusarium species can cause pepper fruit rot, leading to significant yield losses of pepper production and, for some Fusarium species, potential risk of mycotoxin contamination. A total of 106 diseased pepper fruit samples were collected from various pepper cultivars from seven provinces (Gansu, Hainan, Heilongjiang, Hunan, Shandong, Shanghai, and Zhejiang) in China during the 2012 growing season, where pepper production occurs on approximately 25,000 ha. Pepper fruit rot symptom incidence ranged from 5 to 20% in individual fields. Symptomatic fruit tissue was surface-sterilized in 0.1% HgCl2 for 1 min, dipped in 70% ethanol for 30 s, then rinsed in sterilized distilled water three times, dried, and plated in 90 mm diameter petri dishes containing potato dextrose agar (PDA). After incubation for 5 days at 28°C in the dark, putative Fusarium colonies were purified by single-sporing. Forty-three Fusarium strains were isolated and identified to species as described previously (1,2). Morphological characteristics of one strain were identical to those of F. concentricum. Aerial mycelium was reddish-white with an average growth rate of 4.2 to 4.3 mm/day at 25°C in the dark on PDA. Pigments in the agar were formed in alternating red and orange concentric rings. Microconidia were 0- to 1-septate, mostly 0-septate, and oval, obovoid to allantoid. Macroconidia were relatively slender with no significant curvature, 3- to 5-septate, with a beaked apical cell and a foot-shaped basal cell. To confirm the species identity, the partial TEF gene sequence (646 bp) was amplified and sequenced (GenBank Accession No. KC816735). A BLASTn search with TEF gene sequences in NCBI and the Fusarium ID databases revealed 99.7 and 100% sequence identity, respectively, to known TEF sequences of F. concentricum. Thus, both morphological and molecular criteria supported identification of the strain as F. concentricum. This strain was deposited as Accession MUCL 54697 (http://bccm.belspo.be/about/mucl.php). Pathogenicity of the strain was confirmed by inoculating 10 wounded, mature pepper fruits that had been harvested 70 days after planting the cultivar Zhongjiao-5 with a conidial suspension (1 × 106 spores/ml), as described previously (3). A control treatment consisted of inoculating 10 pepper fruits of the same cultivar with sterilized distilled water. The fruit were incubated at 25°C in a moist chamber, and the experiment was repeated independently in triplicate. Initially, green to dark brown lesions were observed on the outer surface of inoculated fruit. Typical soft-rot symptoms and lesions were observed on the inner wall when the fruit were cut open 10 days post-inoculation. Some infected seeds in the fruits were grayish-black and covered by mycelium, similar to the original fruit symptoms observed at the sampling sites. The control fruit remained healthy after 10 days of incubation. The same fungus was isolated from the inoculated infected fruit using the method described above, but no fungal growth was observed from the control fruit. To our knowledge, this is the first report of F. concentricum causing a pepper fruit rot. References: (1) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Ames, IA, 2006. (2) K. O'Donnell et al. Proc. Nat. Acad. Sci. USA 95:2044, 1998. (3) Y. Yang et al. 2011. Int. J. Food Microbiol. 151:150, 2011.

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First Report of Anthracnose on Peach Fruit Caused by Colletotrichum truncatum in South Carolina

Plant Disease ◽

10.1094/pdis-12-13-1215-pdn ◽

2014 ◽

Vol 98 (8) ◽

pp. 1154-1154 ◽

Cited By ~ 4

Author(s):

A. Grabke ◽

M. Williamson ◽

G. W. Henderson ◽

G. Schnabel

Keyword(s):

South Carolina ◽

Causal Agent ◽

Distilled Water ◽

Conidial Suspension ◽

Sterile Water ◽

Colletotrichum Truncatum ◽

Peach Fruit ◽

First Report ◽

Anthracnose Disease ◽

Control Fruit

In July 2013, two diseased peach fruit (Prunus persica (L.) Stokes) of the cv. Sweet Dream were collected from a commercial orchard in Ridge Springs, South Carolina. Affected peaches were at or near maturity and symptoms resembled anthracnose disease caused by Colletotrichum spp. with circular sunken tan to brown lesions that were firm in touch, and had wrinkled concentric rings. The center of the lesion was covered with black acervuli containing setae. To isolate the causal agent, the two symptomatic fruit were surface-sterilized in 10% bleach for 2 min and rinsed with sterile distilled water. Lesions were cut in half, and necrotic tissue from the inside of the fruit was placed on acidified potato dextrose agar (APDA). Flat colonies covered with olive-gray to iron-gray acervuli developed on APDA incubated at 22°C with a 12-h cycle of fluorescent light and darkness. Morphology of acervuli, setae (avg. 90 to 160 μm), conidiophores (up to 90 um long), and conidia (avg. 22 × 3.8 μm) of single spore isolates were consistent with descriptions of Colletotrichum truncatum (Schwein.) Andrus & W.D. Moore (3), a causal agent of anthracnose disease. Genomic DNA was extracted from isolate Ct_RR13_1 using the MasterPure Yeast DNA Purification Kit (Epicentre, Madison, WI). The ribosomal ITS1-5.8S-ITS2 region and a partial sequence of the actin gene were amplified with primer pair ITS1 and ITS4 (4), and primer pair ACT-512F and ACT-783A (2), respectively. A multilocus sequence identification in Q-bank Fungi revealed a 100% similarity with C. truncatum (1). The C. truncatum sequences from the peach isolate were submitted to GenBank (accessions KF906258 and KF906259). Pathogenicity of isolate Ct_RR13_1 was confirmed by inoculating five mature but still firm peach fruits with a conidial suspension of C. truncatum. Peaches were washed with soap and water, surface-disinfected for 2 min with 10% bleach, rinsed with sterile distilled water, and air dried. Dried fruit were stabbed at three equidistant points, each about 2 cm apart, to a depth of 9.5 mm using a sterile 26G3/8 beveled needle (Becton Dickinson & Co., Rutherford, NJ). For inoculation, a 30-μl droplet of conidia suspension prepared in distilled, sterile water (1 to 2 × 104 spores/ml) was placed on each wound; control fruit received sterile water without conidia. Fruit were incubated at 22°C for 2 days at 100% humidity and another 12 days at 70% humidity. Inoculated fruit developed anthracnose symptoms with sporulating areas as described above and the fungus was re-isolated. All control fruit remained healthy. C. truncatum has a wide host range, including legumes and solanaceous plants of the tropics, and is especially common in the Fabaceae family. Its occurrence in a commercial peach orchard is worrisome because control measures may need to be developed that are different from those developed for endemic species, i.e. C. acutatum and C. gloeoporioides, due to differences in disease cycle or fungicide sensitivity. To our knowledge, this is the first report of C. truncatum causing anthracnose on a member of the genus Prunus. References: (1) P. Bonants et al. EPPO Bull. 43:211, 2013. (2) I. Carbone et al. Mycologia 91:553, 1999. (3) U. Damm et al. Fungal Divers. 39:45, 2009. (4) T. J. White et al. Pages 315-322 in: PCR Protocols: A Guide to Methods and Application. Academic Press, NY, 1993.

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First Report of Fruit Rot on Schisandra chinensis Caused by Penicillium glabrum and P. adametzii in China

Plant Disease ◽

10.1094/pdis-08-12-0799-pdn ◽

2013 ◽

Vol 97 (2) ◽

pp. 288-288 ◽

Cited By ~ 2

Author(s):

C. Q. Chen ◽

Y. Zhi ◽

J. Gao

Keyword(s):

Its Region ◽

Causal Agent ◽

Surface Strain ◽

Fruit Rot ◽

Conidial Suspension ◽

Schisandra Chinensis ◽

Sterile Water ◽

Differential Growth ◽

First Report ◽

Sequence Identity

Schisandra (Schisandra chinensis (Turcz.) Baill) is an important medicinal herb in China, which is mainly used for treatment of insomnia and memory decay. In September 2010, rot was observed on approximately 5% of the fruits during ripening of schisandra in several orchards at Linjiang City and Ji'an City, Jilin Province. Watery spots on infected fruits of schisandra appeared at the end of ripening, and then the fruits darkened in color, shrunk, and turned soft. The surface of the lesions became covered with masses of blue-green mycelium, conidiophores, and conidia under high humidity. To isolate the causal agent, conidia and conidiophores were suspended in sterile water and streaked onto the surface of potato dextrose agar (PDA). Single hyphal tips were then transferred to new PDA plates to be purified. The isolates were then cultured on CYA (Czapek yeast extract agar) and two kinds of strains (wwzqm1 and wwzqm2) were established based on differential growth rate, microscopic features, and colony color. Pathogenicity of each strain was tested on 25 healthy mature fruits of schisandra cv. Red Pearl by inoculating the fruit surface with a 15 μl conidial suspension (106 conidia/ml). Control fruits were treated with sterile water. Fruits were kept at 25°C and 90% relative humidity. After a 5-day incubation, symptoms described above were observed on all inoculated fruits, whereas all control fruits were symptomless. The causal agent was reisolated, confirming Koch's postulates. Strain wwzqm1 was identified as Penicillium glabrum (Wehmer) Westling on the basis of its morphology. Conidiophores arose from basal hyphae, with stipes smooth or finely roughened, 50.5 to 300.0 × 2.5 to 3.5 μm, penicillus monoverticillate, and bearing verticils of 8 to 12 phialides. Conidia were globose to subglobose, approximately 2.9 to 3.5 μm in diameter, with smooth or nearly smooth walls, and conidial chains in compact columns. Colonies grown for 7 days on CYA at 25°C attained a diameter of 32.4 to 39.1 mm, with the center of a deep bluish green, plane, velutinous, and heavily sporulated. Margin mycelium of the colony was white, with a yellowish brown under surface. Strain wwzqm2 was identified as P. adametzii K. M. Zalessky according to its morphological features with conidiophores born from funicolose hyphae, stipes smooth, 10.2 to 20.1 × 1.5 to 2.5 μm, penicillus monoverticillate, and bearing verticils of 4 to 6 phialides. Conidia were nearly globose to glubose, 2.0 to 2.7 μm in diameter, with smooth or finely roughened walls, and conidial chains loose and irregular. Colonies on CYA at 25°C for 7 days grew rather fast and reached a diameter of 50.3 to 60.2 mm, and were deep grayish green, near plane, floccose or funicolose, and heavily sporulated. Margin mycelium of the colony was white with a yellowish brown under surface (1). The internal transcribed spacer (ITS) region of ribosomal DNA (rDNA) was amplified for wwzqm1 and wwzqm2 using primers ITS4/ITS5 and sequenced. BLASTn analysis of the 595 bp wwzqm1 (GenBank Accession No. JN887323) amplicon had 99% sequence identity with P. glabrum (DQ681321) and the 568 bp wwzqm2 amplicon (JN887322) had 99% sequence identity with P. adametzii (AF033401). To our knowledge, this is the first report of fruit rot on S. chinensis caused by P. glabrum and P. adametzii in China. Reference: (1). H. Z. Kong. Penicillium et Teleomorphi Cognati. Flora Fungorum Sinicorum. Science Press, Beijing, 35:43, 2007.

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First Report of Alternaria alternata Causing Fruit Rot on Fig (Ficus carica) in Montenegro

Plant Disease ◽

10.1094/pdis-07-13-0770-pdn ◽

2014 ◽

Vol 98 (3) ◽

pp. 424-424 ◽

Cited By ~ 2

Author(s):

N. Latinović ◽

S. Radišek ◽

J. Latinović

Keyword(s):

Alternaria Alternata ◽

Direct Sequencing ◽

Disease Incidence ◽

Morphological Characteristics ◽

Fruit Rot ◽

Conidial Suspension ◽

Culture Collections ◽

First Report ◽

Control Fruit ◽

Rot Disease

In July 2012, a fruit rot disease was observed in several commercial fig tree orchards located in the Podgorica region in Montenegro. Symptoms on fruits initially appeared as small circular to oval, light brown, necrotic, sunken spots located mostly on the areas surrounding the ostiolar canal with an average diameter of 5 to 10 mm, which gradually enlarged in size leading to total fruit rot. Disease incidence on fruit across the fields ranged from 15 to 20% but the disease did not increase further due to hot and dry conditions thereafter. No foliar symptoms were observed. Small pieces (5 mm2) of symptomatic fruits were excised from the junction of diseased and healthy tissue, surface sterilized in 70% ethanol solution for 1 min, washed in three changes of sterile distilled water, air dried, and transferred to potato dextrose agar (PDA). After 2 to 3 days of incubation at 25°C, a fungus was consistently isolated. The isolates had radial growth and produced sooty black colonies. Microscopic observations of the colonies revealed brown septate hyphae and simple or branched conidiophores 30 to 65 μm long and 3 to 4.5 μm wide. Dark brown conidia were in chains (3 to 7), sized 10 to 35 × 5 to 9 μm, ellipsoid to ovoid, with 2 to 5 transverse and a few (1 to 3) to no longitudinal septa. Based on morphological characteristics, the fungus was identified as Alternaria alternata (3). For molecular identification, DNA was extracted from mycelia and conidia of two representative single spore isolates designated as ALT1-fCG and ALT2-fCG. PCR was carried out using internal transcribed spacer (ITS) region primers ITS4/ITS5 and A. alternata species-specific primers AAF2/AAR3 (1). Both primer pairs gave PCR products that were subjected to direct sequencing. BLAST analysis of the 546-bp ITS4/ITS5 (KF438091) and 294-bp AAF2/AAR3 (KF438092) sequences revealed 100% identity with several A. alternata isolates. Pathogenicity tests were conducted on 30 detached almost ripe and healthy fig fruit (cv. Primorka) by spraying them with a conidial suspension of the isolated fungus (106 conidia/ml) with a handheld sprayer. Thirty fruit inoculated with sterile water served as the non-inoculated control. Inoculated and control fruit were kept in a moist chamber at 25°C. Symptoms appeared on inoculated fruit 2 to 3 days after inoculation and all fruit were completely rotted 5 to 6 days after inoculation. Control fruit did not display any symptoms. A. alternata was consistently re-isolated from inoculated fruit, fulfilling Koch's postulates. The fig fruit rot caused by A. alternata has been reported before in California (2) and elsewhere mainly as postharvest pathogen. To our knowledge, this is the first report of fruit rot caused by A. alternata on fig in Montenegro. Considering Podgorica as the largest fig-producing area and the importance of fig as a traditionally grown crop, it could pose a threat to fig production in Montenegro. Voucher specimens are available at the culture collections of the University of Montenegro, Biotechnical Faculty. References: (1) P. Konstantinova et al. Mycol. Res. 106:23, 2002. (2) T. J. Michailides et al. Plant Dis. 78:44-50, 1994. (3) E. G. Simmons. Page 775 in: Alternaria and Identification Manual. CBS Fungal Biodiversity Centre, 2007.

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First Report of Colletotrichum truncatum Causing Anthracnose of Tomato in China

Plant Disease ◽

10.1094/pdis-05-13-0491-pdn ◽

2014 ◽

Vol 98 (5) ◽

pp. 687-687 ◽

Cited By ~ 8

Author(s):

Y. Z. Diao ◽

C. Zhang ◽

D. Lin ◽

X. L. Liu

Keyword(s):

Tomato Fruit ◽

Consensus Sequence ◽

Fruit Rot ◽

Control Treatment ◽

Conidial Suspension ◽

Colletotrichum Truncatum ◽

First Report ◽

Colony Diameter ◽

Control Fruit ◽

Ripe Tomato

Colletotrichum truncatum (syn. C. capsici) is an important plant pathogen that has a wide host range including pepper, eggplant, muskmelon, chickpea, grapes, and many other species of plants (1,2). Anthracnose fruit rot incited by C. coccodes is a prevalent disease in some major tomato (Solanum esculentum) producing regions in China. In October 2012, anthracnose symptoms (circular, sunken lesions with spore masses produced in black acervuli) were observed on ripe tomato fruit in Fuzhou City of Fujian Province, China. Three single-spore isolates (FZ-1, FZ-2, and FZ-3) were derived from fungal cultures isolated from different infected fruits from the same farm. A mycelial plug (5 mm in diameter) from the growing edge of an active colony of each isolate was transferred onto potato dextrose agar (PDA) and incubated at 28°C. Colonies grown on PDA changed from grayish to dark grey with an average colony diameter of 71.8 mm after 7 days. Conidia were falcate and 17.6 to 21.6 × 2.57 to 3.31 μm. Growth rate measured by colony diameter was greater at 25 to 30°C than at other temperatures tested (12, 18, 20, and 37°C). Based upon these morphological and cultural characteristics, the causal agent was identified as C. truncatum (3). To test the three isolates for pathogenicity, detached ripe tomato fruits were each inoculated with 1 μl of a conidial suspension (106 conidia/ml) using either injection or applying a droplet of the spore suspension on the surface of each fruit; the control treatment consisted of fruit that was treated with 1 μl of sterilized water using the two methods of inoculation mentioned above. Five replicate ripe fruits were inoculated with each of the three isolates using each method described above, and incubated in a moist chamber at 25°C. Five ripe fruits were used the negative control treatment for each inoculation method. After 7 days, typical anthracnose symptoms had developed on the inoculated fruit but not on control fruit. To confirm identity of the three isolates, the internal transcribed spacer (ITS) region of ribosomal DNA (rDNA), the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene, a partial sequence of the actin (ACT) gene, and partial histone 3 (HIS3) gene were amplified with ITS1 and ITS4 universal primers, GDF1 and GDR1 primers, ACT-512F and ACT-783R primers, and CYLH3F and CYLH3R primers, respectively (1). The ITS consensus sequence (Accession No. KC460308) of these isolates shared 99% homology with the ITS sequence of C. truncatum in GenBank (AJ301945), and the three other consensus sequences were all 100% homologous with the corresponding sequences of C. truncatum in Genbank (GU228254, GU227960, and GU228058, respectively). The pathogen was re-isolated from the inoculated fruit but not from the control fruit, and the identity of the re-isolates was confirmed to be C. truncatum by morphological features and based on the ITS, GAPDH, ACT, and HIS3 sequences as described above. To our knowledge, this is the first report of C. truncatum causing anthracnose on tomato in China. References: (1) U. Damm et al. Fungal Divers. 39:45, 2009. (2) K. K. Pandey. J. Mycol. Plant Pathol. 36:104, 2006. (3) I. S. Sawant et al. New Dis. Rep. 25:2, 2012.

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Yellow Net of Triumffeta Is Caused by a Geminivirus: A First Report

Plant Disease ◽

10.1094/pdis.1998.82.1.127a ◽

1998 ◽

Vol 82 (1) ◽

pp. 127-127 ◽

Cited By ~ 2

Author(s):

Vipin Hallan ◽

Sangeeta Saxena ◽

B. P. Singh

Keyword(s):

Rainy Season ◽

Blot Hybridization ◽

Southern Blot Hybridization ◽

Viral Disease ◽

Hybridization Experiment ◽

Dorsal Side ◽

Causal Agent ◽

Degenerate Primers ◽

First Report ◽

Initial Symptoms

Triumffeta rhomboidiaceae Jacq. (Tiliaceae family) is an annual rainy season weed that is commonly found throughout India. For the last 3 years, during the rainy season, several plants of T. rhomboidiaceae in and around the gardens of the National Botanical Research Institute have been found with vein yellowing symptoms. The initial symptoms were vein clearing but in later stages the veins became yellow and thickened. In severe cases, the chlorosis extends into interveinal areas, resulting in complete yellowing of the leaves. In a few cases, green leafy or thorny enations could be seen on the dorsal side of the leaf. The disease was investigated to identify the causal agent. Vector transmission studies showed that the causal agent is transmitted by the whitefly, Bemisia tabaci, from infected to healthy seedlings of T. rhomdoidiaceae. Since whitefly transmission of the disease is consistent with a geminivirus as the causal agent, the role of such a virus was investigated. DNA isolated from Triumffeta plants (both from the infected plants in the field as well as from those inoculated experimentally in the greenhouse) showing above mentioned symptoms was amplified with two sets of degenerate primers, PAL1v1978/PAR1c496 (set 1) and PAL1v1978/PCRc1 (set 2), that have been shown to be specific for DNA-A of whitefly transmitted geminiviruses (WTGs), in polymerase chain reaction (1). We could amplify DNA-A fragments of approximately 1.2 kb from set 1 and 0.7 kb from set 2, as expected (1). DNA isolated from healthy seedlings gave no amplification of such fragments. Identification of the amplified DNA fragments (from infected samples) to be of geminiviral in nature was confirmed by Southern blot hybridization carried out under high stringency conditions. DNA-A of Indian tomato leaf curl virus (2) was used as a general probe for WTGs for the above hybridization experiment. Therefore, Triumffeta yellow net disease is caused by a geminivirus. A review of literature revealed that there is no record of a viral disease affecting this weed and, therefore, this is the first report of a viral disease affecting this plant. References: (1) M. R. Rojas et al. Plant Dis. 77:340, 1993. (2) K. M. Srivastava et al. J. Virol. Methods 51:297, 1995.

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First Report of Leptosphaeria biglobosa ‘brassicae’ Causing Blackleg on Brassica juncea var. multisecta in China

Plant Disease ◽

10.1094/pdis-10-20-2173-pdn ◽

2021 ◽

Author(s):

Yuexuan Long ◽

Mingxue Shang ◽

Yue Deng ◽

Chuan Yu ◽

Mingde Wu ◽

...

Keyword(s):

Brassica Juncea ◽

Yellow Pigment ◽

Causal Agent ◽

Control Group ◽

Conidial Suspension ◽

First Report ◽

Fungal Isolates ◽

Dna Fragment ◽

Leptosphaeria Biglobosa ◽

Β Tubulin

Brassica juncea var. multisecta, a leafy mustard, is widely grown in China as a vegetable (Fahey 2016). In May 2018, blackleg symptoms, grayish lesions with black pycnidia, were found on stems and leaves of B. juncea var. multisecta during disease surveys in Wuhan, Hubei Province. Disease incidence was approximately 82% of plants in the surveyed fields (~1 ha in total). To determine the causal agent of the disease, twelve diseased petioles were surface-sterilized and then cultured on potato dextrose agar (PDA) at 20˚C for 5 days. Six fungal isolates (50%) were obtained. All showed fluffy white aerial mycelia on the colony surface and produced a yellow pigment in PDA. In addition, pink conidial ooze formed on top of pycnidia after 20 days of cultivation on a V8 juice agar. Pycnidia were black-brown and globose with average size of 145 × 138 μm and ranged between 78 to 240 × 71 to 220 μm, n = 50. The conidia were cylindrical, hyaline, and 5.0 × 2.1 μm (4 to 7.1 × 1.4 to 2.9 μm, n=100). These results indicated that the fungus was Leptosphaeria biglobosa rather than L. maculans, as only the former produces yellow pigment (Williams and Fitt 1999). For molecular confirmation of identify, genomic DNAs were extracted and tested through polymerase chain reaction (PCR) assay using the species-specific primers LbigF, LmacF, and LmacR (Liu et al. 2006), of which DNA samples of L. maculans isolate UK-1 (kindly provided by Dr. Yongju Huang of University of Hertfordshire) and L. biglobosa ‘brassicae’ isolate B2003 (Cai et al. 2014) served as controls. Moreover, the sequences coding for actin, β-tubulin, and the internal transcribed spacer (ITS) region of ribosomal DNA (Vincenot et al. 2008) of isolates HYJ-1, HYJ-2 and HYJ-3 were also cloned and sequenced. All six isolates only produced a 444-bp DNA fragment, the same as isolate B2003, indicating they belonged to L. biglobosa ‘brassicae’, as L. maculans generates a 331-bp DNA fragment. In addition, sequences of ITS (GenBank accession no. MN814012, MN814013, MN814014), actin (MN814292, MN814293, MN814294), and β-tubulin (MN814295, MN814296, MN814297) of isolates HYJ-1, HYJ-2 and HYJ-3 were 100% identical to the ITS (KC880981), actin (AY748949), and β-tubulin (AY748995) of L. biglobosa ‘brassicae’ strains in GenBank, respectively. To determine their pathogenicity, needle-wounded cotyledons (14 days) of B. juncea var. multisecta ‘K618’ were inoculated with a conidial suspension (1 × 107 conidia/ml, 10 μl per site) of two isolates HYJ-1 and HYJ-3, twelve seedlings per isolate (24 cotyledons), while the control group was only treated with sterile water. All seedlings were incubated in a growth chamber (20°C, 100% relative humidity under 12 h of light/12 h of dark) for 10 days. Seedlings inoculated with conidia showed necrotic lesions, whereas control group remained asymptomatic. Two fungal isolates showing the same culture morphology to the original isolates were re-isolated from the necrotic lesions. Therefore, L. biglobosa ‘brassicae’ was confirmed to be the causal agent of blackleg on B. juncea var. multisecta in China. L. biglobosa ‘brassicae’ has been reported on many Brassica crops in China, such as B. napus (Fitt et al. 2006), B. oleracea (Zhou et al. 2019), B. juncea var. multiceps (Zhou et al. 2019), B. juncea var. tumida (Deng et al. 2020). To our knowledge this is the first report of L. biglobosa ‘brassicae’ causing blackleg on B. juncea var. multisecta in China, and its occurrence might be a new threat to leafy mustard production of China.

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

Plant Disease ◽

10.1094/pdis.2003.87.2.203a ◽

2003 ◽

Vol 87 (2) ◽

pp. 203-203

Author(s):

S. T. Koike ◽

S. A. Tjosvold ◽

J. Z. Groenewald ◽

P. W. Crous

Keyword(s):

Leaf Spot ◽

Sequence Data ◽

Disease Incidence ◽

Leaf Spot Disease ◽

Conidial Suspension ◽

Internal Transcribed Spacer Region ◽

First Report ◽

Spot Disease ◽

Leaf Spots ◽

Initial Symptoms

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

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First Report of Diaporthe hongkongensis Causing Fruit rot on Peach (Prunus persica) in China

Plant Disease ◽

10.1094/pdis-07-20-1505-pdn ◽

2021 ◽

Author(s):

Zhou Zhang ◽

Zheng Bing Zhang ◽

Yuan Tai Huang ◽

FeiXiang Wang ◽

Wei Hua Hu ◽

...

Keyword(s):

Prunus Persica ◽

Fruit Rot ◽

Hunan Province ◽

Post Inoculation ◽

Distilled Water ◽

Conidial Suspension ◽

Internal Transcribed Spacer Region ◽

First Report ◽

Representative Isolate ◽

Rot Disease

Peach [Prunus persica (L.) Batsch] is an important deciduous fruit tree in the family Rosaceae and is a widely grown fruit in China (Verde et al., 2013). In July and August 2018, a fruit rot disease was observed in a few peach orchards in Zhuzhou city, the Hunan Province of China. Approximately 30% of the fruit in more than 400 trees was affected. Symptoms displayed were brown necrotic spots that expanded, coalesced, and lead to fruit being rotten. Symptomatic tissues excised from the margins of lesions were surface sterilized in 70% ethanol for 10 s, 0.1% HgCl2 for 2 min, rinsed with sterile distilled water three times, and incubated on potato dextrose agar (PDA) at 26°C in the dark. Fungal colonies with similar morphology developed, and eight fungal colonies were isolated for further identification. Colonies grown on PDA were grayish-white with white aerial mycelium. After an incubation period of approximately 3 weeks, pycnidia developed and produced α-conidia and β-conidia. The α-conidia were one-celled, hyaline, fusiform, and ranged in size from 6.0 to 8.4 × 2.1 to 3.1 μm, whereas the β-conidia were filiform, hamate, and 15.0 to 27.0 × 0.8 to 1.6 μm. For molecular identification, total genomic DNA was extracted from the mycelium of a representative isolate HT-1 and the internal transcribed spacer region (ITS), β-tubulin gene (TUB), translation elongation factor 1-α gene (TEF1), calmodulin (CAL), and histone H3 gene (HIS) were amplified and sequenced (Meng et al. 2018). The ITS, TUB, TEF1, CAL and HIS sequences (GenBank accession nos. MT740484, MT749776, MT749778, MT749777, and MT749779, respectively) were obtained and in analysis by BLAST against sequences in NCBI GenBank, showed 99.37 to 100% identity with D. hongkongensis or D. lithocarpus (the synonym of D. hongkongensis) (Gao et al., 2016) (GenBank accession nos. MG832540.1 for ITS, LT601561.1 for TUB, KJ490551.1 for HIS, KY433566.1 for TEF1, and MK442962.1 for CAL). Pathogenicity tests were performed on peach fruits by inoculation of mycelial plugs and conidial suspensions. In one set, 0.5 mm diameter mycelial discs, which were obtained from an actively growing representative isolate of the fungus on PDA, were placed individually on the surface of each fruit. Sterile agar plugs were used as controls. In another set, each of the fruits was inoculated by application of 1 ml conidial suspension (105 conidia/ml) by a spray bottle. Control assays were carried out with sterile distilled water. All treatments were maintained in humid chambers at 26°C with a 12-h photoperiod. The inoculation tests were conducted twice, with each one having three fruits as replications. Six days post-inoculation, symptoms of fruit rot were observed on inoculated fruits, whereas no symptoms developed on fruits treated with agar plugs and sterile water. The fungus was re-isolated and identified to be D. hongkongensis by morphological and molecular methods, thus fulfilling Koch’s Postulates. This fungus has been reported to cause fruit rot on kiwifruit (Li et al. 2016) and is also known to cause peach tree dieback in China (Dissanayake et al. 2017). However, to our knowledge, this is the first report of D. hongkongensis causing peach fruit rot disease in China. The identification of the pathogen will provide important information for growers to manage this disease.

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First Report of Curvularia aeria on Etlingera linguiformis from Nagaland, India

Plant Disease ◽

10.1094/pdis-01-14-0060-pdn ◽

2014 ◽

Vol 98 (11) ◽

pp. 1580-1580 ◽

Cited By ~ 2

Author(s):

C. Kithan ◽

L. Daiho

Keyword(s):

Leaf Surface ◽

Agricultural Research ◽

Disease Incidence ◽

Indian Agricultural Research Institute ◽

Mountain Range ◽

Pathogenicity Test ◽

Distilled Water ◽

Conidial Suspension ◽

First Report ◽

Initial Symptoms

Etlingera linguiformis (Roxb.) R.M.Sm. of Zingiberaceae family is an important indigenous medicinal and aromatic plant of Nagaland, India, that grows well in warm climates with loamy soil rich in humus (1). The plant rhizome has medicinal benefits in treating sore throats, stomachache, rheumatism, and respiratory complaints, while its essential oil is used in perfumery. A severe disease incidence of leaf blight was observed on the foliar portion of E. linguiformis at the Patkai mountain range of northeast India in September 2012. Initial symptoms of the disease are small brown water soaked flecks appearing on the upper leaf surface with diameter ranging from 0.5 to 3 cm, which later coalesced to form dark brown lesions with a well-defined border. Lesions often merged to form large necrotic areas, covering more than 90% of the leaf surface, which contributed to plant death. The disease significantly reduces the number of functional leaves. As disease progresses, stems and rhizomes were also affected, reducing quality and yield. The diseased leaf tissues were surface sterilized with 0.2% sodium hypochlorite for 2 min followed by rinsing in sterile distilled water and transferred into potato dextrose agar (PDA) medium. After 3 days, the growing tips of the mycelium were transferred to PDA slants and incubated at 25 ± 2°C until conidia formation. Fungal colonies on PDA were dark gray to dark brown, usually zonate; stromata regularly and abundantly formed in culture. Conidia were straight to curved, ellipsoidal, 3-septate, rarely 4-septate, middle cells broad and darker than other two end cells, middle septum not median, smooth, 18 to 32 × 8 to 16 μm (mean 25.15 × 12.10 μm). Conidiophores were terminal and lateral on hyphae and stromata, simple or branched, straight or flexuous, often geniculate, septate, pale brown to brown, smooth, and up to 800 μm thick (2,3). Pathogen identification was performed by the Indian Type Culture Collection, Division of Plant Pathology, Indian Agricultural Research Institute, New Delhi (ITCC Accession No. 7895.10). Further molecular identity of the pathogen was confirmed as Curvularia aeria by PCR amplification and sequencing of the internal transcribed spacer (ITS) regions of the ribosomal DNA by using primers ITS4 and ITS5 (4). The sequence was submitted to GenBank (Accession No. MTCC11875). BLAST analysis of the fungal sequence showed 100% nucleotide similarity with Cochliobolus lunatus and Curvularia aeria. Pathogenicity tests were performed by spraying with an aqueous conidial suspension (1 × 106 conidia /ml) on leaves of three healthy Etlingera plants. Three plants sprayed with sterile distilled water served as controls. The first foliar lesions developed on leaves 7 days after inoculation and after 10 to 12 days, 80% of the leaves were severely infected. Control plants remained healthy. The inoculated leaves developed similar blight symptoms to those observed on naturally infected leaves. C. aeria was re-isolated from the inoculated leaves, thus fulfilling Koch's postulates. The pathogenicity test was repeated twice. To our knowledge, this is the first report of the presence of C. aeria on E. linguiformis. References: (1) M. H. Arafat et al. Pharm. J. 16:33, 2013. (2) M. B. Ellis. Dematiaceous Hyphomycetes. CMI, Kew, Surrey, UK, 1971. (3) K. J. Martin and P. T. Rygiewicz. BMC Microbiol. 5:28, 2005. (4) C. V. Suberamanian. Proc. Indian Acad. Sci. 38:27, 1955.

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