First report of Phytophthora chlamydospora causing postharvest fruit rot on apples and pears in the Netherlands

Apple (Malus domestica) and pear (Pyrus communis) are important fruit crops in the Netherlands, with total production of 269,000 tons and 402,000 tons in 2018, respectively. In 2018 and 2019 postharvest fruit rots were observed on the apple variety Elstar (one observation) and pear varieties Conference and Doyenné du Comice (multiple observations). The symptoms were found after storage in controlled atmosphere storage facilities on fruits from different orchards across the Netherlands. Disease incidences up to 50% of the stored fruit were observed. The diseased fruits showed circular brown to black spots with irregular and diffuse margins that enlarged rapidly to form distinctive rings, typical of Phytophthora infection. Several Phytophthora species are currently known to cause fruit rot of pome fruit (Sanchez et al. 2019). To isolate the causal agent, small portions of fruit flesh from decayed fruit were excised from the lesion margin and placed on potato dextrose agar (PDA). The plates were incubated at 20°C in the dark, and pure cultures were obtained by transferring hyphal tips on PDA. The colonies were white with petaloid and rosette-shaped patterns. The isolates grown on PDA formed irregularly branched hyphae, produced persistent non-papillate sporangia, usually on unbranched sporangiophores and chlamydospores were produced. The characteristics were similar to those described for Phytophthora chlamydospora Brasier and Hansen sp. nov. (Hansen et al. 2015). The identity of three representative isolates (KP00219, WURR121 and WURR119) from two different pear cultivars (Conference and Doyenné) and one apple cultivar (Elstar), respectively, was confirmed by means of multilocus gene sequencing. Genomic DNA was extracted using the LGC Mag Plant Kit (Berlin, Germany) in combination with the Kingfisher method (Waltham, USA). Sequences of ITS region, COX and EF were amplified and sequenced. The sequences have been deposited in GenBank (Accession Nos. MT125889, MT125891, and MT125890 [ITS], MT153610, MT153612, and MT153611 [COX], MT153613, MT153615, and MT153614 [EF]. MegaBLAST analysis revealed that our ITS, COX and EF sequences matched with 100% identity to Phytophthora chlamydospora isolates in GenBank AF541901 and AF541902 (ITS), JF771548 and JF771549 (COX), JN936005 and JN936006 (EF). In order to perform Koch’s postulates a pathogenicity assay was performed using mycelial plugs of the cultures KP00219, on pear cv. Conference, and WURR119 and WURR121, on apple cv. Elstar and pear cv. Doyenné du Comice. Ten apples and pears per cultivar were disinfected, and wounded using a sterile cork borer in the middle of the fruit surface area. A mycelial plug of a two weeks old fungal culture was then placed onto the fruit. Placement of a PDA plug without fungal growth was used as a control. The fruits were incubated at 18˚C at high relative humidity for 7 days. Symptoms appeared within 3 days on all fruits. Mock-inoculated controls remained symptomless. The fungus was reisolated and confirmed as P. chlamydospora by morphology and sequencing. P. chlamydospora is found in streams and wet soil worldwide, and has only rarely been recovered as a pathogen from ornamental and woody species (Blomquist et al. 2012; Ginetti et al. 2014; Türkölmez et al. 2016). To our knowledge, this is the first report confirming P. chlamydospora as a causal agent of fruit rot of commercially produced apple and pear cultivars in the Netherlands.

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First Report of Phytophthora ramorum on Viburnum bodnantense in Belgium

Plant Disease ◽

10.1094/pdis.2003.87.2.203c ◽

2003 ◽

Vol 87 (2) ◽

pp. 203-203 ◽

Cited By ~ 11

Author(s):

D. De Merlier ◽

A. Chandelier ◽

M. Cavelier

Keyword(s):

The Netherlands ◽

Plant Protection ◽

Morphological Characteristics ◽

Pcr Primers ◽

Selective Medium ◽

Phytophthora Species ◽

Phytophthora Ramorum ◽

Pathogenicity Test ◽

First Report ◽

Root Necrosis

In the past decade, a new Phytophthora species inducing shoot canker on Rhododendron and dieback of Viburnum has been observed in Europe, mainly in Germany and the Netherlands, and California. This new pathogen has been named Phytophthora ramorum (3). In May 2002, a diseased Viburnum plant (Viburnum bodnantense) from the Plant Protection Service (Ministry of Agriculture, Belgium) was submitted to our laboratory for diagnosis. Symptoms included wilting, leaves turning from green to brown, discolored vascular tissues, and root necrosis. The plant came from a Belgian ornamental nursery that obtained supplies of stock plants from the Netherlands. Pieces of necrotic root tissue were excised, surface-disinfected, and transferred aseptically to a Phytophthora selective medium. P. ramorum was identified based on morphological characteristics, including the production of numerous, thin-walled chlamydospores (25 to 70 µm in diameter, average 43 µm) and deciduous, semi-papillate sporangia arranged in clusters. Radial growth after 6 days at 20°C on V8 juice agar was 2.8 mm per day. Random amplified microsatellite markers (RAMS) (2) from the total genomic DNA of the Belgian strain (CBS 110901) were similar to those of P. ramorum reference strains (CBS 101330, CBS 101332, and CBS 101554). Using PCR primers specific for P. ramorum, the identification was confirmed by W. A. Man in't Veld (Plantenziektenkundige Dienst, Wageningen, the Netherlands) (1). A pathogenicity test was carried out on three sterile cuttings of Rhododendron catawbiense (3). Brown lesions were observed on the inoculated cuttings after 6 to 7 days. None of the three uninoculated cuttings showed symptoms of infection. P. ramorum was reisolated from lesion margins on the inoculated cuttings. To our knowledge, this is the first report of the fungus from Belgium. Since our initial observation, we have found P. ramorum in other Belgian nurseries on R. yakusimanum. References: (1) M. Garbelotto et al. US For. Ser. Gen. Tech. Rep. PSW-GRT. 184:765, 2002. (2) J. Hantula et al. Mycol. Res. 101:565, 1997. (3) S. Werres et al. Mycol. Res. 105:1155, 2001.

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First Report of a New Postharvest Fruit Rot on Apple Caused by Sphaeropsis pyriputrescens

Plant Disease ◽

10.1094/pdis.2004.88.2.223a ◽

2004 ◽

Vol 88 (2) ◽

pp. 223-223 ◽

Cited By ~ 14

Author(s):

C. L. Xiao ◽

J. D. Rogers ◽

R. J. Boal

Keyword(s):

Leading Edge ◽

Preliminary Evidence ◽

Apple Fruit ◽

Causal Agent ◽

Fruit Rot ◽

Postharvest Disease ◽

Fluorescent Light ◽

Pathogenicity Test ◽

First Report ◽

Golden Delicious

During March to July 2003, a postharvest fruit rot was observed on ‘Golden Delicious’, ‘Granny Smith’, and ‘Red Delicious’ apples (Malus × domestica Borkh.) sampled from commercial packinghouses in Washington State. Losses as high as 24% in storage bins were observed in July on ‘Red Delicious’. The disease started at the stem bowl area or the calyx end of the fruit. Decayed fruit was apparently not wounded. Decayed areas were brown and firm. Internal decayed flesh appeared yellowish brown. On ‘Red Delicious’ apples, decayed fruit was apparently discolored from red to brown. As the disease advanced, pycnidia of a fungus might form on the stem, sepals, or the surface of decayed fruit. Pycnidia were 0.3 to 0.7 mm in diameter, black, and partially immersed in decayed tissues. To isolate the causal agent, decayed fruit was lightly sprayed with 70% ethanol and air dried. Fragments of diseased tissue were removed from the margin of diseased and healthy tissue and plated on acidified potato dextrose agar (PDA). A fungus was consistently isolated from decayed fruit with the symptoms described above. On PDA, the colonies of the fungus first appeared with dense hyaline mycelium and later turned light yellow to yellow. Black pycnidia of the fungus formed on 2- to 3-week-old oatmeal agar cultures at 20°C under 12-h alternating cycles of fluorescent light and dark. The fungus was identified as Sphaeropsis pyriputrescens Xiao & J. D. Rogers, based on the description of the fungus (1). Voucher specimens were deposited at the WSU Mycological Herbarium. Two isolates of the fungus recovered from decayed apples were tested for pathogenicity on apple. Fruit of ‘Golden Delicious’ and ‘Gala’ were surface-disinfested for 5 min in 0.5% NaOCl, rinsed, and air dried. Fruit was wounded with a sterile 4-mm-diameter nail head. A 4-mm-diameter plug from the leading edge of a 3-day-old PDA culture or plain PDA (control) was placed in the wound of each of 10 replicate fruit for each isolate or control. Fruit was tray packed with polyethylene liners and stored in cardboard boxes in air at 3°C, and decay was evaluated 2 weeks after inoculation. Five decayed fruits from each treatment were selected for reisolation of the causal agent. The experiment was conducted twice. In a separate pathogenicity test, two isolates (one each from apple and pear) were included in the test. Fruit of ‘Red Delicious’ apple was prepared and inoculated as the same manner described above, but fruit was stored in air at 0°C. The experiment was conducted twice. All fruit that were inoculated with the fungus developed decay symptoms. No decay developed on fruit in the controls. The same fungus was reisolated from decayed fruit. This indicates that isolates from apple and pear were pathogenic to apple. S. pyriputrescens is the causal agent of a newly reported postharvest disease on ‘d'Anjou’ pears (1). To our knowledge, this is the first report of this fungus causing postharvest fruit rot on apple. We propose ‘Sphaeropsis rot’ as the name of this new disease on apple and pear. Preliminary evidence suggests that infection of fruit by this fungus occurred in the orchard prior to storage. Reference: (1) C. L. Xiao and J. D. Rogers. Plant Dis. 88:114, 2004.

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First Report of Leaf Blight of Cyperus iria Caused by Fusarium equiseti in India

Plant Disease ◽

10.1094/pdis-07-12-0690-pdn ◽

2013 ◽

Vol 97 (6) ◽

pp. 838-838 ◽

Cited By ~ 1

Author(s):

V. Gupta ◽

V. K. Razdan ◽

D. John ◽

B. C. Sharma

Keyword(s):

Leaf Blight ◽

Spore Suspension ◽

Commonwealth Mycological Institute ◽

Fungal Culture ◽

Distilled Water ◽

Total Production ◽

Conidial Suspension ◽

Internal Transcribed Spacer Region ◽

Fusarium Equiseti ◽

First Report

In India, rice (Oryza sativa L.) plays a major role in national food security, with total production of 102.75 million t, harvested from 44 million ha during 2011 (1). Weeds are one of the major causes of losses in rice. Cyperus iria, locally known as chatriwala dela (rice flat sedge), is an annual weed in the Cyperaceae that can reach 50 to 60 cm tall. A leaf blight of C. iria was observed during August 2010 in a 20-ha rice field (cv. Basmati 370) at the University Research Farm, Chatha, Jammu (32° 43′ N, 74° 54′ E). Symptomatic plants were scattered randomly in the field and had water-soaked spots on the upper leaf surfaces initially, which turned brown after 4 days and developed a yellow halo, resulting in a blighted appearance. The diseased leaves shriveled and infected plants died. Infected C. iria leaf pieces with adjacent healthy tissue were collected, surface-sterilized in 0.1% mercuric chloride for 20 s, then rinsed three times in sterilized distilled water. The pieces were plated onto potato dextrose agar (PDA) and incubated at 27 ± 1°C for 4 days. A pure fungal culture was obtained by single-spore technique on 2% water agar and maintained on PDA at 10°C. The fungus initially produced white mycelium that became brown with age. Dark brown spots or flecks of pigment formed in the agar. Macroconidia were long and slender, with tapered apical cells that were elongated or even whip-like. Basal cells of macroconidia were prominent, foot shaped, and elongated. Macroconidia were 39.55 to 56.74 × 3.75 to 4.5 μm with 3 to 5 septa. Conidiophores were compact, penicillately branched, and arose from lateral branches which initially were one-celled and bore 2 to 4 phialides at the apex. Chlamydospores were intercalary, solitary, in chains or in knots, globose, and 7 to 9 μm in diameter. On the basis of morphological characteristics (2), the fungus was identified as Fusarium equiseti (Corda) Sacc. and deposited in the Indian Type Culture Collection, New Delhi (8424.11). The ITS (internal transcribed spacer) region of rDNA was amplified by PCR with primers ITS1/ITS2 and sequenced. BLASTn analysis of the sequence showed 100% homology with the ITS sequence of F. equiseti in the NCBI database (JN596252.1), and the sequence was deposited in GenBank (KC434458). To confirm pathogenicity of the F. equiseti isolate, 10 seeds of C. iria were planted in five clay pots (each 38 cm in diameter) filled with sterilized soil. Three seedlings were used for the experiment and the remaining seedlings removed from each pot. A total of 15 seedlings (5 pots × 3 seedlings per pot) at the two-leaf stage were spray-inoculated with a 50-ml conidial suspension of the isolate (105 cfu/ml) using a hand atomizer. The control treatment included three seedlings treated similarly with sterile distilled water. The spore suspension was prepared in potato dextrose broth using a culture of the fungus incubated for 10 days and then homogenized at 140 rpm. Tween 20 (1%) was added to the spore suspension. Small spots developed 4 days after inoculation, and the lesions then coalesced into large necrotic areas, resulting in leaf blight 10 days after inoculation. F. equiseti was reisolated from inoculated leaves using the method described above, whereas no fungus was reisolated from control plants, fulfilling Koch's postulates. The isolated fungus displayed the same morphological and cultural features as the original isolate. F. equiseti has been reported to infect Echinochloa spp. in Iran (3), but to our knowledge, this is the first report of F. equiseti infecting C. iria in India. Thus, F. equiseti represents a potential biocontrol agent for managing C. iria in rice fields. References: (1) Anonymous. Direct. Rice Res. Newslett. 10:2, 2012. (2) C. Booth. The Genus Fusarium. Commonwealth Mycological Institute, Kew, Surrey, England, p. 157, 1971. (3) M. R. S. Motlagh. Austral. J. Crop Sci. 4:457, 2010.

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First report of Penicillium olsonii causing postharvest fruit rot on tomato in Serbia

Plant Disease ◽

10.1094/pdis-02-21-0323-pdn ◽

2021 ◽

Author(s):

Svetlana Živković ◽

Danijela Ristić ◽

Stefan Stošić

Keyword(s):

Fruit Rot ◽

Morphological Identification ◽

Malt Extract ◽

Pathogenicity Test ◽

Distilled Water ◽

Total Production ◽

Academic Press ◽

First Report ◽

Initial Symptoms ◽

The Republic

Tomato (Solanum lycopersicum, L.) is one of the most important vegetable crop in Serbia, with a total production of 111,639 t in 2019 (Statistical Office of the Republic of Serbia). In July 2020, six tomatoes (cv. Balkan) with symptoms of fruit rot were collected from market in Belgrade, Serbia. The incidence of disease was about 2%, and the symptomatic samples were stored for 10 days after harvest. The initial symptoms on fruits were small circular, slightly sunken and water-soaked spots with white mycelia, that progressively expanded into larger grey lesions following the occurrence of sporulation. Isolations were conducted from one spot/fruit. Small pieces (2 to 3 mm2) from the margins of lesions were surface sterilized for 1 min in 1% NaOCl, washed twice with sterile distilled water, and cultivated on potato dextrose agar (PDA) at 25°C. The isolation frequency of Penicillium-like colonies was 100%. In total, six monosporic isolates were obtained and two isolates (SZ-20-6 and SZ-20-7) were selected as representative for morphological and molecular identification, and pathogenicity test. Morphological characteristics of both isolates were observed after growth on malt extract agar (MEA) for 7 days at 25ºC. On MEA, mycelia were white and colonies turned greyish-green with abundant sporulation. On the reverse sides colonies were pale yellow. The mean colony diameter on MEA for isolate SZ-20-6 was 25 ± 1.2 mm and 26 ± 1.0 mm for isolate SZ-20-7. The colony texture was velvety, without exudates and pigmentation. The conidiophores of both isolates were terverticillate, unbranched; phialides were flask shaped with a short neck, and conidia were smooth, greenish and subglobose to ellipsoidal. The conidial diameter for isolate SZ-20-6 was 3 to 4 × 2.5 to 3 µm, and for isolate SZ-20-7 was 3.5 to 4 × 2.5 to 3.5 µm (n =50). Based on these characteristics, isolates were identified as Penicillium olsonii (Pitt 1979). To confirm the morphological identification, genomic DNA was extracted from isolates (SZ-20-6 and SZ-20-7), and the rDNA ITS region and partial β-tubulin gene (BenA) were amplified using the primers ITS1/ITS4 (White et al. 1990) and Bt2a/Bt2b (Glass and Donaldson 1995), respectively. All sequences showed 99 to 100% similarity to P. olsonii and were deposited in GenBank (ITS, MW130235 and MW130236; BenA, MW145147 and MW145148). In multilocus phylogenetic analysis (ITS+BenA), isolates from this study clustered together with other P. olsonii sequences with 100% bootstrap support. To complete Koch's postulates, asymptomatic fruits of tomato cv. Balkan (five fruits per isolate) were superficially sterilized with 70% ethanol, wounded with a sterile needle and inoculated with 10 μl of a spore suspension (1 × 106 spores/ml). Five control fruits were inoculated with 10 μl of sterile distilled water. The experiment was repeated twice. After 7 days of incubation in a moisture chamber at 25°C, typical grey lesions developed on inoculated fruits. The control fruits remained symptomless. The isolates recovered from symptomatic fruits showed the same morphological features as the original isolates. P. olsonii was previously reported on tomato fruit only in Canada (Chatterton et al. 2012) and Pakistan (Anjum et al. 2018). To our knowledge, this is the first report of P. olsonii causing postharvest fruit rot on tomato in Serbia, and in Europe, as well. Therefore, it is essential to monitor spreading of P. olsonii on tomato and other crops in storages, and develop efficient disease management strategies. References: Anjum, N. et al. 2018. Plant Dis. 102:451. Chatterton, S., et al. 2012. Can. J. Plant Pathol. 34:524. Glass, N. L. and Donaldson, G. C. 1995. Appl. Environ. Microbiol. 61:1323. Pitt, J. I. 1979. The Genus Penicillium and its Teleomorophic States Eupenicillium and Talaromyces. Academic Press, London, U.K. Statistical Office of the Republic of Serbia. https://www.stat.gov.rs/en-US/ White, T. J., et al. 1990. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA. Funding: This research was financed by the Ministry of Education, Science and Technical Development of the Republic of Serbia, grant 451-03-68/2020-14/200010.

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

Plant Disease ◽

10.1094/pdis-07-13-0740-pdn ◽

2014 ◽

Vol 98 (3) ◽

pp. 424-424 ◽

Cited By ~ 7

Author(s):

Y. K. Kim

Keyword(s):

Aureobasidium Pullulans ◽

Deionized Water ◽

Causal Agent ◽

Fruit Rot ◽

Conidial Suspension ◽

Centraalbureau Voor ◽

First Report ◽

Control Fruit ◽

Initial Symptoms ◽

Sweet Cherries

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 Powdery Mildew of Pumpkin Caused by Golovinomyces cichoracearum in Neelum Valley, Azad Kashmir

Plant Disease ◽

10.1094/pdis-08-11-0642-pdn ◽

2012 ◽

Vol 96 (6) ◽

pp. 906-906

Author(s):

I. Mukhtar ◽

R. Khurram ◽

A. Hannan ◽

Z. Hayat

Keyword(s):

Powdery Mildew ◽

Infected Plant ◽

Causal Agent ◽

Morphological Observation ◽

Fungal Culture ◽

Voucher Specimen ◽

First Report ◽

Golovinomyces Cichoracearum ◽

Azad Kashmir ◽

Basal Septum

During July 2011, a severe powdery mildew outbreak was recorded in pumpkin (Cucurbita moschata Duch.) fields in economically poor areas in the Neelum Valley (Leswa, Mir Pura, Jura, Kundal Shahi, and Bela). Disease symptoms included grayish white circular to irregular patches consisting of epiphytic mycelia and conidia on both surfaces of the leaves of infected vines. Fungal mycelia and conidiophores were also visible in white powdery patches on the main stems, leaves, and petioles of affected plants. Leaves became necrotic with age. Powdery mildew symptoms were more severe on pumpkin vines located in the shade, and severely affected vines collapsed. A voucher specimen (IR00027) has been deposited in First Fungal Culture Bank of the Pakistan (FCBP), Institute of Agriculture Sciences, University of the Punjab, Lahore, Pakistan. For morphological observation, a clear tape strip was used to remove fungal tissue from infected leaves. The tape was mounted on microscope slides with water and examined with a light microscope (2). Conidiophores were unbranched, cylindrical, erect and arose singly on hyphal cells, 110 to 200 × 6 to 12 μm, composed of a foot cell and three to eight barrel-shaped conidia formed in chains with a sinuate edge, followed with a basal septum at the branching point or slightly displaced from the mycelium. Foot cells were 50 to 80 × 6 to 12 μm, and slightly swollen with constriction at the base. Conidia were cylindrical to doliiform, 30 to 50 × 14 to 20 μm and produced in chains. The length/breadth ratio of the conidia was 1.8 to 2.6. Fibrosin bodies were absent in both conidiophores and conidia. No cleistothecia were observed. Identification of the causal agent as Golovinomyces cichoracearum (DC.) V.P. Heluta (synonym Erysiphe cichoracearum) was based on morphology (1). Pathogenicity was determined using field-infected plant leaves transported to a greenhouse. Fresh conidia of field isolates of G. cichoracearum were transferred by a sterile brush from the affected leaves to fully expanded four to five healthy leaves of three 40-day-old vines. For the control, three vines were stroked with a clean sterile brush and control plants were placed 100 m away from the inoculated plants. For disease development 20°C and 80% relative humidity was maintained in the greenhouse. A plastic sheet was placed around each vine for a week and removed later. Inoculated vines developed visible white spots of powdery mildew on the leaves after 10 days in the greenhouse, whereas control plants remained asymptomatic. Fungal colony and conidial morphology on the leaves of inoculated plants were as described above. Previously, G. cichoracearum has been reported on other cucurbits (3) in the plains of Pakistan. To our knowledge, this is the first report of occurrence of G. cichoracearum as a causal agent of powdery mildew on pumpkin in Neelum Valley, Azad Kashmir. References: (1) U. Braun. Beih. Nova Hedwigia 89:1, 1987. (2) J. C. Correll et al. Plant Dis. 71:248, 1987. (3) A. Wahid et al. Pak. J. Agric. Res. 9:209, 1988.

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First Report of Colletotrichum gloeosporioides Causing Anthracnose Fruit Rot of Trichosanthes kirilowii in China

Plant Disease ◽

10.1094/pdis-91-5-0636c ◽

2007 ◽

Vol 91 (5) ◽

pp. 636-636 ◽

Cited By ~ 1

Author(s):

H. Y. Li ◽

Z. F. Zhang

Keyword(s):

Relative Humidity ◽

Colletotrichum Gloeosporioides ◽

Aerial Mycelium ◽

Causal Agent ◽

High Relative Humidity ◽

Fruit Rot ◽

Single Spore ◽

Specific Primers ◽

First Report ◽

Trichosanthes Kirilowii

Trichosanthes kirilowii Maxim., a species within the gourd family, is cultivated in China for its edible seeds and medicinal roots. Since 2000, heavy losses due to fruit rot have been caused by a new disease with typical anthracnose symptoms, i.e., water-soaked, dark brown-to-black, sunken lesions. Signs of the suspected pathogen were usually seen on near-mature fruits and were especially evident after abundant rainfall. The lesions contained numerous black acervuli with black setae that produced abundant, salmon-colored spore masses under high relative humidity. Dark lesions on leaves and stems could also be found in the field that sometimes led to stem girdling and wilting. Conidia produced in acervuli were 14 to 20 × 3 to 6 μm, straight, cylindrical, hyaline, aseptate, with both ends rounded. Conidiophores were 13 to 22 × 4 to 6 μm, aseptate, and cylindrical, while the setae, usually with three to five septa, measured 60 to 86 × 5 to 6 μm. The pathogen was initially identified as Colletotrichum gloeosporioides on the basis of the morphology (2). In culture, the fungus produced a gray-to-black colony with whitish aerial mycelium on potato dextrose agar (PDA) medium. Pathogenicity was tested by inoculating the equator of 10 fruits of T. kirilowii with a 5-day-old mycelia plug from a single-spore colony (0.5 cm in diameter); fruits inoculated with the plugs of PDA medium served as the control. Inoculated fruits were covered with plastic for 24 h to maintain high relative humidity. After 4 days, 100% of the inoculated fruits showed symptoms identical to those observed on T. kirilowii fruit affected in the field, while all fruits inoculated with PDA medium remained free of symptoms. Reisolation of the fungus from fruit lesions confirmed that the causal agent was C. gloeosporioides. To confirm the pathogen to species, the C. gloeosporioides-specific primers CgInt/ITS4 and C. acutatum-specific primers CaInt2/ITS4 (1) were used to amplify the sequence of internal transcribed spacer regions. A fragment of approximately 500 bp was only amplified with primers CgInt/ITS4 and the sequence (GenBank Accession No. AM491334) was 98 to 100% matched to the sequences of several C. gloeosporioides isolates (e.g., GenBank Accession Nos. AJ301919, AB255249, AJ301908), whereas the sequence shares 86 and 91% identity to that of C. orbiculare (GenBank Accession No. AB042308) and C. acutatum (GenBank Accession No. AJ749675), respectively. Thus, we concluded that C. gloeosporioides is the causal agent of anthracnose fruit rot of T. kirilowii. To our knowledge, this is the first report of C. gloeosporioides infecting T. kirilowii. References: (1) A. E. Brown et al. Phytopathology 86:523, 1996. (2) B. C. Sutton. The Coelomycetes. CAB International Publishing, New York, 1980.

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First Report of Fusarium lichenicola as a Causal Agent of Fruit Rot in Pomelo (Citrus maxima)

Plant Disease ◽

10.1094/pdis-10-14-1017-pdn ◽

2015 ◽

Vol 99 (9) ◽

pp. 1278 ◽

Cited By ~ 2

Author(s):

D. B. Amby ◽

T. T. T. Thuy ◽

B. D. Ho ◽

C. Kosawang ◽

T. B. Son ◽

...

Keyword(s):

Causal Agent ◽

Fruit Rot ◽

First Report ◽

Citrus Maxima

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First Report of Diplodia seriata as Causal Agent of Olive Dieback in Croatia

Plant Disease ◽

10.1094/pdis-08-11-0628 ◽

2012 ◽

Vol 96 (2) ◽

pp. 290-290 ◽

Cited By ~ 12

Author(s):

J. Kaliterna ◽

T. Milicevic ◽

D. Ivic ◽

D. Bencic ◽

A. Mesic

Keyword(s):

Elongation Factor ◽

Its Region ◽

Morphological Characters ◽

Molecular Data ◽

Causal Agent ◽

Fruit Rot ◽

Morphological Identification ◽

Foeniculum Vulgare ◽

Diplodia Seriata ◽

First Report

In August 2010, a dieback of young olive (Olea europea L.) trees (cvs. Pendolino and Leccino) occurred in two orchards in Istria, Croatia. According to the producers, low temperatures during the winter severely damaged the plants and led to their decline. Distinctive symptoms, assumed fungal infection, were observed in internal tissue of stems and branches. Elongated brown necrosis, sometimes with black streaks, was visible under the bark, therefore Verticillium wilt was suspected. Of 1,086 trees in two orchards (4 ha), 165 (15%) showed symptoms. To isolate the causal agent, surface-sterilized wood chips of symptomatic tissue were placed on potato dextrose agar (PDA). Fungal colonies resembling Botryosphaeriaceae spp. grew from all wood fragments placed on PDA, and from these colonies, monohyphal isolates were obtained. For morphological identification, pycnidial formation was stimulated by growing the isolates on 2% water agar that included stems of plant species Foeniculum vulgare Mill. at room temperature under diffuse light. Pycnidia contained conidia that initially showed as hyaline, becoming light to dark brown as they matured, ovoid with truncated or rounded base and obtuse apex, aseptate, with wall moderately thick, externally smooth, roughened on the inner surface, and 22.8 to 23.5 × 9.6 to 10.5 μm. On the basis of these morphological characters, fungal species Diplodia seriata (teleomorph “Botryosphaeria” obtusa) was suspected (3). For molecular identification, four isolates (MN3, MN4, MN5, and MN6) were used for PCR to amplify the internal transcribed spacer (ITS) region and partial translation elongation factor 1-alpha (EF1-α) gene, using primers ITS4/ITS5 and EF1-728F/EF1-986R, respectively. Sequencing was performed with those amplified genes, then sequences were deposited in GenBank. Comparison of these sequences with GenBank sequences for referent D. seriata isolate CBS 112555 (AY259094 and AY573220) (3) showed 100% homology. On the basis of molecular data, the isolates were confirmed to be species D. seriata De Not. Pathogenicity tests were performed by inoculation of 2-year-old olive plants, six plants per tested cultivar (Pendolino and Leccino). For every cultivar, four plants were wounded and mycelium plugs from D. seriata cultures on PDA were placed on the wounds and sealed with Parafilm. Two control plants per tested cultivar were inoculated with sterile PDA plugs. After 2 months, six of eight inoculated plants wilted completely, and under the bark, brown necrosis was observed. D. seriata was constantly reisolated from the inoculated plants and fulfilled Koch's postulates and confirmed pathogenicity of D. seriata on olive as causal agent of olive dieback. Control plants showed no symptoms of the disease. This fungus has been recognized as the cause of fruit rot of olive (1) and branch canker or dieback in Spain (2). To our knowledge, this is the first report of D. seriata as a pathogen of olive in Croatia. Also, this is one of the first reports of D. seriata as the cause of olive dieback in the world, while Moral et al. (1,2) mostly reported it as the cause of olive fruit rot. Since the same symptoms of olive dieback were observed at other localities in Croatia, the disease could represent a serious threat, particularly for young olive orchards. References: (1) J. Moral et al. Plant Dis. 92:311, 2008. (2) J. Moral et al. Phytopathology 100:1340, 2010. (3) A. J. L. Phillips et al. Fungal Divers. 25:141, 2007.

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First report of bacterial shot-hole disease caused by Xanthomonas arboricola pv. pruni on plumcot in South Korea

Plant Disease ◽

10.1094/pdis-06-20-1227-pdn ◽

2020 ◽

Author(s):

Walftor Dumin ◽

Mi-Jeong Park ◽

Jong-Han Park ◽

Chang Youl Yang ◽

Chang-Gi Back

Keyword(s):

16S Rdna ◽

Causal Agent ◽

Bacterial Suspension ◽

Pathogenicity Test ◽

Total Production ◽

Sterile Water ◽

First Report ◽

Young Leaves ◽

Leaf Tissues ◽

Xanthomonas Arboricola

Plumcot is an interspecific hybrid product between Japanese plums (Prunus salicina) and apricots (Prunus armeniaca) obtained by the NIHHS, Korea in 1999 [1]. At the early of 2017, black spots-like symptoms were observed on plumcot fruits and leaves at cultivation areas in Naju (34.965595, 126.665853) province. Further investigation shows that approximately 60% of the plumcot leaves in the affected orchard were infected, which caused 40% total production loss. At the early stage of infection, disease symptoms appear as small, angular and water-soaked spots and develop into circular brown spots at the later stages of infection. As the disease progresses, the leaf tissues around the spots became yellow and the lesions enlarged. When the adjacent lesions merged and the necrotic tissues fall off, shot-hole symptoms appear on the leaves. To identify the causal agent of this disease, infected leaf tissues were excised and surface-sterilized with 1% NaOCl for 30 secs prior to rinsing with sterile water, thrice . Tissue samples were then placed in sterile water (0.5 mL) for 5 min before its aliquots were streaked onto Luria-Bertani (LB) agar. Plates then were incubated at 28°C. To obtain pure colonies, bacteria were re-streak into a new LB agar and colonies showing typical Xanthomonas spp. morphology (i.e. convex, smooth, yellow, and mucoid) were subjected to Gram staining assay. For molecular identification, 16S ribosomal DNA (16S-rDNA) and gyrase B (gyrB) genes were amplified using a 9F/1512r and UP-1/UP-2Sr primers [2,3] respectively from 5 gram-negative isolates. PCR products were sequenced and analysed using BLASTN. Result shows that 16S-rDNA and gyrB genes are 99-100% identical to a similar genomic region of Xanthomonas arboricola pv. pruni (Xap) isolated in almond (MK156163), peach (MG049922) and apricot (KX950802) respectively [4,5,6]. 16S-rDNA and gyrB gene sequences were deposited in the GenBank (LC485472 and LC576824), whereas pathogen isolate was deposited into Korean Agricultural Culture Collection (KACC19949). Pathogenicity test was performed using Xap bacterial suspension (108 cfu/mL) inoculated on the abaxial and adaxial surface of plumcot detached leaves. For inoculation, 10 healthy young leaves were used whereas, 5 young leaves mock-inoculated with sterile LB broth were used as a control. Both leaf samples were kept in a closed container to maintain 100% humidity before being incubated at 25°C. The water-soaked symptoms were observed visually on the inoculated leaves 2 to 3 days post-inoculation. No water-soaked symptoms were observed on the control leaves. Morphology and sequences of molecular markers used showed that the 3 bacterial colonies re-isolated from the inoculated leaves were identical to the original isolate, fulfilling Koch’s postulate. Pathogenicity tests were repeated twice and the results obtained were consistent with the first experiment. As a new variety of stone fruit cultivated in Korea, information about pathogens and registered agrochemicals to control disease outbreak in plumcot are still limited. Therefore, the identification of Xap as a causal agent to the black spot disease is critical for the development of disease management strategies and to identify appropriate agrochemicals to control the occurrence of this disease in the field. To our knowledge, this is the first report of Xap as a causal agent to the shot-hole disease on the plumcot in Korea.

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