scholarly journals Occurrence of a Fruit Spot Disease of Pear Caused by Septoria pyricola in Tyrnavos Larissa, Northern Greece

Plant Disease ◽  
2014 ◽  
Vol 98 (6) ◽  
pp. 845-845
Author(s):  
T. Thomidis ◽  
S. Katerinis
Keyword(s):  
South Africa ◽  
Leaf Spot ◽  
Morphological Characteristics ◽  
Northern Greece ◽  
Koch’S Postulates ◽  
Randomized Design ◽  
Fruit Spot ◽  
Koch's Postulates ◽  
Wounded Area ◽  
Septoria Leaf Spot

Pear tree (Pyrus communis) is an important crop in Greece. In 2012, fruits of the pear cv. Kontoula were observed in commercial fields located in Tirnavos, Prefecture of Larrisa, Greece, with symptoms of well-defined brown angular margins and their grayish white centers in which a few pycnidia (about 180 × 150 μm) were produced within the spots. Pycnidia were dark, separate, and globe-shaped with an ostiole (opening at the apex) from which conidia (about 40 to 60 × 3 μm) were extruded, and erupted through the surface of the infected tissue. Conidia were produced on short conidiophores. They were clear, narrowly elongated to filiform, and several-celled. The pathogen was isolated on acidified-PDA (2.5 ml 85% lactic acid per liter of nutrient medium) and incubated at 23°C for 7 days. The pathogen was identified as Septoria pyricola Desm. based on morphological characteristics. Koch's postulates were completed in the laboratory by placing a 40-μl drop of suspension (4 × 105 conidia ml−1 of water) on a wounded area of healthy fruits of cv. Kontoula. Fruits were surface sterilized with dipping in 0.1% chlorine solution, allowed to dry in a laminar flow hood. There were 15 inoculated and 15 control fruits (similarly inoculated with sterile distilled water) in a randomized design. Fruits were covered with perforated polythene bags to maintain a high humidity necessary for infection and these bags were removed 48 h after inoculation and maintained at room temperature (23 ± 2°C). Lesion development was recorded daily for each fruit. Koch's postulates were satisfied after re-isolating the fungus from inoculated fruit that developed symptoms similar to those observed on fruits collected from fields. Symptoms of this disease were found in all pear orchards cultivating the cv. Kontoula located in Tyrnavos (a municipality in the Prefecture of Larissa). Symptoms of septoria leaf spot were also observed in the above pear orchards. In contrast, no symptom of septoria fruit spot and septoria leaf spot was observed in apple orchards of the above regions. To our knowledge, this is the first report of the occurrence of S. pyricola as causal agent of fruit spot of pears in Greece. Fruit spotting is relatively uncommon; nevertheless, Sivanesan (3) gives two reports of conidia infecting pear fruits from Italy and South Africa (1,2). References: (1) G. Florenzano. Int. Bull. Plant Prot. 20:17, 1946. (2) A. J. Louw. Farming in South Africa 23:737, 1948. (3) A. Sivanesan. IMI Descriptions of Fungi and Bacteria, vol. 99, sheet 989. CABI, Wallingford, UK, 1990.

scholarly journals Occurrence of a Fruit Spot Disease of Pomegranates Caused by Colletotrichum gloeosporiodes in the Prefecture of Komotini, Greece

Plant Disease ◽  
2011 ◽  
Vol 95 (7) ◽  
pp. 872-872 ◽  
Author(s):  
T. Thomidis ◽  
E. Exadaktylou
Keyword(s):  
Room Temperature ◽  
Morphological Characteristics ◽  
Punica Granatum ◽  
Lesion Development ◽  
Koch’S Postulates ◽  
Randomized Design ◽  
Fruit Spot ◽  
Koch's Postulates ◽  
Punica Granatum L ◽  
Wounded Area

Pomegranate (Punica granatum L.) is an important crop in Greece. In 2010, fruits of the pomegranate cv. Wonderful, in commercial fields located in the Prefecture of Komotini in eastern Greece, were observed to have symptoms of distinct dark brown spots. The waxy acervuli observed in infected tissue were subepidermal, typically with setae and simple, short, erect conidiophores. Conidia were hyaline, one celled, ovoid to oblong, slightly curved or dumbbell shaped, and 10 to 15 μm long and 5 to 7 μm wide. The pathogen was isolated on acidified potato dextrose agar (PDA) (2.5 ml of 85% lactic acid per liter of nutrient medium) and incubated at 23°C for 7 days. The pathogen was identified as Colletotrichum gloeosporiodes (Penz) Sacc. on the basis of morphological characteristics and internal transcribed spacer (ITS) sequence (100% identity to AJ301912, C. gloeosporioides species complex) (3) by CBS Fungal Biodiversity Centre, Identification Service, Utrecht, the Netherlands, (CG-1 isolate deposited in CBS Collection; Accession No. CBS 129372). Koch's postulates were completed in the laboratory by placing a 40-μl drop of suspension (4 × 105 conidia ml–1 of water) on a wounded area of healthy fruits of cv. Wonderful. Fruits were surface sterilized by dipping in 0.1% chlorine solution and allowed to dry in a laminar flow hood. There were 15 inoculated and 15 control fruits (similarly sprayed with sterile distilled water) in a randomized design. Fruits were covered with perforated polythene bags to maintain a high humidity necessary for infection that were removed 48 h after inoculation and the fruits were maintained at room temperature (23 ± 2°C). Lesion development was recorded daily for each fruit. Koch's postulates were satisfied after reisolating the fungus from inoculated fruit that developed symptoms similar to those observed on fruits collected from fields. To our knowledge, this is the first report of the occurrence of C. gloeosporioides as a causal agent of fruit spot of pomegranates in Greece. Fruit spots caused by C. gloeosporioides have been reported in pomegranate fields of other countries around the world (1,2). References: (1) B. K. M. Lakshmi et al. Trop. Agric. Res. 22:183, 2011. (2) D. S. Patel. Indian Phytopathol. 62:252, 2009. (3) B. S. Weir and P. R. Johnston. Mycotaxon 111:209, 2010.

scholarly journals First Report of Stemphylium botryosum Causing Leaf Blight of Kiwi in the Province Imathia, Northern Greece

Plant Disease ◽  
2008 ◽  
Vol 92 (4) ◽  
pp. 650-650 ◽  
Author(s):  
T. Thomidis ◽  
T. J. Michailides
Keyword(s):  
Apical Cell ◽  
Morphological Characteristics ◽  
Leaf Blight ◽  
Width Ratio ◽  
Northern Greece ◽  
Cell Width ◽  
Koch’S Postulates ◽  
First Report ◽  
Stemphylium Botryosum ◽  
Koch's Postulates

In Greece, kiwi (Actinidia deliciosa) is mostly found in the northern part of the country where approximately 440,000 ha are grown. In the summer of 2006, a Stemphylium sp. was frequently isolated from leaves of kiwi (cv. Hayward) grown in the province of Imathia. Symptomatic leaves were covered with irregular, necrotic, brown areas. Lesions had a distinct margin that, in some cases, covered a wide part of the diseased leaves. Intense symptoms were frequently observed and associated with defoliation. This Stemphylium sp. was consistently isolated from diseased leaves onto potato dextrose agar (PDA) after surface sterilization with 0.1% chlorine solution. On the basis of morphological characteristics of mycelia, dimensions (length 20 to 29 μm and width 14 to 21 μm) and mean length/width ratio (1.42 μm) of conidia, and width and apical cell width of condiophores, the fungus was identified as Stemphylium botryosum (Wallr.) (2,3) Koch's postulates were completed in the laboratory by inoculating leaves of kiwi (cv. Hayward) with an isolate of S. botryosum originated from a symptomatic leaf of a Hayward kiwi. Twenty leaves were surface sterilized by dipping them into 0.1% chlorine solution for 2 to 3 min, washing in sterile distilled water, and allowing them to dry in a laminar flow hood. A leaf was then placed into a petri plate containing a wet, sterilized paper towel. Inoculation was made by transferring a 5-mm-diameter mycelial disc from the margins of a 7-day-old culture onto the center of each leaf surface. Petri plates were closed and incubated at 25°C with 12 h of light for 6 days. Koch's postulates were satisfied when the same S. botryosum was reisolated from 100% of inoculated leaves that developed symptoms similar to those observed in the vineyards. Leaves inoculated with a PDA plug alone (with no S. botryosum) did not develop any symptoms. Previously, Alternaria alternata was reported as the causal agent of a leaf spot pathogen of kiwi (1,4). To our knowledge, this is the first report of the occurrence of S. botryosum causing leaf blight of kiwi in Greece and worldwide. This pathogen can cause a high level of defoliation in diseased plants. References: (1) L. Corazza et al. Plant Dis. 83:487, 1999. (2) M. B. Ellis. Dematiaceous Hyphomycetes. Mycology Institute. London, England, 1971. (3) E. G. Simmons. Mycologia 61:1, 1969. (4) C. Tsahouridou and C. C. Thanassoulopoulos. Plant Dis. 84:371, 2000

scholarly journals First Report of Leaf Spot Disease on Walnut Caused by Colletotrichum fioriniae in China

Plant Disease ◽  
2015 ◽  
Vol 99 (2) ◽  
pp. 289-289 ◽  
Author(s):  
Y. Z. Zhu ◽  
W. J. Liao ◽  
D. X. Zou ◽  
Y. J. Wu ◽  
Y. Zhou
Keyword(s):  
Dna Sequences ◽  
Leaf Spot ◽  
Juglans Regia ◽  
Morphological Characteristics ◽  
Leaf Spot Disease ◽  
Koch’S Postulates ◽  
First Report ◽  
Spot Disease ◽  
Leaf Spots ◽  
Koch's Postulates

In May 2014, a severe leaf spot disease was observed on walnut tree (Juglans regia L.) in Hechi, Guangxi, China. Leaf spots were circular to semicircular in shape, water-soaked, later becoming grayish white in the center with a dark brown margin and bordered by a tan halo. Necrotic lesions were approximately 3 to 4 mm in diameter. Diseased leaves were collected from 10 trees in each of five commercial orchards. The diseased leaves were cut into 5 × 5 mm slices, dipped in 75% ethanol for 30 s, washed three times in sterilized water, sterilized with 0.1% (w/v) HgCl2 for 3 min, and then rinsed five times with sterile distilled water. These slices were placed on potato dextrose agar (PDA), followed by incubating at 28°C for about 3 to 4 days. Fungal isolates were obtained from these diseased tissues, transferred onto PDA plates, and incubated at 28°C. These isolates produced gray aerial mycelium and then became pinkish gray with age. Moreover, the reverse of the colony was pink. The growth rate was 8.21 to 8.41 mm per day (average = 8.29 ± 0.11, n = 3) at 28°C. The colonies produced pale orange conidial masses and were fusiform with acute ends, hyaline, sometimes guttulate, 4.02 to 5.25 × 13.71 to 15.72 μm (average = 4.56 ± 0.31 × 14.87 ± 1.14 μm, n = 25). The morphological characteristics and measurements of this fungal isolate matched the previous descriptions of Colletotrichum fioriniae (Marcelino & Gouli) R.G. Shivas & Y.P. Tan (2). Meanwhile, these characterizations were further confirmed by analysis of the partial sequence of five genes: the internal transcribed spacer (ITS) of the ribosomal DNA, beta-tubulin (β-tub) gene, glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene, chitin synthase 3(CHS-1) gene, and actin (ACT) gene, with universal primers ITS4/ITS5, T1/βt2b, GDF1/GDR1, CHS1-79F/CHS1-354R, and ACT-512F/ACT-783R, respectively (1). BLAST of these DNA sequences using the nucleotide database of GenBank showed a high identify (ITS, 99%; β-tub, 99%; GAPDH, 99%; CHS-1, 99%; and ACT, 100%) with the previously deposited sequences of C. fioriniae (ITS, KF278459.1, NR111747.1; β-tub, AB744079.1, AB690809.1; GAPDH, KF944355.1, KF944354.1; CHS-1, JQ948987.1, JQ949005.1; and ACT, JQ949625.1, JQ949626.1). Koch's postulates were fulfilled by inoculating six healthy 1-year-old walnut trees in July 2014 with maximum and minimum temperatures of 33 and 26°C. The 6-mm mycelial plug, which was cut from the margin of a 5-day-old colony of the fungus on PDA, was placed onto each pin-wounded leaf, ensuring good contact between the mycelium and the wound. Non-colonized PDA plugs were placed onto pin-wounds as negative controls. Following inoculation, both inoculated and control plants were covered with plastic bags. Leaf spots, similar to those on naturally infected plants, were observed on the leaves inoculated with C. fioriniae within 5 days. No symptoms were observed on the negative control leaves. Finally, C. fioriniae was re-isolated from symptomatic leaves; in contrast, no fungus was isolated from the control, which confirmed Koch's postulates. To our knowledge, this is the first report of leaf disease on walnut caused by C. fioriniae. References: (1) L. Cai et al. Fungal Divers. 39:183, 2009. (2) R. G. Shivas and Y. P. Tan. Fungal Divers. 39:111, 2009.

scholarly journals First Report of Aspergillus niger Causing Postharvest Fruit Rot of Cherry in the Prefectures of Imathia and Pella, Northern Greece

Plant Disease ◽  
2012 ◽  
Vol 96 (3) ◽  
pp. 458-458 ◽  
Author(s):  
T. Thomidis ◽  
E. Exadaktylou
Keyword(s):  
Aspergillus Niger ◽  
Fungal Spores ◽  
Morphological Characteristics ◽  
Fruit Rot ◽  
Northern Greece ◽  
Centraalbureau Voor ◽  
Koch’S Postulates ◽  
First Report ◽  
Postharvest Rot ◽  
Koch's Postulates

In June 2011, symptoms of postharvest rot were observed on approximately 3% of all cherries collected from commercial orchards of cultivars Lapen and Ferrovia in the prefectures of Imathia and Pella (northern Greece). Fruit were harvested in a timely manner to avoid overripeness. No wounds or other predisposing injuries were observed on the infected fruits. Lesions enlarged rapidly and separated easily from healthy tissue when pressure was applied. Infected tissues were pale and water soaked and the associated fungal spores were dark and powdery and easily liberated when mature. The fungus grew rapidly and produced black colonies on acidified potato dextrose agar (2.5 ml of 85% lactic acid per liter of nutrient medium) after 5 days at 24°C. Identification of the pathogen was based on morphological characteristics (1). The conidial head was radiate, vesicles were nearly spherical and covered with metulae and phialides (biseriate). Conidia were globose (3 to 5 μm in diameter) and usually very rough with irregular ridges, bars, and verrucae. Koch's postulates were completed in the laboratory by inoculating mature cherry fruits (cv. Lapen). The fruits were surface sterilized by dipping in 10% chloride bleach solution, allowed to dry in a laminar flow hood, and wounded with a sharp glass rod that was 2 mm in diameter. A 40-μl drop of a suspension containing 20,000 conidia per ml of water was placed on each wound. There were 20 inoculated and 20 control fruits (similarly wounded and inoculated with a 40-μl drop of sterile distilled water) in a randomized design and incubated at 24 to 26°C for 6 days. Koch's postulates were satisfied after reisolating the fungus from inoculated fruit that developed symptoms similar to those observed on fruit collected from orchards. Control fruits did not show any symptom of the disease. To our knowledge, this is the first report of the occurrence of Aspergillus niger as the causal agent of postharvest rots of cherries in Greece. Postharvest fruit rots caused by A. niger have been reported in cherry orchards of other countries around the world (2). Because this disease causes postharvest rots of cherry fruits, measures may need to be implemented to manage the pathogen. References: (1) M. A. Klich. Page 12 in: Identification of Common Aspergillus Species. Centraalbureau Voor Schimmelcultures, Utrecht, the Netherlands, 2002. (2) A. Valiuskaite et al. Phytopathol. Pol. 35:197, 2005.

scholarly journals First Report of Leaf Spot Caused by Didymella americana on Cassia nomame in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Weiming Sun ◽  
Lina Feng ◽  
Xiaolei Wen ◽  
Bojia Han ◽  
Danrun Xing ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Leaf Spot ◽  
Disease Incidence ◽  
Morphological Characteristics ◽  
Rrna Gene ◽  
Pathogenicity Test ◽  
Koch’S Postulates ◽  
First Report ◽  
Initial Symptoms ◽  
Koch's Postulates

Cassia nomame (Sieb.) Kitagawa is an annual plant in the Leguminousae family. The aerial parts of C. nomame have been used as tonic and diuretic in Korea and Japan (Syed et al. 2019). A leaf spot was observed on the leaves of a 1-year-old C. nomame landrace in Changli County (39.42°N, 119.10°E), Qinhuangdao City, Hebei Province during August to October in 2018. In many fields (n≥3), the disease incidence over 80% in the middle and late stage of plant growth. Symptoms on leaves in one field began with many small, dark necrotic spot lesions. Later, the lesions spread to round-to-oval, slightly sunken in the center, and large necrotic patches with indefinite margins. Finally, lesions coalesced and resulted in defoliation. Lesions were occasionally observed on the pods. Symptoms on the pods were initially small, dark spots and then expanded to large necrotic patches with irregular edges. Symptomatic tissues (n=32) from pods and leaves were cut into 3 to 8 mm2 squares, surface disinfested with 75% ethanol for 10 s, rinsed with sterile distilled water, then placed on potato dextrose agar (PDA) at 28℃. After 3 days, ten isolates with consistent characteristics were obtained with a frequency 52.6%. The isolates on PDA were round, initially pale and had little aerial mycelium, gradually turned olive green and had dense wool-like dense aerial mycelia after 3 days. Conidia were hyaline, smooth, solitary, and elliptical. The conidia measured 5.4 to 8.2 μm × 2.5 to 3.8 μm (n=50), and has two oil bodies positioning at opposite poles. Pigmented chlamydospores were spherical or nearly pear-shaped, and solitary. Black fructifications (pycnidia) were produced profusely on PDA after subculture for 3 days. All the isolates were similar to Didymella sp. in morphology (Aveskamp et al. 2009). Choice three isolates YSGUO8 YSGGUO8-a and YSGGUO8-b to be further characterized by sequencing of the internal transcribed spacer (ITS), actin gene, and 28S large subunit of the nuclear rRNA gene (LSU) (Zhang et al. 2017). The sequences of three strains (MK836417 MZ484072 and MZ484073 for ITS, MK837604 MZ593675 and MZ593676 for actin, MK843781 MZ836208 and MZ836207 for LSU, respectively) showed 99% to 100% similarity with Didymella americana K-004 (KY070279 for ITS,KY070285 for LSU), Phoma americana CBS 256.65 (FJ426973 for ITS, FJ426871 for actin, MH870196 for LSU) and P. americana CBS 185.85 (FJ426972 for ITS, FJ426870 for actin, GU237990 for LSU) in GenBank. The fungi were identified as D. americana (formerly P. americana or Peyronellaea americana) on the basis of morphological characteristics and sequence analysis. A pathogenicity test was conducted with three times on 1-year-old C. nomame strain at the 4 to 6 compound leaf stage. Conidia were obtained from 7-day-old PDA cultures grown at 28℃ with a 12-h photoperiod. Koch’s postulates were fulfilled by spray inoculating ten healthy young plants with 106 conidia per milliliter of D. americana strain YSGUO8, and sterile water as the control. After inoculation, the plants were managed at 28℃, 60% relative humidity and a 12-h photoperiod. After 5 to 8 days, the inoculated leaves developed small and dark spots lesions similar to those observed on the leaves with initial symptoms in the field. The control leaves remained symptomless. The same fungi were re-isolated from infected leaves by morphology observation and sequence analysis, confirming Koch's postulates. D. americana has caused leaves spot on Table Beet in New York (Vaghefi et al. 2016). To our knowledge, this is the first report of D. americana causing leaf spot of C. nomame in China.

A New Leaf Spot Disease of Calotropis gigantean Caused by Alternaria alternata in Rajasthan, India

2009 ◽  
Vol 10 (1) ◽  
pp. 45
Author(s):  
Satish K. Sain ◽  
H. N. Gour ◽  
P. Sharma ◽  
P. N. Chowdhry
Keyword(s):  
Alternaria Alternata ◽  
Leaf Spot ◽  
First Record ◽  
Leaf Spot Disease ◽  
Koch’S Postulates ◽  
Calotropis Gigantea ◽  
Spot Disease ◽  
Koch's Postulates

Madar (Calotropis gigantea) is a medicinally important wild shrub native to India. The seed floss is used for furniture stuffing and the bark for nets and twine. In early 2005, we observed a leaf spot epidemic of madar growing on wasteland sites near the Sikar district of Rajasthan, India. Koch's Postulates were completed. This is the first record of the disease from the Sikar district of the Rajasthan state of India. Accepted for publication 6 February 2009. Published 31 March 2009.

scholarly journals First Report of a Leaf Spot Disease of Tobacco Caused by Pseudomonas cichorii in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Dayu Lan ◽  
Fangling Shu ◽  
Yanhui Lu ◽  
Anfa Shou ◽  
Wei Lin ◽  
...  
Keyword(s):  
Pseudomonas Syringae ◽  
Leaf Spot ◽  
Leaf Spot Disease ◽  
Pathogenicity Test ◽  
Pseudomonas Cichorii ◽  
Koch’S Postulates ◽  
First Report ◽  
Initial Symptoms ◽  
Wide Range ◽  
Koch's Postulates

Tobacco (Nicotiana tabacum L.), one of the chief commercial crops, is wildly cultivated worldwide. In June 2020 and 2021, an unknown bacterial leaf spot on tobacco was found in Hezhou and Hechi City, Guangxi, China. 30% of the tobacco were affected and the rate of diseased leaves reached about 10% in the field under high temperature and rainstorm. The disease mainly damaged the middle and top leaves of tobacco plants at vigorous growing stage. The initial symptoms were water-soaked spots on the frontal half of a leaf, and then expanded into circular to irregular spots with a yellow halo at the edge. The spots mostly appeared dark brown at high air humidity, while yellow brown at low humidity and exhibited a concentric pattern. In severe cases, the lesions coalesced and the whole leaf was densely covered with lesions, resulting in the loss of baking value. A bacterium was consistently isolated from diseased leaf tissues on nutrient agar (NA). Growth on NA was predominantly grayish white circular bacterial colonies with smooth margins, and the bacterium is rod-shaped, gram-negative and fluorescent on King’s B medium. Seven isolates (ND04A-ND04C and ZSXF02-ZSXF05) were selected for molecular identification and pathogenicity tests. Genomic DNA of the bacterium was extracted and the housekeeping gene of cts (encoding citrate synthase) was amplified with the primers cts-Fs/cts-Rs (forward primer cts-Fs: 5’-CCCGTCGAGCTGCCAATWCTGA-3’; reverse primer cts-Rs: 5’-ATCTCGCACGGSGTRTTGAACATC-3’) (Berge et al. 2014; Sarkar et al. 2004). 409-bp cts gene sequences were deposited in the GenBank database for seven isolates (accession no. OK105110-OK105116). Sequence of seven isolates shared 100% identity with several Pseudomonas cichorii strains within the GenBank database (accession no. KY940268 and KY940271), and the phylogenetic tree of cts genes of the seven isolates clustered with the phylogroup 11 of Pseudomonas syringae (accession no. KJ877799 and KJ878111), which was classified as P.cichorii. To satisfy Koch’s postulates, a pathogenicity test was tested by using a needle to dip a suspension of the bacterium (108 CFU/ml) and pricking three holes in the tobacco leaf. The control plants leaves were needled with sterile water. Each tobacco plant was inoculated with three leaves, and the test was repeated three times. All plants were placed in transparent plastic boxes and incubated in a greenhouse at 25 ± 3°C. The water-soaked spots appeared 24h after inoculation and quickly expanded through leaf veins. Three days after inoculation, all the inoculated leaves showed symptoms similar to those observed in the field. Control plants remained healthy. Only P. cichorii was successfully re-isolated from the lesions, confirming Koch’s postulates. Pseudomonas cichorii can infect eggplant, lettuce, tomatoand other crops, and has a wide range of hosts (Timilsina et al. 2017; Ullah et al. 2015). To our knowledge, this is the first report of P. cichorii causing leaf spot on tobacco in China.

scholarly journals First Report of Ramularia Leaf Spot Caused by Ramularia collo-cygni on Leaves and Seeds of Barley in Estonia

Plant Disease ◽  
2014 ◽  
Vol 98 (7) ◽  
pp. 997-997 ◽  
Author(s):  
P. Sooväli ◽  
M. Tikhonova ◽  
P. Matušinsky
Keyword(s):  
Leaf Spot ◽  
Growth Stage ◽  
Positive Control ◽  
Pathogenicity Test ◽  
Koch’S Postulates ◽  
Ramularia Leaf Spot ◽  
Initial Symptoms ◽  
Two Samples ◽  
Agricultural Regions ◽  
Koch's Postulates

Ramularia leaf spot (RLS) is a disease of barley (Hordeum vulgare) caused by the fungus Ramularia collo-cygni Sutton & Waller (Rcc). Rcc causes necrotic lesions, premature senescence of leaves, and yield reduction. Under Estonian conditions, there are usually no leaf spots on the upper leaves of barley prior to flowering. In 2009, 2010, and 2012, symptoms similar to those of RLS were observed on leaves of spring and winter barley in several Estonian agricultural regions. Approximately 30% of the plants in affected fields were symptomatic. Symptoms were not observed in 2011, which was a dry and hot year. Initial symptoms were small brown spots, beginning on the upper leaves (flag leaf, F-1 leaf) at the flowering growth stage (4). Later, the spots spread to the sheaths, stems, and awns and became necrotic. The lateral margins of the spots were delimited by the leaf veins and spots are surrounded by a chlorotic halo. During summer 2012, two samples of 15 F-1 leaves were collected from spring barley cv. Maali and line SJ111609 from the Estonian Crop Research Institute in eastern Estonia in late July at growth stage 71 (4). In addition, six grain samples, containing 200 seeds each of the cv. Maali, were collected from different agricultural regions in Estonia, along with one grain sample of SJ111609 from Jõgeva. All samples were collected from untreated plots and leaves were observed under a dissecting microscope, revealing white clusters of conidiophores in rows on the undersides of the leaves. Conidia and conidiophores were scraped aseptically from the leaf surface using a sterile needle under a dissecting microscope and transferred to potato dextrose agar (PDA) containing ampicillin sodium salt (50 mg l−1). Plates were incubated at 18°C in the dark for 20 days until fungal mycelia were produced. The fungus was initially identified as Rcc on the basis of morphological characteristics (3). Colorless, 0- to 3-septate conidiophores were 15 to 17 × 2 to 5 μm, with a strongly curved end. Conidia were 7 to 11 × 3 to 6 μm, solitary, subglobose, single-celled, and of a darkish color. To confirm the presence of Rcc, DNA was extracted from the original barley leaf material, milled seeds, and positive control mycelia of Rcc grown on PDA using DNeasy Plant Mini Kit (Qiagen Gmbh, D-40724 Hilden, Germany) following manufacturer's guides. Rcc specific primers RC3 and RC5 (1) were used. A positive control consisting of 1 ng of purified Rcc DNA was included in the PCR. Standard PCR was conducted in a SEE AMP Seegene cycler. PCR were carried out in 20 μl volumes, containing 2 μl of DNA, 10 μl PCR mix, 0.4 μl each of forward and reverse Rcc primers, and 7.2 μl H2O. Qualitative detection analyzed by standard PCR with primers RC3 and RC5 revealed the presence of Rcc in symptomatic leaves and seeds. To complete Koch's postulates, a pathogenicity test was performed. Twenty-five barley seedlings were grown under controlled conditions (15°C/48 h dark, 16 h light/8 h dark, 70% RH) and spray-inoculated with a suspension of Rcc mycelium fragments as described by Macepeace et al. (2). The pathogen was re-isolated from leaves with necrotic lesions similar to those observed in the field, thus fulfilling Koch's postulates. To our knowledge, this is the first confirmed report of Ramularia leaf spot caused by Ramularia collo-cygni on barley in Estonia. References: (1) P. Frei et al. J. Phytopathol. 155:281, 2007. (2) J. C. Makepeace et al. Plant Pathol. 57:991, 2008. (3) B. C. Sutton and J. M. Waller. Trans. Brit. Mycol. Soc. 90:55, 1988. (4) J. C. Zadoks et al. Weed Res. 14:415, 1974.

scholarly journals First report of Stemphylium eturmiunum causing leaf spot and black spot on apple in Zhaotong, Yunnan, China

Plant Disease ◽  
2021 ◽  
Author(s):  
Cong Zhou ◽  
Xin Long Pan ◽  
Bao Hua Kong ◽  
Yueqiu He ◽  
Jun Ma ◽  
...  
Keyword(s):  
Leaf Spot ◽  
Yunnan Province ◽  
Management Practices ◽  
Black Spot ◽  
Parsimony Method ◽  
Koch’S Postulates ◽  
First Report ◽  
The World ◽  
Postharvest Rot ◽  
Koch's Postulates

Apple is the largest fruit tree crop in the world, and China is the largest apple-producing County in the world. Zhaotong, Yunnan Province is a typical cold and mountainous apple-producing area in China. However, apple production is threatened by diseases during the entire growing season, and among them, apple leaf spot and fruit black spot are severe. In previous reports, the main pathogen causing apple leaf spot and fruit black spot was Alternaria sp. (Lior, et al, 2017), while different pathogens were identified. In the current study, seven red Fuji apple fruit with typical black spot samples were collected randomly in Dongda company orchard, Sujiayuan town, Zhaotong, Yunnan on March 25, 2021. The spots on the surface of these apples appear rounded, the diseased parts turn brown or black in colour and the flesh became soften and rotten. The tissues of fruit epidermis at the edge between diseased and healthy parts were cut, soaked in 75% alcohol for 30 s, washed with sterile water three times, and air-dried. Five pieces of tissue were placed on PDA medium amended with rifampicin (50 mg/ml) and incubated in the dark at 25 ℃ for 3-5 days. After colonies grew, mycelial clumps were picked out from the edges of the colonies, transferred to new PDA plates, and incubated at 25 ℃ for 6 days. The diameter of the colonies reached up to 5.7 cm. A representative isolate was retained for further work and was named P6-3-1. The hyphae were white and dense at an early stage, the culture medium on the underside became yellow and the middle parts of the colonies were darker. With maturity, hyphae were clumped, became red with other colors interspersed, and the medium became dark red. Light brown spores were produced, with more vertical septa and fewer transverse septa. Two to three transverse septa were generally observed with obvious constriction at the transverse septa. Average spore size was 22.83 µm ± 2.04 µm × 14.58 µm ± 1.97 µm. DNA was extracted from mycelium, purified and amplified with two pairs of primers, ITS1/ITS4 (White et al. 1990) and gpdF/gpdR (Marcos P. S. Câmara, et al. 2002). The PCR products were sequenced and deposited in GenBank (accession NO.OK560128 and OK627661 ). The similarity of ITS sequences between the isolate and MH843733 (Stemphylium eturmiunum strain ST14) was 100%, and that of gpd sequences between the isolate and MH843728 (Stemphylium eturmiunum strain ST20) was 100%. The maximum parsimony method of Mega7.0 was used and demonstrated that the studied isolate converged to the same branch as Stemphylium eturmiunum. Koch's postulates was applied to identify the pathogenicity of this isolate. A disc of P6-3-1-culture on PDA (5 mm in diameter) was placed on apple leaves and fruit wounds. Sterile PDA was used as a control. All plants were kept in a growth chamber at 25-30 ℃. Four days after inoculation, the disease spot was observed on the inoculated sites and fruit, and with the extension of incubation time, the diseased spots continue to grow, and the leaf spots were not limited by the veins. The pathogen was re-isolated from the inoculated leaves and fruit, satisfying Koch’s postulates. This pathogen can also cause postharvest rot of sweet cherry (Alice Spadoni, et al, 2020), postharvest rot on tomato (Prencipe Simona, et al, 2021), etc. This is the first report that Stemphylium eturmiunum can cause apple leaf spot and fruit black spot in Yunnan province, China. The apple black spot caused by Stemphylium eturmiunum was accurately identified. By distinguishing between the two similar diseases mentioned above, resistance to the host and management practices can be accrued based on the characteristics of the pathogen, its epidemiological pattern and the choice of an effective chemical fungicide.

scholarly journals First Report of Septoria Leaf Spot of Lavandin Caused by Septoria lavandulae in Croatia

Plant Disease ◽  
2014 ◽  
Vol 98 (2) ◽  
pp. 282-282
Author(s):  
K. Vrandečić ◽  
J. Ćosić ◽  
D. Jurković ◽  
I. Stanković ◽  
A. Vučurović ◽  
...  
Keyword(s):  
Leaf Spot ◽  
Antibacterial Properties ◽  
Morphological Characteristics ◽  
Its Region ◽  
Morphological Identification ◽  
Conidial Suspension ◽  
Necrotic Tissue ◽  
First Report ◽  
Initial Symptoms ◽  
Septoria Leaf Spot

Lavandula × intermedia Emeric ex Loiseleur, commonly known as lavandin, is an aromatic and medicinal perennial shrub widely and traditionally grown in Croatia. The lavandin essential oil is primarily used in perfumery and cosmetic industries, but also possesses anti-inflammatory, sedative, and antibacterial properties. In June 2012, severe foliar and stem symptoms were observed on approximately 40% of plants growing in a commercial lavandin crop in the locality of Banovo Brdo, Republic of Croatia. Initial symptoms on lower leaves included numerous, small, oval to irregular, grayish brown lesions with a slightly darker brown margin of necrotic tissue. Further development of the disease resulted in yellowing and necrosis of the infected leaves followed by premature defoliation. Similar necrotic oval-shaped lesions were observed on stems as well. The lesions contained numerous, dark, sub-globose pycnidia that were immersed in the necrotic tissue or partly erumpent. Small pieces of infected internal tissues were superficially disinfected with 50% commercial bleach (4% NaOCl) and placed on potato dextrose agar (PDA). A total of 10 isolates from leaves and five from stems of lavandin formed a slow-growing, dark, circular colonies with raised center that produced pycnidia at 23°C, under 12 h of fluorescent light per day. All 15 recovered isolates formed uniform hyaline, elongate, straight or slightly curved conidia with 3 to 4 septa, with average dimensions of 17.5 to 35 × 1.5 to 2.5 μm. Based on the morphological characteristics, the pathogen was identified as Septoria lavandulae Desm., the causal agent of lavender leaf spot (1,2). Pathogenicity of one selected isolate (428-12) was tested by spraying 10 lavandin seedlings (8 weeks old) with a conidial suspension (106 conidia/ml) harvested from a 4-week-old monoconidial culture on PDA. Five lavandin seedlings, sprayed with sterile distilled water, were used as negative control. After 5 to 7 days, leaf spot symptoms identical to those observed on the source plants developed on all inoculated seedlings and the pathogen was successfully re-isolated. No symptoms were observed on any of the control plants. Morphological identification was confirmed by amplification and sequencing of the internal transcribed spacer (ITS) region of rDNA (3). Total DNA was extracted directly from fungal mycelium with a DNeasy Plant Mini Kit (Qiagen, Hilden, Germany) and PCR amplification performed with primers ITS1F/ITS4. Sequence analysis of ITS region revealed at least 99% identity between the isolate 428-12 (GenBank Accession No. KF373078) and isolates of many Septoria species; however, no information was available for S. lavandulae. To our knowledge, this is the first report of Septoria leaf spot of lavandin caused by S. lavandulae in Croatia. Since the cultivation area of lavandin plants has been increasing in many continental parts of Croatia, especially in Slavonia and Baranja counties, the presence of a new and potentially harmful disease may represent a serious constraint for lavandin production and further monitoring is needed. References: (1) T. V. Andrianova and D. W. Minter. IMI Descriptions of Fungi and Bacteria, 142, Sheet 1416, 1999. (2) R. Bounaurio et al. Petria 6:183, 1996. (3) G. J. M. Verkley et al. Mycologia 96:558, 2004.

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