First Report of Wilt of European hornbeam (Carpinus betulus L.) Caused by Fusarium oxysporum in China

European hornbeam (Carpinus betulus L.) has been used as an important ornamental species for urban landscaping since the Italian Renaissance (Rocchi et al. 2010). In May 2019, 15% of 3000 C. betulus trees with wilted leaves and root rot were observed in a field (about 26 ha) in Pizhou, Jiangsu Province, China. Internal discoloration of the stem began with brown to black discoloration of the vascular system and gradually spread to inward areas. Roots and stems from symptomatic plants were washed free of soil, surface sterilized with 0.8% NaOCl, rinsed three times in sterile H2O, and blotted dry with a paper towel. Small segments (0.5-cm-long) were cut from the discolored vascular tissues, and then put on potato dextrose agar (PDA) at 25°C in darkness. After 4 days, fungal colonies were observed on the PDA. Pure cultures were obtained by monosporic isolation, and 9 morphologically similar fungal isolates (EJ-1 to EJ-9) were obtained. All purified cultures were incubated on PDA at 25°C in darkness as the initial isolation. Colonies of the 9 isolates on PDA displayed entire margins and showed abundant pink aerial mycelia initially and turned to light violet with age. Microconidia were elliptical or oval in shape, 0 septate, (5.2–)8.7(–12.5) × (3.5–)3.6(–5.5) µm. Macroconidia were falciform, 0-4 septate, and straight to slightly curved with a notched foot cell, (17.1–)20.5(–28.4) × (3.8–)4.1(–4.6) µm. These morphological characteristics resemble Fusarium oxysporum (Leslie and Summerell 2006). Genomic DNA of each isolate was extracted from mycelia using a CTAB method (Mo¨ller et al. 1992). The RPB2, TEF1 and cmdA genes were amplified and sequenced with the primers 5f2/7c (Liu et al. 2000), EF-1Ha/EF-2Tb (Carbone and Kohn 1999) and Cal228F/CAL2Rd (Groenewald et al. 2013), respectively. The sequences were deposited in GenBank (Table 1). A maximum likelihood phylogenetic analysis based on RPB2, TEF1 and cmdA sequences using MEGA7 revealed that the isolates were placed in the F. oxysporum species complex with 98% bootstrap support. Based on the morphological and molecular characters, all 9 isolates were identified as F. oxysporum. A pathogenicity experiment was conducted using 30 2-year-old C. betulus seedlings potted in sterile peat, 27 for inoculation (3 replicate plants per isolate) and 3 for a negative control. The treated plants were planted in the peat mixed with 50 ml of a conidial suspension of each isolate respectively. The negative control was inoculated with sterilized water. Conidia were harvested from colonized plates of PDA using sterilized water and adjusted to a concentration of 1×107 conidia/ml. All 30 seedlings were incubated in a greenhouse at 25°C with a relative humidity of 80% and a 12-h photoperiod. The inoculated seedlings displayed wilt symptoms within 30 to 40 days, and eventually died within 75 to 85 days after inoculation. Control plants remained symptomless. F. oxysporum was successfully reisolated from the vascular tissues of symptomatic plants, and sequences of RPB2, TEF1 and cmdA of re-isolates matched those of the original isolates. No pathogen was isolated from the tissues of control plants. The experiment was repeat twice with the similar results, fulfilling Koch's postulates. F. oxysporum is an important soil-borne pathogen and can cause disease in many economic plants, such as yellowwood (Graney et al. 2016), hickory (Zhang et al. 2015) and larch (Rolim et al. 2020). To our knowledge, this is the first report of wilt on C. betulus caused by F. oxysporum in China.

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First Report of Pythium aphanidermatum Causing Stalk Rot on Abelmoschus manihot (L. ) Medic in China

Plant Disease ◽

10.1094/pdis-06-21-1133-pdn ◽

2021 ◽

Author(s):

Qing Qu ◽

Liu Shiwei ◽

Ning Liu ◽

Yunxia Liu ◽

Jia Hui ◽

...

Keyword(s):

Management Practices ◽

Vascular System ◽

Morphological Characteristics ◽

Pythium Aphanidermatum ◽

Negative Control ◽

Perennial Herb ◽

Universal Primers ◽

First Report ◽

Stalk Rot ◽

Aerial Hyphae

Abelmoschus manihot (Linn. ) Medicus (A. manihot) is an annual to perennial herb of the Malvaceae okra, mainly distributed in Guangdong, Guangxi, Fujian, Hunan, Hubei provinces. It can not only be used as an ornamental flower, but also has important economic and medicinal value. Last year, 10% A. manihot in 1,000 acres were observed with stalk rot in the Zhongshang Agricultural Industrial Park, 50 meters east of Provincial Highway 235 in Gaoyang County of Hebei province. Internal discoloration of the stem began brown to black discoloration of the vascular system and became hollow, with the mycelium growing on the surface. Stems from symptomatic plants (approximately 5 mm2) were dissected, washed free of soil, then soaked in 75% ethanol for 16 s to surface-sterilize, and 40 s in HgCl2, then rinsed three times in sterile water. After being dried with blotting paper, five pieces were placed on potato dextrose agar (PDA). After cultured 2 or 3 days, five isolates were purified and re-cultured on PDA in the dark at 25°C. The color of the colony was white. The hyphae were radial in PDA, and the aerial hyphae were flocculent, well-developed with luxuriant branches. The colonies were white and floccus, and the aerial hyphae were well developed, branched and without septum on corn meal agar (CMA). The sporangia were large or petal shaped, composed of irregular hyphae, terminal or intermediate , with the size of (31.6-88.4) μm ×(12.7- 14.6) μm. Vesicles were spherical, terminal or intermediate, ranging from 14.6 to 18.5μm. Oogonia were globose, terminal and smooth which stipe was straight. Antheridia were clavate or baggy and mostly intercalary, sometimes terminal. Oospores were aplerotic, 21.5 to 30.0 μm in diameter, 1.6 to 3.1 μm in wall thickness. The isolates morphological characteristics were consistent with P. aphanidermatum (van der Plaats-Niterink 1981, Wu et al. 2021 ). To identify the isolates, universal primers ITS1/ITS4 (White et al. 1993) were used for polymerase chain reaction–based molecular identification. The amplification region was sequenced by Sangon Biotech (Shanghai, China) and submitted to GenBank (MW819983). BLAST analysis showed that the sequence was 100% identical to Pythium aphanidermatum. Pathogenicity tests were conducted 3 times, with 4 treatments and 2 controls each time. The plants treated were 6 months old. Then the hyphae growing on PDA for 7 days were cut into four pieces. Next, they were inoculated into the soil of the A. manihot. Negative control was inoculated only with PDA for 7 days ( Zhang et al. 2000). The plants were then placed in a greenhouse under 28°C, 90% relative humidity. After inoculated 20 to 30 days, the infected plants showed stalk rot, the same symptoms as observed on the original plants. The control plants didn’t display symptoms. Pythium aphanidermatum was re-isolated from infected stems and showed the same characteristics as described above and was identical in appearance to the isolates used to inoculate the plants. To our knowledge, this is the first report of Pythium aphanidermatum infecting A. manihot stem and causing stalk rot in China. It may become a significant problem for A. manihot. Preliminary management practices are needed for reducing the cost and losses of production.

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First Report on Ovate-leaf Atractylodes Damping-off Caused by Fusarium oxysporum species Complex in South Korea

Plant Disease ◽

10.1094/pdis-05-21-1107-pdn ◽

2021 ◽

Author(s):

Oliul Hassan ◽

Taehyun Chang

Keyword(s):

South Korea ◽

Fusarium Oxysporum ◽

Species Complex ◽

Disease Incidence ◽

Fusarium Species ◽

Morphological Characteristics ◽

Conidial Suspension ◽

Sterile Water ◽

Damping Off ◽

First Report

In South Korea, ovate-leaf atractylodes (OLA) (Atractylodes ovata) is cultivated for herbal medicine. During May to June 2019, a disease with damping off symptoms on OLA seedlings were observed at three farmer fields in Mungyeong, South Korea. Disease incidence was estimated as approximately 20% based on calculating the proportion of symptomatic seedlings in three randomly selected fields. Six randomly selected seedlings (two from each field) showing damping off symptoms were collected. Small pieces (1 cm2) were cut from infected roots, surface-sterilized (1 minute in 0.5% sodium hypochlorite), rinsed twice with sterile water, air-dried and then plated on potato dextrose agar (PDA, Difco, and Becton Dickinson). Hyphal tips were excised and transferred to fresh PDA. Six morphologically similar isolates were obtained from six samples. Seven-day-old colonies, incubated at 25 °C in the dark on PDA, were whitish with light purple mycelia on the upper side and white with light purple at the center on the reverse side. Macroconidia were 3–5 septate, curved, both ends were pointed, and were 19.8–36.62 × 3.3–4.7 µm (n= 30). Microconidia were cylindrical or ellipsoid and 5.5–11.6 × 2.5–3.8 µm (n=30). Chlamydospores were globose and 9.6 –16.3 × 9.4 – 15.0 µm (n=30). The morphological characteristics of present isolates were comparable with that of Fusarium species (Maryani et al. 2019). Genomic DNA was extracted from 4 days old cultures of each isolate of SRRM 4.2, SRRH3, and SRRH5, EF-1α and rpb2 region were amplified using EF792 + EF829, and RPB2-5f2 + RPB2-7cr primer sets, respectively (Carbone and Kohn, 1999; O'Donnell et al. 2010) and sequenced (GenBank accession number: LC569791- LC569793 and LC600806- LC600808). BLAST query against Fusarium loci sampled and multilocus sequence typing database revealed that 99–100% identity to corresponding sequences of the F. oxysporum species complex (strain NRRL 28395 and 26379). Maximum likelihood phylogenetic analysis with MEGA v. 6.0 using the concatenated sequencing data for EF-1α and rpb2 showed that the isolates belonged to F. oxysporum species complex. Each three healthy seedlings with similar sized (big flower sabju) were grown for 20 days in a plastic pot containing autoclaved peat soil was used for pathogenicity tests. Conidial suspensions (106 conidia mL−1) of 20 days old colonies per isolate (two isolates) were prepared in sterile water. Three pots per strain were inoculated either by pouring 50 ml of the conidial suspension or by the same quantity of sterile distilled water as control. After inoculation, all pots were incubated at 25 °C with a 16-hour light/8-hour dark cycle in a growth chamber. This experiment repeated twice. Inoculated seedlings were watered twice a week. Approximately 60% of the inoculated seedlings per strain wilted after 15 days of inoculation and control seedlings remained asymptomatic. Fusarium oxysporum was successfully isolated from infected seedling and identified based on morphology and EF-1α sequences data to confirm Koch’s postulates. Fusarium oxysporum is responsible for damping-off of many plant species, including larch, tomato, melon, bean, banana, cotton, chickpea, and Arabidopsis thaliana (Fourie et al. 2011; Hassan et al.2019). To the best of our knowledge, this is the first report on damping-off of ovate-leaf atractylodes caused by F. oxysporum in South Korea. This finding provides a basis for studying the epidemic and management of the disease.

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First Report of Fusarium Wilt in Blueberry (Vaccinium corymbosum) Caused by Fusarium oxysporum in China

Plant Disease ◽

10.1094/pdis-02-14-0167-pdn ◽

2014 ◽

Vol 98 (8) ◽

pp. 1158-1158 ◽

Cited By ~ 1

Author(s):

Y. H. Liu ◽

T. Lin ◽

C. S. Ye ◽

C. Q. Zhang

Keyword(s):

Fusarium Oxysporum ◽

Infected Plant ◽

Morphological Characteristics ◽

Vaccinium Corymbosum ◽

Morphological Characterization ◽

Internal Transcribed Spacers ◽

Fluorescent Light ◽

Pathogenicity Test ◽

Conidial Suspension ◽

First Report

Blueberry (Vaccinium corymbosum) production is developing quickly in China with about 20,000 ha presently cultivated. In 2010 in Lin'an, Zhejiang Province, plants developed an apparently new disease of blueberry (cv. Duke) with symptoms consisting of wilting of foliage, stunting of plants, and reduced fruit yields. Internal vascular and cortical tissues of plant crowns showed a brown to orange discoloration. Approximately 3% of the plants in the commercial plantings were affected and eventually died after 50 to 60 days. Infected plant samples (stems and roots) collected from different fields were surface sterilized with 1.5% sodium hypochlorite for 2 min, rinsed in water, plated on 2% potato dextrose agar (PDA), and incubated at 25°C in the dark for 1 week. Single conidium cultures were consistently isolated and cultured on acidified PDA (APDA) for morphological characterization (1,2). Colonies were light with purple mycelia, and beige or orange reverse colony colors developed after 7 days incubation at 25°C. Colonies producing abundant microconidia and macroconidia. Microconidia were hyaline and oval-ellipsoid to cylindrical (3.9 to 9.6 × 1.1 to 3.4 μm). Macroconidia were 3 to 5 septate and fusoid-subulate with a pedicellate base (28.6 to 37.5 × 3.3 to 4.2 μm). Morphology and development of macroconidia and microconida were consistent with a description of Fusarium oxysporum Schltdl (1,2). The ribosomal internal transcribed spacers ITS1 and ITS2 of eight isolates were amplified using primers ITS1/ITS4 on DNA extracted from mycelium and nucleotide sequences showed 100% similarity to that of F. oxysporum. To confirm pathogenicity, 20 blueberry plants (cv. Duke) were inoculated by dipping the roots into a conidial suspension (107 conidia per ml) for 30 min. The inoculated plants were transplanted into pots containing sterilized peat and maintained at 25°C and 100% relative humidity in a growth chamber with a daily 12-h photoperiod of fluorescent light. The pathogenicity test was conducted twice. Within 40 days, all inoculated plants developed wilt symptoms similar to that observed in the field. No symptoms were observed on plants dipped into distilled water. The fungus was successfully re-isolated from crowns and roots cultured on APDA, exhibiting morphological characteristics identical to F. oxysporum (1,2), confirming Koch's postulates. To our knowledge, this is the first report of blueberry wilt caused by Fusarium. References: (1) P. M. Kirk et al. The Dictionary of the Fungi, 10th edition, page 159. CABI Bioscience, Wallingford, UK, 2008. (2) W. C. Snyder and H. N. Hansen. Am. J. Bot. 27:64, 1940.

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First Report of Fusarium oxysporum on Lettuce in Europe

Plant Disease ◽

10.1094/pdis.2002.86.9.1052b ◽

2002 ◽

Vol 86 (9) ◽

pp. 1052-1052 ◽

Cited By ~ 42

Author(s):

A. Garibaldi ◽

G. Gilardi ◽

M. L. Gullino

Keyword(s):

Fusarium Oxysporum ◽

Vascular System ◽

The United States ◽

Selective Medium ◽

Northern Italy ◽

Infested Soil ◽

Pathogenicity Test ◽

First Report ◽

Vascular Tissues ◽

Brown Discoloration

In spring 2001, plants of the lettuce cv. Salad Bowl showing symptoms of a wilt disease were observed in several commercial plastic greenhouses near Bergamo, in northern Italy. Wilted plants were first observed during the spring and summer of 2001 when temperatures were between 26 and 35°C. Symptoms were observed in the same area and the same farms in March 2002, in concomitance with a period of high temperatures. Although the distribution of the disease was generally uniform, symptoms were more severe in the central part of the greenhouses where temperatures were warmest. Symptoms were first observed at thinning, when seedlings (30 days old) appeared wilted. Vascular tissues of affected seedlings appeared red or brown. Affected plants were stunted and developed yellow leaves and brown or black streaks in the vascular system. The vascular streaks in the yellow leaves extended from the crown and were continuous with a red-brown discoloration in the vascular system of the crown and upper taproot. Symptoms were typically not visible on the outside of the crowns or roots. Fusarium oxysporum was consistently and readily isolated from symptomatic vascular tissues on a Fusarium-selective medium (2). Seeds of cv. Salad Bowl were planted in steam-sterilized soil artificially infested with 1 × 104 CFU/g soil of each of two isolates of F. oxysporum obtained from infected plants. Plants grown in noninfested soil served as noninoculated control treatments. Plants (25 per treatment) were grown at 25 to 28°C in growth chambers. Wilt symptoms and vascular discoloration of the roots, crown, and veins developed 20 days after seeding for all plants grown in the infested soil, and F. oxysporum was consistently reisolated from infected plants. The pathogenicity test was conducted twice. To our knowledge, this is the first report of F. oxysporum on lettuce in Europe. A wilt of lettuce attributed to F. oxysporum f. sp. lactucae was reported in Japan in 1967 (3) and in the United States in 1993 (1), in the latter case the incitant organism was described as F. oxysporum f. sp. lactucum. References: (1) J. C. Hubbard and J. S. Gerik. Plant Dis. 77:750, 1993. (2) H. Komada. Rev. Plant Prot. Res. 8:114, 1975. (3) T. Matuo and S. Motohashi. Trans. Mycol. Soc. Jpn. 8:13, 1967.

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First report of leaf spot disease caused by Corynespora cassiicola on American sweetgum (Liquidambar styraciflua L.) in China

Plant Disease ◽

10.1094/pdis-08-20-1842-pdn ◽

2021 ◽

Author(s):

Yun-fei Mao ◽

Xiang-rong Zheng ◽

Fengmao Chen

Keyword(s):

Leaf Spot ◽

Morphological Characteristics ◽

Liquidambar Styraciflua ◽

Bootstrap Support ◽

Leaf Spot Disease ◽

Pathogenicity Test ◽

Conidial Suspension ◽

Corynespora Cassiicola ◽

First Report ◽

Leaf Spots

American sweetgum (Liquidambar styraciflua L.) is a forest plant native to North America, which has been introduced into other countries due to its ornamental and medicinal values. In June 2019, symptoms of leaf spots on sweetgum were observed in a field (5 ha) located in Xuzhou, Jiangsu Province, China. On this field, approximately 45% of 1,000 trees showed the same symptoms. Symptoms were observed showing irregular or circular dark brown necrotic lesions approximately 5 to 15 mm in diameter with a yellowish margin on the leaves. To isolate the pathogen, diseased leaf sections (4×4mm) were excised from the margin of the lesion, surface-sterilized with 0.1% NaOCl for 90 s, rinsed 4 times in sterile distilled water, air dried and then transferred on potato dextrose agar (PDA) medium at 25°C in the dark. Pure cultures were obtained by monospore isolation after subculture. Ten purified isolates, named FXI to FXR, were transferred to fresh PDA and incubated as above to allow for morphological and molecular identification. After 7 days, the aerial mycelium was abundant, fluffy and exhibited white to greyish-green coloration. The conidia were dark brown or olive, solitary or produced in chains, obclavate, with 1 to 15 pseudosepta, and measured 45 to 200µm  10 to 18µm. Based on morphological features, these 10 isolates were identified as Corynespora cassiicola (Ellis et al. 1971). Genomic DNA of each isolate was extracted from mycelia using the cetyltrimethylammonium bromide (CTAB) method. The EF-1α gene and ITS region were amplified and sequenced with the primer pairs rDNA ITS primers (ITS4/ITS5) (White et al. 1990) and EF1-728F/EF-986R (Carbone et al.1999) respectively. The sequences were deposited in GenBank. BLAST analysis revealed that the ITS sequence had 99.66% similarity to C. cassiicola MH255527 and that the EF-1α sequence had 100% similarity to C. cassiicola KX429668A. maximum likelihood phylogenetic analysis based on EF-1α and ITS sequences using MEGA 7 revealed that ten isolates were placed in the same clade as C. cassiicola (Isolate: XQ3-1; accession numbers: MH572687 and MH569606, respectively) at 98% bootstrap support. Based on the morphological characteristics and phylogenetic analyses, all isolates were identified as C. cassiicola. For the pathogenicity test, a 10 µl conidial suspension (1×105 spores/ml) of each isolate was dripped onto healthy leaves of 2-year-old sweetgum potted seedlings respectively. Leaves inoculated with sterile water served as controls. Three plants (3 leaves per plant) were conducted for each treatment. The experiment was repeat twice. All seedlings were enclosed in plastic transparent incubators to maintain high relative humidity (90% to 100%) and incubated in a greenhouse at 25°C with a 12-h photoperiod. After 10 days, leaves inoculated with conidial suspension of each isolate showed symptoms of leaf spots, similar to those observed in the field. Control plants were remained healthy. In order to reisolate the pathogen, surface-sterilized and monosporic isolation was conducted as described above. The same fungus was reisolated from the lesions of symptomatic leaves, and its identity was confirmed by molecular and morphological approaches, thus fulfilling Koch’s postulates. Chlorothalonil and Boscalid can be used to effectively control Corynespora leaf spot (Chairin T et al.2017). To our knowledge, this is the first report of leaf spot caused by C. cassiicola on L. styraciflua in China.

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First Report of Cylindrocladium pseudonaviculatum Causing Leaf Spot of Pachysandra terminalis

Plant Disease ◽

10.1094/pdis-03-12-0235-pdn ◽

2012 ◽

Vol 96 (7) ◽

pp. 1069-1069 ◽

Cited By ~ 19

Author(s):

J. A. LaMondia ◽

D. W. Li ◽

R. E. Marra ◽

S. M. Douglas

Keyword(s):

Leaf Spot ◽

Morphological Characteristics ◽

Ground Cover ◽

Soil Surface ◽

Leaf Spot Disease ◽

Terminal Branch ◽

Conidial Suspension ◽

First Report ◽

Buxus Sempervirens ◽

Stem Lesions

Cylindrocladium pseudonaviculatum Crous, J.Z., Groenew. & C.F. Hill 2002 was recently reported infecting common boxwood, Buxus sempervirens L., in Connecticut (2). We isolated the pathogen from leaf and stem lesions of B. sempervirens and obtained single-spored cultures on half-strength potato dextrose agar (½PDA). The pathogen was identified as C. pseudonaviculatum by morphological characteristics (1). Colony size reached 71 mm in diameter after 14 days at room temperature on ½PDA, and was fluffy with white aerial hyphae, mars brown, and reverse color chestnut brown at the center fading to pale brown forming concentric bands. Macroconidiophores were solitary or in a group of up to three, comprised a stipe, a sterile elongation, and one to three penicillate fertile branches. The stipe was up to nine septate, 90 to 250 μm long, colorless, smooth, terminating in a naviculate or broadly ellipsoidal vesicle with a pointed or papillate apex, and 27 to 50 × 6.5 to 9 μm. Primary branches were zero- to one-septate, 20 to 36 × 4 to 5 μm; secondary branches were aseptate and 11 to 20 × 3 to 4.5 μm; tertiary branches were rare, each terminal branch producing two to five phialides; phialides were doliiform or reniform, colorless, 12 to 18 μm. Conidia were cylindrical, rounded at both ends, straight, smooth, colorless, two-celled, 48 to 55 × 4.5 to 5.5 μm, and in colorless slimy cylindrical clusters. Microconidiophores were not observed. Chlamydospores were golden to dark brown, thick-walled, and smooth or rough. Microsclerotia were present on ½PDA. Primers T1 and T22 (3) were used to amplify a portion of the β-tubulin gene from isolates Cps-CT-L1 and Cps-CT-S1. Amplified sequences were used in a BLAST search against the GenBank database to demonstrate 100% sequence identity only with other C. pseudonaviculatum strains. Both sequences were deposited in GenBank (Accession Nos. JQ866628 and JQ866629), using corresponding gene data from C. pseudonaviculatum strain STE-U 3399 (GenBank Accession No. AF449455) to distinguish coding from noncoding regions. Healthy plants of Japanese spurge, Pachysandra terminalis, with three plants per 10 cm diameter pot, were inoculated with water alone or a conidial suspension of C. pseudonaviculatum isolate Cps-CT-L1 (ATCC MYA-4891) (1.0 × 106 conidia/plant) with a handheld sprayer until runoff. Plants were kept moist in a plastic bag for 48 h at laboratory temperature and then transferred to the greenhouse. Circular lesions (1- to 4-mm diameter) were evident on leaves after 10 days. All 12 inoculated plants developed lesions, and no lesions were observed on noninoculated plants. Leaves with lesions were surface sterilized in 0.5% NaOCl for 30 s, rinsed twice in sterile water, and lesion margins plated onto water agar or ½PDA. The pathogen was reisolated from at least one leaf per plant. Koch's postulates were performed again with isolate Cps-CT-S1 (ATCC MYA-4890). After 3 weeks, many of the leaves with lesions yellowed and dropped to the soil surface and heavy sporulation of C. pseudonaviculatum and microsclerotia were observed. To our knowledge, this is the first report of C. pseudonaviculatum causing a leaf spot disease on P. terminalis. Pachysandra is a widely grown ground cover suitable for shady, humid environmental conditions that may be conducive for the development of disease. References: (1) P. Crous, et al. Sydowia 54:23, 2002. (2) K. Ivors et al. Plant Disease. 96:X, 2012. (3) K. O'Donnell and E. Cigelnik Mol. Phylogenet. Evol. 7:103, 1997.

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First Report of Fusarium oxysporum on Eruca vesicaria and Diplotaxis spp. in Europe

Plant Disease ◽

10.1094/pdis.2003.87.2.201a ◽

2003 ◽

Vol 87 (2) ◽

pp. 201-201 ◽

Cited By ~ 9

Author(s):

A. Garibaldi ◽

G. Gilardi ◽

M. L. Gullino

Keyword(s):

Fusarium Oxysporum ◽

Vascular System ◽

Selective Medium ◽

Wild Plants ◽

Pathogenicity Test ◽

First Report ◽

Vascular Tissues ◽

Brown Discoloration ◽

Diplotaxis Erucoides ◽

Wild Rocket

Two types of rocket are available on the market in Italy: (i) Eruca vesicaria (synonym E. sativa) known as ruchetta or cultivated garden rocket; and (ii) several species of Diplotaxis (Diplotaxis erucoides, D. muralis, and D. tenuifolia), which are wild plants now widely cultivated. Rocket is increasingly used in the mediterranean cuisine as salad and or to decorate dishes. In spring 2002, plants of the cultivated (E. vesicaria) and wild (Diplotaxis spp.) rocket showing symptoms of a wilt disease were observed in several commercial plastic greenhouses near Bergamo, in northern Italy. Wilted plants were first observed during the spring and summer of 2001 when temperatures were between 26 and 35°C. In May 2002, symptoms were again observed in the same area, on the same farm as well as other farms. Although the distribution of the disease was generally uniform, symptoms were more severe in the central part of the greenhouses where temperatures were warmest (30 to 60% of plants were affected). Diseased plants were stunted and chlorotic with brown or black streaks in the vascular system. Vascular tissues of affected plants appeared red or brown. Vascular streaks in the chlorotic leaves extended from the crown and were continuous with a red-brown discoloration in the vascular system of the crown and upper taproot. Fusarium oxysporum was consistently and readily isolated from symptomatic vascular tissues when plated on a Fusarium-selective medium (2). Microconidia measured 8.8 × 3.0 µm. E. vesicaria and Diplotaxis spp. were grown in steam-sterilized soil, and 10 days after emergence they were artificially inoculated by root dipping in a spore suspension (1 × 105 CFU/ml) of three F. oxysporum strains collected from infected plants. Uninoculated plants served as control. Plants (60 per treatment) were grown at 25 to 28°C in growth chambers. Wilt symptoms developed on all plants 20 days after inoculation, and F. oxysporum was consistently reisolated from infected plants. The pathogenicity test was conducted twice. To our knowledge, this is the first report of F. oxysporum on cultivated rocket in Europe and the first on wild rocket (Diplotaxis spp.) in the world. A wilt of E. sativa attributed to F. oxysporum f. sp. erucae was previously reported in India in 1973 (1). Studies are being carried out to determine if the Italian isolates of F. oxysporum belong to the same formae speciales. References: (1) C. Chatterjee and J. N. Rai. Indian Phytopathol 28:309, 1973. (2) H. Komada. Rev. Plant Prot. Res. 8:114, 1975.

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First Report of Fusarium Wilt of Strawberry Caused by Fusarium oxysporum in Serbia

Plant Disease ◽

10.1094/pdis-04-14-0346-pdn ◽

2014 ◽

Vol 98 (10) ◽

pp. 1435-1435 ◽

Cited By ~ 4

Author(s):

I. Stanković ◽

D. Ristić ◽

A. Vučurović ◽

K. Milojević ◽

D. Nikolić ◽

...

Keyword(s):

Fusarium Oxysporum ◽

Fusarium Wilt ◽

Fluorescent Light ◽

Fungal Mycelium ◽

Morphological Identification ◽

Post Inoculation ◽

Conidial Suspension ◽

First Report ◽

Vascular Tissues ◽

Initial Symptoms

Strawberry (Fragaria × ananassa Duch.) is the third most important berry crop in Serbia with average production ranging from 30,000 to 35,000 t on approximately 5,000 ha (2). In June 2013, symptoms of wilt and whole plant collapse were observed on approximately 25% plants growing in commercial strawberry crop of cv. Alba in the locality of Zablaće (Moravica district). Initial symptoms included leaf chlorosis and wilt, followed by withering and necrosis of older leaves and reduced fruit production, eventually leading to plant collapse and desiccations. Internal vascular tissues of the crown showed distinct brown reddish discoloration. Three small pieces of infected roots, petioles, or crown vascular tissues were surface disinfested with 2% NaOCl and placed on five potato dextrose agars (PDA) per sample. After 7 days incubation at 23°C under 12 h of fluorescent light, nine monoconidial isolates were obtained (1) forming colonies with light purple mycelia. Colonies produced numerous hyaline, oval to ellipsoid microconidia (5 to 15 × 2.5 to 4.5 μm, average 8.45 × 2.25 μm), 3 to 5 septate fusoid macroconidia with pedicellate bases (20 to 50 × 2.70 to 6 μm, average 32.35 × 3.25 μm from 100 measured) and chlamydospores. Morphological and growth features were similar to the descriptions of Fusarium oxysporum Schlechtend emend. Snyder & Hansen (1). Pathogenicity of one selected isolate (97-13) was tested by dipping for 15 min the roots of five plants of each cultivar: Alba, Arosa, Clery, and Roxana into a conidial suspension (1 × 106 conidia/ml) harvested from a 7-day-old culture on PDA. Control plants were dipped in sterile distilled water. The inoculated plants were transplanted into pots containing sterilized peat and maintained in the greenhouse at 25°C. Thirty to thirty-five days post-inoculation, all plants developed wilt symptoms and vascular discoloration of crown tissues from which F. oxysporum was successfully re-isolated using the same method as for isolation. No symptoms were observed on any of the control plants. Morphological identification was confirmed by amplification and sequencing of a portion of the translation elongation factor-1 alpha (EF-1α) gene. Total DNA was extracted directly from fungal mycelium with a DNeasy Plant Mini Kit (Qiagen, Hilden, Germany) and PCR amplification performed with primers EF-1/EF-2 (4). Sequence analysis of EF-1α region revealed that Serbian isolate 97-13 (GenBank Accession No. KJ647280) shared 99 to 100% identity with the F. oxysporum sequences in GenBank. To our knowledge, this is the first report of Fusarium wilt on strawberry in Serbia. The presence of a new and potentially harmful disease may represent a serious constraint for strawberry production in Serbia. References: (1) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual, Blackwell Publishing, London, UK, 2006. (2) M. Nikolić et al. Acta Hort. 842:615, 2009. (3) K. O'Donnell et al. Proc. Natl. Acad. Sci. USA 95:2044, 1998.

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First Report of Diaporthe longicolla Causing Leaf Spot on Kalanchoe pinnata in China

Plant Disease ◽

10.1094/pdis-12-20-2681-pdn ◽

2021 ◽

Author(s):

Xiaodong Sun ◽

Xinglai Cai ◽

Qiangqiang Pang ◽

Man Zhou ◽

Wen Zhang ◽

...

Keyword(s):

Leaf Spot ◽

Southern China ◽

Elongation Factor ◽

Morphological Characteristics ◽

Negative Control ◽

Hainan Province ◽

Conidial Suspension ◽

Single Spore ◽

First Report ◽

Kalanchoe Pinnata

Kalanchoe pinnata (Lam.) Pers. [syn.: Bryophyllum pinnatum (Lam.) Oken] is an important medicinal agent in southern China. The succulent leaves of this plant are used in the treatment of cholera, bruises, urinary diseases and whitlow. In Oct. 2019, leaf spots were detected on K. pinnata plants in Chengmai County, Hainan Province, China. Lesions with brown to black margins were irregularly shaped and associated with leaf margins. Spots coalesced to form larger lesions (Fig. S1-A), with black pycnidia present in more mature lesions. Symptomatic K. pinnata were found with 10-20% incidence during the humid winters of Hainan Province. Leaf tissues of 10 symptomatic plants were collected and surface sterilized in 70% ETOH for 30s, 0.1% HgCl2 for 30 s, rinsed 3x with sterile distilled water for 30s, placed on potato dextrose agar (PDA) amended with 30mg/L of kanamycin sulfate, and incubated at 25°C in the dark for 3-5 days. Four fungal isolates were obtained using a single-spore isolation method. The colonies were floccose, dense, and white with forming on older colonies grown on PDA (Fig. S1-B-1&2). Alpha conidia exuded from ostiole, rostrate, long-beaked pycnidia in creamy-to-yellowish drops. Alpha conidia were hyaline, ellipsoidal, separated and averaged 6.3μm (SD ± 1.13) long × 1.9μm (SD ± 0.33) wide (n=50). Beta conidia were not seen. The morphological characteristics matched the previous description of Diaporthe longicolla (syn. Phomopsis longicolla) (Hobbs et al. 1985). Mycelial genomic DNA of the representative isolate LDSG3-2 was extracted as template. The internal transcribed spacer (ITS) , translation elongation factor 1α gene (TEF) and β-tubulin (TUB2) regions were amplified. These loci were amplified using primer pairs ITS4/ITS5 (White, et al. 1990), EF1-728F/EF1-986R (Carbone and Kohn 1999) and Bt2a/Bt2b (Glass and Donaldson 1995), respectively. A BLAST search of GenBank showed ITS (MN960195), TEF (MN974483) and TUB2 (MN974482) sequences of the isolate were 99%, 100%, and 99% homologous with D. longicolla strains DL11 (MF125048, 557/563 bp), D55 (MN584792, 347/347 bp) and DPC-HOH-32 (MK161506, 502/504 bp). Maximum likelihood trees based on concatenated nucleotide sequences of the three genes were constructed using MEGA 7.0, and bootstrap values indicated the isolate was D. longicolla (Fig. S1-D). Pathogenicity testing was performed using isolate LDSG3-2 by depositing 5µl droplets of a conidial suspension (1 × 106 ml-1) into 5 artificially wounded leaves (using a sterile needle) of 10 healthy 3-month-old K. pinnata plants. An equal number of artificially wounded control leaves were inoculated with sterile water to serve as a negative control. The test was conducted three times. Plants were kept at 25°C in 80% relative humidity and observed for symptoms. Two weeks after inoculation, no symptoms were observed on control plants (Fig. S1-C-1) and all inoculated plants showed symptoms (Fig. S1-C-2) similar to those observed in the field. The fungus was re-isolated from the infected tissues and showed the same cultural and morphological characteristics of the strain inoculated and could not be isolated from the controls fulfilling Koch’s postulates. To our knowledge, this is the first report of leaf spot on K. pinnata caused by D. longicolla in China. This disease is of concern since Phomopsis diseases are common in K. pinnata fields and can cause significant reduction in yield. References: White, T. J., et al. 1990. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA. DOI: 10.1016/0167-7799(90)90215-J Carbone, I., and Kohn, L. M. 1999. Mycologia. 91:553. DOI: 10.2307/3761358 Glass, N. L., and Donaldson, G. C. 1995. Appl. Environ. Microbiol. 61:1323. DOI: 10.1002/bit.260460112 Hobbs, T. W. et al. 1985. Mycologia. 77: 535. DOI: 10.2307/3793352

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First Report of Fusarium oxysporum Causing Wilt on Jade Plant (Crassula ovata) in Italy

Plant Disease ◽

10.1094/pdis-12-10-0897 ◽

2011 ◽

Vol 95 (9) ◽

pp. 1191-1191 ◽

Cited By ~ 1

Author(s):

A. Garibaldi ◽

D. Bertetti ◽

P. Pensa ◽

A. Poli ◽

M. L. Gullino

Keyword(s):

Fusarium Oxysporum ◽

Vascular System ◽

Apical Cell ◽

Aerial Mycelium ◽

Selective Medium ◽

Fluorescent Light ◽

Pathogenicity Test ◽

Conidial Suspension ◽

Single Spore ◽

First Report

During summer 2010, symptoms of a wilt disease were observed in a commercial farm in northern Italy on Crassula ovata (jade plant). First symptoms consisted of chlorosis and premature drop of still turgid leaves. As the disease progressed, leaves turned yellow and wilted before dropping off and the stem wilted, bent, and eventually rotted starting from the base. In some cases, the stem broke or the basal portion of the leaf rotted. Brown discolorations were observed in the vascular system. Of 10,000 plants, 65% (cv. Mini) and 5% of 600 plants (cv. Magical Tree) were affected. Premature dropping of leaves was more frequent on cv. Magical Tree. Using the Komada's Fusarium-selective medium, a fungus was consistently and readily isolated from symptomatic vascular tissues of plants belonging to both cultivars. Isolates obtained from both cultivars were purified, subcultured on potato dextrose agar (PDA), and single-spore cultures were obtained. On PDA, both isolates produced pale violet, abundant, aerial mycelium, felted in old cultures, with purple pigments in the agar. The isolates were grown on Spezieller Nährstoffarmer agar for characterization of macroconidia and microconidia (1). Both isolates produced sparse, 3 to 5 septate, nearly straight macroconidia measuring 30 to 56 × 3 to 5 (average 40 × 4) μm with a short apical cell and a foot-shaped basal cell. Sporodochia were not observed. Unicellular, oval-elliptical microconidia measuring 5 to 13 × 3 to 4 (average 8 × 3) μm were produced on short monophialides. Chlamydospores were abundant, single and sometime in pairs, terminal and intercalary, rough walled, and measured 6 to 9 μm. Such characteristics are typical of Fusarium oxysporum (3). The ITS region (internal transcribed spacer) of rDNA was amplified with primers ITS1/ITS4 (4) and sequenced. BLASTn analysis of an isolate from C. ovata cv. Mini (515 bp, Accession No. HQ682196) and C. ovata cv. Magical Tree (509 bp, Accession No. HQ682197) showed an E-value of 0.0 with F. oxysporum. For these sequences, pairwise alignment of EMBOSS (E.B.I. - The European Bioinformatics Institute) revealed identity and similarity of 99.0%. To confirm pathogenicity, tests were conducted on 5-month-old plants of cvs. Mini and Magical Tree. Plants (three per treatment) were inoculated by dipping roots in a 1 × 106 CFU/ml conidial suspension of the two isolates of F. oxysporum prepared from 10-day-old cultures grown on casein liquid medium (2), shaken (90 rpm) for 10 days at 24°C ± 1 (12-h fluorescent light, 12-h dark). Inoculated plants were transplanted into pots filled with steamed mix (sphagnum peat/perlite/pine bark/clay; 50:20:20:10) and maintained in a plant growth chamber at 25 ± 1°C under a regimen of 12 h per day of fluorescent light. Inoculated plants belonging to both cultivars showed typical first symptoms of Fusarium wilt after 13 days. In the following days, leaves dropped, stems wilted, and plants died. Noninoculated plants remained healthy. F. oxysporum was reisolated from inoculated plants. The pathogenicity test was conducted twice. This is, to our knowledge, the first report of F. oxysporum on C. ovata in Italy or worldwide. References: (1) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Professional, Ames, IA, 2006. (2) A. Minuto et al. Phytoparasitica 36:294, 2008. (3) B. A. Summerell et al. Plant Dis. 87:117, 2003. (4) T. J. White et al. PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, San Diego, 1990.

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