Branch Blight of Loquat Caused by Pestalotiopsis sydowiana in China

Loquat (Eriobotrya japonica) is a subtropical tree with sour and sweet fruit and its leaf derivatives are often used to treat cough and asthma with phlegm. In 2011, a severe stem and leaf disease, with 30∼60% incidence, was observed in a local loquat cultivar “Ninghaibai” in Ninghai, Zhejiang Province, China. Infected plants exhibited brown spots on the stem and leaves, which became blackish brown to grayish white with some little sporadic black acervuli (137.9 to 189.3 μm in diameter). Leaf symptoms showed large necrotic lesions (more than 20 mm) that frequently, but not exclusively, appeared along the leaf margin or dorsal surface. Necrotic spots were water-soaked, irregularly shaped, and bordered by a tan halo. Fungal isolates were obtained by placing sterilized symptomatic tissue onto acidified potato sucrose agar (PSA) medium and consistently yielded white fungal colonies that produced acervuli (385.6 to 485.4 μm in diameter) containing black, slimy spore masses at 25°C for 7 days. Conidia (20.9 to 29.8 × 7.1 to 14.5 μm) were 5-celled and spindle shaped, and basal cells of the conidia were hyaline, while the other three middle cells were heterochrome. The upper two middle cells were darker than the lower one. The apical cell typically had 2 to 3 apical appendages, was 12.0 to 33.1 μm in length, and the basal appendage was 3.3 to 7.8 μm long. Based on these morphological features, the fungus was identified as Pestalotiopsis sydowiana (2). This characterization was further confirmed by PCR amplification and sequencing of internal transcribed spacer (ITS) of the ribosomal DNA of fungus with primers ITS1: 5′-TCCGTAGGTGAACCTGCGG-3′ and ITS4: 5′-TCCTCCGCTTATTGATATGC-3′ (3). A representative sequence was deposited in NCBI GenBank (Accession No. JX478272). This sequence showed 99% homology with previously deposited sequences of P. sydowiana (HQ248207.1 and FJ478105.1) from Colombia and China. Pathogenicity tests were conducted by inoculating 1-year-old seedlings and detached stems from healthy field trees of cv. Ninghaibai loquat with sclertium (20 × 5 mm) or conidial suspension (106 conidia ml−1). Control seedlings were only encircled the PSA blocks or sprayed with water. Both inoculated and control plants were covered with plastic bags at about 80% relative humidity and kept in a greenhouse at 20 to 30°C for 48 h. Detached stems were incubated to prevent desiccation in a humid chamber at 100% relative humidity and 25°C for 10 days. Four days after inoculation, lesions appeared on sclertium-inoculated seedlings or detached stems, approximately 10 days earlier than those on conidial suspension-inoculated seedlings or detached stems. Symptom observed on artificially inoculated seedlings or detached stems were similar to that observed in naturally infected plants. The control plants remained healthy. P. sydowiana was reisolated from the lesions of the infected stems and leaves but not from control leaves. It had been reported that other species of Pestalotiopsis, such as P. eriobotrifolia and P. eriobotryae-japonica, can only infect leaves of loquat (1); however, to our knowledge, this is the first report that P. sydowiana cause branch blight disease on loquat in China. References: (1) G. G. Chen et al. Acta Agric. Univ. Zhejiangensis 16:153, 1990. (2) Q. X. Ge et al. Flora Fungorum Sinicorum: Pestalotiopsis, p. 185, Science Press, Beijing, 2009. (3) T. J. White, et al. PCR Protocols: A Guide to Methods and Applications, p. 315, Academic Press, San Diego, 1990.

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First Report of Gummosis Disease of Japanese Apricot Caused by Botryosphaeria dothidea in Taiwan

Plant Disease ◽

10.1094/pdis-05-10-0384 ◽

2011 ◽

Vol 95 (1) ◽

pp. 77-77

Author(s):

Y. Ko ◽

C. W. Liu ◽

S. S. Chen ◽

K. Y. Chiu ◽

Y. W. Sun ◽

...

Keyword(s):

Direct Sequencing ◽

Morphological Characteristics ◽

Pcr Amplification ◽

Conidial Suspension ◽

Botryosphaeria Dothidea ◽

Cortical Cells ◽

Japanese Apricot ◽

First Report ◽

Representative Sequence ◽

Plastic Bags

Japanese apricot (Prunus mume Sieb. et Zucc.) is an economically important fruit crop grown on more than 10,000 ha in Taiwan. During May 2008, twigs of Japanese apricot trees in the commercial farms of Renai Region (Nantou County) showed symptoms of gummosis disease, with 12 to 18% of the trees affected. The disease was more severe on trees weakened by drought stress. Limb and twig infections began around lenticles as small, sunken, discolored lesions at the margins of wounds. Following infection, cortical cells collapsed, bark became depressed, and blisters developed, which were often cracked with whitish gummy exudation. Necrotic areas were seen on the cortical tissues. Leaves showed yellowing and drooping. In winter months, numerous black pycnidia or perithecia formed on infected twigs. Single conidial isolates of the pathogen were obtained from diseased twigs on acidified potato dextrose agar (PDA) incubated at 25 ± 1°C for 3 days. On the basis of morphological characteristics, the fungus was identified as Botryosphaeria dothidea (3). Conidia (17 to 22.6 × 4.3 to 6.0 μm) were hyaline, unicellular, and spindle shaped. Asci (78 to 125 × 15 to 17 μm) were hyaline, bitunicate, clavate, and eight spored. Ascospores (18 to 22 × 7.0 to 8.2 μm) were hyaline and spindle shaped or fusoid. The pathogen identity was further confirmed by PCR amplification and sequencing of ribosomal DNA internal transcribed spacer from the fungus with the primers ITS5: 5′-GGAAGTAAAAGTCGTAACAAGG-3′ and ITS4: 5′-TCCTCCGCTTATTGATATGC-3′ (4), and a representative sequence was deposited in NCBI GenBank (Accession No. GU594225). The sequence showed 99 to 100% homology with previously characterized strains of B. dothidea (GenBank Accession Nos. EU441944, DQ177876, and AY786320). Pathogenicity tests were conducted with inoculum prepared by culturing the fungus on PDA under a continuous photoperiod of 128 ± 25 μE·m–2·s–1 at 25°C for 3 days. Shallow cuts (3 × 3 × 3 mm) were made on 12- to 15-month-old healthy twigs with a scalpel and inoculated with either a 5-mm mycelial disc or 0.5 ml of conidial suspension (105 conidia/ml) of the fungus. Two twigs on each of six trees were inoculated. Inoculated areas were covered with moist, sterile cotton and the entire twigs were enclosed in plastic bags. Twigs were inoculated with 5-mm PDA discs or sterile water for controls. The symptoms described above were observed on all inoculated twigs 14 days after inoculation, whereas control twigs remained healthy. Reisolation from the inoculated twigs consistently yielded B. dothidea. In Taiwan, B. dothidea has been reported as the causal agent of gummosis of peach (1) and fruit ring rot of pear (2); however, to our knowledge, this is the first report of B. dothidea causing gummosis on Japanese apricot. References: (1) Y. Ko et al. Plant Pathol. Bull. 1:70, 1992. (2) Y. Ko et al. Plant Prot. Bull. (Taiwan) 35:211, 1993. (3) B. Slippers et al. Mycologia 96:83, 2004. (4) T. J. White et al. In: Amplification and Direct Sequencing of Fungal Ribosomal RNA Genes for Phylogenetics. Academic Press. San Diego, CA, 1990.

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First Report of Pestalotiopsis maculans Causing Necrotic Leaf Spots in Nursery Plants of Arbutus unedo and Ceratonia siliqua in Spain

Plant Disease ◽

10.1094/pdis.2003.87.10.1263a ◽

2003 ◽

Vol 87 (10) ◽

pp. 1263-1263 ◽

Cited By ~ 5

Author(s):

A. Trapero ◽

M. A. Romero ◽

R. Varo ◽

M. E. Sánchez

Keyword(s):

Relative Humidity ◽

Growth Chamber ◽

Leaf Margin ◽

Conidial Suspension ◽

Ceratonia Siliqua ◽

Basal Cells ◽

First Report ◽

Arbutus Unedo ◽

Mean Width ◽

Detached Leaves

Forestation of agricultural lands has led to a great increase in the production of plants in forest nurseries in southern Spain. During a disease survey of several nurseries, performed in 1998 and 1999, a necrotic leaf spot was found causing defoliation in seedlings of two mediterranean forest species, the ericaceous shrub Arbutus unedo and the leguminous tree Ceratonia siliqua. The affected plants were 9 to 24 months old and growing in fertilized peat in containers. Symptoms on both species consisted of large necrotic lesions (up to 20 mm in diameter) that were located mainly, but not exclusively, along the leaf margin. On A. unedo, necrotic spots were bordered by a red halo, while on C. siliqua the halo was dark. The fungus consistently isolated from both hosts was identified as Pestalotiopsis maculans (Corda) Nag Raj (=Pestalotiopsis guepinii), based on morphological characters (1). Acervular conidiomata (up to 200 μm in diameter) developed on the necrotic lesions of leaves incubated at 100% relative humidity and 20 to 24°C, and in cultures on potato dextrose agar (PDA) at 20 to 24°C over the course of 7 days. All isolates had 5-celled smooth conidia. Apical and basal cells were hyaline, while the three median cells were brown; the upper two were darker than the lower one. Conidia were 22 to 30 μm (mean length) and 5 to 9 μm (mean width). There were typically three (range 1 to 4) apical appendages averaging 17 μm long. The average basal appendage was 6 μm long. One-year-old seedlings and detached leaves from healthy field trees of A. unedo and C. siliqua were sprayed to runoff with an aqueous conidial suspension (106 conidia ml-1) of two isolates of the fungus. All inoculated and control plants sprayed with water only were incubated in a growth chamber at 100% relative humidity and 20 to 24°C for 48 h and then in the same growth chamber at 50 to 80% relative humidity or in the greenhouse (10 to 30°C and 40 to 80% relative humidity) until symptoms developed. Detached leaves were incubated to prevent desiccation in a humid chamber at 100% relative humidity and 20 to 24°C for 2 months. After a period of 2 to 4 months, lesions developed on all inoculated leaves but not on noninoculated controls. Lesion morphology on both hosts was similar to that observed in naturally infected plants in the nurseries. P. maculans was reisolated from lesions of all infected leaves but not from control leaves. Although other species of Pestalotiopsis have been reported infecting leaves of C. siliqua (1), to our knowledge, this is the first report of a Pestalotiopsis sp. on A. unedo and of P. maculans on C. siliqua. Reference: (1) T. R. Nag Raj. Coelomycetous Anamorphs with Appendage-Bearing Conidia. Mycologue Publications, Waterloo, Ontario, Canada, 1993.

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First Report of Leaf Blight of Japanese Yew Caused by Pestalotiopsis microspora in Korea

Plant Disease ◽

10.1094/pdis-08-13-0821-pdn ◽

2014 ◽

Vol 98 (5) ◽

pp. 691-691 ◽

Cited By ~ 2

Author(s):

Y. H. Jeon ◽

W. Cheon

Keyword(s):

Dna Sequences ◽

Apical Cell ◽

Morphological Characteristics ◽

Leaf Blight ◽

Economic Damage ◽

Leaf Margin ◽

Conidial Suspension ◽

First Report ◽

Pestalotiopsis Microspora ◽

Large Trees

Worldwide, Japanese yew (Taxus cuspidata Sieb. & Zucc.) is a popular garden tree, with large trees also being used for timber. In July 2012, leaf blight was observed on 10% of Japanese yew seedling leaves planted in a 500-m2 field in Andong, Gyeongsangbuk-do Province, South Korea. Typical symptoms included small, brown lesions that were first visible on the leaf margin, which enlarged and coalesced into the leaf becoming brown and blighted. To isolate potential pathogens from infected leaves, small sections of leaf tissue (5 to 10 mm2) were excised from lesion margins. Eight fungi were isolated from eight symptomatic trees, respectively. These fungi were hyphal tipped twice and transferred to potato dextrose agar (PDA) plates for incubation at 25°C. After 7 days, the fungi produced circular mats of white aerial mycelia. After 12 days, black acervuli containing slimy spore masses formed over the mycelial mats. Two representative isolates were further characterized. Their conidia were straight or slightly curved, fusiform to clavate, five-celled with constrictions at the septa, and 17.4 to 28.5 × 5.8 to 7.1 μm. Two to four 19.8- to 30.7-μm-long hyaline filamentous appendages (mostly three appendages) were attached to each apical cell, whereas one 3.7- to 7.1-μm-long hyaline appendage was attached to each basal cell, matching the description for Pestalotiopsis microspora (2). The pathogenicity of the two isolates was tested using 2-year-old plants (T. cuspidata var. nana Rehder; three plants per isolate) in 30-cm-diameter pots filled with soil under greenhouse conditions. The plants were inoculated by spraying the leaves with an atomizer with a conidial suspension (105 conidia/ml; ~50 ml on each plant) cultured for 10 days on PDA. As a control, three plants were inoculated with sterilized water. The plants were covered with plastic bags for 72 h to maintain high relative humidity (24 to 28°C). At 20 days after inoculation, small dark lesions enlarged into brown blight similar to that observed on naturally infected leaves. P. microspora was isolated from all inoculated plants, but not the controls. The fungus was confirmed by molecular analysis of the 5.8S subunit and flanking internal transcribed spaces (ITS1 and ITS2) of rDNA amplified from DNA extracted from single-spore cultures, and amplified with the ITS1/ITS4 primers and sequenced as previously described (4). Sequences were compared with other DNA sequences in GenBank using a BLASTN search. The P. microspora isolates were 99% homologous to other P. microspora (DQ456865, EU279435, FJ459951, and FJ459950). The morphological characteristics, pathogenicity, and molecular data assimilated in this study corresponded with the fungus P. microspora (2). This fungus has been previously reported as the causal agent of scab disease of Psidium guajava in Hawaii, the decline of Torreya taxifolia in Florida, and the leaf blight of Reineckea carnea in China (1,3). Therefore, this study presents the first report of P. microspora as a pathogen on T. cuspidata in Korea. The degree of pathogenicity of P. microspora to the Korean garden evergreen T. cuspidata requires quantification to determine its potential economic damage and to establish effective management practices. References: (1) D. F. Farr and A. Y. Rossman, Fungal Databases, Syst. Mycol. Microbiol. Lab. Retrieved from http://nt.ars-grin.gov/fungaldatabases/ (2) L. M. Keith et al. Plant Dis. 90:16, 2006. (3) S. S. N. Maharachchikumbura. Fungal Diversity 50:167, 2011. (4) T. J. White et al. PCR Protocols. Academic Press, San Diego, CA, 1990.

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First Report of Mature Citrus Trees Being Affected by Fusarium Wilt in Tunisia

Plant Disease ◽

10.1094/pdis-12-12-1134-pdn ◽

2014 ◽

Vol 98 (4) ◽

pp. 566-566 ◽

Cited By ~ 5

Author(s):

I. Hannachi ◽

S. Rezgui ◽

M. Cherif

Keyword(s):

Fusarium Wilt ◽

Vascular Tissue ◽

Disease Incidence ◽

Pcr Amplification ◽

Its Region ◽

Conidial Suspension ◽

Sour Orange ◽

Basal Cells ◽

First Report ◽

Citrus Trees

Citrus is an important crop in Tunisia and over 98% of trees of all varieties are grafted on sour orange rootstock. Since September 2010, unusual wilt symptoms have been observed in Takilsa, Bni-Khaled, and Manzel Bouzalfa fields that eventually caused tree death. The disease was observed on 10- to 25-year-old trees of sweet orange (Citrus sinensis) ‘Washington Navel’ and on ‘Clementine’ tangerines (C. tangerina) ‘Cassar,’ ‘Hernandina,’ and ‘MA3,’ all grafted on sour orange (C. aurantum) ‘Bigarade Gou Tou.’ The most conspicuous symptoms were wilting of sections of the canopy, chlorosis and epinasty of young leaves, and discoloration of vascular tissue. No root rot was observed. The problem was widespread with a disease incidence of 45 to 67%. Similar symptoms were described by Timmer et al. (2) on Mexican lime (C. aurantiifolia) nursery plants and some other species of citrus. Three representative isolates of Fusarium oxysporum Schlechtend.:Fr. from crown were single-spored and identified by the production of characteristic, three- to five-celled, sickle-shaped macroconidia with foot-shaped basal cells, ellipsoid microconidia borne in false heads on short monophialides, and chlamydospores in culture (1). The internal transcribed spacer (ITS) region of rDNA and the elongation factor (TEF 1-α) were amplified with primers ITS1/ITS4 and (TEF1/TEF2), respectively. GenBank accessions of ITS region are KC282838, KC282839, and KC282840, for TEF 1-α region are KF531633, KF537336, and KF537337, showed 99% homology with isolates of F. oxysporum in Fusarium-ID data. Pathogenicity tests were conducted on 7-month-old seedlings of sour orange using 10 plants for each of the three isolates. Prior to inoculation, roots were scraping with a sterile scalpel and plants were dipped in a conidial suspension of F. oxysporum (106 conidia ml–1) for 10 min. Each seedling was planted in a separate pot containing 0.7 liter of sterile soil. Non-inoculated plants with scraped roots dipped in sterile distilled water served as controls. Plants were irrigated and placed in a greenhouse at 24 ± 2°C and 12-h photoperiod. One month after inoculation, leaf chlorosis was observed and 2 months later, 90% of inoculated plants presented a severe wilt. Symptoms on infected plants were similar to those observed in the field. F. oxysporum was successfully re-isolated from the stems, thereby completing Koch's postulates. Genomic DNA was isolated from the re-isolations and PCR amplification of the ITS region was performed with the same primers. There was 100% nucleotide identity with sequences of the original isolates. To our knowledge, this is the first report of fusarium wilt of citrus trees in Tunisia. The pathogen may represent a new form species because previously, the disease was only reported from lime and lemon. References: (1) J. F. Leslie and B. A. Summerell. Page 256 in: The Fusarium Laboratory Manual. Blackwell Publishing Professional, Hoboken, NJ, 2006. (2) L. W. Timmer et al. Phytopathology 72:698, 1982.

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Screening of Durum Wheat (Triticum turgidum L. subsp. durum (Desf.) Husn.) Italian Cultivars for Susceptibility to Fusarium Head Blight Incited by Fusarium graminearum

Plants ◽

10.3390/plants10010068 ◽

2020 ◽

Vol 10 (1) ◽

pp. 68

Author(s):

Gaetano Bentivenga ◽

Alfio Spina ◽

Karim Ammar ◽

Maria Allegra ◽

Santa Olga Cacciola

Keyword(s):

Durum Wheat ◽

Fusarium Head Blight ◽

Fusarium Graminearum ◽

Triticum Turgidum ◽

Disease Incidence ◽

Morphological Characteristics ◽

Pcr Amplification ◽

Conidial Suspension ◽

Head Blight ◽

Italian Origin

In 2009, a set of 35 cultivars of durum wheat (Triticum turgidum L. subsp. durum (Desf.) Husn.) of Italian origin was screened for fusarium head blight (FHB) susceptibility at CIMMYT (Mexico) and in the 2019–20 cropping season, 16 of these cultivars, which had been included in the Italian National Plant Variety Register, were tested again in southern and northern Italy. Wheat cultivars were artificially inoculated during anthesis with a conidial suspension of Fusarium graminearum sensu lato using a standard spray inoculation method. Inoculum was a mixture of mono-conidial isolates sourced in the same areas where the trials were performed. Isolates had been characterized on the basis of morphological characteristics and by DNA PCR amplification using a specific primer set and then selected for their virulence and ability to produce mycotoxins. The susceptibility to FHB was rated on the basis of the disease severity, disease incidence and FHB index. Almost all of the tested cultivars were susceptible or very susceptible to FHB with the only exception of “Duprì”, “Tiziana” and “Dylan” which proved to be moderately susceptible. The susceptibility to FHB was inversely correlated with the plant height and flowering biology, the tall and the late heading cultivars being less susceptible.

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Veronica sibirica Leaf Spots Caused by Phacellium veronicae, a New Disease in China

Plant Disease ◽

10.1094/pdis-11-12-1052-pdn ◽

2013 ◽

Vol 97 (12) ◽

pp. 1662-1662 ◽

Cited By ~ 1

Author(s):

Q. R. Bai ◽

S. Han ◽

Y. Y. Xie ◽

J. Gao ◽

Y. Li

Keyword(s):

Morphological Characteristics ◽

Its Region ◽

Control Treatment ◽

Perennial Herb ◽

Conidial Suspension ◽

New Disease ◽

Medical College ◽

Leaf Spots ◽

Plastic Bags ◽

Insect Bites

Veronica sibirica (Veronicastrum sibiricum) is an erect perennial herb, an ornamental, and a traditional Chinese medicine plant distributed mostly in northeastern, northern, and northwestern China. It has dehumidifying and detoxifying properties, and is mainly used for the treatment of cold, sore throat, mumps, rheumatism, and insect bites (4). In June 2008 through 2012, leaf spots of V. sibirica were observed in the Medicinal Herb Garden of Jilin Agricultural University (43°48′N, 125°23′E) and the medicinal plantations of Antu County (43°6′N, 128°53′E), Jilin Province. Leaf spots were amphigenous, subcircular, angular-irregular, brown, and 1 to 10 mm in diameter; they occasionally merged into a larger spot with an indefinite margin or with a pale center and dark border. Pale conidiomata were hypophyllous and scattered on the spots. The conidiophores were 100 to 400 μm high and clustered together to form synnemata 20 to 50 μm in diameter, which splayed out apically and formed loose to dense capitula. Conidiophores occasionally emerged through the stomata individually and produced conidia on the surface of the infected leaves. The conidiogenous cell terminal was geniculate-sinuous with somewhat thickened and darkened conidial scars. Conidia were solitary or catenulate, ellipsoid-ovoid or subcylindric-fusiform, hyaline and spinulose, 4.01 to 7.18 × 11.16 to 20.62 μm with obtuse to somewhat attenuated ends, and slightly thickened, darkened hila. Six isolates were obtained from necrotic tissue of leaf spots and cultured on potato dextrose agar at 25°C. After incubation for 14 days, colony surfaces were white to pinkish. The colony diameter increased by 12 mm after 21 days' incubation. Hyphae were hyaline, septate, and branched. Conidiophores grew individually or fascicularly. The symptoms and morphological characteristics were consistent with previous descriptions (1,2), and the fungus was identified as Phacellium veronicae (Pass.) (U. Braun 1990). The internal transcribed spacer (ITS) region of the nuclear rDNA was amplified using primers ITS4/ITS5 (3). The ITS was identical among all six isolates (HE995799) and 98% identical to that of P. veronicae (JQ920427, HQ690097). Pathogenicity was confirmed by spraying five 1-year-old V. sibirica seedlings with a conidial suspension (106 conidia/ml) of each isolate and five seedlings with sterile water as a control treatment. Plants were grown in the greenhouse at 20 to 25°C and were covered with plastic bags to maintain humidity on the foliage for 72 h. After 15 days, the same symptoms appeared on the leaves as described earlier for the field-grown plants; the control plants remained healthy. The same fungus was reisolated from the leaf spots of inoculated plants. Currently, the economic importance of this disease is limited, but it may become a more significant problem, as the cultivated area of V. sibirica is increasing. To our knowledge, although P. veronicae was recorded on the other species of Veronica (V. austriaca, V. chamaedrys, V. grandis, V. longifolia, V. paniculata, and V. spicata ssp. incana) in Europe (Germany, Denmark, Ireland, Romania) and V. wormskjoldii in North America (Canada) (1), this is the first report of V. sibirica leaf spots caused by P. veronicae in the world, and it is a new disease in China. References: (1) U. Braun. A monograph of Cercosporella, Ramularia and allied genera (phytopathogenic Hyphomycetes) 2, IHW-Verlag, Germany, 1998. (2) U. Braun. Nova Hedwigia 50:499, 1990. (3) D. E. L. Cooke et al. Mycol. Res. 101:667, 1997. (4) Jiangsu New Medical College. Dictionary of Chinese Materia Medica. Shanghai: Shanghai Scientific and Technical Publishers, China, 1977.

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Production, Survival, and Evaluation of Solid-Substrate Inocula of Penicillium oxalicum, a Biocontrol Agent Against Fusarium Wilt of Tomato

Phytopathology ◽

10.1094/phyto.2002.92.8.863 ◽

2002 ◽

Vol 92 (8) ◽

pp. 863-869 ◽

Cited By ~ 27

Author(s):

I. Larena ◽

P. Melgarejo ◽

A. De Cal

Keyword(s):

Fusarium Wilt ◽

Biocontrol Agent ◽

Solid Substrate ◽

Penicillium Oxalicum ◽

Conidial Suspension ◽

Conidial Production ◽

Plastic Bags ◽

General Decline ◽

Leguminous Seeds ◽

Solid Fermentation

Production of conidia of Penicillium oxalicum (ATCC number pending), a biocontrol agent of Fusarium oxysporum f. sp. lycopersici, was tested in liquid and solid fermentation. P. oxalicum produced 250-fold more conidia in solid than in liquid fermentation at 30 days after inoculation of substrate. Solid fermentation was carried out in plastic bags (600 cm3) especially designed for solid fermentation (VALMIC) containing 50 g of peat/vermiculite (PV) (1:1, wt/wt) with 40% moisture, sealed, sterilized, and then inoculated with 1 ml of a conidial suspension of P. oxalicum (105 conidia g-1 dry substrate), sealed again, and incubated in darkness at 20 to 25°C for 30 days. Addition of amendments to PV in a proportion of 0.5 (wt/wt) significantly increased conidial production of P. oxalicum. The best production was obtained on PV plus meal of cereal grains (barley) or leguminous seeds (lentil) (100-fold higher). Conidial production obtained after 5 days of inoculation was similar to that obtained at 30 days. However, viability of conidia produced in PV plus lentil meal was 35% higher than that of conidia produced in PV plus barley meal. Changes in proportions (1:1:0.5, wt/wt/wt; 1:1:1, wt/wt/wt; 1:0.5:0.5, wt/wt/wt; 1:1:0.5, vol/vol/vol) of components of the substrate (peat/vermiculite/lentil meal) did not enhance production or viability of conidia. Optimal initial moisture in the substrate was 30 to 40%. At lower moistures, significant reductions of production of conidia were observed, particularly at 10%. There was a general decline in the number of conidia in bags with time of storage at -80, -20, 4, and 25°C, or at room temperature (range from 30 to 15°C), with the highest decline occurring from 60 to 180 days. Conidial viability also was reduced with time, except for conidia stored at -20°C. Fresh conidia produced in solid fermentation system or those conidia stored at -20°C for 180 days reduced Fusarium wilt of tomato by 49 and 61%, respectively.

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

Plant Disease ◽

10.1094/pdis-09-21-2001-pdn ◽

2021 ◽

Author(s):

Yanxiang Qi ◽

Yanping Fu ◽

Jun Peng ◽

Fanyun Zeng ◽

Yanwei Wang ◽

...

Keyword(s):

Leaf Spot ◽

Disease Incidence ◽

Morphological Characteristics ◽

Universal Primers ◽

Leaf Spot Disease ◽

Leaf Margin ◽

Conidial Suspension ◽

Tropical Fruit ◽

First Report ◽

Spot Disease

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

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First Report of the Pathogenicity of Colletotrichum gloeosporioides on Invasive Ferns, Lygodium microphyllum and L. japonicum, in Florida

Plant Disease ◽

10.1094/pdis.2003.87.1.101b ◽

2003 ◽

Vol 87 (1) ◽

pp. 101-101 ◽

Cited By ~ 2

Author(s):

K. A. Jones ◽

M. B. Rayamajhi ◽

P. D. Pratt ◽

T. K. Van

Keyword(s):

Colletotrichum Gloeosporioides ◽

First Record ◽

South Florida ◽

Growth Chamber ◽

Fluorescent Light ◽

Distilled Water ◽

Conidial Suspension ◽

Potted Plants ◽

Plastic Bags ◽

Fern Species

Lygodium microphyllum (Cav.) R.Br. (Old World climbing fern) and L. japonicum (Thunb.) Sw. (Japanese climbing fern), in the family Schizaeaceae, are among the most invasive weeds in Florida (1). L. microphyllum invades fresh water and moist habitats in south Florida, while L. japonicum has spread in relatively well-drained habitats from Texas to North Carolina and central Florida. Some potted plants of both Lygodium spp. grown in shadehouse as well as in full sunlight developed discolored spots on pinnules (foliage), which coalesced and resulted in browning and dieback of severely infected vines. Symptomatic foliage obtained from these plants was surface-sterilized by immersing in a 15% solution of commercial bleach for 90 s, followed by a series of four rinses with sterile deionized distilled water. Disks (4 mm in diameter) of pinnules were cut from the junction of discolored and healthy tissues and placed on potato dextrose agar (PDA). A fungus, Colletotrichum gloeosporioides (Penz.) Penz. & Sacc. was consistently isolated from these disks. Fungal colonies produced abundant conidia on PDA. Conidia were hyaline, straight, cylindrical, averaging 14.7 μm (range 12.5 to 17.5 μm) × 5.0 μm (range 3.8 to 7.5 μm), and similar to those described for C. gloeosporioides (2). To confirm the pathogenicity of C. gloeosporioides on L. microphyllum and L. japonicum, Koch's postulates were performed. A fungal isolate was grown on PDA for 3 weeks, after which 10 ml of sterile deionized distilled water was added to the culture and agitated to dislodge conidia. The conidial suspension was strained through three layers of cheesecloth to remove hyphal fragments, and its concentration was adjusted to 1.7 × 106 conidia/ml. Foliage of healthy L. microphyllum and L. japonicum plants grown in 500-ml containers was sprayed with the conidial suspension until runoff. Plants were covered with plastic bags whose inner sides were misted with water to maintain high humidity and placed in a growth chamber under 12 h of fluorescent light per day. Temperature and relative humidity in the chamber ranged from 26 to 29°C and 44 to 73%, respectively. Plastic bags were removed after 3 days, and plants were further incubated for 3 weeks in the same growth chamber. Control plants were sprayed with sterile water, covered with plastic bags, and exposed to the same temperature, light, and humidity regime as those of the fungus-inoculated plants. Small, discolored foliar spots appeared 3 days after fungus inoculation. These spots were similar to those observed on pinnules of potted plants that originated from shadehouse and outdoor environments. Within 3 weeks after inoculation, the foliage of L. japonicum developed abundant discolored spots that led to edge browning and wilting of the pinnules. L. microphyllum had similar but more severe symptoms, with plants suffering as much as 50% dieback. C. gloeosporioides was consistently reisolated from the symptomatic tissues of both fern species. No symptoms appeared on the water-inoculated plants. To our knowledge, this is the first record of C. gloeosporioides pathogenicity on L. microphyllum and L. japonicum. References: (1) R. W. Pemberton and A. P. Ferriter. Am. Fern J. 88:165, 1998. (2) B. C. Sutton. Colletotrichum: Biology, Pathology and Control. CAB International, Wallingford, Oxon, UK, 1992.

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First Report of Leaf Spot Caused by Botrytis cinerea on Cardamine hupingshanensis in China

Plant Disease ◽

10.1094/pdis-04-21-0698-pdn ◽

2021 ◽

Author(s):

Jun Guo ◽

Jin Chen ◽

Zhao Hu ◽

Jie Zhong ◽

Jun Zi Zhu

Keyword(s):

Botrytis Cinerea ◽

Morphological Characteristics ◽

Gray Mold ◽

Hunan Province ◽

Conidial Suspension ◽

Heat Shock Protein 60 ◽

Basal Cells ◽

Single Spore ◽

Internal Transcribed Spacer Region ◽

First Report

Cardamine hupingshanensis is a selenium (Se) and cadmium (Cd) hyperaccumulator plant distributed in wetlands along the Wuling Mountains of China (Zhou et al. 2018). In March of 2020, a disease with symptoms similar to gray mold was observed on leaves of C. hupingshanensis in a nursery located in Changsha, Hunan Province, China. Almost 40% of the C. hupingshanensis (200 plants) were infected. Initially, small spots were scattered across the leaf surface or margin. As disease progressed, small spots enlarged to dark brown lesions, with green-gray, conidia containing mold layer under humid conditions. Small leaf pieces were cut from the lesion margins and were sterilized with 70% ethanol for 10 s, 2% NaOCl for 2 min, rinsed with sterilized distilled water for three times, and then placed on potato dextrose agar (PDA) medium at 22°C in the dark. Seven similar colonies were consistently isolated from seven samples and further purified by single-spore isolation. Strains cultured on PDA were initially white, forming gray-white aerial mycelia, then turned gray and produced sclerotia after incubation for 2 weeks, which were brown to blackish, irregular, 0.8 to 3.0 × 1.2 to 3.5 mm (n=50). Conidia were unicellular, globose or oval, colourless, 7.5 to 12.0 × 5.5 to 8.3 μm (n=50). Conidiophores arose singly or in group, straight or flexuous, septate, brownish to light brown, with enlarged basal cells, 12.5 to 22.1 × 120.7 to 310.3 μm. Based on their morphological characteristics in culture, the isolates were putatively identified as Botrytis cinerea (Ellis 1971). Genomic DNA of four representative isolates, HNSMJ-1 to HNSMJ-4, were extracted by CTAB method. The internal transcribed spacer region (ITS), glyceraldehyde-3-phosphate dehydrogenase gene (G3PDH), heat-shock protein 60 gene (HSP60), ATP-dependent RNA helicaseDBP7 gene (MS547) and DNA-dependent RNA polymerase subunit II gene (RPB2) were amplified and sequenced using the primers described previously (Aktaruzzaman et al. 2018) (MW820311, MW831620, MW831628, MW831623 and MW831629 for HNSMJ-1; MW314722, MW316616, MW316617, MW316618 and MW316619 for HNSMJ-2; MW820519, MW831621, MW831627, MW831624 and MW831631 for HNSMJ-3; MW820601, MW831622, MW831626, MW831625 and MW831630 for HNSMJ-4). BLAST searches showed 99.43 to 99.90% identity to the corresponding sequences of B. cinerea strains, such as HJ-5 (MF426032.1, MN448500.1, MK791187.1, MH727700.1 and KX867998.1). A combined phylogenetic tree using the ITS, G3PDH, HSP60 and RPB2 sequences was constructed by neighbor-joining method in MEGA 6. It revealed that HNSMJ-1 to HNSMJ-4 clustered in the B. cinerea clade. Pathogenicity tests were performed on healthy pot-grown C. hupingshanensis plants. Leaves were surface-sterilized and sprayed with conidial suspension (106 conidia/ mL), with sterile water served as controls. All plants were kept in growth chamber with 85% humidity at 25℃ following a 16 h day-8 h night cycle. The experiment was repeated twice, with each three replications. After 4 to 7 days, symptoms similar to those observed in the field developed on the inoculated leaves, whereas controls remained healthy. The pathogen was reisolated from symptomatic tissues and identified using molecular methods, confirming Koch’s postulates. B. cinerea has already been reported from China on C. lyrate (Zhang 2006), a different species of C. hupingshanensis. To the best of our knowledge, this is the first report of B. cinerea causing gray mold on C. hupingshanensis in China and worldwide. Based on the widespread damage in the nursery, appropriate control strategies should be adopted. This study provides a basis for studying the epidemic and management of the disease.

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