Sterile White Basidiomycete Fungus Marasmius graminum: A New Pathogen Causing Seedling Blight in Rice

In April 2018, damping-off of rice (Oryza sativa L.) seedlings at the 2-to-3-leaf stage was observed in three fields in the counties of Wharton and Matagorda of Texas and Jefferson-Davis Parish of Louisiana. All affected areas were 1 ha or greater, with 10 to 20% of the seedlings showing the symptoms. Infected seedlings showed dark-brown necrotic lesions on the roots and/or mesocotyls where white superficial mycelium was usually present. Symptomatic tissues excised from 10 diseased seedlings of each field were surface sterilized with 1% NaOCl, double rinsed in sterilized distilled water, and plated on potato dextrose agar (PDA). The plates were incubated at 25°C with a 12-h photoperiod in a growth chamber. After 48 h, hyphal tips of fungal colonies were transferred onto PDA and 12 isolates were obtained. Clamp connections and dolipore septa were observed in young hyphae, indicating that these isolates were a basidiomycete fungus. Young hyphal cells were binucleate based on safranin O stain (Bandoni 1979). No fruiting bodies or sclerotia produced on PDA after one month of incubation. Based on these morphological characteristics, these isolates were identified as belonging to sterile white basidiomycetes (SWB) (Howard et al. 1977). To further identify the isolates into the species level, the internal transcribed spacer (ITS) region of ribosomal DNA (rDNA) of a representative isolate was sequenced with primer ITS1 and ITS4 (Vinnere et al. 2005). The ITS sequence (GenBank acc. no. MT524457) had more than 97% sequence similarity with known Marasmius graminum strains from Denmark (JN943595) (Schoch et al. 2012) and Sweden (MH857692) (Vu et al. 2019). Pathogenicity was tested with three representative isolates in a growth chamber using a modified method (Carling and Leiner 1990). Pots (6.5 cm in diameter x 7.5 cm height) were filled with 100 g of sterilized sand and watered to field capacity. Five PDA plugs (4 mm in diameter) from 5-day-old growing culture were placed on the sand surface of each pot. Pots inoculated with PDA plugs without fungus served as the controls. Five seeds of rice cv. Presidio were planted into each pot and covered with 10 g of sterilized sand. Pots were maintained at 25±2°C in a growth chamber with a 12-h photoperiod for 14 days. There were four replicated pots for each treatment and the experiment repeated twice. After 2 weeks, severe damping-off and associated symptoms similar to those observed in the field appeared in the inoculated pots. No symptoms developed in the control pots. The same fungus was consistently re-isolated from infected plants. Based on morphological characteristics and rDNA-ITS sequencing, these isolates were identified as M. graminum. The SWB fungus was first reported as a causal agent of stem rot of snap bean in Florida (Howard et al. 1977) and Nebraska (Harveson 2002), root or hypocotyl rot of corn, snap bean, squash and peanut in Georgia (Sumner et al. 1979; Bell and Sumner 1984), and crown rot of pigeon pea (Cajanus cajan) in Puerto Rico (Kaiser et al. 1987). Later, the SWB strain (ATCC 28344) causing stem rot of snap bean in Florida was further identified as M. graminum based on nuclear large subunit rRNA gene (Vinnere et al. 2005). Comparing the ITS region of this isolate (AY445120) with our isolate revealed a 99% similarity. To our knowledge, this is the first report that the SWB fungus M. graminum causes seedling blight in rice. Identification of this new disease will help to develop management strategies for control of stand loss in rice.

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First Report of Pythium schmitthenneri Causing Maize Seedling Blight in Iowa

Plant Disease ◽

10.1094/pdis-08-13-0892-pdn ◽

2014 ◽

Vol 98 (7) ◽

pp. 994-994

Author(s):

R. L. Matthiesen ◽

A. A. Ahmad ◽

M. L. Ellis ◽

A. E. Robertson

Keyword(s):

Morphological Characteristics ◽

Its Region ◽

Corn Meal ◽

Maize Seedlings ◽

Damping Off ◽

Seedling Blight ◽

Root Tips ◽

First Report ◽

South Central ◽

Paper Towels

In spring 2012, maize farmers in southeast and south central Iowa reported stand losses due to pre- and post-emergence damping-off, and many of the fields had to be replanted. Symptoms of the disease included rotted seed, or brown, rotted, water-soaked mesocotyls and root tips. Maize seedlings with severe root and mesocotyl symptoms were yellow and wilted, stunted, or dead. The disease occurred approximately 2 weeks after cool, wet conditions. Symptomatic mesocotyls and roots were washed for 30 min, rinsed with sterile distilled water, and blotted dry on sterile paper towels. Isolation of the pathogen was performed by aseptically cutting 2- to 3-mm sections of tissue from the edge of a lesion, placing the segments under corn meal agar (CMA) containing pimaricin, ampicillin, rifampicin, and pentachloronitrobenzene (PARP), and incubating at 22°C in the dark. Colonies that developed were putatively identified as Pythium species based on morphological characteristics and cultural features when compared to published descriptions (2,3). Characteristics of isolate IAC12F21-3 included spherical and smooth-walled oogonia 18 to 26 μm in diameter, monoclinous or usually diclinous antheridia 10 to 22 μm long and 5 to 10 μm wide with one or occasionally two per oogonium, and plerotic oospores 15 to 25 μm in diameter. Sporangia were globose to ellipsoidal, 22 to 41 μm in diameter, and zoospores were 7 to 10 μm long. Primers ITS1 and ITS4 were used to amplify the ITS region within clade E1 of 88 isolates. The resultant amplicons were sequenced and a BLAST search in GenBank confirmed isolate IAC12F21-3 as Pythium schmitthenneri based on 100% similarity with GenBank accession numbers JF836869 and JF836870. Pathogenicity testing was conducted using seed and seedling assays (1,4). Koch's postulates was performed by sampling pieces of symptomatic mesocotyl and root tissue from the inoculated pots, placing segments under CMA + PARP, and incubating at 22°C. Symptoms were similar to those observed in the field and P. schmitthenneri was re-isolated successfully. Non-inoculated control plants showed no symptoms. This is the first report of P. schmitthenneri causing seedling blight on maize in Iowa. Previously, P. schmitthenneri was reported as a pathogen on maize in Ohio (2). References: (1) K. Broders et al. Plant Dis. 91:727, 2007. (2) M. Ellis et al. Mycologia, 104:477, 2012. (3) J. Middleton. Memoirs of the Torrey Botanical Club 20:171, 1943. (4) A. Rojas et al. Phytopathology, 102(Suppl):S5.8, 2012.

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First Report of Basal Stem Rot and Foliar Blight Caused by Pythium sylvaticum on Miscanthus sinensis in Illinois

Plant Disease ◽

10.1094/pdis-08-10-0592 ◽

2011 ◽

Vol 95 (5) ◽

pp. 616-616 ◽

Cited By ~ 9

Author(s):

M. O. Ahonsi ◽

B. O. Agindotan ◽

M. E. Gray ◽

C. A. Bradley

Keyword(s):

Perennial Grass ◽

Morphological Characteristics ◽

Its Region ◽

The United States ◽

Stem Rot ◽

Miscanthus Sinensis ◽

Growth Media ◽

Bioenergy Feedstock ◽

First Report ◽

Pythium Sylvaticum

Miscanthus sinensis Anderss., a perennial grass, is native to eastern Asia. It has been widely grown as an ornamental in temperate regions of the world, including the United States, and recently has become an important component of public and private sector bioenergy feedstock Miscanthus selection programs. In August 2008, stem rot and blight was observed on M. sinensis plants in two irregular patches, ~2 to 2.5 × 1 to 1.5 m each in a trial plot that was preceded by corn, at the University of Illinois Energy Farm near Urbana, IL. At the time of the observation, most plants were dead and the wilted tillers had black, soft rotted basal stems. A few plants were stunted and the crowns of the tillers had black-to-brown soft rot. Some tillers' leaves were dead and others had turned light brown. Sample tissue fragments were surface disinfested in 0.5% NaOCl and plated on 1% water agar (WA). After 3 days of incubation in the dark at 23°C, colonies were transferred to corn meal agar (CMA), potato dextrose agar (PDA), or 10% V8 juice agar and incubated at 23°C under continuous white light for up to 2 weeks. Morphological characteristics of the isolates correspond to those originally described for Pythium sylvaticum W.A. Campb. & J.W. Hendrix (1). The mycelia grew and covered the 10-cm-diameter plates within 5 days. On PDA, the culture was a creamy white mycelial mat of coenocytic hyphae. The isolates produced only globose, terminal or intercalary hyphal swellings ranging from 28 to 48 μm in diameter, but no oogonia were produced on any of the three growth media. No zoospores were produced when agar blocks bearing mycelium were flooded with distilled water or 1% soil water. Sequence analysis was performed with the internal transcribed spacer (ITS) region of the rDNA amplified with primer pair ITS1/ITS4 (3) and the mitochondrially encoded cytochrome c oxydase subunit II (cox II) gene using primers FM58/FM66 (2). The resulting 871-bp ITS nucleotide sequence (Accession No. HM991706) was identical among all three isolates analyzed and 99% identical (100% coverage) to ITS sequences of multiple isolates of P. sylvaticum in GenBank. Likewise, the 544-bp cox II sequence (Accession No. HQ454429) was 99% identical (97% coverage) to cox II sequences of multiple isolates of P. sylvaticum. Six pots of M. sinensis seedlings were inoculated by placing two CMA plugs of a 2-week-old culture of isolate F71 at the crown. The control pots were mock inoculated with sterile CMA plugs. The plants were incubated at ~90% relative humidity (RH) and 25°C day and 22°C night for 3 days, and thereafter left on the greenhouse bench at ~65% RH with alternating 9 h of darkness and 15 h of light. Three weeks after inoculation, two of the inoculated seedlings wilted, others were stunted with leaves wilting from the tip downwards and the stems rotting from the crown upward. A thick mat of mycelia was seen on the rotted basal stems. No symptoms were observed in the control. P. sylvaticum was reisolated from both the rotted basal stems and the wilted foliage. To our knowledge, this is the first report of P. sylvaticum on M. sinensis. Infestation of farm soils with P. sylvaticum could limit M. sinensis biomass production significantly by limiting seedling establishment. References: (1) W. A. Campbell and F. F. Hendrix. Mycologia 59:274, 1967. (2) F. M. Martin. Mycologia 92:711, 2000. (3) T. J. White et al. Page 38 in: PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, San Diego, 1990.

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First Report of Soybean Neocosmospora Stem Rot Caused by Neocosmospora vasinfecta var. vasinfecta in China

Plant Disease ◽

10.1094/pdis-04-11-0287 ◽

2011 ◽

Vol 95 (8) ◽

pp. 1031-1031

Author(s):

Y. Gai ◽

R. Pan ◽

D. Xu ◽

C. Ji ◽

M. Deng ◽

...

Keyword(s):

Soil Surface ◽

Its Region ◽

The United States ◽

Crown Rot ◽

Stem Rot ◽

Universal Primers ◽

Conidial Suspension ◽

Fruiting Bodies ◽

Gentle Pressure ◽

Stem Lesions

During October 2009, the occurrence of a disease on soybean (Glycine max) was observed in several fields in Boluo County and Zengcheng City, Guangdong Province. Top leaves of infected plants initially turned yellow and plants eventually were defoliated, while stems and roots became black and rotted. The stem lesions sometimes extended 10 to 15 cm upward from the soil surface. Orange-to-brown spherical fruiting bodies, which were very similar with those of the soybean red crown rot pathogen, scattered or congregated on the stem lesions. Plants with symptoms were sampled from fields. Fruiting bodies were excised from diseased tissues. Microscopic examination revealed that they were perithecia, globose to pyriform, ostiolate with a short neck, and measured 160 to 298 × 151 to 235 μm. Under gentle pressure, asci and ascospores were exuded from these perithecia. Asci were eight spored, narrowly cylindrical to clavate, thin walled, with a short stalk, and measured 58 to 124 μm long and 8 to 15 μm in diameter. Ascospores were uniseriately arranged, globose to ellipsoid, thick walled, one celled, hyaline to pale, and measured 14 to 17 × 8 to 12 μm. Isolation was made from stem tissues at the edge of disease lesions on potato dextrose agar (PDA) amended with streptomycin sulfate and incubated at 25°C. Mycelia were white and floccose. Conidia were cylindrical to oblong-ellipsoidal, hyaline, one celled, and measured 6 to 22 × 2 to 5 μm, aggregating in a slimy mass on the apex of the conidiogenous cell. Abundant orange-to-brown spherical perithecia were produced on the colony. Ascospores had walls with a rugose ornamentation that could be clearly seen under a scanning electron microscope. The fungus was identified as Neocosmospora vasinfecta var. vasinfecta (anamorph Acremonium sp.) (1). The internal transcribed spacer (ITS) region of rDNA of two isolates were amplified with universal primers ITS1/ITS4 and sequenced (GenBank Accession No. JF705861 and JF705862), and comparisons with GenBank accessions showed 99% similarity with N. vasinfecta strain Pec070 (Accession No. FJ940902) and strain NRRL22497 (Accession No. AY381142). Pathogenicity tests were conducted. Five, 3-week-old seedlings of soybean cv. Huaxia No. 3 planted in plastic pots (20 cm in diameter) were wounded with a needle at the base of the stem below the soil line and near the root system, and then inoculated by drenching the soil with a conidial suspension (105 per ml). Control plants were inoculated with sterile water. There were six replicates for each treatment. The treated plants were incubated at 25 ± 2°C in a greenhouse. All inoculated plants exhibited symptoms of leaf yellowing and black rot of stems and roots 3 weeks after inoculation. N. vasinfecta var. vasinfecta was reisolated from the diseased plants. All control plants remained healthy. To our knowledge, this is the first observation of Neocosmospora stem rot of soybean in China. The pathogen could pose a threat to soybean, which is a major crop in China. This disease has been previously reported in the United States though the anamorph of the pathogen has either not been identified or has been identified as a Cylindrocarpon sp. (2,4). This fungus is also associated with human infections (3). References: (1) P. F. Cannon and D. L. Hawksworth. Trans. Br. Mycol. Soc. 82:673, 1984. (2) F. A. Gray et al. Plant Dis. 64:321, 1980. (3) P. Manikandan et al. Med. Mycol. 46:279, 2008. (4) D. V. Phillips. Phytopathology 62:612, 1972.

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First Report of Damping-Off of Rhodiola sachalinensis Caused by Rhizoctonia solani AG-4 HG-II in China

Plant Disease ◽

10.1094/pdis-07-11-0559 ◽

2012 ◽

Vol 96 (1) ◽

pp. 142-142 ◽

Cited By ~ 1

Author(s):

Q. Bai ◽

Y. Xie ◽

X. Wang ◽

Y. Li ◽

J. Gao ◽

...

Keyword(s):

Rhizoctonia Solani ◽

Morphological Characteristics ◽

Its Region ◽

Herbaceous Plant ◽

Protective Effects ◽

Damping Off ◽

First Report ◽

Rhodiola Sachalinensis ◽

Perennial Herbaceous Plant ◽

Safranin O

Rhodiola sachalinensis A. Bor (family Crassulaceae), a perennial herbaceous plant, is distributed mainly in the mountainous areas of China, Japan, Korea, and Russia. It is widely used as a traditional Chinese medicine with adaptogenic properties, cardiopulmonary protective effects, and central nervous system activities (3). Currently, it is extensively cultivated in northeastern China. In August 2010, widespread (>60% of plants were symptomatic) damping-off was observed in a seedling field in Linjiang, China. Leaves and stems near the ground were affected first, with dark lesions forming on the stem and the lowest leaves exhibiting wilt. The wilt spread rapidly over the entire plant with leaves becoming grayish brown and water soaked and then turned black and died. Root rot, defoliation, and damping-off were also observed. Six isolates with morphological characteristics of Rhizoctonia solani Kühn were isolated from symptomatic stems when plated on potato dextrose agar (PDA). Mycelium was branched at right angles with a septum near the branch and a slight constriction at the branch base. Fungal colonies were initially white, turned brown with age, and produced irregularly shaped, brown sclerotia after 8 days on PDA. Hyphal cells removed from cultures grown at 25°C on 2% water agar were determined to be multinucleate when stained with 1% safranin O and 3% KOH solution (1) and examined at ×400 magnification. The internal transcribed spacer (ITS) region of the nuclear rDNA was amplified by using the primers ITS4/ITS5 (2). The ITS sequences (715 bp) were identical in these six isolates (GenBank Accession No. FR878087) and had 100% sequence identity with R. solani AG-4 HG-II (GenBank Accession No. HQ629873) along with numerous other accessions from this AG subgroup. Pathogenicity tests were performed on healthy, potted seedlings of R. sachalinensis. Twenty plants were inoculated near the base of the stem with a 0.6-cm-diameter mycelial plug from 3-day-old PDA cultures for each isolate. Twenty plants inoculated with only PDA plugs served as controls. The plants were covered with plastic bags and kept in a greenhouse at 20 to 25°C for 72 h. All inoculated plants showed characteristic symptoms as previously observed in the seedling field 13 days after inoculation, while control plants remained healthy. R. solani AG-4 HG-II was reisolated from symptomatic tissues on inoculated plants. To our knowledge, this is the first report of R. solani AG-4 HG-II causing damping-off on R. sachalinensis in China. References: (1) R. J. Bandoni. Mycologia 71:873, 1979. (2) D. E. L. Cooke et al. Mycol. Res. 101:667, 1997. (3) T. F. Yan et al. Conserv. Genet. 4:213, 2003.

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First Report of Cherry Stem Rot and Leaf Necrosis Disease Caused by Phytophthora nicotianae in Yantai, China

Plant Disease ◽

10.1094/pdis-05-14-0489-pdn ◽

2015 ◽

Vol 99 (2) ◽

pp. 284-284

Author(s):

X. L. Yu ◽

X. Q. Liu ◽

P. S. Wang ◽

Y. Z. Wang

Keyword(s):

Average Length ◽

Morphological Characteristics ◽

Its Region ◽

Phytophthora Species ◽

Phytophthora Nicotianae ◽

Control Measures ◽

Stem Rot ◽

Economic Losses ◽

First Report ◽

Leaf Necrosis

Cherry (Cerasus avium (Linn.) Moench) is the third most economically important fruit in Yantai, Shandong Province, China. In August 2012, brown spots or necrosis on cherry seedling leaves, with an incidence of 8.2 to 34.3%, were observed in some fields of cherry seedlings in Yantai. Our survey indicated that the economic losses could reach up to 15.3% if disease conditions, such as a cool rainy summer season, were favorable. Conspicuous watery lesions on the stems turned to brown streaks; the leaves all wilted; and finally the plants collapsed. Diseased stem and leaf samples were surface-disinfected in 1% sodium hypochlorite for 1 min, rinsed three times in sterile water, which was absorbed with filter paper, and then transferred to 10% V8 juice agar medium containing 50 μg/ml ampicillin and 5 μg/ml carbendazim (1). The plates were incubated at 22°C in the dark for 5 days. The colonies consisted of white, loose, fluffy aerial mycelia. Eight isolates were obtained, and all were identified as Phytophthora nicotianae based on morphological characteristics and the sequence of the internal transcribed spacer (ITS) region of rDNA. The sporangia were ovoid/spherical, obturbinate with rounded bases and prominent papillae that were 37.5 to 62.5 × 30 to 50 μm (average 46.4 × 37.8 μm, n = 100) in size, with an average length-to-breadth ratio of 1.2. Chlamydospores were terminal, intercalary, and measured 19 to 42 μm (average 30.4 μm), which is typical of P. nicotianae (2). The genomic DNA of the eight isolates was extracted from mycelia. The ITS region of all eight isolates was amplified using primers ITS1 and ITS4, producing specific products that were directly sequenced. The sequence of a representative isolate P1401 (895 bp) was submitted to GenBank (Accession No. KJ754387). It was 100% similar to P. nicotianae strains NV-20T and TARI 22073 (KC768775 and GU111667). To confirm the pathogenicity, at least 10 cherry leaves and new stems were inoculated with mycelial plugs (5 × 5 mm) from each isolate. Necrosis of leaves and stems was observed 4 and 7 days after inoculation, respectively. No symptoms were observed on the control leaves and stems that were inoculated with blank agar plugs. P. nicotianae was re-isolated from the infected leaves, and the ITS sequence was analyzed to confirm its identity. Phytophthora species, such as P. cambivora, P. megasperma, and P. drechsleri, had been previously isolated from cherry (3), but to the best of our knowledge this is the first report of stem rot and leaf necrosis disease caused by P. nicotianae on cherry. Since the economic loss caused by this disease could reach 15% if an outbreak occurred in a rainy summer, control measures should be implemented. References: (1) Y. Balci et al. Mycol. Res. 112:906, 2008. (2) D. C. Erwin and O. K. Ribeiro. Phytophthora Diseases Worldwide. The American Phytopathological Society, St Paul, MN, 1996. (2) S. M. Mircetich and M. E. Matheron. Phytopathology 66:549, 1976.

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First Report of Nectria haematococca Causing Stem Rot of Soybean in China

Plant Disease ◽

10.1094/pdis-07-11-0626 ◽

2012 ◽

Vol 96 (3) ◽

pp. 457-457 ◽

Cited By ~ 1

Author(s):

Y. Gai ◽

R. Pan ◽

D. Xu ◽

M. Deng ◽

W. Chen ◽

...

Keyword(s):

Elongation Factor ◽

Morphological Characteristics ◽

Its Region ◽

Stem Rot ◽

Universal Primers ◽

Conidial Suspension ◽

Fruiting Bodies ◽

Nectria Haematococca ◽

First Report ◽

Soybean Seedlings

In October 2010, soybean (Glycine max) plants growing in commercial soybean fields in Zengcheng City, Guangdong Province developed symptoms consisting of stem and root rot, yellowing, and defoliation of leaves. Reddish, spherical fruiting bodies appeared in lesions that developed on stems. Plants with symptoms were sampled from fields. Fruiting bodies were excised from diseased tissues. Microscopic examination revealed that they were perithecia, globose to pyriform, and measured 197 to 260 μm in diameter and 226 to 358 μm long. When squeezed gently, cylindrical to clavate asci, 7.2 to 9.6 μm in diameter and 75.4 to 92.0 μm long, containing eight ascospores were exuded from the perithecia. Ascospores were ellipsoid to obovate, two celled, slightly constricted at the septum, had longitudinal striations, and measured 4.9 to 6.0 μm in diameter and 10.6 to 15.0 μm long. The fungus was isolated from the basal stem tissues of diseased soybean plants and cultured on potato dextrose agar (PDA) medium amended with streptomycin sulfate. On PDA, the culture developed into blue-pigmented colonies with whitish mycelium that produced oval to cylindrical microconidia. Microconidia had 0 to 1 septum, ranged from 2.5 to 5.2 × 7.6 to 29.4 μm, and were produced on monophialides. Macroconidia were cylindrical to falcate, thick walled, 2 to 5 septa, and 3.5 to 6.0 × 25.4 to 66.8 μm. Chlamydospores were present and ranged from 6.8 to 13.6 × 5.5 to 9.5 μm. Orange-to-reddish perithecia were readily formed in old culture. These morphological characteristics were consistent with descriptions of Nectria haematococca (anamorph Fusarium solani) (1). The rDNA internal transcribed spacer (ITS) region and the fragment of translation elongation factor 1-alpha (EF1-α) genes of the fungus were amplified, respectively, with universal primers ITS1/ITS4 and ef1/ef2 primers and sequenced. BLAST searches showed that the ITS sequences of three isolates (GenBank Accession Nos. JN015069, JN190942, and JN190943) had 99% similarity with those of N. haematococca(GenBank Accession Nos. DQ535186, DQ535185, and DQ535183) and the EF1-α sequences of three isolates (GenBank Accession Nos. JN874641, JN874642, and JN874643) had 100% similarity with those of F. solani (GenBank Accession Nos. DQ247265 and DQ247327). Completion of Koch's postulates confirmed the pathogenicity of the isolates in a replicated experiment. Thirty-day-old soybean seedlings of cultivar Huaxia No. 3 were inoculated by soaking their root systems in a conidial suspension (106 conidia per ml) for 30 min and then transplanted in plastic pots (20 cm in diameter) and incubated at 25 ± 2°C in a greenhouse. Control plants were treated with sterile water in the same way. There were four plants per pot and there were six replicates for each treatment. Within 3 weeks, more than 70% of the inoculated plants exhibited symptoms of leaf yellowing, stem rot, and root rots while control plants were symptomless. N. haematococca was reisolated from the diseased plants. To our knowledge, this is the first report of N. haematococca causing stem rot of soybean in China and the first description of sexual reproduction of F. solani causing soybean stem rot in nature. This pathogen may pose a serious threat to soybean production in China where soybean is a main crop. Reference: (1) C. Booth. The Genus Fusarium. CAB International, Wallingford, UK, 1971.

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First Report of Damping-off on Sedum plumbizincicola Caused by Rhizoctonia solani AG 2-1 in China

Plant Disease ◽

10.1094/pdis-08-20-1849-pdn ◽

2020 ◽

Author(s):

Ai Guo He ◽

Jin Chen ◽

Zhi Xin Hu ◽

Jie Zhong ◽

Jun Zi Zhu

Keyword(s):

Rhizoctonia Solani ◽

Management Strategies ◽

Morphological Characteristics ◽

Its Region ◽

Hunan Province ◽

Disease Spread ◽

Damping Off ◽

First Report ◽

Sedum Plumbizincicola

Sedum plumbizincicola X.H. Guo et S.B. Zhou sp. nov. is a plant species of the family Crassulaceae that has the ability to hyperaccumulate cadmium and zinc in high concentrations (Liu et al. 2017). In April of 2018 and 2019, a disease of damping-off was observed on S. plumbizincicola seedlings in a nursery in Changsha (28°13’N; 112°56’E), the Hunan Province of China, in which nearly 1 million seedlings were planted. Approximately 40% of the surveyed plants were infected. The affected plants displayed water-soaking on the shoots and stems, and chlorosis on the leaves. As the disease spread upward, leaf stalks or the whole plants became wilted and collapsed. Five diseased stem and leaf samples were collected. Symptomatic tissues were excised and surface sterilized with 70% ethanol for 10 s, and 0.1% HgCl2 for 2 min, washed with sterile distilled water for three times, and then cultured on potato dextrose agar (PDA) at 26°C in darkness. Fungal colonies were similar in morphology: white, light gray to brown, with hyphae branched at nearly right angles, septa near the branching point and constrictions at the base of hyphal branches. After 10 days, white-gray to brown sclerotia were produced. The morphological characteristics were consistent with those of Rhizoctonia solani J.G. Kühn (Sneh et al. 1991). Genomic DNA of a representative isolate was extracted using the cetyltrimethylammonium bromide method. The internal transcribed spacer (ITS) region of rDNA was amplified and sequenced with the primer pairs ITS4/ITS5 (White et al. 1990). When analyzed by the BLASTn program, the ITS sequence (GenBank Accession No. MN961664) had 100% identity to the corresponding gene sequence of R. solani anastomosis group (AG) 2-1(Accession Nos: LC202869.1 and MH862641.1). In addition, primers Rhsp1/ITS4B and Rhsp2/ITS1F specific for R. solani, and AG21sp/ITS4B specific for R. solani AG 2-1 were also used (Salazar et al. 2000). Results revealed that our isolate was R. solani AG 2-1. Pathogenicity was confirmed via in vivo inoculation of one-month-old S. plumbizincicola seedlings in sterilized nursery soil with four representative isolates. For each pot, five 5-mm-diameter mycelial plugs from 7-days old colonies on PDA were placed in the soil near the base of the stems. Plants inoculated with agar plugs without mycelium served as controls. The inoculated plants were kept in a growth chamber at 25°C with a 12/12 h light/dark cycle. Pathogenicity tests were performed twice, with three replicative potted plants for each isolate in each test. Approximately 25 days after inoculation, the damping-off symptoms resembling those observed in the field were displayed on the inoculated plants, while no obvious symptoms were observed on the control plants. R. solani was re-isolated from all infected plants and molecularly characterized, thus confirming Koch’s postulates. R. solani has been previously reported as the pathogen of damping-off disease in many plants, such as canola (Paulitz et al. 2006) and oat (Zhang et al. 2016). However, to the best of our knowledge, this is the first report of R. solani causing damping-off of S. plumbizincicola in China. S. plumbizincicola is widely planted for heavy metal pollution treatment in China. The occurrence of this disease could seriously affect the production of the seedlings, and management strategies should be developed.

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First Report of Stem Rot on Cereus peruvianus monstruosus Caused by Bipolaris cactivora (Petr.) Alcorn in Italy

Plant Disease ◽

10.1094/pdis-06-13-0614-pdn ◽

2014 ◽

Vol 98 (1) ◽

pp. 159-159 ◽

Cited By ~ 4

Author(s):

A. Garibaldi ◽

D. Bertetti ◽

P. Pensa ◽

A. Poli ◽

M. L. Gullino

Keyword(s):

United States ◽

Morphological Characteristics ◽

Its Region ◽

The United States ◽

Stem Rot ◽

Pathogenicity Test ◽

First Report ◽

Plastic Bags ◽

Blastn Analysis ◽

Dark Green

Cereus peruvianus monstruosus, known as “monster cactus,” family Cactaceae, is grown as a potted plant. In the winter of 2013, a stem rot was observed on a farm located near Ventimiglia (northern Italy) on 80% of 4,000 9-month-old plants grown in trays in a peat substrate. Symptoms consisted of a rapid rot of the upper portion of the stem. Affected stems at first showed yellowish spots that became brown irregular necrotic lesions with well-defined margins. The tissues below the affected areas were blackened and dry but became soft in the presence of high relative humidity. Fungal sporulation on rotted tissues consisted of caespitose, non-branched, septate conidiophores, olivaceous to brown at the base, paler above, measuring 89.0 to 196.9 × 6.2 to 8.7 (average 124.8 × 7.0) μm. Single conidia were borne on terminal cells. At maturity, conidia with 2 to 5 (average 3) septa were brownish-olivaceous, varying in shape from obclavate, fusiform, ellipsoid or sometimes furcate, and measuring 23.4 to 48.6 × 8.0 to 12.6 (average 38.8 × 10.3) μm. Symptomatic tissues were immersed in 1% sodium hypochlorite for 2 to 3 s and rinsed in sterile distilled water, then fragments excised from the margin of internal lesions were cultured on potato dextrose agar (PDA) medium amended with 25 mg/l of streptomycin sulfate and incubated at 20 to 23°C under alternating daylight and darkness (10 h light, 14 h dark). A fungus that was consistently isolated was subcultured on PDA. At maturity, dark green floccose colonies comprised of light brown septate hyphae, 4.2 to 8.1 (average 5.6) μm in width, produced non-branched, pale to dark brown, septate conidiophores, measuring 99.6 to 176.1 × 4.5 to 6.5 (average 146.7 × 5.4) μm. The conidia produced on PDA were similar to those observed on infected tissues and measured 20.6 to 40.7 × 7.5 to 11.4 (average 32.0 × 9.7) μm, with 1 to 3 septa (average 2). On the basis of the morphological characteristics, the fungus was identified as Bipolaris cactivora (Petr.) Alcorn [Syn.: Drechslera cactivora (Petr.) M. B. Ellis] (4). The internal transcribed spacer (ITS) region of the ribosomal DNA (rDNA) was amplified for one isolate using ITS1/ITS4 primers and sequenced (GenBank Accession No. KF041822). BLASTn analysis (1) of the 557-bp segment showed a 99% similarity with the ITS sequence of Bipolaris cactivora HM598679. For pathogenicity tests, 8 mm diameter mycelial disks removed from 15-day-old PDA cultures of the fungus were placed at the wounded stem apexes of three 7-month-old healthy plants (three disks per plant). Three plants inoculated with non-inoculated PDA disks served as controls. Plants were covered with plastic bags and maintained in a growth chamber at 23 ± 1°C with 12 h light/dark. By 8 days after inoculation, all the inoculated stems were rotted and 10 colonies of B. cactivora were re-isolated from infected tissues. Control plants remained healthy. The pathogenicity test was carried out twice with similar results. Several hosts are listed for B. cactivora including C. peruvianus, and the pathogen has been reported in the United States (2) and in South Korea (3). To our knowledge, this is the first report of B. cactivora on C. peruvianus monstruosus in Italy. References: (1) S. F. Altschul et al. Nucleic Acids Res. 25:3389, 1997. (2) D. F. Farr et al. Fungi on Plants and Plant Products in the United States. APS Press, St Paul, MN, 1989. (3) I. H. Hyun et al. Res. Plant Dis. 7:56, 2001. (4) A. Sivanesan. Mycopathologia 111:125, 1990.

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First report of Curvularia gladioli causing leaf spots on Gladiolus gandavensis in China

Plant Disease ◽

10.1094/pdis-01-21-0016-pdn ◽

2021 ◽

Author(s):

Jingxin Zhang ◽

Huifang Shen ◽

Yongqiang Zhang ◽

Xiaoming Pu ◽

Qiyun Yang ◽

...

Keyword(s):

Management Strategies ◽

Morphological Characteristics ◽

Guangdong Province ◽

Its Region ◽

Diagnostic Methods ◽

Morphological Observation ◽

Its Sequencing ◽

First Report ◽

The Third ◽

Leaf Spots

Gladiolus (Gladiolus gandavensis Van Houtte) is a perennial plant in the family Iridaceae, which shows sword-shaped leaves and spikes of brilliantly colored irregular flowers arising from corms. It is one of the most important fresh cut flowers and is widely cultivated worldwide, including in China. In September 2020, white pinpoints were first observed on gladiolus leaves in Jiangmen City, Guangdong Province, China. The white spots eventually turned brown. The lesions then developed into oval to circular spots, which were surrounded with an obvious yellow halo. The spots expanded and coalesced, causing leaf blight. These symptoms were observed on approximately 10% of gladiolus plants in fields measuring ca. 70 ha. Symptomatic leaves were sampled from fields, surface sterilized in 75% ethanol for 30 s, submerged in a 2% NaOCl solution for 10 min, and rinsed three times with sterile water. The samples were then cut into pieces (5 × 5 mm) and incubated for 4 d on potato dextrose agar (PDA) at 25°C. A representative fungal colony was subcultured onto new PDA and grown for another 7 d, and its mycelium appeared to be grayish-black and villiform. This strain was named as Cg_TS. Its conidiophores were simple, septate, cylindrical in shape, and moderate brown in color. They occurred singly or in groups. They were straight or slightly flexuous and ranged in size from 57.0 to 80.0 μm × 4.0 to 8.0 μm. Conidia were 3-distoseptate and curved at the third cell from the base. The third cell was swollen to one side and larger than other cells. These conidia ranged in size from 23.5 to 32.0 μm × 11.5 to 16.0 μm. These morphological characteristics were consistent with the description of Curvularia gladioli Boerema & Hamers (Boerema and Hamers 1989). Using primer pair ITS1 and ITS4, PCR was applied to amplify the internal transcribed spacer (ITS) region of rDNA. This sequence (GenBank accession No. MW426196.1) was subjected to BLAST in GenBank, suggesting that it was most similar to C. gladioli sequences, LT631345.1 and HG778987.1, with both of 99.49% of similarity. To fulfill Koch's postulates, healthy two-month-old gladiolus plants were used for pathogenicity testing, and the leaves were wounded by pressing slightly with a pipette tip. Mycelium disks (3 mm diameter) were applied onto wounded leaves of 10 plants. Another 10 healthy plants were inoculated with PDA disks which served as control. Inoculated samples were placed in a greenhouse at 25°C and 90% relative humidity. After 3 d, brown leaf spots appeared on all of pathogen-inoculated leaves. The symptoms were consistent with those initially observed and C. gladioli was re-isolated from the symptomatic tissue. Identification was confirmed by morphological observation and ITS sequencing. Control leaves remained symptomless. The curvularia fungus was firstly reported on gladiolus in Florida in 1947 and spread globally via infected corms (Torre et al. 2015), it was also reported to cause leaf spots on gladiolus in Brazil in 2013 (Torres et al. 2013). Although C. gladioli had been recorded as a Curvularia species occurring in China (Zhang et al. 2006), it was not reported to cause leaf spots on gladiolus in Guangdong Province and elsewhere in China. To our knowledge, this is the first report of Curvularia gladioli causing leaf spots on gladiolus in China. Identification of this pathogen will help develop diagnostic methods for corms and seedlings, and may lead to the development of appropriate chemical management strategies.

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Identification and Characterization of Isolates of Pythium and Phytophthora spp. from Snap Beans with Cottony Leak

Plant Disease ◽

10.1094/pdis-06-15-0662-re ◽

2016 ◽

Vol 100 (7) ◽

pp. 1446-1453 ◽

Cited By ~ 5

Author(s):

J. D. Olson ◽

J. P. Damicone ◽

B. A. Kahn

Keyword(s):

Morphological Characteristics ◽

Pythium Ultimum ◽

The United States ◽

Phytophthora Nicotianae ◽

Damping Off ◽

Internal Transcribed Spacer Region ◽

Snap Bean ◽

Snap Beans ◽

Phytophthora Spp ◽

Phytophthora Drechsleri

Cottony leak is an important disease of snap bean in Oklahoma and nearby states. Oomycete pathogens isolated from diseased pods collected from commercial fields and research plots consisted of both Pythium spp. (n = 131) and Phytophthora spp. (n = 46). Isolates were identified to species by morphological characteristics and by sequencing a portion of the internal transcribed spacer region of representative isolates. The most common Pythium spp. were Pythium ultimum var. ultimum; Pythium ‘group HS’, a self-sterile form of P. ultimum that produces hyphal swellings in lieu of sporangia (n = 74); and P. aphanidermatum (n = 50). Phytophthora spp. included Phytophthora drechsleri (n = 41) and P. nicotianae (n = 5). Nearly all of the isolates (95%) and all species were pathogenic on detached pods but Pythium ultimum var. ultimum and Pythium ‘group HS’ were most aggressive. Phytophthora drechsleri was most aggressive on seedlings, causing preemergence damping off and seed rot. Pythium ultimum var. ultimum, Pythium ‘group HS’, and P. aphanidermatum were intermediate in virulence to seedlings, causing root rot, stunting, and limited postemergence damping off. Phytophthora nicotianae and Pythium diclinum (n = 4) were not pathogenic on seedlings. Most (87%) isolates were sensitive to metalaxyl-M (concentration that caused a 50% reduction in mycelial growth [EC50] < 1 μg/ml) and the rest were intermediate in sensitivity (EC50 > 1 to < 100 μg/ml). Phytophthora drechsleri was the most sensitive species (EC50 = 0.06 μg/ml) compared with Pythium aphanidermatum, which was least sensitive (EC50 = 1.3 μg/ml). Cottony leak is a disease complex caused by several oomycete species that should include Phytophthora drechsleri, a newly reported pathogen of snap bean in the United States.

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