First Report of Fusarium graminearum Causing Flower Blight On Hemp (Cannabis sativa) in Kentucky

In October of 2020, a grower in Boyle County, KY, reported mold and blight symptoms on flowers of field-grown hemp. Plants were approaching harvest, and the mold was affecting 100% of the cultivar ‘White CBG’ being grown for cannabinoid (CBD) extraction. Mycelium colonized the flower heads and any seeds within bracts. Affected flower bracts were necrotic, and mycelium and necrosis in the most severe cases also encompassed adjacent (sugar) leaves. Necrotic symptomatic tissue was collected, disinfested in 10% bleach for one minute, and cultured on acidified potato dextrose agar (APDA). Each isolate was single-spored, transferred to PDA, stored in 15% glycerol at -80°C and maintained at room temperature under blacklight blue and fluorescent bulbs on a 12-hour light-dark cycle. Colonies produced white-pink mycelia with a dark red pigment on the undersides. Conidia collected after 7-9 days were falcate and septate (5 to 6). No microconidia were produced. Macroconidia measured 35.4-49.7 µm x 3.4-5.8 µm (n=50). The strains produced blue-black fertile perithecia on carrot agar when induced according to the method of (Bowden and Leslie, 1999). To confirm pathogenicity, flowers of hemp cultivars ‘Lifter’, ‘Trump Towers’, ‘Wife’ and ‘White CBG’ were inoculated in the greenhouse with a representative fungal strain (20Hemp010). Plants were inoculated at two different stages: when the styles were still green or after they had become senescent. Macroconidia were collected from 7- to 9-day-old cultures grown under a 12-hour light-dark cycle. Plants were spray-inoculated with a 5 x 105 per ml conidial suspension in 0.05% Tween 20 until runoff. Flower heads were individually covered with clear plastic bags and incubated for 72 h at 95-100% humidity under greenhouse benches to avoid direct light. Bags were removed after 72 h and returned to the bench. Greenhouse conditions were 23-25°C with a 14-hour photoperiod and 50% RH. Symptoms developed 7 dai in 1% of the flowers inoculated when styles were green, and 36% of the flowers that had senescent styles. Symptoms were similar to those initially noticed in Boyle County, including necrotic flower bracts and sugar leaves, and visible fungal growth. Symptoms were more severe on plants inoculated when styles were necrotic. Recovered fungi were morphologically similar to 20Hemp010. Genomic DNA was extracted from the mycelium with the Zymo Research Quick-DNA Fungal/Bacterial Miniprep Kit. A fragment of the translation elongation factor 1-alpha 1 gene was amplified with primers EF1 and EF2 as described by (O’Donnell et al. 1998). Amplicons were sequenced and the consensus (MZ407909) was compared with the NCBI GenBank Refseq database by BLASTn. The top hit was Fusarium graminearum with 100% identity (JF270185.1). Pairwise alignments via MycoBank Fusarium MLST and Fusarium-ID also revealed a top hit of F. graminearum with 100% identity (AY452957.1). Conidial and colony morphology were also consistent with F. graminearum (Leslie and Summerell, 2006), thus we conclude that this species was the causal agent of the flower blight and mold. The same disease was subsequently confirmed on hemp in Breathitt and Franklin Counties in KY in 2020. This is the first report of this disease in KY, although F. graminearum has been reported previously causing a similar flower blight on hemp in NY and NC (Bergstrom et al., 2019, Thiessen et al. 2020). Fusarium graminearum is common in KY as a cause of Fusarium head blight on wheat and Gibberella ear rot on corn. In cereals, fungal infection is facilitated by the production of the mycotoxin deoxynivalenol (DON), which is harmful to humans and livestock (Desjardins and Hohn, 1997). As hemp production in Kentucky continues to rise for production of CBD products and edible grains, accumulation and concentration of DON in these products could become a concern.

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First report of Trichoderma afroharzianum causing seed rot on maize in Italy

Plant Disease ◽

10.1094/pdis-12-21-2697-pdn ◽

2022 ◽

Author(s):

Martina Sanna ◽

Massimo Pugliese ◽

Maria Lodovica GULLINO ◽

Monica Mezzalama

Keyword(s):

Rna Polymerase Ii ◽

Elongation Factor ◽

Aerial Mycelium ◽

Light Cycle ◽

Conidial Suspension ◽

Ear Rot ◽

Trichoderma Spp ◽

Trichoderma Species ◽

First Report ◽

Pathogenicity Tests

Maize (Zea mays L.) is a cereal crop of great economic importance in Italy; production is currently of 60,602,320 t, covering 588,597 ha (ISTAT 2021). Trichoderma species are widespread filamentous fungi in soil, well known and studied as biological control agents (Vinale et al., 2008). Seeds of a yellow grain hybrid (class FAO 700, 132 days) were collected in September 2020 from an experimental field located in Carmagnola (TO, Italy: GPS: 44°53'11.0"N 7°40'60.0"E) and tested with blotter test (Warham et al., 1996) to assess their phytosanitary condition. Over the 400 seeds tested, more than 50% showed rotting and development of green mycelium typical of the genus Trichoderma. Due to the high and unexpected percentage of decaying kernels, ten colonies were identified by morphological and molecular methods. Single conidia colonies of one Trichoderma (T5.1) strain were cultured on Potato Dextrose Agar (PDA) for pathogenicity tests, and on PDA and Synthetic Nutrient-Poor Agar (SNA) for morphological and molecular identification. The colonies grown on PDA and SNA showed green, abundant, cottony, and radiating aerial mycelium, and yellow pigmentation on the reverse. Colony radius after 72 h at 30°C was of 60-65 mm on PDA and of 50-55 mm on SNA. The isolates produced one cell conidia 2.8 - 3.8 µm long and 2.1 - 3.6 µm wide (n=50) on SNA. Conidiophores and phialides were lageniform to ampulliform and measured 4.5 – 9.7 µm long and 1.6 – 3.6 µm wide (n=50); the base measure 1.5 – 2.9 µm wide and the supporting cell 1.4 – 2.8 µm wide (n=50). The identity of one single-conidia strain was confirmed by sequence comparison of the internal transcribed spacer (ITS), the translation elongation factor-1α (tef-1α), and RNA polymerase II subunit (rpb2) gene fragments (Oskiera et al., 2015). BLASTn searches of GenBank using ITS (OL691534) the partial tef-1α (OL743117) and rpb2 (OL743116) sequences of the representative isolate T5.1, revealed 100% identity for rpb2 to T. afroharzianum TRS835 (KP009149) and 100% identity for tef-1α to T. afroharzianum Z19 (KR911897). Pathogenicity tests were carried out by suspending conidia from a 14-days old culture on PDA in sterile H2O to 1×106 CFU/ml. Twenty-five seeds were sown in pots filled with a steamed mix of white peat and perlite, 80:20 v/v, and maintained at 23°C under a seasonal day/night light cycle. Twenty primary ears were inoculated, by injection into the silk channel, with 1 ml of a conidial suspension of strain T5.1 seven days after silk channel emergence (BBCH 65) (Pfordt et al., 2020). Ears were removed four weeks after inoculation and disease severity, reaching up to 75% of the kernels of the twenty cobs, was assessed visually according to the EPPO guidelines (EPPO, 2015). Five control cobs, inoculated with 1 ml of sterile distilled water were healthy. T. afroharzianum was reisolated from kernels showing a green mold developing on their surface and identified by resequencing of tef-1α gene. T. afroharzianum has been already reported on maize in Germany and France as causal agent of ear rot of maize (Pfordt et al. 2020). Although several species of Trichoderma are known to be beneficial microorganisms, our results support other findings that report Trichoderma spp. causing ear rot on maize in tropical and subtropical areas of the world (Munkvold and White, 2016). The potential production of mycotoxins and the losses that can be caused by the pathogen during post-harvest need to be explored. To our knowledge this is the first report of T. afroharzianum as a pathogen of maize in Italy.

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First Report of Fusarium Maize Ear Rot Caused by Fusarium meridionale in China

Plant Disease ◽

10.1094/pdis-01-14-0108-pdn ◽

2014 ◽

Vol 98 (8) ◽

pp. 1156-1156 ◽

Cited By ~ 4

Author(s):

H. Zhang ◽

W. Luo ◽

Y. Pan ◽

J. Xu ◽

J. S. Xu ◽

...

Keyword(s):

Fusarium Species ◽

Elongation Factor ◽

Morphological Characteristics ◽

Pathogenicity Test ◽

Conidial Suspension ◽

Fusarium Ear Rot ◽

Ear Rot ◽

Translation Elongation Factor ◽

Translation Elongation ◽

First Report

Fusarium is an important genus of fungal pathogens that are responsible for devastating diseases, such as Fusarium ear rot on maize, which may result in yield losses and/or mycotoxin contamination. In September 2013, a survey to determine population composition of Fusarium species on maize was conducted at 22 fields in 18 counties in Gansu Province. Maize ears with clear symptoms (with a white to pink- or salmon-colored mold at the ear tip) were collected. Symptomatic seeds were surface-sterilized with 70% ethanol and 10% sodium hypochlorite and rinsed three times with sterile water to eliminate hypochlorite residues. After drying on sterile filter paper, the seeds were placed on potato dextrose agar (PDA) and incubated at 25°C in the dark for 3 days. Mycelium that was characteristic of Fusarium spp. (2) was purified by transferring single spores to fresh PDA. Fusarium species were identified by morphological characteristics (2), multilocus genotyping assay (MLGT) (3), and sequence analysis of the translation elongation factor-1α (TEF) gene. Several Fusarium species were identified and Fusarium verticillioides and F. proliferatum were the predominant species. Based on MLGT, two strains from Chenghong County were identified as F. meridionale with NIV chemotype, a species in F. graminearum species complex (FGSC). Morphological characteristics were also identical to FGSC. Colonies grew rapidly on PDA and produce relatively large amounts of dense mycelia and red pigments. Slender, thick-walled, and moderately curved or straight macroconidia were observed with 5- to 6-septate. Furthermore, conidia on SNA also showed typical characteristics of F. meridionale, as the dorsal and ventral lines were often parallel and gradually curved. Sequences comparison of the partial translation elongation factor (TEF-1α, 644 bp) gene (1) was used to validate these observations. BLASTn analysis with the FUSARIUM-ID database revealed 100% sequence identity to F. meridionale (GenBank Accession No. KJ137017). Thus, both morphological and molecular criteria supported identification of the strains as F. meridionale. A pathogenicity test was performed on Zhengdan958, the maize variety with the largest planted acreage in China. Four days after silk emergence, 2 ml conidial suspension (105 macroconidia/ml) of each isolate were injected into each of 10 maize ears through silk channel. Control plants were inoculated with sterile distilled water. Typical FER symptoms (reddish-white mold) was observed on inoculated ears and no symptoms were observed on water controls. Koch's postulates were fulfilled by re-isolating the same fungus from the infected seeds. F. meridionale was one of the pathogens causing Fusarium head blight on wheat and barley in China and produced nivalenol (4,5) and it also has been isolated from maize in Korea and Nepal. To our knowledge, this is the first report of F. meridionale causing Fusarium ear rot on maize in China. Further studies on biological characteristics such as temperature sensibility and fungicide resistance are needed to gain a better understanding of this new pathogen. References: (1) D. M. Geiser et al. Eur. J. Plant Pathol. 110:473, 2004. (2) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Ames, IA, 2006. (3) T. J. Ward et al. Fungal Genet. Biol. 45:473, 2008. (4) L. Yang et al. Phytopathology 98:719, 2008. (5) H. Zhang et al. Plos one 7:e31722, 2012.

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First report of Head blight of wheat caused by Fusarium vorosii in Serbia

Plant Disease ◽

10.1094/pdis-04-21-0715-pdn ◽

2021 ◽

Author(s):

Ana Obradović ◽

Jelena Stepanovic ◽

Vesna Krnjaja ◽

Aleksandra Bulajic ◽

Goran Stanković ◽

...

Keyword(s):

Fusarium Graminearum ◽

Species Recognition ◽

Histone H3 ◽

Elongation Factor ◽

Phylogenetic Species ◽

Head Blight ◽

Single Spore ◽

First Report ◽

Genealogical Concordance ◽

Β Tubulin

The cosmopolitan species Fusarium graminearum Schwabe directly reduces yield, as well as grain quality of cereals, due to its ability to synthesize mycotoxins. Previously it was considered to be one species occurring on all continents. However, phylogenetic analysis employing the GCPSR method (Genealogical Concordance Phylogenetic Species Recognition) revealed the existence of 15 phylogenetic species within what is now recognised as the Fusarium graminearum Species Complex (FGSC) (Sarver et al. 2011). During 1996-2008, a MRIZP collection of FGSC isolates was established and isolates originating from wheat (5), maize (3) and barely (2) were selected for further study. Morphological features including the appearance of colonies and macroconidia (average size 38.5-53.1 × 4.6-5.4 µm, No 50) of all 10 isolates on PDA were consistent with descriptions of F. graminearum (O’Donnell et al. 2004, Leslie and Summerell 2006). Total DNA was isolated from mycelium removed from 7-day old colonies of single-spore isolates grown on PDA using the DNeasy Plant Mini Kit (Qiagen, Hilden). Further identification was based on amplification and sequencing of elongation factor TEF−1α, histone H3 and β−tubulin in both directions, with primers ef1/ef2, H3-1a/H3-1b and T1/T22, respectively (Jacobs et al. 2010). The sequences were deposited in NCBI under accession numbers MF974399 - MF974408 (TEF−1α), MG063783 - MG063792 (β−tubulin) and MF999139 - MF999148 (histone H3). Sequence analysis was performed using BLAST while genetic similarity was calculated using MEGA 6.0 software. Isolate 1339 originating from wheat (collected at the locality of Kikinda in 2006), shared 100% nucleotide identity with TEF−1α (DQ459745), histone H3 (DQ459728) and β−tubulin (DQ459643) of F. vorosii isolate NRRL37605 (Starkey et al. 2007). The remaining nine isolates were identified as F. graminearum as they shared 99% to 100% nucleotide similarity with F. graminearum NRRL 28439 (O’Donnell et al. 2004). Pathogenicity was tested using artificial inoculations of spikes during wheat flowering (Mesterhazy et al. 1999). Thirty classes were inoculated with each isolate, in three replicates. Inoculum was prepared from 7-day colonies on PDA, and 30 ml of a conidia suspension (1x105 conidia/ml) was used. Control plants were inoculated with sterile water. Three weeks after inoculation, typical Fusarium head blight symptoms were visible on inoculated plants, from which all 10 isolates were successfully reisolated. Control spikes remained symptomless. Disease severity was estimated on the 1-7 scale (Blandino et al. 2012). Average pathogenicity of the F. vorosii isolate 1339 was 1.9, and 2.4 -5.1 of F. graminearum isolates. Toxin production was determined using gas chromatography-tandem mass spectrometry. Kernels inoculated with the 10 isolates were ground and tested for the presence of deoxynivalenol (DON) and its acetyl derivatives 3ADON, 15ADON and NIV. F. vorosii isolate 1339 possessed the 15ADON chemotype, as well as eight F. graminearum isolates, while only one F. graminearum isolate was 3ADON chemotype. To date, F. vorosii has only been detected in Hungary on wheat (Toth et al. 2005) and Korea on barley, corn and rice (Lee et al. 2016). This is the first report of F. vorosii in Serbia, which is of great importance, because it indicates the spread of this toxigenic species. Further studies should be focused on determining the distribution, aggressiveness and toxicological profile of F. vorosii.

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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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First Report of Fusarium stem and root rot of Gerbera jamesonii caused by Fusarium incarnatum in China

Plant Disease ◽

10.1094/pdis-02-21-0440-pdn ◽

2021 ◽

Author(s):

Shuning Chen ◽

Wei Sun ◽

Huizhu Yuan ◽

Xiaojing Yan

Keyword(s):

Root Rot ◽

Vascular System ◽

Elongation Factor ◽

Morphological Observation ◽

Conidial Suspension ◽

Gerbera Jamesonii ◽

Gene Sequences ◽

Plastic Container ◽

First Report ◽

Brown Discoloration

Gerbera (Gerbera jamesonii Bolus) is an important cut flower grown globally. In 2020, gerbera plants (Redaicaoyuan, Baimawangzi, and Hongditan cultivars) with roots, crowns, and stems rot were found in a greenhouse in Nanping, Fujian, China. Approximately 30% of the 60,000 plants showed symptoms. Diseased plants were stunted with chlorotic leaves. The leaves and flower heads were wilted and withered. Brown discoloration with red to black streaks occurred in the vascular system of the crown and stem. The stem pieces (3×3 mm) showing the symptom were surface-disinfected with 1% NaClO for 1 min and washed three times with sterilized water. The stem pieces were then dried and placed on potato dextrose agar (PDA) at 25℃ inside a dark chamber. Ten single-spored isolates were identified as Fusarium incarnatum based on morphological features. White to light brown mycelia were observed among the isolates on PDA medium. Falculate, multicelluar, straight to slightly curved macroconidia produced in monophialide sporodochia without distinctive foot shaped basal cell; and chlamydospores produced in some isolates (Leslie and Summerell). The size of macroconidia was 36.4 ± 5.20 × 4.6 ± 1.3 μm (n = 100) with 3 to 5 septates. Microconidia were mostly 0 to 1 septate measured 14.6 ± 1.9 × 2.6 ± 0.5 μm (n=100). Based on the morphological observation, isolates were further identified by molecular method. The ITS1/4 region combined with partial gene fragments of translation elongation factor (EF-1α, primer EF1/EF2, Geiser et al.) and calmodulin (CAM, primer CL1/CL2A, O’Donnell.) from the isolates were amplified and sequenced. All of the three tested isolates showed identical gene sequences. Sequences amplified from one represented isolate FIN-1 were submitted to Genbank. BLAST searches revealed that ITS1/4 (MW527088), EF-1α (MW556488), and CAM (MW556487) had 99.22%, 99.53%, 99.42% identity compared to F. incarnatum (MN480497, MN233577, and LN901596, respectively) in GenBank. FUSARIUM-ID (Geiser et al. 2004) analysis also showed 99 to 100% similarity with sequences of the F. incarnatum-equiseti species complex (FIESC) (FD_01636 for CAM, FD_01643 for EF-1α). The phylogenetic analysis was conducted using neighbor-joining algorithm based on the ITS, EF-1α, and CAM gene sequences. The isolate was clustered with F. incarnatum clade. Then, the pathogenicity of the fungus was confirmed by performing Koch’s postulates. Pure single-spored cultures were grown on carboxymethyl-cellulose (CMC) medium for sporulation. G. jamesonii plants used for pathogenicity tests were grown on sterilized potting soil in a plastic container to the ten-leaf stage prior to inoculation. Spores harvested from the CMC medium were adjusted to a concentration of 1×105 conidial/ml. Twelve healthy rooted gerbera seedlings were inoculated by drenching 10 ml of the conidial suspension onto roots. Twelve gerbera seedlings treated with 10 ml sterile water served as control treatments. Plants were grown in the glasshouse at temperatures of 23°C, relative humidity >70%, and 16 h light per day. After 10 days, blackening stems and withered leaf edges began to appear on inoculated seedlings, whereas control seedlings remained healthy. F. incarnatum was consistently re-isolated from the symptomatic stems, whereas no isolates were obtained from the control seedlings. The assay was conducted twice. To the best of our knowledge, this is the first report of F. incarnatum causing stem and root rot on G. jamesonii.

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First Report of Anthracnose of Onion Caused by Colletotrichum coccodes in Ohio

Plant Disease ◽

10.1094/pdis-01-14-0032-pdn ◽

2014 ◽

Vol 98 (9) ◽

pp. 1271-1271 ◽

Cited By ~ 2

Author(s):

F. Baysal-Gurel ◽

N. Subedi ◽

D. P. Mamiro ◽

S. A. Miller

Keyword(s):

Leaf Tissue ◽

Its Region ◽

The United States ◽

Tween 20 ◽

Colletotrichum Coccodes ◽

Academic Press ◽

Conidial Suspension ◽

First Report ◽

Allium Cepa L ◽

Total Dna

Dry bulb onion (Allium cepa L. cvs. Pulsar, Bradley, and Livingston) plants with symptoms of anthracnose were observed in three commercial fields totaling 76.5 ha in Huron Co., Ohio, in July 2013. Symptoms were oval leaf lesions and yellowing, curling, twisting, chlorosis, and death of leaves. Nearly half of the plants in a 32.8-ha field of the cv. Pulsar were symptomatic. Concentric rings of acervuli with salmon-colored conidial masses were observed in the lesions. Conidia were straight with tapered ends and 16 to 23 × 3 to 6 μm (2). Colletotrichum coccodes (Wallr.) S. Hughes was regularly isolated from infected plants (2). Culturing diseased leaf tissue on potato dextrose agar (PDA) amended with 30 ppm rifampicin and 100 ppm ampicillin at room temperature yielded white aerial mycelia and salmon-colored conidial masses in acervuli. Numerous spherical, black microsclerotia were produced on the surface of colonies after 10 to 14 days. To confirm pathogen identity, total DNA was extracted directly from a 7-day-old culture of isolate SAM30-13 grown on PDA, using the Wizard SV Genomic DNA Purification System (Promega, Madison, WI) following the manufacturer's instructions. The ribosomal DNA internal transcribed spacer (ITS) region was amplified by PCR using the primer pair ITS1 and ITS4 (2), and sequenced. The sequence, deposited in GenBank (KF894404), was 99% identical to that of a C. coccodes isolate from Michigan (JQ682644) (1). Ten onion seedlings cv. Ebenezer White at the two- to three-leaf stage of growth were spray-inoculated with a conidial suspension (1 × 105 conidia/ml containing 0.01% Tween 20, with 10 ml applied/plant). Plants were maintained in a greenhouse (21 to 23°C) until symptoms appeared. Control plants were sprayed with sterilized water containing 0.01% Tween 20, and maintained in the same environment. After 30 days, sunken, oval lesions each with a salmon-colored center developed on the inoculated plants, and microscopic examination revealed the same pathogen morphology as the original isolates. C. coccodes was re-isolated consistently from leaf lesions. All non-inoculated control plants remained disease-free, and C. coccodes was not re-isolated from leaves of control plants. C. coccodes was reported infecting onions in the United States for the first time in Michigan in 2012 (1). This is the first report of anthracnose of onion caused by C. coccodes in Ohio. Unusually wet, warm conditions in Ohio in 2013 likely contributed to the outbreak of this disease. Timely fungicide applications will be necessary to manage this disease in affected areas. References: (1) A. K. Lees and A. J. Hilton. Plant Pathol. 52:3. 2003. (2) L. M. Rodriguez-Salamanca et al. Plant Dis. 96:769. 2012. (3) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990.

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First Report of Nigrospora osmanthi Causing Leaf Spot on Tartary Buckwheat in China

Plant Disease ◽

10.1094/pdis-08-20-1773-pdn ◽

2020 ◽

Author(s):

Quan Shen ◽

Xixu Peng ◽

Feng He ◽

Shaoqing Li ◽

Zuyin Xiao ◽

...

Keyword(s):

Relative Humidity ◽

Leaf Spot ◽

Plant Pathogens ◽

Hunan Province ◽

Tartary Buckwheat ◽

Tween 20 ◽

Pathogenicity Test ◽

Conidial Suspension ◽

Sterile Water ◽

First Report

Buckwheat (Fagopyrum tataricum) is a traditional short-season pseudocereal crop originating in southwest China and is cultivated around the world. Antioxidative substances in buckwheat have been shown to provide many potential cardiovascular health benefits. Between August and November in 2019, a leaf spot was found in several Tartary buckwheat cv. Pinku1 fields in Xiangxiang County, Hunan Province, China. The disease occurred throughout the growth cycle of buckwheat after leaves emerged, and disease incidence was approximately 50 to 60%. Initially infected leaves developed a few round lesions, light yellow to light brown spots. Several days later, lesions began to enlarge with reddish brown borders, and eventually withered and fell off. Thirty lesions (2×2 mm) collected from three locations with ten leaves in each location were sterilized in 70% ethanol for 10 sec, in 2% sodium hypochlorite for 30 sec, rinsed in sterile water for three times, dried on sterilized filter paper, and placed on a potato dextrose PDA with lactic acid (3 ml/L), and incubated at 28°C in the dark for 3 to 5 days. Fungal colonies were initially white and later turned black with the onset ofsporulation. Conidia were single-celled, black, smooth, spherical to subspherical, and measured 9.2 to 15.6 µm long, and 7.1 to 11.6 µm wide (n=30). Each conidium was terminal and borne on a hyaline vesicle at the tip of conidiophores. Morphologically, the fungus was identified as Nigrospora osmanthi (Wang et al. 2017). Identification was confirmed by amplifying and sequencing the ITS region, and translation elongation factor 1-alpha (TEF1-α) and partial beta-tublin (TUB2) genes using primers ITS1/ITS4 (Mills et al. 1992), EF1-728F/EF-2 (Carbone and Kohn 1999; O’Donnell et al. 1998) and Bt-2a/Bt-2b (Glass et al. 1995), respectively. BLAST searches in GenBank indicated the ITS (MT860338), TUB2 (MT882054) and TEF1-α (MT882055) sequences had 99.80%, 99% and 100% similarity to sequences KX986010.1, KY019461.1 and KY019421.1 of Nigrospora osmanthi ex-type strain CGMCC 3.18126, respectively. A neighbor-joining phylogenetic tree constructed using MEGA7.0 with 1,000 bootstraps based on the concatenated nucleotide sequences of the three genes indicated that our isolate was closely related to N. osmanthi. Pathogenicity test was performed using leaves of healthy F. tataricum plants. The conidial suspension (1 × 106 conidia/ml) collected from PDA cultures with 0.05% Tween 20 buffer was used for inoculation by spraying leaves of potted 20-day-old Tartary buckwheat cv. Pinku1. Five leaves of each plant were inoculated with spore suspensions (1 ml per leaf). An equal number of control leaves were sprayed with sterile water to serve as a control. The treated plants were kept in a greenhouse at 28°C and 80% relative humidity for 24 h, and then transferred to natural conditions with temperature ranging from 22 to 30°C and relative humidity ranging from 50 to 60%. Five days later, all N. osmanthi-inoculated leaves developed leaf spot symptoms similar to those observed in the field, whereas control leaves remained healthy. N. osmanthi was re-isolated from twelve infected leaves with frequency of 100%, fulfilling Koch’s postulates. The genus Nigrospora has been regarded by many scholars as plant pathogens (Fukushima et al. 1998) and N. osmanthi is a known leaf blight pathogen for Stenotaphrum secundatum (Mei et al. 2019) and Ficus pandurata (Liu et al. 2019) but has not been reported on F. tataricum. Nigrospora sphaerica was also detected in vegetative buds of healthy Fagopyrum esculentum Moench (Jain et al. 2012). To our knowledge, this is the first report of N. osmanthi causing leaf spot on F. tataricum in China and worldwide. Appropriate strategies should be developed to manage this disease.

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Investigating Useful Properties of Four Streptomyces Strains Active against Fusarium graminearum Growth and Deoxynivalenol Production on Wheat Grains by qPCR

Toxins ◽

10.3390/toxins12090560 ◽

2020 ◽

Vol 12 (9) ◽

pp. 560

Author(s):

Elena Maria Colombo ◽

Andrea Kunova ◽

Claudio Gardana ◽

Cristina Pizzatti ◽

Paolo Simonetti ◽

...

Keyword(s):

Fusarium Graminearum ◽

Plant Pathogens ◽

Fungal Growth ◽

Toxin Production ◽

Fungal Mycelium ◽

Head Blight ◽

Wheat Grains ◽

Streptomyces Spp ◽

Dual Plate

Streptomyces spp. can be exploited as biocontrol agents (BCAs) against plant pathogens such as Fusarium graminearum, the main causal agent of Fusarium head blight (FHB) and against the contamination of grains with deoxynivalenol (DON). In the present research, four Streptomyces strains active against F. graminearum in dual plate assays were characterized for their ability to colonize detached wheat grains in the presence of F. graminearum and to limit DON production. The pathogen and BCA abundance were assessed by a quantitative real-time PCR, while DON production was assessed by HPLC quantification and compared to ergosterol to correlate the toxin production to the amount of fungal mycelium. Fungal growth and mycotoxin production were assessed with both co-inoculation and late inoculation of the BCAs in vitro (three days post-Fusarium inoculation) to test the interaction between the fungus and the bacteria. The level of inhibition of the pathogen and the toxin production were strain-specific. Overall, a higher level of DON inhibition (up to 99%) and a strong reduction in fungal biomass (up to 71%) were achieved when streptomycetes were co-inoculated with the fungus. This research enabled studying the antifungal efficacy of the four Streptomyces strains and monitoring their development in DON-inducing conditions.

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First Report of Leaf Blight of Rice Caused by Cochliobolus lunatus in China

Plant Disease ◽

10.1094/pdis-03-13-0303-pdn ◽

2014 ◽

Vol 98 (5) ◽

pp. 686-686 ◽

Cited By ~ 4

Author(s):

L. M. Liu ◽

S. W. Huang ◽

L. Wang ◽

E. Q. Hou ◽

D. F. Xiao

Keyword(s):

Leaf Blight ◽

Green Water ◽

Tween 20 ◽

Curvularia Lunata ◽

Conidial Suspension ◽

First Report ◽

Rice Plants ◽

Rdna Its ◽

Cochliobolus Lunatus ◽

Wide Range

Leaf-streak symptoms were observed on rice (Oryza sativa L.) starting at the booting stage through harvest in Zhejiang Province, China, in 2012. Based on Fuyang County, only 15% of the rice fields were estimated to show these symptoms. However, incidence could be 40 to 80% when the rice got infected. Typical symptoms started as green water-soaked streaks from the tip or edge of leaf blades, similar to bacterial leaf blight caused by Xanthomonas oryzae. Infected leaves turned yellow, then eventually became wilted and dry. No bacterial streaming was observed and no bacteria were isolated. Pieces of infected leaf tissue were surface sterilized using 0.1% (v/v) mercuric chloride, rinsed with sterilized water, then placed on water agar (WA). After 2 or 3 days on WA at 28°C, only fungal growth was observed from surface sterilized tissues. Fungi were isolated, purified by single spore separation process, and subcultured to potato dextrose agar (PDA) plates. Growing on PDA, the surface of the colony was circular, fluffy, and shiny velvety-black, whereas the under surface was dark Prussian blue. Conidiophores were single or fascicled, brown to dark brown, rarely branched, multiseptate, and straight or often geniculate near the apex. Conidia were brown, smooth, fusiform, geniculate or hook-shaped, 17.5 to 28.5 × 8.5 to 14.0 μm, and 3-septate, with the third cell from the base larger and darker than the others. Molecular identification was performed by analysis of the rDNA internal transcribed spacer region (ITS1-5.8S-ITS2). The rDNA-ITS region was amplified with primer pair ITS1 and ITS4 (5), sequenced, and deposited in GenBank (Accession No. KC462186). The sequence of rDNA-ITS (KC462186) showed 100% identity with Cochliobolus lunatus R.R. Nelson & Haasis (JN943422) after BLAST. Based on the results of morphological and molecular analyses, the fungus isolated from infected leaves was identified as C. lunatus (anamorph: Curvularia lunata (Wakk.) Boedijn) (3). Pathogenicity tests were conducted three times by spraying a conidial suspension (1 × 105 spores/ml) with 0.1% (v/v) Tween 20 on 12 healthy rice plants at late tillering stage. The same number of the healthy rice plants sprayed with sterilized water with 0.1% (v/v) Tween 20 were used as control. All plants were kept at 30°C and 75 to 85% relative humidity (RH) under a 12-h light/dark rotation. About 5 to 7 days after inoculation, green water-soaked streaks began to appear on inoculated plants. From 7 to 14 days after inoculation, the lesions developed quickly and the leaves began to wilt. After 14 days, inoculated plants showed symptoms similar to those originally observed in the field, while control plants (sprayed with sterilized water) remained healthy. C. lunatus was re-isolated from all inoculated plants, and re-identified by the same methods (morphological and molecular methods) as described above, thereby satisfying Koch's postulates, and confirming C. lunatus as the cause of the disease. C. lunatus is a pathogen of a wide range of plants and is common in paddy environments. It was reported as one of the causal agents of black kernel of rice (4) and rice spikelet rot disease (SRD) (1,2). The level of incidence observed in the affected fields suggest that this disease could potentially cause major losses under favorable weather conditions if susceptible cultivars are grown. To our knowledge, this is the first report of C. lunatus causing leaf blight of rice in China. References: (1) S. W. Huang et al. Crop Prot. 30:1, 2011. (2) S. W. Huang et al. Crop Prot. 30:10, 2011. (3) D. S. Manamgoda et al. Fungal Divers. 51:3. (4) S. H. Ou. Rice diseases [M]. CABI, 1985. (5) T. J. White et al. PCR Protocols: a Guide to Methods and Application. Academic Press, San Diego, CA, 1990.

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Fusarium graminearum Isolates from Wheat and Maize in New York Show Similar Range of Aggressiveness and Toxigenicity in Cross-Species Pathogenicity Tests

Phytopathology ◽

10.1094/phyto-07-14-0208-r ◽

2015 ◽

Vol 105 (4) ◽

pp. 441-448 ◽

Cited By ~ 16

Author(s):

Paulo R. Kuhnem ◽

Emerson M. Del Ponte ◽

Yanhong Dong ◽

Gary C. Bergstrom

Keyword(s):

New York ◽

Fusarium Graminearum ◽

Rank Order ◽

Ear Rot ◽

Head Blight ◽

Mycotoxin Production ◽

Pathogenicity Tests ◽

Similar Range ◽

Maize Kernels ◽

Wheat Kernels

This study aimed to assess whether pathogenic Fusarium graminearum isolates from wheat and maize were more aggressive on their host of origin and whether aggressiveness was influenced further by B-trichothecene chemotype. Fifteen isolates were selected from a contemporary collection of isolates surveyed in New York in 2011 to 2012 to represent diversity of host of origin and chemotype. Three pathogenicity assays were used to evaluate and compare these isolates. Fusarium head blight (FHB) severity and trichothecene production in wheat, and maize seedling blight were evaluated in greenhouse inoculation experiments, and Gibberella ear rot (GER) severity and trichothecene production were evaluated in maize ears inoculated in the field. Our results showed among F. graminearum isolates a wide variation in aggressiveness and mycotoxin production toward wheat and maize and these isolates could not be structured by their host of origin or by chemotype. Moreover, aggressiveness rank order changed according to the host/organ evaluated. This indicates that relative susceptibility at the seedling stage may not predict susceptibility of ears. Significant correlations were observed of total trichothecenes (deoxynivalenol [DON] and its acetylated derivatives) produced with FHB and GER severity on wheat and maize, respectively. One isolate did not produce DON or ADON in wheat or maize kernels, yet was aggressive on both hosts. Nine of the fifteen isolates produced small amounts of zearalenone (ZON) in maize kernels, but not in wheat kernels, and ZON level was not correlated with GER severity. F. graminearum isolates from New York showed wide variation in aggressiveness and mycotoxin production toward susceptible wheat and maize. Neither host of origin nor trichothecene chemotype appeared to structure the populations we sampled.

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