First Report of Fusarium venenatum causing foot and root rot of wheat (Triticum aestivum) in Germany

Field experiments were established in the 2018/19 and 2019/20 growing seasons at the experimental station in Neu-Eichenberg, Hessen, Germany to examine the suitability of multiple wheat cultivars for intercropping with pea and the effect of mixtures on diseases. Approximately 600 wheat tillers (BBCH 49-61, depending on the year and cultivar) were sampled in each year and assessed for severity of foot (lower stem) rot symptoms. Fungi from plants with Fusarium-like symptoms exhibiting reddish-brown discolorations on the stems were isolated following the methods described in Šišić et al. (2018). Surface disinfected (3% NaOCl for 10s) wheat stems and roots were cut into three 1 cm long pieces and placed on COONS agar (Coons, 1916). Following 7 – 12 days of incubation under constant blacklight blue fluorescent light, pure cultures were generated on potato dextrose and synthetic nutrient-poor agar (Nirenberg, 1976) using the hyphal tip transfer technique, and the resulting colonies examined microscopically. Based on morphology ca 15% of all Fusarium isolates recovered belonged to a distinct taxonomic unit and were initially identified as F. sambucinum-like (Leslie and Summerell, 2006). The identity of 16 randomly selected isolates (GenBank accession numbers MW085924 - MW085939) was confirmed by sequencing a portion of the translation elongation factor 1-alpha gene region (O’Donnell et al., 1998). Blast analysis in the FUSARIUM-ID (Geiser et al., 2004) and the NCBI databases revealed >99 to 100% identity match with the Fusarium venenatum accession numbers NRRL 22196, FRC R-09186 and MRC 2394. Pathogenicity tests were conducted on wheat cultivar Torborzo using six F. venenatum isolates. Inoculum was prepared using a sterile sand-millet mix infested with six agar plugs of each of the isolates. Once fully colonized, the inoculum was mixed with sterilized sand in a 1:7 ratio (by volume), transferred to 300 ml pots and 4 surface sterilized wheat seeds (5 min 70% alcohol) were sown in each pot. Non-inoculated controls were amended with sterilized inoculum. The experiment was conducted in a greenhouse in a completely randomized design with 5 replicates. Disease symptoms were assessed after four weeks. All isolates induced reddish-brown discolorations on the lower stems similar to those observed in the field grown plants. In addition, the infected plants developed dark brown to black discolorations on the crowns and roots and showed clear signs of stunted root growth. These symptoms were further accompanied by chlorosis (yellowing) of the lower leaves starting from the leaf tip. All isolates were successfully reisolated from the infected wheat plants but not from the controls. To the best of our knowledge, this is the first report of F. venenatum causing foot and root rot of wheat in Germany. Results from field and greenhouse inoculation experiments indicate that F. venenatum may be an important pathogen of wheat in Germany. Further studies on distribution and relative abundance of the species in the Fusarium foot and root rot complex of wheat in Germany are warranted. In addition, it is important to note that our results contrast the results from previous studies (Farr and Rossman, 2021) which reported F. venenatum primarily as a soil saprophyte and presumably non-pathogenic fungus in its nature. Our findings thus, also warrant the need to further investigate pathogenic potential of this species and the role it may play on other common rotational crops in Germany.

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First Report of Calonectria hongkongensis Causing Fruit Rot of Rambutan (Nephelium lappaceum)

Plant Disease ◽

10.1094/pdis-01-13-0008-pdn ◽

2013 ◽

Vol 97 (8) ◽

pp. 1117-1117 ◽

Cited By ~ 2

Author(s):

L. M. Serrato-Diaz ◽

E. I. Latoni-Brailowsky ◽

L. I. Rivera-Vargas ◽

R. Goenaga ◽

P. W. Crous ◽

...

Keyword(s):

Mycelial Growth ◽

Elongation Factor ◽

Type Specimen ◽

Fruit Rot ◽

Fluorescent Light ◽

Mycelium Growth ◽

Tropical Fruit ◽

First Report ◽

Nephelium Lappaceum ◽

Pure Cultures

Fruit rot of rambutan is a pre- and post-harvest disease problem of rambutan orchards. In 2011, fruit rot was observed at USDA-ARS orchards in Mayaguez, Puerto Rico. Infected fruit were collected and 1 mm2 tissue sections were surface disinfested with 70% ethanol followed by 0.5% sodium hypochlorite. Infected fruit were rinsed with sterile, deionized, double-distilled water and transferred to acidified potato dextrose agar (APDA). Plates were incubated at 25 ± 1°C for 6 days. Three isolates of Calonectria hongkongensis (Cah), CBS134083, CBS134084, and CBS134085, were identified morphologically using taxonomic keys (2,3). In APDA, colonies of Cah produced raw sienna to rust-colored aerial mycelial growth. Conidiophores of Cah had a penicillate arrangement of primary to quaternary branches of 2 to 6 phialides. Conidia (n = 50) were cylindrical, hyaline, 1-septate, rounded at both ends, and 44 to 52 μm × 3.5 to 4.5 μm. Conidiophores produced terminal and lateral stipe extensions with terminal sphaeropedunculate vesicles that were 8 to 12 μm wide. Subglobose to ovoid perithecia, 300 to 500 μm × 200 to 350 μm and orange to red-brown, were produced in groups of 3. Asci were clavate and contained 8 ascospores aggregated at the top of the ascus. Ascospores (n = 50) were hyaline, guttulate, fusoid with rounded ends, straight to curved, 1-septate with constriction at the septum, and 28 to 36 μm × 4 to 7 μm. For molecular identification, the ITS rDNA, fragments of β-tubulin (BT), histone H3 (HIS3), and elongation factor (EF1-α) genes were amplified by PCR, sequenced, and compared using BLASTn with Calonectria spp. submitted to the NCBI GenBank. The sequences of Cah submitted to GenBank include accessions KC342208, KC342206, and KC342207 for ITS; KC342217, KC342215, and KC342216 for BT; KC342211, KC342209, and KC342210 for HIS3; and KC342214, KC342212, and KC342213 for EF1α. The sequences were >99% or identical with the ex-type specimen of Cah CBS 114828 for all genes used. Pathogenicity tests were conducted on 5 healthy superficially sterilized fruits per isolate. Both scalpel-wounded and unwounded fruit tissues were inoculated with 5-mm mycelial disks from 8-day-old pure cultures grown in APDA. Untreated controls were inoculated with APDA disks only. Fruits were kept in a humid chamber for 8 days at 25°C under 12 h of fluorescent light. The test was repeated once. Three days after inoculation (DAI), white mycelial growth was observed on the fruit. Five DAI, the fruit changed color from red to brown and yellowish mycelia colonized 50 to 62% of the fruit surface. Eight DAI, all the fruit turned brown, the mycelium growth covered the entire fruit, and conidiophores were produced on spinterns (hairlike appendages). Fruit rot of spinterns, exocarp (skin), endocarp (aril), and light brown discoloration were observed inside the fruit. Untreated controls showed no symptoms of fruit rot and no fungi were reisolated from tissue. Cah was reisolated from diseased tissue, fulfilling Koch's postulates. Calonectria spp. (or their Cylindrocladium asexual states) have been associated with lychee decline syndrome in North Vietnam (1). Both fruits belong to the Sapindaceae family. To our knowledge, this is the first report of Cah causing fruit rot of rambutan. References: (1) L. M. Coates et al. Diseases of Longan, Lychee and Rambutan. Pages 307-325 in: Diseases of Tropical Fruit Crops. R. C. Ploetz, ed. CABI Publishing, Cambridge, MA, 2003. (2) P. W. Crous. Taxonomy and Pathology of Cylindrocladium (Calonectria) and Allied Genera. APS Press, St Paul, MN, 2002. (3) P. W. Crous, et al. Stud. Mycol. 50:415, 2004.

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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 Crown and Root Rot Caused by Rhizoctonia solani AG-4 on Orange Jessamine in Italy

Plant Disease ◽

10.1094/pdis-93-2-0204b ◽

2009 ◽

Vol 93 (2) ◽

pp. 204-204 ◽

Cited By ~ 1

Author(s):

D. Aiello ◽

A. Vitale ◽

E. Lahoz ◽

R. Nicoletti ◽

G. Polizzi

Keyword(s):

Rhizoctonia Solani ◽

Root Rot ◽

Morphological Characteristics ◽

Ruthenium Red ◽

Fluorescent Light ◽

First Report ◽

Pectic Enzymes ◽

Electrophoretic Patterns ◽

Crown And Root Rot ◽

Hyphal Diameter

Murraya paniculata (L.) Jack, commonly called orange jessamine or orange jasmine (Rutaceae), is a small tropical tree that is native to Asia. This species, closely related to Citrus, is grown as an ornamental tree or hedge. During October of 2007, crown and root rot was observed on approximately 12,000 pot-grown, 4-month-old plants in a nursery in eastern Sicily, Italy. Basal leaves turned yellow and gradually became necrotic, and infected plants often died. Disease symptoms were observed on 1,800 (15%) plants. Isolations from affected tissues on potato dextrose agar (PDA) amended with streptomycin sulfate at 100 mg/liter recovered a fungus with mycelial and morphological characteristics consistent with Rhizoctonia solani Kühn. Fungal colonies were initially white, turned brown with age, and produced irregularly shaped, brown sclerotia. Microscopic examination revealed that hyphae had a right-angle branching pattern, were constricted at the base of the branch near the union with main hyphae, and were septate near the constriction. The nuclear condition of hyphal cells was determined on cultures grown at 25°C on 2% water agar (WA) when stained with 3% safranin O solution and examined at ×400. Anastomosis groups were determined by pairing isolates on 2% WA in petri plates (4). Pairings were made with tester strains AG-1 IA, AG-2-2-1, AG-2-2IIIB, AG-2-2IV, AG-3, AG-4, AG-5, AG-6, and AG-11. Anastomosis was observed only with tester isolates of AG-4 producing both C2 and C3 reactions. The hyphal diameter at the point of anastomosis was reduced, the anastomosis point was obvious, and cell death of adjacent cells was observed. These results were consistent with other reports on anastomosis reactions (1). The identification of group AG-4 within R. solani has been confirmed by electrophoretic patterns of pectic enzymes (polygalacturonases) in vertical pectin-acrylamide gel stained with ruthenium red (2). Pathogenicity tests were conducted on potted, healthy, 6-month-old seedlings of orange jessamine. Twenty-five plants were inoculated by placing 1-cm2 PDA plugs from 5-day-old mycelial cultures near the base of the stem. The same number of plants inoculated with PDA plugs served as controls. Plants were maintained at 25°C and 95% relative humidity on a 12-h fluorescent light/dark regimen. Wilt symptoms, identical to ones observed in the nursery, developed 3 months after inoculation because of crown and root rot. Control plants remained disease free. The pathogen was reisolated from symptomatic tissues, completing Koch's postulates. Collar rot due to R. solani was previously detected on M. koenigii (3). To our knowledge, this is the first report of R. solani causing disease on M. paniculata. References: (1) D. E. Carling. Page 37 in: Grouping in Rhizoctonia solani by Hyphal Anastomosis Reactions. Kluwer Academic Publishers, the Netherlands, 1996. (2) R. H. Cruickshank and G. C. Wade. Anal. Biochem. 107:177, 1980. (3) A. C. Jain and K. A. Mahmud. Rev. Appl. Mycol. 32:460, 1953. (4) C. C. Tu and J. W. Kimbrough. Mycologia 65:941, 1973.

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First Report of Root Rot of Soybeans Caused by Corynespora cassiicola in Wisconsin

Plant Disease ◽

10.1094/pdis.1999.83.7.696b ◽

1999 ◽

Vol 83 (7) ◽

pp. 696-696 ◽

Cited By ~ 5

Author(s):

S. J. Raffel ◽

E. R. Kazmar ◽

R. Winberg ◽

E. S. Oplinger ◽

J. Handelsman ◽

...

Keyword(s):

Root Rot ◽

Growth Stage ◽

Lateral Roots ◽

Seedling Emergence ◽

Fluorescent Light ◽

Corynespora Cassiicola ◽

First Report ◽

Seedling Roots ◽

Symptomatic Tissue ◽

Soybean Plants

Corynespora cassiicola (Berk. & M. A. Curtis) C. T. Wei was isolated from diseased soybean plants (Glycine max) collected in two fields near Racine and Arlington, WI. Plants sampled at seedling emergence (VC), late vegetative (V5), and mid-reproductive (R5) stages exhibited reddish to dark brown longitudinal lesions on the exterior of the tap root extending vertically on the hypocotyl to the soil line, and extensive necrosis of lateral roots. Sample size at each growth stage was 144 plants per site. Roots were surface sterilized in 0.5% sodium hypochlorite for 2 min and sections of symptomatic tissue placed on water agar (12 g/liter) containing 100 μg of streptomycin per ml. Sporulation occurred on lesions and on mycelium that had grown out from the plant tissue onto the water agar following a 2-week incubation at 24°C under fluorescent light (280 μmol s-1 m-2). Incidence of isolation of C. cassiicola at both sites was 40% of plants sampled at growth stage VC, 67% at V5, and 78% at R5. Conidia characteristic of C. cassiicola were particularly abundant on the surface of necrotic lateral root tissue. Elongated conidia produced on water agar were 151 ± 5 μm × 15 ± 0.5 μm with an average of 13 ± 0.4 cells separated by hyaline pseudosepta (1). To confirm pathogenicity, a 1-cm lateral slice into each of four 5-day-old soybean seedling roots was made and a plug of agar taken from the margin of a colony of C. cassiicola grown on potato dextrose agar was placed in each wound and incubated for 14 days at 24°C in a growth chamber. Symptoms similar to those of diseased field plants were observed and C. cassiicola was reisolated from all plants inoculated with C. cassiicola; all controls treated with agar alone had no symptoms and C. cassiicola was recovered from none of the noninoculated controls. This is the first report of root rot caused by C. cassiicola on soybean in Wisconsin. Reference: (1) W. L. Seaman and R. A. Shoemaker. Can. J. Bot. 43:1461, 1965.

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First report of Nigrospora sphaerica causing fruit dried-shrink disease in Akebia trifoliata from China

Plant Disease ◽

10.1094/pdis-11-20-2471-pdn ◽

2021 ◽

Author(s):

Xiujing Hong ◽

Shijia Chen ◽

linchao Wang ◽

Bo Liu ◽

Yuruo Yang ◽

...

Keyword(s):

Elongation Factor ◽

Morphological Characteristics ◽

Its Region ◽

Average Diameter ◽

Ethanol Solution ◽

Likelihood Method ◽

Conidial Suspension ◽

Multiple Sequence ◽

First Report ◽

Pure Cultures

Akebia trifoliata, a recently domesticated horticultural crop, produces delicious fruits containing multiple nutritional metabolites and has been widely used as medicinal herb in China. In June 2020, symptoms of dried-shrink disease were first observed on fruits of A. trifoliata grown in Zhangjiajie, China (110.2°E, 29.4°N) with an incidence about 10%. The infected fruits were shrunken, colored in dark brown, and withered to death (Figure S1A, B). The symptomatic fruits tissues (6 × 6 mm) were excised from three individual plants, surface-disinfested in 1% NaOCl for 30s and 70% ethanol solution for 45s, washed, dried, and plated on potato dextrose agar (PDA) containing 50 mg/L streptomycin sulfate in the dark, and incubated at 25℃ for 3 days. Subsequently, hyphal tips were transferred to PDA to obtain pure cultures. After 7 days, five pure cultures were obtained, including two identical to previously reported Colletotrichum gloeosporioides causing leaf anthracnose in A. trifoliata (Pan et al. 2020) and three unknown isolates (ZJJ-C1-1, ZJJ-C1-2, and ZJJ-C1-3). The mycelia of ZJJ-C1-1, ZJJ-C1-2 and ZJJ-C1-3 were white, and formed colonies of approximate 70 mm (diameter) in size at 25℃ after 7 days on potato sucrose agar (PSA) plates (Figure S1C). After 25 days, conidia were formed, solitary, globose, black, shiny, smooth, and 16-21 μm in size (average diameter = 18.22 ± 1.00 μm, n = 20) (Figure S1D). These morphological characteristics were similar to those of N. sphaerica previously reported (Li et al. 2018). To identify species of ZJJ-C1-1, ZJJ-C1-2 and ZJJ-C1-3, the internal transcribed spacer (ITS) region, β-tubulin (TUB2), and the translation elongation factor 1-alpha (TEF1-α) were amplified using primer pairs including ITS1/ITS4 (Vilgalys and Hester 1990), Bt-2a/Bt-2b (Glass and Donaldson 1995), and EF1-728F/EF-2 (Zhou et al. 2015), respectively. Multiple sequence analyses showed no nucleotide difference was detected among genes tested except ITS that placed three isolates into two groups (Figure S2). BLAST analyses determined that ZJJ-C1-1, ZJJ-C1-2 and ZJJ-C1-3 had 99.73% to N. sphaerica strains LC2705 (KY019479), 100% to LC7294 (KY019397), and 99.79-100% to LC7294 (KX985932) or LC7294 (KX985932) based on sequences of TUB2 (MW252168, MW269660, MW269661), TEF-1α (MW252169, MW269662, MW269663), and ITS (MW250235, MW250236, MW192897), respectively. These indicated three isolates belong to the same species of N. sphaerica. Based on a combined dataset of ITS, TUB2 and TEF-1α sequences, a phylogenetic tree was constructed using Maximum likelihood method through IQ-TREE (Minh et al. 2020) and confirmed that three isolates were N. sphaerica (Figure S2). Further, pathogenicity tests were performed. Briefly, healthy unwounded fruits were surface-disinfected in 0.1% NaOCl for 30s, washed, dried and needling-wounded. Then, three fruits were inoculated with 10 μl of conidial suspension (1 × 106 conidia/ml) derived from three individual isolates, with another three fruits sprayed with 10 μl sterilized water as control. The treated fruits were incubated at 25℃ in 90% humidity. After 15 days, all the three fruits inoculated with conidia displayed typical dried-shrink symptoms as those observed in the farm field (Figure S1E). The decayed tissues with mycelium and spores could be observed on the skin or vertical split of the infected fruits after 15 days’ inoculation (Figure S1F-H). Comparably, in the three control fruits, there were no dried-shrink-related symptoms displayed. The experiment was repeated twice. The re-isolated pathogens were identical to N. sphaerica determined by sequencing the ITS, TUB2 and TEF-1α. Previous reports showed N. sphaerica could cause postharvest rot disease in kiwifruits (Li et al. 2018). To our knowledge, this is the first report of N. sphaerica causing fruits dried-shrink disease in A. trifoliata in China.

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First Report on the Presence of Leptosphaeria maculans Pathogenicity Group-3, the Causal Agent of Blackleg of Canola in Manitoba

Plant Disease ◽

10.1094/pdis.2003.87.10.1268a ◽

2003 ◽

Vol 87 (10) ◽

pp. 1268-1268 ◽

Cited By ~ 13

Author(s):

W. G. D. Fernando ◽

Y. Chen

Keyword(s):

Leptosphaeria Maculans ◽

Fluorescent Light ◽

Sodium Hypochlorite Solution ◽

Distilled Water ◽

Brassica Napus L ◽

First Report ◽

Pure Cultures ◽

Pathogenic Variability ◽

Serious Disease ◽

Highly Virulent

Blackleg, caused by Leptosphaeria maculans (Desmaz.) Ces. & De Not. (anamorph = Phoma lingam) (Tode:Fr.) Desmaz.), is an economically important and serious disease of canola (Brassica napus L.) in Australia, Europe, and Canada. L. maculans isolates can be categorized into four pathogenicity groups (PGs) on the basis of the interaction phenotypes (IP) on the differential canola cvs. Westar, Glacier, and Quinta (1) by using a standard screening protocol in the greenhouse. PG1 isolates are weakly virulent and PG2, PG3, and PG4 isolates are highly virulent. In Manitoba, L. maculans population consists mainly of PG2 (virulent on cv. Westar; avirulent on cvs. Glacier and Quinta) and a few PG1 isolates (avirulent on all three differentials). The Oilseed Pathology Lab in the Department of Plant Science, University of Manitoba examines the pathogenic variability of blackleg isolates obtained from Manitoba each year. In 2002, the blackleg-resistant cv. Q2, was found to be severely infected in Roland, Manitoba. The canola stubble collected from a coop trial plot (Roland, Manitoba) and a farm in East Selkirk (60 km northeast of Winnipeg, Manitoba) was isolated for the blackleg fungus. Small pieces of stubble were cut from the pseudothecia forming section and surface sterilized with 1% sodium hypochlorite solution for 3 to 5 min and then rinsed in sterile distilled water. V8 agar medium containing 1% streptomycin sulphate was used to culture the isolates under continuous cool-white fluorescent light for 14 days. Pure cultures of the pathogen were isolated and characterized as L. maculans by means of colony morphology, pycnidia, and microscopic observations of pycnidiospores. Pycnidiospores that formed on V8 plates were flooded with 10 ml of sterile distilled water and then harvested by filtering through sterilized Miracloth and kept at -20°C. The isolates were passed once through cv. Westar to maintain their virulence. The PG test was performed with the three differential cultivars. Two additional cultivars, Q2 (resistant to PG2 isolates) and Defender (moderately resistant to PG2 isolates), were included for comparisons. Twelve 7-day-old cotyledons of each differential cultivar grown in Metro Mix were wound inoculated with a 10-μl droplet of pycnidiospore suspension (1 × 107 pycnidiospores per ml). Inoculated cotyledons were maintained in the greenhouse (16/21°C night/day and a 16-h photoperiod). The experiment was repeated twice. Disease severity on cotyledons was assessed 12 days postinoculation by using a 0 to 9 scale (2). All five isolates from Roland and East Selkirk were highly virulent on Glacier (6.4 to 7.7), Q2 (7.1 to 8.2), and Defender (7.2 to 8.4), but intermediately virulent on Quinta (4.5 to 5.4). This clearly indicated that these isolates were of PG3. Isolates of PG2 have been predominant in Manitoba for the past 25 years, and highly virulent isolates belonging to PG3 had not been detected previously. To our knowledge, this is the first report of the presence of PG3 in L. maculans in Manitoba. References: (1) A. Mengistu et al. Plant Dis. 75:1279, 1991. (2) P. H. Williams. Crucifer Genetics Cooperatives (CrGC) Resource Book, University of Wisconsin—Madison, 1985.

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First Report of Root Rot Caused by Sclerotium rolfsii on Daucus carota var. sativa in Southern Korea

Plant Disease ◽

10.1094/pdis-03-11-0233 ◽

2011 ◽

Vol 95 (12) ◽

pp. 1585-1585

Author(s):

J.-H. Kwon ◽

Y. H. Lee ◽

H.-S. Shim ◽

J. Kim

Keyword(s):

Daucus Carota ◽

Root Rot ◽

Pathogenic Fungus ◽

Sclerotium Rolfsii ◽

Agricultural Science ◽

Control Treatment ◽

Pathogenicity Test ◽

Optimum Temperature ◽

First Report ◽

Causal Fungus

Carrot (Daucus carota var. sativa DC.), an important root vegetable, is cultivated widely because of its dietary fiber and beta carotene. In June 2009 and June 2010, a disease suspected as root rot of carrot caused by Sclerotium rolfsii occurred in a 5-ha field in Jinju, Korea. Early symptoms consisted of water-soaked lesions on root and lower stem tissue near the soil line. Infected plants gradually withered and white mycelial mats appeared on the surface of roots. Numerous sclerotia were often produced on stem and root surfaces in contact with the soil. The heavily infected carrots became rotted and blighted and the whole plant eventually died. The freshly isolated pathogenic fungus was grown on potato dextrose agar (PDA) and examined microscopically. Optimum temperature for mycelia growth or sclerotia formation was 25 to 30°C. Numerous globoid sclerotia formed on the PDA after 18 days of mycelial growth. The sclerotia (1 to 3 mm in diameter) were white at first and then gradually turned dark brown. Aerial mycelia usually formed, consisting of many narrow hyphal strands 3 to 9 μm wide. The white mycelium formed a typical clamp connection after 5 days of growth at optimum temperature. To fulfill Koch's postulates, 10 carrot seedlings were inoculated with colonized agar discs (6 mm in diameter) of the causal fungus directly on the root and incubated in a humid chamber at 25°C for 24 h. Ten carrot seedlings were inoculated similarly with agar discs as the control treatment. After this period, the inoculated and noninoculated plants were maintained in a greenhouse. Eight days after inoculation, the disease symptoms seen in the field were reproduced and the fungus was reisolated from the artificially inoculated plants. To confirm identity of the causal fungus, the complete internal transcribed spacer (ITS) rDNA region of the causal fungus was amplified using the primers ITS1 and ITS4 (2) and sequenced. The resulting sequence of 684 bp was deposited in GenBank (Accession No. JF342557). The sequence was 99% similar to sequences of Athelia rolfsii (Sclerotium rolfsii) in GenBank. Cultures of S. rolfsii have been deposited with the Korean Agricultural Culture Collection (KACC 45154), National Academy of Agricultural Science, Korea. On the basis of symptoms, fungal colonies, the ITS sequence, and the pathogenicity test on the host plant, this fungus was identified as S. rolfsii Saccardo (1). To our knowledge, this is the first report of root rot of carrot caused by S. rolfsii in Korea. This disease is highly dependent upon environmental conditions, including warm weather and high humidity. Recent occurrence of the disease suggests that S. rolfsii could spread widely. References: (1) J. E. M. Mordue. CMI Descriptions of Pathogenic Fungi and Bacteria. No. 410, 1974. (2) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, Inc., New York, 1990.

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First Report of Diaporthe amygdali Associated with Twig Canker and Shoot Blight of Nectarine in Spain

Plant Disease ◽

10.1094/pdis-10-20-2283-pdn ◽

2021 ◽

Author(s):

Francisco Beluzán ◽

Diego Olmo ◽

Maela León ◽

Paloma Abad-Campos ◽

Josep Armengol

Keyword(s):

Prunus Persica ◽

Tap Water ◽

Elongation Factor ◽

Its Region ◽

Sodium Hypochlorite Solution ◽

First Report ◽

Potted Plants ◽

Randomized Design ◽

Shoot Blight ◽

One Year

Nectarine (Prunus persica (L.) Batsch var. nucipersica (Suckow) C. K. Schneid.) is a fruit crop widely cultivated throughout the Mediterranean basin. In Spain, it is mainly grown in eastern regions of the country. In March 2018, 5-year-old nectarine trees showing twig canker symptoms were observed after a rainy spring period in a 0.5 ha orchard located at Alaior, Menorca island (Spain). Cankers were frequent on affected trees (approximately, 80% of the total trees), thus leading to shoot blight. Ten twig segments of one-year old wood with cankers were cut, washed under running tap water, surface disinfected for 1 min in a 1.5% sodium hypochlorite solution and rinsed twice in sterile distilled water. Small pieces (2 mm) of affected tissues were taken from the margin of the cankers and plated on potato dextrose agar (PDA) supplemented with 0.5 g/L of streptomycin sulphate (PDAS). The plates were then incubated at 25 ºC in the dark for 7 to 10 d. Actively growing colonies were first hyphal-tipped and then transferred to PDA and 2% water agar supplemented with sterile pine needles and incubated at 21-22ºC under a 12h/12h near UV / darkness cycle during 21 d (León et al. 2020). Colonies were white at first, becoming light cream, with visible solitary and aggregate pycnidia at maturity. Alpha conidia were aseptate, fusiform, hyaline, multi-guttulated (mean ± SD = 7.4 ± 0.7 × 2.8 ± 0.4 µm, n = 100). Beta and gamma conidia were not observed. The morphological and cultural characteristics of the isolates were congruent with those of Diaporthe spp. (Gomes et al. 2013). The ITS1-5.8S-ITS2 (ITS) region and fragments of β-tubulin (tub2), the translation elongation factor 1-alpha (tef1-α) gene regions, histone H3 (his3) and calmodulin (cal) genes of representative isolate DAL-59 were amplified and sequenced (Santos et al. 2017). The BLASTn analysis revealed 100% similarity with sequences of D. mediterranea (Synonym D. amygdali) (Hilário et al. 2021) isolate DAL-34 from almond (ITS: MT007489, tub2: MT006686, tef1-α: MT006989, his3: MT007095 and cal: MT006761). Sequences of isolate DAL-59 were deposited in GenBank Database (ITS: MT007491, tub2: MT006688, tef1-α: MT006991, his3: MT007097 and cal: MT006763). Pathogenicity tests were conducted using one-year-old potted plants of nectarine cv. Boreal, which were inoculated with isolate DAL-59. In each plant, a 3 mm wound was made in the center of the main branch (about 30 cm length) with a scalpel. Colonized agar plugs with 3 mm diameter, which were obtained from active 10-day-old colonies growing on PDA, were inserted underneath the epidermis and the wounds sealed with Parafilm. Inoculated plants were incubated in a growth chamber at 23 ºC with 12 h of light per day. Controls were inoculated with uncolonized PDA plugs. There were twelve plants per treatment, which were arranged in a completely randomized design. Five days after inoculation necrosis development was observed in the area of inoculation. Wilting and twig blight symptoms over the lesion occurred 3-wk after inoculation and pycnidia were detected, while the controls remained asymptomatic. Diaporthe amygdali was re-isolated from symptomatic tissues and identified as described above to satisfy Koch’s postulates. To our knowledge, this is the first report of D. amygdali causing twig canker and shoot blight disease on nectarine in Spain.

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First Report of Fusarium Wilt of Lettuce Caused by Fusarium oxysporum f. sp. lactucae in Arizona

Plant Disease ◽

10.1094/pdis.2003.87.10.1265c ◽

2003 ◽

Vol 87 (10) ◽

pp. 1265-1265 ◽

Cited By ~ 23

Author(s):

M. E. Matheron ◽

S. T. Koike

Keyword(s):

Fusarium Oxysporum ◽

Root Rot ◽

The United States ◽

San Joaquin Valley ◽

Fluorescent Light ◽

Lettuce Seedling ◽

Disease Symptoms ◽

First Report ◽

Root Rot Disease ◽

Rot Disease

A new wilt and root rot disease was observed in 6 and 11 commercial fields of lettuce (Lactuca sativa) in western Arizona during the fall of 2001 and 2002, respectively. Distance between infested sites ranged from approximately 0.5 to 39 km. Five head lettuce cultivars as well as a red leaf lettuce cultivar were affected. Disease symptoms included yellowing and wilting of leaves, as well as stunting and plant death. The cortex of the crown and upper root of infected plants usually was decayed and reddish brown. Disease symptoms first appeared at the time of plant thinning and continued to develop up to plant maturity. Fusarium oxysporum was consistently isolated from symptomatic plant roots. Seeds of cv. Lighthouse were planted in nonsterile vermiculite within 3.0-cm-square × 7.0-cm-deep cells in a transplant tray and thinned to a single plant per cell. When the first true leaves were emerging, 10 individual seedlings were inoculated with a single-spore isolate of F. oxysporum recovered from diseased lettuce root cortex tissue. Inoculum was prepared by growing the fungus on potato dextrose agar in 100-mm-diameter × 15-mm-deep plastic petri dishes at 28°C with a 12-h photoperiod under fluorescent light. Once the fungus completely covered the agar surface, 50 ml of sterile distilled water was added to the dish, and the mycelia and conidia on the surface were scraped off the agar and suspended in the water. This fungal suspension was decanted, and a 2-ml aliquot containing 1.8 × 105 CFU was pipetted into the vermiculite near the stem of each lettuce seedling. Ten plants grown in noninfested vermiculite served as uninoculated controls. After inoculation, plants were maintained in a growth chamber at 28°C with a 12-h photoperiod under fluorescent light for 3 weeks. Symptoms of yellowing, wilt, vascular decay, and often plant death developed during the incubation period on all inoculated plants but not on control plants. Fusarium oxysporum was consistently reisolated from inoculated plants but not from uninoculated plants. The experiment was repeated and yielded the same results. A wilt and root rot disease of lettuce attributed to F. oxysporum f. sp. lactucae was first reported in Japan in 1967 (3) and subsequently in the United States (San Joaquin Valley of California) in 1993 (2), and Italy in 2002 (1). The researchers of the U.S. report did not cite the earlier work from Japan and described the pathogen as F. oxysporum f. sp. lactucum. The Arizona isolate used to demonstrate pathogenicity was of the same vegetative compatibility group as an isolate of the pathogen from lettuce in California reported in 1993. Several companies grow and harvest lettuce in Arizona and California. At the end of production and harvest in the fall, tractors, implements, and harvesting equipment are transported from the San Joaquin Valley in California to western Arizona. The similarity between the isolate of F. oxysporum f. sp. lactucae from western Arizona and the San Joaquin Valley of California suggest a possible introduction of the pathogen into Arizona from California, perhaps on soil adhering to farm equipment. To our knowledge, this is the first report of F. oxysporum f. sp. lactucae infecting lettuce in Arizona. References: (1) A. Garibaldi et al. Plant Dis. 86:1052, 2002. (2) J. C. Hubbard and J. S. Gerik. Plant Dis. 77:750, 1993. (3) T. Matuo and S. Motohashi. Trans. Mycol. Soc. Jpn. 8:13, 1967.

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Macrophomina vaccinii Causes a Basal Stem and Root Rot of Patchouli (Pogostemon cablin) in China

Plant Disease ◽

10.1094/pdis-06-21-1300-pdn ◽

2021 ◽

Author(s):

Ying Xin Fang ◽

Jin Ling Li ◽

Xiao Fei Li ◽

Min Zhong Chen ◽

Jia Yi Lin ◽

...

Keyword(s):

Root Rot ◽

Elongation Factor ◽

Morphological Characteristics ◽

Perennial Herb ◽

Bootstrap Support ◽

Molecular Characteristics ◽

Plastic Bags ◽

Pogostemon Cablin ◽

Pure Cultures ◽

Rot Disease

Patchouli (Pogostemon cablin Benth.) is a perennial herb native to South and South East Asia, which widely used as a traditional herbal medicine against indigestion, diarrhea, cold, fever, vomiting and headache in China (Swamy and Sinniah, 2015). In September 2020, a new basal stem and root rot disease of patchouli was observed in three cultivated fields of Zhanjiang City, Guangdong Province. Symptoms included sudden discoloration, chlorosis and wilting of the leaves and severe rot associated with external and internal browning at the basal part of stems and roots that results in the death of approximately 2 to 5% of plants in each field. To determine the causal agent, symptomatic roots with typical lesions were cut into small pieces, then surface sterilized in 2.5% NaClO for 1 min, rinsed three times in distilled water, and then inoculated on potato dextrose agar (PDA) medium. A fungus with same morphological characteristics was consistently isolated from disease tissue. The mycelia initially white to cream, later turning pale brown to black with age on PDA. To induce the sporulation, the isolated fungi were transferred to synthetic nutrient-poor agar (SNA) with autoclaved pine needles and ten pure cultures were obtained by single spores. Masses of black, hard and oblong microsclerotia (av. 171.2×136.5 µm, n=50) were observed on SNA. Conidia hyaline, ellipsoid to obovoid, smooth, enclosed in a mucous sheath, 15.9~32.5×7.7~11.9 µm (av. 23.9 × 9.6 µm, n=100). The internal transcribed spacer (ITS) regions as well as the partial translation elongation factor (EF-1α) and β-tubulin (TUB) genes of two representative isolates GHX-1 and CHX-2 were sequenced, using primer pairs ITS1/ITS4 (White et al. 1990), EF-688F/EF-986R (Carbone and Kohn 1999), and Bt2a/Bt2b (Glass and Donaldson 1995), respectively. The obtained sequences were submitted to GenBank under the following accession numbers: ITS, MZ375360 and MZ375361; EF-1α, MZ393804 and MZ393805; and TUB, MZ393806 and MZ393807. BLASTn searches revealed that 100% identity with the existing sequences of ex-type culture CGMCC3.19503 of Macrophomina vaccinii (ITS, MK687450; EF-1α, MK687426; and TUB, MK687434), respectively. Phylogenetic analysis using Neighbour-Joining method in Mega7.0 with concatenated sequences of ITS, EF-1α and TUB showed that the isolates clustered in the clade of M. vaccinii with high bootstrap support values. Based on both of the morphological and molecular results, the isolates were identified as M. vaccinii (Zhao et al., 2019). To confirm the pathogenicity of the two isolates, 5-mm-diameter mycelial agar plus from the margins of 5-day-old PDA cultures were placed on the wounded basal stem of 2-month-old patchouli seedlings in each pot. Five separate pots were used for each isolate and plants inoculated with sterile agar plus were served as controls. All plants were covered with plastic bags to maintain 90% relative humidity and kept at 26°C with a photoperiod of 12 h in a greenhouse conditions. The patchouli plants developed similar necrotic symptoms on basal stem and root to those observed in the field after 3 weeks after inoculation, whereas the control plants were asymptomatic. M. vaccinii was reisolated from inoculated plants and identify by morphological and molecular characteristics, fulfill the Koch’s postulates and identify. Recently, stem blight on blueberry caused by M. vaccinii was reported in China (Zhao et al. 2019). To our knowledge, this is the first report of M. vaccinii causing basal stem and root rot on patchouli in China and worldwide.

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