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

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 Fusarium falciforme (FSSC 3+4) causing root rot on chickpea in Mexico

Plant Disease ◽

10.1094/pdis-05-21-1123-pdn ◽

2021 ◽

Author(s):

Sixto Velarde Felix ◽

Victor Valenzuela ◽

Pedro Ortega ◽

Gustavo Fierros ◽

Pedro Rojas ◽

...

Keyword(s):

Fusarium Solani ◽

Root Rot ◽

Species Complex ◽

Elongation Factor ◽

Morphological Characteristics ◽

Aerial Mycelium ◽

Pathogenicity Test ◽

Conidial Suspension ◽

Single Spore ◽

First Report

Chickpea (Cicer aretinium L.) is a legume crop of great importance worldwide. In January 2019, wilting symptoms on chickpea (stunted grow, withered leaves, root rot and wilted plants) were observed in three fields of Culiacan Sinaloa Mexico, with an incidence of 3 to 5%. To identify the cause, eighty symptomatic chickpea plants were sampled. Tissue from roots was plated on potato dextrose agar (PDA) medium. Typical Fusarium spp. colonies were obtained from all root samples. Ten pure cultures were obtained by single-spore culturing (Ff01 to Ff10). On PDA the colonies were abundant with white aerial mycelium, hyphae were branched and septae and light purple pigmentation was observed in the center of old cultures (Leslie and Summerell 2006). From 10-day-old cultures grown on carnation leaf agar medium, macroconidias were falciform, hyaline, with slightly curved apexes, three to five septate, with well-developed foot cells and blunt apical cells, and measured 26.6 to 45.8 × 2.2 to 7.0 μm (n = 40). The microconidia (n = 40) were hyaline, one to two celled, produced in false heads that measured 7.4 to 20.1 (average 13.7) μm × 2.4 to 8.9 (average 5.3) μm (n = 40) at the tips of long monophialides, and were oval or reniform, with apexes rounded, 8.3 to 12.1 × 1.6 to 4.7 μm; chlamydospores were not evident. These characteristics fit those of the Fusarium solani (Mart.) Sacc. species complex, FSSC (Summerell et al. 2003). The internal transcribed spacer and the translation elongation factor 1 alpha (EF1-α) genes (O’Donnell et al. 1998) were amplified by polymerase chain reaction and sequenced from the isolate Ff02 and Ff08 (GenBank accession nos. KJ501093 and MN082369). Maximum likelihood analysis was carried out using the EF1-α sequences (KJ501093 and MN082369) from the Ff02 and Ff08 isolates and other species from the Fusarium solani species complex (FSSC). Phylogenetic analysis revealed the isolate most closely related with F. falciforme (100% bootstrap). For pathogenicity testing, a conidial suspension (1x106 conidia/ml) was prepared by harvesting spores from 10-days-old cultures on PDA. Twenty 2-week-old chickpea seedlings from two cultivars (P-2245 and WR-315) were inoculated by dipping roots into the conidial suspension for 20 min. The inoculated plants were transplanted into a 50-hole plastic tray containing sterilized soil and maintained in a growth chamber at 25°C, with a relative humidity of >80% and a 12-h/12-h light/dark cycle. After 8 days, the first root rot symptoms were observed on inoculating seedlings and the infected plants eventually died within 3 to 4 weeks after inoculation. No symptoms were observed plants inoculated with sterilized distilled water. The fungus was reisolated from symptomatic tissues of inoculated plants and was identified by sequencing the partial EF1-α gene again and was identified as F. falciforme (FSSC 3 + 4) (O’Donnell et al. 2008) based on its morphological characteristics, genetic analysis, and pathogenicity test, fulfilling Koch’s postulates. The molecular identification was confirmed via BLAST on the FusariumID and Fusarium MLST databases. Although FSSC has been previously reported causing root rot in chickpea in USA, Chile, Spain, Cuba, Iran, Poland, Israel, Pakistan and Brazil, to our knowledge this is the first report of root rot in chickpea caused by F. falciforme in Mexico. This is important for chickpea producers and chickpea breeding programs.

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First report of Diaporthe eres causing root rot of Coptis chinensis Franchet

Plant Disease ◽

10.1094/pdis-12-20-2564-pdn ◽

2021 ◽

Author(s):

PengYing MEI ◽

Xuhong Song ◽

Zhiyu Zhu ◽

Longyun Li

Keyword(s):

Root Rot ◽

Elongation Factor ◽

Its Region ◽

Small Samples ◽

Molecular Characteristics ◽

Conidial Suspension ◽

Coptis Chinensis ◽

Base Pairs ◽

First Report ◽

Surface Sterilization

Chongqing coptis (Coptis chinensis Franchet) industry produces more than 60% of the Chinese coptis crop, and has been exported to many countries and regions. Since 2008, root rot has become a serious and widespread disease on coptis plants in Shizhu county with an average incidence of 40%, and yield losses up to 67%. Symptomatic coptis plants showed stunted growth, with the fibrous roots and main roots having brown or black, rotten, necrotic lesions. To our knowledge, Fusarium solani, F. carminascens, F. oxysporum and F. tricinctum have been previously reported as pathogens of coptis root rot (Luo et al. 2014; Cheng et al. 2020; Wu et al. 2020), but non Fusarium pathogens has not been reported yet. In order to identify new pathogens, 33 diseased roots were collected from Shizhu (30°18'N, 108°30'E) in October 2019. Small samples (0.5 cm in length) were cut from the border between diseased and healthy tissue, and then put on PDA after surface sterilization. Cultures were incubated at 25°C in dark until fungal colonies were observed. After subculturing for 3 times, 3 out of 21 isolates yielded a similar type of fungal colony. White, aerial, fluffy mycelium were formed and reached 8.3 cm diameter within 7 days, and dark pigmentation developed in the centre. Colonies turned to gray with age, and abundant dark brown pycnidia and black stromata were formed at maturity. Alpha conidia were aseptate, hyaline, fusiform to ellipsoidal, often biguttulate, measuring (6.0-8.5)×(2.0-3.0) μm. Beta conidia were aseptate, hyaline, linear to hooked, measuring (18-30)×(1.0-1.5) μm (Figure S1). For further identification, a multigene phylogenetic analysis was carried out. The internal transcribed spacer (ITS), translation elongation factor 1ɑ (tef1-ɑ), histone H3 (his3), calmodulin (cal), and β-tubulin (tub2) gene regions were amplified with ITS1/ITS4, EF1-728F/EF1-986R, CYLH3F/H3-1b, CAL228F/CAL737R, T1/Bt2b (White et al. 1990; Glass and Donaldson 1995; Carbone and Kohn 1999; Crous et al. 2004). GenBank accession numbers of isolate H13 were MT463391 for the ITS region, MT975573 for tef1-ɑ, MT975574 for his3, MT975575 for cal, and MT975576 for tub2. BLAST results showed the ITS, tef1-ɑ, his3, cal and tub2 sequences revealed 99.82% (553/554 base pairs), 100% (347/347 base pairs), 100% (474/474 base pairs), 99.39% (486/489 base pairs), and 99.14% (803/810 base pairs) homology respectively with those of Diaporthe eres (MN816416.1, KU557616.1, KC343564.1, KU557595.1, and KY569366.1). Thus, H13 were identified as D. eres based on its morphological and molecular characteristics. Pathogenicity of D. eres in coptis was investigated using the H13 isolate (1 of the 3 isolates). The roots of 10 healthy 2-year-old coptis plants were individually inoculated with 5 ml of a 106 conidia/mL conidial suspension and sterilized water was used to mock inoculate. Thirty days after inoculation, most of the inoculated coptis roots showed dark brown and rotten root, similar to those observed in the field, whereas mock inoculated roots showed healthy. D. eres was recovered from symptomatic roots and identified based on morphology. To our knowledge, this is the first report of D. eres causing root rot of coptis not only in China but anywhere in the world.

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First report of root rot caused by Fusarium armeniacum on American ginseng in China

Plant Disease ◽

10.1094/pdis-12-19-2554-pdn ◽

2020 ◽

Author(s):

Ximei Zhang ◽

Yanmeng Bi ◽

Junfei LI ◽

Huihui SHAO ◽

XIAO LIN JIAO ◽

...

Keyword(s):

Root Rot ◽

Apical Cell ◽

Elongation Factor ◽

American Ginseng ◽

Ginseng Root ◽

Conidial Suspension ◽

First Report ◽

Tubulin Genes ◽

One Year ◽

Whole Plants

American ginseng (Panax quinquefolius) is an important medicinal plant cultivated in China since the 1980s. Its dried roots are used for food, health care products, and medicine in China (Yuan et al. 2010). Root rot caused by Fusarium spp. was a major disease, with 33 to 41% incidence surveyed in main production areas of Wendeng County (121.80 °E, 37.09 °N) in Shandong Province, China in 2016 to 2019. Symptoms included soft, water-soaked, dark brown to black lesions on the roots. Lesions progressed and the inner parts gradually disintegrated. One-year-old diseased roots were collected in September 2016. Symptomatic tissues were surface-sterilized in 75% ethanol for 30 s and 0.8% NaOCl for 3 min, rinsed in sterile water, plated on potato dextrose agar (PDA), and incubated at 25°C in darkness. Single colonies were then obtained and transferred to carnation leaf agar (CLA) (Burgess et al. 1993) for growth at 25°C with a 12-h photoperiod. Colonies cultured on PDA for 7 days were white to light pink, turning to apricot pigmentation in color. After 30 days on CLA, the colonies produced elongate, falcate macroconidia having 3 to 5 septa, with a long, tapering and curved apical cell, and having the size ranging from 31.1 to 45.6 μm long x 4 to 4.6 μm wide. Microconidia were zero to 1septate, ellipsoid to ovoid and varied in size from 9.5 to 16.8 μm long x 3 to 3.2 μm wide. Chlamydospores formed abundantly, in chains or clusters. This fungus was identified as F. armeniacum (Burgess et al. 1993). Identification was confirmed by sequencing three DNA regions including the internal spacer ribosomal DNA (ITS), elongation factor 1α and β-tubulin genes (Lu et al. 2019). The three DNA regions (MN417271, MG457199, and MN427653) had 100% homology to the sequences of F. armeniacum (KJ737378, HM744664 and HQ141640) (Wang et al. 2015, Yli-Mattila et al. 2011). Pathogenicity tests were conducted on 1- to 2-year-old bare roots and 2-year-old whole plants. For root inoculation, 14 healthy roots were inoculated with two mycelial PDA plugs/root. After 3 to 10 days at 25°C, all the inoculated roots showed water-soaked and root rot symptoms while no lesions were observed in the control roots. For plant inoculation, eight seedlings planted in pots filled with sterilized soil were inoculated by pouring a conidial suspension of 1×105 conidia/ml at 30 ml/pot. Eight seedlings inoculated with sterilized water served as the controls. After 90 days, only 37.5% of the roots survived with typical root rot symptoms whereas the control plants remained symptomless. F. armeniacum was re-isolated from symptomatic roots but not from the control roots. Besides F. armeniacum, F. solani and F. oxysporum that have been reported to be associated with American ginseng root rot in China and Canada (Reeleder et al. 2002; Punja et al. 2008) were also obtained from the diseased root samples in this study. However, the development of root rot caused by F. armeniacum was much more rapid and its symptoms were more severe. Moreover, F. armeniacum could directly infect American ginseng with no wound requirement. F. armeniacum was previously reported on Glycine max (Leguminosae) (Ellis et al. 2012), Platycodon grandiflorus (Campanulaceae) (Wang et al. 2015) and natural grasses (Poaceae) (Nichea et al. 2015). This is the first report of F. armeniacum causing root rot on American ginseng in China. As this species is more virulent to American ginseng, more research is needed to work on this disease.

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First Report of Fusarium solani Causing Crown and Root Rot on Strawberry Crops in Southwestern Spain

Plant Disease ◽

10.1094/pdis-07-13-0682-pdn ◽

2014 ◽

Vol 98 (1) ◽

pp. 161-161 ◽

Cited By ~ 10

Author(s):

A. M. Pastrana ◽

N. Capote ◽

B. De los Santos ◽

F. Romero ◽

M. J. Basallote-Ureba

Keyword(s):

Fusarium Solani ◽

Elongation Factor ◽

Fruit Production ◽

Morphological Characterization ◽

Soil Samples ◽

Fluorescent Light ◽

Distilled Water ◽

Conidial Suspension ◽

First Report ◽

Brown Discoloration

Spain is the fourth largest strawberry (Fragaria × ananassa) producing country in the world. Since April 2010, stunted and dead strawberry plants have been detected in four strawberry fruit production fields in Huelva (southwestern Spain) affecting less than 1% of plants. Symptoms consisted of foliage wilt, plant stunting and drying, and death of older leaves. Internal vascular and cortical tissues of plant crowns showed an orange to brown discoloration. Crowns and roots of symptomatic plants were surface sterilized in 1% sodium hypochlorite for 2 min, rinsed in sterile distilled water for 2 min, and air-dried in a laminar flow cabinet. Small disinfested pieces were transferred to petri dishes containing potato dextrose agar (PDA) and incubated for 10 days at 25°C with a 12-h photoperiod. Cultures derived from single spores were obtained, and morphological characterization was performed by microscopic examination. White to pale cream colonies developed after 10 days of incubation. Unbranched monophialides with microconidia in false heads, micro- (0 to 3 septa) and macroconidia (5 to 7 septa) wide and robust in shape, and chlamydospores were consistent with descriptions of Fusarium solani (Martius) Appel & Wollenweber emend. Snyder & Hansen (2). In addition, the fungus was isolated from asymptomatic runner plants from nurseries by the same method described above, and from soil samples from six fruit-producing fields. Soil samples were analyzed by dilution plating on Fusarium-selective agar medium (1). Genomic DNA from three isolates (FPOST-81 from dead plant ‘Sabrina,’ TOR-11 from runner plant ‘Camarosa,’ and TOR-1 from soil) was obtained with a DNA extraction kit (Isolate Plant DNA MiniKit, Bioline). A portion of the translation elongation factor-1 alpha (EF-1α) gene was sequenced using EF-1/-2 primers (3) (GenBank Accession Nos. KF275032, KF275033, and KF275034). The sequence comparison revealed a 99 to 100% match with F. solani sequences in GenBank and Fusarium-ID databases. To confirm the pathogenicity of the fungi, runner strawberry plants ‘Camarosa’ were inoculated by dipping crowns and roots into a conidial suspension (106 to 107 conidia per ml) for 30 min (8 plants per F. solani isolate) or into sterile distilled water for the controls. Plants were potted in 13-cm diameter pots with peat and maintained at 25/18°C and 70/40% relative humidity (day/night) in a growth chamber with a daily 16-h photoperiod of fluorescent light. Three plants inoculated with isolates TOR-11 and FPOST-81, and four plants inoculated with isolate TOR-1, died within 10 days after inoculation. After 8 to 12 weeks, all of the remaining inoculated plants were stunted and developed symptoms similar to those observed in the field. Production of new feeder roots was lacking or scarce. Control plants remained healthy and formed feeder roots. All plants inoculated with isolates TOR-1 and FPOST-81, and 50% of plants inoculated with TOR-11, showed brown discoloration in the crown. F. solani was re-isolated from symptomatic plants at frequencies of 100% and 80 to 100% from root and crown tissues, respectively. Although F. solani has been reported as a pathogen in other crops, to our knowledge, this is the first report of the occurrence of F. solani causing disease in strawberry plants in Spain. References: (1) D. Bouhot and F. Rouxel. Ann. Phytopathol. 3:251, 1971. (2) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual, Blackwell Publishing, London, 2006. (3) K. O'Donnell et al. Proc. Natl. Acad. Sci. USA 95:2044, 1998.

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First Report of Lily Blight and Wilt Caused by Fusarium tricinctum in China

Plant Disease ◽

10.1094/pdis-11-12-1010-pdn ◽

2013 ◽

Vol 97 (7) ◽

pp. 993-993 ◽

Cited By ~ 10

Author(s):

Y. Y. Li ◽

Y. J. Wang ◽

Z. K. Xie ◽

R. Y. Wang ◽

Y. Qiu ◽

...

Keyword(s):

Root Rot ◽

Elongation Factor ◽

Aerial Mycelium ◽

Its Region ◽

Universal Primers ◽

Conidial Suspension ◽

Root Tips ◽

First Report ◽

Horticultural Crops ◽

Blastn Analysis

Lily (Lilium spp.) is one of the most well-known horticultural crops, and plays an important economic role in China. In September 2011, wilted plants were observed on Lilium oriental hybrid cultivar ‘Sorbonne’ growing in Longde County, Ningxia Hui Autonomous Region, China. Disease symptoms included wilting, stem and root rot, brown spots of bulbs and then bulbs rotting and spalling from the basal disc, plus a progressive yellowing and defoliation of the leaves from the base. Diseased plants were sampled from fields. Small pieces of symptomatic bulbs, stems, and roots from 10 different plants were surface disinfected with 75% ethanol for 30 s, 3% sodium hypochlorite for 5 min, and then washed three times in sterilize distilled water. The tissues were placed onto Martin Agar (2) at 25°C for 7 days. Nine isolates with morphology similar to Fusarium were obtained from the diseased tissues. Isolates were transferred to potato dextrose agar (PDA) and carnation leaf agar (CLA) and incubated at 25°C. Seven were identified as Fusarium oxysporum and one was F. solani, which have been reported as pathogens of lily in China (1). The other isolate, when grown on PDA, rapidly produced dense, white aerial mycelium that became pink with age and formed red pigments in the medium. On CLA, macroconidia with three to five septate were abundant, relatively slender, and curved to lunate. Microconidia were abundant, oval or pyriform, and one to two celled. Chlamydospores were in chains with smooth exine. The rDNA internal transcribed spacer (ITS) region and a portion of the translation elongation factor 1-alpha (EF-1α) gene of the fungus were amplified, with universal primers ITS1/ITS4 and EF1/EF2 primers respectively (3) and sequenced. In addition, the β-tubulin gene (β-tub) of the fungus was amplified with modified primers Btu-F-F01 (5′-CAGACMGGTCAGTGCGTAA-3′) and Btu-F-R01 (5′-TCTTGGGGTCGAACATCTG-3′) (4). BLASTn analysis showed that the ITS sequences of the isolate (GenBank Accession No. JX989827) had 98.9% similarity with those of F. tricinctum (EF611092, JF776665, and HM776425) and the EF-1α sequences of the isolate (JX989828) had 98.1% similarity with those of F. tricinctum (EU744837 and JX397850). The β-tub sequences of the isolate (JX989829) had 99.0% similarity with those of F. tricinctum (EU490236 and AB587077). The isolate was tested for pathogenicity. Two-month-old ‘Sorbonne’ seedlings were inoculated by placing 5 ml of conidial suspension (about 106 conidia per ml) over the roots of plants in each pot. Control plants were treated with sterile water in the same way. Plants were placed in a greenhouse at 22 to 25°C with a 15-h photoperiod. There were eight plants per pot and three replicates for each treatment. After 3 weeks, 87.5% of the inoculated plants exhibited browning of the root tips, root rot, and yellowing of the leaves, while control plants were symptomless. The pathogen was reisolated from the infected roots and identified as F. tricinctum, thus fulfilling Koch's postulates. To our knowledge, this is the first report of Fusarium wilt of lily caused by F. tricinctum. This information will provide guidance for the control of lily wilt disease and add information useful for the production of lilies. References: (1) C. Li and J. J. Li. Acta Phytopathol. Sin. (in Chinese) 26:192, 1995. (2) J. P. Martin. Soil Sci. 38:215, 1950. (3) K. O'Donnell et al. Proc. Nat. Acad. Sci. U. S. A. 95:2044, 1998. (4) M. Watanabe et al. BMC Evol. Biol. 11:322. 2011.

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First report of Fuasium wilt caused by Fusarium kyushuense in China

Plant Disease ◽

10.1094/pdis-06-21-1280-pdn ◽

2021 ◽

Author(s):

Shuning Chen ◽

Wei Sun ◽

Huizhu Yuan ◽

Xiaojing Yan

Keyword(s):

Management Practices ◽

Vascular System ◽

Elongation Factor ◽

Sterile Water ◽

Neighbor Joining ◽

Gerbera Jamesonii ◽

Translation Elongation Factor ◽

Dark Chamber ◽

Translation Elongation ◽

First Report

Gerbera daisy, Gerbera jamesonii H. Bolus ex. Hooker, is an important flower grown globally. In September 2020, gerbera seedlings in a greenhouse farm in the region of Fujian, China, developed symptoms of severe wilting and stunting. The main stem exhibited reddish to light brown vascular discoloration. Approximately 30% of the 60,000 plants showed symptoms. To isolate the causal agent, necrotic tissue pieces (3×3 mm) from the symptomatic stem were surface-disinfected with 1% NaClO for 1 min and washed three times with sterile water. The disinfected pieces were dried and placed on potato dextrose agar (PDA) at 25°C in the darkness for 4 days inside a dark chamber. Reddish-white and floccose mycelia developed on PDA after 3 days incubation. Ten single-spored isolates were identified as Fusarium kyushuense based on morphological features (Aoki & O'Donnell, 1998). Hyaline and straight or slightly curved macroconodia were observed with 3 to 5 septate, 24.5 - 46.6 × 3.6 - 5.7 μm (n = 100). Microconidia were ellipsoidal to clavate, 0 to 1 septate, and 6.3 to 19.5 × 3.2 to 5.3 μm (n = 100). No chlamydospores were observed. In order to validate this result, partial RNA polymerase second largest subunit (RPB2) combined with translation elongation factor (EF-1α) gene regions were amplified and sequenced from three isolates with primers 5f2/11ar (Liu et al., 1999) and primers EF1/EF2 (Geiser et al. 2004), respectively. Fusarium MLST analysis showed that the RPB2 (Genbank accession No. MZ130468, No. MZ130469, No. MZ130470) matched 99.72% (MH582170) to F. kyushuense, and the EF-1α (MZ130471, MZ130472, MZ130473) matched 99.84% (MH582297) to F. kyushuense in the Fusarium MLST. Besides, a phylogenetic analysis was conducted using the neighbor-joining algorithm based on the RPB2 and EF-1α gene sequences. The isolates clustered with F. kyushuense. To assess pathogenicity, the three molecularly identified isolates were used. The isolates were grown on carboxylmethyl cellulose (CMC) medium (carboxymethyl-cellulose (Sigma C-4888) 15.0 gram, NH4NO3 1.0 gram, KH2PO4 monobasic 1.0 gram, MgSO4·7H2O 0.5 gram, yeast extract 1.0 gram, distilled water filled to 1.0 liter) for sporulation. The roots of 12 healthy 30-day-old gerbera plants were inoculated by treating them with 10 mL of conidia suspension (1×106 conidial/mL). A group of 12 seedlings of the same age was treated with sterile water to serve as the control. Plants were grown in a glasshouse at 23 °C, relative humidity >70%, and 16 h light per day. Typical symptoms of wilt and discoloration of the vascular system in roots and stems developed within 10 days. Uninoculated plants remained healthy. Isolates were consistently re-isolated from the symptomatic stem and the recovered isolates were identified as F. kyushuense by amplifing the EF-1α gene. The assays were conducted twice. F. kyushuense has been reported to cause wilt and rot of tobacco (Wang et al., 2013), maize ears (Wang et al., 2014) and rice (Zhao et al., 2007) in China. To the best of our knowledge, this is the first report of F. kyushuense causing stem and root wilt on G. jamesonii. The disease must be considered in existing management practices.

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First Report of Pythium Root Rot of Rau Ram (Polygonum odoratum)

Plant Disease ◽

10.1094/pd-89-0340a ◽

2005 ◽

Vol 89 (3) ◽

pp. 340-340

Author(s):

E. N. Rosskopf ◽

C. B. Yandoc ◽

B. Stange ◽

E. M. Lamb ◽

D. J. Mitchell

Keyword(s):

Root Rot ◽

Vascular System ◽

Pond Water ◽

Centraalbureau Voor ◽

First Report ◽

Pathogenicity Tests ◽

Symptomatic Tissue ◽

Pythium Helicoides ◽

Production House ◽

Test Plants

Polygonum odoratum (= Persicaria odorata), known as rau ram or sang hum, is native to southeastern Asia and is a common herb in Vietnamese cuisine (1). It has been studied most extensively for its aromatic compound content (2). In Florida, rau ram commonly is grown hydroponically in greenhouses using large, cement beds with recirculated water. The plants form dense mats from which new growth is trimmed for market. During January of 2002, a severe dieback was observed in one production house in Saint Lucie County, FL. Plants with less severe symptoms were yellowed and stunted. Roots of symptomatic plants were largely decayed with root symptoms beginning as a tip necrosis. The cortex of severely affected roots slipped off easily, leaving a stringy vascular system. Plating of symptomatic tissue from 20 randomly selected plant samples was performed with multiple general and selective media including potato dextrose agar, corn meal agar with pimaricin, ampicillin, rifampicin, and pentachloronitrobenzene (PARP) (3). All colonies produced were identified as Pythium helicoides Drechsler on the basis of sporangial, oogonial, and antheridial characteristics (4). Isolates had proliferous, obovoid, papillate sporangia, and were homothallic with smooth-walled oogonia and thick-walled, aplerotic oospores. Multiple antheridial attachments per oogonium were common with the antheridium attached along its entire length. Pathogenicity tests were conducted using P. odoratum plants grown from commercial transplants. Two tests were performed. Each test was conducted using eight inoculated and eight control plants. In the first test, plants were maintained in 10-cm pots immersed in sterilized pond water for the duration of the test. Plants were inoculated with five 7- × 70-mm sections of freshly growing mycelial culture per plant using 10-day-old cultures of Pythium helicoides grown on water agar. Chlorosis was observed at approximately 2 months after inoculation. Root necrosis was observed in inoculated plants approximately 5 months after inoculation. This test was performed in the greenhouse with temperatures ranging from 20 to 30°C. The second test was performed in growth chambers at 35 to 40°C. Plants were maintained in 10-cm pots immersed in Hoagland's solution and were inoculated with four 6-mm plugs per plant. Symptoms were observed on inoculated plants at this temperature within 1 week of inoculation. No chlorosis or root decay was observed in noninoculated, immersed plants. The pathogen was reisolated from inoculated, symptomatic tissue. To our knowledge, this is the first report of root rot of P. odoratum caused by Pythium helicoides. References: (1) R. E. Bond. Herbarist 55:34, 1989. (2) N. X. Dung et al. J. Essent. Oil Res. 7:339, 1995. (3) M. E. Kannwischer and D. J. Mitchell. Phytopathology 68:1760, 1978. (4) A. J. van der Plaats-Niterink. Monograph of the Genus Pythium. Vol. 21, Studies in Mycology. Centraalbureau voor Schimmelcutltures, Baarn, The Netherlands, 1981.

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First Report of Fusarium solani Causing Root Rot on Coptis chinensis in Southwestern China

Plant Disease ◽

10.1094/pdis-02-14-0164-pdn ◽

2014 ◽

Vol 98 (9) ◽

pp. 1273-1273 ◽

Cited By ~ 2

Author(s):

X.-M. Luo ◽

J.-L. Li ◽

J.-Y. Dong ◽

A.-P. Sui ◽

M.-L. Sheng ◽

...

Keyword(s):

Fusarium Solani ◽

Root Rot ◽

Southwestern China ◽

Molecular Characteristics ◽

Distilled Water ◽

Conidial Suspension ◽

Storage Roots ◽

Coptis Chinensis ◽

Causal Organism ◽

First Report

China is the world's largest producer country of coptis (Coptis chinensis), the rhizomes of which are used in traditional Chinese medicine. Since 2008, however, root rot symptoms, including severe necrosis and wilting, have been observed on coptis plants in Chongqing, southwestern China. Of the plants examined from March 2011 to May 2013 in 27 fields, 15 to 30% were covered with black necrotic lesions. The leaves of infected plants showed wilt, necrotic lesions, drying, and death. The fibrous roots, storage roots, and rhizomes exhibited brown discoloration and progressive necrosis that caused mortality of the infected plants. Infected plants were analyzed to identify the causal organism. Discoloration of the internal vascular and cortical tissues of the rhizomes and taproots was also evident. Symptomatic taproots of the diseased coptis were surface sterilized in 1% sodium hypochlorite for 2 min, rinsed in sterile distilled water for 2 min, and then air-dried in sterilized atmosphere/laminar flow. Small pieces of disinfested tissue (0.3 cm in length) were transferred to petri dishes containing potato dextrose agar (PDA) supplemented with 125 μg ml–1 streptomycin sulfate and 100 μg ml–1 ampicillin, and incubated for 5 days at 25°C with a 12-h photoperiod. Four distinct species of fungal isolates (HL1 to 4) derived from single spores were isolated from 30 plants with root rot symptoms collected from the study sites. To verify the pathogenicity of individual isolates, healthy coptis plants were inoculated by dipping roots into a conidial suspension (106 conidia/ml) for 30 min (15 plants per isolate), as described previously (1). Inoculated plants were potted in a mixture of sterilized quartz sand-vermiculite-perlite (4:2:1, v/v) and incubated at 25/18°C and 85 to 90% relative humidity (day/night) in a growth chamber with a daily 16-h photoperiod of fluorescent light. Plants dipped in sterile distilled water were used as controls. After 15 days, symptoms similar to those observed in the field were observed on all plants (n = 15) that were inoculated with HL1, but symptoms were not observed on plants inoculated with HL2, HL3, and HL4, nor on control plants. HL1 was re-isolated from symptomatic plants but not from any other plants. Morphological characterization of HL1 was performed by microscopic examination. The septate hyphae, blunt microconidia (2 to 3 septa) in the foot cell and slightly curved microconidia in the apical cell, and chlamydospores were consistent with descriptions of Fusarium solani (2). The pathogen was confirmed to be F. solani by amplification and sequencing of the ribosomal DNA internal transcribed spacer (rDNA-ITS) using the universal primer pair ITS4 and ITS5. Sequencing of the PCR product revealed a 99 to 100% similarity with the ITS sequences of F. solani in GenBank (JQ724444.1 and EU273504.1). Phylogenetic analysis (MEGA 5.1) using the neighbor-joining algorithm placed the HL1 isolate in a well-supported cluster (97% bootstrap value based on 1,000 replicates) with JQ724444.1 and EU273504.1. The pathogen was thus identified as F. solani based on its morphological and molecular characteristics. To our knowledge, this is the first report of root rot of coptis caused by F. solani in the world. References: (1) K. Dobinson et al. Can. J. Plant Pathol. 18:55, 1996. (2) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Oxford, 2006.

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First Report of Medical Tree Peony Root Rot Caused by Fusarium solani in Tongling, China

Plant Disease ◽

10.1094/pdis-02-12-0142-pdn ◽

2012 ◽

Vol 96 (6) ◽

pp. 909-909 ◽

Cited By ~ 10

Author(s):

M. Guo ◽

Y. M. Pan ◽

Z. M. Gao

Keyword(s):

Fusarium Solani ◽

Root Rot ◽

Tap Water ◽

Blood Stasis ◽

The United States ◽

Effective Control ◽

Conidial Suspension ◽

Tree Peony ◽

First Report ◽

Peony Root

Tree peony bark, a main component of Chinese traditional medicine used for alleviating fever and dissipating blood stasis, is mainly produced in Tongling, China. Recently, tree peony cultivation in this area was seriously affected by root rot, with approximately 20 to 30% disease incidence each year. The disease severely affects yield and quality of tree peony bark. During the past 2 years, we collected 56 diseased tree peony plants from Mudan and Fenghuang townships in Tongling. We found reddish brown to dark brown root rot in mature roots, especially on those with injuries. Plant samples collected were disinfected with 2% sodium hypochlorite and isolations were conducted on potato sucrose agar (PSA). Eleven isolates were obtained and all had white fluffy aerial hypha on PSA. Two types of conidia were produced; the larger, reaphook-shaped ones had three to five septa and the smaller, ellipse-shaped ones had one or no septum. The reaphook-shaped conidia were 20.15 to 37.21 × 3.98 to 5.27 μm and the ellipse-shaped conidia were 6.02 to 15.52 × 2.21 to 5.33 μm in size. Chlamydospores were produced, with two to five arranged together. Biological characteristics of the fungi indicated that the optimum temperature for the mycelial growth on PSA was 25 to 30°C and the optimum pH range was 5.5 to 7.0. The above morphological characteristics point the fungal isolates to be Fusarium solani. To confirm pathogenicity, 30 healthy 1-year-old tree peony seedling plants were grown in pots (25 cm in diameter) with sterilized soil and a conidial suspension from one isolate (FH-1, 5 × 105 conidia/ml) was used for soil inoculation. Inoculated seedlings were maintained at 28°C in a greenhouse with a 12-h photoperiod of fluorescent light. Seedlings inoculated with distilled water were used as controls. After 3 weeks, the roots were collected and rinsed with tap water. Dark brown lesions were observed in the inoculated mature roots but not in the control roots. To confirm the identity of the pathogen, F. solani strains were reisolated from the lesions and total genomic DNA was extracted with the cetyltriethylammnonium bromide method from the mycelia of the reisolated strains (1). PCR was performed using the fungal universal primers ITS4 (5′-TCCTCCGCTTATTGATATGC-3′) and ITS5 (5′-GGAAGTAAAAGTCGTAACAAGG-3′) to amplify a DNA fragment of approximately 590 bp. The purified PCR products were sequenced (Invitrogen Co., Shanghai, China) and shared 100% sequence identity with each other. A comparison of the sequence (JQ658429.1) by the Clustal_W program (2) with those uploaded in GenBank confirmed with the fungus F. solani (100% sequence similarity to isolate S-0900 from the Great Plains of the United States; EU029589.1). To our knowledge, this is the first report of F. solani causing medical tree peony root rot in China. The existence of this pathogen in China may need to be considered for developing effective control strategies. References: (1). C. N. Stewart et al. Biotechniques 14:748, 1993. (2). J. D. Thompson et al. Nucleic Acids Res. 22:4673, 1994.

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First Report of Stem Blight Caused by Calonectria colhounii (Anamorph Cylindrocladium colhounii) on Greenhouse-Grown Blueberries in the United States

Plant Disease ◽

10.1094/pdis-02-11-0117 ◽

2011 ◽

Vol 95 (9) ◽

pp. 1187-1187

Author(s):

J. J. Sadowsky ◽

T. D. Miles ◽

A. M. C. Schilder

Keyword(s):

United States ◽

North Carolina ◽

Root Rot ◽

The United States ◽

Vaccinium Corymbosum ◽

Conidial Suspension ◽

Centraalbureau Voor ◽

First Report ◽

Stem Lesions ◽

Β Tubulin

Necrotic stems and leaves were observed on 2- to 4-month-old, rooted microshoot plants (Vaccinium corymbosum L. ‘Liberty’ and ‘Bluecrop’, V. angustifolium Aiton ‘Putte’, and V. corymbosum × V. angustifolium ‘Polaris’) in a Michigan greenhouse in 2008 and 2009. As the disease progressed, leaves fell off and 80 to 100% of the plants died in some cases. Root rot symptoms were also observed. A fungus was isolated from stem lesions. On potato dextrose agar (PDA), cultures first appeared light tan to orange, then rusty brown and zonate with irregular margins. Chains of orange-brown chlamydospores were abundant in the medium. Macroconidiophores were penicillately branched and had a stipe extension of 220 to 275 × 2.5 μm with a narrowly clavate vesicle, 3 to 4 μm wide at the tip. Conidia were hyaline and cylindrical with rounded ends, (1-)3-septate, 48 to 73 × 5 to 7 (average 60 × 5.5) μm and were held together in parallel clusters. Perithecia were globose to subglobose, yellow, 290 to 320 μm high, and 255 to 295 μm in diameter. Ascospores were hyaline, 2- to 3-septate, guttulate, fusoid with rounded ends, slightly curved, and 30 to 88 × 5 to 7.5 (average 57 × 5.3) μm. On the basis of morphology, the fungus was identified as Calonectria colhounii Peerally (anamorph Cylindrocladium colhounii Peerally) (1,2). The internal transcribed spacer region (ITS1 and ITS2) of the ribosomal DNA and the β-tubulin gene were sequenced (GenBank Accession Nos. HQ909028 and JF826867, respectively) and compared with existing sequences using BLASTn. The ITS sequence shared 99% maximum identity with that of Ca. colhounii CBS 293.79 (GQ280565) from Java, Indonesia, and the β-tubulin sequence shared 97% maximum identity with that of Ca. colhounii CBS 114036 (DQ190560) isolated from leaf spots on Rhododendron sp. in North Carolina. The isolate was submitted to the Centraalbureau voor Schimmelcultures in the Netherlands (CBS 129628). To confirm pathogenicity, 5 ml of a conidial suspension (1 × 105/ml) were applied as a foliar spray or soil drench to four healthy ‘Bluecrop’ plants each in 10-cm plastic pots. Two water-sprayed and two water-drenched plants served as controls. Plants were misted intermittently for 2 days after inoculation. After 7 days at 25 ± 3°C, drench-inoculated plants developed necrotic, sporulating stem lesions at the soil line, while spray-inoculated plants showed reddish brown leaf and stem lesions. At 28 days, three drench-inoculated and one spray-inoculated plant had died, while others showed stem necrosis and wilting. No symptoms were observed on control plants. Fungal colonies reisolated from surface-disinfested symptomatic stem, leaf, and root segments appeared identical to the original isolate. Cy. colhounii was reported to cause a leaf spot on blueberry plants in nurseries in China (3), while Ca. crotalariae (Loos) D.K. Bell & Sobers (= Ca. ilicicola Boedijn & Reitsma) causes stem and root rot of blueberries in North Carolina (4). To our knowledge, this is the first report of Ca. colhounii causing a disease of blueberry in Michigan or the United States. Because of its destructive potential, this pathogen may pose a significant threat in blueberry nurseries. References: (1) P. W. Crous. Taxonomy and Pathology of Cylindrocladium (Calonectria) and Allied Genera. The American Phytopathological Society, St. Paul, MN, 2002. (2) L. Lombard et al. Stud. Mycol. 66:31, 2010. (3) Y. S. Luan et al. Plant Dis. 90:1553, 2006. (4) R. D. Milholland. Phytopathology 64:831, 1974.

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