scholarly journals First Report of Vascular Wilt Caused by Fusarium redolens on Lentil in Italy

Plant Disease ◽  
2008 ◽  
Vol 92 (7) ◽  
pp. 1132-1132 ◽  
Author(s):  
L. Riccioni ◽  
A. Haegi ◽  
M. Valvassori

Lentil (Lens culinaris Medik.) is a traditional crop in Sicily, Italy. Near Villalba (Caltanissetta), a local lentil landrace, “Lenticchia di Villalba”, is commonly grown. From 2002 to 2004, wilt was observed in five lentil fields (≈1 ha each) at rates from 5 to 20%. Affected plants were yellow and stunted with discoloration in the vascular tissue of stems and crowns. Pieces of brown vascular tissue from stems were disinfested in 2% sodium hypochlorite for 2 min, rinsed with sterile distilled water, placed on potato dextrose agar, and incubated at 23°C. Isolates with morphological characteristics of Fusarium oxysporum Schlecht.:Fr. (2) were consistently recovered from affected plants. For molecular identification of five isolates, the rDNA internal transcribed spacer (ITS) region and a portion of the elongation factor EF-1α were sequenced using ITS5/4 and EF1/2 primers, respectively (1). Two sequences of the ITS region were obtained: a 468-bp sequence from isolates ER1259, ER1260, and ER1275 (submitted as GenBank Accession No. EU159118) and a 483-bp sequence from isolates ER1274 and ER1276 (submitted as GenBank Accession No. EU281661). The two sequences shared 93% similarity. A sequence homology search using the NCBI BLAST program revealed that the first sequence had 100% homology with the ITS sequences of more than 50 F. oxysporum isolates of various formae speciales in GenBank and the second shared 100% homology with the ITS sequences of five isolates of F. redolens Wollenw. (e.g., GenBank Accession No. X94169 of the strain CBS 360.87). Amplification of the EF-1α produced a sequence from isolates ER1274 and ER1276 (submitted as GenBank Accession No. EU281660) with 99 to 100% homology to sequences of F. redolens and a sequence from strains ER1259, ER1275, and ER1260 (submitted as GenBank Accession No. EU281659) with 100% homology to that of more than 50 F. oxysporum strains in GenBank. Although F. redolens and F. oxysporum are morphologically similar, recent molecular studies have shown that they are distinct and phylogenetically distant species (3). On the basis of genetic sequences, isolates ER1274 and ER1276 were identified as F. redolens. These isolates were evaluated for pathogenicity on lentil. For each isolate, 10 2-week-old seedlings of “Lenticchia di Villalba” were inoculated by submerging roots in a suspension of 2.5 × 106 conidia/ml for 10 min. Plants were put into separate tubes containing 70 ml of a nutritional liquid medium (7 ml of HydroPlus Olikani per liter; Yara, Nanterre, France) and incubated in a growth chamber at 20°C with 12 h of light per day. Seedlings dipped in sterile water served as the control treatment. The pathogenicity test was repeated twice. Inoculated seedlings started to wilt 1 week after inoculation and developed root rot and vascular discoloration. After 2 weeks, 70% of the inoculated plants were affected by both isolates and 40 and 10% died when inoculated with ER1274 and ER1276 isolates, respectively. F. redolens was consistently reisolated from the stems of wilted plants. Noninoculated plants remained healthy. Currently, only F. oxysporum f. sp. lentis Vasud. and Sriniv. has been reported as the cause of Fusarium wilt of lentil. To our knowledge, this is the first report of F. redolens as a pathogen on lentil. References: (1) R. P. Baayen et al. Phytopathology 91:1037, 2001. (2) P. E. Nelson et al. Fusarium Species: An Illustrated Manual for Identification. The Pennsylvania State University Press, University Park, 1983. (3) K. O'Donnell et al. Mycologia 90:465, 1998.

Plant Disease ◽  
2014 ◽  
Vol 98 (8) ◽  
pp. 1156-1156 ◽  
Author(s):  
H. Zhang ◽  
W. Luo ◽  
Y. Pan ◽  
J. Xu ◽  
J. S. Xu ◽  
...  

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.


Plant Disease ◽  
2014 ◽  
Vol 98 (9) ◽  
pp. 1278-1278 ◽  
Author(s):  
S. E. Cho ◽  
J. H. Park ◽  
S. H. Hong ◽  
I. Y. Choi ◽  
H. D. Shin

Agastache rugosa (Fisch. & C.A. Mey.) Kuntze, known as Korean mint, is an aromatic plant in the Lamiaceae. It is widely distributed in East Asian countries and is used as a Chinese traditional medicine. In Korea, fresh leaves are commonly added to fish soups and stews (3). In November 2008, several dozen Korean mints plants growing outdoors in Gimhae City, Korea, were found to be severely infected with a powdery mildew. The same symptoms had been observed in Korean mint plots in Busan and Miryang cities from 2008 to 2013. Symptoms first appeared as thin white colonies, which subsequently developed into abundant hyphal growth on stems and both sides of the leaves. Severe disease pressure caused withering and senescence of the leaves. Voucher specimens (n = 5) were deposited in the Korea University Herbarium (KUS). Appressoria on the mycelium were nipple-shaped or nearly absent. Conidiophores were 105 to 188 × 10 to 13 μm and produced 2 to 4 immature conidia in chains with a sinuate outline, followed by 2 to 3 cells. Foot-cells of the conidiophores were straight, cylindrical, slightly constricted at the base, and 37 to 58 μm long. Conidia were hyaline, ellipsoid to barrel-shaped, measured 25 to 40 × 15 to 23 μm (length/width ratio = 1.4 to 2.1), lacked distinct fibrosin bodies, and showed reticulate wrinkling of the outer walls. Primary conidia were obconically rounded at the apex and subtruncate at the base. Germ tubes were produced at the perihilar position of conidia. No chasmothecia were observed. The structures described above were typical of the Oidium subgenus Reticuloidium anamorph of the genus Golovinomyces. The measurements and morphological characteristics were compatible with those of G. biocellatus (Ehrenb.) V.P. Heluta (1). To confirm the identification, molecular analysis of the sequence of the internal transcribed spacer (ITS) region of ribosomal DNA (rDNA) of isolate KUS-F27200 was conducted. The complete ITS rDNA sequence was amplified using primers ITS5 and P3 (4). The resulting 514-bp sequence was deposited in GenBank (Accession No. KJ585415). A GenBank BLAST search of the Korean isolate sequence showed >99% similarity with the ITS sequence of many G. biocellatus isolates on plants in the Lamiaceae (e.g., Accession Nos. AB307669, AB769437, and JQ340358). Pathogenicity was confirmed by gently pressing diseased leaf onto leaves of five healthy, potted Korean mint plants. Five non-inoculated plants served as a control treatment. Inoculated plants developed symptoms after 7 days, whereas the control plants remained symptomless. The fungus present on inoculated plants was identical morphologically to that observed on the original diseased plants. The pathogenicity test was repeated with identical results. A powdery mildew on A. rugosa caused by G. biocellatus was reported from Romania (2). To our knowledge, this is the first report of powdery mildew caused by G. biocellatus on A. rugosa in Korea. The plant is mostly grown using organic farming methods with limited chemical control options. Therefore, alternative control measures should be considered. References: (1) U. Braun and R. T. A. Cook. Taxonomic Manual of the Erysiphales (Powdery Mildews), CBS Biodiversity Series No. 11. CBS, Utrecht, 2012. (2) D. F. Farr and A. Y. Rossman. Fungal Databases. Syst. Mycol. Microbiol. Lab., online publication, USDA ARS, retrieved 17 February 2014. (3) T. H. Kim et al. J. Sci. Food Agric. 81:569, 2001. (4) S. Takamatsu et al. Mycol. Res. 113:117, 2009.


Plant Disease ◽  
2020 ◽  
Author(s):  
Fangmin Hao ◽  
Quanyu Zang ◽  
Weihong Ding ◽  
Erlei Ma ◽  
Yunping Huang ◽  
...  

Melon (Cucumis melo L.) is a member of the Cucurbitaceae family, an important economical and horticultural crop, which is widely grown in China. In May 2020, fruit rot disease with water-soaked lesions and pink molds on cantaloupe melons was observed in several greenhouses with 50% disease incidence in Ningbo, Zhejiang Province in China. In order to know the causal agent, diseased fruits were cut into pieces, surface sterilized for 1 min with 1% sodium hypochlorite (NaClO), 2 min with 75% ethyl alcohol, rinsed in sterile distilled water three times (Zhou et al. 2018), and then placed on potato dextrose agar (PDA) medium amended with streptomycin sulfate (100 μg/ml) plates at 25°C for 4 days. The growing hyphae were transferred to new PDA plates using the hyphal tip method, putative Fusarium colonies were purified by single-sporing. Twenty-five fungal isolates were obtained and formed red colonies with white aerial mycelia at 25°C for 7 days, which were identified as Fusarium isolates based on the morphological characteristics and microscopic examination. The average radial mycelial growth rate of Fusarium isolate Fa-25 was 11.44 mm/day at 25°C in the dark on PDA. Macroconidia were stout with curved apical and basal cells, usually with 4 to 6 septa, and 29.5 to 44.2 × 3.7 to 5.2 μm on Spezieller Nährstoffarmer agar (SNA) medium at 25°C for 10 days (Leslie and Summerell 2006). To identify the species, the internal transcribed spacer (ITS) region and translational elongation factor 1-alpha (TEF1-α) gene of the isolates were amplified and cloned. ITS and TEF1-α was amplified using primers ITS1/ITS4 and EF1/EF2 (O’Donnell et al. 1998), respectively. Sequences of ITS (545 bp, GenBank Accession No. MT811812) and TEF1-α (707 bp, GenBank Acc. No. MT856659) for isolate Fa-25 were 100% and 99.72% identical to those of F. asiaticum strains MSBL-4 (ITS, GenBank Acc. MT322117.1) and Daya350-3 (TEF1-α, GenBank Acc. KT380124.1) in GenBank, respectively. A phylogenetic tree was established based on the TEF1-α sequences of Fa-25 and other Fusarium spp., and Fa-25 was clustered with F. asiaticum. Thus, both morphological and molecular characterizations supported the isolate as F. asiaticum. To confirm the pathogenicity, mycelium agar plugs (6 mm in diameter) removed from the colony margin of a 2-day-old culture of strain Fa-25 were used to inoculate melon fruits. Before inoculation, healthy melon fruits were selected, soaked in 2% NaClO solution for 2 min, and washed in sterile water. After wounding the melon fruits with a sterile needle, the fruits were inoculated by placing mycelium agar plugs on the wounds, and mock inoculation with mycelium-free PDA plugs was used as control. Five fruits were used in each treatment. The inoculated and mock-inoculated fruits were incubated at 25°C with high relative humidity. Symptoms were observed on all inoculated melon fruits 10 days post inoculation, which were similar to those naturally infected fruits, whereas the mock-inoculated fruits remained symptomless. The fungus re-isolated from the diseased fruits resembled colony morphology of the original isolate. The experiment was conducted three times and produced the same results. To our knowledge, this is the first report of fruit rot of melon caused by F. asiaticum in China.


Phytotaxa ◽  
2016 ◽  
Vol 260 (2) ◽  
pp. 101 ◽  
Author(s):  
CHANG SUN KIM ◽  
JONG WON JO ◽  
YOUNG-NAM KWAG ◽  
GI-HO SUNG ◽  
JAE-GU HAN ◽  
...  

Thirty-four Lycoperdon specimens from Korea were examined with the internal transcribed spacer (ITS) region of ribosomal DNA sequence data. The result of the ITS sequences phylogenetic analysis indicated that the Korean specimens represented nine different species. To confirm the taxonomic position of these species, we conducted an intensive morphological investigation, and additional phylogenetic investigation of the protein coding regions RNA polymerase subunit II (RPB2) and translation elongation factor 1-alpha (TEF1). We discovered two new species (L. albiperidium and L. subperlatum) and one (L. ericaeum) newly discovered in Korea. Lycoperdon albiperidium is closely related to L. ericaeum based on ITS, RPB2 and TEF1 sequence data, but these species were distinguishable by morphological characteristics, especially the shape of the basidiocarps, the diameter of the eucapillitial threads and the size of the basidospores. Lycoperdon subperlatum is quite similar to the European and American L. perlatum based on morphological characteristics. However, L. subperlatum is clearly distinct from European and American L. perlatum based on ITS, RPB2 and TEF1 sequence data, and somewhat differs from them in macro- and microscopic characteristics. Based on morphological characteristics, L. ericaeum is related to L. subumbrinum and L. lividum but it is distinguishable by the presence of fragile, eucapillitial threads, the diameters of the threads and ITS sequences. Here, we describe four Lycoperdon species collected in Korea.


Plant Disease ◽  
2013 ◽  
Vol 97 (10) ◽  
pp. 1377-1377 ◽  
Author(s):  
H.-W. Choi ◽  
S. K. Hong ◽  
Y. K. Lee ◽  
H. S. Shim

In July 2010, fusarium wilt symptoms of tomato (Lycopersicon esculentum Mill.) plants were found in two commercial greenhouses in the Damyang area of Korea. Approximately 1% of 7,000 to 8,000 tomato plants were wilted and chlorotic in each greenhouse. The vascular tissue was usually dark brown and the discoloration extended to the apex. Fragments (each 5 × 5 mm) of the symptomatic tissue were surface-sterilized with 1% NaOCl for 1 min, then rinsed twice in sterilized distilled water (SDW). The tissue pieces were placed on water agar and incubated at 25°C for 4 to 6 days. Nine Fusarium isolates were obtained from four diseased plants, of which three isolates were identified as F. oxysporum based on morphological characteristics on carnation leaf agar medium and DNA sequences of the translation elongation factor 1-alpha (EF-1α) gene (2). Macroconidia were mostly 3- to 5-septate, slightly curved, and 28 to 53 × 2.8 to 5.2 μm. Microconidia were abundant, borne in false heads or short monophialides, generally single-celled, oval to kidney shaped, and 5 to 23 × 3 to 5 μm. Chlamydospores were single or in short chains. The EF-1α gene was amplified from three isolates by PCR assay using ef1 and ef2 primers (3), and the amplification products were sequenced. The nucleotide sequences obtained were deposited in GenBank (Accession Nos. KC491844, KC491845, and KC491846). BLASTn analysis showed 99% homology with the EF-1α sequence of F. oxysporum f. sp. lycopersici MN-24 (HM057331). Pathogenicity tests and race determination were conducted using root-dip inoculation (4) on seedlings of tomato differential cultivars: Ponderosa (susceptible to all races), Momotaro (resistant to race 1), Walter (resistant to races 1 and 2), and I3R-1 (resistant to all races). A spore suspension was prepared by flooding 5-day-old cultures on potato dextrose agar with SDW. Plants at the first true-leaf stage were inoculated by dipping the roots in the spore suspension (1 × 106 conidia/ml) for 10 min. Inoculated plants were transplanted into pots containing sterilized soil, and maintained in the greenhouse at 25/20°C (12/12 h). Twenty-four seedlings of each cultivar were arranged into three replications. An equal number of plants of each cultivar dipped in water were used as control treatments. Disease reaction was evaluated 3 weeks after inoculation, using a disease index on a scale of 0 to 4 (0 = no symptoms, 1 = slightly swollen and/or bent hypocotyl, 2 = one or two brown vascular bundles in the hypocotyl, 3 = at least two brown vascular bundles and growth distortion, 4 = all vascular bundles brown and the plant either dead or very small and wilted). All isolates caused symptoms of fusarium wilt on all cultivars except I3R-1, indicating that the isolates were race 3. The pathogen was reisolated from the discolored vascular tissue of symptomatic plants. Control plants remained asymptomatic, and the pathogen was not reisolated from the vascular tissue. Fusarium wilt of tomato caused by isolates of F. oxysporum f. sp. lycopersici races 1 and 2 has been reported previously; however, race 3 has not been reported in Korea (1). To our knowledge, this is the first report of isolates of F. oxysporum f. sp. lycopersici race 3 on tomato in Korea. References: (1) O. S. Hur et al. Res. Plant Dis. 18:304, 2012 (in Korean). (2) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Ames, IA, 2006. (3) K. O'Donnell et al. Proc. Nat. Acad. Sci. 95:2044, 1998. (4) M. Rep et al. Mol. Microbiol. 53:1373, 2004.


Plant Disease ◽  
2021 ◽  
Author(s):  
Francisco Bruno da Silva Café ◽  
Rhannaldy Benício Rebouças ◽  
Juvenil H. Cares ◽  
Cristiano Souza Lima ◽  
Francisco de Assis Câmara Rabelo Filho ◽  
...  

During a survey in 2018 for plant nematodes associated with roots and soil in cactus cultivation areas in Ceará State (3°44'48"S, 38°34'29"W), cysts were found on roots of mandacaru, Cereus jamacaru DC. This cactus is native to Brazil, can grow to 6-10 meters in height, and is widely distributed in the Northeast region (Romeiro-Brito et al. 2016) where it is used in construction, in disease remedies, as forage, and as an ornamental (Sales et al. 2014). Several cysts, second-stage juveniles (J2) and eggs extracted from the soil and roots, using sucrose centrifugation, were examined by scanning electron microscopy (SEM) and light microscopy (LM) to determine morphological and morphometric characteristics. Molecular characteristics were determined by DNA extraction from J2 and embryonated eggs using a protocol specific for Heteroderidae (Subbotin et al., 2018). The internal transcribed spacer sequence (ITS) region of the rDNA and D2-D3 regions of the 28S rDNA were amplified using the universal primers TW81 (5′-GTTTCCGTAGGTGAACCTGC-3′) and AB28 (5′-ATATGCTTAAGTTCAGCGGGT-3′), D2A(5′-ACAAGTACCGTGAGGGAAAGTTG-3′) and D3B(5′-TCGGAAGGAACCAGCTACTA-3′), respectively. To confirm that mandacaru is a host for C. cacti, six plantlets of mandacaru were inoculated with 1,800 eggs of the nematode, and kept in a greenhouse at 31 ± 3 ºC and irrigated daily. Six non inoculated mandacaru plantlets served as control treatment. Morphometric characteristics of cysts (n=35) were body length, excluding neck, 555.8 ± 87.8 (354,9 - 727,6) μm, body width 392.1 ± 63.4 (297.9 - 553.7) μm, neck length 63.5 ± 25.8 (49.8-105.0) μm, length to width ratio 1.4 ± 0.2 (1.0-1.8) μm and vulval cone length 48.4 ± 15.2 (40.7 –53.6) μm. Cysts had a rough surface, were lemon-shaped to rounded and had a zigzag cuticular pattern with a protruding vulval cone. They were circumfenestrate without underbridge and bullae, but with the presence of vulval denticles. Measurements of second-stage juveniles (n = 13) included the body length 511.2 ± 33.7 (452.7 - 551.5) μm, stylet length 28.0 ± 2.8 (25.4 - 34.0) μm, tail length 50.7 ± 5.1 (40.6 - 57.4) μm, tail hyaline region 22.7 ± 2.2 (18.9 – 27.1), with a = 20.9 ± 2.2 (17.7-24.3) μm, b = 5.4 ± 0.4 (5.1-5.8) μm, b'= 3.4 ± 0.4 (3.1-3.9) μm, c = 10.2 ± 1.3 (8.9-13.3) μm and c' = 3.8 ± 0.4 (3.0-4.5) μm. The observations of essential morphological characteristics for identification indicated that the species found on C. jamacaru was Cactodera cacti (Filipjev & Schuurmans-Stekhoven, 1941) Krall & Krall, 1978. The sequences of the studied rDNA regions were submitted to GenBank (ITS: MW562829 and D2–D3 regions of 28S: MW562830). The samples used for molecular analysis showed a high degree of sequence identity (99.59%) with C. cacti, from China, Iran and USA for the ITS region. The identity of the D2-D3 regions of 28S sequence was 99.54% with C. cacti isolates from Germany and 99.41% with isolates from USA. Phylogenetic analyses were performed using Maximum likelihood (ML) method for both individual loci, confirming the species as Cactodera cacti. All inoculated mandacaru plantlets showed C. cacti cysts on the roots after 60 days, confirming that mandacaru is a host for C. cacti. This species was reported in São Paulo State, in 2001, associated with ornamental cactus cultivated in pots, but plant species were not identified (Santos et al., 2001). The second report in Brazil was to Schlumbergera sp., an ornamental plant (Oliveira et al. 2007). In both studies, the nematode was not morphologically nor molecularly characterized. Cactodera cacti has been commonly associated with cactus worldwide (Esser, 1992). It has been reported in association with C. jamacaru was first reported in 2011 in China (Duan et al. 2012). This is the first report of the occurrence of C. cacti on C. jamacaru in field conditions in Brazil, and its presence in cactus cultivation areas with agricultural importance represents a threat to cactus production in the country.


Plant Disease ◽  
2014 ◽  
Vol 98 (12) ◽  
pp. 1742-1742
Author(s):  
J. H. Park ◽  
S. E. Cho ◽  
S. H. Lee ◽  
C. K. Lee ◽  
H. D. Shin

The Republic of Korea (South Korea) is the second largest chestnut producer in the world. Major cultivars planted in Korea, including cv. Daebo, Hyogo57, and Okkwang, are hybrids of Japanese chestnut (Castanea crenata) and Chinese chestnut (C. mollissima). Because of high perishability, most chestnuts harvested in September and October are preserved in cold rooms (0°C) for marketing. During a survey of postharvest diseases in April to August 2013, chestnut rots were continuously observed in cold rooms located in Buyeo County, Korea. Preliminary studies revealed that the most common agent of rot appeared to be a species of Mucor. When cut open, infected chestnuts showed partial interior discoloration varying from chalky white to dark brown. About 3 to 10% of chestnuts showed symptoms. Hyogo57 seemed to be the most susceptible variety with higher infection rates, up to 30% in some piles. Isolation was done by placing infected tissues on potato dextrose agar. A representative isolate was deposited in the Korean Agricultural Culture Collection (Accession No. KACC47727). Sporangiophores were mostly erect, branched sympodially, and hyaline. Sporangia were globose, pale yellow at first, then grayish brown at maturity. Columellae were obovoid to globose, subhyaline to pale brown, and usually with truncate base and collars. Sporangiospores were globose to irregular, and 4 to 10 μm in diameter. Chlamydospores were cylindrical to globose with oil drops. The fungus was identified as Mucor racemosus f. sphaerosporus (Hagem) Schipper based on the morphological characteristics and growth at low temperature (3). To conduct molecular analyses, genomic DNA was extracted with DNeasy Plant Mini Kits (Qiagen Inc., Valencia, CA). The primers ITS1/ITS4 and NL1/LR3 were used to amplify the internal transcribed spacer (ITS) region of rDNA and the D1/D2 region of the large subunit (4). The resulting 595-bp ITS sequences and 678 bp D1/D2 sequences were deposited in GenBank (Accession Nos. KJ769665 and KF769666). BLAST searches revealed that both the ITS sequences and D1/D2 sequences showed more than 99% similarity with those of M. racemosus f. sphaerosporus, respectively (JN939201 and AJ878775). To perform a pathogenicity test, a suspension of sporangiospores (1 × 105 spores/ml) was sprayed over 10 chestnuts cv. Hyogo57 wounded with a sewing needle and kept in plastic containers (0°C, 100% RH). Another 10 chestnuts wounded with a sewing needle and treated with sterile water served as controls. After 5 days, typical rots appeared on the inoculated chestnuts, whereas no symptoms were observed on controls. Koch's postulates were fulfilled with the re-isolation of M. racemosus from inoculated chestnuts. The pathogenicity test was carried out twice with similar results. M. hiemalis and M. mucedo have been recorded on chestnuts as postharvest pathogens in Switzerland (2) and Chile (1). To our knowledge, this is first report of postharvest rot of chestnut caused by M. racemosus f. sphaerosporus worldwide as well as in Korea. Further studies are necessary for control measures during cold storage of fresh chestnuts. References: (1) D. F. Farr and A. Y. Rossman. Fungal Databases. Syst. Mycol. Microbiol. Lab., online publication, ARS, USDA, Retrieved May 23, 2014. (2) M. Jermini et al. J. Sci. Food Agric. 86:877, 2006. (3) M. A. A. Schipper. Stud. Mycol. 12:1, 1976. (4) G. Walther et al. Persoonia 30:11, 2013.


Plant Disease ◽  
2021 ◽  
Author(s):  
Xiujing Hong ◽  
Shijia Chen ◽  
linchao Wang ◽  
Bo Liu ◽  
Yuruo Yang ◽  
...  

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.


Plant Disease ◽  
2008 ◽  
Vol 92 (5) ◽  
pp. 832-832 ◽  
Author(s):  
A. Aroca ◽  
R. Raposo ◽  
D. Gramaje ◽  
J. Armengol ◽  
S. Martos ◽  
...  

A field of Richter 110 rootstock mother plants in Valencia Province (eastern Spain) was surveyed during November 2006 to study the mycoflora of declining plants. Two canes with stunted leaves were collected from a plant with a reduced number of shoots. No cankers or vascular lesions were observed in the collected canes. Six wood chips (1 to 2 mm thick) were taken from one basal fragment (3 to 4 cm long) of each cane, surface sterilized in 70% ethanol for 1 min, and plated on malt extract agar supplemented with 0.5 g L–1 of streptomycin sulfate. Petri dishes were incubated for 7 days at 25°C. A fungus was consistently isolated from all samples that showed the following characteristics: colonies grown on potato dextrose agar (PDA) at 25°C developed a white, aerial mycelium that turned gray after 4 to 6 days and produced pycnidia after 1 month on sterile grapevine slivers of twigs placed on the PDA surface; conidia from culture were ellipsoidal, thick walled, initially hyaline, nonseptate, and measuring 20 to 25 (22.5) × 12 to 14 (13) μm; aged conidia were brown, 1-septate with longitudinal striations in the wall; and pseudoparaphyses variable in form and length were interspersed within the fertile tissue. The fungus was identified as Lasiodiplodia theobromae (Pat.) Griffon & Maubl. from the above characteristics (2). Identity was confirmed by analysis of the nucleotide sequences of the internal transcribed spacer (ITS) region from the rRNA repeat and part of the translation elongation factor 1-alpha (EF1-α) and the β-tubulin (B-tub) genes, as done elsewhere (1,3). BLAST searches at GenBank showed a high identity with reference sequences (ITS: 100%, EF1-α: 97%; B-tub: 99%). Representative sequences of the studied DNA regions were deposited at GenBank (Accession Nos.: ITS: EU254718; EF1-α: EU254719; and B-tub: EU254720). A pathogenicity test was conducted on 1-year-old grapevine plants cv. Macabeo grafted onto Richter 110 rootstocks maintained in a greenhouse. A superficial wound was made on the bark of 10 plants with a sterilized scalpel, ≈10 cm above the graft union. A mycelial plug obtained from the margin of an actively growing fungal colony (isolate JL664) was placed in the wound and the wound was wrapped with Parafilm. Ten additional control plants were inoculated with sterile PDA plugs. All control plants grew normally, and the inoculation wound healed 3 months after inoculation. Plants inoculated with L. theobromae showed no foliar symptoms in the same period, but developed cankers variable in size surrounding the inoculation sites. Vascular necroses measuring 8.4 ± 1.5 cm (mean ± standard error) developed in the inoculated plants that were significantly longer than the controls (0.3 ± 0.2 cm). The pathogen was reisolated from all inoculated plants and no fungus was reisolated from the controls. These results confirmed the pathogenicity of L. theobromae to grapevine and points to a possible involvement of L. theobromae in the aetiology of grapevine decline as previously reported (3,4). To our knowledge, this is the first report of L. theobromae isolated from grapevine in Spain. References: (1) J. Luque et al. Mycologia 97:1111, 2005. (2) E. Punithalingam. No. 519 in: Descriptions of Pathogenic Fungi and Bacteria. CMI, Kew, Surrey, UK, 1976. (3) J. R. Úrbez-Torres et al. Plant Dis. 90:1490, 2006. (4) J. M. van Niekerk et al. Phytopathol. Mediterr. 45(suppl.):S43, 2006.


Plant Disease ◽  
2021 ◽  
Author(s):  
Zong-ming Sheu ◽  
Ming-hsueh Chiu ◽  
Lawrence Kenyon

Mungbean (Vigna radiata L.) is routinely grown in the experimental fields at the headquarters of the World Vegetable Center (23°6'30.88"N, 120°17'51.31"E) for breeding, research and germplasm multiplication. In a spring 2016 mungbean trial, about 50% of the plants were affected with powdery mildew. The white, powdery-like patches first appeared on the upper leaf surfaces, and soon developed to grey patches on both sides of the leaves. Purple to brown discoloration appeared on the underside of the infected leaf. Microscopy examination revealed that the causal organism was not Erysiphe polygoni, which had previously been documented as the powdery mildew pathogen on mungbean in Taiwan (Hartman et al. 1993). The fungus produced typical structures of the powdery mildew Euoidium, anamorph of the genus Podosphaera. The mycelium consisted of septate, flexuous hyphae with indistinct appressoria. The erect conidiophores arising from superficial hyphae varied from straight or slightly curved to curled. Three to ten conidia were borne in long chains with crenate edges. Foot-cells were straight, cylindrical and measured 30 to 52 µm long. Conidia were hyaline, ellipsoid-ovoid to barrel-shaped, with fibrosin bodies, and measured 27 to 33 (mean = 30.4) × 15 to 20 (mean = 16.6) µm. Germ tubes were clavate and occasionally forked, and were produced from the lateral sites of the conidia. No chasmothecia were found in the samples. The morphological characteristics were consistent with P. xanthii (Castagne) U. Braun & Shishkoff (Braun & Cook 2012). To confirm the identity, the internal transcribed spacer (ITS) region of rDNA and partialβ-tubulin gene (TUB2) for the isolate MG3 were amplified with the primers ITS4/ITS5 (White et al. 1990) and BtuF5/BtuR7a (Ellingham et al. 2019), respectively. BLASTn analysis revealed the ITS sequence (MN833717) was 100% identical to many records of P. xanthii whereas the TUB2 sequence (MW363957) was 100% identical to a record of P. fusca (syn. P. xanthii; KC333362) in NCBI GenBank. A pathogenicity test was conducted by dusting conidia from an infected leaf onto six healthy four-week-old mungbean plants (cv ‘Tainan No. 3’). Another three plants were not inoculated and were used as control. All the plants were maintained in a greenhouse at 25 to 28°C. All inoculated plants developed powdery mildew symptoms after 10 days, whereas the control plants remained symptomless. To our knowledge, this is the first report of P. xanthii causing disease on mungbean in Taiwan. P. xanthii also has been reported on mungbean in Thailand (Meeboon et al. 2016), while other records referring to E. polygoni infecting Vigna spp. are from Brazil and Fiji (Farr & Rossman 2020). Although both P. xanthii and E. polygoni have now been reported as causing powdery mildew on mungbean in Taiwan, which species predominates or is more important remains unclear. A comprehensive survey with accurate species identification is required to develop effective management of the disease, particularly for resistance breeding.


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