Sung Kee Hong

Morphology, Molecular Phylogeny and Pathogenicity of Colletotrichum panacicola Causing Anthracnose of Korean Ginseng

Colletotrichum panacicola isolates were obtained from anthracnose lesions of Korean ginseng and compared with four Colletotrichum species in morphology, molecular phylogeny and pathogenicity. Based on morphological characteristics, C. panacicola was easily distinguished from Colletotrichum gloeosporioides but not from Colletotrichum higginsianum, Colletotrichum destructivum and Colletotrichum coccodes. A phylogenetic tree generated from ribosomal DNA-internal transcribed spacer sequences revealed that C. panacicola is remarkably distinguished from C. gloeosporioides and C. coccodes but not from C. higginsianum and C. destructivum. However, molecular sequence analysis of three combined genes (actin + elongation factor-1α + glutamine synthatase) provided sufficient variability to distinguish C. panacicola from other Colletotrichum species. Pathogencity tests showed that C. panacicola is pathogenic to Korean ginseng but not to other plants. These results suggest that C. panacicola is an independent taxon distin-zguishable from C. gloeosporioides and other morphologically similar Colletotrichum species.

Identification and Distribution of Two Fungal Species Causing Sclerotial Disease on Mulberry Fruits in Korea

A total of 520 overwintered sclerotia were collected from surface of soil under mulberry trees in six locations in Korea during February in 2006 and 2007. The collected sclerotia were tested for their germination in vitro and identified based on their morphological characteristics. Out of all sclerotia tested, 523% of the sclerotia germinated and produced two types of apothecia. The two types of fungi occurred from the sclerotia at the ratio of 49.8 vs. 50.2. The fungal type with cup-shaped apothecia was identified as Ciboria shiraiana and another type of fungus with club-shaped apotheeia as Scleromitrula shiraiana. Taxonomy and distribution of the two sclerotial fungi were described and discussed.

Leaf Spot and Stem Rot on Wilford Swallowwort Caused by Stemphylium lycopersici in Korea

In June 2012, leaf spot and stem rot were observed on Wilford Swallowwort plants grown in Cheonan, Korea. Three fungal isolates obtained from the diseased leaves and stems were identified as Stemphylium lycopersici, based on morphological, cultural, and molecular characteristics and pathogenicity. This is the first report of leaf spot and stem rot on Wilford Swallowwort caused by S. lycopersici.

Mating Types and Optimum Culture Conditions for Sexual State Formation of Fusarium fujikuroi Isolates.

Twenty-five isolates of Fusarium fujikuroi acquired from rice seeds and rice plants evidencing symptoms of Bakanae disease were evaluated to determine their mating types and characterize the formation of their sexual state. The mating types of the isolates were evaluated via multiplex PCR with the diagnostic primers of the mating-type (MAT) region: GFmat1a, GFmat1b, GFmat2c, and GFmat2d. Among the 25 isolates, 11 were identified as MAT-1 (male), and 14 as MAT-2 (female). Four MAT-1 isolates and three MAT-2 isolates were mated and cultured to evaluate the optimal culture conditions for the production of their sexual states. Among four tested media, 10% V8 juice agar proved optimal for the perithecial production of the isolates. The isolates also generated the largest numbers of perithecia when incubated at 23°C in alternating cycles of 12 hr fluorescent light and NUV fluorescent light and 12 hr darkness.

Identification of Microdochium bolleyi Associated with Basal Rot of Creeping Bent Grass in Korea

Symptoms of basal rot occurred sporadically on creeping bent grasses growing at a golf course in Hampyeong, Korea in April 2007. Ten isolates of Microdochium sp. were obtained from leaves and crowns of the diseased bent grasses. All isolates were identified as Microdochium bolleyi based on morphological, cultural, and molecular characteristics. This is the first report on M. bolleyi associated with basal rot on creeping bent grass in Korea.

Occurrence of Stem-end Rot on Mango Fruits Caused by Lasiodiplodia theobromae in Korea

Mango ( Mangifera indica L.), an economically important fruit cropin the subtropics and tropics, was recently introduced into Korea. Itis being cultivated over an area of 20 ha with an annual productionof approximately 300 tons in Jeju Island. In July 2010, stem-end rotswere found on fruits of mango (cv. Irwin) grown in a greenhouse inJeju. In the early stage, the affected fruits appeared as small brownspots. As they enlarged, the lesions became circular, dark brown toblack and water-soaked patches and progressed from the surface toinner part of the fruit (Fig. 1A). Two fungal isolates were obtained from lesions on mango fruits.Cultures were initially white to smoke grey, with fluffy aerialmycelium on potato dextrose agar and became gray or black with afew medium grays in laterstage (F ig.1B) . The pycnidia produced onthe 14-day-old cultures (Fig. 1C) were simple, often aggregated,stromatic, ostiolate and hairy. Paraphyses within the pycnidia werehyaline,cy lindrica septate, occasionallyl, branched. Conidiogenouscells were hyaline, thin-walled, smooth, cylindrical, holoblastic(Fig. 1D). Young conidia were hyaline, unicellular and subovoid toellipsoid, with a granular content (Fig. 1E). Mature conidia wereone-septate, dark brown, thick walled, ellipsoidal, with longitudinalstriations (Fig. 1F) and measured 17.5 −26.8 × 12.3 −17.1 μm (mean22.7 × 14.7 μm). No teleomorph was observed in culture. The twofungal isolates were identified as Lasiodiplodia theobromae (Pat.)Griffon & Maubl. [Synonym: Botryodipodl ia theobromae Pat].bas edon their morphological and cultural characteristics, and correspondedto previous description for the species (Alves et al., 2008).To confirm the identification, the complete internal transcribedspacer (ITS) rDNA regions and translation elongation factor 1-alpha (EF1-α) of the two fungal isolates, ML1001 and ML1005were amplified using the primers ITS1/ITS4 and EF1-688F/EF1-1251R asd escribed by Alvese at . (20l 08) and sequencedTh . ere sultingsequences were deposited in GenBank (Accession numbersJN542561 and JN542562 for ITS rDNA, JN542563 and JN542564for EF1-α). Sequences of reference isolates were retrieved fromGenBank. A phylogenetic tree derived from combined sequences ofITS rDNA and EF1-α was constructed by the neighbor-joiningmethod with Kimura’s two-parameter distance model using MEGAversion 5.0. In the phylogenetic tree, the present isolates placed inthe same clade with L. theobromae in GenBank and clearlydistinguished from the closely related Lasiodiplodia species, L.parva and L. pseudotheobromae (Fig. 2).Pathogenicity tests were made on mango fruits. The fruits wereperforated with 3 mm cork borer, followed by drop-inoculated with100 μl of conidial suspensions (5×10

Occurrence of Gray Mold on Blueberry Trees Caused by Botrytis cinerea in Korea

Gray mold symptoms were frequently observed on green twigs, blossoms, leaves, and fruits of blueberry trees grown in greenhouses in Cheongyang, Dangjin, Daejeon, and Jeju during disease survey in eight locations of Korea from 2007 to 2010. The disease symptoms were not observed in the fields of the other locations investigated. The disease incidence ranged 1~30% in the greenhouses investigated. A total of 27 single spore isolates of Botrytis species were obtained from the gray mold symptoms, and all the isolates were identified as Botrytis cinerea based on their morphological and cultural characteristics. Four isolates of the fungus were tested for pathogenicity to leaves of four varieties of blueberry trees by artificial inoculation with conidial suspensions. All the tested isolates caused gray mold symptoms on the leaves, which were similar to those observed in the greenhouses. This is the first report that B. cinerea causes gray mold of blueberry trees grown in greenhouses in Korea.

Occurrence of Anthracnose on Fruits of Asian Pear Tree Caused by Colletotrichum acutatum.

Anthracnose symptoms often occurred on fruits of Asian pear trees grown in Anseong, Naju, Seonghwan and Pyeongtaek in Korea during the harvesting period from 2000 to 2005. A total of 28 isolates of Colletotrichum sp. were obtained from the anthracnose symptoms. Ail the isolates were identified as Colletotrichum acutatum based on their morphological and cultural characteristics. Four isolates of the fungus were tested for pathogenicity to fruits of Asian pear tree by artificial inoculation. All the isolates induced anthracnose symptoms on the fruits by wound inoculation but not by unwound inoculation. The anthracnose symptoms induced by artificial inoculation were similar to those observed in the orchard. This is the first report of anthracnose of Asian pear tree caused by Colletotrichum acutatum.

Characterization of Bacillus amyloliquefaciens DA12 Showing Potent Antifungal Activity against Mycotoxigenic Fusarium Species

In an attempt to develop a biological control agent against mycotoxigenic Fusarium species, we isolated Bacillus amyloliquefaciens strain DA12 from soil and explored its antimicrobial activities. DA12 was active against the growth of mycotoxigenic F. asiaticum, F. graminearum, F. proliferatum, and F. verticillioides both in vitro and in planta (maize). Further screening using dual culture extended the activity range of strain DA12 against other fungal pathogens including Botrytis cinerea, Colletotrichum coccodes, Endothia parasitica, Fusarium oxysporum, Raffaelea quercus-mongolicae, and Rhizoctonia solani. The butanol extract of the culture filtrate of B. amyloliquefaciens DA12 highly inhibited the germination of F. graminearum macroconidia with inhibition rate 83% at a concentration of 31.3 μg/ml and 100% at a concentration of 250 μg/ml. The antifungal metabolite from the butanol extract was identified as iturin A by thin layer chromatography-bioautography. In addition, volatile organic compounds produced by DA12 were able to inhibit mycelial growth of various phytopathogenic fungi. The volatile compounds were identified as 2-heptanone, 5-methyl heptanone and 6-methyl heptanone by gas chromatography-mass spectrometry (GC-MS) analysis. These results indicate that the antagonistic activity of Bacillus amyloliquefaciens DA12 was attributable to iturin A and volatile heptanones, and the strain could be used as a biocontrol agent to reduce the development of Fusarium diseases and mycotoxin contamination of crops.

Fungicide Resistance of Gibberella fujikuroi Isolates Causing Rice Bakanae Disease and Their Progeny Isolates

A total of 25 isolates of Fusarium fujikuroi were obtained from diseased rice plants in Korea from 2006 to 2007 to assess their resistance against fungicides prochloraz and benomyl + thiram. Minimal inhibitory concentration (MIC) values of F. fujikuroi isolates were examined by agar dilution method. Most of the isolates were sensitive to the fungicides. Out of 25 isolates, six were resistant to prochloraz and three to benomyl + thiram. In addition, the isolates CF245, CF249 and CF337 showed resistant to both fungicides. The progenies ( isolates) obtained through two different crosses between sensitive parental isolates(CF202, CF232 and CF179) and resistant parental isolate (CF337) were evaluated for their mycelial growth at different temperatures and resistance against fungicides. Mycelial growth rate of isolates originated from CF202 CF232 was similar to the parental isolates. However mycelial growth rate of isolates originated from CF179 CF337 was faster than their parent isolates. In case of prochloraz, distribution ratio of sensitivity(S) to resistance(R) against to the fungicide of isolates originated from CF202 CF232 and CF179 CF337 was 86 : 14 and 78 : 22, respectively. In case of benomyl+thiram, all the isolates originated from CF202 CF232 were sensitive to the fungicide, however ratio of sensitivity(S) to resistance(R) against to the fungicide of isolates originated from CF179 CF337 was 35 : 65.