Min Woo Hyun

Fungal and Plant Phenylalanine Ammonia-lyase

Abstract L-Phenylalanine is one of the essential amino acids that cannot be synthesized in mammals in adequate amounts to meet the requirements for protein synthesis. Fungi and plants are able to synthesize phenylalanine via the shikimic acid pathway. L-Phenylalanine, derived from the shikimic acid pathway, is used directly for protein synthesis in plants or metabolized through the phenylpropanoid pathway. This phenylpropanoid metabolism leads to the biosynthesis of a wide array of phenylpropanoid secondary products. The first step in this metabolic sequence involves the action of phenylalanine ammonia-lyase (PAL). The discovery of PAL enzyme in fungi and the detection of 14CO2 production from 14C-ring-labeled phenylalanine and cinnamic acid demonstrated that certain fungi can degrade phenylalanine by a pathway involving an initial deamination to cinnamic acid, as happens in plants. In this review, we provide background information on PAL and a recent update on the presence of PAL genes in fungi.

Fungi Isolated from Pine Wood Nematode, Its Vector Japanese Pine Sawyer, and the Nematode-Infected Japanese Black Pine Wood in Korea

Pine tree death caused by pine wood nematode (PWN) involves phoretic relationships between PWN and its vector Japanese pine sawyer beetle (JPS). In an effort to understand the diversity of fungi involved in PWN life cycle, a total of 176 fungal isolates were collected from PWNs, adults and larvae of JPS, PWN-diseased Japanese black pine that was cut down in 2005 at Jinju, Korea. Based on microscopic observation and colony morphology, and sequence analysis of the ITS rDNA, the fungal isolates were identified at the level of genus. Three genera including Mucor, Ophiostoma, and Penicillium were identified from PWN. Two genera of Ophiostoma and Penicillium were discovered from JPS larvae. From JPS adult beetles, nine genera of Aspergillus, Gibberella, Hypocrea, Irpex, Leptosphaeria, Ophiostoma, Penicillium, and Plectosphaerella and unknown basidio-myeetes were found. Ten genera from PWN-infeeted wood were confirmed as Bionectria, Botrytis, Camarops, Fusarium, Hypocrea, Nectrtia, Mucor, Ophiostoma, Penicillium, and Trichoderma. Penicillium and Ophiostoma were commonly distributed on PWN and its vector and host. This is first report of the fungi associated with PWN and its vector and host in Korea.

Identification and Characterization of Eurotium rubrum Isolated from Meju in Korea.

We isolated and identified a strain of Eurotium rubrum from Meju that has not been reported in Korea. This fungus is yellowish brown; reverse dark brown on CYA and PDA while yellow on 2% MEA at 25°C. Cleistothecia are first bright yellow and gradually turned brown. Mycerial growth on CYA attained a diameter of 30 mm at 20°C, 37 mm at 25°C and 32 mm at 30°C after 15 days. The isolate grew slower on 2% MEA (< 20 mm 15 days at 25°C) compared to CYA and PDA (< 40 mm 15 days at 25°C). Cleistothecia are superficial, yellow to light brown, globose to subglobose, 40~75 μm in diameter. Asci are 8-spored and globose to subglobose 8~11 μm. Ascospores are disciform, 4.0~5.0 μm in length and 4.2~4.5 μm in width. Conidia are ovate or bacillar, finely roughened to densely spinulose, 4.6~6.0 μm in length and 3.0~4.3 μm in width. Compared to known Eurotium rubrum, the Korean isolate showed 99% sequence similarity in ITS rDNA (554 bp) and calmodulin (750 bp) gene and 100% in β-tubulin (1016 bp) gene. The E. rubrum isolate also had weak β-glucosidase and protease activities.

Detection of Cellulolytic Activity in Ophiostoma and Leptographium species by Chromogenic Reaction

To understand the ability of producing cellulolytic enzyme activity in the sapstaining fungi, four species of Ophiostoma and two species of Leptographium were investigated in the culture media containing each of cellulose substrates such as CM-cellulose, Avieel and D-cellobiose and each of chromogenic dyes such as Congo-Red, Phenol Red, Remazol Brilliant Blue and fry plum Blue. When the fungi were grown for 5~7 days at 25°C, the formation of clear zone by chromogenic reaction around the margin of the fungal colony was demonstrated in all the culture media Congo-Red containing CM-cellulose. There was difference in the formation of clear zone among the dyes. Only Ophiostoma setosum and Leptographium spp. showed cellulolytic activity to the three substrates. Overall, the results of this study show that ophiostomatoid sapstaining fungi can produce cellulolytic enzymes.

Ophiostoma ips from Pinewood Nematode Vector, Japanese Pine Sawyer Beetle (Monochamus alternatus), in Korea

Abstract Japanese pine sawyer beetle (Monochamus alternatus) is an economically important pest in coniferous trees. Ophiostoma ips was isolated from the beetle and identified based on analysis of morphological properties and the β-tubulin gene sequence. The fungus easily produced perithecia with a long neck on malt extract agar and its ascospores were rectangular shaped. This is first report of Ophiostoma species associated with the pinewood nematode vector beetle in Korea.

Filamentous Fungi Isolated from Platypus koryoensis, the Insect Vector of Oak Wilt Disease in Korea.

The ambrosia beetle, Platypus koryoensis, is a serious pest of oak trees in Korea. In this study we investigated filamentous fungi present in the body of the beetle. Fourteen genera of filamentous fungi belonging to Ascomycota and Basidiomycota were isolated. Among the isolated fungi, some were able to produce wood degrading enzymes. This is first report of fungi associated with P. koryoensis.

New Record of Mariannaea elegans var. elegans in Korea

A Mariannaea fungus was isolated during investigation of an elm tree infested with unidentified beetles. Based on morphological characteristics and molecular analysis of the internal transcribed spacer rDNA sequence, the fungus was identified as Mariannaea elegans var. elegans. Fungal growth was better on malt extract agar than on potato dextrose agar and oatmeal agar. Optimal temperature and pH for growth of the fungus were 30°C and pH 7.0, respectively. The fungus was found to have the ability to produce extracellular enzymes such as amylase, β-glucosidase, cellulase, and protease. This is first report on M. elegans var. elegans in Korea.

Identification and Characterization of Gliocladium viride Isolated from Mushroom Fly Infested Oak Log Beds Used for Shiitake Cultivation

Abstract A green mold species that has not previously been reported in Korea was isolated from oak log beds used for shiitake (Lentinula edodes) cultivation that were infested by mushroom flies. In this study, we identify the mold species as Gliocladium viride (an anamorph of Hypocrea lutea) and describe its myeologieal properties. The fungus was cottony on both potato dextrose agar (PDA) and Czapek yeast extract agar (CYA), but was colored white on PDA and became yellowish green and brown on CYA. Mycelial growth on PDA attained a diameter of 73 mm at 30°C after 5 days. The fungus grew faster on malt extract agar (> 80 mm, 5 days at 25°C) compared to CYA and PDA (< 68 mm, 5 days at 25°C). Penieillate eonidiophores of the fungus are hyaline, smooth walled, branching above ty pically in four stages, and 120–240 pm in length. Club-shaped or slender phialides are formed on the metulae. Conidia of the fungus were ovate and elliptic, yellowish brown and green, and 2.5~3.0 μm × 1.8–2.3 μm in size. Typically, slimy conidia are formed in a mass and colored brown to dark green to almost black. The internal transcribed spacer rDNA and translation elongation factor 1 alpha gene sequences of the fungus isolated here show 99% identity with previously identified G viride strains.

Coleopteran and Lepidopteran Hosts of the Entomopathogenic Genus Cordyceps sensu lato

Entomophthoralean and ascomycetous fungi are the two major groups known to parasitize arthropods in almost every terrestrial habitat of the earth. Within Ascomycota, Cordyceps sensu lato is a large genus with more than 400 spp. described on numerous orders of Arthropoda. Among the hosts of Cordyceps, Coleoptera and Lepidoptera are the two major orders. Out of the estimated 200 Cordyceps spp. recorded on coleopteran and lepidopteran hosts, we have documented 92 spp. based on the available information of their host species. Among coleopteran hosts, Scarabaeidae and Elateridae are the two major families. Similarly, among lepidopterans, Hepialidae is the largest host family. Cordyceps militaris shows the widest host range, extending to 2 orders, 13 families, and 32 spp. We hope such accumulative work will be useful as a quick reference for interested biologists, forest ecologists, biocontrol researchers, and fungal and insect taxonomists to apprehend host range and host specificities of Cordyceps fungi.

Mariannaea samuelsii Isolated from a Bark Beetle-Infested Elm Tree in Korea.

During an investigation of fungi from an elm tree infested with bark beetles in Korea, one isolate, DUCC401, was isolated from elm wood. Based on morphological characteristics and phylogenetic analysis of the internal transcribed spacer and 28S rDNA (large subunit) sequences, the isolate, DUCC401, was identified as Mariannaea samuelsii. Mycelia of the fungus grew faster on malt extract agar than on potato dextrose agar and oatmeal agar media. Temperature and pH for optimal growth of fungal mycelia were 25°C and pH 7.0, respectively. The fungus demonstrated the capacity to degrade cellobiose, starch, and xylan. This is the first report on isolation of Mariannaea samuelsii in Korea.