Pheromone (PHB)-receptor (RCB) interaction in the mating pheromone response pathway of Lentinula edodes was investigated using synthetic PHBs. Functionality of the C-terminally carboxymethylated synthetic PHBs was demonstrated by concentration-dependent induction of a mating-related gene (znf2) expression and by pseudoclamp formation in a monokaryotic strain S1-11 of L. edodes. Treatment with synthetic PHBs activated the expression of homeodomain genes (HDs) residing in the A mating type locus, and of A-regulated genes, including znf2, clp1, and priA, as well as genes in the B mating type locus, including pheromone (phb) and receptor (rcb) genes. The synthetic PHBs failed to discriminate self from non-self RCBs. PHBs of the B4 mating type (B4 PHBs) were able to activate the mating pheromone response pathway in both monokaryotic S1-11 and S1-13 strains, whose B mating types were B4 (self) and B12 (non-self), respectively. The same was true for B12 PHBs in the B4 (non-self) and B12 (self) mating types. The synthetic PHBs also promoted the mating of two monokaryotic strains carrying B4-common incompatible mating types (A5B4 × A1B4). However, the dikaryon generated by this process exhibited abnormally high content of hyphal branching and frequent clamp connections and, more importantly, was found to be genetically unstable due to overexpression of mating-related genes such as clp1. Although synthetic PHBs were unable to discriminate self from non-self RCBs, they showed a higher affinity for non-self RCBs, through which the mating pheromone response pathway in non-self cells may be preferentially activated.
Lentinula* ; Pheromones* ; Shiitake Mushrooms*

The objectives of this study were to evaluate applicability of food waste compost (FWC) as a substrate for cultivation of Ganoderma lucidum, Lentinula edodes, and Pholiota adipose, and to determine contents of Ca, Mg, Na, and K in fruiting bodies (FB). FB yield per substrate in FWC-free controls was 53 +/- 4 g/kg for G. lucidum, 270 +/- 90 g/kg for L. edodes, and 1,430 +/- 355 g/kg for P. adipose. Substrates supplemented with FWC showed the highest FB production at FWC content of 10% for G. lucidum (64 +/- 6 g/kg), and 13% for L. edodes (665 +/- 110 g/kg) and P. adipose (2,345 +/- 395 g/kg), which were 1.2~2.5 times higher than the values for the controls. P. adipose contained higher amounts of mineral elements than the other species. Ca, Mg, Na, and K content in FB did not show a significant relation to FWC content.
Fruit* ; Ganoderma* ; Lentinula* ; Pholiota* ; Reishi* ; Shiitake Mushrooms* ; Soil*

The effects of aeration through lid filters on the hyphal growth of Lentinula edodes (oak mushroom) in sawdust cultivation bags were investigated. The aeration treatment levels were traditional 27 mm hole cotton plugs, cotton balls and combinations of seven hole sizes x two hole positions (up and under) in the lids covering plastic bags containing 1.4 kg sawdust medium at 63% moisture that had been autoclaved for one hour and inoculated with sawdust spawn of L. edodes strain 921. Aeration treatment effects were measured based on the CO2 concentration at the 15th wk, as well as the hyphal growth rate and degree of weight loss of bags every 14 days for 15 wk. In bags with traditional cotton plugs, the CO2 concentration was 3.8 +/- 1.3%, daily mean hyphal growth was 2.3 +/- 0.6 mm and daily mean weight loss was 0.84 +/- 0.26 g. In the bags with 15 mm diameter holes, the CO2 concentration was 6.0 +/- 1.6%, daily hyphal growth was 2.8 +/- 0.2 mm and daily weight loss was 0.86 +/- 0.4 g. The bags with 15 mm holes had a higher CO2 concentration and lower water loss than bags with other hole sizes, but the hyphal growth was not significantly different from that of other bags. The weight loss of bags increased proportionally relative to the lid hole sizes. Taken together, these results indicate that traditional cotton plugs are economically efficient, but 15 mm hole lids are the most efficient at maintaining hyphal growth and controlling water loss while allowing CO2 emissions.
Lentinula ; Plastics ; Shiitake Mushrooms ; Sprains and Strains ; Weight Loss

Protoplast fusion is a useful technique for establishing fungal hybrids to overcome the natural barriers. The ultrastructure of protoplast and its fusion process were observed using a scanning electron microscopy(SEM) and a transmission electron microscopy(TEM). The protoplasts were variable in size from 0.5~15microm in diameter, and the mean diameter was about 3~5microm. It was impossible to discriminate protoplasts of Lentinula edodes from protoplasts of Coriolus versicolor by size and surface structure. Big aggregates of the dehydrated protoplasts were observed, after polyethylene glycol 4000 treatment. Nucleus, mitochondria, lipid granules and various vesicles having granules were scattered in the cytoplasm. The vesicles were heterogeneous in size and vary from one protoplast to another. The fused membrane layer of the two protoplasts was observed. Time protoplast membrane contact and reorganization of membrane components were essential condition for protoplast fusion. Transmission electron micrograph showed fused protoplasts and flattening of the cells in the area of the membrane contact. We hope that our electron microscopic observations provide some insights into the understanding of the fusion process of protoplast in fungi.
Cytoplasm ; Fungi ; Hope ; Lentinula* ; Membranes ; Mitochondria ; Polyethylene Glycols ; Protoplasts* ; Shiitake Mushrooms*

Sikhae is a Korean traditional beverage of saccharified rice. Its factory waste (SFW) is usually thrown away instead of being used. We developed a cheap substrate of SFW for use in liquid spawn that is known for its higher fruit body yields than grain spawn in sawdust cultivation. Mycelia of Lentinula edodes ASI 3046, which is regarded as the most suitable strain for sawdust cultivation, were cultured on six kinds of previous known media and SFW. As the seven kinds of media were applied, a Sikhae Factory Waste (SFW) was most excellent in growth. The dried mycelial weight in SFW was almost four times as much as that in the other media. In the flask culture, optimum culture conditions for the mycelial growth were obtained after 13 days of cultivation at media volume of 100 ml, 100 rpm, initial pH 4.5, and 25degrees C. The best mycelial growth was observed when MgSO4 . 7H2O and D-sucrose were added as a supplement in SFW. SWM must be a remarkable medium for L. edodes because of its simple preparation and low cost.
Beverages ; Edible Grain ; Fruit ; Hydrogen-Ion Concentration ; Lentinula* ; Shiitake Mushrooms* ; Waste Water*

Differential display of reverse transcription (DDRT)-PCR was conducted to have a profile of the differentially expressed genes during the formation of fruiting body of Lentinus edodes. The lines of L. edodes (ImHyup-1) employed were cultivated in the artificial blocks of sawdust, and the fruiting body was induced from the mycelia or the mass protruded from the brown surface of the sawdust blocks. RNAs were prepared from the four different developmental stages; mycelial, primordial, and stipes and pileus of fruiting body. The fragments of cDNA were synthesized from the combinations of the arbitrary primers and 3' one anchored Oligo-dT primer. Twelve combinations using the primers have been tested, and among them nineteen bands were identified as differentially expressed. Those genes were further analyzed by DNA sequencing and followed by homology search. Characterization of one clone was conducted as a preliminary data and more are under investigation.
Clone Cells ; DNA, Complementary ; Fruit ; Lentinula* ; Morphogenesis* ; Reverse Transcription ; RNA ; Sequence Analysis, DNA ; Shiitake Mushrooms*

Sixteen genomic DNA simple sequence repeat (SSR) markers of Lentinula edodes were developed from 205 SSR motifs present in 46.1-Mb long L. edodes genome sequences. The number of alleles ranged from 3–14 and the major allele frequency was distributed from 0.17–0.96. The values of observed and expected heterozygosity ranged from 0.00–0.76 and 0.07–0.90, respectively. The polymorphic information content value ranged from 0.07–0.89. A dendrogram, based on 16 SSR markers clustered by the paired hierarchical clustering' method, showed that 33 shiitake cultivars could be divided into three major groups and successfully identified. These SSR markers will contribute to the efficient breeding of this species by providing diversity in shiitake varieties. Furthermore, the genomic information covered by the markers can provide a valuable resource for genetic linkage map construction, molecular mapping, and marker-assisted selection in the shiitake mushroom.
Alleles ; Breeding ; DNA* ; Gene Frequency ; Genetic Linkage ; Genetic Variation ; Genome ; Lentinula* ; Methods ; Microsatellite Repeats ; Shiitake Mushrooms*

This study was conducted to understand how osmotic potentials in Lentinula edodes tissues are related to water contents and how they change while a mushroom matures. Water content and osmotic potential of L. edodes mushroom tissues from log cultivation and sawdust cultivation were measured and the relationships were analyzed. Osmotic potentials in the tissues were exponentially proportional to their moisture contents and there were strain differences in the potentials. Strain 290 has lower osmotic potential than strain 302, in the tissues at the same water content. As the mushrooms mature, tissue water content maintained ca 94% in head tissues and ca 90% in gills, but significantly decreased from ca 90% to 82% in the stipe tissues. Osmotic potential changes were similar to the tissue water content changes as the mushrooms mature. While osmotic potentials maintained -0.25 to -0.45 MPa in head and gill tissues, the potentials greatly decreased from -0.65 to -1.33MPa in stipe tissues. Our results show that osmotic potentials in L. edodes tissues are exponentially proportional to tissue water contents, that strains differ in osmotic potential related to water, and that stipe tissues can still have nutritional value when they mature.
Agaricales ; Animals ; Gills ; Head ; Lentinula ; Nutritive Value ; Shiitake Mushrooms ; Sprains and Strains

Trichoderma spp. cause large crop losses of the cultivated shiitake mushroom, Lentinula edodes. We bred several shiitake strains that are resistant to Trichoderma spp. using di-mon mating to establish a useful method for controlling the greenmold disease. We examined the competitive ability of L. edodes against Trichoderma spp. using a dual culture system to select resistant strains. By screening Trichoderma-resistant strains, we found that among 11 parental strains, 4 strains, including KFRI 36, were confirmed resistant strains. They showed especially strong resistance to T. harzianum, which formed deadlock after mycelial contact and then invaded into the territory of T. harzianum. KFRI 171 also showed resistance to T. atroviride strains. Among 13 strains, which were made by hybridization of shiitake strains, 5 were confirmed to be resistant to Trichoderma, including KFRI 58-1. Their resistance was not correlated to the resistant activity of their parents' strains. Two strains lose resistance and two strains acquire resistance compared to their parents' strains. In SEM observation, the mycelium of L. edodes at the interaction zone of Lentinula-Trichoderma was rugged and swollen by T. harzianum.
Breeding ; Chimera ; Humans ; Lentinula ; Mass Screening ; Mycelium ; Parents ; Shiitake Mushrooms ; Trichoderma

The culture filtrate of Lentinula edodes shows potent antimicrobial activity against the plant pathogenic bacteria Ralstonia solanacearum. Bioassay-guided fractionation was conducted using Diaion HP-20 column chromatography, and the insoluble active compound was not adsorbed on the resin. Further fractionation by high-performance liquid chromatography (HPLC) suggested that the active compounds were organic acids. Nine organic acids were detected in the culture filtrate of L. edodes; oxalic acid was the major component and exhibited antibacterial activity against nine different phytopathogenic bacteria. Quantitative analysis by HPLC revealed that the content of oxalic acid was higher in the water extract from spent mushroom substrate than in liquid culture. This suggests that the water extract of spent L. edodes substrate is an eco-friendly control agent for plant diseases.
Agaricales ; Bacteria* ; Chromatography ; Chromatography, High Pressure Liquid ; Chromatography, Liquid ; Lentinula* ; Oxalic Acid* ; Plant Diseases ; Plants ; Ralstonia solanacearum ; Shiitake Mushrooms* ; Water