A Flammulina velutipes CDNA library was constructed using mRNA from 7-day-old culture after fruiting induction, and a cDNA clone for the FVFD30 gene was selected by differential screening. The FVFD30 transcript was detected from 1 to 10day-old culture which forms primordia and particularly expressed at higher level in 7-day-old culture, but not in cultures before fruiting induction, several days after primordium formation and after that. It is suggested that the FVFD30 is involved in the initial stages of fruiting body development in F. velutipes. The FVFD30 cDNA contains an open reading frame encoding a putative protein of 319 amino acid residues (MW=34,592), which might belong to the oxidoreductase family.

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The factors producing the white line were studied in an attempt to develop the simplified methods of diagnosing of cultivated mushrooms infected with Pseudomonas tolaasii. White line were produced when P. tolaasii and Pseudomonas sp. (white line reacting organism : WLRO) were dually cultured on Pseudomonas agar F, King's B and TMGA medium. White lines produced on a PAF, and TMGA medium were clear and stable, but strains that can not be produced on a King's B medium had existed, and it was observed that white lines formed, if any, were not quite clear and tended to have disappeared when incubation was prolonged. When a distance between P. tolaasii and WLRO was within 20mm, clear white lines were produced, and the shorter the distance, the earlier the production was achieved. Only strains of P. tolaasii having pathogenicity in mushroom fruit bodies produced white lines, while none pathogenic mutants no longer had their capability of producing white lines. The capability of producing white lines was recognized in neither Erwinia spp nor epiphytic bacteria grown in mushroom species. When the tissue of diseased fruit bodies were dually cultured on PAF with WLRO, approximately 80% of tissues had definitely produced white lines 2days thereafter, and it was possible to diagnose the bacterial blotch disease directly without isolation of P. tolaasii.

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Fruit body production from a submerged culture mycelia of Favolus arcularius was examined. A glass bottle, 3 L in capacity, was used as the culture vessel, which contained 2 L of liquid medium composed of maltose and peptone as carbon and nitrogen sources, respectively. A large number of small pieces of mycelia was inoculated, and incubated at 27℃ for 120 h with agitation. The mycelial pellets were collected by filtration, placed into Petri dishes, and then the culture filtrates were poured to immerse up to a half of the height of mycelium pellets. The dishes were incubated to induce fruiting at 27℃ under light exposure. By incubating the dishes for 1-2 d, the aerial mycelia appeared on the surface of mycelium pellets. It grew primordia, that would develop into mature fruit bodies. However, the filamentous mycelia by submerged culture did not exhibit fruit body formation during incubation for 2 weeks. The optimal pellet size was estimated to be around 10 mm in diameter. One pellet could produce only one mature fruit body, although two or more numbers of primordia could arise on a pellet. Fruiting was completed within 10 days after the treatment of mycelium pellets for fruit body production.

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To obtain a better understanding of how metal proteinases (Mpase) function in vegetative mycelial maturation and fruit-body growth of Hypsizygus marmoreus, we examined changes in Mpase activity. Intracellular Mpase with pI 7.7 and pi 8.4 that were separated by fractionation were assayed during cultivation of this mushroom in sawdust rice bran medium. The activity of the Mpase of pI 8.4 increased markedly during vegetative mycelial growth and was maximal 25 days after inoculation. However, upon initiation of mycelial maturation, a decrease the activity of this Mpase was observed. The activity then increased again upon initiation of primordium formation of fruit-body. On the other hand, the level of Mpase with pI 7.7 was lower throughout vegetative mycelial growth and mycelial maturation. Then, we studied whether the Mpase of pI 8.4 produced in mycelia during vegetative mycelial growth is immunologically identical to the Mpase of pI 8.4 produced during development of fruit-body. Western blotting analysis revealed that those Mpases were identical.

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Shiitake mushroom, Lentinula edodes, was cultivated by using the steamed and hot water-extracted leaf residue of bamboo grass, Sasa senanensis, as a substitute for hardwood sawdust which is generally used as a basal substrate for the culture medium. The effects of the ratio of the leaf residue of bamboo grass to the birch sawdust on the fruit body production were investigated. Influences of the addition of the leaf residue on the mycelial growth and the shape of fruit bodies could not be observed. No significant difference in the fruit body yield was detected among the media up to 50% substitution of the residue for the sawdust. The cultures containing the residue at 25% and 50% of the basal substrate tended to produce relatively larger fruit bodies. These results indicate that replacement up to a half amount of hardwood sawdust by steamed and extracted residue of bamboo grass leaves can be applied to the commercial cultivation of L. edodes.

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