Miyato Higaki

Regeneration of protoplasts from hyphal strands of Volvariella volvacea

A series of experiments on the preparation and regeneration of protoplasts from hyphal strands ofVolvariella volvacea (Bull. ex. Fr.) Singer were conducted with the aim of optimizing the conditions for its efficient regeneration. One commercial (Vvcl) and two wild (EAAC-0001 and EAAC-0002) strains ofV. volvacea from the Philippines were used and subjected to varying conditions to determine the most efficient means for regeneration of their protoplasts. The effects of age and type of strain, pH, type and concentration of osmotic stabilizer, enzymatic composition, treatment time, temperature, reciprocal frequency during enzymatic lysis of the cell wall, and centrifugation conditions were investigated. Results showed that the three strains ofV. volvacea had varying responses in terms of yield, size, and ability of their protoplasts to regenerate into the protoplast regeneration medium. Among the three strains, EAAC-0002 had the highest rate of regeneration. The 5-day-old culture ofV. volvacea, when subjected to a combination of 2% Novozyme 234 and 0.2% chitinase in 0.6M mannitol (pH 6.0) for 3 h at 30°C, 90 strokes/min and centrifuged at 1100 g for 10 min; produced an efficient yield of protoplasts with a relatively high regeneration rate.

Physiological considerations for efficient mycelial colonization of Philippine strains of Volvariella volvacea

The nutritional and physical requirements for the efficient mycelial colonization ofVolvariella volvacea (Bull. ex. Fr.). Singer were elucidated with the percentage mycelial colonization and density as references. This investigation was limited to the evaluation of two commercial strains (designated Vvc1 and Vvc2) and two wild strains (designated EAAC-0001 and EAAC-0002) ofV. volvacea from the Philippines with the aim of providing baseline data on their physiological requirements. The four strains ofV. volvacea had varying preferences for carbon. Vve1 preferred polysaccharides (starch and cellulose), whereas Vvc2 grew luxuriantly at a relatively rapid rate in sugar alcohol (sorbitol). The two wild strains preferred starch as a carbon source. In terms of nitrogen utilization, soytone, peptone, and glycine supported efficient mycelial colonization of the four strains. The vitamin utilization test revealed that ascorbic acid, calcium pantothenate, and biotin are good sources. The mycelial growth performances of the strains were also evaluated on six dehydrated mycological media. Efficient colonization of Vvc1, Vvc2, and EAAC-0002 with dense mycelial growth was noted in mycological agar. EAAC-0001, on the other hand, grew more efficiently in malt extract agar. The Philippine strains ofV. volvacea grew luxuriantly when incubated at 35°C and pH 8.0 under dark and sealed conditions. Moreover, the relatively higher moisture content (70%) of the oolong tea leaf formulation favorably stimulated efficient mycelial colonization. Under optimum physiological conditions, Vvc1, Vvc2, and EAAC-0002 were fast-growing strains, whereas EAAC-0001 was a moderately growing type.

Termiticidal Performance of the Entomogenous Fungus Metarhizium anisopliae ESC1. (I). Growth Environment and Termiticidal Effect of M. anisopliae ESC1.

昆虫寄生菌Metarhizium anisopliae ESC 1分生胞子を製剤化した「バイオブラスト」の防蟻効果を研究した。滅菌土壌に処理したM.anisopliae ESC 1株の分生胞子は,イエシロアリとヤマトシロアリの両種に対して強い殺蟻効果を示した。その殺蟻効果は,処理土壌中の分生胞子数が多くなるほど強く,25℃から30℃で本菌は良く発芽・生育し,強い殺蟻効果を発現した。