The synergistic effects of high hydrostatic pressure (HHP), mild heating, and amino acids on the germination of Clostridium sporogenes spores were examined by determining the number of surviving spores that returned to vegetative growth after pasteurization following these treatments. Pressurization at 200 MPa at a temperature higher than 40°C and treatment with some of the 19 L-amino acids at 10 mM or higher synergistically facilitated germination. When one of these factors was omitted, the level of germination was insignificant. Pressures of 100 and 400 MPa were less effective than 200 MPa. The spores were effectively inactivated by between 1.8 and 4.8 logs by pasteurization at 80°C after pressurization at 200 MPa at 45°C for 120 min with one of the amino acids with moderate hydrophobicity, such as Leu, Phe, Cys Met, Ala, Gly, or Ser. However, other amino acids showed poor inactivation effects of less than 0.9 logs. Spores in solutions containing 80 mM of either Leu, Phe, Cys, Met, Ala, Gly, or Ser were successfully inactivated by pasteurization by more than 5.4 logs after pressurization at 200 MPa at 70°C for 15 to 120 min. Ala and Met reduced the spore viability by 2.8 and 1.8 logs, respectively, by pasteurization at a concentration of 1 mM under 200 MPa at 70°C. These results indicate that germination of the spores is facilitated by a combination of high hydrostatic pressure, mild heating, and amino acids.

Scales excised from lily bulblets were cultured on MS medium supplemented with 0.044 or 4.4 lM BA in the dark for 180 days. The culture period was divided into stage 1 (day 0-30), stage 2 (day 31-90) and stage 3 (day 91-180). The scales were cultured at 25°C in stage 1, 25°C or 8°C in stage 2, and 25°C in stage 3. When the scales were cultured on medium with 4.4 lM BA at 25°C for 180 days, bulblets with and without an elongated stem were produced. The percentage of bulblets with elongated stems greatly increased when the scales had been cultured at 8°C in stage 2. On medium with 0.044 lM BA, only bulblets without elongated stems were produced. The diameter of shoot primodia significantly enlarged in bulblets produced on medium with 4.4 lM BA at 8°C in stage 2 and no such enlargement occurred under the other conditions. Nearly square parenchyma cells were observed in the non-elongated shoot primodia in the former bulblets but not in the latter. These cells changed into longitudinally rectangular ones in the internode of elongated stems. Procambium was arranged almost parallel to the shoot axis in the stem of bulblets in the medium with 4.4 lM BA, but not in the medium with 0.044 lM BA.

Factors affecting the initiation and elongation of stems from bulblets developed on the scales of 'White Aga' (Lilium x formolongi) were examined by low-temperature treatment of bulbscales cultured in darkness or by culturing them in a medium with colchicine at 25°C. Scale explants excised from in vitro-cultured bulblets were cultured for a total of 180 days in darkness. First, they were cultured in MS medium with 0.54 µ µ µ µM NAA and 4.4 or 44 µ µ µ µM BA at 25°C for 30 days, and then further subcultured under the following conditions: (1) continuously cultured at 25°C for 150 days (control) on MS medium with 4.4 µ µ µ µM BA; (2) cultured at 8°C for 60 days, followed by culture at 25°C for 90 days (low-temperature treatment) on MS medium with 4.4 µ µ µ µM BA; (3) cultured on MS medium with 4.4 µ µ µ µM BA and 1 µ µ µ µM or 10 µ µ µ µM of colchicine at 25°C for 30 days, followed by culture on MS medium with 4.4 µ µ µ µM BA at 25°C for 120 days (colchicine treatment). Stem elongation from bulblets developing on the scales was observed in bulblets subjected to both low-temperature and colchicine treatment. Since both treatments, which inhibit the polymerization of microtubules (MTs) and stimulate the depolymerization of MTs, affected stem elongation from bulblets, there might be some relationship between the inhibition of polymerization as well as the stimulation of depolymerization of MTs and the induction and elongation of stems in bulblets.

Considering the demand for transforming poultry waste into eco-friendly manure, we carbonized chicken manure at 402, 449 and 528°C and determined the physicochemical properties. We evaluated the effectiveness of the ensuing carbonized chicken manure (CCM) as a fertilizer using Brassica rapa, var. komatsuna and Oryza sativa L., var. japonica, cv. Koshihikari, in the upland and the paddy field soil, respectively. Herein, we carried out duplicate treatment of CCM (carbonized at 528°C) either alone or in combination with nitrogen and measured the growth from three or more plants. The plots with the recommended chemical fertilizers containing nitrogen (N), phosphorus (P) and potassium (K) and with no fertilizers were used as controls for growth measurement. The vessel tests indicated that when applied alone, the CCM plot could not support the normal growth of both the plants. However, the CCM plot restored the plant growth with simultaneous application of nitrogen, and the growth in the CCM ? N plot was comparable to that observed in the NPK plot. By estimating the growth in the NPK plots as 100%, the growth percentages of fresh weights of komatsuna were 12% in the CCM plot, 79% in the N plot and 99% in the CCM ? N plot. Likewise, the plant heights of Koshihikari were 43% in the CCM plot, 77% in the N plot and 99% in the CCM ? N plot. Our data suggest CCM supplementation to N fertilizers as a potential replacement of inorganic fertilizers like P and K for effective environmental management.

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