群体ボヤ Botrylloides simodensis の内陸飼育法の確立

抄録

Colonial growth of the ascidian, Botrylloides simodensis, has been investigated in laboratory conditions. Two strains, S81 and S85, were used. Animals attached on glass slides were held vertically in the culture tank that contained five liters of natural sea water (20°C, pH 8.2-8.4). The sea water was filtered every one or two days and recycled for about a month. The aeration was not performed. The diatom, Nitzschia closterium, was added to the culture tank twice a day at the final concentration of 0.2-4×10⁴cells/ml. The S81 strain showed a zooid proliferation value of 2.3 on an average. The life span of individual zooids was 16-17 days. The gonads became mature periodically and oozooids (F1) could be obtained by means of autogamy. On the other hand, the S85 strain did not grow well when it was fed on only Nitzschia. The artificial diet, Liquifry marine (40-80μl/tank/day), was very effective for keeping high proliferation ratio (maximal value; 2.9). However, the sea water needed to be exchanged with new one every two or three weeks. The permissive number of colonies in a single tank was examined in the last series of this work. When two or three colonies coexisted, they proliferated well (maximal value; 2.4). But, in case of four to five colonies, the value did not exceed 2.0. The results showed that when natural sea water is available periodically the colonies of Botrylloides simodensis can be maintained easily in any laboratory.
MATERIALS and METHODS
Animals Botrylloides simodensis has been recently separated from B. violaceus taxonomically (SAITOH et al., 1981). The original stocks of B. simodensis were collected in Uranouchi Inlet, Usa, Kochi Prefecture, in 1981 (strain No. S81) and in 1985 (strain No. S85). Ever since their asexual progenies have been reared on glass slides in culture boxes floating in the field as described previously (SUGIMOTO and NAKAUCHI, 1974).
The S81 was brought to our laboratory and reared from March to August in 1983 and from October, 1984 to February, 1985. It was very tough and proliferated well in laboratory conditions. Unfortunately, the S81 was extinguished in summer of 1985 because of red tide. The S85 was kept in the laboratory from November of 1985 onward.
Only a single colony was attached to a single glass plate which was held vertically in a culture tank. Respective series of experiments were always performed using clonal colonies derived from a large parental colony. The parental colony was separated into many small pieces and they were put on new glass slides in a moist chamber for about one hour in order for them to adhere firmly to the substratum.
Sea water About 10 liters of natural sea water were prepared for a single culture tank. It was halved and each (five liters) was used alternately every one or two days. It was renewed immediately after use by passing through cotton filter. The sea water that contained animals was kept at 20°C and adjusted to pH 8.2-8.4 with 0.2M Tris, if necessary. The aeration was not performed. The sea water could be recycled for about a month. But, it needed to be exchanged with a new one every two or three weeks when the artificial diet was administered.
Foods Animals were fed on the diatom, Nitzschia closterium. The diatom was permitted to proliferate in the artificial sea water, Jamarin (Jamarin Lab. Japan), containing 9μg/ml KH₂PO₄, 72μg/ml KNO₃, and 0.5μg/ml Fe·EDTA. Under continuous light condition, it became saturated in about a week. Twenty mililiters of this diatom-containing Jamarin were added to the tank (the final conc. 0.2-4×10⁴cells/ml) twice a day. The cell number of the diatom was consistent roughly with that applied previously to Symplegma reptans (NAKAUCHI et al., 1979).