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Antifreeze Water-Rich Dormant Cysts of the Terrestrial Ciliate Colpoda cucullus Nag-1 at −65 ℃: Possible Involvement of Ultra-Antifreeze Polysaccharides

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Matsuoka T., Sogame Y., Nakamura R., Hasegawa Y., Arikawa M., Suizu F.

Acta Protozoologica

2020

vol. 59

issue 3-4

141-147

We found that the water-rich (osmolality below 0.052 Osm/l) wet resting cysts of the soil ciliate Colpoda cucullus Nag-1 were tolerant to extremely low temperature (−65℃). When cell fluid obtained from the resting cysts was cooled at −65℃, small particles of ice crystals did not grow into large ice crystals. At −65℃, the cysts shrank due to an outflow of water, because a vapor pressure difference was produced between the cell interior and freezing surrounding medium. The osmolality of these shrunk cells was estimated 0.55 Osm/l, and the freezing point depression of the shrunk cell fluid was estimated to be 1.02℃. Hence, the antifreeze ability of wet cysts at −65℃can not be explained by freezing point depression due to elevation of cytoplasmic osmolality. The cytoplasm of resting cysts was vividly stained red with periodic acid-Schiff (PAS) and stained purple with toluidine blue. On the other hand, the excystment-induced cysts were not stained with PAS, and exhibited a loss of the antifreeze activity. PAS staining of SDSPAGE gel obtained from encysting Colpoda cells showed that a large amount of PAS-positive macromolecules accumulated as the encystment stage progressed. These results suggest that antifreeze polysaccharides may be involved in the antifreeze activity of C. cucullus Nag-1 dormant forms.

Tolerance of Colpoda cucullus Nag-1 Resting Cysts and Presumed Structure for Protection against UV Light

84%

Yamane S., Watanabe M., Funadani R., Miyazaki R., Hasegawa Y., Arikawa M., Suizu F., Matsuoka K., Matsuoka T.

Acta Protozoologica

2020

vol. 59

issue 1

55-60

Resting cysts of the terrestrial ciliate Colpoda cucullus (Nag-1 strain) are highly resistant to UV light. It has been speculated that auto-fluorescent (blue fluorescent) particles surrounding the nuclei and yellowish fluorescent layers of the cyst wall are the candidate structures for the protection of the cellular components from UV light. The UV resistance of encysting cells was quickly acquired up to 5 h after the onset of encystment induction, and then gradually increased for several days. The less fluorescent ectocyst layer, yellowish fluorescent first-synthesized endocyst layer (en-1) and the NSPs were formed within 5 h after the onset of encystment induction, and thereafter endocyst layers became gradually thicker for several days. The cyst wall sample (ectocyst and endocyst layers) markedly absorbed a broad range of UV light. This result indicates that the cyst wall evidently has UV-cut function. These results support that the cyst wall and NSPs of C. cucullus play a role in the shielding of the cell components from UV light.

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2 Acta Protozoologica

2 2020

2 Arikawa M.

2 Hasegawa Y.

2 Matsuoka T.

2 Suizu F.

1 Funadani R.

1 Matsuoka K.

1 Miyazaki R.

1 Nakamura R.

1 Sogame Y.

1 Watanabe M.

1 Yamane S.

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Antifreeze Water-Rich Dormant Cysts of the Terrestrial Ciliate Colpoda cucullus Nag-1 at −65 ℃: Possible Involvement of Ultra-Antifreeze Polysaccharides

Tolerance of Colpoda cucullus Nag-1 Resting Cysts and Presumed Structure for Protection against UV Light