Assays of protein contained in water-soluble fraction of encysting cells Colpoda cucullus Nag-1 by two-dimensional electrophoresis (2-D PAGE) and mass spectrometry (MS) revealed that the amount of β-tubulin abruptly increased in 2.5–10 h after encystment induction. Judging from the results that total α-tubulin content did not decrease much until 12 h after encystment induction, the result indicates that disassembly of microtubules may occur soon after encystment is induced. Therefore, we tried to visualize dynamics of microtubules. Immunofluorescence microscopy using anti-α-tubulin antibody indicated that disassembly of axonemal microtubules of cilia became within 1.5 h after encystment induction, and resorbed in 3 days. Although the cytoplasmic microtubules failed to be visualized clearly, encystmentdependent globulation of cells was promoted by taxol, an inhibitor of disassembly of microtubules. It is possible that a temporary formation of cytoplasmic microtubules may be involved in cell globulation.

The phosphorylation level of actin (43 kDa) became slightly elevated just after encystment induction. Lepidosomes, the sticky small globes surrounding encysting cells, were vividly stained with Acti-stain 555 phalloidin, suggesting that 43-kDa actin or its homologues may be contained in lepidosomes.

The genus Anthosphaera Kamptner emend. Kleijne is one of the most taxonomically confusing modern coccolithophores and its species level taxonomy has long been in a state of flux. Based on the review of imaged specimens from our collections, we attempt to rectify the nomenclatural problems and elucidate the obfuscated taxonomy of the genus. Review of included formally and informally described species shows that they are a distinctive group with shared characters, including ten different morphotypes of probable species level. Two of these, including the type species A. fragaria, have been shown to form life-cycle associations with heterococcoliths of the Syracosphaera molischii type. Hence, all species are transferred to Syracosphaera and the new combinations S. periperforata, S. lafourcadii, and S. origami are proposed. In addition, various informally described morphotypes are now formally described as Syracosphaera molischii var. pertusa, S. periperforata var. cylindrata, S. periperforata var. tridentata, S. rotaconica, and S. elevata.

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.

This paper is a first report on species of endosymbiotic ciliates (Litostomatea, Trichostomatia) inhabiting the intestine of zebras in South Africa. Ciliates from Mountain Zebra were investigated for the first time in the world. The wild population of mountain zebras in general and the Cape Mountain Zebra subspecies in particular is low in numbers: this species is included as vulnerable in the IUCN Red List. Approximately 15 species of trichostome ciliates from 9 different genera were found in the samples collected from wild zebras in Western Cape, South Africa. Some of the ciliate species are also common to horses and other equids, while others are unique for zebras. The ciliates of Triplumaria genus common to elephants and rhinoceroses, and the species Blepharosphaera ceratotherii previously described in rhinoceroses were found in equids for the first time.