Phialina pupula ( Müller, 1773 ) Foissner, 1983

Rajter, Ľubomír, Bourland, William & Vďačný, Peter, 2019, Morpho-molecular Characterization of the Litostomatean Predatory Ciliate Phialina pupula (Müller, 1773) Foissner, 1983 (Haptoria, Lacrymariidae), Acta Protozoologica 58 (2), pp. 53-68 : 55-57

Phialina pupula ( Müller, 1773) Foissner, 1983

Improved diagnosis (based on Boise population): In vivo size about 60–130 × 20–50 µm. Body shape highly variable depending on state of contraction, ranging from clavate in extended condition through fusiform, pyriform, elliptical to almost globular in semi-contracted and contracted state. Macronucleus elliptical with a single micronucleus. Highly refractive dumbbell-shaped inclusions scattered throughout cytoplasm and usually concentrated in anterior body part. Contractile vacuole subterminal in extended condition, terminal in contracted state. Extrusomes about 10 µm long, rod-shaped, attached to oral bulge and forming bundles in cytoplasm. On average 15 ciliary rows, each row anteriorly differentiated into a dorsal brush composed of one to four dikinetids.

Type locality: Müller (1773) did not specify the type locality. He mentioned only that he found the species in water and ice from dunghills during November and December.

Type material and voucher slides: No type material is available from Müller’s (1773) specimens. Three voucher slides containing protargol-impregnated specimens from the Boise population have been deposited at Department of Zoology, Comenius University in Bratislava.

Material studied: Specimens from lower microaerobic layers of the interstitial sandy sediments from the floodplain area of the Boise River near the Glenwood Bridge, Boise, Idaho, U.S.A.

Etymology: Not given in the original description. The feminine Latin noun pupula is a diminutive form of pupa (doll, puppet or pupa of an insect), obviously referring to the doll- or pupa-like body shape of the ciliate. The name is treated as a noun in the nominative singular standing in apposition to the generic name [Art. 11.9.1.2 of the International Commission on Zoological Nomenclature (1999)].

Description of Boise population: Size in vivo 60– 130 × 20–50 µm, usually about 85 × 30 µm, as calculated from some in vivo measurements and morphometric data adding 15% preparation shrinkage; length:width ratio on average 2.2: 1 in vivo and 2.9:1 (n = 32) in protargol preparations ( Table 1 ). Body shape highly variable depending on state of contraction, ranging from clavate in extended condition through fusiform, pyriform, elliptical to almost globular in semi-contracted and contracted state. Head barrel-shaped, about 8.5 × 6.0 µm in size after protargol impregnation, distinct from trunk but without neck-like region, sometimes retracted into trunk creating an impression of shoulders. Posterior body end tapered and tail-like in extended condition, narrowly to broadly rounded in semi-contracted and contracted state ( Figs 2A, E, F , 3A, C, E, F, H–M ). Contraction occurs slowly.

Nuclear apparatus located in or slightly posterior to mid-body, usually slightly lateral of cell center. Macronucleus elliptical, on average 15 × 10 µm (n = 32) in size after protargol impregnation. Micronucleus adjacent to macronucleus, usually attached to anterior pole of macronucleus, elliptical and about 2 µm long in vivo ( Table 1 ; Figs 2A , 3A, D, E, F ). Contractile vacuole subterminal in extended specimens while terminal in semi-contracted and contracted cells, excretory pore(s) not recognizable in vivo or after protargol impregnation ( Figs 2A, F , 3A, F ). Only one type of extrusomes, rod-shaped, about 10 × 0.5 µm in size in vivo, attached to oral bulge and in bundles scattered throughout cytoplasm, impregnate well with the protargol method used ( Figs 2A, C , 3C, F, G ). Cortex very flexible, distinctly furrowed by ciliary rows, sometimes dotted by tips of cortical granules in SEM ( Fig. 4A–C ). Cortical granules colorless, broadly elliptical to elliptical and about 0.8 × 0.4 µm in size in vivo, oriented perpendicularly to cell surface, rather irregularly and narrowly spaced forming seven or eight rows between adjacent ciliary rows, impregnate deeply with the protargol method used often making observations of the ciliary pattern difficult ( Figs 2D , 3A, F ). Cytoplasm colorless, packed with few to many lipid droplets, some extrusome bundles, and many highly refractive inclusions. Individual inclusions dumbbell-shaped, about 2 µm long and usually numerous in anterior body half, rendering the cell dark in appearance at low magnifications ( Figs 2A, B , 3A–C, E–M ). Swims fast along helical trajectory by rotation about main body axis.

Somatic cilia about 8 µm long in vivo, arranged in an average of 15 rows, each row composed of about 22 monokinetids with some dikinetids (dividing basal bodies) irregularly interspersed. Somatic kineties ordinarily spaced, extend meridionally to slightly helically depending on state of contraction ( Table 1 ; Figs 2A , 4A, C ). Dorsal brush at anterior end of all somatic kineties, very inconspicuous not only in vivo but also in protargol preparations and in SEM because composed of only two to five dikinetids (SEM measurements): first brush dikinetid bears a short, 1.5–2.0 µm-long, rod-like cilium followed by an ordinary cilium about 6.5 µm long; second dikinetid associated with a minute, 0.3 µm-long, stump-like cilium followed by an ordinary cilium; all following brush dikinetids with anterior basal body unciliated and posterior basal body bearing an ordinary cilium ( Table 1 ; Figs 2E , 4A, B ).

Oral apparatus occupies apical end of head. Oral bulge contains tip of extrusomes, posteriorly delimited by circumoral kinety as usual in congeners. Circumoral kinety and its structure very difficult to recognize in protargol preparations, very likely composed of dikinetids. Head kineties helical and narrowly spaced, extend between circumoral kinety and dorsal brush, composed of densely arranged monokinetids bearing about 10 µm long cilia in vivo and almost completely covering head in SEM ( Figs 2A, E , 3A , 4A ).

Figures

Fig. 2. A–F. Phialina pupula from life (A‒D, F) and after protargol impregnation (E). (A) Overview of a representative semi-contracted specimen. (B) Details of dumbbell-shaped inclusions from various views. (C) Extrusomes are rod-shaped and about 10 µm long. (D) Surface view showing cortical granulation. (E) Ciliary pattern. (F) Variability of body shape in extended, semi-contracted and contracted cells. CK – circumoral kinety; CV – contractile vacuole; DB – dorsal brush; DI – dumbbell-shaped inclusions; EB – extrusome bundle; EX – ex- trusomes; G – cortical granules; OB – oral bulge; MA – macronucleus; MI – micronucleus; SK – somatic kineties. Scale bars: 20 μm.

Fig. 3. Phialina pupula from life under differential interference contrast (A–G) and bright field (H–M) illumination. (A) Overview of a semi-contracted specimen, showing the general body organization. The head is attached directly to the broadly fusiform trunk. Note that the contractile vacuole is located terminally due to the body contraction. The macronucleus is elliptical and situated slightly below the midbody. (B) Detail of the highly refractive dumbbell-shaped inclusions scattered throughout the cytoplasm. (C) A semi-contracted specimen, showing an accumulation of the dumbbell-shaped inclusions in the anterior body half. (D) Detail of the nuclear apparatus. The macronucle- us is elliptical, and the micronucleus is attached to the anterior pole of the macronucleus. (E) A contracted specimen, showing many refrac- tive, dumbbell-shaped inclusions scattered throughout the cytoplasm and an elliptical macronucleus accompanied by a single micronucleus. (F) A strongly squeezed specimen, showing the nuclear apparatus, multiple extrusome bundles and some lipid droplets scattered throughout the cytoplasm. Left inset shows optical section through the cortex (opposed arrowhead), containing inconspicuous elliptical granules. (G) Detail of a cytoplasmic rod-shaped extrusome. (H, J) Fusiform, slightly curved cells with narrowly rounded posterior body end. (I) A cylindrical cell. (K) An extended, fusiform exemplar with tail-like posterior end. (L) A sigmoid cell with narrowly rounded ends. (M) A semi-contracted, pyriform specimen with broadly rounded posterior body end. CV – contractile vacuole; DI – dumbbell-shaped inclusions; EB – extrusome bundles; EX – extrusomes; G – cortical granules; H – head; LD – lipid droplets; MA – macronucleus; MI – micronucleus; OB – oral bulge; T – trunk. Scale bars: 20 μm.

Fig. 4. Phialina pupula in the scanning electron microscope (SEM). (A) Detail of the anterior body half. The head is localized at the anterior body end and is attached directly to the trunk, as typical of the genus Phialina. The head is covered by very narrowly spaced cilia arranged in helically extending rows. Note that the cortex of the trunk is distinctly furrowed by slightly helically extending ciliary rows. According to protargol preparations, each somatic ciliary row has two to five brush dikinetids at its anterior end (see Fig. 2E). SEM observations show that the anterior basal body of a brush dikinetid bears a minute to short cilium or is unciliated, while the posterior basal body bears an or- dinary somatic cilium. Therefore, the brush is very difficult to recognize in the SEM and in vivo. (B) Detail of the anterior end of somatic ciliary rows, showing that the anterior basal body of a brush dikinetid bears a short cilium (arrowheads) or is unciliated. The posterior basal body of a brush dikinetid bears an ordinary somatic cilium. Such an inconspicuous brush is a typical feature of lacrymariids and also of the possibly related chaeneids. (C) Detail of a somatic ciliary row, showing a dikinetid (dividing basal bodies) followed by monokinetids that bear ordinary cilia. As typical for haptorians, the anterior cilium of dividing basal bodies is short and stump-like while the posterior cilium is ordinarily long. AC – anterior stump-like cilium of dividing basal bodies; G – tips of cortical granules; H – head; HC – head cilia; SC – somatic cilia; T – trunk.