The Fine Structure of the Colonial Colorless Flagellates Rhipidodendron splendidum Stein and Spongomonas uvella Stein with Special Reference to the Flagellar Apparatus

SYNOPSIS. The cell structure of the colorless colonial flagellates Rhipidodendron splendidum Stein and Spongomonas uvella Stein has been examined by electron microscopy to assertain their phylogenetic affinities. The cylindrical cells of R. splendidum have 2 smooth flagella of equal length, an asymmetrical flagellar pocket supported by microtubules, and a curved pit between the latter and an anterior prolongation of the cell. The matrix of the branched tubes comprising the fanshaped colony is composed largely of dense spherules which are produced in special cytoplasmic vesicles some of which contain symbiotic bacteria. The anterior nucleus has a flattened sac pressed closely against its posterior end. The sac has a long tail extending deep into the cytoplasm, a single bounding membrane and homogeneous contents. Several types of vesicle are described but food vacuoles and contractile vacuoles could not be positively identified. A kinetoplast mitochondrion is not present. The various cross-banded, microtubular and amorphous components of the complex and highly asymmetrical flagellar root system are described in detail and a 3-dimensional reconstruction is provided.
The ovoid cells of S. uvella are basically similar to those of R. splendidum ; though a nuclear sac is missing, there are some detailed differences in the structure of the flagellar root system, and bacteria are never present in the vesicles producing the matrix granules. Notwithstanding much similarity, Rhipidodendron is not combined with Spongomonas because of the basic difference in colony structure.
The possible relationships of R. splendidum and S. uvella with other groups are examined and it is concluded that they cannot be considered as colorless chrysomonads as previously thought or considered to be related to any of the other orders comprising the class Phytomastigophorea. They do not, however, appear to be related to any of the orders at present comprising the Zoomastigophorea.     

The Fine Structure of the Centriolar Apparatus and Associated Structures in the Flagellates Deltotrichonympha and Koruga. I. Interphase

SYNOPSIS. An electronmicroscopic study was made of the centriolar apparatus in the rostrum of Deltotrichonympha operculata and Koruga bonita , 2 closely related hypermastigote flagellates from the Australian termite, Mastotermes darwiniensis. In interphase flagellates, the centriolar apparatus consists of 2 similar parts with a mutually perpendicular orientation. Each part contains a large, club-shaped centriolar body consisting of fibrillar and granular material, without recognizable internal symmetry or microtubules. The anterior centriolar body extends from the inner rostral wall, which is structurally related to the fibrous wall surrounding the posterior centriolar body. The 2 centriolar bodies are joined by connecting branches, which meet at 3 barren kinetosome-like structures located inside the rostrum. Thus, an interphase flagellate has 2 centriolar bodies oriented at a 90° angle to each other, like a pair of typical centrioles in an interphase metazoan cell.     

Isolation of the Kinetoplast DNA of Leishmania tarentolae in the Form of a Network*

The major kinetoplast DNA complement of Leishmania tarentolae promastigotes has been isolated as a single sheet of interconnected molecules on the basis of its relative stability to shear forces and its high sedimentation coefficient. Two successive differential centrifugations were sufficient to recover 50 ± 10% of the total kinetoplast DNA free of nuclear DNA contamination. We use the term “network” to describe this unusual type of DNA configuration. Leishmania networks have a molecular weight of ～10¹⁰ daltons and an S_20,W in neutral sucrose gradients of 1729 7plusmn; 189 [n = 19] and exhibit an extremely low specific viscosity due to the compactness of packing of the DNA. The networks were visualized in the electron microscope, and in the light microscope either by fluorescence in solution after staining with acridine orange or in dried smears after staining with Giemsa. Purified networks from stationary phase cells banded in the position characteristic of closed monomeric minicircles in ethidum bromide-CsCl equilibrium gradients, and were stable in alkaline sucrose. Treatment of the closed networks with RNase and pronase had no effect on the ethidium bromide-CsCl banding pattern. However, treatment of closed networks with DNase I or II, X-irradiation or γ-irradiation changed the banding pattern by introducing single strand and double strand breaks, yielding an upper band and in some cases a intermediate band.     

Fine Structure of the Macronuclear Anlage of the Hypotrichous Ciliate Keronopsis rubra

In exconjugants of the hypotrich Keronopsis a large, highly polyploid macronuclear anlage is formed from which condensed chromatin bodies are passed into the cytoplasm where they are thought to give rise to the numerous small macronuclei of the vegetative cell. Electron microscopy shows that the chromatin bodies within the macronuclear anlage are separated from each other by sheets of low contrast lamellar material. The anlage appears therefore as a composite nucleus containing prepacked units which are extruded into the cytoplasm following condensation.     

Fine Structure of the Cell Surface and Golgi Apparatus of Ochromonas*

SYNOPSIS. Ochromonas danica has an unusually flexible cell surface capable of producing projections of varying sizes and shapes: large projections, 340–360 nm long, and small projections, 50–110 nm long. These projections have been demonstrated by transmission and scanning electron microscopy; some of them may break off into the medium and be the source of extracellular membranes and vesicles reported in the cell-free O. danica growth medium. Ruthenium red stained the acid mucopolysaccharide layer just outside the cell surface as well as small blebs at the cell surface. The Golgi complex of O. danica, Ochromonas malhamensis, Ochromonas sociabilis and Ochromonas sp. produced small coated vesicles which may move toward and fuse with the plasma membrane. The role of the several vesicles is unknown but possible functions are discussed.     

The Structure and Origin of Mastigonemes in Ochromonas minute and Monas sp.*

Structure, function, and development of mastigonemes (flagellar hairs) of 2 chrysophycean flagellates were examined with light and electron microscopy in whole mount and sectioned preparations. Mastigonemes of both organisms are identical, consisting of a tapered base 0.25–0.3 μm long, maximum width of 0.03 μm; a hollow shaft 0.85 μm × 23 nm; and 2 types of laterally projecting filaments. Two rows of mastigonemes are attached to the long flagellum, one on each side in the same plane as the central pair of microtubules. One row is composed of single mastigonemes while the other bears them in “tufts.” The primary mastigonemal attachment is on the flagellar membrane. Developmental sequences as supported by electron micrographs and kinetic studies demonstrate the intracellular location of promastigonemes during reflagellation, colchicine-inhibited reflagellation, and release from inhibition. The promastigonemes first appear in the peri-nuclear space in association with the outer nuclear membrane and several dozen may accumulate there. These may pinch off as bundles and move into the cytoplasm, or if mastigonemes are being utilized rapidly by the cell, the promastigonemes are channeled a few at a time from the perinuclear space into the Golgi apparatus where some structural modifications are made. The mastigonemes are then transported in Golgi-derived secretory-type vesicles to the cell surface near the base of the growing flagellum where the vesicle membrane fuses with the plasma membrane and the mastigonemes become extracellular, although the membrane association is retained. The origin of the asymmetric arrangement of mastigonemes on the flagellum is discussed.     

The Structure and Morphogenesis of the Ventral Ciliature in Paraurostyla hymenophora*

SYNOPSIS. The structure and morphogenesis of the ventral ciliature of Paraurostyla hymenophora (Stokes) are described. The oral primordium apparently originates in association with transverse cirrus #6, from which it migrates anteriorly simultaneous with kinetosomal proliferation. The primordium eventually forms an elongate ciliary field from which the future opisthe's fronto-ventro-transverse (FVT) and undulating membrane primordial fields arise. Concomitantly, the future proter's FVT primordial field is initiated by the disaggregation of frontal cirri #4, #5, and #6. Primordia then develop simultaneously within marginal and ventral cirral rows by a disaggregation of cirri within the respective rows, and do not give rise to new cirri until the FVT fields complete segregation into discrete cirri. Near the completion of cirral production from the FVT primordia, each ventral cirral primordium (VCP) forms the 2 rightmost transverse cirri. Segregation of new cirri within the marginal cirral primordia and VCP then occurs, eventually replacing all old cirri within their respective marginal and ventral cirral rows. At the end of cortical morphogenesis, all old ciliary organelles, with the exception of the adoral zone of membranelles, are either reorganized or replaced. These results suggest an evolutionary affinity between the ventral and marginal cirral rows and raise questions about the control of the developmental competence of individual primordia.     

Fine Structure of the Oocyst Wall of Eimeria nieschulzi

SYNOPSIS. Oocysts of Eimeria nieschulzi from the laboratory rat, Rattus, norvegicus , were studied by scanning and transmission electron microscopy. Oocysts had a rough outer wall with apparent random depressions. The oocyst wall is composed of 2 layers: an osmiophilic outer layer consisting of a rough external and smooth internal surface, and a relatively thick, electron-lucent inner layer. The outer layer is composed of a dense, coarsely granular matrix. The inner layer consists of homogeneous fine granular material interspersed with coarse osmiophilic granules and contains one closely applied membrane on the outermost surface. Several raised lenticular areas are seen on the coarse outer surface of the inner layer. These layers are 102 (75–128) and 176 (135–204) nm thick, respectively.
The sporocyst wall is thin, consisting of 3 to 4 unit membranes, and measures 27 (18–34) nm thick.     

Fine Structure of Haemoproteus columbae Kruse During Macrogametogenesis and Fertilization*

When blood is withdrawn from a pigeon (Columba livia) infected with gametocytes of Haemoproteus columbae, differentiation of the gametes begins immediately. This study examines the formation of the macrogamete and its fertilization. The first visible signs of differentiation are the elongation of the nucleus along with the appearance of an intranuclear spindle and atypical centrioles. Then maturation bodies, the products of nuclear reduction, form in both erythrocytic macrogametocytes and macrogametocytes free of their host cells. Penetration of the macrogamete by the microgamete occurs rapidly. Their plasma membranes fuse, and the microgamete's nucleus, axonemes and cytoplasm enter the macrogamete. The nucleus of the microgamete expands and migrates to lie at an angle to the macrogamete nucleus. The 2 fuse across a small area. The nuclear envelope and the plasma membrane of the zygote are a mosaic of the membranes of the 2 gametes.     

The Fine Structure of Four “Species” of Amoeba*

The fine structure of Amoeba discoides, Amoeba dubia, and Amoeba amazonas was studied and compared with that of Amoeba proteus. The different kinds of amebas showed general similarities but differed in the ultrastructural details of their organelles. With respect to fine structure, A. discoides was indistinguishable from A. proteus, while both A. dubia and A. amazonas had distinctive features. The nuclei of all had a prominent honeycomb-like fibrous lamina, but A. dubia differed from the others in the distribution of nucleoli within the nucleus. The mitochondria of A. amazonas were unusual in having a variable pattern of cristae, some being plate-like and others tubular. Golgi bodies in A. amazonas had a greater proportion of vesicles and a smaller number of cisternae than those of the others, while Golgi bodies in A. dubia had highly flattened cisternae without a lining of filamentous material such as is found in the other types. The plasma membrane of A. dubia also lacked the prominent filamentous cell coat common to A. proteus and other amebas. The relation between the Golgi apparatus and the cell coat and the significance of the degree of development of the cell coat for pinocytosis and other phenomena is considered. The experimental use of these cells, including the formation of hybrids by nuclear transplantation is discussed.     

Fine Structure of Penetration of Cultured Cells by Isospora canis Sporozoites

SYNOPSIS Monolayers of Embryonic Bovine Trachea (EBTr) cells were inoculated with Isospora canis Nemeséri spcrozoites. As penetration commenced, they were fixed, stained with OsO₄-ruthenium red, dehydrated, embedded and sectioned in situ. Examination by electron microscopy revealed that host cell membranes remained intact during penetration. The sporozoites caused an invagination of the cell's plasmalemma until the parasites were entirely within the cell, after which the invagination was sealed by short pseudopodia enclosing the parasite within a membrane-lined vacuole inside the cells. Rhoptries and micronemes, which appeared as branched elements of the same network, became less tortuous near the conoid and often became empty or partially empty during penetration. Concurrent with the appearance of these partially empty rhoptries, vesiculations were seen in the host cell cytoplasm opposite the apical tip of the sporczoite. Constrictions of the sporozoite during entry were probably due to bands of microfilaments beneath the plasmalemma and elsewhere in the cytoplasm of the host cell.     

Fine Needle Tissue Aspiration for Accurate Localization of Histological Specimens from Complex Structures

A procedure is presented for exact, detailed comparison of light and electron microscopic analyses of tissues with complex architecture. Earlier techniques require one to make drawings of tissue pieces to be analyzed by electron microscopy to permit rough localization of the origin of the tissue pieces. Specifically, exact analysis of fetal cartilage and bone is hampered by the complicated arrangement of both tissue components, severely limiting the assessment of electron microscopic analyses. The advantage of the technique described here is that it allows precise localization of the tissue sample in the original tissue area. Punches 1 mm in diameter were obtained from femora and coxae with a syringe and embedded for light and electron microscopy. The remaining tissue with its exactly defined punctures is prepared for standard histology. Human fetal cartilage and bone tissue were used to demonstrate this technique, but this procedure may be used for other kinds of tissues.     

Fine Structure of the Malaria Parasite Plasmodium lophurae Developing Extracellularly in vitro*

Duck malaria parasites (Plasmodium lophurae), synchronized at the uninucleate trophozoite stage, were freed from their host erythrocytes by immune lysis and cultured extracellularly in duck erythrocyte extract medium. At 0 time, 1, 2, and 3 days, samples were taken for light and electron microscopy and for measurement of incorporation of [¹⁴C]-methionine or [¹⁴C]-proline. For 2 days the parasites developed fairly normally, progressing from large trophozoites-early schizonts at 1 day to segmenters-forming merozoites at 2 days. However, the 3-day samples showed signs of deterioration: incorporation of amino acids dropped; the percentage degenerate cells rose; the progression of developmental stages slowed. At the fine structure level 2 abnormalities were observed which may indicate the limits of extracellular cultivation in vitro. Through 2 days of culture all parasites were surrounded by 2 membranes. The 3-day samples contained some organisms with only one membrane, which may have arisen from merozoites produced extracellularly. The 2nd alteration was in the food vacuoles, which were progressively fewer, smaller, and less dense in the cultured samples and may indicate an abnormality in the extracellular parasite's feeding mechanism.     

A Study of the Structure and Gliding Movement of Gregarina garnhami

SYNOPSIS The structure and gliding movement of Gregarina garnhami Canning, a eugregarine found in the midgut of the desert locust, Schistocerca gregaria , have been studied by light microscopy and transmission and scanning electron microscopy (EM). Ultrastructural studies revealed that the cytoplasm of G. garnhami is separated from the epicyte folds by a basal lamina. The pellicle consists of 3 membrane layers. At the tips of the epicyte folds there are 2 sets of longitudinally oriented filaments. An ectoplasmic network is present in the ectoplasm and the endoplasm contains numerous paraglycogen granules. The effect of cytochalasin B on G. garnhami was studied. Examination of scanning EM preparations of gliding and stationary gregarines yielded inconclusive results. In some instances the epicyte folds were thrown into waves; in others the folds were straight, regardless of treatment before fixation. Gregarina garnhami glides through its environment without any apparent deformation in shape. As it moves, a mucus trail is left behind it. Phase-contrast observations were made of centrifuged gregarines in which the endoplasm was displaced. Centrifuged gregarines continued to glide. Displacement of the endoplasm allows visualization of the epicyte folds in gliding animals. No lateral waves were seen in the epicyte folds of gliding centrifuged animals.     

Fine Structure of the Schizont Stage of the Testate Marine Ameba,Trichosphaerium sp.*

SYNOPSIS. The schizont stage of 3 strains of the testate marine ameba, Trichosphaerium sp., was examined in the transmission electron microscope. The cytoplasm of this multinucleate organism contained the usual organelles; the test was covered by spicules. Pseudopodial types included broad ectoplasmic lobopods that assisted in locomotion, and thin dactylopods that probably had a sensory function. Nuclear division, observed in one of the strains, was characterized by an intact nuclear envelope (at least through anaphase), and the absence of centrioles. Nuclei in an ameba divided synchronously. An unusual intranuclear body of unknown function was found in another of the strains examined.     

The Fine Structure of the Phoront of the Apostomatous Ciliate,Hyalophysa chattoni*

The phoront of the apostomatous ciliate, Hyalophysa chattoni, is an encysted stage that is carried on the exoskeleton of its crustacean host until the ecdysis of the host. At molting the phoront rapidly metamorphoses to the feeding stage, excysts, and immediately begins to feed on exuvial fluid trapped in the cast-off exoskeleton. The fine structure of the resting phoront resembles that of the preceding migratory stage, the tomite. A prominent ventral tuft of cilia, the ogival field, has vanished, and the trichocysts that paralleled the kinetics have disappeared. The dense inclusion bodies that were concentrated around the mouth and falciform fields have dispersed and greatly decreased in number. The cytoplasm and its membranous organelles do not appear visibly condensed or altered from the preceding stage in the life cycle. The phoront is merely quiescent instead of dormant. Unlike the few ciliate cysts previously examined by electron microscopy, the phoront's cyst is not divisible into separable layers. It resembles the loricae of certain suctoria in being formed principally of a fibrous substance, the outer surface of which has a paracrystalline pattern. The peduncle attaching the cyst to the crab's gill is a continuation of the cyst wall although its structure is somewhat modified. The most conspicuous innovation in the phoront's fine structure is the massive tracts of microtubules that run longitudinally through the macronucleus. The microtubules are in intimate contact with Feulgen-positive chromatin masses which are crowded toward the periphery of the macronucleus.     

Discotricha papillifera: Structure and Morphogenesis of a Marine Interstitial Ciliate*

SYNOPSIS. Discotricha papillifera Tuffrau, a marine interstitial ciliate, is redescribed with the aid of light, scanning, and transmission electron microscopy from cells collected at a New Hampshire beach. Presence of primitive membranelles as well an an advanced stomatogenesis is demonstrated. The ultrastructure, including a unique membrane-bounded septate structure, is described. The cell is tentatively placed in the nassulid suborder Microthoracina, but affinities with other groups are discussed.     

Light microscopy and scanning electron microscopy studies on the reduction of the tongue microstructures in the white stork (Ciconia ciconia,Aves)

The structure of the tongue in the white stork (Ciconia ciconia) is observed macroscopically and under light and scanning electron microscopy. Our observations of the tongue reveal a rare terminal reduction of the size of the tongue and microstructures of the lingual mucosa among the investigations of birds published so far. The short, triangular tongue with a pointed tip is approximately 2.5 cm long in the adult and is situated in the caudal part of the oral cavity close to the laryngeal prominence. On the dorsal surface of the tongue, no typical mucosa microstructures like lingual papillae, median groove or lingual prominence are observed. The main structure of the tongue is composed of rostral part of hyoid apparatus, that is, entoglossal cartilage connects with basihyoid. Very thin mucosa is composed of fibrous connective tissue covered with orthokeratinized epithelium. No lingual glands and muscles are observed in the lamina propria of mucosa. Even though the triangular shape of the tongue in the white stork is typical for birds, the inner structure of the reduced organ is composed only of flat cartilagineous entoglossum of hyoid apparatus. During feeding behaviour of the white stork, the food transportation in oral cavity called cranio‐inertial transport is undoubtedly affected by structural reduction of the tongue.     

The Fine Structure of the Cytostome of the Apostomatous Ciliate Hyalophysa chattoni*

SYNOPSIS. The fine structure of the organelles concerned with the ingestion of exuvial fluid by the trophont of the apostome ciliate, Hyalophysa chattoni, has been examined. One of the taxonomic characteristics of the order Apostomatida is that cytostomes of ciliates within the taxon are reduced and evolving toward astomy. When examined by electron microscopy the cytostome of H. chattoni appears as a small region of active pinocytosis which is continuous with a very large cortical area, the extended cytostome. The fine structure of the extended cytostome resembles that of the cytostomes of ciliates from other orders in that it is covered by a single membrane underlain with microtubular ribs. Beneath the extended cytostome are accumulations of peculiar organelles that may represent stored membrane for recycling during food vacuole formation. Associated with the site of pinocytosis is a complex fiber that may be contractile.