Tracer Experiments in Feeding Littoral Foraminifera*†‡

SYNOPSIS. Tracer technic has proved to be an excellent tool in the study of predator-prey relationships among the foraminifera. More than fifty axenic species of protists including diatoms, dinoflagellates, chlorophytes, chrysophytes, cyanophytes, bacteria and yeasts were tested as potential food for Allogromia sp (NF), A. laticollaris, Am. monia beccarii, Quinqueloculina spp, Rosalina floridana, Anomalina sp, Elphidium sp, Spiroloculina hyalina, Globigerina bulloides, and Globorotalia truncatulinoides. Although many types of potential food are present in the environment, foraminifera select only certain organisms. The yeasts, cyanophytes, dinoflagellates, chrysophytes and most bacteria tested were not eaten. Selected species of diatoms, chlorophytes and bacteria were eaten in large quantity. Three additional factors affect feeding: the “age” of the food organism, the “age” of the foraminifer or its position in the life cycle, and the concentration of the food. Feeding by foraminifera on most food is erratic below a concentration of 10³ organisms and is approximately directly proportional to concentration within a range of 10³-10⁶ organisms per 10 ml experimental tube. A natural bloom of Protelphidium tisburyensis was analyzed. A high concentration of 6 species of diatoms characterized the community. A “bloom”-feeder hypothesis for foraminiferal nutrition is presented.     

Biosynthesis of Phosphatidylcholine in Euglena gracilis*†

SYNOPSIS. Extracts of Euglena gracilis carry out a very rapid but limited synthesis of phosphatidylcholine when S-adenosylmethionine or ATP and methionine are supplied. Cytidinediphosphocholine apparently is not utilized. Qualitatively the same results are obtained whether the cells are light- or dark-grown.     

Phylogenetic Origin of the Chloroplast*†

SYNOPSIS. The 16S ribosomal RNA of the chloroplast of Euglena gracilis strain Z has been characterized in terms of its 2-dimensional electrophoretic “fingerprint” (T1 ribonuclease). Over 100 spots were resolved on the “fingerprint” and each spot was characterized as to which RNA oligonucleotide fragment(s) it contained. When compared to similar analyses of prokaryotic 16S rRNAs and eukaryotic cytoplasmic 18S rRNAs, the chloroplast 16S rRNA was a typically prokaryotic RNA, but bore little if any relationship to eukaryotic 18S rRNAs. Therefore, the cistrons for chloroplast 16S rRNA are related to the equivalent prokaryotic cistrons, but, apparently, are not related to the equivalent eukaryotic cistrons. Among the organisms available for comparison, the Euglena chloroplast 16S rRNA appears most closely related to the 16S rRNA of the eukaryote, Porphyridium cruentum (a red alga), and at least distantly related to the 16S rRNAs of the blue-green algae and perhaps also to the bacilli.     

Fine-Structural Aspects of Macrogametogenesis in Eimeria magna*†

SYNOPSIS. Macrogamonts in tissues from rabbits killed 5 1/2 days after inoculation with Eimeria magna oocysts were studied with the electron microscope. In young macrogamonts, parts of cytoplasm, sometimes including micronemes, were pinched off into the parasitophorous vacuole. In all stages of development, small segments of the inner membrane complex were present beneath the limiting membrane. Micropores also were seen in all stages, and some apparently functional ones were present in mature macrogametes. Wall-forming bodies of Type I and Type II were observed in relatively early stages. The former were less numerous than the latter, which had a more compact appearance than in other species. Usually, several Golgi complexes were present and several Golgi adjuncts occurred in the vicinity of the nucleus in all stages of development. Microgametes were observed in the cytoplasm of host cells harboring immature macrogametes.     

Bioelectric Control of Locomotion in the Ciliates*†

SYNOPSIS. Locomotor behavior in the ciliate protozoa is controlled by the cell membrane through electrophysiological principles already familiar in receptor, nerve, and effector cells of the metazoa. This is illustrated by the avoiding reaction (15). When the membrane of the anterior part of the ciliate receives a mechanical stimulus, as during collision, it permits a local influx of Ca⁺⁺. This constitutes a receptor current which depolarizes the remaining cell membrane by electrotonic spread. Depolarization causes a secondary transient increase in the calcium conductance of the entire cell membrane, and a general influx of Ca⁺⁺ occurs. The resulting increase in concentration of intracellular Ca⁺⁺ activates a reorientation (“reversal”) of the ciliary power stroke, causing the organism to swim backward. Forward locomotion is restored as the resting concentration of intracellular Ca⁺⁺ in the cell cortex is restored by diffusion, active extrusion, or intracellular sequestering. The control and coordination of locomotion in ciliates depend on several factors in addition to the excitable properties of the membrane. These include the sensitivities of the ciliary apparatus to intracellular concentrations of calcium and other regulating substances, the anatomical distribution of sensory receptor properties of the cell membrane, and the cable properties of the cell which permit electrotonic spread of graded potential signals without need of all-or-none conducted signals.     

Temperature-Enhanced Osmotic Growth Requirement of Crithidia*†

SYNOPSIS. An osmotic growth requirement for the trypanosomatid Crithidia fasciculata became conspicuous at 32 C. Osmo-supportive compounds were surveyed at 28 vs. 33 C, in “low-osmotic” defined medium. Effective osmotic support was provided by many compounds, e.g., glycerophosphate, sorbitol, mannitol, glycerol, Na isethionate, glycine, arginine, KCl, NaCl, NH₄Cl, and K₂SO₄. The nonspecificity of the requirement was thus evident, but inactivity of the presumably poorly adsorbable pentaerythritol indicated that osmotic pressure was a likely but insufficient condition for satisfying the temperature-enhanced growth requirement most clearly expressed as a need for osmotic support. Fortification of the medium with a combination of glycerophosphate, glycine, glycerol, and Tween 80 permitted good growth at 35 C. Possible relations between the temperature-enhanced osmotic requirement cell membrane damage, and morphological phases of Trypanosoma and Leishmania are discussed.     

Algal Flagellate Symbiosis in the Foraminifer Archaias*†

SYNOPSIS. A symbiotic alga from the foraminifer Archaias angulatus was isolated axenically. Algae from crushed hosts were coccoid and highly vacuolated; division stages within an envelope were common. Biflagellate motile piriform organisms predominated in newly transferred cultures and were gradually replaced by the coccoid, highly vacuolated stage. Incorporation of ¹⁴C label in intact Archaias was greatest for organisms fed and incubated in light. Starved symbiotized organisms incubated in the light incorporated ～30% as much label as fed counterparts, There was no obvious difference in ⁴⁵Ca incorporation between fed and starved organisms. Light significantly enhanced calcification. The Archaias symbiont infected Rosalina leei but not Quinqueloculina spp.     

Contractility and its Control in Peritrich Ciliates*†

A review of the recent literature on the structure and physico-chemical properties of the myoneme and its specialization, the spasmoneme, of peritrich ciliates was made. Myonemes are composed of tightly packed bundles of 3–5 nm microfibrils which parallel, more or less, the long axis of the bundle and are of indefinite length. The presence of contractility in these ciliates is correlated with the presence of myonemes. Associated with the microfibrils is a system of membrane-bound tubules and saccules continuous with the endoplasmic reticulum (ER). This system is known to accumulate calcium. Myonemes differ from muscles in their structure, solubility properties, birefringence pattern, and in the time during the contraction-relaxation cycle at which they require energy. They may be related more closely to the cytochalasin B-sensitive microfibrils of higher organisms than to muscles. Contraction of extracted stalks can be induced solely by raising the calcium ion concentration above a certain threshold. Thus the calcium-accumulating myoneme-associated ER would appear to play an important role in the control of myoneme contractility. A specialization at the interface between the myoneme and the ER membranes is described as it appears in Vorticella and Opercularia. This structure, called a linkage complex, is found both in the body myonemes and the spasmoneme and links the membranes of the ER to the microfibrils. It also has microfilaments that pass from the ER-myoneme interface to the surface membranes. The uniqueness of this structure and its location suggests that it may play a role in controlling the movements of calcium between the ER and myoneme and also in transmission of messages from the pellicular membranes, possibly the alveolar sacs, to the ER.     

Anemia in Ducks Infected with Leucocytozoon simondi*†

SYNOPSIS. Thirty-four white Pekin ducklings were used to study the anemia associated with infection by Leucocytozoon simondi. The first appearance of gametocytes in the peripheral blood, as detected by thin smears, marked the onset of anemia. This anemia lasted for the duration of the initial parasitemia, usually reaching a low point early in the infection (1 to 5 days post patency) and returning slowly to normal as the parasitemia decreased. Greatest gametocyte density occurred 5 to 8 days post patency. In a number of cases recovery from anemia began simultaneously or even prior to the highest level in the gametocyte density. In low level parasitemias a fluctuation occurred in erythrocyte numbers which corresponded with the peaks of gametocyte density. In none of the infections was a sufficient number of parasite observed to account for the existing anemia. Haemopoietic activity was observed for a brief period at the time of maximum erythrocyte loss in only a few birds. The overcompensation for erythrocyte loss at the end of the primary parasitermia favors the view that increased erythrocyte production may account for the short duration of haemopoiesis.     

What the Peritrichs Have Taught Us*†

SYNOPSIS. The peritrichs have taught naturalists, biologists, zoologists, and other kinds of scientists many of their secrets. Consequently, we know many details regarding their gross structure, ultrastructure, ecology, and life history. Their sexual phenomena are rather clearly understood. While general agreement has been attained as regards the forementioned topics, the taxonomic position of peritrichs is still vigorously debated. In this address representative examples of advances in our knowledge of peritrichs will be reviewed … representative instead of all-inclusive examples. A plea for the restoration of peritrichs to a taxonomic position above spirotrichs will be made. The Society's members will be assured that all protozoologists will have the opportunity to help design a completely International Commission on Protozoology in the International Union of Biological Sciences and they will be assured that such a Commission can and should be planned at the 1969 International Conference in Leningrad.     

Dinoflagellate Evolution: Speculation and Evidence*†

SYNOPSIS. Nuclear features of dinoflagellates that were used originally to support the Mesocaryota concept are reviewed. The fibrillar diameter of dinoflagellate chromatin, low level of chromosomal basic proteins, membrane attachment of chromosomes and swirl pattern observed in sectioned chromosomes are features that support a prokaryotic affinity. The presence of repeated and highly complex DNA, a S-phase of DNA synthesis in the cell cycle, presence of basic proteins, and the reinterpretation of extranuclear microtubules as a spindle support the contention that dinoflagellates are eukaryotes. This combination of prokaryotic and eukaryotic features suggests that dinoflagellates are a geologically old group and that perhaps they diverged from the higher eukaryotic lineage before evolution of eukaryotic chromatin but after the evolution of repeated DNA. The 2 patterns of carotenoid composition exemplified by the presence of peridinin or fucoxanthin suggest separate origins of dinoflagellate plastids, perhaps by prokaryotic and eukaryotic capture. It is suggested that the species possessing fucoxanthin obtained their plastids by capture of photosynthetic eukaryotes. A new class and order, Syndiniophyceae and Syndiniales, are proposed for the dinoflagellates with low chromosome numbers, V-shaped chromosomes, chromosomes containing a sufficient quantity of basic proteins detectable histochemically, possession of centrioles associated with mitosis, intracellular parasitism as a mode of nutrition, and lack of a cellular covering containing plates. Ultrastructural and paleontologic evidence indicates that the thecate is more primitive than the nonthecate condition. The Prorocentrales are considered to be primitive and their thecal construction is reinterpreted as having epithecal and hypothecal regions surrounding a flagellar pore region containing 7 plates. Acritarchs resemble cysts of modern dinoflagellates in size, structure, and chemical composition except for the absence of a polygonal excystment aperture and lack of any indication of transverse and longitudinal flagellar grooves on the acritarchs. The suggestion that some acritarchs may have dinoflagellate affinities is supported by the occurrence of modern dinoflagellates (Prorocentrales) which lack a theca of numerous polygonal plates and lack transverse and longitudinal flagellar arrangement. The Prorocentrales, as opposed to the more typical Dinophyceae, perhaps represent the type of organism that produced some acritarchs.     

Synthetic Requirements of Chloroplast Development in Chlamydomonas reinhardi*†

SYNOPSIS. The synthesis of chlorophyll by chlorotic cultures of Chlamydomonas reinhardi is stimulated by a utilizable carbon source. This synthesis is almost completely inhibited by cycloheximide in concentrations as low as 0.50 μg/ml. The period of chloroplast development during which stimulation by carbon source and inhibition by cylcoheximide are most effective corresponds to a period of rapid formation of chloroplast lamellae.     

Poly(A)+ RNA from Tetrahymena: Stimulation of Protein Synthesis in Vitro*†

SYNOPSIS Cell-free synthesis of high molecular weight polypeptides, programmed by RNA from Tetrahymena pyriformis strain W is reported, and methods for preparation of the RNA are described. The RNA was extracted by the SDS-phenol-chloroform-isoamyl alcohol technic. The bulk of extracted RNA was ribosomal and on sucrose gradients peaked at -17S and 25S. After heat denaturation all the 25S RNA was converted to 17S. indicating the presence of hidden breaks, possibly the result of nuclease activity during extraction. Nevertheless, when poly(A)–RNA was collected using oligo-(dT)-cellulose column chromatography, it promoted a 15–fold increase in incorporation of [35S] methionine into TCA-precipitable material. Slab-gel electrophoresis and autoradiography of the product revealed 12 different major polypeptides, varying in weight from 28.000 to 65,000 Daltons. A method for preparation of translatable RNA from Tetrahymena will make possible the comparison of messenger RNAs associated with specific cell structures and with different developmental events.     

The Positioning of Ciliary Organelles in Hypotrich Ciliates*†

It is commonly observed in hypotrichs that new ciliary rudiments arise directly from or in close juxtaposition to certain pre-existing ciliary elements. Oral primordia often are initiated near specific cirri, cirral rudiments frequently arise as a result of the disaggregation of certain old cirri, and new dorsal ciliature is formed within pre-existing ciliary rows. In the first 2 situations it has been demonstrated experimentally that neither the old ciliature in question nor the specific cortical site marked by that ciliature is essential for the appearance of the new cirral rudiment. The experimental analysis done thus far suggests that the positions of oral and cirral primordia are determined by interacting gradients established in relation to certain reference points. The nature of the reference points is not fully elucidated; in some cases at least these points appear to be more closely related to topographic features of the cell than to specific pre-existing cortical structures. In the dorsal ciliary rows of Euplotes new ciliary units are formed usually and perhaps invariably in close proximity to old ones, and are generally oriented along the axis of the pre-existing row. The result is a tendency to perpetuate the preexisting row number across cell generations. Changes in row number, however, can occur as a result of occasional formation of new units at right angles to the row, a process that is much enhanced in certain homozygous segregants (basal body deficient). The optimal row number (stability range) as well as the number of ciliary units are under genic control. In addition, the spatial pattern of distribution of ciliary units among rows is invariant in all of the material examined. This pattern is presumed to result from an underlying field whose geometry is independent of both the number of units and the number of rows.     

Fine Structure of Human Malaria In Vitro*†

SYNOPSIS. The erythrocytic cycle of the human malaria parasite, Plasmodium falciparum, was examined by electron microscopy. Three strains of parasites maintained in continuous culture in human erythrocytes were compared with in vivo infections in Aotus monkeys. The ultrastructure of P. falciparum is not altered by continuous cultivation in vitro. mitochondria contain DNA-like filaments and some cristae at all stages of the erythrocytic life cycle. The Golgi apparatus is prominent at the schizont stage and may be involved in the formation of rhoptries. In culture, knob-like protrusions first appear on the surface of trophozoite-infected erythrocytes. The time of appearance of knobs on cells in vitro correlates with the life cycle stage of parasites which are sequestered from the peripheral circulation in vivo. Knob material of older parasites coalesces and forms extensions from the erythrocyte surface. Some of this material is sloughed from the host cell surface. The parasitophorous vacuole membrane breaks down in erythrocytes containing mature merozoites both in vitro and in vivo. Merozoite structure is similar to that of P. knowlesi. The immature gametocytes in culture have no knobs.     

Biochemical Approaches to Problems of Cellular Patterning*†

SYNOPSIS. The expression of intracellular patterning is perhaps nowhere more impressive than in the arrangements of structural elements associated with the cell surface in protozoa. The view is proposed that biochemical studies of protozoan plasma membranes and associated surface structures represent important contributions of potential significance for the understanding of the perpetuation, and expression of positional information at the intracellular level. Some recent work dealing with the isolation, identification, and metabolism of pellicular proteins in Tetrahymena is presented and discussed. Some integral membrane proteins have been identified by iodination and polyacrylamide gel electrophoresis. Labeling studies suggest heterogeneous turnover rates within the group of presently identified membrane proteins. High molecular weight proteins with some similarity to spectrin have been isolated from Tetrahymena epiplasm. It is suggested that the ciliate epiplasm is one example of membrane-associated, actomyosin-like systems found in a variety of cell types. The epiplasm may play a role in the positioning of surface-associated structures and in the control of cell shape.     

Morphometric Variation in the Hypotrich Ciliate Genus Euplotes*†

SYNOPSIS. A quantitative method for the analysis of cirral patterns in the genus Euplotes is developed and contrasted with alternative approaches. Inherent variations in cirral pattern, as they are reflected in frequency distributions of intercirral distances, are ascertained through subclonal analyses of Euplotes harpa samples grown under different conditions. This assessment of the extent of variation within a form having 10 frontoventral cirri provides a basis for a comparative study of other Euplotes of cirrotype 10. These results suggest that all marine cirrotype 10 forms with a single dargyrome have the same configuration of frontoventral and transverse cirri. Similarly, most marine double dargyrome forms of cirrotype 10 have this same cirral pattern, and thus constitute variations upon the same morphometric theme.     

Cholesterol Inhibition of Pentacyclic Triterpenoid Biosynthesis in Tetrahymena pyriformis*†

SYNOPSIS. Tetrahymena pyriformis synthesizes tetrahymanol and “diplopterol” from acetate, mevalonate or squalene. The formation of these pentacyclic triterpenoid alcohols is inhibited by the addition of cholesterol to the culture fluid of the ciliates. Cholesterol also inhibits the biosynthesis of squalene from acetate or mevalonic acid. The synthesis of other terpene derivatives from acetate and mevalonate continues in the presence of cholesterol, thus suggesting that a major block occurs after “isoprene” formation and before squalene formation. Further, inhibition of squalene conversion to the pentacyclic alcohols by cholesterol has been established.