Growth of Particle-Bearing and Particle-Free Paramecium aurelia in Axenic Culture*

SYNOPSIS. Several strains of particle-bearing and particle-free Paramecium aurelia have been cultivated in an axenic medium composed of proteose peptone, trypticase, yeast nucleic acid, MgSO⁴.7H²O, TEM-4T (diacetyl tartaric acid esters of tallow monoglycerides), stigmasterol and a mixture of vitamins. The “yeast fraction,” an indispensable component of previous media used for the cultivation of these ciliates has been replaced by a mixture of trypticase, yeast nucleic acid and TEM-4T. Particle-bearing animals of stock 299 lambda, 138 mu, and 139 pi maintain their particles when cultivated in the medium, whereas particle-bearing animals of stock 51 kappa, 225 kappa and 114 signia do not. With the exception of stock 92 (syngen 3) the medium appears to be selective in its ability to support the growth of animals of the even- but not odd-numbered syngens of P. aurelia. Maintenance of the particles was dependent only to a small degree upon environmental conditions brought about by changes in pH and temperature. Division of the particles was found to be comparable with the division of the protozoan. Methods for the growth, maintenance and mass cultivation of particle-bearing P. aurelia are given in detail.     

Aerobic Respiration of Kappa Particles from Paramecium aurelia,Stock 51*

SYNOPSIS. Kappa particles from killer cultures of stock 51 Paramecium aurelia were purified and their respiration measured polarographically. The slight bacterial contaminations in the kappa preparations were not significant. Freshly collected kappa in dilute buffer at room temperature had an endogenous Q_O2 of 17.0 ± 1.6 μl/mg dry weight/hr (mean ± standard error). The Q_O2 decayed 50% in 5 hr. Among the sugars tested only glucose and sucrose increased the respiratory rate of kappa. The di- and tri-carboxylates of the tricarboxylic acid cycle stimulated the respiration of kappa. KCN, CO and 2-heptyl-4-hydroxyquinoline N-oxide (HOQNO) inhibited respiration. These findings ensure an organismic status for kappa and justify the belief that it is bacterial in origin.     

The Axenic Culture of Strains of the Various Syngens of Paramecium aurelia*

SYNOPSIS Strains of the various syngens of Paramecium aurelia respond differently to the culture media developed for Strain 299 of syngen 8. Not only is there a variety of response between the syngens, but strains belonging to the same syngen respond differently to the 3 growth media.     

Abnormalities in Nuclear Behavior and Mating Type Determination in Cytoplasmically Bridged Exconjugants of Doublet Paramecium aurelia*

Exconjugant clones of Paramecium aurelia stock 51S, syngen 4, which fail to separate prior to the 1st fission have numerous cytologic and mating type determination anomalies. The doublets have abnormal distribution of macronuclear anlagen, fewer macronuclear fragments per cell, and abnormalities in numbers of micronuclei. Despite apparent cell fusion and mixture of cytoplasm, the singlets arising from each side of the doublet may be of opposite mating types, and mating type determination may remain unstable for 1 or more fissions in contrast to the usual pattern of mating type determination before the 1st postconjugation fission.     

The Nutrition of Paramecium aurelia, Stock 299

SYNOPSIS. Paramecium aurelia , stock 299 (symbiote-free) was cultivated in a synthetic medium consisting of amino acids, vitamins, purine and pyrimidine derivatives, fatty acids, stigmasterol, sodium acetate and salts. The medium supported the continued growth of this stock in serial subculture. Populations up to 17,000 organisms/ml were obtained in 9 or 10 days in the medium supplemented with a phospholipid. Synthetic 1-oleoyl-2-stearoyl-dl-phosphatidyl serine, phosphatidyl ethanolamine, phosphatidyl inositol, phosphatidyl serine and cephalin were comparable in growth-promoting activity. The nutritional need for each of the components of the medium was examined. The following were determined to be essential nutrilites for P. aurelia : arginine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, threonine, tryptophan, tyrosine, valine, folic acid, nicotinamide, calcium pantothenate, pyridoxal, riboflavin, thiamine, DL-6-thioctic acid, guanosine, uridine (or cytidine), oleic acid, stigmasterol, calcium and magnesium. Serine replaced glycine for growth in the presence of thymidine. In the absence of thymidine, comparatively high levels of folic acid were required for optimal growth. Sodium acetate did not replace DL-6-thioctic acid. Populations were reduced in the absence of the non-essential amino acids, alanine, asparagine, aspartic acid and glutamic acid. These were restored to optimal levels by the addition of sodium acetate to the medium. Pyruvate was about as effective as acetate in this respect; glucose and certain other carbohydrates were not.     

Axenic Paramecium caudatum. I. Mass Culture and Structure*

SYNOPSIS. To establish and grow Paramecium caudatum in mass axenic culture the culture medium of Soldo, Godoy & van Wagtendonk was modified by substituting phosphatidylethanolamine (PE) for TEM-4T and by a 10-fold increase in folic acid. Population densities of 4000 to 6000 cells/ml and a generation time of 20–26 h are regularly obtained. Optimal growth is obtained with PE-stigmasterol ratios between 40:1 to 400:1. Cells from 1-day-old axenic cultures have many lipid bodies aggregated in clumps (which disappear in 2 to 3 days) as well as foci of rough endoplasmic reticulum bordered by dictyosomes. The latter suggests a very active metabolism. Crystalline sheets found in both food vacuoles and lysosomes presumably play a role in digestion. Axenically grown cells also have abundant Golgi bodies (dictyosomes) and by late log phase become filled with lysosomes.     

Antigenic transformations in Paramecium aurelia,variety 4, stock 51 under the influence of trypsin and chymotrypsin

Exposure of P. aurelia to trypsin and chymotrypsin results in the death of a large percentage of the population. Not all antigenic types are equally affected; antigen types 51B and 51E are most affected by exposure to trypsin, while antigen type 51B is most affected by the action of chymotrypsin. Type 51A does not transform when exposed to either trypsin or chymotrypsin, or after the animals have been removed from the enzyme preparations. Type 51B shows little transformations. Types 51C, 51D and 51E transform readily during and after exposure to these two enzymes. In every instance exposure to the enzymes, followed by growth at 20 °, 27 ° or 32 ° results in the transformation to more type 51A than growth at these temperatures alone. Exposure to chymotrypsin leads invariably to more transformation to type 51A than does exposure to trypsin. Exposure of type 51D to either trypsin or chymotrypsin gives rise to type 51J. The obtained results are analogous with those obtained by transformations, resulting from exposure to antiserum or exposure to different temperatures.     

Induction of Temperature Sensitivity in Paramecium aurelia by Nitrosoguanidine*

Paramecium aurelia cells were exposed to N-methyl-N-nitroso-N′-nitroguanidine for periods of 15–30 min. The lethality in homozygous clones derived from treated cells depends on the time of treatment within the cell cycle. Exposures at interfission ages 0.04, 0.40, and 0.80 were tested yielding lethalities of 12.5, 44 and 2%, respectively. These results correlate with the period of DNA synthesis in the micronuclei. A temperature sensitive mutant has been found which cannot live at 31 C but divides at ～1 fission per day at 19 and 25 C. The rise in temperature from 19–25 C does not significantly change the fission rate whereas in normal cells it would be doubled. Genetic analysis shows that this mutation is caused by a single recessive gene.     

The Feeding of Amoeba proteus on Paramecium aurelia*

SYNOPSIS. Density of prey (Paramecium aurelia) and predator (Amoeba proteus) were varied while volume of inorganic medium was kept constant. Variations in density of prey had little effect on the rates of feeding and reproduction of the amoebae; but with increasing predator density the amoebae captured the paramecia less rapidly and ingested fewer before dividing, altho division size did not change appreciably. Therefore, amoebae of a low density population with a constant food supply carry more nutritional reserves from generation to generation than do those in a denser population.     

Chemical Induction of Autogamy in Paramecium multimicronucleatum,Syngen 21

The combination of KCl + acriflavine + Ca²⁺-poor condition, known as conjugation-inducing-chemicals, was found to be autogamy-inducing-chemicals as well. Suspension of a single cell of Paramecium multimicronucleatum, syngen 2 in this medium for three hours or more resulted in autogamy that was evidenced by cytological similarity to conjugation.     

Subcellular Effects of Cytochalasin B and Dimethylsulfoxide on Paramecium aurelia*

SYNOPSIS. Paramecium aurelia syngen 4, stock 57 (sensitive) cultivated in Cerophyl infusion were exposed to cytochalasin B CB and dimethylsulfoxide (DMSO), the solvent for CB, to distinguish between the effects of these agents on a cellular system. DMSO significantly inhibited survival, fission rate, [³H]leucine incorporation, and cell size. CB-treated cells generally had slower division and poorer survival rates than cells exposed to the equivalent DMSO concentration, although the [3H]leucine incorporation was generally greater at the lower CB concentrations than for DMSO alone. As seen by electron microscopy and a new grycerination technic for observing polysomes, DMSO caused nuclear (nucleolar, chromatin) abnormalities as well as membrane degradation and polysomal breakdown; CB caused the formation of aberrant membrane structures and ribosomal tetramers, crystals, and tubes.     

Paramecium tredecaurelia of the Paramecium aurelia complex in Israel

The paper concerns the finding of a new habitat (Kiryat Motzkin, north of Haifa, Israel) of Paramecium tredecaurelia from the P. aurelia complex. This is only the forth known locality of the species in the world. Previously, its strains were obtained from widely separated localities: the River Seine, Paris, France; Benenitra, Madagascar, and the Cuernavaca Valley, Taxco, Mexico. The studied strain originating from Israel was identified as P. tredecaurelia on the basis of the strong (90%) conjugation between the complementary mating type of the examined clones with the appropriate standard strain 209 of P. tredecaurelia from Paris, France (restricted to odd mating type). However, the strain from Israel is restricted to the even mating type.