Further studies on Nephroselmis and its allies (Prasinophyceae). I. The question of the genus Bipedinomonas

Based on a series of collections made in New Zealand it is concluded that Nephroselmis longifilis and Bipedinomonas pyriformis are identical species, for which the correct name is Nephroselmis pyriformis (N. Carter) comb. nov. A detailed examination by light and electron microscopy is given, using material from geographically widely separated areas, including Greenland, the North Sea, a shallow Danish Fiord, Thailand and New Zealand. N. pyriformis is a temperature– and halotolerant species, as shown by its known range of occurrence at temperatures from 2.3°C (West Greenland) to ca. 28°C (Thailand) and at salinities between 3 and 36%o. Features of fine structural interest include details of the scaly covering on the flagellar and cell surfaces, the former very similar to those on the flagella of Tetraselmis cordiformis , a quadriflagellate prasinophyte. Within the cell, the pyrenoid shows an unusual fine structure, being penetrated by a number of membrane–lined pockets, in which the membranes are continuous with thylakoids. N. pyriformis is compared with other members of the genus, and a list of the few known species is included.     

AGRILUS CUPRIPES (CARTER) COMB. N. (COLEOPTERA: BUPRESTIDAE)

i
Synechocera cupripes Carter is transferred to Agrilus Stephens.
Subfamily BUPRESTINAE
Tribe AGRILINI
Genus Agrilus Stephens
Agrilus Stephens, 1830, Ill. Brit Ent. 3: 239.
Agrilus cupripes (Carter) comb.n.
Synechocera cupripes Carter, 1924, Proc. Linn. Soc. N.S.W. 49: 31.     

Determination of bioavailability of some long-chain N-substituted derivatives of L-methionine and L-lysine

The bioavailability of N-acyl-L-methionine derivatives has been determined using microbiological assay with Tetrahymena pyriformis. It was found that palmitoyl- and stearoyl-L-methionine, stearoyl-L-methionine ethyl ester, and stearoyl-L-methionine sodium salt were partially utilized (14-38%) for growth of the microorganism. These compounds partially inhibited utilization of free methionine added to the media. The shorter derivatives, acetyl-, hexanoyl-, lauroyl-, and myristoyl-L-methionine completely inhibited the growth of T. pyriformis. This effect was not reversed when DL-methionine was added to the media. N6-fatty acyl L-lysine derivatives gave low availability values (3-18%) in microbiological assessment with T. pyriformis. N2-Acetyl-and N6-acetyl-lysine did not inhibit the utilization of the added parent amino acid. Nutritional evaluation of L-methionine derivatives by the rat growth method using net protein ratio (NPR) as the performance index indicated complete availability of stearoyl-L-methionine, stearoyl-L-methionine sodium salt, and partial availability of stearoyl-L-methionine ethyl ester (52%), stearoyl-L-methionylglycine (32%), and lauroyl-L-methionine (75%).     

A specific uncoupler-binding protein in Tetrahymena pyriformis and Paracoccus denitrificans

The uncoupler of mitochondrial oxidative phosphorylation, 2-nitro-4-azido-carbonylcyanide phenylhydrazone (N3CCP) which is capable of photoaffinity labeling has been used to examine the effect of uncouplers on the energy conserving membrane of Paracoccus denitrificans and Tetrahymena pyriformis. The N3CCP uncouples respiration in P. denitrificans and T. pyriformis cells with U1/2 values of 1.05 microM and 0.24 microM, respectively. Binding studies show the presence of 0.65 +/- 0.05 high affinity sites per cytochrome alpha with Kd of 0.5 +/- 0.1 microM in P. denitrificans membranes and 1.4 +/- 0.2 sites per cytochrome alpha 2 with a Kd of 0.4 +/- 0.1 microM in T. pyriformis membranes. Irradiation of [3H]-N3CCP bound to the membranes leads to a covalent linking of the radioactive uncoupler to a peptide of 10--15 kdaltons as analyzed by SDS-polyacrylamide gel electrophoresis. It is concluded that these two microbial systems contain a specific high affinity uncoupler binding site very similar to that of mammalian mitochondria (Katre, N.V. and Wilson, D.F. (1978) Arch. Biochem. Biophys. 191, 647--656).     

Microelectrode of the Thomas type using a liquid membrane electrode

By replacing the glass-based pH electrode (L. R. Pucacco, S. K. Corona, H. R. Jacobson, and N. W. Carter (1986) Anal. Biochem. 153, 251-261) with a liquid membrane-based pH electrode, a relatively easy-to-manufacture modified Thomas electrode has been developed. The liquid membrane-based modified Thomas electrode can be manufactured without the special equipment (forge) and materials (glass) required to make the glass membrane pH microelectrode (L. R. Pucacco and N. W. Carter (1976) Anal. Biochem. 73, 501-512). The sensitivity (57.4 +/- 0.22 mV/pH unit), response time (20.0 +/- 2.67 s), and electrical resistance (3.48 +/- 0.67 X 10(11) ohm) of this electrode are similar to those of the glass-based version.     

Antiprotozoan activity of nonspecific cytotoxic cells (NCC) from the channel catfish (Ictalurus punctatus)

Nonspecific cytotoxic cells (NCC) obtained from channel catfish (Ictalurus punctatus) kill Tetrahymena pyriformis, an opportunistic parasite in fish. Based upon this fact, a new mechanism for nonspecific cellular anti-parasitic immunity in fish is proposed. Optimum in vitro conditions for NCC killing of deciliated T. pyriformis were first obtained. Lysis of T. pyriformis by NCC occurred by 10 hr of cocultivation of effector and target cells. During this time period, 50 to 60% cytotoxicity occurred. Fish anti-T. pyriformis serum enhanced NCC killing of T. pyriformis either by prolonging immobilization (after the cilia regeneration period) or by delaying cilia regeneration. Shared antigenic determinants between T. pyriformis, Ichthyophthirius multifiliis, and NC-37 target cells were demonstrated by binding-depletion experiments. For these studies, NCC were depleted from anterior kidney cells (the hemopoetic organ in fish) by preincubating formalin-treated T. pyriformis, I. multifiliis, or viable NC-37 target cells with NCC for 3 hr. Conjugates of effector and target cells were removed by overlaying on fetal bovine serum. Unconjugated fish anterior kidney cells were tested for cytotoxic activity against NC-37 or T. pyriformis target cells. Cold target inhibition experiments by using a 4-hr 51chromium cytotoxicity assay also demonstrated these shared antigenic determinants. Target-specific antisera, used to mediate the killing of T. pyriformis by NCC, were required only for immobilizing the targets, and did not function in an antibody-dependent cell-mediated (ADCC)-like mechanism. Scanning electron micrographs of NCC-T. pyriformis conjugates additionally demonstrated NCC binding to both cilia and cell surface determinants.     

Recognition of Prey by Suctoria: The Role of Cilia

ABSTRACT. The interaction between the predator, Discophrya collini , and the prey, Tetrahymena pyriformis , was investigated in order to determine what prey factors(s) may be recognized, facilitating the feeding response of the suctorian predator. When viable, deciliated T. pyriformis were offered to starved D. collini , only 12% of the suctoria captured a ciliate compared to 98% when presented with ciliated T. pyriformis . Starved suctoria were also offered three types of tanned sheep erythrocytes used as artificial prey. After exposure to erythrocytes coated with sonicates of T. pyriformis , 16% of the D. collini displayed one or more attached erythrocytes but after incubation with erythrocytes coated with T. pyriformis cilia, 72% of the suctoria had tentacles with attached cells. No untreated, tanned erythrocytes were captured. When T. pyriformis were treated with ciliary antiserum and then offered to starved D. collini , the resulting capture rate was low (13%), but T. pyriformis treated with normal rabbit serum were captured at a rate comparable to controls (92%). These results suggest that the prey's cilia and/or their surface macromolecules may facilitate prey capture.     

Complete sequence of the mitochondrial genome of Tetrahymena pyriformis and comparison with Paramecium aurelia mitochondrial DNA

We report the complete nucleotide sequence of the Tetrahymena pyriformis mitochondrial genome and a comparison of its gene content and organization with that of Paramecium aurelia mtDNA. T. pyriformis mtDNA is a linear molecule of 47,172 bp (78.7 % A+T) excluding telomeric sequences (identical tandem repeats of 31 bp at each end of the genome). In addition to genes encoding the previously described bipartite small and large subunit rRNAs, the T. pyriformis mitochondrial genome contains 21 protein-coding genes that are clearly homologous to genes of defined function in other mtDNAs, including one (yejR) that specifies a component of a cytochrome c biogenesis pathway. As well, T. pyriformis mtDNA contains 22 open reading frames of unknown function larger than 60 codons, potentially specifying proteins ranging in size from 74 to 1386 amino acid residues. A total of 13 of these open reading frames ("ciliate-specific") are found in P. aurelia mtDNA, whereas the remaining nine appear to be unique to T. pyriformis; however, of the latter, five are positionally equivalent and of similar size in the two ciliate mitochondrial genomes, suggesting they may also be homologous, even though this is not evident from sequence comparisons. Only eight tRNA genes encoding seven distinct tRNAs are found in T. pyriformis mtDNA, formally confirming a long-standing proposal that most T. pyriformis mitochondrial tRNAs are nucleus-encoded species imported from the cytosol. Atypical features of mitochondrial gene organization and expression in T. pyriformis mtDNA include split and rearranged large subunit rRNA genes, as well as a split nad1 gene (encoding subunit 1 of NADH dehydrogenase of respiratory complex I) whose two segments are located on and transcribed from opposite strands, as is also the case in P. aurelia. Gene content and arrangement are very similar in T. pyriformis and P. aurelia mtDNAs, the two differing by a limited number of duplication, inversion and rearrangement events. Phylogenetic analyses using concatenated sequences of several mtDNA-encoded proteins provide high bootstrap support for the monophyly of alveolates (ciliates, dinoflagellates and apicomplexans) and slime molds.     

A cytophotometric study of the influence exerted by epidermal growth factor on RNA and protein synthesis in the ciliate Tetrahymena pyriformis

The influence of EGF (10(-8) M) on RNA and protein synthesis in the ciliate Tetrahymena pyriformis was studied. It was found that EGF stimulated RNA and proteins synthesis in T. pyriformis within 3 h after addition of EGF.     

Dissimilar effects of L and D amino acids on the growth of Tetrahymena

Of the L and D configurations of four amino acids (phenylalanine, valine, tryptophan, tyrosine) tested for influence on the growth rate of Tetrahymena, only L-tyrosine was able to induce imprinting in Tetrahymena pyriformis Zeuthen. D-valine stimulated the division of T.pyriformis NT-1, but failed to induce imprinting. The experiments have substantiated the selectivity of the amino acid receptors of Y.pyriformis, and the extraordinary imprinting potential of tyrosine as well, as judged by its influence on the growth rate.     

Poliovirus and echovirus survival in Tetrahymena pyriformis culture in vivo

Axenic cultures of Tetrahymena pyriformis, strain I MT IV, grown in a defined medium at room temperature, were used to study interactions of these protozoa with vaccination strain L Sc 2ab of poliovirus type 1, vaccination strain P 712 of poliovirus type 2 and with type 30 echovirus, strain 480/78. T. pyriformis cultures in media containing 10(3.0) TCD50/1 ml of type poliovirus, 10(3.0) TCD50/1 ml of type 2 poliovirus or 10(2.5) TCD50/1 ml echovirus 30 and in virus-free medium did not differ one from another in their growth and die-away kinetics during the 21 days of observation. Two-day T. pyriformis cultures were infected with poliovirus 1 (initial concentration 10(3.2) TCD50/1 ml), and poliovirus 2 and echovirus 30 (initial concentrations 10(3.0) TCD50/1 ml). Viruses were titrated in test tube cultures of BGM cells. The supernatant fluid, standardized sediment and samples of control virus suspension free of protozoa were titrated after 0, 2, 6, 10, 13, 18, 28 and 30 days. Most of the virus in culture was found associated with the sediment, both in the period of active growth and during the die-away phase of T. pyriformis protozoa. The virus in sediment was present at higher titres and its survival time was longer than in virus in liquid phase. Thirteen days after the first contact between T. pyriformis and virus the sediment and supernatant fluid of the old protozoan culture and the T. pyriformis-free control viral suspension were taken and used as inocula for new two-day T. pyriformis cultures.(ABSTRACT TRUNCATED AT 250 WORDS)     

Proliferation of Legionella pneumophila as an intracellular parasite of the ciliated protozoan Tetrahymena pyriformis.

In a series of experiments, we have determined that Legionella pneumophila will proliferate as an intracellular parasite of the ciliated holotrich Tetrahymena pyriformis in sterile tap water at 35 degrees C. After 7 days of incubation, serpentine chains of approximately 10(3) L. pneumophila cells were observed throughout the cytoplasm of the protozoan infected initially with 1 to 30 L. pneumophila cells. The overall L. pneumophila population increased from ca. 1.0 X 10(2) to ca. 5.0 X 10(4) cells per ml in the coculture within this time frame. The interactions between the protozoan and the bacterium appear to depend upon their concentrations as well as temperature of incubation. L. pneumophila did not multiply in sterile tap water alone, in suspensions of lysed T. pyriformis, or in cell-free filtrates of a T. pyriformis culture. In addition to establishing an ecological model, we found that addition of T. pyriformis to environmental specimens served as an enrichment method that improved isolation of legionella from the specimens.     

Comparative study of DNA methylation in three unicellular eucaryotes.

We have analyzed the nature/content of methylated bases in the nuclear DNA of three unicellular eucaryotes. The pattern of methylation was different for each of the three organisms studied: Saccharomyces cerevisiae contained only 5-methylcytosine; Tetrahymena pyriformis contained only N6-methyladenine; and Chlamydomonas reinhardi contained both modified bases.     

Use of bioluminescent Escherichia coli O157:H7 to study intra-protozoan survival of bacteria within Tetrahymena pyriformis

A method was developed that enabled real-time monitoring of the uptake and survival of bioluminescent Escherichia coli O157 within the freshwater ciliate Tetrahymena pyriformis. Constitutively bioluminescent E. coli O157 pLITE27 was cocultured with T. pyriformis in nutrient-deficient (Chalkley's) and in nutrient-rich (proteose peptone, yeast extract) media. Non-internalised bacteria were inactivated by addition of colistin, indicated by a decline in bioluminescence. Protozoa were subsequently lysed with Triton X-100 which lead to a further drop in bioluminescence, consistent with release of live internal bacteria from T. pyriformis into the colistin-containing environment. Bioluminescence measurements for non-lysed cultures indicated that internalised E. coli O157 pLITE27 cells were only slowly digested by T. pyriformis, in both media, over the time period studied. The results suggest that bioluminescent bacteria are useful tools in the study of bacterial intra-protozoan survival.     

Proliferation of Legionella pneumophila as an intracellular parasite of the ciliated protozoan Tetrahymena pyriformis

In a series of experiments, we have determined that Legionella pneumophila will proliferate as an intracellular parasite of the ciliated holotrich Tetrahymena pyriformis in sterile tap water at 35 degrees C. After 7 days of incubation, serpentine chains of approximately 10(3) L. pneumophila cells were observed throughout the cytoplasm of the protozoan infected initially with 1 to 30 L. pneumophila cells. The overall L. pneumophila population increased from ca. 1.0 X 10(2) to ca. 5.0 X 10(4) cells per ml in the coculture within this time frame. The interactions between the protozoan and the bacterium appear to depend upon their concentrations as well as temperature of incubation. L. pneumophila did not multiply in sterile tap water alone, in suspensions of lysed T. pyriformis, or in cell-free filtrates of a T. pyriformis culture. In addition to establishing an ecological model, we found that addition of T. pyriformis to environmental specimens served as an enrichment method that improved isolation of legionella from the specimens.     

Mitochondrial deoxyribonucleic acid from Tetrahymena pyriformis and its kinetic complexity

1. Mitochondrial DNA from Tetrahymena pyriformis strain T has a buoyant density (rho) of 1.685 compared with rho1.688 for whole cell DNA. Mitochondrial preparations from T. pyriformis strain W show an enrichment of a light satellite (rho1.686), although this is not obtained free from nuclear DNA (rho1.692). 2. T. pyriformis mitochondrial DNA renatures rapidly and the kinetics of this process indicate a complexity of approx. 3x10(7) daltons. 3. The base-pairing in the renaturation product is of a precise nature, since the ;melting' temperature (80.5 degrees C) is indistinguishable from that of the native DNA (80.5 degrees C). 4. Centrifugation of mitochondrial DNA in an alkaline caesium chloride density gradient gives two bands, implying the separation of the complementary strands.     

The methylated purines of Tetrahymena pyriformis transfer ribonucleic acid.

By phenol extraction and DEAE cellulose column chromatography, tRNA was isolated from Tetrahymena pyriformis strain GL. Following acid hydrolysis of the tRNA, the methylated purine content was determined by Dowex 50 column chromatography and paper chromatography. The most abundant methylated guanine derivative was found to be N2-DMG. Also present were 1-MG, N2-MG, and 7-MG. The most abundant methylated adenine was found to be 1-MA; no 2-MA was detected. Small amounts of the N6-methyladenines were detected.