Paramecium
- Paramecium aurelia
- Paramecium bursaria
- Paramecium calkinsi
- Paramecium caudatum
- Paramecium multimicronucleatum
- Paramecium putrinum
- Paramecium tetraurelia
[ref. ID; 4047]
Test system
To distinguish between the effect of CB and DMSO
Strains
Syngen 4, stock 57 (sensitive).
Toxicants
Dimethylsulfoxide (DMSO), Cytochalasin B.
Test design
- Lethality: 10 cells (log-phase culture) were isolated into 20, 40, 60 and 70 ug/ml CB (+DMSO); 2, 4, 5, 7, 10, and 20% (v/v) DMSO; and into bacterized (Aerobacter cloacae) culture fluid. The dead cells were counted and removed every 30 min for the first 8 hr or exposure, and then once in every 12 hr. Cells which survived the first 24 hr were reisolated into fresh concentrations of the test reagent.
- Reversibility: Mass cultures (60-80 cells) were placed in 20, 30, 40, 50, 60 and 70 ug/ml CB (+DMSO); 2, 3, 4, 5, 6, and 7% (v/v) DMSO; and bacterized culture fluid. At 30-min intervals, 6 cells were removed at random from each concentration of CB (+DMSO), DMSO, and from the controls, washed once, and then isolated into bacterized culture fluid (1 cells/depression). After 24 hr, the cells in each depression were counted.
- Division rate: Single cells were isolated into 20, 40, 60 and 100 ug/ml CB (+DMSO); 2, 4, 6, and 10% DMSO; and bacterized culture fluid. After 24 hr, the cells were counted and average fission rates culculated.
- [3H]Leucine incorporation: To obtain synchronized cells, dividing ciliates were selected within a 15-min interval from a culture 3-5 fissions after autogamy. 4 hr after division (time within the cell cycle or interfission age ~0.65-0.7), at least 10 cells were isolated into each concentration of CB (+DMSO) (10, 20, 30 and 40 ug/ml), DMSO (1, 2, 3, and 4%) and bacterized culture fluid. 20 min after introduced into the different media, the cells were exposed 20 min to 10 ug/ml [3H]leucine in the appropriate concentration of CB (+DMSO), DMSO, or culture fluid. These cells were then rinsed twice and left for 1 hr in the original dilutions of CB (+DMSO), DMSO, or culture fluid. Following this chase period, cells were affixed by drying onto gell coated microscope slides, fixed in Carnoy's solution.
- Ribosomal tetramers, tubes, and crystals: Log-phase cells were exposed to 70, 100, and 150 ug/ml CB (+DMSO); 7, 10 and 15% DMSO; and bacterized culture fluid for 20-80 min before being fixed for electron microscopy.
Measurements/observations
Ultrastructure of cell organelles.
[ref. ID; 6111]
Test system
Acute toxicity (24-hr LC50)
Strains
The organisms was obtained, initially, from Stirone Stream (northern Italy), a freshwater environment free from known toxicants and, in particular, from heavy metals. The culture medium was constituted of boiled rice and wheat grain in 10 ml of filtered natural water. 20+/-1 degrees C, photoperiod of 16:8 h light:dark.
Toxicant/concentrations
NiCl2/6H2O (0.25, 0.33, 0.45, 0.62 ppm).
Test design
Tissue culture plates with 24 wells, 3 replicates.
Measurements
Mortality or the survivorship.
Evaluations
LC50 using the probit method.
[ref. ID; 7301]
Test system
Aminotransferase and the production of alanine during hyperosmotic stress
Strains
SM-3 from a small marsh that drained into the eastern edge of the Eel Pond in Woods Hole, Massachusetts.
Stress
- Osmolality 10, 70, 200, 600 and 700 mosm.
- Aminotransferase inhibitor: Aminooxyacetic acid (AOA).
Temperature
Room temperature (about 23 degrees C).
Measurements/observations
Alanine.
[ref. ID; 991]
Test system
The passage of heavy metals from bacteria to protozoa (predator-prey relationship)
Strains
Bacteria (Sphaeotilus sp. from activated-sludge sample) - Protozoa (bacteria-free Paramecium caudatum)
Toxicants
Cr, Mn, Ni, Zn, and Pb.
Test design
Sphaeotilus sp. was grown in 150 ml of SMM (Stokes' Modified Medium) containing no metals, 1 mg/l, and 2 mg/l metals at 25 degrees C for 7 days. Paramecium (100 cells) transferred to Petri dish containing hay infusion, and then metal-containing Sphaeotilus sp. were added. After 10 days the ciliate were washed to remove Sphaeotilus sp.
Measurements/observations
Metals concentration using by Atomic Absorption Spectrophotometer.
[ref. ID; 1072]
Test system
Effect of copper the interaction between Didinium (predator) - Paramecium (prey)
Strains
Escherichia coli, Paramecium caudatum (Ward's Biological Company) and Didinium nasutum (Ward's Biological Company).
Toxicants/concentrations
CuSO4/5H2O, Cu concentrations 0, 150, 300, 600, 900, 1200 and 1500 ug/L.
Test design
pH 6.8-7.0. 24-hr dark photoperiod at 25+/-0.2 degrees C. Paramecium or Didinium were added to a 50-mL glass beaker containing Cerophyl medium.
- Intrinsic rates of natural increase:
- Isocline analysis: Determination of the equilibrium point of the predatory-prey interaction under control (0 ug/L Cu).
- Acute lethality tests:
Measurements/observations
At 4 intervals, over a 24-hr period, a 2.0-mL was taken from each beaker and all animals were counted.
[ref. ID; 1926]
Test system
48-hr acute toxicity test
Strains
PW2, axenically.
Toxicants
Toxin producing blue-green algae: Anabaena flos-aquae (NRC 44), Nostoc linckia (Roth) Bornet & Thuret (NT 69-43), Gloeotrichia echinulata (Smith) Richter (UTEX 1303), Fischerella epiphytica Ghose (NT 69-32) (from the personal collection of Dr. J.T. Wyatt, U.S. Army Environmental Hygiene Agency).
Experimental condition
Peter's osmotic solution. Temperature 25 degrees C.
Measurements/observations
Mortality.
[ref. ID; 4827]
Test system
Effects of O2 on Gravitaxis and Gravikinesis
Strains
Line G3 was bacterized with Aerobactor aerogenes.
Toxicants
Oxygen.
Test design
Two peripheral 1.5 ml-spaces of the recording chamber (outer dimensions: 11x5x1.5 cm) were filled with 1.5% (w/v) agar in experimental solution to incresae the chemical buffering volume of the chamber. A sample of about 200 cells in experimental solution was infused into the central space of 2 ml (depth: 1.6 mm) avoiding the inclusion of air bubbles. Fresh experimental solution calibrated between 5% and 100% air saturation was perfused using hydrostatic pressure difference. The solution passed filtering pads at the entrance and exit limiting cell loss. Perfusion with fresh solution occurred during a period of 35 min following 20 min of recording at defined O2 tension and time after incubation. During each perfusion period, the total fluid volume of the chamber was replaced fresh solution (2 ml). Tests using suspensions of poylstyrene microballs (diameter 10 um) in the experimental solution showed a steady parallel flow (at or below 10 um/sec), which did not agitate the paramecia. Perfusion of the chamber was stopped 5 min prior to a recording period. Experimental temperature 22 degrees C.
Experimental protocol
Horizontal and vertical swimming data were recorded at: 0 hr, 1 hr, 2 hr, 3 hr, 4 hr, 5 hr, 6 hr after cell incubation, and this sequence was repeated for air saturations of 5%, 10%, 20%, 50% and 100% using new cell samples. At 0 hr, after mounting the chamber in horizontal position, cells were continuously recorded during two periods of 2 min, separated by a 1-min nonrecording interval in the dark. The chamber was then slowly reoriented to the vertical position in the course of 1 min, and one more min was allowed for the decay of possible mechanical disturbance effects. Three more 2-min recording periods followed, the latter one separated from the former by a 1-min nonrecording time interval. The same protocol applied to the following recordings (1 hr to 6 hr). During the 20-min recording sequence the perfusion of the chamber was interrupted.
Measurements
Cell number, swimming velocity.
Evaluations
Gravikinesis (um/s).
[ref. ID; 6011]
Test system
Acute toxicity (24-hr LC50)
Strains
From the aeration tank of activated sludge works designated for the treatment of domestic wastes in the district of Reggio Emilia, northern Italy.
Toxicants
Hydrated cadmium chloride, hydrated copper chloride, zinc chloride, mercury chloride.
Test design
Costar tissue culture plates with 24 wells were employed. Two replicates of 12 organisms each were run of each test concentration.
Evaluations
LC50 using the probit method.
[ref. ID; 6111]
Test system
The acute toxicity (24-hr LC50)
Strains
The organisms was obtained, initially, from Stirone Stream (northern Italy), a freshwater environment free from known toxicants and, in particular, from heavy metals. The culture medium was constiuted of boiled rice and wheat grain in 10 ml of filtered natural water. 20+/-1 degrees C, photoperiod of 16:8 hr light:dark.
Toxicant/concentrations
NiCl2/6H2O (0.4, 0.5, 0.75, 1 ppm).
Test design
Tissue culture plates with 24 wells, 3 replicates.
Measurements
Mortality or the survivorship.
Evaluations
LC50 using the probit method.
[ref. ID; 7449]
Test system
Inhibition of phagosome formation
Strains
From the Carolina Biological Supply Co., Burlington, NC.
Inhibitors
Cytochalasin A, B, C, D, E, J and dihydrochalasin B (Sigma Chemical Co., St. Louis, MO).
Test design
P. multimicronucleatum were cultured in the dark with several stems of timothy hay and boiled wheat grains in Carolina spring water. All experiments were carried out using randomly selected, rapidly feeding cultures, and no attempt was made to selet cultures in specific growth phases. Ciliate suspensions were obtained by pipet collection of "piles" of feeding cells that had congregated around wheat seeds. These were diluted into 35x10 mm plastic Petri dishes containing the assay mixture. Room temperature (approx. 24 degrees C).
Measurements/observations
Phagosomes per cell.
Evaluations
Using the two-tailed Mann-Whitney test (Microsoft Statmost 2.5).
[ref. ID; 1268]
Test system
Acute and chronic toxicity
Strains
CCAP 1660/14 from the Culture Collection of Algae and Protozoa (Cumbria, UK).
Toxicants
Selenite (Na2SeO3) and selenate (Na2SeO4).
Test design
All experiments were run in the Hyco medium: CaCl2/H2O 32.2, MgSO4/7H2O 36.9, NaHCO3 25.2, K2HPO4 8.7, NaNO3 85, Na2SiO3/9H2O 28.4, KBr 0.12, NaF 0.4, FeEDTA 4.3, CuSO4/5H2O 0.01, ZnSO4/7H2O 0.022, CoCl2/6H2O 0.01, MnCl2/4H2O 0.18, Na2MoO4/2H2O 0.006, H3BO3 0.62 (in milligrams per liter of distilled water). HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid) buffer was used at 2.5 mM. pH 7.0.
- Toxicity: Final concentrations of 0, 10, 100, and 1,000 ug of selenite or selenate liter-1. Three replicates. Experimental period 120 hr.
- Uptake:
Run 1. Dissolved Na2[75]SeO3 or Na2[75]SeO4 was added directly to mixed bacterial-ciliate cultures in Cerophyll-enriched (0.1%) Hypo medium. Incubation temperature 0 and 24 degrees C.
Run 2. Bacteria were prelabeled with [75]selenium before addition to P. putrinum in unenriched Hyco medium. 25 degrees C in the dark.
Measurements/observations
- Toxicity: Number.
- Uptake: Gamma counting and number.
Evaluations
Toxicity: Population growth rate.
[ref. ID; 6111]
Test system
The acute toxicity (24-hr LC50)
Strains
The organisms was obtained, initially, from Stirone Stream (northern Italy), a freshwater environment free from known toxicants and, in particular, from heavy metals. The culture medium was constituted of boiled rice and wheat grain in 10 ml of filtered natural water. 20+/-1 degrees C, photoperiod of 16:8 hr light:dark.
Toxicant/concentrations
NiCl2/6H2O (1.2, 1.3, 1.4, 1.5 ppm).
Test design
Tissue culture plates with 24 wells, 3 replicates.
Measurements
Mortality or the survivorship.
Evaluations
LC50 using the probit method.
[ref. ID; 7337]
Test system
Capillary tube chemoresponse assay
Strains
Axenic Paramecium tetraurelia Wild type (type 51s) and the trichocyst non-discharge mutant (strain nd6, from B.H. Satir).
Chemorepellents
External GTP (Guanosine 5'-Triphosphate), the oxidant NBT (Nitro Blue Tetrazolium), the secretagogue Alcian Blue, the dye Cibacron Blue, the secretagogue AED (Aminoethyldextran), the oxidant Cytochrome C.
Test design
Samples were concentrated by spinning in a desk top centrifuge at 750 g for 1-2 min and washed by resuspending the cells in a buffer and repelleting the cells. 0.175 ml of washed cells (about 4,000-5,000 cells/ml) were added to a capillary tube and placed on a horizontal surface under a dissecting microscope to equilibrate. The capillary tubes used for the chemorepellent assays were 10.16 cm long with 1.65 mm outer diameter and 1.1 mm inner diameter. A 10 ul volume of either a test solution or a control solution was added to each end. The number of cells at each end of the capillary tube (within 1.0 cm of the end) were then counted, typically after 5 min. Temperature 25 degrees C.
Measurements/observations
Cell number.
Evaluations
EC50
[ref. ID; 7424]
Test system
CABS (Computer-Assisted Backward Swimming) assay (5-min)
Strains
P. tetraurelia trichocyst non-discharge mutant nd-6, behaviorally wild type except for a deffect in trichocyst discharge. Late logarithmic phase, axenically.
SERCA (sarcoplasmic/endoplasmic reticulum Ca2+-dependent ATPase) Pump Inhibitors
2,5-di-tert-butylhydroquinone (BHQ, 10 uM), cyclopiazonic aicd (CPA, 100 uM), thapsigargin (10 uM).
Test design
Room temperature.
Cells were transferred to either resting buffer or resting buffer containing a SERCA inhibitor and allowed to adapt to the new buffer for 15 min. After this time, cells were transferred individually to microscope slide depression wells containing the buffer to which they had adapted, supplemented with 2 uM GTP. Cells were then observed individually for five minutes under a low power dissecting microscope. Each backward swimming event was registered by depressing a key on a computer keyboard for the duration of the backward swimming event.
Measurements/observations
Backward swimming time and electrophysiology (depolarization).