Method for toxicity test of titanium dioxide nanoparticles in ciliate protozoan Tetrahymena.

Titanium dioxide (TiO2) nanoparticles have a high surface-to-mass ratio and rapidly aggregate in water causing great difficulties for toxicity test exposed to aquatic organisms or other cell lines. This study uses a cell viability kit for routine toxicity test of TiO2 as well as other nanoparticles which accumulate in the aquatic environment. Tetrahymena immediately endocytoses TiO2 nanoparticles and stores them in food vacuoles until the particles undergo exocytosis as larger aggregates. However, during the process of endocytosis and exocytosis, TiO2 particles interfere with cell growth and consequently induce acute toxicity. It exerted high cell growth inhibition at 20 h incubation and induces significant cytotoxic effects. Surprisingly, the effect of TiO2 decreases at 40 h incubation, due to the recovery of cell growth and reduction of the cytotoxicity of the particles.

5-Fluorouracil accumulation in green microalgae and its biogenetic transfer into ciliate protozoan.

The study has demonstrated that anticancer drug 5-fluorouracil causes acute toxicity and interferes with the growth of green microalgae, Scenedesmus vacuolatus. It accumulates in microalgae biomass with bioaccumulation factor of 1.84 × 10(4) and further integrates into the DNA and RNA of microalgae. In addition, the labelled microalgae genome is transferred into protozoan Tetrahymena pyriformis on feeding and is retained in the food vacuoles of predator organisms. This biotransfer of labelled 5-fluorouracil via genomic material was evaluated using radioactivity in Tetrahymena cell pellets though radioactivity did not detect anticancer drug in the genome of the predator organism.

Isolation, Identification and Genetic Characterization of Antibiotic Resistant Escherichia coli from Frozen Chicken Meat Obtained from Supermarkets at Dhaka City in Bangladesh.

Antimicrobials have been used to improve animal welfare, food security, and food safety that promote the emergence, selection, and dissemination of antimicrobial-resistant (AMR) bacteria. In this study, 50 E. coli were isolated from frozen chicken meat samples in Dhaka city. Antibiotic sensitivity patterns were assessed through the disk diffusion method and finally screened for the presence of antimicrobial resistance genes (ARG) using the polymerase chain reaction (PCR). Among the 160 samples, the prevalence of E. coli was observed in fifty samples (31.25%). All of these isolates were found resistant to at least one antimicrobial agent, and 52.0% of the isolates were resistant against 4-7 different antimicrobials. High resistance was shown to tetracycline (66.0%), followed by resistance to erythromycin (42.0%), ampicillin and streptomycin (38.0%), and sulfonamide (28.0%). In addition, the most prevalent ARGs were tet(A) (66.0%), ereA (64.0%), tet(B) (60.0%), aadA1 and sulI (56.0%), blaCITM (48.0%) and blaSHV (40.0%). About 90.0% of isolates were multidrug resistant. This study reveals for the first time the current situation of E. coli AMR in broilers, which is helpful for the clinical control of disease as well as for the development of policies and guidelines to reduce AMR in broilers production in Bangladesh.

ISTA13-catecholamine toxicity and metabolism in the ciliated protozoan, Tetrahymena pyriformis.

A high throughput culture methodology of unicellular eukaryote Tetrahymena pyriformis, strain GL were used for the determination of catecholamines toxicity and their metabolism. Catecholamines exhibited acute toxicity to Tetrahymena cells where dopamine and L-DOPA showed higher toxic potential of EC(10) (0.39 and 0.63 mg/L, respectively) and EC(20) (1.1 and 1.0 mg/L, respectively). All the testing catecholamines were highly degradable in the PPY-medium due to the oxidizing environment during incubation. They were also naturally synthesized and released by Tetrahymena cells into the culture medium and increasingly accumulated with time where as noradrenalin demonstrated significant results. Cells were exposed with physiological concentration (0.12 mg/L) and one higher concentration (8.0 mg/L) of catecholamines, resulting noradrenalin depletion and in vivo generation of a metabolite in response to dopamine with higher concentration treatment. This dopamine metabolite was relatively nonpolar compared with the catecholamines and was eluted later from the reverse phase C-18 column.

Identification of dopamine receptor in Tetrahymena thermophila by fluorescent ligands.

Two types dopamine receptor present in the cell membrane of vertebrates. But in this study D1 receptor was identified in the invertebrate ciliates protozoan, Tetrahymena thermophila by use of fluorescent ligands. D1 specific agonist SKF-38393 binds specifically to Tetrahymena. The specific binding of SKF-38393 was encountered by equimolar addition of D1 antagonist thus showed no binding of ligands. In addition, it was also proved that the D1 specific agonist did not cross bind with the D2 type receptor due to the equimolar addition of D2 selective antagonist spiperone. Interestingly this study also showed that the dopamine receptor present in the endoplasmic reticulum and endosomes of Tetrahymena as well as cell membrane which was revealed by laser scanning microscope. Therefore, this evidence supports the existence of a D1 receptor in the ciliate protozoan.

Impact of fluorotelomer alcohols (FTOH) on the molecular and macroscopic phenotype of Tetrahymena thermophila.

BACKGROUND, AIM, AND SCOPE: As possible precursors of PFOA, fluorotelomer alcohols are a class of highly fluorinated and volatile compounds. Although they are widespread in the environment, little toxicity data is available. The present study focused on testing the population growth impairment potential of FTOH. Moreover, certain efforts were made to find the possible effect mechanism of these compounds. MATERIALS AND METHODS: The growth inhibition test was made both in an open system (96-well microplates) and in a closed system (closed flasks). In addition, cells were stained with acridine orange and observed under fluorescence microscopy at 488 nm. Furthermore, direct membrane damage was checked by measuring LDH leakage. RESULTS: For 8:2 FTOH and 10:2 FTOH, no growth inhibition was found in either of the systems. In contrast, 4:2 FTOH interfered with population growth in the closed system (EC(50) = 276.1 mg/L), whereas, 6:2 FTOH had an influence on population growth both in the closed system (EC(50) = 64.3 mg/L) and in the open system. Macronucleus destruction was observed with 6:2 FTOH. No direct membrane damage was detectable. DISCUSSION: With a closed system, 4:2 and 6:2 FTOH were found to be capable of impairing population growth. However, this potential was to a certain extent underestimated. With the help of the air-water distribution coefficient, the real EC(50) was estimated within the interval [203.2, 276.1] mg/L and [14.7, 64.3] mg/L for 4:2 and 6:2 FTOH, respectively. Some evidence, such as the absence of direct membrane or macronucleus damage, indicate that certain FTOH could likely cause apoptosis. But the exact effect mechanism could not be determined on the basis of the present results. CONCLUSIONS: Comparing the results from the two test systems, tests in a closed system are more reliable for testing these volatile compounds with Tetrahymena thermophila than in an open system. RECOMMENDATIONS AND PERSPECTIVES: The present study has highlighted several future research directions. For ecotoxicological risk assessment of FTOH, their distribution and environmental fate should be determined. To understand the effect mechanism, more tests could be conducted to test whether apoptosis is caused. Finally, in order to standardize test procedure in a closed system, more compounds should be investigated in the closed system to clarify the sensitivity of the test procedures.

Growth inhibition and biodegradation of catecholamines in the ciliated protozoan Tetrahymena pyriformis.

A 96-well plate culture methodology for the unicellular eukaryote Tetrahymena pyriformis, strain GL was used for the determination of toxicity and metabolism of catecholamines. Catecholamines exhibited moderate acute toxicity to Tetrahymena cells where dopamine and L-DOPA showed higher toxic potential at EC(10) (0.39 ppm and 0.63 ppm, respectively) and EC(20) (1.1 ppm and 1.0 ppm respectively) after 48 h exposure. All tested catecholamines were highly degradable in the PPY-medium due to the oxidizing environment during incubation. Also the catecholamines were naturally synthesized and released by Tetrahymena cells into the culture medium and increasingly accumulated with time where noradrenalin exhibited the highest degree of accumulation. However, the exogenous exposure of catecholamines to the cells caused the depletion of natural noradrenalin synthesis even with the addition of very low physiological concentration (0.12 ppm). Dopamine caused the higher effect on inhibiting noradrenalin synthesis. Treatment with a higher concentration (8.0 ppm) of dopamine in 96-well plates caused strong excitation of the cells and ascertained a new metabolite in vivo while the other representative catecholamines were not responsible for the production of this metabolite. This dopamine metabolite is relatively non-polar as compared to noradrenalin, adrenaline and dopamine and eluting later through the reverse phase C-18 column.