Cytotoxicity of Potassium Salts of (+)- and (-) Usnic Acid for Paramecium caudatum.

We analyzed cytotoxicity of water-soluble potassium salts of (+)- and (-) usnic acid (UA) for ciliates P. caudatum. The median lethal concentrations for (+)- and (-) enantiomers did not significantly differ and were 7.5±0.5 and 6.7±0.4, respectively. In a concentration of 8 µM, (+)-UA and (-)-UA salts increased the content of TBA-reactive products, which indicates the formation of oxidative stress under the action of high UA concentrations. In the presence of (+)-UA and (-)-UA salts in a concentration range from 2 to 8 µM, the number of food vacuoles in ciliates decreased, which attested to a decrease in phagocytosis activity. The concentrations of UA enantiomers >0.5 µM affected macronucleus morphology (shape and size). The cytotoxic activity of (+)-UA and (-)-UA salts against P. caudatum did not differ.

Individual and mixture toxicity of carbofuran and diuron to the protozoan Paramecium caudatum and the cladoceran Ceriodaphnia silvestrii.

The toxicity of the insecticide carbofuran and herbicide diuron (individually and in mixture) to the invertebrates Paramecium caudatum and Ceriodaphnia silvestrii was evaluated. Acute and chronic toxicity tests were carried out with the diuron and carbofuran active ingredients and their commercial products, Diuron Nortox® 500 SC and Furadan® 350 SC, respectively. Individual toxicity tests showed that C. silvestrii was more sensitive to both carbofuran and diuron than P. caudatum. In single exposures, both pesticides caused adverse effects to C. silvestrii in environmentally relevant concentrations (48 h EC50 = 0.001 mg L-1 and 8 d LOEC = 0.00038 mg L-1 for formulated carbofuran; 8 d LOEC < 0.05 mg L-1 for formulated diuron). For P. caudatum, carbofuran and diuron in single exposures were only slightly toxic (24 h IC50 = 5.1 mg L-1 and 6.9 mg L-1 for formulated carbofuran and diuron, respectively). Acute and chronic exposures to diuron and carbofuran mixtures caused significant deviations of the toxicity predicted by the Concentration Addition and Independent Action reference models for both test species. For the protozoan P. caudatum, a dose-dependent deviation was verified for mortality, with synergism caused mainly by carbofuran and antagonism caused mainly by diuron. For protozoan population growth, however, an antagonistic deviation was observed when the active ingredient mixtures were tested. In the case of C. silvestrii, antagonism at low concentrations and synergism at high concentrations were revealed after acute exposure to active ingredient mixtures, whereas for reproduction an antagonistic deviation was found. Commercial formulation mixtures presented significantly higher toxicity than the active ingredient mixtures. Our results showed that carbofuran and diuron interact and cause different toxic responses than those predicted by the individually tested compounds. Their mixture toxicity should therefore be considered in risk assessments as these pesticides are likely to be present simultaneously in edge-of-field waterbodies.

Combined effects of food resources and exposure to ammonium nitrogen on population growth performance in the bacterivorous ciliate Paramecium caudatum.

Ciliated protozoa (ciliates) play vital roles in biological wastewater-treatment processes, however, combined effects of abiotic and biotic factors as well as the importance of species-specificity of bacterial food organisms on population growth dynamics remain poorly understood, which are hampering the management and optimization of biological wastewater treatment processes. This study investigated the effects of food resources and ammonium nitrogen (NH4+) exposure, both independently and in combination, on the population growth of the bacterivorous ciliate Paramecium caudatum. Results showed that, when fed with two different bacterial food organisms, population growth performance of P. caudatum differed significantly and increased with the addition of protozoa pellet medium. When exposed to NH4+ population growth declined and metabolic enzyme activities were altered. The negative effects of NH4+ on population growth could be weakened by supplementing the food resource with protozoa pellet media. In brief, it was confirmed that the existence of interactive effect of food resources and ammonium nitrogen, as well as the importance of species-specificity of bacterial food organisms on the population growth performance of ciliates. These findings might lead to the development of a valuable strategy for improving the performance of biological wastewater-treatment processes.

Investigation of the effect of biologically active threo-Ds-isocitric acid on oxidative stress in Paramecium caudatum.

The effect of biologically active form (threo-Ds-) of isocitric acid (ICA) on oxidative stress was studied using the infusorian Paramecium caudatum stressed by hydrogen peroxide and salts of some heavy metals (Cu, Pb, Zn, and Cd). ICA at concentrations between 0.5 and 10 mM favorably influenced the infusorian cells with oxidative stress induced by the toxicants studied. The maximal antioxidant effect of ICA was observed at its concentration 10 mM irrespective of the toxicant used (either H2O2 or heavy metal ions). ICA was found to be a more active antioxidant than ascorbic acid. Biologically active pharmaceutically pure threo-Ds-ICA was produced through cultivation of the yeast Yarrowia lipolytica and isolated from the culture liquid in the form of crystalline monopotassium salt with a purity of 99.9%.

Changes in the ecological properties of organic wastes during their biological treatment.

Organic wastes, such as the organic fractions of municipal solid waste (OFMSW) or sewage sludge (SS), have become a serious environmental problem in Russia as well as in other countries. The use of these wastes as soil amendments allows their negative impact on the environment to be minimized. However, before these wastes can be used, they need to be treated appropriately in order to decrease their level of hazard. In this study, composting of raw SS, OFMSW, a mixture of these two wastes (OFMSW+SS) at a ratio 1:2 as well as the anaerobically digested variants of these wastes (SSd, OFMSWd and OFMSWd+SSd) mixed with oiled sawdust was performed. Composting was conducted in the containers containing 20kg of the wastes. The results of three elutriate bioassays (with water flea Daphnia magna, infusoria Paramecium caudatum and radish plant, Raphanus sativus) and one contact bioassay (with oat plant Avena sativa) were used to indirectly estimate changes in the hazardous properties of the biological treatments. Besides, Corg, Ntot content and pH were analyzed in the process of composting. Within the study stability tests to determine maturation process completion were not carried out. It was revealed, that in the process of anaerobic pretreatment for 15days, the toxicity increased by a mean of 1.3-, 1.9- and 1.1-fold for OFMSW, SS and OFMSW+SS, respectively. During composting, the toxicity level of these pretreated samples decreased more rapidly as compared with those which were not pretreated. As a result, the toxicity levels of the elutriates from the final composts made of pretreated wastes OFMSW, SS and OFMSW+SS were three-, two- and 17-fold lower for D. magna and 15-, 21- and 12-fold lower for P. caudatum. As follows from phytotoxicity estimations, composts from digested substrates became mature on the 60th day and had a stimulation effect on the plants after the 90th day of incubation. For the composts prepared from raw substrates, a significantly longer period was needed for maturation. On the basis of ecotoxicity changes of the wastes treated, it can be concluded that anaerobic pretreatment of the municipal solid wastes is an effective acceleration tool for further composting and that waste mixtures can be treated more efficient as compared with raw wastes.

Effects of diuron and carbofuran pesticides in their pure and commercial forms on Paramecium caudatum: The use of protozoan in ecotoxicology.

Toxic effects of diuron and carbofuran on Paramecium caudatum were evaluated. Acute and chronic tests were conducted with diuron and carbofuran active ingredients and their commercial formulations, Diuron Nortox(®) 500 SC and Furadan(®) 350 SC, respectively. The sensitivity range of P. caudatum to reference substance sodium chloride was established. A preliminary risk assessment of diuron and carbofuran for Brazilian water bodies was performed. The tests indicated that toxicity of pure diuron and its commercial formulation was similar, while the commercial product carbofuran was more toxic than its pure form. In acute tests, readings were carried out at 2, 3, 4 and 6 h and showed an increase of mortality with increasing exposure time. The sensitivity of P. caudatum to NaCl ranged from 3.31 to 4.44 g L(-1), averaging 3.88 g L(-1). For diuron, the 6 h LC50 was 64.6 ± 3.3 mg L(-1) for its pure form and 62.4 ± 2.5 mg L(-1) for its commercial formulation. Carbofuran active ingredient was less toxic than that of diuron, presenting a 6 h LC50 of 142.0 ± 2.4 mg L(-1) for its pure form and 70.4 ± 2.2 mg L(-1) for its commercial product. Chronic tests showed that these pesticides cause significant decrease on population growth, generation number and biomass of P. caudatum. The 24 h IC50 was 7.10 ± 0.58 mg L(-1) for pure diuron, 6.78 ± 0.92 mg L(-1) for commercial diuron, 22.95 ± 3.57 mg L(-1) for pure carbofuran and 4.98 ± 0.62 mg L(-1) for commercial carbofuran. Preliminary risk assessment indicated that diuron and carbofuran present potential ecological risks for Brazilian water bodies. P. caudatum was a suitable and sensitive test organism to evaluate diuron and carbofuran toxicity to freshwater protozooplankton and, taking into account the relevant role of protozoans in aquatic environments, we strongly recommend its inclusion in ecotoxicological studies.

Transgenic Bacillus thuringiensis (Bt) rice is safer to aquatic ecosystems than its non-transgenic counterpart.

Rice lines genetically modified with the crystal toxin genes from Bacillus thuringiensis (Bt) have experienced rapid development, with biosafety certificates for two Bt rice lines issued in 2009. There has still been no commercial release of these lines yet due to public concerns about human health and environmental risks. Some studies confirmed that Bt rice was as safe as conventional rice to non-target organisms when pesticides were not applied, however, pesticides are still required in Bt rice to control non-lepidopteran pests. In this study, we assessed the environmental effects of two Bt rice lines expressing either the cry1Ab/1Ac or cry2A genes, respectively, by using zooplanktons as indicator species under normal field management practices using pesticides when required. In the whole rice growing season, non-Bt rice was sprayed 5 times while Bt rice was sprayed 2 times, which ensured both rice achieved a normal yield. Field investigations showed that rice type (Bt and non-Bt) significantly influenced zooplankton abundance and diversity, which were up to 95% and 80% lower in non-Bt rice fields than Bt rice fields. Laboratory rearing showed that water from non-Bt rice fields was significantly less suitable for the survival and reproduction of Daphnia magna and Paramecium caudatum in comparison with water from Bt rice fields. Higher pesticide residues were detected in the water from non-Bt than Bt rice fields, accounting for the bad performance of zooplankton in non-Bt field water. Our results demonstrate that Bt rice is safer to aquatic ecosystems than non-Bt rice, and its commercialization will be beneficial for biodiversity restoration in rice-based ecosystems.

[Hygienic assessment of waste of soda production].

The object of investigations was soda industry waste. Slimes are formed at slimes storage which occupy considerable areas and are considered to be the source of permanent impact on the hydrosphere objects. Slimes storage placement within settlement boundaries and water protection zone of large watercourses leads to the deterioration of sanitary, hygienic and environmental situation and to the rising of risks to health of communities. Waste processing with getting new materials on the base of soda industry waste with wide application is seems to be one of the way for problem solving. It is essential to take into account sanitary and hygienic characteristics of slimes within justifying possible directions of its use. Thus, researches concerning assessment of physical, chemical and toxicological waste characteristics are considered to be actual. The aim of researches is to examine physical, chemical and toxicological characteristics of soda production slimes for justifying directions of its use including delivery of new materials respondent to the all regulatory sanitary and hygienic requirements. Experimental investigations of assessment physical, chemical and toxicological characteristics of slimes were carried out according to standard methods. Within assessment of toxicological slimes characteristics the following test-objects were used: Ceriodaphnia affinis, Paramecium caudatum. As a result of investigations watered slime samples were determined to be referred to the 4th hazard level (low-hazard) waste; samples with preliminary mechanical dehydration are referred to the 5th hazard level (practically nonhazardous) waste for environment. These are correspond to the 3rd and 4th hazard level according to sanitary regulations, respectively.

Antimicrobial activity of metal oxide nanoparticles supported onto natural clinoptilolite.

The antimicrobial activity of Cu(2)O, ZnO and NiO nanoparticles supported onto natural clinoptilolite was investigated in the secondary effluent under dark conditions. After 24h of contact the Cu(2)O and ZnO nanoparticles reduced the numbers of viable bacterial cells of Escherichia coli and Staphylococcus aureus in pure culture for four to six orders of magnitude and showed consistent 100% of antibacterial activity against native E. coli after 1h of contact during 48 exposures. The antibacterial activity of NiO nanoparticles was less efficient. The Cu(2)O and NiO nanoparticles showed 100% of antiprotozoan activity against Paramecium caudatum and Euplotes affinis after 1h of contact, while ZnO nanoparticles were less efficient. The morphology and crystallinity of the nanoparticles were not affected by microorganisms. The metal oxide nanoparticles could find a novel application in the disinfection of secondary effluent and removal of pathogenic microorganisms in the tertiary stage of wastewater treatment.

Design, engineering and utility of biotic games.

Games are a significant and defining part of human culture, and their utility beyond pure entertainment has been demonstrated with so-called 'serious games'. Biotechnology--despite its recent advancements--has had no impact on gaming yet. Here we propose the concept of 'biotic games', i.e., games that operate on biological processes. Utilizing a variety of biological processes we designed and tested a collection of games: 'Enlightenment', 'Ciliaball', 'PAC-mecium', 'Microbash', 'Biotic Pinball', 'POND PONG', 'PolymerRace', and 'The Prisoner's Smellemma'. We found that biotic games exhibit unique features compared to existing game modalities, such as utilizing biological noise, providing a real-life experience rather than virtual reality, and integrating the chemical senses into play. Analogous to video games, biotic games could have significant conceptual and cost-reducing effects on biotechnology and eventually healthcare; enable volunteers to participate in crowd-sourcing to support medical research; and educate society at large to support personal medical decisions and the public discourse on bio-related issues.

Perfluorooctanoic acid (PFOA) but not perfluorooctane sulfonate (PFOS) showed DNA damage in comet assay on Paramecium caudatum.

Persistent perfluorinated organic compounds such as perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) are distributed widely in the global environment including wildlife and human. In this study, we investigated the genotoxicity of PFOS and PFOA using the novel in vivo comet assay developed for Paramecium caudatum. For the comet assay, large nuclei squeezed out of the paramecia with 0.25 M sucrose containing 0.6% Triton X-100 were embedded in a layer of agarose gel placed over the slide glass. N-methyl-N´-nitro-N-nitrosoguanidine (MNNG) and 2-aminoanthracene (2-AA) were successfully used for positive controls. Productions of 8-hydroxydeoxyguanosine (8-OH-dG) and intracellular reactive oxygen species (ROS) were also measured in paramecia. PFOS did not cause DNA damage on any conditions examined. On the other hand, 12 and 24 hr exposure to PFOA (100 µM) increased DNA migration in electrophoresis condition at pH 13, but not at pH 12.1, suggesting that the DNA damage may be alkali labile site (such as apurinic/apyrimidinic (AP) site). Exposure of paramecia to 100 µM PFOA for 1, 3 and 24 hr and to 10 µM PFOA for 24 hr significantly increased intracellular ROS. Under the same condition, however, 8-OH-dG level was not affected by PFOA. The PFOA-induced DNA damage was not abolished by the application of 100 µM GSH which completely inhibited the increase of intracellular ROS. In conclusion, the PFOA-induced in vivo DNA damage was first shown in paramecia, and the DNA damage might not be directly attributable to increase in intracellular ROS.

Cytotoxicity assessment of monocrotophos in Paramecium caudatum and Oxytricha fallax.

Experiments were conducted to evaluate the toxic effects ofmonocrotophos in ciliate models Paramecium caudatum and Oxytricha fallax. In acute toxicity studies higherconcentrations of monocrotophos caused marked increase in mobility of cells exhibiting rocking movements within two mins of exposure but were decreased after 30 mins. LC50 value by mortality curve for 3 hr acute toxicity test of Oxytricha fallax and Paramecium caudatum was found 307.744 +/- 33.27 mg l(-1) and 332.284 +/- 57.52 mg l(-1) respectively. Oxytricha fallax was found sensitive than Paramecium caudatum to monocrotophos. In acute exposure cells showed deformities such as swelling, oval shaped deformity and in higher concentrations shortening of longitudinal axis with blackening of cytoplasm occurred. The length of paramecia was reduced prominently. Similarly enlargement of contractile vacuole and stress egestion of food vacuoles was also observed. The morphological studies showed the changes in shape, size, colour and width of Paramecia and Oxytricha. Frequencies of macronuclear aberrations were significant showing deformities such as rod shaped, elongation, fragmentation, diffusion and total absence of nucleus and were concentration dependent. The data provided in the present study on interaction of pesticides with nuclear structure can be of immense value because most of these pesticides have been reported to have carcinogenic, mutagenic and teratogenic properties.

[Biological effects of electromagnetic radiation of extremely high frequencies combined with physiologically active compounds].

The study of the action of the electromagnetic radiation (EMR) of low intensity (10 microW/cm2) in the range of frequencies 120-170 GHz at the test-reaction of Infusoria Paramecium caudatum was carried out. The resonant character of the effects was established. The EMR action at 156.6 and 161.3 GHz caused the increase of infusorians mobility, the action at frequencies 151.8, 155.7, 167.1 GHz caused the mobility reduction. Isolated and combined with EMR effects of nicotine (10(-4)-10(-15) mol/l) and antimicrobial drug metronidazole (10(-5), 10(-8), 10(-9) mol/l) were investigated. The radiation at the frequency 167.1 GHz was shown to reduce the effect of nicotine (10(-9) mol/l) and to enhance the effect of metronidazole (10(-9) mol/l). This phenomenon may be explained by different effects of the substances in low concentration at the water hydrogen bonds net structure.

[Motor activity of infusoria: fundamental and applied aspects].

The article considers morpho-functional organization of the cilia, locomotor organelle of the infusoria, and demonstrates the complicity of locomotor behavior of these protista. The problem of control of locomotion of infusoria is whole organism in discussed; and conclusion is drawn that system of control of movements could be multilevel and include receptor, afferent, central, efferent and effector units. In this context the macronucleus, could act as a central integrator and coordinator of the locomotor behavior being closely connected with periphery by dynamic elements of cytoskeleton. The eradication of infusoria parasitizing in humans and animals by interrupting of locomotion of the protista is also discussed.

Effects of perfluorinated amphiphiles on backward swimming in Paramecium caudatum.

PFOS and PFOA are ubiquitous contaminants in the environment. We investigated the effects of fluorochemicals on calcium currents in Paramecium caudatum using its behavioral changes. Negatively charged amphiphiles prolonged backward swimming (BWS) of Paramecium. PFOS significantly prolonged BWS, while PFOA was less potent (EC(50): 29.8+/-4.1 and 424.1+/-124.0microM, respectively). The BWS prolongation was blocked by cadmium, indicating that the cellular calcium conductance had been modified. The positively charged amphiphile FOSAPrTMA shortened BWS (EC(50): 19.1+/-17.3). Nonionic amphiphiles did not affect BWS. The longer-chain perfluorinated carboxylates PFNA and PFDA were more potent than PFOA (EC(50): 98.7+/-20.1 and 60.4+/-10.1microM, respectively). However, 1,8-perfluorooctanedioic acid and 1,10-perfluorodecanedioic acid did not prolong BWS. The critical micelle concentration (CMC) and BWS prolongation for negatively charged amphiphiles showed a clear correlation (r(2)=0.8008, p<0.001). In summary, several perfluorochemicals and PFOS and PFOA had similar effects in Paramecium, while chain length, CMC, and electric charge were major determinants of BWS duration.

Membrane potential responses of paramecium caudatum to external Na+.

The membrane potential responses of Paramecium caudatum to Na+ ions were examined to understand the mechanisms underlying the sensation of external inorganic ions in the ciliate by comparing the responses of the wild type and the behavioral mutant. Wild-type cells exhibited initial continuous backward swimming followed by repeated transient backward swimming in the Na+-containing test solution. A wild-type cell impaled by a microelectrode produced initial action potentials and a sustained depolarization to an application of the test solution. The prolonged depolarization, the depolarizing afterpotential, took place subsequently after stimulation. The ciliary reversal of the cell was closely associated with the depolarizing responses. When the application of the test solution was prolonged, the wild-type cell produced sustained depolarization overlapped by repeated transient depolarization. A behavioral mutant defective in the Ca2+ channel, CNR (caudatum non reversal), produced a sustained depolarization but no action potential or depolarizing afterpotential. The mutant cell responded to prolonged stimulation with sustained depolarization overlapped by transient depolarization, although it did not show backward swimming. The results suggest that Paramecium shows at least two kinds of membrane potential responses to Na+ ions: a depolarizing afterpotential mediating initial backward swimming and repeated transient depolarization responsible for the repeated transient backward swimming.