Cyto-Genotoxic and Behavioral Effects of Flubendiamide in Allium cepa Root Cells, Drosophila melanogaster and Molecular Docking Studies.

Flubendiamide (FLB) is an insecticide that is commonly employed to control pests on a variety of vegetables and fruits, with low toxicity for non-target organisms. However, due to its widespread use, the environmental risks and food safety have become major concerns. In this study, the toxicity potential of FLB was studied in the model organisms, Allium cepa and Drosophila melanogaster. The cyto-genotoxic effects of FLB on the root growth, mitotic index (MI), chromosomal aberrations (CAs) and deoxyribonucleic acid (DNA) damage in A. cepa root meristematic cells were investigated using the root growth inhibition Allium test and Comet assays. FLB caused CAs in the form of disturbed ana-telophase, chromosome laggards, stickiness, anaphase-bridge and polyploidy depending on the concentration and the exposure time. The toxicity and genotoxicity of FLB at various doses (0.001, 0.01, 0.1 and 1 mM) on D. melanogaster were investigated from the point of view of larval weight and movement, pupal formation success, pupal position, emergence success and DNA damage, respectively. FLB exposure led to a significant reduction of the locomotor activity at the highest concentration. While DNA damage increased significantly in the FLB-treated onions depending on the concentration and time, DNA damage in the FLB-treated D. melanogaster significantly increased only at the highest dose compared to that which occurred in the control group. Moreover, to provide a mechanistic insight into the genotoxic and locomotion-disrupting effects of FLB, molecular docking simulations of this pesticide were performed against the DNA and diamondback moth (DBM) ryanodine receptor (RyR) Repeat34 domain. The docking studies revealed that FLB binds strongly to a DNA region that is rich in cytosine-guanine-adenine bases (C-G-A) in the minor groove, and it displayed a remarkable binding affinity against the DBM RyR Repeat34 domain.

Ames and random amplified polymorphic DNA tests for the validation of the mutagenic and/or genotoxic potential of the drinking water disinfection by-products chloroform and bromoform.

Chloroform and Bromoform are two abundant trihalomethanes found in Algerian drinking water. The investigation of the mutagenic hazard of these disinfection by-products was studied by Ames test as prokaryotic bioassay to show their mutagenic effects. For this, Salmonella typhimurium TA98 and TA100 strains were employed. Both chloroform and bromoform showed a direct mutagenic effect since the number of revertant colonies gradually increase in dose-dependent manner with all concentrations tested with the two bacterial strains and these were both in the absence and presence of S9 metabolic activation. The genotoxic hazard was also studied by random amplified polymorphic DNA test on the root cells of Allium cepa as eukaryotic bioassay. DNA extracted from the roots of the onion were incubated at different concentrations of chloroform and bromoform and then amplified by polymerase chain reaction. This was based on demonstrating a major effect of disappearance of bands compared to roots incubated in the negative control (distilled water). The results showed that these two compounds affected genomic DNA by breaks although by mutations.

Genotoxic evaluation of Halfenprox using the human peripheral lymphocyte micronucleus assay and the Ames test.

The genotoxicity and mutagenicity of Halfenprox, a synthetic pyrethroid insecticide and acaricide, was assessed using two standard genotoxicity assays of the Salmonella typhimurium mutagenicity assay (Ames test) and in vitro micronucleus (MN) assay in human peripheral lymphocytes. In the Ames test, Salmonella strains TA98 and TA100 were treated with or without S9 fraction. The doses of Halfenprox were 6.25, 12.5, 25, 50, and 100 µg/plate and test materials were dissolved in DMSO. The concentrations of Halfenprox did not show mutagenic activity on both strains with and without S9 fraction. The MN assay was used to investigate the genotoxic effects of Halfenprox in human peripheral lymphocytes treated with 250, 500, 750, and 1000 µg/ml concentrations of Halfenprox for 24 and 48 h, and at 1000 µg/ml the concentration was significantly increased and the MN formation was compared with the negative control for both treatment periods. In addition, a significant decrease of the nuclear devision index (NDI) values at the higher concentrations of Halfenprox and at both treatment periods was observed.

Evaluation of cytotoxic and genotoxic effects of Benodanil by using Allium and Micronucleus assays.

The aim of this study was to evaluate the potential cytotoxic effects of Benodanil fungicide by employing both mitotic index (MI) and mitotic phases on the root meristem cells of Allium cepa and genotoxic effects by using in vitro micronucleus assay (MN) in human peripheral blood lymphocyte. In the Allium root growth inhibition test, the EC50 value was first determined as 25 ppm. Then, 2 × EC50 value (50 ppm), EC50 value (25 ppm), and 1/2 × EC50 value (12.5 ppm) were tested with different treatment periods (24, 48, and 72 h). Both negative and positive controls were also used in parallel experiments. We obtained that mitotic index and prophase index decreased when compared with the control in all concentrations. In the micronucleus assay, lymphocytes were treated with various concentrations (250, 500, 750, and 1000 µg/ml) of Benodanil for 24 and 48 h. The results showed that Benodanil did not induce MN frequency in all concentrations of both treatment periods. Additionally, it was determined that this pesticide decreased nuclear division index (NDI) significantly. It was concluded that Benodanil has a cytotoxic effects depending on decreasing of MI and NDI.

Determination of mutagenicity and genotoxicity of indium tin oxide nanoparticles using the Ames test and micronucleus assay.

In this study, the mutagenicity and genotoxicity of indium tin oxide (ITO) nanomaterial were assessed using two standard genotoxicity assays, the Salmonella reverse mutation assay (Ames test) and the in vitro micronucleus (MN) assay. Seven different concentrations (12.5, 25, 50, 75, 100, 125, and 150 µg/plate) of this nanomaterial were tested using the Ames test on the TA98 and TA100 strains in the presence and absence of the S9 mixture. At all the concentrations tested, this substance did not significantly increase the number of revertant colonies compared with the control with or without S9 mixture. The genotoxic effects of ITO were investigated in human peripheral lymphocytes treated with 125, 250, 500, and 750 µg/ml concentrations of this substance for 24- and 48-h treatment periods using an MN test. Nuclear division index (NDI) was also calculated in order to determine the cytotoxicity of ITO. It was determined that ITO increased MN frequency in the 750 µg/ml concentration in 24- and 48-h treatments. In addition, ITO dose dependently decreased the NDI significantly for two treatment periods.

Assessment of cytotoxic and genotoxic potential of pyracarbolid by Allium test and micronucleus assay.

The present study evaluates the cytotoxic and genotoxic potential of pyracarbolid using both micronuleus (MN) assay, in human lymphocytes, and Allium cepa assay, in the root meristem cells. In Allium test, EC50 value was determined in order to selecting the test concentrations for the assay and the root tips were treated with 25 ppm (EC50/2), 50 ppm (EC50) and 100 ppm (EC50 × 2) concentrations of pyracarbolid. One percent of dimethyl sulphoxide (DMSO) and methyl methane sulfonate (MMS) were used as negative and positive controls, respectively. In the micronucleus assay, the cultures were treated with four concentrations (250, 500, 750 and 1000 µg/ml) of pyracarbolid for 24 and 48 h, negative and positive controls were also used in the experiment parallely. The results showed that mitotic index (MI) significantly reduced with increasing the pyracarbolid concentration at each exposure time. It was also obtained that prophase and metaphase index decreased significantly in all concentration at each exposure time. Anaphase index decreased as well and results were found to be statistically significant, except 24 h. A significant increase was observed in MN frequency in all concentrations and both treatment periods when compared with the controls. Pyracarbolid also caused a significant reduction in the cytokinesis block proliferation index (CBPI) in all concentration and both exposure time.

Detection of genotoxicity and mutagenicity of chlorthiophos using micronucleus, chromosome aberration, sister chromatid exchange, and Ames tests.

Potential mutagenic and genotoxic effects of Chlorthiophos, an organophosphate pesticide, were evaluated using four standard assays. Five different concentrations of the pesticide were tested by an Ames test using Salmonella typhimurium strains TA97, TA98, TA100, and TA102, with and without S9 metabolic activation. No concentrations of Chlorthiophos showed mutagenic activity on the TA97, TA100, and TA102 strains, with and without S9 fraction, but were all mutagenic to the TA98 strain without S9. Sister chromatid exchange (SCE), chromosome aberration (CA), and micronucleus (MN) tests were used to investigate the genotoxic effects of Chlorthiophos in human peripheral lymphocytes treated with 25, 50, 100, and 200 µg/mL concentrations of Chlorthiophos for 24 and 48 h. The nuclear division index (NDI), replication index (RI), and mitotic index (MI) were also calculated to determine the cytotoxicity of Chlorthiophos. No increase in SCE frequency was seen for any treatment period or concentration, but Chlorthiophos at 200 µg/mL increased the frequency of CAs. Increases in MN formation were only observed at Chlorthiophos concentrations of 200 µg/mL following 24 and 48 h treatments. Chlorthiophos treatment reduced the MI and NDI significantly, but had no effect on the RI.

Synthesis of Moringa oleifera coated silver-containing nanocomposites of a new methacrylate polymer having pendant fluoroarylketone by hydrothermal technique and investigation of thermal, optical, dielectric and biological properties.

Firstly, silver nanoparticles were synthesized by green synthesis method from Moringa oleifera extract. Nanocomposites containing newly synthesized methacrylate polymer, poly 2-(4-fluorophenyl)-2-oxoethyl-2-methylprop-2-enoate (PFPAMA) and Ag nanoparticles from M. oleifera in different mass ratios (1, 3, and 5 wt%) were synthesized using the hydrothermal method. The morphological and structural properties of the materials have been examined by SEM, FTIR, UV, TGA, and XRD techniques. The activation energies (Ea) related to thermal decomposition of the nanocomposites were estimated by the Flynn-Wall-Ozawa and Kissinger-Akahira-Sunose methods by using non-isothermal TGA experiments. The thermal stability, glass transition temperature (Tg), and the thermal decomposition activation energy (Ea) values of nanocomposites were increased by increasing the Ag nanoparticles amount on the composite. The dielectric constant (ε'), the dielectric loss factor (ε″) and ac conductivity of neat PFPAMA and nanocomposites were also measured for the frequency range of 100 Hz to 2 kHz at 25 °C. It was seen that the frequency dependence of the dielectric constant and dielectric loss factor decreased with increasing frequency. The biological activities of nanocomposites against gram-positive (Staphylococcus aureus), gram-negative (Escherichia coli) bacteria and Candida krusei yeast were also tested. The antibacterial effect increased against both bacterial species as the amount of Ag nanoparticles from M. oleifera in the nanocomposites increased. In addition, the wound healing properties of nanocomposites were investigated by the scratch wound test.

Evaluation of germination, root growth and cytological effects of wastewater of sugar factory (Afyonkarahisar) using Hordeum vulgare bioassays.

In this study, the effect of Afyonkarahisar Sugar Factory's discharge water on germination percentage, root growth and mitotic divisions of the root tip cells of Hordeum vulgare L. were investigated. Six concentrations of wastewater and ranging from 10(0), 10( -1), 10( -2), 10( -3), 10( -4), 10( -5), were applied for 6, 12, 18 and 24 h, respectively. It was observed that the treatments reduced the germination percentages of H. vulgare grains and inhibited the root growth as well as reduced mitotic index compared to the control group at all concentrations. It was also observed that the increase of the concentrations of wastewater decreased the cell division, and several mitotic anomalies such as c-mitosis, lagging chromosomes, multipolar anaphases and chromosome bridges increased.

Mutagenic and genotoxic effects of Anilofos with micronucleus, chromosome aberrations, sister chromatid exchanges and Ames test.

We aimed to evaluate the mutagenic effect of Anilofos, organophosphate pesticide, by using Ames/Salmonella/microsome test. Its cytotoxic and genotoxic effects were also determined by chromosome aberration (CA), sister chromatid exchange (SCE) and micronucleus (MN) test in human peripheral blood lymphocytes. In the Ames test, five different concentrations of Anilofos were examined on TA97, TA98, TA100 and TA102 strains in the absence and presence of S9 fraction. According to the results all concentrations of this pesticide have not shown any mutagenic activity on TA97, TA100 and TA102 strains in the absence and presence of S9 fraction. But, 10, 100 and 1000 µg/plate concentrations of Anilofos were determined to be mutagenic on TA98 strain without S9 fraction. Lymphocytes were treated with various concentrations (25, 50, 100 and 200 µg/ml) of Anilofos for 24 and 48 h. The results of the assays showed that Anilofos did not induce SCE frequency, replication index and MN formation at all concentrations for both treatment periods. Anilofos significantly increased CA frequency at 100 and 200 µg/ml concentrations at 24 h treatment periods and at 50, 100 and 200 µg/ml concentrations in 48 h treatment periods. Additionally, it was determined that this pesticide decreased mitotic index and nuclear division index significantly. It was concluded that Anilofos has genotoxic and cytotoxic effects in human peripheral lymphocytes.

Mutagenic and cytotoxic activities of benfuracarb insecticide.

Benfuracarb is a carbamate insecticide used to control insect pests in vegetables and it has anti-acetylcholinesterase activity lower than other carbamates. Cytotoxic effects of benfuracarb were evaluated by using root growth inhibition (EC50), mitotic index (MI), and mitotic phase determinations on the root meristem cells of Allium cepa and mutagenic effects were determined in Salmonella typhymurium Ames test by TA98 and TA100 strains with and without metabolic activation. In Allium test, 1 % DMSO was used as negative control group and 10 ppm MMS was used as positive control group. 75 ppm concentration of benfuracarb was found as EC50. In MI and mitotic phases determination study, 37.5, 75 and 150 ppm doses of benfuracarb were used. Dose-dependent cytotoxic activity was found by root growth inhibition and MI studies. It was identified that mitotic inhibition activity of benfuracarb was higher than 10 ppm MMS. In Ames test, mutagenic activity was not observed and over 200 µg/plate of benfuracarb was determined as cytotoxic to S. typhymurium strains. Benfuracarb can be called as "mitotic inhibitor" but not called as mutagen.

Potential cytotoxic effect of Anilofos by using Allium cepa assay.

Cytogenetic effects of Anilofos which was widely used in agriculture, was evaluated in Allium cepa root meristematic cells. In the Allium root growth inhibition test EC50 value was determined 50 ppm and 1/2× EC50 (25 ppm), EC50 (50 ppm) and 2 × EC50 (100 ppm) concentrations of Anilofos were applied to onion roots. A negative and positive control were used in the experiment in parallel. According to results mitotic index decreased with increasing the Anilofos concentrations in all application groups and each exposure time, while disturbed anaphase-telophase, choromosome laggard(s), stickiness and anaphase bridge(s) were observed. In anaphase-telophase cells, c-metaphase, disturbed nucleus and binuclear cells were observed in other anomalies. The results were also analyzed statistically by using Dunnett t test (2-tailed) and all concentrations of Anilofos were found significant.

Cytotoxic and genotoxic effects of dioxacarb by human peripheral blood lymphocytes CAs and Allium test.

Dioxacarb (Elecron, Famid) is a phenyl methylcarbamate insecticide and in vitro cytotoxic and genotoxic effects of this pesticide on human peripheral blood lymphocytes and Allium root meristematic cells were investigated by chromosomal aberrations (CAs) and Allium test. Human lymphocytes were treated with 62.5, 125, 250 and 500 ppm doses of dioxacarb for CAs. CA/cell, abnormal cell % and mitotic index % (MI %) data were obtained from these concentrations in 24 and 48 h treatment periods. Dioxacarb did not increase the CA/cell frequency significantly, so this insecticide was not identified as genotoxic. But it was found cytotoxic especially at 250 and 500 ppm concentrations because of the reduced the MI % and increased the abnormal cell %. In Allium test, 25 ppm (EC50/2), 50 ppm (EC50) and 100 ppm (EC50 × 2) concentrations were used for root growth inhibition (EC50 determination) and Allium mitotic index (MI) determination tests. The used concentrations of dioxacarb induced dose-dependent inhibition of MI and root growth on root meristems. Mitotic inhibition of dioxacarb was found significantly higher than for the positive control. These Allium results indicated the high cytotoxicity of dioxacarb. The present study is the first research on cytotoxicity and genotoxicity of dioxacarb by human lymphocyte CAs and Allium test.

Micronucleus assay in human lymphocytes after exposure to alloxydim sodium herbicide in vitro.

This study evaluates the cytotoxic and genotoxic potential of alloxydim sodium using micronucleus (MN) assay, in human peripheral lymphocytes. MN assay was used to investigate the genotoxic effects of alloxydim sodium in human peripheral lymphocytes treated with 250, 500, 750, 1,000 µg/ml concentrations of alloxydim sodium for 24 and 48 h. Solvent, negative and positive controls were also used in the experiments in parallel. The obtained results were evaluated in statistical analyses by using Dunnett-t test (two sided) and p < 0.05 was accepted as significant. Alloxydim sodium significantly increased the MN formation compared with the negative control, at both 750 and 1,000 µg/ml concentrations and treatment periods. We also evaluated the nuclear division index (NDI) for cytotoxicity of this pesticide in the experiment, and finally observed a significant decrease of the NDI values at all concentrations of alloxydim sodium and at both treatment periods.

Testing of the mutagenicity and genotoxicity of metolcarb by using both Ames/Salmonella and Allium test.

Mutagenic and genotoxic effects of metolcarb were investigated by both bacterial reverse mutation assay in Salmonella typhimurium TA97, TA98, TA100 and TA102 strains with or without metabolic activation system (S9) and Allium cepa root meristematic cells, respectively. Metolcarb was dissolved in DMSO in Ames/Salmonella test system. 0.1, 1 and 10 microg/plate doses of metolcarb were found to be mutagenic S. typhimurium TA98 without S9. In Allium root growth inhibition test, EC50 value was determined 200 ppm and 0.5xEC50, EC50 and 2xEC50 concentrations of metolcarb were introduced to onion tuber roots and distilled water used as a negative control. Mitotic index (MI), increased in all concentrations compared to control at each exposure time. While disturbed anaphase-telophase, chromosome laggards, stickiness and bridges were observed in anaphase-telophase cells, pro-metaphase, C-mitosis, polyploidy, binuclear cells and disturbed nucleus were observed in other cells. The results were also analyzed statistically by using SPSS for Windows, Mann-Whitney test and Duncan's multiple range tests were performed respectively.