Genotoxic Hazard and Oxidative Stress Induced by Wastewater Irrigated Soil With Special Reference to Pesticides and Heavy Metal Pollution

Enhancement of industrial growth and urbanization, soil contamination is increasingly prominent. Therefore, it’s important to examine possible adverse effects of industrial waste. Samples were found to be polluted with several heavy-metals and pesticides. Gas chromatographic results showed occurrence of high-level of organochlorine and organophosphate pesticides. Genotoxicity of soil extracts was assessed using environmental-risk assessment models. Soil samples were extracted in hexane and dichloromethane solvents and were evaluated for genotoxic potential by prokaryotic (Ames test, plasmid nicking assay and E. coli K-12 DNA repair defective mutants) and eukaryotic (Allium cepa root chromosomal aberration and Vigna radiata seed-germination test) bioassays. Strain TA98 was found most susceptible among soil extracts. Hexane soil extract of wastewater irrigation was determined more mutagenic than DCM samples in terms of mutagenic index, mutagenic potential and induction factor for Ames strains. The damage in DNA repair defective mutants of hexane extracts were found higher compared to DCM extracts at dose of 20 μl/ml of culture. Survival in polA, lexA and recA mutants were 39%, 47% and 55% while treated with hexane extract. Allium cepa test, mitotic index was decreased in dose-dependent way and various kinds of chromosomal aberrations were found. Vigna radiata seeds germination and other parameters were also affected when treated with contaminated soil. Oxidative stress in V. radiata roots were also showed under CLS microscope. Genotoxicity of contaminated soil extract was also confirmed by plasmid nicking test. Our study provides possible explanation for the assessment of potential health and environmental hazards of the industrial region.

Reconstitution of a Staphylococcal Plasmid-Protein Relaxation Complex In Vitro

ABSTRACT The isolation of plasmid-protein relaxation complexes from bacteria is indicative of the plasmid nicking-closing equilibrium in vivo that serves to ready the plasmids for conjugal transfer. In pC221 and pC223, the components required for in vivo site- and strand-specific nicking at oriT are MobC and MobA. In order to investigate the minimal requirements for nicking in the absence of host-encoded factors, the reactions were reconstituted in vitro. Purified MobA and MobC, in the presence of Mg2+ or Mn2+, were found to nick at oriT with a concomitant phosphorylation-resistant modification at the 5′ end of nic. The position of nic is consistent with that determined in vivo. MobA, MobC, and Mg2+ or Mn2+ therefore represent the minimal requirements for nicking activity. Cross-complementation analyses showed that the MobC proteins possess binding specificity for oriT DNA of either plasmid and are able to complement each other in the nicking reaction. Conversely, nicking by the MobA proteins is plasmid specific. This suggests the MobA proteins may encode the nicking specificity determinant.

The acid deoxyribonuclease of neutrophils: a possible participant in apoptosis-associated genome destruction

Human neutrophils are terminally differentiated cells that spontaneously undergo apoptosis in tissue culture. Apoptosis in these cells can be delayed by culture in the presence of granulocyte colony- stimulating factor or other inflammatory mediators. Neutrophils were found to contain an acid endonuclease that appeared to be responsible for the internucleosomal DNA cleavage that accompanies apoptosis. As measured by a plasmid nicking assay, this endonuclease had a molecular weight (M(r)) of 35,000, a pH optimum of 5.5, and a threshold for activity of pH 6.6 to 6.8. It was weakly inhibited by divalent cations (Ca2+, Mg2+, and Zn2+) and more strongly inhibited by aurintricarboxylic acid and N-bromosuccinimide. DNA from neutrophils treated with nigericin in buffers of defined pH displayed nucleosomal ladders whose prominence varied with pH in a manner that paralleled the pH dependence of the plasmid cleavage assays, consistent with internucleosomal DNA cleavage by the acid endonuclease. We have previously shown that neutrophils undergo acidification to a pH value as low as 6.0 during apoptosis; we suggest that this endonuclease may be responsible for the DNA cleavage seen in apoptotic neutrophils.