Bio-safety assessment of nanozeolites of varying size and doses on soil beneficial microorganisms

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Aim: To evaluate the toxic effects of nano-zeolites on soil beneficial microorganisms. Methodology: This study involved dry milling of zeolites at varying parameters to obtain dry-milled zeolites of four size regimes such as 10-100 nm, 200-400 nm, 500-1000 nm and 1000-2000 nm that were tested at incremental concentrations such as 100 ppm, 1000 ppm and 2000 ppm along with control on various soil beneficial microorganisms. Three replications were done for each treatment with factorial completely randomized design. The poison food technique, growth curve assessment followed by cytotoxicity and genotoxicity studies on all four bacterial genera treated with zeolites of varying sizes and doses were undertaken. The effects of zeolite on average linear growth rate (ALGR) of biocontrol agent, Trichoderma viride were also studied. Results: The effect of zeolites tested on four bacterial genera viz. Azotobacter chroococcum, Rhizobium leguminosarum, Bacillus megaterium and Pseudomonas fluorescens using poison food technique and growth curve revealed that zeolites regardless of size or concentration had positively influenced the growth dynamics of all four bacteria tested. The effect of zeolite on average linear growth rate (ALGR) of Trichoderma viride also indicated that incremental dose of zeolite had a positive effect. Lactose dehydrogenase revealed that 2000 ppm nano-zeolite exhibited cytotoxic effects on soil beneficial micro-organisms tested. On the other hand, comet assay demonstrated no quantifiable DNA damage in nano-zeolite treated cells in comparison to control cultures. Interpretation: This study unequivocally demonstrated that zeolites of size greater than 200-400 nm, irrespective of doses even up to 2000 ppm are quite safe for soil beneficial microbes.