Ecotoxicity of as-synthesised copper nanoparticles on soil bacteria

Release of metallic nanoparticles in soil poses a serious threat to the ecosystem as they can affect the soil properties and impose toxicity on soil microbes that are involved in the biogeochemical cycling. In this work, in vitro ecotoxicity of as-synthesised copper nanoparticles (CuNPs) on Bacillus subtilis (MTCC No. 441) and Pseudomonas fluorescens (MTCC No. 1749), which are commonly present in soil was investigated. Three sets of colloidal CuNPs with identical physical properties were synthesised by chemical reduction method with per batch yield of 0.2, 0.3 and 0.4 gm. Toxicity of CuNPs against these soil bacteria was investigated by MIC (minimum inhibitory concentration), MBC (minimum bactericidal concentration), cytoplasmic leakage and ROS (reactive oxygen species) assay. MIC of CuNPs were in the range of 35-60 µg/ml and 35-55 µg/ml for B. subtilis and P. fluorescens respectively, while their MBC ranged from 40-70 µg/ml and 40-60 µg/ml respectively. MIC and MBC tests reveal that Gram-negative P. fluorescens was more sensitive to CuNPs as compared to Gram positive B. subtilis mainly due to the differences in their cell wall structure and composition. CuNPs with smaller hydrodynamic size (11.34 nm) were highly toxic as revealed by MIC, MBC tests, cytoplasmic leakage and ROS assays, which may be due to the higher active surface area of CuNPs and greater membrane penetration. Leakage of cytoplasmic components and generation of extra-cellular oxidative stress by reactive oxygen species (ROS) causes cell death. The present study realizes in gauging the negative impact of inadvertent release of nanoparticles in the environment, however, in situ experiments to know its overall impact on soil health and soil microflora can help in finding solution to combat ecotoxicity of nanoparticles.