Heterogenic response of prokaryotes toward silver nanoparticles and ions is facilitated by phenotypes and attachment of silver aggregates to cell surfaces.

Abstract

Tons of anthropogenic silver nanoparticles (AgNPs) are assumed to be released into the environment due to their use in many consumer products. AgNPs are known to be toxic toward microorganisms and thus may harm their specific functions in ecosystems. Here we explore the impact of AgNPs on functioning of single cells in microbial populations at doses typically found in anthropogenic environments. The response of single cells to AgNPs was analyzed by flow cytometry and using the fluorescent dyes propidium iodide and DiBAC4 (3) as markers for cell membrane disintegration and depolarization, respectively. The effects of 10-nm and 30-nm AgNPs on three bacterial species (Mycobacterium frederiksbergense, Pseudomonas putida, and Escherichia coli) showed that the populations split into affected cells and others not showing any malfunction, with varying abundances depending on strains and cell growth states. Further, the dissolution of AgNPs measured with 3 KDa ultrafiltration and inductively coupled plasma-mass-spectrometry to distinguish particle-related effects from toxicity of dissolved Ag revealed that Ag ions were the principal toxicant. AgNP aggregate formation was followed by dynamic light scattering and the aggregates' attachment to cell surfaces was visualized by transmission electron microscopy and scanning electron microscopy-energy dispersive X-ray spectroscopy. An increased AgNP-affected cell fraction relative to the Ag ion impact was identified. The study shows that individual cells in a population cope differently with AgNP induced stress by evolving heterogeneous phenotypes. The response is linked to cell death and cell energy depletion depending on cell type and cell growth states. The attachment of AgNP aggregates to cell surfaces seems to amplify the heterogeneous response. © 2017 International Society for Advancement of Cytometry.