Rapid evolution of silver nanoparticle resistance in Escherichia coli.

Joseph L Graves,Adero Campbell,Mehrdad Tajkarimi,Scott H Harrison,Quincy Cunningham,Herve Nonga,Jeffrey E Barrick

doi:10.3389/fgene.2015.00042

Abstract

The recent exponential increase in the use of engineered nanoparticles (eNPs) means both greater intentional and unintentional exposure of eNPs to microbes. Intentional use includes the use of eNPs as biocides. Unintentional exposure results from the fact that eNPs are included in a variety of commercial products (paints, sunscreens, cosmetics). Many of these eNPs are composed of heavy metals or metal oxides such as silver, gold, zinc, titanium dioxide, and zinc oxide. It is thought that since metallic/metallic oxide NPs impact so many aspects of bacterial physiology that it will difficult for bacteria to evolve resistance to them. This study utilized laboratory experimental evolution to evolve silver nanoparticle (AgNP) resistance in the bacterium Escherichia coli (K-12 MG1655), a bacterium that does not harbor any known silver resistance elements. After 225 generations of exposure to the AgNP environment, the treatment populations demonstrated greater fitness vs. control strains as measured by optical density (OD) and colony forming units (CFU) in the presence of varying concentrations of 10 nm citrate-coated silver nanoparticles (AgNP) or silver nitrate (AgNO3). Genomic analysis shows that changes associated with AgNP resistance were already accumulating within the treatment populations by generation 100, and by generation 200 three mutations had swept to high frequency in the AgNP resistance stocks. This study indicates that despite previous claims to the contrary bacteria can easily evolve resistance to AgNPs, and this occurs by relatively simple genomic changes. These results indicate that care should be taken with regards to the use of eNPs as biocides as well as with regards to unintentional exposure of microbial communities to eNPs in waste products.

Highlights

Experimental evolution is routinely used to study the causes and consequences of natural selection
This study indicates that despite previous claims to the contrary bacteria can evolve resistance to AgNPs, and this occurs by relatively simple genomic changes
Population growth of the control (C1—C5) and treatment (T1— T5) populations were measured at generation 162 and 250 in the presence of 10 nm citrate-coated AgNPs by cocktailing the control and treatment replicates

Summary

Introduction

Experimental evolution is routinely used to study the causes and consequences of natural selection. Experimental evolution can predict how phenotypes and their underlying genomic architecture change in response to new environments. This approach may be particular fertile for bacterial evolution. While these organisms have persisted and diversified for billions of years, they often must respond to novel environments. Increasing amounts of metallic/metallic oxide nanoparticles are being used in consumer products. Nano-titanium oxide (nanoTiO2) is produced on a large scale for applications in paints, cosmetics, sunscreens, photo-catalysts and solar cells, as well as water purification devices. The predicted concentration of nanoTiO2 in European waters for 2009 was 20 ng/L (Gottschalk et al, 2009). Values for nano-silver were calculated at 6.6 μg/kg/year, 526 μg/kg/year, 0.088 μg/L/year, 16.40 μg/L/year, 1.29 mg/kg/year, and 153 μg/kg/year for American soil, sludge, surface water, Sewage Treatment Plant (STP) effluent, STP sludge, and sediment respectively

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