Linking NE1545 gene expression with cell volume changes in Nitrosomonas europaea cells exposed to aromatic hydrocarbons

Nitrosomonas europaea, a model ammonia oxidizing bacterium, was exposed to a wide variety of aromatic hydrocarbons in 3 h batch assays. The expression of NE1545, a phenol sentinel gene involved in fatty acid metabolism, was monitored via quantitative real-time polymerase chain reaction (qRT-PCR) and a Coulter Counter technique was used to monitor changes in cell volume. Decreases in cell volume and NE1545 gene expression correlated strongly with exposure to aromatic hydrocarbons that possessed a single polar group substitution (e.g. phenol and aniline). Aromatic hydrocarbons that contain no polar group substitutions (e.g. toluene) or multiple polar group substitutions (e.g. p-hydroquinone) caused negligible changes in NE1545 expression and cell volume. The oxidation of aromatic hydrocarbons by N. europaea from configurations without a single polar group to one with two polar groups (e.g. p-cresol oxidized to 4-hydroxybenzyl alcohol) and from configurations with no polar groups to one with a single polar group (e.g. ethylbenzene oxidized to 4-ethylphenol) greatly influenced NE1545 gene expression and observed changes in cell volume. Nitrification inhibition in N. europaea by the aromatic hydrocarbons was found to be completely reversible; however, the decreases in cell volume were not reversible suggesting a physical change in cell membrane composition. Ammonia monooxygenase blocking studies showed that the chemical exposure that was responsible for the cell volume decrease and up-regulation in gene expression and not the observed inhibition. N. europaea is the first bacterium shown to experience significant changes in cell volume when exposed to μM concentrations of aromatic hydrocarbons, three orders of magnitude lower than previous studies with other bacteria.     

Influence of liberated silver from silver nanoparticles on nitrification inhibition of Nitrosomonas europaea

The ecotoxicity of silver nanoparticles (Ag-NPs) to wastewater biota, including ammonia oxidizing bacteria (AOB), is gaining increasing interest as the number of products containing Ag-NPs continues to rise exponentially and they are expected to accumulate in wastewater treatment plants. This research demonstrated that the addition order of Ag-NP and the media constituents had a profound influence on the stability of the Ag-NP suspension and the corresponding repeatability of results and sensitivity of Nitrosomonas europaea. N. europaea, a model AOB, was found to be extremely sensitive to ionic silver (Ag⁺) and two sizes of Ag-NPs (20 and 80 nm). Ag⁺ exposures resulted in the highest level of toxicity with smaller Ag-NPs (20 nm) being more toxic than larger Ag-NPs (80 nm). The increased sensitivity of N. europaea to smaller Ag-NPs was caused by their higher rates of dissolved silver (dAg) release, via dissolution, due to a greater surface area to volume ratio. dAg was shown to be responsible for the vast majority of the observed Ag-NP toxicity, as determined by abiotic Ag-NP dissolution tests. For the sizes of Ag-NP studied (20 and 80 nm), there appears to be a negligible nanoparticle-specific toxicity. This was further supported by similarities in inhibition mechanisms between Ag⁺ and Ag-NP, with both causing decreases in AMO activity and destabilization of the outer-membrane of N. europaea. Finally, equal concentrations of total silver were found to be tightly associated to both Ag⁺ and Ag-NP-exposed cells despite Ag-NP concentrations being five times greater, by mass, than Ag⁺ concentrations.