Huajun Zhen

Potential for inhalation exposure to engineered nanoparticles from nanotechnology-based cosmetic powders.

Background: The market of nanotechnology-based consumer products is rapidly expanding, and the lack of scientific evidence describing the accompanying exposure and health risks stalls the discussion regarding its guidance and regulation. Objectives: We investigated the potential for human contact and inhalation exposure to nanomaterials when using nanotechnology-based cosmetic powders and compare them with analogous products not marketed as nanotechnology based. Methods: We characterized the products using transmission electron microscopy (TEM) and laser diffraction spectroscopy and found nanoparticles in five of six tested products. TEM photomicrographs showed highly agglomerated states of nanoparticles in the products. We realistically simulated the use of cosmetic powders by applying them to the face of a human mannequin head while simultaneously sampling the released airborne particles through the ports installed in the mannequin’s nostrils. Results: We found that a user would be exposed to nanomaterial predominantly through nanoparticle-containing agglomerates larger than the 1–100-nm aerosol fraction. Conclusions: Predominant deposition of nanomaterial(s) will occur in the tracheobronchial and head airways—not in the alveolar region as would be expected based on the size of primary nanoparticles. This could potentially lead to different health effects than expected based on the current understanding of nanoparticle behavior and toxicology studies for the alveolar region.

Release of Free DNA by Membrane-Impaired Bacterial Aerosols Due to Aerosolization and Air Sampling

ABSTRACT We report here that stress experienced by bacteria due to aerosolization and air sampling can result in severe membrane impairment, leading to the release of DNA as free molecules. Escherichia coli and Bacillus atrophaeus bacteria were aerosolized and then either collected directly into liquid or collected using other collection media and then transferred into liquid. The amount of DNA released was quantified as the cell membrane damage index (ID ), i.e., the number of 16S rRNA gene copies in the supernatant liquid relative to the total number in the bioaerosol sample. During aerosolization by a Collison nebulizer, the ID of E. coli and B. atrophaeus in the nebulizer suspension gradually increased during 60 min of continuous aerosolization. We found that the ID of bacteria during aerosolization was statistically significantly affected by the material of the Collison jar (glass > polycarbonate; P < 0.001) and by the bacterial species (E. coli > B. atrophaeus; P < 0.001). When E. coli was collected for 5 min by filtration, impaction, and impingement, its ID values were within the following ranges: 0.051 to 0.085, 0.16 to 0.37, and 0.068 to 0.23, respectively; when it was collected by electrostatic precipitation, the ID values (0.011 to 0.034) were significantly lower (P < 0.05) than those with other sampling methods. Air samples collected inside an equine facility for 2 h by filtration and impingement exhibited ID values in the range of 0.30 to 0.54. The data indicate that the amount of cell damage during bioaerosol sampling and the resulting release of DNA can be substantial and that this should be taken into account when analyzing bioaerosol samples.

Development of a dual-internal-reference technique to improve accuracy when determining bacterial 16S rRNA:16S rRNA gene ratio with application to Escherichia coli liquid and aerosol samples.

Accurate enumeration of rRNA content in microbial cells, e.g. by using the 16S rRNA:16S rRNA gene ratio, is critical to properly understand its relationship to microbial activities. However, few studies have considered possible methodological artifacts that may contribute to the variability of rRNA analysis results. In this study, a technique utilizing genomic DNA and 16S rRNA from an exogenous species (Pseudomonas fluorescens) as dual internal references was developed to improve accuracy when determining the 16S rRNA:16S rRNA gene ratio of a target organism, Escherichia coli. This technique was able to adequately control the variability in sample processing and analysis procedures due to nucleic acid (DNA and RNA) losses, inefficient reverse transcription of RNA, and inefficient PCR amplification. The measured 16S rRNA:16S rRNA gene ratio of E. coli increased by 2-3 fold when E. coli 16S rRNA gene and 16S rRNA quantities were normalized to the sample-specific fractional recoveries of reference (P. fluorescens) 16S rRNA gene and 16S rRNA, respectively. In addition, the intra-sample variation of this ratio, represented by coefficients of variation from replicate samples, decreased significantly after normalization. This technique was applied to investigate the temporal variation of 16S rRNA:16S rRNA gene ratio of E. coli during its non-steady-state growth in a complex liquid medium, and to E. coli aerosols when exposed to particle-free air after their collection on a filter. The 16S rRNA:16S rRNA gene ratio of E. coli increased significantly during its early exponential phase of growth; when E. coli aerosols were exposed to extended filtration stress after sample collection, the ratio also increased. In contrast, no significant temporal trend in E. coli 16S rRNA:16S rRNA gene ratio was observed when the determined ratios were not normalized based on the recoveries of dual references. The developed technique could be widely applied in studies of relationship between cellular rRNA abundance and bacterial activity.

Analysis of airborne microbial communities using 16S ribosomal RNA: Potential bias due to air sampling stress

A limited number of studies have been conducted to analyze ribosomal RNA (rRNA, present in the ribosome) in bioaerosol samples to identify currently or potentially active airborne microbes, although its genomic counterpart, the rRNA gene (on the chromosome) has been frequently targeted for airborne microbial community analysis. A knowledge gap still exists regarding whether the bioaerosol rRNA abundances are affected by the bioaerosol collection process. We investigated the effect of air sampling stress on the measurement and characterization of 16S rRNA for bioaerosols in the laboratory and field experiments using quantitative polymerase chain reaction (qPCR) and high-throughput sequencing techniques. In a laboratory study, known quantities of freshly grown Escherichia coli cells were spiked onto the filter of a Button Aerosol Sampler and into liquids of BioSampler and SpinCon air samplers and then exposed to sampling stress when the samplers were operated for 2h. We found that the recovered cellular 16S rRNA abundance as determined by qPCR was dependent on sampler type. Further, two devices (Button Aerosol Sampler and BioSampler) that exhibited markedly different efficiency in preserving 16S rRNA were employed in an outdoor environment to collect bioaerosols simultaneously on eight days in two different seasons. The abundance of 16S rRNA in the outdoor air sample (1.3×106-4.9×107copies/m3) was about two orders of magnitude higher than that of 16S rRNA gene (6.9×103-1.5×105copies/m3). The 16S rRNA sequences revealed a different bacterial community compared with 16S rRNA gene-based results across all samples, and this difference depended on the sampling device. In addition, a number of bacterial taxa exhibited higher abundance in the 16S rRNA gene sequences than in 16S rRNA sequences, which suggests the potential activities of certain microbes in airborne phase. Overall, this study highlights the importance of sampling device selection when analyzing RNA in bioaerosols.

Diastereoisomer-specific biotransformation of hexabromocyclododecanes by a mixed culture containing Dehalococcoides mccartyi strain 195

Hexabromocyclododecane (HBCD) stereoisomers may exhibit substantial differences in physicochemical, biological, and toxicological properties. However, there remains a lack of knowledge about stereoisomer-specific toxicity, metabolism, and environmental fate of HBCD. In this study, the biotransformation of (±)α-, (±)β-, and (±)γ-HBCD contained in technical HBCD by a mixed culture containing the organohalide-respiring bacterium Dehalococcoides mccartyi strain 195 was investigated. Results showed that the mixed culture was able to efficiently biotransform the technical HBCD mixture, with 75% of the initial HBCD (∼12 μM) in the growth medium being removed within 42 days. Based on the metabolites analysis, HBCD might be sequentially debrominated via dibromo elimination reaction to form tetrabromocyclododecene, dibromocyclododecadiene, and 1,5,9-cyclododecatriene. The biotransformation of the technical HBCD was likely diastereoisomer-specific. The transformation rates of α-, β-, and γ-HBCD were in the following order: α-HBCD > β-HBCD > γ-HBCD. The enantiomer fractions of (±)α-, (±)β-, and (±)γ-HBCD were maintained at about 0.5 during the 28 days of incubation, indicating a lack of enantioselective biotransformation of these diastereoisomers. Additionally, the amendment of another halogenated substrate tetrachloroethene (PCE), which supports the growth of strain 195, had a negligible impact on the transformation patterns of HBCD diastereoisomers and enantiomers. This study provided new insights into the stereoisomer-specific transformation patterns of HBCD by anaerobic microbes and has important implications for microbial remediation of anoxic environments contaminated by HBCD using the mixed culture containing Dehalococcoides.