Research ArticleEARTH SCIENCES

Effects of silver nanoparticles on nitrification and associated nitrous oxide production in aquatic environments

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Science Advances  02 Aug 2017:
Vol. 3, no. 8, e1603229
DOI: 10.1126/sciadv.1603229
  • Fig. 1 Percentage reduction of nitrification rate in AgNP or Ag+ treatments compared to the no-silver control (incubation time = 12 hours; n = 3).

    EC10 and EC50 represent the concentrations that produced a 10 or 50% reduction in nitrification rate relative to the control, respectively. Nonlinear fitted curves (ExpDec1) and equations are given (P < 0.01).

  • Fig. 2 Effect of AgNPs or Ag+ on N2O emission during nitrification (incubation time = 12 hours).

    Data show the percentage changes of N2O emission in the AgNP or Ag+ treatments compared to the no-silver control (n = 3). Nonlinear fitted curves (Gauss) and equations are given (P < 0.01).

  • Fig. 3 Identification of key N2O production pathways.

    (A) SP values. (B) Contribution of NH2OH oxidation pathway to N2O emission. (C) Isotopomer ratios at the central site of N2O. (D) Isotopomer ratios at the end site of N2O. Horizontal lines indicate the median, five-point stars show the mean, asterisks indicate outlier, the boxes give the 25th and 75th percentiles, and whiskers show range from the 5th to 95th percentile. Control group represents the incubation without silver. The 10 nm, 30 nm, 100 nm, and Ag+ in the “N2O stimulation” area represent the incubations with AgNPs (100 μg liter−1, 10 nm; 500 μg liter−1, 30 nm; and 1000 μg liter−1, 100 nm) and Ag+ (5 μg liter−1), wherein 43.0, 84.1, 121.5, and 73.9% of N2O emission enhancement were detected, respectively. The 10 nm, 30 nm, 100 nm, and Ag+ in the “N2O inhibition” area represent the incubations with AgNPs (2000 μg liter−1, 10 nm; 3000 μg liter−1, 30 nm; and 3000 μg liter−1, 100 nm) and Ag+ (500 μg liter−1), wherein 89.9, 48.3, 19.0, and 59.3% of N2O emission inhibition were detected, respectively. The incubation time was 12 hours.

  • Fig. 4 Response of nitrifying communities to AgNP exposure.

    (A) Schematic model depicting the effects of AgNPs on the expression of gene families involved in nitrification. N2O can be produced through NO2 reduction (the bold pink arrows) or incomplete NH2OH oxidation (the bold blue arrows). Upward green arrows indicate that the gene expressions were up-regulated when exposed to AgNPs, the downward red arrows indicate that the gene expressions were down-regulated, and “N” denotes that gene expression was not affected by AgNP exposure. CusA, Cu(I)/Ag(I) efflux system membrane protein cusA; CusB, Cu(I)/Ag(I) efflux system periplasmic protein cusB; amo, ammonia monooxygenase; hao, hydroxylamine oxidase; Cyt554, cytochrome c554; mCyt552, cytochrome cm552; nir, nitrite reductase (NO-forming) nirK; nor, nitric oxide reductase norQ; nxr, nitrite oxidoreductase; Cyt551, cytochrome c551; Cyt552, cytochrome c552; Cyt553, cytochrome c553; Q, ubiquinone; QH2, ubiquinol. The roman numbers refer to the enzyme complex I (NADH-ubiquinone reductase), complex III (ubiquinol-cytochrome c reductase), complex IV (cytochrome c oxidase), and complex V (F-type ATPase) in the respiratory chain (related gene expression regulations by AgNPs are shown in fig. S12). Colored proteins were detected in the cDNA libraries, whereas those in dark gray were not identified but are included in the model of electron transport for reference (74). Dotted blue arrows show the movement of electrons, and white arrows show movement of protons. The membrane was broken by dotted line, as nitrite oxidation did not often occur in the same organism with ammonia oxidation, with the exception of recently discovered comammox Nitrospira (55, 56). (B) Fold change (FC) of transcripts encoding proteins involved in heavy metal stress response, oxidative stress release, and the nitrogen transformation process of nitrifying organisms when exposed to 30-nm AgNP (500 μg liter−1) for 12 hours. FC in relative gene expression was calculated by comparing AgNP-treated samples to the no-silver control. Gene expression levels were calculated on the basis of FPKM. (C) Contribution of different pathways to N2O emission in the no-silver control and the 30-nm AgNP (500 μg liter−1) treatment. (D) TEM image of the no-silver control at 12 hours. (E) TEM image of the 30-nm AgNP (500 μg liter−1) treatment at 12 hours. No apparent physical damage to the cell surface was observed.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/3/8/e1603229/DC1

    Supplementary Materials and Methods

    fig. S1. Nitrate (NO3) concentrations in the glass vials during the 30-hour incubation period.

    fig. S2. Percentage reduction in nitrification rate in AgNP or Ag+ treatments compared to no-silver control during the 30-hour incubation period.

    fig. S3. Pearson correlation between the dissolved Ag+ concentrations released from AgNPs and the percentage reduction in nitrification rate at the end of the 12-hour exposure period.

    fig. S4. Percentage effects of PVP coating on nitrification activity and N2O production compared to the control group (incubation time = 12 hours, n = 3).

    fig. S5. N2O concentrations in the headspace of the glass vials during the 30-hour incubation period.

    fig. S6. Effects of AgNPs and Ag+ on N2O emission during nitrification in the 30-hour incubation period.

    fig. S7. Effects of the dissolved Ag+ released from AgNPs on N2O emission during nitrification (incubation time = 12 hours).

    fig. S8. DO concentration at the end of the incubation period (incubation time = 12 hours).

    fig. S9. Bioreactor performance.

    fig. S10. Community compositions of the nitrifying bioreactor during the incubation period and the original sediment samples collected from the intertidal flat of the Yangtze Estuary based on 16S rRNA gene sequencing.

    fig. S11. Community compositions of the nitrifying organisms in the no-silver control and the AgNP (500 μg liter−1 of 30 nm AgNPs) treatment at the end of the 12-hour incubation, based on the metatranscriptome sequencing.

    fig. S12. FC of transcripts encoding the enzymes complex I (NADH-ubiquinone reductase), complex III (ubiquinol-cytochrome c reductase), complex IV (cytochrome c oxidase), and complex V (F-type ATPase) in the respiratory chain of the nitrifying organisms under 30-nm AgNP (500 μg liter−1) exposure for 12 hours.

    fig. S13. Neighbor-joining phylogenetic tree of ammonia monooxygenase based on protein sequences.

    fig. S14. TEM imaging of AgNPs used in this study.

    table S1. Dissolved Ag+ concentration released from AgNPs after a 12-hour exposure period in the nitrification inhibition experiment.

    table S2. Sequencing read statistics of the metatranscriptomic libraries.

    table S3. Expression of genes encoding proteins involved in nitrogen transformation, heavy metal stress response, and oxidative stress release of nitrifying organisms in the no-silver control and 30-nm AgNP (500 μg liter−1) treatment based on qPCR method (incubation time = 12 hours).

    table S4. Primers and qPCR protocols used in this study.

    References (7583)

  • Supplementary Materials

    This PDF file includes:

    • Supplementary Materials and Methods
    • fig. S1. Nitrate (NO3) concentrations in the glass vials during the 30-hour incubation period.
    • fig. S2. Percentage reduction in nitrification rate in AgNP or Ag+ treatments compared to no-silver control during the 30-hour incubation period.
    • fig. S3. Pearson correlation between the dissolved Ag+ concentrations released from AgNPs and the percentage reduction in nitrification rate at the end of the 12-hour exposure period.
    • fig. S4. Percentage effects of PVP coating on nitrification activity and N2O production compared to the control group (incubation time = 12 hours, n = 3).
    • fig. S5. N2O concentrations in the headspace of the glass vials during the 30-hour incubation period.
    • fig. S6. Effects of AgNPs and Ag+ on N2O emission during nitrification in the 30-hour incubation period.
    • fig. S7. Effects of the dissolved Ag+ released from AgNPs on N2O emission during nitrification (incubation time = 12 hours).
    • fig. S8. DO concentration at the end of the incubation period (incubation time = 12 hours).
    • fig. S9. Bioreactor performance.
    • fig. S10. Community compositions of the nitrifying bioreactor during the incubation period and the original sediment samples collected from the intertidal flat of the Yangtze Estuary based on 16S rRNA gene sequencing.
    • fig. S11. Community compositions of the nitrifying organisms in the no-silver control and the AgNP (500 μg liter−1 of 30 nm AgNPs) treatment at the end of the 12-hour incubation, based on the metatranscriptome sequencing.
    • fig. S12. FC of transcripts encoding the enzymes complex I (NADH-ubiquinone reductase), complex III (ubiquinol-cytochrome c reductase), complex IV (cytochrome c oxidase), and complex V (F-type ATPase) in the respiratory chain of the nitrifying organisms under 30-nm AgNP (500 μg liter−1) exposure for 12 hours.
    • fig. S13. Neighbor-joining phylogenetic tree of ammonia monooxygenase based on protein sequences.
    • fig. S14. TEM imaging of AgNPs used in this study.
    • table S1. Dissolved Ag+ concentration released from AgNPs after a 12-hour exposure period in the nitrification inhibition experiment.
    • table S2. Sequencing read statistics of the metatranscriptomic libraries.
    • table S3. Expression of genes encoding proteins involved in nitrogen transformation, heavy metal stress response, and oxidative stress release of nitrifying organisms in the no-silver control and 30-nm AgNP (500 μg liter−1) treatment based on qPCR method (incubation time = 12 hours).
    • table S4. Primers and qPCR protocols used in this study.
    • References (75–83)

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