Impedance changes caused by the PA14ΔphzM mutant.

<p>Changes to the impedance spectrum can be clearly observed when a mutant of <i>P. aeruginosa</i>, unable to produce pyocyanin is measured. Changes to the (A) phase, (B) modulus and (C) normalised resistance are similar to those found in the PA14 wild type. (D) the UV-visible spectrum shows a peak at 367 nm, suggesting the presence of phenazine-1-carboxylic acid. (E) Indicates the biosynthesis pathway for phenazines and shows the effect that the phzM knockout is anticipated to have. Background shading represent +/−1 SD. PA14ΔphzM <i>n</i> = 3; Neg Ctrl <i>n</i> = 3.</p

Indicative cell densities of planktonic cells at the end of the experiment and timepoints during the experiment.

<p>Indicative cell densities of planktonic cells at the end of the experiment and timepoints during the experiment.</p

Attachment of <i>P. aeruginosa</i> PA14 to the electrode surface could affect the impedance.

<p>Electrode coupons were incubated in LB media (aerobic conditions) for 48 Hrs. (A) At the air-liquid interface, extensive biofilm formation can be seen indicating that <i>P. aeruginosa</i> will attach to the electrode surface. (B) Some limited microbial attachment can be seen on the electrode surface at the bottom of the a well after 48 hours. (C) Negative control showing that the electrode surface does not autofluoresce. (D) A small change can be observed in the impedance signature (contrasted to the negative control) at the end of a microaerophilic experiment when the media is exchanged with a fresh, sterile aliquot. Scale bars represent 50 um. Shading in (D) represents +/−1SD, <i>n = </i>3.</p

Impedance changes caused by the PA14ΔphzS mutant.

<p>Similar changes are observable in the (A) phase, (B) modulus and (C) normalised resistance to the wild type and the PA14ΔphzS mutant. (D) the presence of phenazine-1-carboxylic acid can also be seen. (E) Indicates the biosynthesis pathway for phenazines and shows the effect that the phzS knockout is anticipated to have. Background shading represent +/−1 SD. PA14ΔphzS <i>n</i> = 3; Neg Ctrl <i>n</i> = 3.</p

Polymicrobial growth of <i>P. aeruginosa</i> and <i>S. aureus</i>.

<p>When RN4220 and PA14 are grown together, the impedance is similar to when PA14 is grown alone. (A) and (B) a change is clearly visible the overall phase angle and a drop in the impedance modulus, (C) the normalised resistance shows a peak, and a change in the peak frequency as a result of growth. Background shading represent +/−1 SD. Polymicrobial <i>n = </i>5; neg ctrl <i>n</i> = 5.</p

Impedance changes as consequence of growth with <i>S. aureus</i> RN4220.

<p>Shows that there is no discernible different in (A) the phase, (B) the modulus and (C) the normalised reactance under the same conditions as those used for <i>P. aeruginosa</i> PA14. The arrow indicates a slight peak at 125 Hz at 24 hours moving to 400 Hz at later timepoints. Background shading represent +/−1 SD. RN4220 <i>n</i> = 4; Neg Ctrl <i>n</i> = 5.</p

Measurement of P. aeruginosa PA14ΔphzA1-G1/A2-G2 double mutant.

<p>(A and B) no significant different can be seen in the phase or modulus of the impedance. (C) a small difference in the normalised resistance can be seen, contrasted to the negative control. (D) The UV-visible spectrum indicates that the phenazines present in the wild type or other mutants tested are no present. (E) Indicates the point on the biosynthesis pathway that the double knockout is anticipated to affect. Background shading represent +/−1 SD. PA14ΔphzA1-G1/A2-G2 <i>n</i> = 4; Neg Ctrl <i>n = </i>4.</p

Changes in the impedance resulting from <i>P. aeruginosa</i> PA14.

<p>(A) phase plot, showing a drop in the absolute phase angle as a result of microbial growth, (B) the final impedance modulus at low frequency is more than one and a half orders of magnitude lower than the starting impedance, (C) the normalised resistance shows a peak for the electrode chambers containing PA14 that is not present in the control data. Background shading and error bars represent +/−1 SD. PA14 <i>n</i> = 4; Neg Ctrl <i>n</i> = 5.</p

Bacterial strains used in this study.

<p>Bacterial strains used in this study.</p

Mechanisms through which microorganisms could affect the impedance.

<p>(A) The impedance signature of a given electrochemical system is defined by the interplay of solution resistance, redox compounds, diffusion gradients and the electrolyte composition adjacent to the electrode surface. (B) Microorganisms could affect this impedance signature through: (1) production of electroactive secondary metabolites that facilitate a charge transfer at the electrode/electrolyte interface; (2) biofilm matrix attached to the electrode surface that affects capacitance and/or charge transfer; (3) direct microbial attachment, through pili, flagella and outer membrane proteins facilitating charge transfer; (4) Outer cell membrane contact at high cell densities that affect capacitance; (5) Breakdown of nutrients within the media reducing solution resistance; (6) Protein/macromolecule adsorption on the electrode surface influencing double layer capacitance.</p