Virulence expression in response to k-opioid at high and low phosphate concentrations.

<p>(A) MvfR expression measured by β-galactosidase activity in PAO1/<i>mvfR'-lacZ</i>. (B) pyocyanin production normalized to cell density. (C) pyoverdin production normalized to cell density. (D) Mortality in <i>C. elegans</i> incubated with PAO1 grown in 0.1xTY media plus 200 µM U-50,488 with and without supplementation with Pi. (E–I) Heat maps of gene expression related to (E) low phosphate signaling/acquisition, (F) low iron signaling/acquisition, (G) quorum sensing, (H) multi-drug resistance, and (I) stress response.</p

Kappa opioid induces activation of <i>pqsABCDE</i> synergistically with HHQ.

<p>Luminescence normalized to cell density was measured dynamically in the reporter strain ΔPqsA/<i>pqsA::luxCDABE</i> grown in 0.1xTY in the presence of (A) varying doses of U-50,488; (B) varying doses of HHQ; (C) varying doses of U-50,488+200 µM HHQ; and (D) varying doses of PQS.</p

QRT-PCR analysis demonstrating involvement of PqsE and PqsH in the response of <i>P. aeruginosa</i> to the k-opioid.

<p>Average values with standard deviations are represented. n = 3/variant, *p<0.05 (wtPAO1 vs ΔPqsE); #p<0.05 (ΔPqsE vs ΔPqsH). Results were reproduced in two independent experiments.</p

Proposed circumstances under which <i>P. aeruginosa</i> is exposed to opioids and phosphate depletion during stress and mechanisms of phosphate-dependent virulence activation or suppression.

<p>Critical care treatment destabilizes the indigenous microbiota which becomes replaced by pathogens such as <i>P. aeruginosa</i> that colonize the distal gut. Processed foods or parenteral nutrition leads to nutrient limitation in the distal gut where pathogenesis develops. Physiologic stress, pain, and use of analgesics (opioids) coupled with nutrient limitation, promotes premature activation of the MvfR regulated <i>pqsABCDE</i> operon leading to the expression of multiple virulent and antibiotic resistant genes mediated by PqsE. When phosphate is abundant expression of these genes is suppressed in <i>P. aeruginosa</i>. Mechanisms may include suppression of the PstS-PhoB phosphosensory/phosphoregulatory system that involves phosphate binding to PstS, stabilization of Pst-PhoU-PhoR complex, inability of PhoR to phosphorylate PhoB, and inactivation of the PHO operon including MvfR.</p

<i>P. aeruginosa</i> PAO1 response to k-opioid during growth in nutrient poor medium.

<p>(<b>A</b>) Effect of 200 µM U-50,488 on PAO1 cell density, n = 3. (<b>B</b>) Pyoverdin production normalized to cell density, n = 3, *p<0.01. (<b>C</b>) Pyocyanin production normalized to cell density, n = 3, *p<0.05. (<b>D–D″</b>) Transmission electron microscopy images of PAO1 cells grown in (<b>D</b>) 0.1xTY, and (<b>D′, D″</b>) grown in 0.1xTY supplemented with 200 µM U-50,488. Flagella (<b>D′</b>) and vesicles (<b>D″</b>) are shown by arrows. (<b>E</b>) Kaplan-Meier survival curves in <i>C. elegans</i> feeding in <i>P. aeruginosa</i> PAO1 grown in poor nutrient medium vs poor nutrient medium containing 50 µM U-50,488. Cell cultures were collected at 7 hrs. Cumulative survival is represented as n = 7/plate, 3 plates/experiment, p<0.01.</p

Acoustic telemetry detection data

This file explains the variables in the acoustic telemetry detection dataset that accompanies: Kraus et al. 2018, Evaluation of Acoustic Telemetry Grids for Determining Aquatic Animal Movement and Survival, in Methods in Ecology and Evolution