Article Figures & Data
Figures
- FIG. 1.
Growth curve for P. aeruginosa strain H103 in the presence or absence of ciprofloxacin. ⧫, untreated PAO1-H103; □, PAO1-H103 plus 0.01 μg/ml ciprofloxacin; ▵, PAO1-H103 plus 0.03 μg/ml ciprofloxacin; and ×, PAO1-H103 plus 0.1 μg/ml ciprofloxacin.
- FIG. 2.
Electron micrographs of supernatants from P. aeruginosa cells that were untreated or treated with 1× MIC of ciprofloxacin. (A) Untreated strain H103 cells. (B) 1× MIC of ciprofloxacin-treated strain H103 cells. Arrows point to the presence of pyocin/phage tail structures. (C) Untreated strain PA0620::luxCDABE cells. (D) 1× MIC of ciprofloxacin-treated PA0620::luxCDABE cells showing absence of R-type pyocin tail structures.
- FIG. 3.
Induction of PA0620::luxCDABE fusions in LB media alone (black bars), with 0.1× MIC of ciprofloxacin (0.01 μg/ml; grey bars), with 0.3× MIC of ciprofloxacin (0.03 μg/ml; white bars), or with 1× MIC of ciprofloxacin (0.1 μg/ml; patterned bars).
Tables
- TABLE 1.
Pseudomonas aeruginosa strains used in this study
Strain Genotype Abbreviation Reference or source H103 Wild-type P. aeruginosa PAO1 H103 57 PA0611::ISlacZ/hah derivative of PAO1 prtR::lacZ 15 6322 PA0613::ISlacZ/hah derivative of PAO1 PA0613::lacZ 15 PAO1_lux_22_E4 PA0620::luxCDABE derivative of H103; Tcr PA0620::lux This study 3501 PA0621::ISlacZ/hah derivative of PAO1 PA0621::lacZ 15 PAO1_lux_26_H2 PA0641::luxCDABE derivative of H103; Tcr PA0641::lux This study 43080 PA3617::ISphoA/hah derivative of PAO1 recA::phoA 15 PAO1_lux_24_A3 PA3866::luxCDABE derivative of H103; Tcr PA3866::lux This study - TABLE 2.
Primer sequences used in this study
Primer Sequence PA0610_sense 5′-TAGCACTCCGATTCCACGC-3′ PA0610_antisense 5′-CCGAAGATGCGGTAGACCA-3′ PA0611_sense 5′-AGCTTCAACCGCGAGGAATA-3′ PA0611_antisense 5′-CATGTCCTCCGGCGAGTACT-3′ PA0621_sense 5′-TTTCCCGTCAGCAACGTAGC-3′ PA0621_antisense 5′-GCTGACTATCCCGCCATCTC-3′ PA0623_sense 5′-CCGAGAAGCGCTGAATTTCT-3′ PA0623_antisense 5′-CCATTGAAAGCGCTCTGGTC-3′ PA3617_sense 5′-GTGAAGAACAAGGTTTCCCCG-3′ PA3617_antisense 5′-GAGGATCTGGAACTCGGCCT-3′ PA3866_sense 5′-CCACTTGTCGTGACCAGAGGA-3′ PA3866_antisense 5′-CATCGACCCAGGCTCGTAA-3′ rplF_sense 5′-AGGTTGCTGCCGAAATTCG-3′ rplF_antisense 5′-CTTGCCTTTGTAAGGCTCCG-3′ - TABLE 3.
MICs of different antimicrobials for various strains of P. aeruginosa grown in LB medium
Strain MIC (μg/ml)a CIP ENO NOR NAL AMK FEP GAT H103 0.06 0.3 0.15 15.6 0.4 0.6 1.75 prtR::lacZ 0.06 0.3 0.15 31.25 0.4 0.6 0.875 PA0613::lacZ 0.5 1.6 >2.5 250 0.4 0.6 0.875 PA0620::lux 0.5 1.6 >2.5 250 0.4 1.25 0.875 PA0621::lacZ 0.5 >5 >2.5 250 0.2 0.3 0.875 PA0641::lux 0.5 1.6 1.25 125 0.4 1.25 1.75 recA::phoA 0.5 3.2 >2.5 250 0.4 0.6 0.875 PA3866::lux 0.06 0.3 0.15 31.25 0.4 0.3 ND ↵a Results shown are the mode of four independent experiments. CIP, ciprofloxacin; ENO, enofloxacin; NOR, norfloxacin; NAL, naldixic acid; AMK, amikacin; FEP, cefepime; GAENT, gentamicin; ND, not determined.
- TABLE 4.
Pyocin/phage operon and related genes induced by 0.3× and 1× MIC of ciprofloxacina
ORF Gene nameb Change (n-fold) with 0.3× MIC Change (n-fold) with 1× MIC Description PA0610 prtN 2.0 6.8 Transcriptional activator PA0611 prtR 1.3 3.0 Transcriptional repressor PA0612 —c 8.5 Homolog of Zn2+ finger protein PA0613 4.0 19.3 Conserved hypothetical PA0614 Hol 3.3 15.3 Holin PA0615 1.7 7.2 Conserved hypothetical PA0616 VR2 2.9 13.0 Homologous to baseplate assembly protein V PA0617 WR2 3.0 13.9 Homologous to baseplate assembly protein W PA0618 JR2 2.1 10.0 Homologous to baseplate assembly protein J PA0619 IR2 3.2 14.7 Homologous to tail protein I PA0620 HR2 2.7 12.1 Homologous to tail fiber protein H PA0621 4.3 16.7 Homologous to tail fiber assembly protein PA0622 FIR2 2.4 10.3 Homologous to contractile sheath protein FI PA0623 FIIR2 2.9 12.8 Homologous to tail tube protein FII PA0624 2.7 12.8 Conserved hypothetical PA0625 2.5 14.1 Homologous to tail length determination protein PA0626 UR2 1.8 9.8 Homologous to tail formation protein U PA0627 XR2 2.3 10.8 Homologous to tail protein X PA0628 DR2 1.8 10.0 Homologous to tail formation protein D PA0629 Lys 2.2 8.7 Lytic protein; homology to predicted chitinase PA0630 1.6 10.6 Hypothetical protein PA0631 2.4 16.2 Unique hypothetical protein PA0632 3.4 21.2 Unique hypothetical protein PA0633 VF2 4.3 16.4 Homologous to major tail protein V PA0634 3.2 15.8 Unique hypothetical protein PA0635 2.0 15.9 Conserved hypothetical protein PA0636 HF2 2.1 10.0 Homologous to tail length determination protein H PA0637 MF2 2.7 11.0 Homologous to tail fiber protein M PA0638 LF2 — 8.5 Homologous to tail fiber protein L PA0639 KF2 3.0 15.9 Homologous to tail assembly protein K PA0640 IF2 3.1 16.4 Homologous to tail assembly protein I PA0641 JF2 2.4 9.8 Homologous to tail fiber protein J PA0642 2.7 12.9 Hypothetical protein PA0643 1.9 8.9 Homologous to tail fiber domain protein PA0644 4.0 18.0 Hypothetical protein PA0645 1.8 15.8 Hypothetical protein PA0646 3.1 13.4 Homologous to putative tail fiber protein PA0647 3.8 23.0 Conserved hypothetical protein PA0648 — 8.2 Conserved hypothetical protein PA0985 pys5 4.5 18.1 Pyocin S5 PA1150 pys2 — 5.4 Pyocin S2 PA3617 recA 2.8 4.6 Recombinase for DNA recombination and repair PA3866 pys4 13.9 51.3 Pyocin S4 ↵a Only genes identified as being affected by subinhibitory (0.3 μg/ml) or inhibitory (1 μg/ml) ciprofloxacin relative to untreated strain H103 and that were of interest in this study are included. A list of all genes identified as being affected by this concentration of ciprofloxacin and exhibiting a statistically significant change (P ≤ 0.05) is available in the supplemental material. Genes are identified by ORF designation, gene name or alternative gene name, and homology description based on the Pseudomonas genome project (www.pseudomonas.com).
↵b Genes were named as per Table 1 of Nakayama et al. (21) and reflect the homology observed by these authors to phages P2 and λ.
↵c —, no significant change in expression.
- TABLE 5.
Comparison of expression changes for various pyocin/phage-related open reading frames as analyzed by relative real-time PCR and custom P. aeruginosa microarray
ORF Gene Name Change (n-fold) in PAO1 strain H103 at 0.3× MIC of ciprofloxacin Custom microarray Real-time PCR PA0610 prtN 2.0 10.0 PA0611 prtR 1.3 11.4 PA0621 4.3 3.0 PA0623 FIIR2 2.9 3.7 PA3617 recA 2.8 3.9 PA3866 pys3 13.9 22.5
HTML Page - index.htslp
Files in this Data Supplement:
- Supplemental file 1
-
List of up- and down-regulated genes induced by ciprofloxacin in Pseudomonas aeruginosa
Excel document, 186K.
- Supplemental file 1
-
List of up- and down-regulated genes induced by ciprofloxacin in Pseudomonas aeruginosa
