TABLE 1.

Bacterial strains and plasmids used in this study

Strain or plasmidPhenotype or genotypeaSource or reference
Strains
    P. aeruginosa strains
        PAO1BesWild-type reference strainK. Stover
        CMZ091PAO1Bes ΔmexZThis study
        CM090PAO1Bes ΔPA5470This study
        CM093PAO1Bes ΔPA5471This study
        CM095PAO1Bes ΔparSThis study
        CM096PAO1Bes ΔparRSThis study
        CM099PAO1Bes ΔpmrABThis study
        CM106CM096 cis complemented with parRS genes from PAOW2, inserted at the attB site; TcrThis study
        PAOW1PAO1Bes spontaneous mutant; AmkrThis study
        PAOW2PAO1Bes spontaneous mutant with Met59-to-Ile change in ParR; AmkrThis study
        CMZ089PAOW2 ΔmexZThis study
        CM092PAOW2 ΔPA5470This study
        CM094PAOW2 ΔPA5471This study
        CM097PAOW2 ΔparSThis study
        CM098PAOW2 ΔparRSThis study
        CM100PAOW2 ΔpmrABThis study
        CMCWild-type susceptible clinical strainThis study
        CMCW2CMC spontaneous mutant with Leu14-to-Gln change in ParS; AmkrThis study
        CM107CM096 cis complemented with parRS genes from CMCW2, inserted at the attB site; TcrThis study
    E. coli strains
        S17.1recA thi pro hsdR Tra+ Smr44
        DH5αF supE44 endA1 hsdR17(rK mK) thi-1 recA1 Δ (argF-lacZYA)U169 φ80dlacZΔM15 phoA gyrA96 relA1 deoR λInvitrogen
        CC118A(ara-leu) araD ΔlacX74 galE galK phoA20 thi-1 rpsE rpoB argE(Am) recAl29
        CC118 λpirCC118 lysogenized with λpir phage12
        HB101supE44 hsdS20(rB mB) recA13 ara-14 proA2 lacY1 galK2 rpsL20 xyl-5 mtl-1 leuB6 thi-125
Plasmids
    pCR2.1Cloning vector for PCR products; lacZΔColE1 f1 ori Apr KmrInvitrogen
    pKNG101Suicide vector in P. aeruginosa; sacB Smr21
    pRK2013Helper plasmid; ColE1 ori Tra+ Mob+ Kmr6
    Mini-CTX1Self-proficient integration vector with tet, V-FRT-attPMCS, ori, int, and oriT; Tcr14
    pKNGΔmexZBamHI/ApaI 1.212-kb fragment composed of sequences flanking 5′ and 3′ ends of mexZ, cloned into pKNG101; SmrThis study
    pKNGΔ5470BamHI/ApaI 1.194-kb fragment composed of sequences flanking 5′ and 3′ ends of PA5470, cloned into pKNG101; SmrThis study
    pKNGΔ5471BamHI/ApaI 1.241-kb fragment composed of sequences flanking 5′ and 3′ ends of PA5471, cloned into pKNG101; SmrThis study
    pKNGΔparSApaI/ApaI 1.079-kb fragment composed of sequences flanking 5′ and 3′ ends of parS, cloned into pKNG101; SmrThis study
    pKNGΔparRSApaI/ApaI 1.045-kb fragment composed of sequences flanking 5′ and 3′ ends of parRS, cloned into pKNG101; SmrThis study
    pKNGΔpmrABBamHI/ApaI 1.241-kb fragment composed of sequences flanking 5′ and 3′ ends of pmrAB, cloned into pKNG101; SmrThis study
    CTX-W2parRS from PAOW2 cloned into mini-CTX1 at SpeI/EcoRV sites; TcrThis study
    CTX-CW2parRS from CMCW2 cloned into mini-CTX1 at SpeI/EcoRV sites; TcrThis study
  • a Antibiotic resistance phenotypes; Tcr, tetracycline resistance; Amkr, amikacin resistance; Smr, streptomycin resistance; Apr, ampicillin resistance; Kmr, kanamycin resistance. MCS, multiple cloning sites.