TABLE 1.

Bacterial strains and mutants used in this study

Strain or plasmidRelevant characteristicsReference or source
P. aeruginosa strains
    PA99Secretes ExoS, ExoT, and ExoU; clinical isolate8
    PA99SSecretes only ExoS; PA99ΔexoTexoUThis work
    PA99TSecretes only ExoT; PA99null complemented with mini-CTX1-ExoTThis work
    PA99USecretes only ExoU; PA99ΔexoSexoTThis work
    PA99nullSecretes no known effectors, makes functional secretion apparatus; PA99ΔexoSexoTexoUThis work
    PA99secrSecretes no effectors, does not make secretion apparatus; PA99ΔpscJThis work
    PA99null+SSecretes only ExoS; PA99ΔexoSexoT exoU + mini-CTX1-ExoSThis work
    PA99null+S(R146A)Secretes only ExoS(R146A); no GAP activity; PA99ΔexoSexoTexoU +mini-CTX1-ExoS-R146AThis work
    PA99null+S(E379A/E381A)Secretes only ExoS (E379A/381A); no ADPRT activity: PA99ΔexoS exoTexoU+mini-CTX1-ExoS-E379A/E381AThis work
    PA103Secretes ExoT and ExoU; laboratory isolate31
    388Secretes ExoS and ExoT; laboratory isolate3
E. coli strains
    XL1-BluerecA1 endA1 gyrA96 thi-1 hsdR17 supE44 relA1 lac [F′ proAB lac1qZ ΔM15 Tn10 (Tetr)]Stratagene
    GM2163F ara-14 leuB6 fhuA31 lacY1 tsx-78 glnV44 galK2 galT22 mcrA dcm-6 hisG4 rfbD1 rpsL136 (Strr) dam-13::Tn9 (Camr) xylA5 mtl-1 thi-1 mcrB1 hsdR2New England Biolabs
    TOP10F mcrA Δ(mrr-hsdRMS-mcrBC) Φ80lacZ ΔM15 ΔlacX74 deoR recA1 araD139 Δ(ara-leu)7697 galU galK rpsI (Strr) endA1 nupGInvitrogen
    S17.1thi pro hsdR recA RP4-2 (Tet::Mu)(Km::Tn7)53
Plasmids
    Mini-CTX1Plasmid for chromosomal gene integration into attB locus; TetrH. Schweizer; 21
    Mini-CTX1-ExoSexoS from strain 388 in mini-CTX1; TetrThis work
    Mini-CTX1-ExoS-R146AGAP point mutant in mini-CTX1-ExoS; TetrThis work
    Mini-CTX1-ExoS-E379A/E381AADPRT point mutant in mini-CTX1-ExoS; TetrThis work
    Mini-CTX1-ExoTexoT from PA103 in mini-CTX1; TetrThis work
    pCM104ΔexoT in pEX100T (in frame); Apr (Cbr)aThis work
    pCM111ΔpscJ in pEX100T (in frame); Apr (Cbr)This work
    pEX100TPlasmid for homologous recombination; Apr52
    pFlp2Plasmid carrying recombinase to excise mini-CTX1 plasmid sequences; Apr (Cbr)H. Schweizer; 21
    pGS003ΔexoS in pEX100T (in frame); Apr (Cbr)This work
    pGS013ΔexoU in pEX100T (in frame); Apr (Cbr)This work
    pGS015ΔexoU in pEX100T with xylE aacC1 cassette inserted into the SalI site of pGS013; Apr (Cbr) GmrThis work
  • a Apr in E. coli, Cbr in P. aeruginosa.