Table 3 Plasmids used in this study.

pUC19

• High copy plasmid [41]; accession # L09137

• Amp^R

• E. coli DH5α

pGP704

• Suicide vector [33]

• Backbone from pBR322, ori R6K instead of ori E1 from pBR322, mob region from pRP4

• Replicates in E. coli SM10λpir but not in EHEC strains used because ori R6K needs π-Protein coded by the pir gene for replication

• Amp^R

• E. coli SM10λpir

pMHH1

• Derivative of pGP704

• Has a positive selection system based on the sacB gene from Bacillus subtilis [38]

• E. coli SM10λpir

pMHH6*

• Derivative of pUC19

• Harbors a 990 bp rpoS fragment from EHEC 86-24 amplified with primers RpoS 3/RpoS 4 and cloned via Xba I und Sac I restriction sites

• E. coli DH5α

pMHH7*

• Derivative of pMHH6

• Restriction digest with Dra III und Bsa AI leads to a 390 bp deletion in rpo S

• E. coli DH5α

pMHH8*

• Derivative of pMHH1

• Contains the mutagnized 600 bp rpoS fragment from pMHH7 cloned via Xba I and Sac I restriction sites

• E. coli SM10λpir

pBR322

• Low copy plasmid [36]; accession # J01749

• Amp^R, Tet^R

• E. coli DH5α

• EHEC 126814

pSC1*

• Derivative of pBR322

• Harbors nlpD-rpoS from EHEC 126814 accession # DQ272417 amplified by PCR using primers RpoS 8/RpoS 9 and cloned via Hind III restriction sites

• E. coli DH5α

• E. coli MHH126-5

pSC2*

• Derivative of pBR322

• Harbors nlpD-rpoS from EHEC EDL933_a (rpoS*) accession # DQ272419 amplified by PCR with primers RpoS 8/RpoS 9 and cloned via Hind III restriction sites

• E. coli DH5α

• E. coli MHH126-5

pMH33*

• Made from pSC2 by PCR mutagenesis using Primers RpoS 17a/RpoS 17b

• Has point mutation rpoS_T721G which resolves the TAA stop codon at position 723 in $rpo{S}_{EDL933_a}$

• E. coli MHH126-5

pUD2*

• Derivative of pBR322

• Harbors nlpD-rpoS from EHEC EDL933_b accession # DQ272421 amplified by PCR with primers RpoS 8/RpoS 9 and cloned via Hind III restriction sites

• E. coli MHH126-5

pUD4*

• Derivative of pBR322

• Harbors nlpD-rpoS from EHEC 86-24 accession # DQ272418 amplified by PCR with primers RpoS 8/RpoS 9 and cloned via Hind III restriction sites

• E. coli MHH126-5

pUD8*

• Derivative of pBR322

• Harbors nlpD-rpoS from STEC E-D53 accession # DQ272423 amplified by PCR with primers RpoS 8/RpoS 9 and cloned via Hind III restriction sites

• E. coli MHH126-5

pUD6*

• Derivative of pBR322

• Harbors nlpD-rpoS from STEC E-D68 accession # DQ272424 amplified by PCR with primers RpoS 8/RpoS 4c and cloned via Hind III restriction sites

• E. coli MHH126-5

pUD9*

• Derivative of pBR322

• Harbors nlpD-rpoS from STEC E-D68 accession # DQ272424 amplified by PCR with primers RpoS 8/RpoS 4c and cloned via Hind III restriction sites

• E. coli MHH126-5

pUD10*

• Derivative of pBR322

• Harbors nlpD-rpoS from EHEC 288597 accession # DQ272422 amplified by PCR with primers RpoS 8/RpoS 9 and cloned via Hind III restriction sites

• E. coli MHH126-5

pDS4*

• Derivative of pBR322

• Harbors nlpD-rpoS from EcN amplified by PCR with primers RpoS 8/RpoS 22 and cloned via Hind III restriction sites

• E. coli MHH126-5