Construction of strains and plasmids

Template vector p3773 was generated by PCR amplification of tetR P_tetA from pWRG99 using TetR-PtetA-For-SacI and TetR-PtetA-Rev-XhoI. The resulting fragment was digested by XhoI and SacI and subcloned in XhoI/SacI digested plasmid p2795. For introduction of a tetracycline-inducible promoter element upstream of the coding region for Bcf, Fim, Lpf, Pef, Saf, Stb, Stc, Std, Stf, Sti and Stj fimbriae as well as the Bap T1SS, an expression cassette was amplified from p3773 using oligonucleotides as listed in Table ^S4 and introduced into the genome of S. Typhimurium NCTC 12023 by λ Red mutagenesis harboring pWRG730 for the expression of Redαβγ. The coding regions for the respective fimbriae including the aph tetR P_tetA cassette and the vector pWSK29 were amplified with oligonucleotides as listed in Table ^S4 and purified by PCR purification (Qiagen). The reverse-amplified PCR product from vector pWSK29 and the respective PCR products encompassing the coding sequence for fimbrial adhesins were then assembled by Gibson assembly according to manufacturer’s protocols (NEB).

The genes encoding for MisL, SadA, Rck, PagN, CsgBAC, CsgEFG and the operon encoding for the bapABCD operon were amplified from S. Typhimurium NCTC12023. shdA and the lpf operone were amplified from S. Typhimurium LT2 with oligonucleotides listed in Table ^S4. The vector p4392 including the aph tetR P_tetA but excluding the fimAICDHF operon was then reverse amplified using oligonucleotides listed in Table ^S4. After PCR purification, reverse-amplified PCR product from p4392 and the respective PCR products containing the coding regions for the respective adhesins were assembled by Gibson assembly. To generate the construct for expression of curli fimbriae, both PCR products encompassing the operons csgBAC and csgEFG were assembled with the reverse-amplified PCR product from p4392 by Gibson assembly in a single step to generate one hybrid operon under control of P_tetA.