Escherichia coli enteropatogênica (EPEC) é uma importante causa de diarreia infantil. EPEC adere no epitélio intestinal e causa uma lesão conhecida como attaching and effacing (A/E). Cepas recombinantes de Mycobacterium smegmatis (Smeg) e Mycobacterium bovis (BCG) foram construídas para expressar BfpA ou Intimina. Os genes dessas proteínas foram amplificadas por PCR do DNA genômico de EPEC e inseridos nos sítios de BamHI/KpnI do vetor pMIP12. Os plasmídeos construídos pMIP-bfpA e pMIP-intimina foram introduzidos, separadamente,nas cepas de Smeg e BCG. Clones recombinantes foram selecionados baseado na resistência a kanamicina e nomeados como rSmeg pMIP (bfpA ou intimina) and rBCG pMIP (bfpA ou intimina). A expressão dos genes bfpA e intimina foram detectados por Western blotting usando anticorpos primários policonais anti-BfpA e anti-intimina. A imunogenicidade dessas proteínas foi avaliada em camundongos C57BL/6 pela presença de anticorpos anti-BfpA e anti-intimina IgA e IgG nas fezes e soro, respectivamente. TNF-a e INF-g foram produzidas in vitro por céulas do baço de camundongos imunizados com a vacina recombinante com BfpA e Intimina. A adesão de EPEC (E2348/69) em células alvo Hep-2 foi bloqueada pelos anticorpos IgA ou IgG de camundongos imunizados com as vacinas recombinantes, mas não por anticorpos de animais não imunizados. Vacinas recombinantes contendo as proteínas BfpA e Intimina de EPEC podem ser candidatas promissoras.

Enteropathogenic Escherichia coli (EPEC) is an important cause of diarrhea in children. EPEC adheres to the intestinal epithelium and causes attaching and effacing (A/E) lesions. Recombinant Mycobacterium smegmatis (Smeg) and Mycobacterium bovis BCG strains were constructed to express either BfpA or intimin. The entire bfpA gene and a portion of the intimin gene were amplified by PCR from EPEC genomic DNA and inserted into the pMIP12 vector at the BamHI/KpnI sites. The pMIP bfpA and pMIP intimin vectors were introduced separately into Smeg and BCG. Recombinant clones were selected based on kanamycin resistance and designated rSmeg pMIP (bfpA or intimin) and rBCG pMIP (bfpA or intimin). The expres-sion of bfpA and intimin was detected by Western blotting using polyclonal anti-BfpA and anti-intimin primary antibodies. The immunogenicity of these proteins was assessed in C57BL/6 mice by assaying the feces and serum for the presence of anti-BfpA and anti-intimin IgA and IgG antibodies. TNF-a and INF-g were produced in vitro by spleen cells from mice immunized with recombinant BfpA, whereas TNF-g was produced in mice immunized with recombinant intimin. The adhesion of EPEC (E2348/69) to Hep-2 target cells was blocked by IgA or IgG antibodies from mice immunized with recombinant BfpA or intimin but not by antibodies from non-immunized mice. Immunogenic non-infectious vectors containing relevant EPEC virulence genes may be promising vaccine candidates.