Molecular investigation of tRNA genes integrity and its relation to pathogenicity islands in Shiga toxin-producing Escherichia coli (STEC) strains

RESUMO

tRNA genes are known target sites for the integration of pathogenicity islands (PAI) and other genetic elements, such as bacteriophages, into bacterial genome. In most STEC (Shiga toxin-producing Escherichia coli), the PAI called LEE (locus of enterocyte effacement) is related to bacterial virulence and is mostly associated to the tRNA genes selC and pheU. In this work, we first investigated the relationship of LEE with tRNA genes selC and pheU in 43 STEC strains. We found that 28 strains (65 percent) had a disrupted selC and/or pheU. Three of these strains (637/1, 650/5 and 654/3) were chosen to be submitted to a RAPD-PCR technique modified by the introduction of specific primers (corresponding to the 5'end of genes selC and pheU) into the reaction, which we called "anchored RAPD-PCR". The PCR fragments obtained were transferred onto membranes, and those fragments which hybridized to selC and pheU probes were isolated. One of these fragments from strain 637/1 was partially sequenced. An 85-nucleotide sequence was found to be similar to the cfxA2 gene that encodes a beta-lactamase and is part of transposon Tn4555, a pathogenicity island originally integrated into the Bacteroides genome.