RESUMO
Since no recent data characterizing Shiga toxin-producing E. coli (STEC) from human infections in Brazil are available, the present study aimed to investigate serotypes, stx genotypes, and accessory virulence genes, and also to perform pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST) of 43 STEC strains recovered from 2007 to 2017. Twenty-one distinct serotypes were found, with serotype O111:H8 being the most common. However, serotypes less frequently reported in human diseases were also found and included a hybrid STEC/ETEC O100:H25 clone. The majority of the strains carried stx1a as the sole stx genotype and were positive for the eae gene. Regarding the occurrence of 28 additional virulence genes associated with plasmids and pathogenicity islands, a diversity of profiles was found especially among the eae-harboring strains, which had combinations of markers composed of up to 12 distinct genes. Although PFGE analysis demonstrated genetic diversity between serotypes such as O157:H7, O111:H8, O26:H11, O118:H16, and O123:H2, high genetic relatedness was found for strains of serotypes O24:H4 and O145:H34. MLST allowed the identification of 17 distinct sequence types (STs) with ST 16 and 21 being the most common ones. Thirty-five percent of the strains studied were not typeable by the currently used MLST approach, suggesting new STs. Although STEC O111:H8 remains the leading serotype in Brazil, a diversity of other serotypes, some carrying virulence genes and belonging to STs incriminated as causing severe disease, were found in this study. Further studies are needed to determine whether they have any epidemiological relevance.
RESUMO
O sorotipo O26:H11 associado às aEPEC e STEC tem sido frequentemente implicado com doenças entéricas em diversos países. Análises comparativas de cepas O26:H11 STEC e aEPEC de vários países indicam que as aEPEC O26:H11 representam clones que perderam os genes stx através da excisão fágica. No Brasil, o isolamento de aEPEC O26:H11 de casos de infecção humana é frequente, sendo este um dos mais importantes sorotipos de aEPEC em nosso meio. O objetivo deste estudo foi investigar e fazer uma análise comparativa sobre a ocorrência de vários genes de virulência, alguns deles altamente específicos para o patotipo STEC, em cepas O26:H11 STEC e aEPEC, além de avaliar a diversidade clonal através das técnicas de PFGE e sequenciamento MLST. Oito das 10 cepas STEC apresentaram o genótipo stx1a, enquanto duas cepas apresentaram stx2a. Este é o primeiro relato sobre a ocorrência de STEC O26:H11 albergando o genótipo stx2a no Brasil. Os genes plasmidiais ehxA, katP, espP e toxB foram encontrados em 8 (80%), 7 (70%), 8 (80%) e 8 (80%) das STEC. Todas as STEC abrigaram os genes efa, escN, nleB, nleE, sen, z2098, z2099, z2121, ureD e terE. Os genes espK e espM1 foram encontrados em igual frequência. Dentre as aEPEC, todas foram positivas para os genes efa, escN, nleB, nleE, sen e z2121. Os genes ehxA, espP, espM1, iha, katP, toxB, z2098, z2099, espK, espV, espN, ureD e terE estiveram presentes em 25 (66%), 25 (66%), 35 (92%), 31(82%), 16 (42%), 18 (47%), 26 (68%), 26 (68%), 31 (82%), 13 (34%), 19 (50%), 26 (68%) e 30 (79%) das cepas, respectivamente. O gene astA foi encontrado em apenas três (8%) aEPEC e nenhuma das cepas estudadas...(AU)
The O26: H11 serotype which is associated with pathotypes aEPEC and STEC, is implicated with enteric diseases in several countries. Comparative analyzes of O26:H11 STEC and aEPEC strains from several countries indicated that many aEPEC strains represent STEC clones that lost the stx genes through phage excision. The aim of this study was to investigate the occurrence of several virulence genes, some of which highly specific for the STEC pathotype, in strains O26:H11 STEC and aEPEC, and to evaluate clonal diversity employing of PFGE and MLST aproches. Eight of the 10 STEC strains presented stx1a subtype, while two strains presented stx2a. This is the first report on the occurrence of STEC O26: H11 harboring the stx2a genotype in Brazil. The plasmidial genes ehxA, katP, espP and toxB were found in 8 (80%), 7 (70%), 8 (80%) and 8 (80%) of STEC. All STECs harbored the efa, escN, nleB, nleE, sen, z2098, z2099, z2121, ureD and terE genes. The espK and espM1 genes were found at the same frequency. Among aEPEC, all were positive for the efa, escN, nleB, nleE, sen and z2121 gene. The genes ehxA, espP, espM1, iha, katP, toxB, z2098, z2099, espK, espV, espN, ureD and terE were present in 25 (66%), 25 (66%), 35 (92%), 31 ( 82%), 16 (42%), 18 (47%), 26 (68%), 26 (68%), 31 (82%), 13 (34%), 19 (50%), 26 (68%) ) and 30 (79%). The astA gene was found in only three (8%) aEPEC and none of these strains showed the presence of cdt-V, etpD and pagC. All STECs presented CRISPR sequences, being C+D CRISPR types. Among the aEPEC in addition to these two polymorphisms associated with CRISPR sequences, type E was also found. PFGE typing revealed a wide genetic diversity both between STEC and aEPEC. MLST typing was performed on 24 strains of aEPEC and STEC, ten STECs and 14 aEPEC, but not all strains had a valid ST through the Enterobase bank, only 18 strains presented ST, and all these ST belonged to the clonal complex 29. ST21 and ST29 occurred equally, in four STECs each. Among the aEPEC, two were ST21 and eight belonged to ST29. Most strains of aEPEC O26: H11 in our study demonstrated several characteristics compatible with STEC and are therefore derived from this pathotype. (AU)