Caracterização molecular, formação de biofilme e susceptibilidade a antimicrobianos de isolados de E. coli de aderência difusa Afa/Dr e não Afa/Dr

• Autor(a) principal: Fagundes, Laura Kreuz
• Data de Publicação: 2013
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: Repositório Institucional da Universidade Federal do Espírito Santo (riUfes)
• Texto Completo: http://repositorio.ufes.br/handle/10/1344
• topic: Doenças Infecciosas e Parasitárias; Escherichia coli; Biofilme; 61

The diffusely adherent Escherichia coli (DAEC), a diarrheagenic pathotype of E. coli, represent a heterogeneous group without a virulence marker common to all isolates and with a controversial role in childhood diarrhea. The aim of this study was to characterize phenotypic and genotypic samples of DAEC, with or without Afa/Dr adhesins, isolated from children with and without diarrhea. In 70 samples DAEC, PCR was performed to search for genes described in DAEC, EAEC or UPEC, encoding: (i) eight fimbrials and afimbrials (fimH, papC, sfa, aggA, aafA, agg3A, aida/aah, afaC ), (ii) five toxins (pet, astA, set1A, sat, hlyA), (iii) three iron-chelators (irp2, yucA, chuA/shuA), (iv) invasin (daad) and; antigen 43 (agn43). Biofilm formation assay was carried out from the bacteria grown in Luria-Bertani broth and inoculated in microtiter plates with DMEM 0.4% glucose. Optical density (OD490) was measured after safranin staining. Seroagglutination for 23 O antigens (Probac Brazil) was performed in 50% of DAEC strains. Disk diffusion method was conducted to test the susceptibility to 13 antimicrobial agents. The presence of at least one gene encoding for adhesins, toxins, iron chelators or invasin were found in 58.6%, 51.4%, 80% and 48.6%, respectively, with the genes fimH, irp2, agn43, yucA, chuA/shuA, present in over 50% of the strains. afaC+ gene (PCR+) and/or afaBC+ probe (colony hybridization) classified 50% of DAEC as Afa/Dr, being pet, sat, irp2, yucA, chuA/shuA and agn43 significant in these strains (p<0,05). Ou of the 70 DAEC, 44.3% were biofilm former, equally present among Afa/Dr and non Afa/Dr, and no gene has been associated with this phenotype. Serology of 35 strains showed the following serogroups: 1 O29, 2 O125, 2 O127 and 7 O86. All O86 were DAEC Afa/Dr. Higher frequency of antimicrobial resistance were found for ampicillin (55.7%), trimethoprim/sulfamethoxazole (35.7%) and tetracycline (28.6%) and the pattern resistant/intermediate to amoxicillin/clavulanic acid, ampicillin, sulfamethoxazole/trimethoprim was significant in Afa/Dr DAEC, as well as the multi-drug resistance (p <0.05). In conclusion, we observed: (i) a high frequency of fimH and pet and the presence of agn43, hitherto not described in DAEC, at similar frequencies to those found in EAEC, UPEC and EAEC/UPEC, respectively; (ii) that the samples Afa/Dr and non Afa/Dr DAEC constituted groups with different genetic profiles to each other; (iii) a few serogroups were found among DAEC; (iv) smaller resistance frequencies when compared with the few descriptions of DAEC, suggesting a lower selective pressure of the population of the present study and; (v) DAEC Afa/Dr strains may represent an important reservoir of antibiotic resistance genes, beyond several virulence factors.