RAPD fingerprinting of enterohaemorrhagic Escherichia coli O157:H7 isolates in Santafe de Bogota, DC, Colombia

Enterohaemorrhagic Escherichia coli (EHEC) is an important, food-born pathogen increasingly causing diarrhoea, haemorrhagic colitis and haemolytic uraemic syndrome. The objectives of our study were to determine the genetic diversity and molecular typing of EHEC O 157:H7 isolated in Bogota, Colombia, by using random amplification of polymorphic DNA (RAPD). We investigated the genetic diversity of 16 enterohaemorrhagic E coli (EHEC) strains of serotype O 157:H7, previously isolated from diarrhoeic patients in Bogota over a period of 1 year, and from bovine faeces. Informative band arrays were obtained with a 10-mer primer (OPA-07) with 60% C + G content. Polymerase chain reaction (PCR) analyses were performed in 25 μl reaction volumes containing 1X PCR buffer, 1.5 mM MgCl₂, 0.2 mM (each) dNTPs, 40 pmol of primers, 1 U of Taq DNA polymerase and 25 ng of DNA. Temperature cycling was controlled by a PCR system programmed as follows: 94 °C for 5 min, followed by 30 cycles of 94°C for 0.5 min, 36°C for 0.5 min, 72 °C for 0.5 min and a final extension step at 72 °C for 4 min. RAPD reactions generated informative band arrays composed of a minimum of 1 band and a maximum of 15 bands, with molecular size ranging approximately from 1,018 to 5,090 bp. RAPD profiles exhibited an overall rate of genetic disagreement of 4 - 79%. Stronger analysis of genetic similarity between different RAPD profiles demonstrates that Colombian strains are closely related and represent two clonal lineages. (C) 2000 Science Reviews.