[Direct in situ PCR method for the detection of verotoxin-producing Escherichia coli]

Rapid detection of verotoxin-producing Escherichia coli at a single-cell level under an epifluorescence microscope without culturing processes was accomplished by using the direct in situ PCR technique. We used a DNA primer set for amplification of the slt-I and slt-II genes encoding respectively verotoxin 1 and 2 (EVT and EVS primers). The bacterial cells were detected specifically by the HNPP (2-hydroxy-3-naphthoic acid-2'-phenylanilide phosphate)/Fast Red TR reaction technique. The direct in situ PCR with HNPP/Fast Red TR technique is applicable to the detection of verotoxin-producing bacteria with the slt-I or slt-II gene in not only Escherichia coli O157 but also VTEC of other serotypes.