The aim of our work was to develop a new molecular method for the simultaneous detection of Salmonella spp., Listeria monocytogenes and Escherichia coli O157 directly in milk samples.jrm_170 195..213 Three specific target sequences were chosen for a multiplex polymerase chain reaction (PCR) assay: a 155-bp region of the Salmonella spp. tetrathionate reductase (ttr) locus; a 173-bp region of the L. monocytogenes listeriolysin O gene (hlyA); a 217-bp region of the E. coli O157 lipopolysaccharide gene (rfbE). An internal amplification control was also included to detect PCR inhibition. In addition, a magnetic-based extraction method was also developed to isolate PCR-ready DNA from milk. The assay was able to detect, whether alone or mixed, also with a difference of 2 log units, as few as 102 cells of each pathogen per 10 mL of spiked milk.
Simultaneous direct detection of Salmonella spp., Listeria monocytogenes and Escherichia coli O157 in milk samples by magnetic exctraction and multiplex PCR
AMAGLIANI, GIULIA;BRANDI, GIORGIO;MAGNANI, MAURO
2009
Abstract
The aim of our work was to develop a new molecular method for the simultaneous detection of Salmonella spp., Listeria monocytogenes and Escherichia coli O157 directly in milk samples.jrm_170 195..213 Three specific target sequences were chosen for a multiplex polymerase chain reaction (PCR) assay: a 155-bp region of the Salmonella spp. tetrathionate reductase (ttr) locus; a 173-bp region of the L. monocytogenes listeriolysin O gene (hlyA); a 217-bp region of the E. coli O157 lipopolysaccharide gene (rfbE). An internal amplification control was also included to detect PCR inhibition. In addition, a magnetic-based extraction method was also developed to isolate PCR-ready DNA from milk. The assay was able to detect, whether alone or mixed, also with a difference of 2 log units, as few as 102 cells of each pathogen per 10 mL of spiked milk.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
