Selection of primers for specific detection of Clostridium botulinum types B and E neurotoxin genes using PCR method

Improved oligonucleotide primers were designed to flank 370- and 307-bp fragments of the bont genes encoding botulinum neurotoxins types B and E, respectively. Primer specificity was confirmed for reference strains of Clostridium botulinum types B and E for strains representing bacterial species common in food, and for the DNA mixtures of C. botulinum types B and E in the presence of background DNA isolated from cold smoked salmon and ham. The detection limit of template DNAs of C. botulinum types B and E from the DNA mixtures increased from 1 to 0.1 ng by raising annealing temperature from 50°C to 62°C.