After optimizing the PCR (amplification) conditions, all
positive controls (reference strains and clinical isolates)
yielded amplicons of the predicted sizes and confirmed
the specificity of the primers used (Figure 1). The primer
paris were tested in mixed (Table 1, Figure 1) and individual
reactions (data not shown). The two multiplex PCR assays
were 6 validated with a panel of fifteen characterized
Gram-negative bacterial strains. This collection includes
5 class A carbapenemases (5 KPC), 8 class B carbapenemases
(2 IMP, 2 NDM and 4 VIM) and 2 class D carbapenemases
(2 OXA). The resistance genes of the control isolates were
correctly determined by each multiplex PCR (accuracy
100%; Figure 1). Non-specific amplification generating
fragments of unexpected size was not observed. In addition,
eleven pairs of primers have been proposed but not
tested experimentally in PCR and/or multiplex reactions