Rapid Detection of bla(KPC-9) Allele from Clinical Isolates

The emergence of Klebsiella pneumoniae carbapenemase (KPC) nosocomial outbreaks related to specific bla(KPC) gene variants dictates the need for applicable diagnostic methods for allele discrimination. We report here a simple method of bla(KPC-9) allele recognition based on a combination of endonuclease digestion analysis and PCR amplification using unique primers. K. pneumoniae isolates carrying the bla(KPC) gene were tested. Digestion with RsaI restriction endonuclease was found to efficiently differentiate the bla(KPC-2) from the bla(KPC-9) variants into two distinct groups of digestion patterns named KPC-2-like and KPC-9-like, respectively. An additional procedure, the amplification refractory mutation system (ARMS) method, was applied to identify the variant within the same group. The principles of this procedure could be developed to identify several bla(KPC) gene variants, as well as monitoring the spread and evolution of specific KPC variants within local geographical regions.