Characterisation of extended-spectrum beta-lactamases of the SHV family using a combination of PCR-single strand conformational polymorphism (PCR-SSCP) and PCR-restriction fragment length polymorphism (PCR-RFLP)

FEMS Microbiol Lett. 2000 Mar 1;184(1):85-9. doi: 10.1111/j.1574-6968.2000.tb08995.x.

Abstract

Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) has been developed to extend the identification of SHV beta-lactamases previously characterised by PCR-single strand conformational polymorphism (PCR-SSCP) analysis alone. Eight bacteria, each producing a different SHV beta-lactamase, were used in this study. These bacteria harbour bla(SHV-1), bla(SHV-2a), bla(SHV-3), bla(SHV-4), bla(SHV-5) (two strains), bla(SHV-11) and bla(SHV-12). All isolates were characterised by PCR-SSCP and PCR-RFLP with DdeI and NheI digestion. By a combination of these techniques, the genes encoding these beta-lactamases could be differentiated from each other. In addition, the PCR-RFLP technique theoretically can be applied to distinguish the genes encoding SHV-7, SHV-9, SHV-10, SHV-15, SHV-17 and SHV-24 from those encoding other SHV variants. We report a simple PCR-RFLP technique that can be used in epidemiological studies to enable the rapid characterisation of known SHV beta-lactamases in a combination with the previously published PCR-SSCP analysis.

MeSH terms

  • DNA, Bacterial / genetics
  • Enterobacter / enzymology
  • Enterobacter / genetics
  • Enterobacteriaceae / enzymology
  • Enterobacteriaceae / genetics*
  • Escherichia coli / enzymology
  • Escherichia coli / genetics
  • Klebsiella / enzymology
  • Klebsiella / genetics
  • Microbial Sensitivity Tests
  • Mutation
  • Polymerase Chain Reaction
  • Polymorphism, Restriction Fragment Length
  • Polymorphism, Single-Stranded Conformational
  • beta-Lactamases / genetics*

Substances

  • DNA, Bacterial
  • beta-Lactamases