Background: Extended-spectrum β-lactamases (ESBLs) belonging to the SHV family remain a major cause of ESBL-positive phenotypes in Klebsiella pneumoniae. The blaSHV gene is a normal constituent of the K. pneumoniae chromosome. However, most ESBL-encoding blaSHV genes found in K. pneumoniae are plasmid borne. The objective was to determine the contribution of promoter variants to the expression of plasmid-borne blaSHV genes. Methods: K. pneumoniae clinical isolates were analysed for the presence of IS26 insertions characteristic of plasmid-borne blaSHV, and differences in their blaSHV promoter sequences and expression levels. A high resolution melting (HRM)-based method for rapid promoter analysis was developed. Results: An IS26 insertion characteristic of the plasmid-borne blaSHV-1/blaSHV-2/blaSHV-5 family was 100% linked to a promoter mutated in the –10 region, a mutation previously only found on the chromosome. The mutation was shown by real-time reverse transcriptase PCR to be associated with increased blaSHV expression. Conclusions: Plasmid-borne blaSHV is associated with strong promoters. It is likely that an SHV-dependent ESBL-positive phenotype requires both a strong promoter and a coding sequence mutation. An HRM assay can indicate blaSHV expression. ; Mark S. Turner, Patiyan Andersson, Jan M. Bell, John D. Turnidge, Tegan Harris and Philip M. Giffard