Genetic Factors Associated with Enhanced blaKPC Expression in Tn3/Tn4401 Chimeras

The expression of the bla(KPC) gene plays a key role in carbapenem resistance in Enterobacteriaceae. However, the genetic regulators of the bla(KPC) gene have not been completely elucidated, especially the genes in Tn3-Tn4401 chimeras. Two novel Tn3-Tn4401 chimera isoforms were characterized in our hospital, isoform A (CTA), which harbors a 121-bp deletion containing the PX promoter and was present in 22.6% (54/239) of isolates, and isoform C (CTC), which harbors a 624-bp insertion and a P1 promoter deletion and was present in only 1 isolate. The carbapenem MICs of both isoforms were 2-fold or more higher than those of the wild type (Tn3-n4401 chimera, CTB), and bla(KPC) was most highly expressed in CTA. Bioinformatics and 5' rapid amplification of cDNA ends (5' RACE) experiments indicated a novel strong putative promoter, PY, at the 3' end of the ISKpn8 gene. PY mutation nearly abrogated bla(KPC) expression (P < 0.01) and restored carbapenem susceptibility in all 3 isoforms. Although the mutation of PX or P1 halved bla(KPC) expression in CTB (P < 0.05), PX deletion caused a 68% increase in bla(KPC) expression (P = 0.037) in CTA. The level of bla(KPC) mRNA in CTC was 8-fold higher than that in InCTC, which harbors P1 (P = 0.011). These results suggest that PY is a core promoter of the blaKPC gene in the chimeras and that the deletion of the PX and P1 promoters enhanced gene expression in CTA and CTC, respectively.