Characterization of Unexpressed Extended-Spectrum Beta-Lactamase Genes in Antibiotic-Sensitive Klebsiella pneumoniae Isolates.

OBJECTIVE

The current investigation explores whether extended-spectrum β-lactamase (ESBL) genes exist in clinical non-ESBL-producing Klebsiella pneumoniae isolates.

METHODS

A total of 202 clinical isolates with non-ESBL-producing K. pneumoniae were collected from southern and middle of China. Thirteen β-lactamase genes (blaSHV, CTX-M, TEM, OXA-2, OXA-10, VEB, PER, SFO, GES, CSP, TLA, BEL, and IBC) were screened by PCR and their identity confirmed by sequencing of PCR products. The ESBL-producing phenotype of the isolates that carried ESBL genes was tested and confirmed in 9 of the 18 isolates by a double-disc synergy test. The sequences upstream of ESBL genes of isolates with ESBL-producing genotype (+)/phenotype (-) were also subjected to PCR and sequencing. The ESBL genes and their upstream regions were cloned into Escherichia coli DH5α for functional evaluation.

RESULTS

A total of 8.9% (18/202) isolates carried ESBL genes. All of them harbored only one ESBL gene, including 33.3% (6/18) blaSHV and 66.7% (12/18) blaCTX-M. Among the isolates carrying ESBL genes, nine isolates were confirmed as ESBL phenotype (-). The ESBL genotype (+)/phenotype (-) isolates had blaSHV-27,38,41,42 (66.7%, 6/9) and blaCTX-M-3,15,24 (33.3%, 3/9). The upstream gene sequences, including promoters of these unexpressed ESBL genes, were intact without any mutations or spacers and effective among eight strains. The ISEcp1 element in the upstream region was not found in one isolate carrying an unexpressed blaCTX-M-15 gene.

CONCLUSIONS

Clinical non-ESBL-producing K. pneumoniae isolates could carry ESBL genes with intact promoter, but without the correlated phenotype. Specific silencing mechanisms may play an important role in regulating ESBL gene expression. This kind of isolates has the potential to transfer their ESBL genes to other bacteria with effective promoters, resulting in ESBL phenotype.