Genotypic detection of the bla_CTX-M-1 gene among extended-spectrum β-lactamase-producing Enterobacteriaceae

Extended-spectrum β-lactamases (ESBLs), a group of β-lactamase enzymes produced by bacteria in the family Enterobacteriaceae, are becoming a major problem in the healthcare community worldwide. Although many attempts have been made in the detection of ESBL-producing bacteria, the cost and speed of detection remains an important challenge. Therefore, this study aimed to develop a rapid, effective and affordable method for detection of the bla_CTX-M-1 ESBL gene by a loop-mediated isothermal amplification (LAMP) technique.

Clinical ESBL-producing Enterobacteriaceae, including Escherichia coli and Klebsiella pneumoniae, were isolated and were used as representative strains. The double-disk synergy method was performed to detect ESBL-producing Enterobacteriaceae. Performance of the LAMP method in the detection of bla_CTX-M-1 was compared with conventional PCR in terms of sensitivity and specificity.

The developed LAMP method efficiently identified the presence of the bla_CTX-M-1 gene in ESBL-producing Enterobacteriaceae. It provided similar results to conventional PCR, but the LAMP technique required only 20min of testing time. The accuracy of the LAMP method was confirmed by restriction digestion, which showed the predicted size of the bla_CTX-M-1 gene. In addition, the developed method was comparable with PCR that amplified only the target bla_CTX-M-1 gene in terms of specificity, but LAMP was ca. 1000-fold more sensitive than PCR.

A rapid assay to detect ESBL-producing Enterobacteriaceae by a LAMP technique was developed in this study. The developed method is sensitive and suitable for rapid screening of bla_CTX-M-1 in routine laboratories with limited resources.