New Trimodal Phenotypic Reporter of Extended-Spectrum β-Lactamase Activity.

Abstract

Bacterial resistance to β-lactam antibiotics continues to grow as misadministration presents evolutionary pressure that drives bacteria to develop improved resistance enzymes. Known as extended-spectrum β-lactamases (ESBLs), these enzymes are capable of hydrolyzing advanced β-lactam antibiotics such as third-generation (and higher) cephalosporins. Phenotypic detection substrates can be used to rapidly identify a cultured patient sample prior to confirmation by more exhaustive but slower means, critically aiding in the antibiotic stewardship essential in maintaining the effectiveness of not only the cephalosporins but also indirectly the carbapenems, our last-resort β-lactams. To enhance the phenotypic detection arsenal, we have designed an ESBL detection substrate that releases a glucose molecule upon β-lactamase hydrolysis. Because many forms of detection for glucose exist, the substrate enables ESBL quantification via three modalities commonly found in the clinical laboratory: optical absorbance, for use with the most common microbiology platforms; fluorescence, for enhanced sensitivity; and electrochemistry, which offers the potential for integration into a hand-held platform similar to a personal glucometer. Moreover, we demonstrate that, as opposed to currently available phenotypic detection substrates, our new substrate is engineered to be resistant to older and narrower β-lactamases, thus enabling specific identification of newer and more dangerous ESBLs.