AUTHOR=Nakano Ryuichi , Yamada Yuki , Nakano Akiyo , Suzuki Yuki , Saito Kai , Sakata Ryuji , Ogawa Miho , Narita Kazuya , Kuga Akio , Suwabe Akira , Yano Hisakazu 

TITLE=The Role of nmcR, ampR, and ampD in the Regulation of the Class A Carbapenemase NmcA in Enterobacter ludwigii

JOURNAL=Frontiers in Microbiology

VOLUME=12

YEAR=2022

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2021.794134

DOI=10.3389/fmicb.2021.794134

ISSN=1664-302X

ABSTRACT=<p>Various carbapenemases have been identified in the Enterobacteriaceae. However, the induction and corresponding regulator genes of carbapenemase NmcA has rarely been detected in the <italic>Enterobacter cloacae</italic> complex (ECC). The NmcA-positive isolate ECC NR1491 was first detected in Japan in 2013. It was characterized and its induction system elucidated by evaluating its associated regulator genes <italic>nmcR</italic>, <italic>ampD</italic>, and <italic>ampR</italic>. The isolate was highly resistant to all β-lactams except for third generation cephalosporins (3GC). Whole-genome analysis revealed that <italic>bla</italic><sub>NmcA</sub> was located on a novel 29-kb putatively mobile element called EludIMEX-1 inserted into the chromosome. The inducibility of β-lactamase activity by various agents was evaluated. Cefoxitin was confirmed as a strong concentration-independent β-lactamase inducer. In contrast, carbapenems induced β-lactamase in a concentration-dependent manner. All selected 3GC-mutants harboring substitutions on <italic>ampD</italic> (as <italic>ampR</italic> and <italic>nmcR</italic> were unchanged) were highly resistant to 3GC. The <italic>ampD</italic> mutant strain NR3901 presented with a 700 × increase in β-lactamase activity with or without induction. Similar upregulation was also observed for <italic>ampC</italic> and <italic>nmcA</italic>. NR1491 (pKU412) was obtained by transforming the <italic>ampR</italic> mutant (135Asn) clone plasmid whose expression increased by ∼100×. Like NR3901, it was highly resistant to 3GC. Overexpression of <italic>ampC</italic>, rather than <italic>nmcA</italic>, may have accounted for the higher MIC in NR1491. The <italic>ampR</italic> mutant repressed <italic>nmcA</italic> despite induction and it remains unclear how it stimulates <italic>nmcA</italic> transcription via induction. Future experiments should analyze the roles of <italic>nmcR</italic> mutant strains.</p>