Abstract
A biochemical test (NitroSpeed-Carba NP test) was developed to identify carbapenemase production in Enterobacterales and to discriminate between the different types of clinically significant carbapenemases (Ambler classes A, B, and D). It is based on two main features, namely, the hydrolysis by all β-lactamases, including carbapenemases of the nitrocefin substrate, and the capacity of ertapenem to prevent this hydrolysis for all β-lactamases except carbapenemases. Specific carbapenemase inhibitors of class A (avibactam, vaborbactam), class B (dipicolinic acid), and class D (avibactam) were used to inhibit the nitrocefin hydrolysis and to allow the identification of the carbapenemase types with a turnaround time of ca. 30 min. The test was evaluated with a collection of 248 clinical enterobacterial isolates, including 148 carbapenemase producers and 100 non-carbapenemase producers. Its overall sensitivity and specificity were 100% and 97%, respectively, including detection of all types of OXA-48-like carbapenemases. For the detection of the carbapenemase type, including strains that produce double carbapenemases, the sensitivity was 100%, 97%, and 100% for the detection of classes A, B, and D, respectively. This easy-to-implement test may contribute to optimization of the choice of the β-lactam/β-lactamase inhibitor combinations for treating infection due to carbapenemase producers.
Highlights
A biochemical test (NitroSpeed-Carba NP test) was developed to identify carbapenemase production in Enterobacterales and to discriminate between the different types of clinically significant carbapenemases (Ambler classes A, B, and D)
The main clinically important carbapenemases identified in clinical isolates are grouped into three different classes according to their amino acid identity, corresponding to molecular class A (e.g., Klebsiella pneumoniae carbapenemase [KPC] enzymes), molecular class B (e.g., New Delhi metallo--lactamase [NDM], Verona integron-encoded metallo--lactamase [VIM], and imipenemase [IMP] enzymes), and class D (e.g., OXA-48 and its derivatives) (9, 11)
Most of the phenotype-based techniques suffer from some specificity and sensitivity issues but are time-consuming and lack guidance regarding the specific carbapenemase being produced; they are poorly adapted to the clinical need for isolating patients rapidly to prevent nosocomial outbreaks (12, 13)
Summary
A biochemical test (NitroSpeed-Carba NP test) was developed to identify carbapenemase production in Enterobacterales and to discriminate between the different types of clinically significant carbapenemases (Ambler classes A, B, and D). Four isolates producing enzymes that are not detected by using the Carba NP test (namely, CTX-M-33, SHV-38, and overexpressed OXY -lactamase from K. oxytoca) were tested. One hundred non-carbapenemase producers were tested that included ESBL and non-ESBL (n ϭ 50) producers of plasmid-mediated AmpC enzymes, and isolates combining production of their chromosomally encoded AmpC together with a decreased membrane permeability (n ϭ 49) were included as well as a single -lactamase-negative E. coli reference strain ATCC 25922 (Table 1).
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