October 2023 – Clinical Microbiology

Production of a carbapenemase.

Acquisition of mecA gene leading to production of altered penicillin binding protein (PBP2a) is incorrect because acquisition of the gene mecA leads to the production of an altered penicillin binding protein (PBP2a), which is associated with methicillin resistant Staphylococcus aureus (MRSA). PBP2a binds less avidly to oxacillin (and methicillin), leading to resistance.

Change in cell wall peptidoglycan architecture from D-Ala-D-Ala to D-Ala-D-Lac is incorrect because changes in cell wall peptidoglycan architecture from D-Ala-D-Ala to D-Ala-D-Lac are found in vancomycin-resistant Enterococcus faecium and are often due to mutations in the vanA gene cluster.

Induction of erm methyltransferase is incorrect, because induction of erm methyltransferase leads to methylation of RNA polymerase and macrolide resistance. This induction is associated with inducible clindamycin resistance in staphylococci and streptococci illustrated by the D-test.

Modification in lipopolysaccharide (LPS) structure, altering cell envelope permeability is incorrect because modifications in lipopolysaccharide (LPS) structure alter cell envelope permeability affecting a range of antimicrobial susceptibilities, particularly last resort antimicrobials such as polymyxins in gram-negative bacteria.

The prevalence of carbapenemases has been increasing with the rise in use of carbapenem antimicrobials (i.e., imipenem, ertapenem, meropenem). Carbapenems have become the drug of choice for treating bacteria causing multidrug-resistant gram-negative bacterial infections. Carbapenem resistance has been the result of rising prevalence of extended-spectrum β-lactamase (ESBL)-producing Enterobacterales. The first carbapenemase, NmcA, an Ambler class A carbapenemase (Naas & Nordmann, 1994), was reported in 1993 and since that time a wide array of carbapenemases have been identified in Enterobacterales and non-fermenting gram-negative bacteria, such as Acinetobacter and Pseudomonas aeruginosa. Carbapenemases are typically plasmid encoded and are easily transmitted; thus, patients with isolates positive for carbapenemases are placed on strict infection control precautions such as contact precautions when in hospitals or other healthcare settings (van Loon et al., 2018). Carbapenem-resistant Enterobacterales (CREs) are classified by the CDC as an “urgent” public health threat (CDC, 2019).

The most clinically significant carbapenemase is KPC (Ambler class A), which hydrolyzes a broad range of β-lactams while being inhibited by clavulanic acid and tazobactam (Nordmann, et al., 2012). Class B carbapenemases (e.g., IMP, NDM) exhibit a broad spectrum of activity against all penicillins, cephalosporins, and carbapenems, resistance to available β-lactam inhibitors, and susceptibility to aztreonam and metal ion chelators, as the mechanism of action is dependent on zinc ions in the active site. Class D carbapenemases (OXA) are penicillinases capable of hydrolyzing oxacillin that are poorly inhibited by clavulanic acid and ethylenediaminetetraacetic acid (EDTA) with variable carbapenemase activity. As an alternative to carbapenemases, carbapenem resistance can be due to changes in bacterial permeability or increased efflux (Li, et al., 2015; Szabó et al., 2006). Mechanistic differences in carbapenem resistance present challenges in patient management, infection control precautions, and diagnostic testing. Given the rise in carbapenem resistance, the Clinical and Laboratory Standards Institute (CLSI) lowered the breakpoints of carbapenem for better detection of carbapenemase-producing isolates in 2010 (CLSI, 2020; Hombach et al., 2012). However low-level resistance and even susceptibility to carbapenems have been observed for some producers of carbapenemases, although resistance against both carbapenems was seen in this case.

Carbapenemase activity is commonly detected using the CarbaNP test, which can rapidly detect carbapenem hydrolysis by carbapenemases within two hours. Hydrolysis of the carbapenem acidifies the medium, which results in a color change of the pH indicator. This method can detect any carbapenemases. CarbaNP testing for carbapenemases should be performed for any Enterobacterales or P. aeruginosa isolate with decreased carbapenem susceptibility. This phenotypic test only detects transmissible carbapenem resistance (plasmid mediated), and has a high specificity and sensitivity (>95%, Dortet et al., 2015; Poirel & Nordmann, 2015).

Phenotypic testing of the P. rettgeri was positive for carbapenemase by CarbaNP. P. rettgeri, of the order Enterobacterales, is a gram-negative bacterium that can produce inducible AmpC-β-lactamases, plasmid-mediated ESBLs, and IMP carbapenemases. Treatment regimens depend on the susceptibility of the isolate (Walters et al., 2018).

PCR for KPC, NDM, OXA-48-like, and VIM carbapenemases performed at Mayo Clinic was negative. Due to the discrepancy between phenotypic (CarbaNP) and genotypic tests for carbapenemases, the isolate was sent to the Minnesota Department of Health, where it was found to be positive by PCR for IMP carbapenemase. In part due to the rising incidence of IMP carbapenemases (Lowe et al., 2020; Woodworth et al., 2018), Mayo Clinic has begun offering the Cepheid Xpert Carba-R carbapenem resistance assay, a molecular PCR for the rapid detection and differentiation of bacteria producing the KPC, NDM, VIM, and OXA carbapenemases.

This patient remained asymptomatic during his hospitalization and was placed on contact precautions. His hospital course included 2 days of cefdinir followed by 2 days of cefepime, after which he was discharged on a 7-day course of trimethoprim-sulfamethoxazole. The A. faecalis isolate, which was resistant only to trimethoprim-sulfamethoxazole was covered by the cephalosporin therapy.

References

1. CDC, A. (2019). Antibiotic resistance threats in the United States. US Department of Health and Human Services: Washington, DC, USA.
2. CLSI. (2023). Performance Standards for Antimicrobial Susceptibility Testing. Clinical and Laboratory Standards Institute, 33^rd ed. CLSI supplement M100. 
3. Dortet L, Agathine A, Naas T, Cuzon G, Poirel L, Nordmann P. Evaluation of the RAPIDEC® CARBA NP, the Rapid CARB Screen® and the Carba NP test for biochemical detection of carbapenemase-producing Enterobacteriaceae. J Antimicrob Chemother. 2015;70(11):3014-3022. doi:10.1093/jac/dkv213
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7. Naas T, Nordmann P. Analysis of a carbapenem-hydrolyzing class A beta-lactamase from Enterobacter cloacae and of its LysR-type regulatory protein. Proc Natl Acad Sci U S A. 1994 Aug 2;91(16):7693-7. doi:10.1073/pnas.91.16.7693. PMID: 8052644; PMCID: PMC44468.
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