MDR Gram Negative Sepsis and CREs

β-lactam antibiotics (beta-lactam antibiotics) are antibiotics that contain a beta-lactam ring in their chemical structure. This includes penicillin derivatives, cephalosporins and cephamycins, monobactams (aztreonam), carbapenems and carbacephems. These β-lactam antibiotics are cornerstone of treatment of gram-negative bacterial (GNB) sepsis. However, resistance to β-lactam antibiotics is now not uncommon and is major cause of sepsis deaths. 

What is single most important mechanism of resistance in Gram Negative Bacteria’s?

Gram-negative resistance has spread worldwide via a variety of mechanisms. The most problematic mechanism of resistance is production of beta-lactamase enzymes by these bacteria. 

There is variety of beta-lactamase enzymes however three broad groups are particularly problematic. These are –

• [1] AmpC enzymes
• [2] Extended-spectrum β-lactamases (ESBLs)
• [3] Carbapenemases

These are the primary cause of resistance to beta-lactam antibiotics in GNB.

How beta-lactamase enzymes are classified?

Ambler classification of β-lactamases by main antibacterial substrate. 

What is mean by (CRE) Carbapenem-Resistant Enterobacterales?

The CDC defines CRE (Carbapenem-Resistant Enterobacterales) as members of the Enterobacterales which are resistant to at least one carbapenem antibiotic or producing a carbapenemase enzyme. For practical purposes, CRE refers to organisms displaying resistance to either meropenem or imipenem, or those Enterobacterales isolates producing carbapenemase enzymes.

Are all CREs due to carbapenemase production?

No, CRE comprise a heterogenous group of pathogens encompassing multiple mechanisms of resistance, broadly divided into these two groups.

[1] Not carbapenemase-producing.

CRE that are not carbapenemase-producing may be the result of amplification of non-carbapenemase β-lactamase genes (e.g., ESBL genes) with concurrent outer membrane porin disruption. 

Carbapenem resistance in Enterobacterales can be due to other mechanisms such as porin mutations and efflux pump upregulation.

[2] Carbapenemase-producing.

Carbapenem resistance is particularly serious given that carbapenems are often the last resort in treating infections resistant to other drugs.

What are the three most common and clinically important types of Carbapenemase producing CREs?

A wide range of enzymes have been identified among carbapenemase-producing Enterobacterales however following three types are commonly isolated. 

• [1] Klebsiella pneumoniae carbapenemase (KPC)
• [2] Metallo-β-lactamase (MBL) including New Delhi MBL (NDM) or Verona integron-encoded MBL (VIM), imipenemase (IMP)
• [3] OXA-48-like carbapenemases

KPC, NDM and OXA-48 enzymes are among the carbapenem resistance mechanisms of greatest concern.

How to detect whether the bacterial strain is carbapenemase-producing strain or not? If yes then which specific subtype of carbapenamase?

Detection of carbapenemase enzyme production is very crucial for early initiation of targeted definitive therapy among carbapenem resistant organism infections. Various phenotypic and gene based diagnostic tests are available for carbapenemase detection, however these tests fails to detect non- carbapenemase producing chromosomal mechanisms like poring loss, efflux pumps or alteration of membrane permeability.

Phenotypic tests: Phenotypic methods determine the presences of carbapenemase enzymes by hydrolysis of imipenem or meropenem drugs by culture isolates. Some phenotypic method (Carba NP) can be performed on clinical samples, detecting calorimetric positive signals in less than an hour. The Clinical and Laboratory Standards Institute (CLSI) has formulated recommendations and guidelines for detecting CREs based on phenotypic methods as Carba NP test, Carbapenem Inactivation Method (CIM) and modified CIM (m-CIM).

Numerous rapid immunochromatographic assays are available for detection of carbapenemase based on its reaction with labelled anti-carbapenemase monoclonal antibodies. This assay is required to be performed on fresh bacterial growth. Its sensitivity and specificity varies between 88-100 % and 95-100 % respectively.

Molecular testing can identify specific carbapenemase gene families (e.g., differentiating KPC from OXA-48-like genes). Molecular technique like Qualitative Real Time – PCR is also commonly used to detection of most common five carbapenemases (KPC, IMP, VIM, NDM and OXA-48 like variants) with the short turnaround time of less than 24hrs and with a sensitivity of 80-100%. Microarray techniques are more sensitive then PCR at identifying a large number of target genes for simultaneous detection of organism and resistant indicators. Whole Genome Sequencing (WGS) improves understanding about chromosomal mutations linked to resistance and acquired resistant determinants. In recent years, WGS had a substantial impact over Multi-Drug Resistance (MDR) genetic level testing and surveillance.

Rapid gene based assay have an advantage of early reporting about MDR status of an organism by avoiding the need for culture isolate and allowing it directly over samples, which in turns support to start targeted therapy and lowers the risk of treatment failure associated with empiric antimicrobial therapy.

It is important to consider that most of the commercially available diagnostic panels focus only on five targets of carbapenemase encoding genes (KPC, IMP, VIM, NDM, and OXA-48 like). As a result, the negative result does not always suggest an organism to be susceptible to carbapenem; which could be due to presence of other mechanisms like alteration of membrane permeability, porin loss or efflux pump. Even, the presence of a gene does not always indicates carbapenem resistance as it also depends upon the percentage expression of the resistant gene in that organism.

Tell us some salient points about carbapenamase?

[1] New Delhi metallo-β–lactamase (NDM) and coproduction of NDM with Oxacillinase-48 like (OXA-48–like) enzymes are the most predominant mechanisms of carbapenem-resistant Enterobacteriaceae (CRE) infections in India.

[2] KPC is most common CRE in USA.

[3] Plasmids carrying carbapenemase genes like NDM-1 are diverse and can harbor a high number of additional resistance genes (e.g., ESBL-alleles) as well as other carbapenemase genes like Oxacillinase-48 types, VIM types, and so forth, as the source of multidrug resistance in one single bacteria

[4] OXA-48 gene for carbapenem resistance has been found in ESBL producers especially those harboring CTX-M.

What is role of beta-lactamase inhibitors?

Beta-lactamase inhibitors are drugs that are co-administered with beta-lactam antimicrobials to prevent antimicrobial resistance by inhibiting serine beta-lactamases.

For the most part, beta-lactam inhibitors, such as clavulanic acid, tazobactam and sulbactam, are inactive against carbapenemases.

Avibactam is a relatively newer beta-lactam inhibitor which inhibits AmpC beta-lactamases, ESBLs and the serine carbapenemases, including Klebsiella pneumoniae carbapenemases (KPCs) and some OXA carbapenemases, but is not active against the metallo-beta-lactamases.

What are newer beta-lactam antibiotic and beta lactamase inhibitor combination and their activity against CREs?

	KPC	NDM	OXA-48 like
Ceftazidime-avibactam	Active	Not Active	Active
Meropenem-vaborbactam	Active	Not Active	Not Active
Imipenem-cilastatin-relebactam	Active	Not Active	Not Active

Neither ceftazidime-avibactam, meropenem-vaborbactam, nor imipenem-cilastatin-relebactam have activity against metallo-β-lactamase (e.g., NDM) producing Enterobacterales.

What is inhibitory activity of NDM? What is 1st choice of treatment when NDM is detected in clinical isolates?

NDMs are a type of metalloproteinases whcih hydrolyze penicillins, cephalosporins, and carbapenems, but not aztreonam. Although aztreonam is active against NDMs, it can be hydrolyzed by ESBLs, AmpC β-lactamases, KPCs, or OXA-48-like carbapenemases which are frequently co-produced by NDM-producing isolates. Avibactam generally remains effective at inhibiting the activity of these latter β-lactamase enzymes.

A combination of Ceftazidime/avibactam + aztreonam is considered optimal for NDM strains. 

References:

1. Matteo Bassetti, Javier Garau, Current and future perspectives in the treatment of multidrug-resistant Gram-negative infections, Journal of Antimicrobial Chemotherapy, Volume 76, Issue Supplement_4, November 2021, Pages iv23–iv37, https://doi.org/10.1093/jac/dkab352
2. Tamma PD, Aitken SL, Bonomo RA, Mathers AJ, van Duin D, Clancy CJ. Infectious Diseases Society of America Antimicrobial-Resistant Treatment Guidance: Gram-Negative Bacterial Infections. Infectious Diseases Society of America 2023; Version 3.0. Available at https://www.idsociety.org/practice-guideline/amr-guidance/.
3. Bassetti M, Peghin M, Vena A and Giacobbe DR (2019) Treatment of Infections Due to MDR Gram-Negative Bacteria. Front. Med. 6:74. doi: 10.3389/fmed.2019.00074

Article Compiled by:

Dr Sujeet Kumar, Associate Professor, Hemat-oncology and BMT, HBCH Varanasi

Dr Rahul Sarode, Associate Professor, Microbiology, HBCH Varanasi

 

Carbapenem-Resistant Enterobacterales (CRE) – A thread
What are CRE?
CRE (Carbapenem-Resistant Enterobacterales) are members of the Enterobacterales which are resistant to at least one carbapenem antibiotic or which produce a carbapenemase enzyme.

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