Molecular diversity of extended-spectrum β-lactamases and carbapenemases, and antimicrobial resistance

Abstract

Along with the recent spread of multidrug-resistant bacteria, outbreaks of extended-spectrum β-lactamase (ESBL) and carbapenemase-producing bacteria present a serious challenge to clinicians. β-lactam antibiotics are the most frequently used antibacterial agents and ESBLs, and carbapenemases confer resistance not only to carbapenem antibiotics but also to penicillin and cephem antibiotics. The mechanism of β-lactam resistance involves an efflux pump, reduced permeability, altered transpeptidases, and inactivation by β-lactamases. Horizontal gene transfer is the most common mechanism associated with the spread of extended-spectrum β-lactam- and carbapenem resistance among pathogenic bacterial species. Along with the increase in antimicrobial resistance, many different types of ESBLs and carbapenemases have emerged with different enzymatic characteristics. For example, carbapenemases are represented across classes A to D of the Ambler classification system. Because bacteria harboring different types of ESBLs and carbapenemases require specific therapeutic strategies, it is essential for clinicians to understand the characteristics of infecting pathogens. In this review, we summarize the current knowledge on carbapenem resistance by ESBLs and carbapenemases, such as class A carbapenemases, class C extended-spectrum AmpC (ESAC), carbapenem-hydrolyzing class D β-lactamases (CHDLs), and class B metallo-β-lactamases, with the aim of aiding critical care clinicians in their therapeutic decision making.

Keywords: Carbapenemase; Classification; Multidrug resistance; β-Lactam; β-Lactamase.

Fig. 1

The antimicrobial action of β-lactamase against the peptidoglycan structure of the bacterial cell membrane. Peptidoglycan possesses a basic structural unit in which two amino sugars of N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) alternate, and a longitudinal peptide chain linked to NAM forms a pillar. This peptide chain is a pentapeptide consisted of alternating d- and l-forms of l-alanine, γ-d-glutamic acid, l-lysine, d-alanine, and d-alanine that forms a bacterial enzyme known as penicillin-binding protein (PBP). PBP recognizes alanyl-alanine, which is an alanine dimer formed by d-alanine-d-alanine present at the end of the pentapeptide, and exerts the enzymatic action of cross-linking. Since penicillin is structurally similar to alanyl-alanine at the terminal region of the pillar structure, PBP captures penicillin, thereby inhibiting the cross-linking reaction

Fig. 2

The genes encoding penicillin-binding proteins in P. aeruginosa PAO1. Eight types of PBP genes and three chromosomal β-lactamase genes for PIB-1 (class A), AmpC (class C), and PoxB (class D) exist in the genome of P. aeruginosa reference strain PAO1 [15]

Fig. 3

The enzymatic action of a β-lactamase to penicillins. The β-lactam ring is the basic structure of β-lactam antibiotics. β-lactamase inhibits the antibacterial action by dissociating the –CO–NH structure of the β-lactam ring. The –CO–NH structure of the β-lactam ring is similar to the –CO–NH peptide in alanyl-alanine, from which the peptidoglycan cross-links are formed. Thus, β-lactams mimic alanyl-alanine of the peptidoglycan pillar structure. Thus, β-lactamase is a protease (peptidase) that dissociates the –CO–NH structure of β-lactams

Fig. 4

The chemical structure of β-lactam antimicrobials. Clinically used β-lactams can be classified into five basic structures capable of exerting different antimicrobial activities

Fig. 5

The classification of β-lactamases. Molecular structure classification using the Ambler method [16] and functional classification using the Bush-Jacobi-Medeiros method [17, 18] have both been employed. In the Ambler classification, β-lactamases are grouped into four classes A, B, C, and D by motifs composed of primary sequences constituting the protein molecules. β-lactamases of classes A, C, and D use a serine at the enzyme active center, whereas β-lactamases of class B use metal zinc ions. In functional classification using the Bush-Jacobi-Medeiros method, β-lactamases are classified into groups 1 to 3 depending on the degradation of β-lactam substrates and the effect of the inhibitor

Fig. 6

A phylogenetic tree of class A carbapenemases. A phylogenetic tree of six types of class A capbapenemases: GES (Guiana extended-spectrum β-lactamase), SME (Serratia marcescens enzyme), SHV-38 (sulfhydryl variable lactamase), KPC (Klebsiella pneumoniae carbapenemase), IMI/NMC-A (imipenemase/non-metallocarbapenemase-A), and SFC-1 (Serratia fonticola carbapenemase). Based on the primary sequences, the tree was generated using Clustal Omega [–34] of GenomeNet at Kyoto University Bioinformatics Center (https://www.genome.jp/tools-bin/clustalw)

Fig. 7

A phylogenetic tree of the class D β-lactamase OXA family (oxiacillinase). A phylogenetic tree of the OXA types of β-lactamases. OXA-10, OXA-11, and OXA-15 are recognized as extended-spectrum cephalosporinases, whereas OXA-23 and OXA-48 are recognized as carbapenem-hydrolyzing class D β-lactamases (CHDLs). Based on the primary sequences, the tree was generated using Clustal Omega [–34] of GenomeNet at Kyoto University Bioinformatics Center (https://www.genome.jp/tools-bin/clustalw)

Fig. 8

The genomic structure of the class 1 integron of P. aeruginosa NCGM2.S1. The intl gene, which encodes integrase Int1, catalyzes recombination between the attl1 site of the integron and the attC site of an antimicrobial gene cassette [56]. In multidrug-resistant P. aeruginosa NCGM2.S1 [58], three antimicrobial gene cassettes (blaIMP-1, aac(6′)-1ae, and aadA1) are integrated into the region between attI and attC. The Int1 integrase gene, the attI integration site, and qacE partially deleted in the sulI gene, which encodes resistance to sulfonamide are present

Fig. 9

A phylogenetic tree of the class B metallo-β-lactamase family. MBLs can be classified into three subclasses (B1, B2, B3) based on their amino acid sequences. Subclasses B1 and B3 are characterized by two Zn²⁺ molecules in the enzyme active center (Zn₁, Zn₂), implying a broader range of substrates, whereas target substrates of subclass B2, which have a single Zn²⁺ at the active center, are narrower in range than those of classes 1 and 3 [54]. Based on the primary sequences, the tree was generated using Clustal Omega [–34] of GenomeNet at Kyoto University Bioinformatics Center (https://www.genome.jp/tools-bin/clustalw)

Fig. 10

The classification of carbapenemases. Carbapenemases is represented in all classes, A to D, of the Ambler classification system [16]. Functional classification, using the Bush-Jacobi-Medeiros method [17, 18], indicated that the class A carbapenemases were represented by GES and KPC. The β-lactamases belonging to class C, which function as cephalosporinases, are encoded by the AmpC gene carried on the chromosome of many Enterobacteriaceae. ESAC enzymes are known as ESACs. In class D, OXA enzymes, which were originally oxacillinases, have mutated to become CHDLs. Class B β-lactamases are characterized by possessing a metal Zn²⁺ as the enzyme activity center. IMP- and VIM-type β-lactamases are the main MBLs that fit into the integron structure. MBLs can be classified into three subclasses (B1, B2, B3) based on their amino acid sequence. Subclasses B1 and B3 are characterized by two zinc molecules (Zn₁, Zn₂) in the enzyme active center, demonstrating more extensive substrate degradation, while subclass B2 has a single Zn²⁺ at the active center and displays a narrower spectrum