β-Lactam Antibiotics

In gram-positive organisms, capsular materials may hinder access to the cytoplasmic membrane, but this rarely limits the diffusion of the cell-wall inhibitors. Gram-negative bacteria have a restricting sieving mechanism (porins) in their outer membranes (external cell wall), which reduces the penetration of several types of antibiotics. Different species of gram-negative bacteria exhibit varying permeability barriers to β-lactam antibiotics, and these impair access of the antibiotics to the membrane-associated binding proteins. For example, the permeability barrier of Haemophilus influenzae is readily crossed by β-lactam antibiotics, Escherichia coli presents a greater obstacle to these agents, and the outer membranes of Pseudomonas aeruginosa are penetrated with great difficulty by most β-lactam compounds. Penicillins, aminopenicillins, first- and second-generation cephalosporins, and selected other β-lactams cannot penetrate the outer membrane of P aeruginosa. In addition, porins are frequently associated with efflux proteins that effectively remove drug that has successfully penetrated the lipopolysaccharide covering of gram-negative organisms.

The chemical nature of β-lactams (penicillins, cephalosporins, and the β-lactamase inhibitors), as well as their concentration gradients, also greatly influence their penetration of bacteria to their targets at the surface of the cytoplasmic membrane, giving rise to the differences between antibacterial spectra of the various classes of penicillin. β-Lactams are often used in combination with other antibiotics that disrupt the integrity of the membranes and thereby facilitate access by β-lactams. The genetic loci controlling permeability generally have been considered to be chromosomally located, but they also may be plasmid-specified genes.

The most important mechanism of bacterial resistance to β-lactam antibiotics is enzymatic inactivation by β-lactamases by cleavage of the 4-member β-lactam ring. Cleavage results in the inability of the drugs to bind to the target PBPs. There currently are >800 different β-lactamases, representing six major classes, with the enzyme varying with the organism and drugs targeted varying with the enzyme. The increase in the number of enzymes reflects, in part, pressure brought with the increasingly widespread use of β-lactams and the continued manipulation of the drugs in an attempt to circumvent bacterial β-lactamase production. For example, the addition of larger R groups on the β-lactam structure rendered cephalosporins to be resistant to penicillinases. However, cephalosporinases emerged with continued use of first-generation cephalosporins. Second- and third-generation cephalosporins reflect modifications, including larger R groups that hindered β-lactamase access to the β-lactam ring. Inhibitors of β-lactamases (clavulanic acid, sulbactam) were added to minimize penicillin destruction. As a result, newer β-lactamases emerged. Approval and use of third-generation cephalosporins have been associated with emergence of extended-spectrum β-lactamases (ESBLs), particularly by E coli, Klebsiella, and Proteus, that target third-generation cephalosporins (but not cephamycins such as cefoxitin). In contrast, carbapenems (imipenem and meropenem) are not subject to ESBLs that target third-generation cephalosporins, but they are subjected to carbapenemases. β-Lactamases do not discriminate among the drugs within class, meaning both human and veterinary drugs will be targeted. Interestingly, clavulanic acid is not susceptible to ESBLs; susceptibility data indicating resistance to cephalosporins but susceptibility to amoxicillin-clavulanic acid indicates ESBL formation.

β-Lactamases are produced by both gram-positive (Staphylococcus aureus, S epidermidis, S pseudintermedius but generally not enterococci) and gram-negative organisms. Some of these enzymes are active exclusively against penicillins, others are principally active against cephalosporins, and several types hydrolyze both equally. The type and concentration of β-lactamases are also specific to bacterial species. Gram-positive β-lactamases generally are excreted into the external environment as exoenzymes, produced in large quantity, plasmid mediated (single determinant), usually inducible (rarely constitutive), unable to initiate self-transmission (rely principally on transduction), and are active primarily against penicillins. Staphylococcal strains are the main gram-positive bacteria in which β-lactamase resistance develops, often very quickly. Gram-negative β-lactamases generally are heterogeneous (wide range), retained within the periplasmic space, produced in small quantity, often constitutive (less often inducible), able to initiate self-transmission (conjugation mechanisms), and active against both penicillins and cephalosporins. The impact of β-lactamase protectors such as clavulanic acid may not be as positive for treatment of gram-negative versus gram-positive organisms. Gram-negative bacteria capable of resistance as a result of β-lactamase production include Escherichia, Haemophilus, Klebsiella, Pasteurella, Proteus, Pseudomonas, and Salmonella spp; resistance may take longer to develop in some of these strains.

Resistance to β-lactam antimicrobial agents can be acquired by alterations in the PBP targets of these drugs. A loss or decrease in affinity of crucial PBP can lead to a significant increase in resistance to β-lactams. For example, resistance of enterococci to cephalosporins appears to reflect the lack of affinity of a PBP to this subclass of drugs. Changes in PBP-2 of Staphylococcus spp render the organism resistant to all β-lactams. Methicillin resistance in Staphylococcus spp reflects acquisition of the mec gene, which results in a mutation in PBP-2. As such, no β-lactam can bind to this protein, resulting in resistance to all β-lactam drugs. Problematically, genes conferring methicillin resistance may be accompanied by genes conferring multidrug resistance.

Organisms that have no cell wall, such as Mycoplasma, are intrinsically resistant to β-lactams. A phenotypic form of resistance can occur when spheroplasts (incomplete cell wall) or protoplasts (absence of cell wall) are present. These so-called “L-forms” must be present in a hyperosmotic environment (eg, the renal medulla) to survive; otherwise, they will lyse. The clinical significance of this form of resistance is unclear.