A common mechanism of antibiotic resistance

By Swati Kumar, MD

Antibiotic resistance is a significant problem encountered in the treatment of infections with both Gram-positive and Gram-negative bacteria. A brief discussion of one of the most common mechanisms of resistance – production of beta-lactamases (BLs) – is presented here.

Beta-lactamases are enzymes that hydrolyse beta-lactam antibiotics (penicillins, cephalosporins, monobactams and carbapenems). They are of several kinds depending on their characteristics, such as which antibiotics they lyse and other properties summarized in Table 1. As shown in the table, they are grouped into classes A, B, C and D. Classes A and D include BLs that range from narrow spectrum BLs, which lyse only a narrow spectrum of drugs (such as E. Coli resistant to only amplicillin but susceptible to other antibiotics), to extended spectrum beta-lactamases (ESBLs), that lyse a broad spectrum of drugs including not only pencillins but also extended spectrum (third generation) cephalosporins. ESBLs are defined as BLs that confer resistance to penicillins, first-, second- and third-generation cephalosporins and aztreonam, but not to cephamycins (cefoxitin) and carbapenems, and that are inhibited by beta-lactamase inhibitors (BLIs) (such as clavulanate, sulbactam, tazobactam). ESBLs are produced by various Gram-negative bacteria, but most commonly Klebsiella species and E Coli. Class B BLs hydrolyse penicillins, cephalosporins and carbapenems, are not inhibited by BLIs and are classically produced by Klebsiella pneumoniae. Class C enzymes, also known as ampC BLs, are similar to ESBLs in their spectrum of resistance in that they both spare carbapenems, but ampC BLs can be distinguished from ESBLs by their lysis of cephamycins and resistance to BLIs. In addition, ampC BLs are associated with inducible resistance. So, pathogens producing these enzymes may not display phenotypic resistance initially but do so after exposure to certain antibiotics such as ampicillin, third-generation cephalosporins and imipenem, which lead to expression of resistance.

Diagnosis

Recognizing the presence of resistance due to ESBL production or ampC BL production is important because of its implications on therapy of these Gram-negative infections. Not all microbiology labs perform testing to specifically detect the production of these enzymes. One way of figuring out which, if any, of these various groups of BLs are present in a Gram-negative isolate is to look at the susceptibility report. Thus:

• Resistance to ampicillin alone suggests the presence of a narrow spectrum BL.
• Resistance to pencillins, first-, second- and third-generation cephalosporins and aztreonam, but not to carbapenems and cephamycins (cefoxitin), suggests the presence of ESBLs.
• Resistance to penicillins, first-, second- and third-generation cephalosporins, aztreonam and cephamycins, but not to carbapenems, suggests ampC BLs.
• Resistance to all beta-lactams including carbapenems, but not to monobactams, suggests carbapenemases.

In addition, ESBLs are by definition susceptible to inhibition by beta-lactamase inhibitors, and therefore, the pathogen will show susceptibility to beta-lactam-BLI combinations (such as ampicillin-sulbactam, ticarcillin-clavulanate, piperacillin-tazobactam) in vitro (except when another mechanism conferring resistance to these combinations is concurrently present in the same isolate). Whereas, AmpC BLs are resistant to inhibition by BLIs and therefore will confer resistance to antimicrobials containing BLIs.

Therapy implications

The drugs of choice for treatment of serious infections with ESBL-producing organisms are carbapenems. Beta-lactam-BLI combinations, although usually susceptible in vitro, have been associated with therapeutic failures in vivo. Reasons for this discrepancy include:

1) The inoculum effect – the minimum inhibitory concentrations of these drugs rise with increasing inoculum of the pathogen, so that susceptibility as indicated by in vitro testing using a standard inoculum may not necessarily hold true in clinical situations with much higher loads of bacteria.

2) Production of multiple ESBLs (versus a single ESBL) by an isolate, which lowers the effectiveness of these drugs. 

More importantly, third-generation cephalosporins should not be used for treatment of these infections even if they appear to be susceptible in vitro. Variablity in hydrolysis of the various third-generation cephalosporins can lead to some drugs being reported as susceptible and others resistant (such as cefotaxime S, ceftazidime R). Treatment with susceptible-appearing cephalosporins has been associated with therapeutic failures and thus these drugs should not be used, regardless of their susceptibility on the antibiogram. Similarly, cephamycins (cefoxitin) should not be used for treating these pathogens.

Treatment of ampC BL producers is complicated by the lack of apparent resistance on initial susceptibility testing (before drug exposure), and thus use of third-generation cephalosporins for the treatment of an apparently susceptible isolate. This initial exposure of the patient to antibiotics known to induce expression of the normally suppressed resistance leads to subsequent failure of therapy. Isolates obtained during this phase now show phenotypic resistance to all beta-lactams except carbapenems. Knowledge of bacteria that are known to be associated with this mechanism of resistance (Enterobacter, Citrobacter, Serratia, Pseudomonas and Morganella) should prevent continued use of third-generation cephalosporins once the lab reports one of these Gram-negatives as being present, even if the initial testing reported susceptibility to these drugs.

Table 1 - Classification of beta-lactamases
Enzyme Class	A, D	B	C
Substrate (drugs lysed)	Variable, often many groups of beta-lactams	All beta-lactams except monobactams	All but carbapenems
Enzymes	Include Gram + beta-lactamases, G-ve beta-lactamases including, narrow spectrum beta-lactamases, ESBL and carbapenemases	Carbapenemases	AmpC lactamases
Chromosomal/Plasmid	Plasmid	Chromosomal/Plasmid	Chromosomal/Plasmid
Organisms	G+ and -ve	G-ve	G-ve
Clavulanate Inhibition	Yes	No	No
Inducible	No	No	Yes
Prototype Pathogens	E. Coli, Klebsiella and other Gram-negatives, Staphylococcus aureus	Klebsiella	Enterobacter species, Serratia, Pseudomonas, Morganella and Citrobacter

Swati Kumar, MD, is a pediatric infectious disease specialist at Children's Hospital of Wisconsin. She also is an assistant professor of Pediatrics (Infectious Diseases) at the Medical College of Wisconsin and a member of Children's Specialty Group.

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