AmpC ENZYMES

AmpC β-lactamases are a class of enzymes produced by certain bacteria that confer resistance to a wide range of β-lactam antibiotics, including penicillins, cephalosporins (especially cephamycins), and monobactams. These enzymes hydrolyze the β-lactam ring of these antibiotics, rendering them ineffective. AmpC enzymes belong to the Ambler class C and are typically encoded on the chromosome of several Gram-negative bacteria such as Enterobacter spp., Citrobacter freundii, Serratia marcescens, and Pseudomonas aeruginosa. However, plasmid-mediated AmpC enzymes have also emerged, particularly in Escherichia coli and Klebsiella pneumoniae, which increases the potential for horizontal gene transfer and widespread resistance.

Chromosomal AmpC expression can be inducible or derepressed (constitutively expressed at high levels), especially following exposure to certain β-lactam antibiotics. In inducible organisms, the enzyme is produced at low levels but can be upregulated in the presence of β-lactam drugs, often leading to treatment failure. Plasmid-mediated AmpC enzymes, by contrast, are usually constitutively expressed and not regulated by the bacterial chromosomal mechanisms, posing additional challenges in clinical settings.

Unlike extended-spectrum β-lactamases (ESBLs), AmpC enzymes are not inhibited by traditional β-lactamase inhibitors such as clavulanic acid, sulbactam, or tazobactam. However, some newer inhibitors like avibactam can inhibit certain AmpC enzymes. Detection of AmpC-producing organisms in clinical microbiology is crucial but can be challenging, as standard susceptibility tests may not always reveal resistance, especially if inducible mechanisms are involved.

The presence of AmpC enzymes in bacteria complicates treatment options, often necessitating the use of carbapenems or non-β-lactam antibiotics. Continued surveillance, rapid diagnostic tools, and antimicrobial stewardship are essential strategies to monitor and manage the spread of AmpC-mediated resistance in both healthcare and environmental settings.

AmpC enzymes are broad-spectrum beta-lactamase enzymes that are usually encoded on bacterial chromosome, and which are active on cephamycins (e.g. cefoxitin and cefotetan) and oxyimino-β-lactam agents. They are bacterial enzymes that hydrolyze third-generation extended spectrum cephalosporins and cephamycins (e.g. cefoxitin), thus engendering antimicrobial resistance to these categories of antibiotics. AmpC strains are resistant to the cephamycins including cefoxitin and cefotetan but they are susceptible to the fourth generation cephalosporin, cefepime. High level AmpC producers cause resistance to all 1^st, 2^nd and 3^rd generation cephalosporins, the beta lactam-inhibitor drugs such as amoxicillin-clavulanic acid and the monobactams (e.g. aztreonam).

AmpC beta-lactamase enzymes began to gain importance in the health sector in the early 1970s as one of the mediators of antimicrobial resistance in Gram negative bacteria including Escherichia coli, Klebsiella speciesand Pseudomonas aeruginosa. The genes that confer or mediate the production of AmpC beta-lactamases in Gram-negative bacteria such as in members of the Enterobacteriaceae family and Pseudomonas are often chromosomally-borne (i.e. they are chromosomally encoded) rather than being plasmid encoded but these genes may still be plasmid-encoded in some bacteria.

Chromosomal AmpC beta-lactamases are usually inducible while plasmid-mediated AmpC enzymes are not inducible. Instead, plasmid-mediated AmpC enzymes arise as a result of point mutations in the AmpC genes. Chromosomal AmpC enzymes are AmpC beta-lactamases that are mediated by ampC genes – which is usually located on the chromosomal DNA of the host bacteria. Plasmid-mediated AmpC beta-lactamases are beta-lactamases produced by bacterial pathogens devoid of chromosomal ampC genes but that may have acquired resistance plasmids of the resistance traits via antibiotic selection pressure, mutation or through gene transfer mechanisms such as conjugation, transformation and transduction.

The expression of AmpC enzymes in members of the Enterobacteriaceae family such as Escherichia coli and Klebsiella species is usually low but AmpC enzymes can be overproduced in these organisms if they are exposed to the beta-lactam antibiotics especially the bet-lactamase inhibitors such as clavulanic acid – which is known to mask the production of extended spectrum β-lactamases (ESBLs) in organisms harbouring both AmpC and ESBL genes. Thus, exposure to beta-lactam antibiotics can induce the production of AmpC enzymes in a bacterium.

The AmpC β-lactamases are clinically important beta-lactamases because they confer antimicrobial resistance to the narrow-spectrum, expanded-spectrum and the broad-spectrum cephalosporins including cefotaxime, ceftazidime, ceftriaxone, aztreonam and the penicillins; and their resistance is also expressed towards the β- lactamase inhibitors such as amoxycillin-clavulanic acid. The genes encoding AmpC β-lactamases are much more frequently chromosomal than plasmid-mediated.

However, the genes for AmpC-β-lactamases can be transferred to organisms that do not harbour them (such as Klebsiella pneumoniae) through plasmids and other genetic transfer elements such as transposons. Klebsiella speciesespecially K. pneumoniae is one of the few Gram negative bacteria which do not possess a chromosomal AmpC β-lactamase, but the organism can acquire this resistant gene through the transfer of AmpC-containing plasmids from other Enterobacteriaceae harbouring it.

The chromosomally mediated β-lactamase production of AmpC enzymes by Gram negative bacteria takes place through the expression of the AmpC gene which is either constitutive or inducible. As aforementioned, most of the genera of the family Enterobacteriaceae produce AmpC enzymes through an inducible mechanism – in which case the presence of broad-spectrum antibiotics sparks the production of the enzyme in the organism. AmpC enzymes confer resistance to all classes of beta-lactams, except the carbapenems including ertapenem, meropenem and imipenem; and they are also not inhibited by clavulanic acid and other beta-lactamase inhibitors.

Though they are mostly inducible, AmpC enzymes can also be produced in high levels through mutation in Enterobacteriaceae and other Gram negative organisms especially in the presence of an inducing agent (which can be an antibiotic). The production of some multidrug resistant enzymes such as extended spectrum beta-lactamases (ESBLs) by Gram negative bacteria can be masked by the co-expression of AmpC in the same organism. AmpC production in some Gram negative bacteria including P. aeruginosa, E. coli and Klebsiella species is usually induced when these organisms are exposed to certain array of beta-lactam agents such as amoxicillin, imipenem, clavulanic acid or amoxicillin-clavulanic acid and ampicillin.

Chromosomal AmpC enzymes (which can also be called inducible AmpC enzymes) and plasmid-borne AmpC enzymes are the two main types of AmpC beta-lactamases that exist amongst bacteria especially in Gram negative organisms – in which these multidrug resistant enzymes are produced. Inducible AmpC beta-lactamases are usually seen in Citrobacter species, Morganella species, Enterobacter species and Serratia marcescens while the plasmid-mediated AmpC enzymes are seen in other enteric and non-enteric bacteria including Escherichia coli, Klebsiella species and Pseudomonas aeruginosa.

They can also be plasmid encoded; and AmpC enzymes like other extended or expanded beta-lactamases such as ESBLs and MBLs confer on pathogenic Gram positive and Gram negative bacteria the exceptional ability to be resistant to a wide array of beta-lactam drugs and non-beta-lactams. AmpC beta-lactamases are differentiated from extended spectrum beta-lactamases (ESBLs) by the ability of the former (i.e. AmpC enzymes) to hydrolyze cephamycins (e.g. cefoxitin) and their lack of inhibition by clavulanic acid. The failure to detect AmpC beta-lactamase enzymes has no doubt contributed to the uncontrolled spread of AmpC enzymes; and in most of the cases these has also contributed to the treatment failures experienced in patients infected with such organisms.

Of particular concern are the limited treatment options for infections caused by Gram negative resistant bacteria – leading to antibiotic selection pressure and consequent risk of the emergence of antibiotic resistant pathogens. Studies have shown that the onslaught of AmpC resistance represents a major challenge for physicians as these high-profile beta-lactamase hydrolyzing enzymes renders third-generation cephalosporins and the cephamycins increasingly inefficacious in the treatment of bacterial related infections caused by AmpC-producing bacteria.

The detection of AmpC beta-lactamases in bacterial isolates still remains problematic especially in those organisms that produce or have extended spectrum beta-lactamases (ESBLs); and this is due in part to the fact that ESBL-producing bacteria that also harbour genes for AmpC enzyme production mask the production of AmpC enzymes. This is why the Clinical Laboratory Standard Institute (CLSI) and other researchers recommend the use of several antimicrobial agents that also incorporates chelating agents such as ethylene diamine tetraacetic acid (EDTA) and boronic acid for the detection of AmpC enzymes from both environmental and hospital isolates.

It is even more worrisome when clinical microbiology laboratories in some hospitals fail to detect these pathogens from clinically important specimens and/or other environmental samples – since the presence of an AmpC-producing isolate may impair treatment options. Bacterial resistance to the cephalosporins (especially 3^rd-generation cephalosporins) and the cephamycins should raise a suspicion for possible production of AmpC beta-lactamases – that warrants phenotypic confirmation. The confirmation of AmpC production in clinical and/or environmental pathogens is important for the patient’s welfare because it will support the susceptibility test result by permitting the reservation of broad spectrum antibiotics (e.g. carbapenems) for more serious and complicated bacterial disease.

And such a confirmatory test coupled with the antimicrobial susceptibility test results will help to select targeted narrow spectrum antibiotics for treatment rather than using drugs with broad spectrum activity, thereby minimizing the risk of selecting for, or promoting the development of antimicrobial resistant bacteria pathogens. Treatment options for infections caused by AmpC-producing bacteria are usually limited because of the multidrug resistant nature of such organisms. However, there is no consensus on the actual therapeutic measure for infections caused by AmpC-producing bacteria since AmpC-positive bacteria are multidrug resistant and thus are resistant to most first-line and second-line drugs.

Bacterial strains with ampC genes are often resistant to multiple antibiotics; and this makes the selection of an effective antibiotic difficult in the face of an infection by AmpC-producing bacteria. As a general rule and due to the multidrug resistant nature of AmpC-producing bacteria, beta-lactam/beta-lactamase inhibitor combinations and most cephalosporins and penicillins should be totally avoided because of the in vitro resistance, and the potential for AmpC induction or selection of high-enzyme-level mutants, and recorded poor clinical outcomes with these drugs.

However, cefepime can be used for the treatment of bacterial infection caused by AmpC-producing bacteria because cefepime is a poor inducer of AmpC beta-lactamase production in bacteria; and the drug rapidly penetrates through the outer cell membrane of the target organism. Cefepime is also poorly hydrolyzed by AmpC beta-lactamase. Temocillin, a 6-α-methoxy derivative of ticarcillin is active in vitro against many Enterobacteriaceae irrespective of whether the AmpC gene (ampC) is chromosomally-borne or plasmid-mediated.

The carbapenems can also be used for the treatment of infections caused by AmpC-producing bacteria. But the use of the carbapenems is usually followed by the emergence of carbapenem-resistant bacteria especially carbapenem-resistant K. pneumoniae. Fluoroquinolones can also be used to treat infections caused by AmpC-producing bacteria if the isolate shows in vitro susceptibility to the test agent; and fluoroquinolones is usually used for no-life-threatening diseases such as urinary tract infections. Tigecycline has also been reported to show good activity in vitro against AmpC-producing bacteria and thus could be used to treat infections caused by AmpC-producing bacteria.       

References

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