CE: Development of resistance to antibiotics

This second article in a three-part series provides healthcare representatives with an understanding of the concept of bacterial resistance to antibiotics in terms of how bacteria acquire resistance and the mechanisms of resistance.

J. Peter Rissing, M.D., FACP, FIDSA, chief of Infectious Diseases Section, Sydenstricker Professor of Medicine, Medical College of Georgia, Augusta, GA; and John C. Rotschafer, Pharm.D., FCCP, Section of Clinical Pharmacy, Regions Hospital, and professor, College of Pharmacy, University of Minnesota, St. Paul, MN, served as consultants for this article for the Certified Medical Representatives Institute Inc.

Learning Objectives

* Describe chromosome mutations, transduction, transformation and conjugation, and explain the role they play in the development and spread of antibiotic resistance.

* Describe the types of antibiotic-resistant bacteria currently causing concern in the medical and scientific communities.

* Explain how alterations in antibiotic efflux, as well as alterations in the drug's target site and metabolic bypass, may result in bacterial resistance to antibiotics.

* List the common classes of antibiotics and typical mechanisms of resistance associated with them.

* Describe the multiple factors that have contributed to the rise of antibiotic resistance.

The number of antibiotic-resistant bacterial strains is on the rise, both in the United States and around the world. This increase results in major therapeutic issues and creates a need for new antibiotics.

Transmission of resistance

Bacteria have proved remarkably adept at developing resistance to new antimicrobial agents. Resistance to an antibiotic may be an intrinsic property of a bacterial species, or it may be an acquired characteristic of an individual organism.

Once bacteria do become resistant, they can (and usually do) pass this property from one generation to the next. They may also donate the property of resistance to other strains. If it were not for the movement between strains of genes that confer resistance, resistance properties would be restricted to the strains that possess them. While chromosome mutation can be a cause of resistance, it is more frequently acquired through transfer of genetic material from one bacterium to another by the following mechanisms: transduction, transformation and conjugation.

Alterations in or mutations of the bacterial chromosomes were the first identified mechanisms of resistance. For example, in Escherichia coli or Staphylococcus aureus, rifampin resistance is mediated by a chromosomal mutation in the structural gene for the RNA group, such that it no longer effectively binds rifampin. Parallel chromosomal mutations are responsible for pneumococci developing resistance to penicillin and the sulfonamides, as well as streptomycin resistance in the enterococci and Gram-negative resistance to fluoroquinolones.

In the method of transduction, a bacteriophage (a virus that infects bacteria) carries bacterial DNA within its protein coat. If this genetic material includes a gene for resistance, a newly infected bacterial cell may become resistant to a drug and be capable of passing the trait of resistance to its offspring. Transduction is particularly important in the transfer of antibiotic resistance among strains of S. aureus, where some bacteriophages can carry plasmids that code for penicillinase. Others transfer genetic information encoding resistance to erythromycin, tetracycline or chloramphenicol. Plasmids are self-replicating extrachromosomal DNA elements containing R-factors or genetic elements that encode for synthesis of enzymes to functionally inactivate or modify antibiotics. This confers resistance to similar macrolide antibiotics in succeeding generations of the bacteria.

Transformation involves incorporating DNA that is roaming free in the environment into bacteria. Some bacterial cells are capable of excreting transforming DNA during certain phases of growth.

Conjugation, the passage of genes from cell to cell by direct contact through a sex pilus or bridge, is an extremely important mechanism for the spread of antibiotic resistance, particularly since DNA that codes for resistance to multiple drugs is transferred via this method. The transferable genetic material consists of two different sets of DNA sequences contained in plasmids. The first set of sequences codes for the actual resistance and is termed the R-determinant plasmid. The second plasmid, termed the resistance transfer factor or RTF, contains the information needed for bacterial conjugation to occur. Each of these two plasmids can exist independently, or they can combine to form a complete R-factor, which is disseminated by bacterial conjugation.

The spread of resistance

The proportion of enteric bacteria that carry plasmids for multiple-drug resistance has slowly risen in the past 30 years.

For example, multiple-drug-resistant enterococci (e.g., resistant to penicillin, aminoglycosides and vancomycin) have become a significant problem throughout the world, causing treatment problems for physicians and creating a need for new antibiotics. The degree of enterococci resistance may be categorized as follows:

• Ampicillin/penicillin-sensitive enterococci.

• Ampicillin/penicillin-resistant enterococci.

• Low-level aminoglycoside resistance (<500 mg/L).

• High-level aminoglycoside resistance (>2,000 mg/L).

• Vancomycin-sensitive enterococci.

• Vancomycin-resistant enterococci.

The emergence of beta-lactamase-producing strains of Haemophilus influenzae is another major therapeutic problem. The gene for production of this enzyme is carried on small plasmids. However, effective enzyme production is dependent on a number of factors, and actual production of the resistance enzyme is extremely variable depending on the organism.

Since 1987, a rise in the number of Streptococcus pneumoniae strains resistant to penicillin has been reported in the United States. This is a significant concern, as S. pneumoniae is estimated to cause over 500,000 cases of pneumonia, 55,000 cases of bacteremia, and 6,000 cases of meningitis in the United States annually.

During the 1990s, strains of S. pneumoniae showed increasing resistance to penicillin and related drugs, including cephalosporins and other classes such as macrolides. Initially, most of these resistant organisms were said to be "intermediate" (or nonsusceptible) in their susceptibility (MIC = 0.125 to 1.0 µg/mL). During 1997 and 1998, Thornsberry reported 36% of 3,340 S. pneumoniae U.S. strains to be resistant. While 22% were "intermediate" (or nonsusceptible), 14% were fully resistant to penicillin (>2.0 µg/mL). Many strains were multiple-drug-resistant.

Drug-resistant S. pneumoniae may be underreported, as cultures are not often obtained for otitis media, bronchitis, community-acquired pneumonia or sinusitis, and if cultures are tested, the automated equipment may not detect the resistant organisms. Recently, resistance to third-generation cephalosporins, such as cefotaxime or ceftriaxone, as well as other antibiotic classes, has also appeared among pneumococcal strains in the United States, Spain and South Africa.

Mechanisms of resistance

One of the most worrisome mechanisms of resistance involves the ability of bacteria to exclude antimicrobial agents from the cell. For example, high-level, multiple-drug-resistant strains of bacteria have altered their membrane transport system so that the membrane is impermeable to all aminoglycosides, and possibly other antibiotics.

Some Gram-negative and Gram-positive pathogens have an efflux pump that can remove fluoroquinolones fast enough to protect the bacteria from the drug's antibiotic effects. Many types of Gram-positive and Gram-negative bacteria are also capable of resisting tetracyclines through the efflux mechanism.

Resistance to an antibiotic may also result from a change or alteration in the target site at which the drug acts. Some strains of bacteria alter the protein composition of their outer cell membrane to prevent a drug from reaching its target site. This is a common method of resistance to the penicillins.

Metabolic bypass is another common way in which bacteria can become resistant to antibiotics. In this case, the antibiotic interferes with the metabolic pathway by blocking or interfering with a critical step. The bacteria substitute an "antidote" that bypasses the block.

Bacterial enzymes that destroy or inactivate antibiotics. Some resistant bacteria produce enzymes that cleave or alter the molecular structure of an antibiotic, thereby rendering it ineffective. The most common enzymes that mediate bacterial resistance are the beta-lactamases. As shown in the above diagram, these enzymes open the beta-lactam ring, a molecular structure that is characteristic of the penicillins and cephalosporins and linked to the activity of both classes of drugs. Thus, the greater the stability of the beta-lactam ring, the less chance there is of resistance developing. A beta-lactamase that specifically attacks the beta-lactam ring in penicillins is known as a penicillinase, while a beta-lactamase that attacks a cephalosporin is known as a cephalosporinase. Resistance to the penicillins has increased rapidly.

Chromosomally mediated beta-lactamases (Richmond-Sykes type I or Bush type I) are present in many Enterobacter, Citrobacter, Proteus, Providencia, Serratia and Pseudomonas species, and in all Klebsiella species. At least 35% of Haemophilus influenzae strains possess a plasmid-mediated beta-lactamase. Extended-spectrum beta-lactamases are newer plasmid-mediated enzymes that have been identified predominantly in Klebsiella pneumoniae and in other Enterobacteriaceae. Extended-spectrum beta-lactamases are capable of destroying cephalosporins that were previously impervious to plasmid-mediated beta-lactamase.

The aminoglycosides are important antibiotics in the treatment of infections caused by Gram-negative bacilli. Increased use by physicians, however, has led to the increased development of resistance to aminoglycosides. A growing number of bacterial strains have become capable of producing plasmid-mediated enzymes (different from beta-lactamases) that modify and thus inactivate the ability of aminoglycosides to inhibit protein synthesis.

The following are common classes of antibiotics and typical mechanisms of resistance associated with each:

• Beta-lactams (penicillins, cephalosporins, carbapenems, carbacephems).

– Destruction by beta-lactamase.

– Alteration of penicillin-binding proteins.

• Aminoglycosides.

– Enzyme modification.

– Absence of oxygen-dependent membrane transport.

• Chloramphenicol.

– Altered target site (ribosome group).

• Erythromycin, clindamycin.

– Ribosomal.

– Altered efflux.

• Fluoroquinolones.

– Altered target site (DNA synthesis).

– Altered influx/efflux.

• Sulfonamides.

– Altered target site.

Susceptibility testing is becoming increasingly important. Susceptibility testing traditionally has been performed more frequently in the hospital for inpatients. However, due to increasing numbers of resistant bacteria appearing in community-acquired infections, primary care physicians may be likely to order more susceptibility tests from their local microbiology laboratories.

Increase in antimicrobial resistance

A major factor contributing to the rise of antibiotic resistance has been the overuse of antibiotics, often in situations where they are inappropriate or unnecessary. United States production of antibiotics has increased from two million pounds in 1954 to over 50 million pounds in 1998. About one half of the antibiotics consumed each year in the United States is used for human treatment. Only about half of that, or about one quarter of the total, is prescribed to cure bacterial infections and administered in ways that do not promote antibiotic resistance. According to the Centers for Disease Control and Prevention, U.S. physicians write about 150 million outpatient prescriptions for antibiotics every year, and about 50 million of these prescriptions are unnecessary. This excessive and routine use of antibiotics has contributed to the increase in antibiotic resistance by selectively encouraging the survival and growth of resistant strains of bacteria. Hospitals and surgeons may also promote antibiotic resistance through the routine and sometimes overextended use of prophylactic antibiotics with surgery.

Patients contribute to antibiotic resistance when they do not take the drugs according to directions. Many patients stop taking their antibiotics before they have finished the entire course of treatment. An insufficient dose of antibiotic usually fails to kill all of the disease-causing bacteria. The survivors are the most resistant strains, which may later cause infectious illnesses that are difficult to treat.

In many Asian, African and South American countries, potent antibiotics are sold without prescription. The antibiotics available in third-world countries may be less than fully potent because they are past their expiration dates or have been stored under poorly controlled conditions, or they may be counterfeits, adulterated products or generic formulations that are not equivalent to the original. These and other factors help resistant bacterial strains flourish in developing countries. Because international travel has become common, these resistant strains may easily spread to other countries as well.

The same antibiotics prescribed for human illnesses are also widely used in animal husbandry and agriculture. Animals usually receive low doses of antibiotics over long periods of time, creating an environment where resistant bacteria thrive. Farmers also spray antibiotics on fruit trees, encouraging the growth of resistant bacteria on the fruit and in the surrounding environment.

Article Summary

* Once bacteria become resistant, they can pass this property of resistance to other strains.

* While chromosome mutation can be a cause of resistance, it is more frequently acquired through transfer of genetic material from one bacterium to another by the mechanisms of transduction, transformation and conjugation.

* Antibiotic resistance in common bacterial organisms, such as enterococci, Haemophilus influenzae and Streptococcus pneumoniae, have caused treatment problems and created a need for new antibiotics.

* One mechanism of resistance involves bacterial exclusion of antimicrobial agents from the cell.

• Some bacteria alter their membrane transport system so that the membrane is impermeable to certain antibiotics.

– Some pathogens have an efflux pump that can remove antibiotics fast enough to protect the bacteria from the drug's effects.

– Some bacteria alter the protein composition of their outer cell membrane to prevent a drug from reaching its target site.

* Metabolic bypass is another common way in which bacteria can become resistant.

* Some bacteria produce beta-lactamase enzymes, including ESBLs, that cleave or alter the beta-lactam ring of penicillins and cephalosporins.

* A growing number of bacterial strains are capable of producing plasmid-mediated enzymes that modify and, thus, inactivate the bactericidal capability of the aminoglycosides.

* Key factors contributing to increased resistance include: overuse and misuse of antibiotics, patients' noncompliance, social and economic factors in developing countries, and agricultural use of antibiotics.

© 2002 The Certified Medical Representatives Institute Inc., Roanoke, VA 24018. All rights reserved. No part of this article may be reproduced by any method or in any form without written permission from the CMR Institute. Reprints of this article are available from the CMR Institute. Request Continuing Education article DR-2.