Antibiotic resistance occurs through a variety of mechanisms that alter the binding site of the drug to the microorganism or prevent the antibiotic from penetrating into the cell. Some organisms develop “efflux” pumps that actually pump the drug out of the cell after it enters, explained Geraldine Hall, PhD, medical director, Bacteriology, Mycobacteriology and Specimen Processing, Cleveland Clinic, Cleveland.
In other instances, some organisms can bypass the function that the drug interferes with, making the organism no longer susceptible to its inhibition. “Organisms can be either intrinsically or innately resistant to a particular antibiotic; they are never susceptible to it,” Hall said. “Or, organisms can acquire resistance mechanisms due to the pressure of exposure to the particular antibiotic or class of antibiotics.” This acquired resistance often occurs in a hospital environment where many antibiotics are used and organisms are constantly being exposed to them.
Detecting and Monitoring Antibiotic Resistance
Most bacteriology laboratories serve two functions: to identify organisms that might be pathogens in clinical samples (by culture or antigen detection, or by molecular means) and to test those organisms for the presence of antibiotic resistance mechanisms in vitro – by subjecting the organisms to increasing concentrations of the antibiotics in agar, in broth, or antimicrobial pattern via disk diffusion or E-test, Hall said.
Molecular methods that can pick up specific resistance mechanisms via detection of the gene responsible for the mutation are rapidly developing in clinical microbiology and these will further enable susceptibility testing to be reported as quickly as possible.
The lab surveys resistance patterns by storing data of individual organism resistance results and then displaying that data annually or more frequently on an antibiogram. The latter is a chart demonstrating the percent susceptibility (or resistance) of the most common bacteria (and also fungi) to commonly used drugs to treat these organisms when they cause infections. Clinicians access the antibiogram to monitor resistance patterns so they can gauge which empiric therapy might be needed for a patient’s infection before the patient’s specific pattern of resistance is known.
The Surveillance Network (TSN) is the largest and longest standing electronic surveillance network specifically designed to track in vitro antimicrobial activity patterns in the United States.1 More than 500 hospitals contribute data to the network daily, which contains more than 40 million pieces of information, reports John G. Thomas, MS, PhD, HCLD, director, Biofilm Research Laboratory for Translational Studies in Medicine, Dentistry, and Industry, West Virginia University (WVUH), Morgantown, W. Va., and senior consultant to clinical microbiology, WVUH.
Issues in Antibiotic Resistance
Overuse of antibiotics is the largest contributing factor to increasing antibiotic resistance throughout the world. Pharmaceutical companies are rarely developing and marketing new antibiotics.
Another challenge is the ability to detect resistance with automated systems. “Kirby-Bauer disk diffusion is manual and time consuming, but it is still used a lot to back up questionable results from automated systems,” said Beverley L. Orr, MT(ASCP), microbiology technical supervisor, Boston Medical Center, Boston, Mass.
Testing Types
Both phenotypic tests and molecular tests are now available to detect antibiotic resistance. Phenotypic tests are actual disk diffusion or minimum inhibitory concentration (MIC) tests that give the physician a panel of drugs that are either susceptible or resistant. These panels give the physician multiple classes of drugs to include or exclude from a patient’s therapy. The MIC will give a relative amount of antibiotic that is required in the patient’s serum to keep the organism from multiplying. E-test, a product manufactured and distributed by bioMerieux (Marcy l’Etoile), France, is a gradient diffusion method for determining an MIC.
Molecular tests, new to the clinical microbiology lab, detect genes that are usually key for resistance. “These tests are usually quicker than phenotypic tests, however they typically only test for one class of drugs,” Orr said. These tests are used primarily for preliminary results. Phenotypic tests usually follow for confirmation and results of a greater variety of antibiotics.
Signs of Growing Resistance
Molecular detection is a growing field for detecting resistance along with more rapid identification of microbes. This will most certainly continue along with increased use of matrix assisted laser desorption ionization — time of flight (MALDI-TOF) as well as mass spectrophotometric analysis of organisms for identification and resistance detection, as well as sequencing.
New Advances
Automatic systems are continuously improving their detection of resistance patterns. Orr said the Microphage Keypath system is a good choice because it does not require instrumentation. “We are no longer specifically reporting whether an isolate is resistant due to an extended spectrum beta-lactamases (ESBL) or Klebsiella pneumoniae carbapenemase (KPC), but rather just that it is resistant to a drug or class of drugs without reporting a specific mechanism. The breakpoints for resistance to β-lactam antibiotics in the Enterobacteriacae have been lowered by Clinical Laboratory Standards Institute (CLSI) recently as have the breakpoints for carbapenems versus Pseudomonas aeruginosa in order to capture all of the possible resistance mechanisms that could be occurring.
Karen Appold is an editorial consultant. Visit www.WriteNowServices.com
Reference
The Surveillance Network (TSN). Available at: http://www.thetsn.com/. Accessed Aug. 24, 2012.
