Mayo Clinic’s antimicrobial susceptibility testing (AST) section of the Bacteriology Laboratory in Rochester, Minnesota, is one of the few clinical laboratories in the country that routinely uses agar dilution for aerobic and anaerobic bacterial susceptibility testing.
Antimicrobial susceptibility testing determines whether these bacteria are susceptible or resistant to a particular antibiotic. Bacteria are added onto plates of solid agar, where each plate has a different and increasing concentration of antibiotic than the previous plate. The minimum inhibitory concentration (MIC) is the lowest concentration of an antimicrobial that inhibits growth of the bacteria.
Each result generally falls into one of three interpretive categories:
Using this method, “Our results can help the health care provider predict what antibiotics may work best for treating the patient’s infection,” says Audrey Schuetz, M.D., Co-Director of the Bacteriology Laboratory and Head of the AST section.
As the primary method of testing in the Bacteriology Laboratory at Mayo Clinic, agar dilution is one of the gold standard methods of AST. However, it is a laborious method requiring exceptional expertise. In contrast, most other clinical microbiology labs use other, more automated methods for their AST.
“We use agar dilution because it’s reproducible and accurate, and it predicts how well an antibiotic may work for treating a person’s infection,” says Dr. Schuetz. “This method is also used to set breakpoints (specific antibiotic concentrations that define isolates as Susceptible, Intermediate, or Resistant) that are determined by breakpoint-setting organizations like the Clinical & Laboratory Standards Institute (CLSI).”
An additional benefit to using agar dilution for bacterial AST at Mayo is that the Bacteriology Laboratory does not need to rely on a commercial method for the majority of its testing. This allows the laboratory to be less affected by uncontrollable events (such as back orders and recalls) that come with relying exclusively on outside companies. Instead, the laboratory has a section dedicated to making the majority of its media in-house and therefore can prepare its own antibiotic plates for AST testing.
“It’s nice to have that independence,” says Dr. Schuetz. “Plus, we can be a comparator for other laboratories that want to send us their bacteria for validation of a new AST method.”
Being self-reliant also means that the AST section can validate new drugs quickly without having to wait for commercial systems to add new drugs to their automated panels. “We validate and offer new antibiotics as soon as possible after they hit the market so that we can offer susceptibility testing for patients for whom the drug may be considered as appropriate treatment,” says Dr. Schuetz.
Additionally, turnaround time of AST results is an important factor for patient care of which agar dilution testing can be of benefit. Turnaround time for agar dilution AST results is typically two days for aerobic bacteria and about five days for anaerobes due to stricter growth conditions.
Currently, the AST section of the laboratory is working on a novel application of a test method through an NIH-funded clinical trial with the Antibacterial Resistance Leadership Group (ARLG) and CLSI.
“In this trial, we’re studying an AST method that will decrease the turnaround time for susceptibility results when performed directly from a clinical sample,” says Dr. Schuetz, who is the principal investigator for this multisite study. “Instead of waiting for bacterial growth on a plate prior to agar dilution susceptibility testing, we’re using a novel application of an AST method directly from positive blood culture broth (i.e., positive blood culture specimens). Although some laboratories have used this approach in the past, this large multi-center study will aid CLSI in setting appropriate breakpoints for this method which can be uniformly adopted by laboratories.”
In this method, a blood culture flags positive for microorganism growth by an automated detection instrument. Further analysis of that blood culture broth confirms the presence of a microorganism (e.g., gram-negative bacteria on a routine Gram stain). A certain amount of this positive broth is then inoculated onto the surface of an agar plate, and antibiotic-impregnated disks are added to the plate. After a certain incubation period, which is still being assessed, bacterial growth around this disk can be read and interpreted according to CLSI guidelines as Susceptible, Intermediate, or Resistant to that particular antibiotic.
If validated, this approach would be less expensive, produce results in less time, and be relatively easy for clinical laboratories to adopt, requiring less labor and expertise than agar dilution.
Dr. Schuetz and her Co-Director of the Bacteriology Laboratory, Robin Patel, M.D., are involved in other studies as well, and they are always considering trials and antimicrobial testing methods for early adoption.
“There are also molecular tools that we are developing that will predict resistance to certain antimicrobials,” Dr. Patel says. “These molecular tools detect the presence of the gene target rather than examining bacterial growth in reaction to the antimicrobial. There’s not always a one-to-one correlation between the presence of a gene and its association to clinical resistance, but it’s another tool for us to use alongside phenotypic susceptibility testing.”
Validating and adopting new AST methods will allow microbiology laboratories to deliver results closer to point of care (e.g., the patient’s bedside) and, therefore, improve patient outcomes.
“The sooner we can let a physician know if a bacterium is susceptible or resistant to a particular antibiotic, the faster the provider can cater appropriate antibiotic therapy for the patient,” says Dr. Schuetz. “The work we do here can be compared to personalized medicine because with each bacterial infection coming from a particular source in an individual, we’re trying to gather as much information as possible about this microorganism to guide that patient’s treatment.”
|Antimicrobial Susceptibility, Aerobic Bacteria, MIC (Mayo ID: ZMMLS)||Antimicrobial Susceptibility, Anaerobic Bacteria, MIC (Mayo ID: MMLSA)|
|Aerobic Gram-Negative Bacilli Antimicrobials|
Additional Gram-Negative Bacteria Antimicrobials
Additional Gram-Positive Bacteria Antimicrobials
Staphylococcus, Enterococcus, Bacillus, and Related Genera Antimicrobials
|Anaerobic Bacteria Antimicrobials|