Clinical implications of resistance (detecting in the lab)

Antimicrobial resistance (AMR) is a huge clinical concern worldwide and is making infections harder to treat. It poses such a problem, that the WHO has declared AMR to be one of the top 10 global public health threats facing humanity, with implications ranging from loss of life to huge economic burdens.

Antimicrobial stewardship (AMS) is a coordinated approach, aiming to reduce resistance by encouraging appropriate usage of antimicrobials – the right drug, at the right dose, at the right time, for the right duration. The ultimate goal of AMS is to achieve the best possible outcome for the patient. Antimicrobial susceptibility testing (AST) can be performed on pathogens isolated from clinical samples and is used to ensure the correct antimicrobials are used against a particular infection by measuring the pathogen’s susceptibility and resistance to antimicrobial agents, including aminoglycosides.

AST is performed in the laboratory, and there are several types of techniques that can be employed:

• Phenotypic AST – based on measuring the activity of the antimicrobial agent against the microorganism. The minimum inhibitory concentration (MIC) is typically measured and compared against breakpoints.
• Genotypic AST – detection of a resistance gene (mecA, vanA/vanB, etc) in the genome or on transferable genetic elements.
• Mechanistic AST – detection of a resistance mechanism – by detecting the product of the resistance gene (enzyme).
• Expert rules AST – susceptibility based on empiric knowledge.

Phenotypic AST & MIC measurement

MIC is the lowest concentration of a drug needed to inhibit cell division in the environment in which the test was performed. It is a broth microdilution and is the basis for all susceptibility tests. Clinical AST should produce a clinical recommendation, to determine whether to treat or not and whether standard dosing will suffice or if increased exposure (i.e. a higher dose) is required.

The MIC is not an absolute value and is affected by factors such as:

• Inoculum
• Incubation time
• The broth used and its composition
• Atmosphere
• pH

Testing many isolates of the same species against a defined agent can determine the wild-type distribution of the species for that particular agent. Wild-type isolates are those that lack an acquired resistance mechanism. Once the wild-type distribution is established and epidemiological cut-off values (ECOFF) are determined, the highest MIC of the wild-type distribution, successive isolates can be categorised as wild-type or non-wild-type.

Gentamicin MIC

The current threshold for detecting high-level resistance to gentamicin in vitro is an MIC of 500 μg/ml or greater. These levels of resistance are surrogate markers for predicting synergism based on the presence or absence of the aac(6’)-Ie-aph(2”)-Ia bifunctional gene.

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