Antibiotic sensitivity testing

The Etest, an antibiotic impregnated strip, has been available since the 1980s, and genetic methods such as polymerase chain reaction (PCR) testing have been available since the early 2000s.

In clinical medicine, antibiotics are most frequently prescribed on the basis of a person's symptoms and medical guidelines.

[1] Antibiotic sensitivity testing is also conducted at a population level in some countries as a form of screening.

[4] This is to assess the background rates of resistance to antibiotics (for example with methicillin-resistant Staphylococcus aureus), and may influence guidelines and public health measures.

[6] There are many factors that can affect the results of antibiotic sensitivity testing, including failure of the instrument, temperature, moisture, and potency of the antimicrobial agent.

[5] To ensure that the results are accurate, the concentration of bacteria that is added to the agar or broth (the inoculum) must be standardized.

Once an appropriate concentration (most commonly an 0.5 McFarland standard)[10] has been reached, which can be determined by visual inspection or by photometry, the inoculum is added to the growth medium.

The bacteria are classified as sensitive, intermediate, or resistant to an antibiotic by comparing the diameter of the zone of inhibition to defined thresholds which correlate with MICs.

[5] The minimum inhibitory concentration can be identified based on the intersection of the teardrop-shaped zone of inhibition with the marking on the strip.

[18] In agar and broth dilution methods, bacteria are placed in multiple small tubes with different concentrations of antibiotics.

[14] Whether a bacterium is sensitive or not is determined by visual inspection or automatic optical methods, after a period of incubation.

[14] Automated instruments, such as the VITEK 2, BD Phoenix, and Microscan systems, are the most common methodology for AST.

The specifications of each instrument vary, but the basic principle involves the introduction of a bacterial suspension into pre-formulated panels of antibiotics.

The panels are incubated and the inhibition of bacterial growth by the antibiotic is automatically measured using methodologies such as turbidimetry, spectrophotometry or fluorescence detection.

[22] Genetic testing, such as via polymerase chain reaction (PCR), DNA microarray, and loop-mediated isothermal amplification, may be used to detect whether bacteria possess genes which confer antibiotic resistance.

[25] Polymerase chain reaction is a method of identifying genes related to antibiotic susceptibility.

[9] Using magnetic nanoparticles studded with a beta-2-glycoprotein I peptide imitating a plasma protein, microbial pathogens could selectively be retrieved from blood culture specimens within hours, in a study published September 2024.

[27] Matrix-assisted laser desorption ionisation-time of flight mass spectrometry (MALDI-TOF MS) is another method of susceptibility testing.

[6] This is a form of time-of-flight mass spectrometry, in which the molecules of a bacterium are subject to matrix-assisted laser desorption.

The MIC is compared to standard threshold values (called "breakpoints") for a given bacterium and antibiotic.

[31][32] Antibiograms help the clinician to select the best empiric antimicrobial therapy based on the local resistance patterns until the laboratory test results are available.

[1] These samples are transferred to the microbiology laboratory where they are added to culture media, in or on which the bacteria grow until they are present in sufficient quantities for identification and sensitivity testing to be carried out.

[41] Quantitative PCR, with the view of determining the percent of a detected bacteria that possesses a resistance gene, is being explored.

[9] Whole genome sequencing of isolated bacteria is also being explored, and likely to become more available as costs decrease and speed increases over time.

[9] These involve labelled proteins targeted at biomarkers, nucleic acid sequences present within cells that are found when the bacterium is resistant to an antibiotic.

See caption.
Example of antibiotic sensitivity testing. Thin paper discs containing an antibiotic have been placed on an agar plate growing bacteria . Bacteria are not able to grow around antibiotics to which they are sensitive. This is called "the zone of inhibition".
Three clear test tubes containing solutions of successively increasing turbidity.
From left to right: 0.5, 1, and 2 McFarland standards
Bacteria are growing on an agar plate upon which a strip containing varying concentrations of antibiotics has been placed. An elliptical zone without growth is present around areas with higher concentrations of the antibiotic.
Example of an Etest , which uses a plastic strip impregnated with an antibiotic at a range of concentrations
See caption.
Antibiotic resistance tests : Bacteria are streaked on dishes with white disks, each impregnated with a different antibiotic. Clear rings, such as those on the left, show that bacteria have not grown—indicating that these bacteria are not resistant. The bacteria on the right are fully resistant to all but two of the seven antibiotics tested. [ 33 ]