Antibiotic sensitivity testing: Difference between revisions

Antibiotic sensitivity testing or antibiotic susceptibility testing is the measurement of the susceptibility of bacteria to antibiotics. It is used because bacteria may have resistance to some antibiotics. Knowledge about what antibiotics a bacterium is sensitive to can change the choice of antibiotics from empiric therapy to directed therapy.^[1]

Sensitivity testing usually occurs in a laboratory setting, and may be based on culture methods that exposure bacteria to antibiotics, or genetic methods that test to see if a bacterium possesses genes that confer resistance. Culture methods are often based on measuring the diameter of the zones of inhibition on agar culture dishes of bacterial growth around paper discs that are impregnated with antibiotics. The minimum inhibitory concentration of the antibiotic can be determined from this.

Antibiotic sensitivities are reported to clinicians in a tabular format.

In clinical medicine, antibiotics are most frequently prescribed on the basis of a person's symptoms and general guidelines, called empiric therapy.^[1] These are based around knowledge about what bacteria cause an infection, and also what antibiotics bacteria may be sensitive or resistant to in a geographical area.^[1] For example, a simple urinary tract infections might be treated with trimethoprim/sulfamethoxazole.^[2] This is because Escherichia coli is the most likely causative bacterium, and may be sensitive to that combination antibiotic.^[2] However, bacteria can be resistant to several classes of antibiotics.^[2] This resistance might be because of the type of bacteria,^[2] because of resistance following past exposure to with antibiotics,^[2] or because resistance may be transmitted from other sources such as plasmids.^[3] Antibiotic sensitivity testing provides information about what antibiotics are more likely to be successful and are used in this context to provide information about what antibiotics should be used.^[4][1]

Antibiotic sensitivity testing is also conducted at a population level in some countries as a form of screening.^[5] 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.^[5]

Testing for antibiotic sensitivity usually occurs in a laboratory setting.^[6] Methods of testing include:

• Methods based on exposing bacteria to antibiotics:
  • Kirby-Bauer method. Small paper discs containing antibiotics are placed onto a plate upon which bacteria are growing. If the bacteria are sensitive to the antibiotic, a clear ring, or zone of inhibition, is seen around the wafer indicating poor growth.^[7] Müeller-Hinton agar is frequently used in this antibiotic susceptibility test.^[8]
  • Automated methods ^[9]
  • Etest (also based on antibiotic diffusion) ^[9]
  • Agar and Broth dilution methods for minimum inhibitory concentration (MIC) determination.^[10][11]
• Genetic testing, such as via polymerase chain reaction, DNA microarray, DNA chips, and loop-mediated isothermal amplification, which may be used to detect whether bacteria possess genes which confer antibiotic resistance.^[4][12]

Methods used include:

• a semi-quantitative way based on diffusion (Kirby-Bauer method); small discs containing different antibiotics, or impregnated paper discs, are dropped in different zones of the culture on an agar plate, which is a nutrient-rich environment in which bacteria can grow. The antibiotic will diffuse in the area surrounding each tablet, and a disc of bacterial lysis will become visible. Since the concentration of the antibiotic was the highest at the centre, and the lowest at the edge of this zone, the diameter is suggestive for the Minimum Inhibitory Concentration, or MIC, (conversion of the diameter in millimeter to the MIC, in μg/ml, is based on known linear regression curves).
• a quantitative way based on dilution: a dilution series of antibiotics is established (this is a series of reaction vials with progressively lower concentrations of antibiotic substance). The last vial in which no bacteria grow contains the antibiotic at the Minimal Inhibiting Concentration.
• Automated analysers^[13]

The results of the testing are reported as a table, sometimes called an antibiogram.^[14] Bacteria might be marked as sensitive, resistant, or having intermediate resistance to an antibiotic.^[6] Specific patterns pf drug resistance or multi drug resistance may be noted, such as the presence of an extended-spectrum beta lactamase.^[6]

The sensitive, resistant or intermediate resistance to antibiotics is reported base don the minimum inhibitory concentration. It is compared to known values for a given bacterium and antibiotic.^[6] For example, Streptococcus pneumoniae isolates are considered susceptible to penicillin if MICs are ≤0.06 μg/ml, intermediate if MICs are 0.12 to 1 μg/ml, and resistant if MICs are ≥2 μg/ml.^[15][16] Such information may be useful to the clinician, who can change the empirical treatment, to a more custom-tailored treatment that is directed only at the causative bacterium.^[1] Sometimes, whether an antibiotic is marked as resistant is also based on bacterial characteristics that are associated with known methods of resistance such as the potential for beta lactamase production.^[17][6]

Ideal antibiotic therapy is based on determining the causal agent and its antibiotic sensitivity. Empiric treatment is often started before laboratory microbiological reports are available when treatment should not be delayed due to the seriousness of the disease. The effectiveness of individual antibiotics varies with the location of the infection, the ability of the antibiotic to reach the site of infection, and the ability of the bacteria to resist or inactivate the antibiotic.^[19]

Often clinical specimens are sent to the clinical laboratory for culture and sensitivity, which is culture and antibiotic sensitivity testing offered as one combined service.^[6] This is done through collecting samples from affected body sites, with greatest results if the samples are taken before antibiotics are given. For example, a person in an intensive care unit may develop a hospital-acquired pneumonia. The patients are vulnerable and there is a chance the causal bacteria may be different to community-acquired pneumonia, involving bacteria such as Pseudomonas aeruginosa. Treatment is generally started empirically, on the basis of surveillance data about the local common bacterial causes. This first treatment, based on statistical information about former patients, and aimed at a large group of potentially involved microbes, is called empirical treatment.^[20]

Before starting this treatment, the physician will collect a sample from a suspected contaminated compartment: a blood culture sample when bacteria possibly have invaded the bloodstream, a sputum sample in the case of a ventilator associated pneumonia, and a urine sample in the case of a urinary tract infection. These samples are transferred to the microbiology laboratory, which examines the sample under the microscope, and grows the bacteria on culture media.^[21]

When antibiotic sensitivity testing is reported, it will provide useful information the organisms present in the sample, and which drugs will work against them.^[6] Antibiotic sensitivity testing is done in a laboratory (in vitro), but the correlation of this testing to the sensitivity of the antibiotics in a person (in vivo) is often high enough for the test to be clinically useful.^[22]

Point-of-care testing is being developed to speed up the time for testing, and to help practitioners avoid prescribing unnecessary antibiotics in the style of precision medicine.^[23] Traditional techniques typically take 12 to 48 hours,^[24] to up to five days.^[6] In contrast, rapid testing using molecular diagnostics is defined as "being feasible within an 8-h working shift".^[24] Progress has been slow due to a range of reasons including cost and regulation.^[25]

In 2016, the United States National Institutes of Health announced the Antimicrobial Resistance Diagnostic Challenge as a $20 million prize to encourage the development of diagnostic tests to identify highly resistant bacterial infections.^[26] Also, the US Congress wants drug companies that are not working on new resistance antibiotics to do so and if not may pay into a fund if they are not working on new antibiotics to combat the antibiotic resistance crisis.^[27]

As of 2017, point-of-care resistance diagnostics was available for methicillin-resistant Staphylococcus aureus (MRSA), rifampin-resistant Mycobacterium tuberculosis (TB), and Vancomycin-resistant enterococci (VRE) through GeneXpert by molecular diagnostics company Cepheid.^[28]

• Antibiotic resistance
• Miles-Misra method
• Bacterial culture
• Throat culture
• Laboratory specimen

• Burnett, David (2005). The science of laboratory diagnosis. Chichester, West Sussex, England Hoboken, NJ: Wiley. ISBN 978-0-470-85912-4. OCLC 56650888.

• Research Data
• Raw Data
• Antibiogram technique video (diffusion method)