Detecting antimicrobial resistance

Phenotypic methods

Broth dilution tests

• Gold standard
• Uses serial two-fold dilutions of the antimicrobial agent
• Macrobroth has a final volume of 1-2 mL
• Microbroth has a final volume of 50 µL
• Determines an MIC and MBC (min. bactericidal concentration)
  • Read at lowest concentration that inhibits growth (i.e. no obvious growth)
  • Trailing can occur; the MIC is read at the first spot where trailing begins

Agar dilution

• Concentration of antibiotic is incorporated into the agar
• Each agar plate has a different concentration
• Multiple inocula are set up on each plate
  • Final inoculum of 10^4^ CFU per spot

Disk diffusion

• Drug released from disk with an antimicrobial agent diffuses through the agar
• Zone diameter is inversely proportional to the log of the MIC
• Interpretation usually uses reflected light reading above a black, nonreflective background; measured using a ruler on the back of an inverted dish
• If on blood agar, must be read from the front

Other

• Potentiation using standard methods
• Colorimetric methods
• Lateral flow assay

Genotypic methods

• Detected resistance may not predict antibiotic failure if genes are non-functional or not expressed, or may be expressed in insignificant levels
• Similarly, a negative results may not indicate susceptibility given genetic variation, novel mechanisms of resistance, or inhibition of amplification
• Genotypic methods
  • PCR
  • Sequencing