Ceftolozane-tazobactam susceptibility testing evaluates the effectiveness of the cephalosporin antibiotic ceftolozane combined with the beta-lactamase inhibitor tazobactam against bacterial isolates. This combination is used to treat infections caused by multidrug-resistant Gram-negative bacteria.

Testing Procedures

Preparation:

• Bacterial Isolate: Use a pure culture of the microorganism.
• Inoculum Preparation: Adjust the bacterial suspension to a 0.5 McFarland turbidity standard for accurate and reproducible results.

Broth Microdilution Method:

• Preparation: Prepare a series of ceftolozane-tazobactam concentrations in a microtiter plate. Typically, this involves preparing ceftolozane-tazobactam in a fixed ratio (e.g., 4:1, ceftolozane) across a range of concentrations.
• Inoculation: Add the standardized bacterial suspension to each well containing ceftolozane-tazobactam.
• Incubation: Incubate at 35-37°C for 16-24 hours.
• Reading Results: Determine the minimum inhibitory concentration (MIC), the lowest concentration of ceftolozane-tazobactam that inhibits visible bacterial growth.
• Interpretation: Compare MIC values to established breakpoints. Common breakpoints are:
  • Susceptible: MIC ≤ 4/4 µg/mL
  • Intermediate: MIC = 8/4 µg/mL
  • Resistant: MIC ≥ 16/8 µg/mL Breakpoints may vary based on guidelines from organizations like the Clinical and Laboratory Standards Institute (CLSI) or the European Committee on Antimicrobial Susceptibility Testing (EUCAST).

Disk Diffusion Method:

• Agar Plate: Plate Mueller-Hinton agar with the standardized bacterial suspension.
• Ceftolozane-Tazobactam Disk: Place a disk impregnated with ceftolozane-tazobactam (e.g., 30 µg of ceftolozane and 15 µg of tazobactam) on the agar surface.
• Incubation: Incubate the plate at 35-37°C for 16-18 hours.
• Measurement: Measure the diameter of the zone of inhibition around the disk.
• Interpretation: Compare the zone diameter to standardized breakpoints:
  • Susceptible: ≥ 20 mm
  • Intermediate: 17-19 mm
  • Resistant: ≤ 16 mm Note that disk diffusion testing for ceftolozane-tazobactam may not be as commonly standardized as for other antibiotics; consult specific guidelines from CLSI or EUCAST for the most accurate breakpoints.

Automated Systems (e.g., VITEK 2, Phoenix):

• Procedure: Automated systems can test ceftolozane-tazobactam susceptibility using predefined panels designed for combination antibiotics.
• Incubation and Reading: The system automates incubation and growth measurement, providing susceptibility results based on predefined algorithms and breakpoint criteria.