Lecture 3 Antimicrobials and Susceptibility tests Dr. Abdelraouf A. Elmanama Islamic University-Gaza Medical Technology Department
Lecture outlines Kirby-Bauer susceptibility test Antimicrobial profiles selection Reporting susceptibility test
What Does the Laboratory Need to Know about Antimicrobial Susceptibility Testing (AST) ? Which organisms to test? What methods to use? What antibiotics to test? How to report results?
What Does a Laboratory Need to Know about AST? (con’t) How to determine the clinical significance of results? How to ensure accuracy of results? –Quality control / quality assurance When to call the MD, infection control, public health?
What Does a Laboratory Need to Know about AST? (con’t) When to ask for help? Where to go for help?
Brief Review of Routine AST Methods
Routine Susceptibility Tests Disk diffusion (Kirby Bauer) Broth micro-dilution MIC –NCCLS reference method Etest
Disk Diffusion Test
Select colonies Prepare inoculum suspension
Mix well Standardize inoculum suspension
Swab plate Remove sample
Add disks Incubate overnight
Measure Zones Transmitted LightReflected Light
Zone Interpretive Criteria (mm) Drug Disk content (ug) ResIntSusc Cefazolin30 18 Gentamicin10 15 Flash presentation for summary
Qualitative results –Susceptible –Intermediate – may respond if infection is at body site where drug concentrates (e.g. urine) or if higher than normal dose can be safely given –Resistant Disk Diffusion Test
Modify methods for fastidious bacteria
Clinical Conditions when MICs are Useful Endocarditis Meningitis Septicemia Osteomyelitis Immunosuppressed patients (HIV, cancer, etc.) Prosthetic devices Patients not responding despite “Sensitive results”
MIC Minimal inhibitory concentration The lowest concentration of antimicrobial agent that inhibits the growth of a bacterium Interpret: –Susceptible –Intermediate –Resistant
Inoculum Preparation MIC Testing (NCCLS Reference Method) Standardize inoculum suspension Final inoculum concentration –3 – 5 x 10 5 CFU/ml –(3 – 5 x 10 4 CFU/well)
Microdilution MIC tray Prepare inoculum suspension
Dilute & mix inoculum suspension
Pour inoculum into reservoir and inoculate MIC tray
Inoculate purity plate Incubate overnight
Examining purity plate Reflected light Transmitted light
Read MICs
> MICs >64
MIC on a strip
S. pneumoniae Penicillin MIC = 3 g/ml
MIC Interpretive Criteria ( g/ml) DrugSuscIntRes cefazolin 8 16 32 gentamicin 4 4 8 16
Empirical Treatment Infants 1-3 mos Ampicillin + cefotaxime or ceftriaxone Immunocompetent children > 3 mos and adults <55 Cefotaxime or ceftriaxone + vancomycin Adults > 55 and adults of any age with alcoholism or other debilitating illnesses Ampicillin + cefotaxime or ceftriaxone + vancomycin Hospital-acquired meningitis, posttraumatic or postneurosurgery meningitis, neutropenic patients, or patients with impaired cell-mediated immunity Ampicillin + ceftazidime + vancomycin
Ceftazidime should be substituted for ceftriaxone or cefotaxime in neurosurgical patients and in neutropenic patients
Specific treatment N. meningitidis –Penicillin sensitive Penicillin G or Ampicillin –Penicillin-resistant Ceftriaxone or cefotaxime
Chemoprophylaxis for N. meningitidis Rifampin 600 mg every 12 h for 2 days in adults and 10 mg/kg every 12 h for 2 days in children >1 year Or One dose of ciprofloxacin (750 mg) One dose of azithromycin (500 mg) One intramuscular dose of ceftriaxone (250 mg) Rifampicin is not recommended in pregnant women.
Pneumococci –Penicillin-sensitive Penicillin G –Penicillin-intermediate Ceftriaxone or cefotaxime –Penicillin-resistant (Ceftriaxone or cefotaxime) + vancomycin
Gram-negative bacilli (except Pseudomonas spp.) Ceftriaxone or cefotaxime Pseudomonas aeruginosa Ceftazidime
Staphylococci spp. –Methicillin-sensitive Nafcillin –Methicillin-resistant Vancomycin Listeria monocytogenes Ampicillin + gentamicin Haemophilus influenzae Ceftriaxone or cefotaxime Streptococcus agalactiae Penicillin G or ampicillin Bacteroides fragilis Metronidazole Fusobacterium spp. Metronidazole
Local Data and protocols should be observed and reviewed periodically Thank you