Microbiology: Specimens, Identification and Susceptibility
Microbiology questions reward a disciplined sequence: was the right specimen collected, what does the direct stain show, what grew, how is it identified, and what should be reported. The exam also tests rejection criteria and biosafety judgement.
Core rules
- Specimen quality drives the result. Sputum is screened by the Murray-Washington criteria: acceptable when fewer than 10 squamous epithelial cells and more than 25 neutrophils per low-power field. Reject saliva-like samples.
- Swabs are poor for anaerobes and for most bacterial culture beyond throat, wound surface and genital sites; aspirates or tissue are preferred.
- Gram stain reporting includes quantity, morphology, arrangement, Gram reaction and presence of inflammatory cells.
- Identification logic: catalase and coagulase for staphylococci; bacitracin, optochin, bile solubility and bile esculin for streptococci and enterococci; oxidase, lactose fermentation and TSI for gram-negative bacilli; MALDI-TOF or molecular methods for definitive identification.
- Susceptibility testing follows CLSI: disk diffusion (Kirby-Bauer), broth microdilution for MIC, and categories S, I (susceptible, increased exposure) and R. MRSA is confirmed with a cefoxitin disk, which induces mecA-mediated resistance more reliably than oxacillin.
Worked example
CSF from a 3-year-old: WBC 1,200 x10^6/L with 88% neutrophils, glucose 1.4 mmol/L against a serum glucose of 5.6 mmol/L, protein 2.6 g/L, and Gram stain showing Gram-negative diplococci both inside and outside neutrophils. Reasoning: a neutrophilic pleocytosis with a CSF:serum glucose ratio below 0.4 and high protein indicates bacterial meningitis. The morphology points to Neisseria meningitidis; Moraxella and Acinetobacter are the other Gram-negative diplococci to mention. Action: phone the result as critical, ensure blood cultures were collected, and set up susceptibility testing per CLSI.
Traps that cost marks
- Reading beta-haemolysis on a blood agar plate held under anaerobic conditions, where Streptococcus pyogenes haemolysis is poorly expressed; read in air.
- Confusing coagulase-negative staphylococci from a single blood culture bottle with true bacteraemia; multiple bottles and clinical context are needed.
- Ignoring that beta-lactamase testing and inducible clindamycin resistance (D-zone) change therapy for Staphylococcus aureus.
- Forgetting that stool culture for E. coli O157:H7 requires sorbitol-MacConkey and that non-O157 Shiga toxin producers need immunoassay or molecular testing.
- A Gram stain is not a substitute for culture, and a negative culture does not exclude infection after antibiotics.