Sputum Culture


1. Objective

To detect, isolate, and identify pathogenic microorganisms from lower respiratory tract secretions, especially in patients with pneumonia, bronchitis, tuberculosis, or lung abscess.

2. Principle

Sputum is collected from deep cough (not saliva), inoculated onto selective and differential media, and incubated. Colonies are analyzed by Gram stain, biochemical tests, and antibiotic susceptibility testing to identify the etiological agent of infection.

3. Materials

  • Sterile wide-mouth sputum container

  • Glass slides and cover slips

  • Inoculating loop

  • Blood agar, Chocolate agar, MacConkey agar

  • Lowenstein-Jensen (if TB suspected)

  • Incubator (35–37°C, with CO₂ if needed)

  • Gram staining reagents

  • Mueller-Hinton agar (for sensitivity test)

  • PPE, sputum collection instructions

4. Procedure

A. Collection

  1. Instruct patient to cough deeply and collect early morning sputum (before food).

  2. Avoid contamination with saliva.

B. Direct Microscopy

  1. Prepare smear and Gram stain.

  2. Examine for WBCs, epithelial cells, and bacteria.

C. Culture

  1. Inoculate sputum onto:

    • Blood agar: detects Streptococcus pneumoniae, Haemophilus

    • MacConkey agar: detects Klebsiella, Pseudomonas, E. coli

    • Chocolate agar: for fastidious organisms

  1. Incubate at 35–37°C for 18–24 hours.

  2. Examine growth and identify bacteria using biochemical tests.

  3. Perform antibiotic susceptibility testing.

5. Result Interpretation

  • Acceptable sputum: >25 WBCs and <10 epithelial cells per low power field

  • Contaminated sample: high epithelial cells → reject/repeat

  • Positive culture: Pure or predominant pathogenic colony growth

  • Sensitivity: Report antibiotics as Sensitive (S), Intermediate (I), or Resistant (R)

6. Uses

  • Diagnose bacterial lower respiratory infections

  • Guide targeted antibiotic therapy

  • Detect hospital-acquired pneumonia organisms

  • Differentiate bacterial vs viral pneumonia

7. Conclusion

Sputum culture is a key diagnostic tool for evaluating respiratory tract infections. Correct sample collection, timely processing, and interpretation using clinical correlation ensure accurate and meaningful results.

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