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Microbiology

Healthcare-Associated

Healthcare-associated pneumonia (HCAP), including ventilator-associated pneumonia (VAP), poses a significant threat to hospitalized patients. Timely and accurate identification of the causative agents and detection of antimicrobial resistance are essential for guiding appropriate treatment and improving patient outcomes. Understanding the colony morphology and identification of these organisms is vital for clinical microbiology

General Principles

  • High Risk of Multidrug-Resistant Organisms (MDROs): HCAP is often caused by MDROs
  • Infection Control: Meticulous infection control practices are essential
  • Specimen Collection
    • Endotracheal Aspirate (ETA): Most common source
    • Bronchoalveolar Lavage (BAL): More invasive but yields better quality specimens
    • Bronchial Wash (BW)
    • Protected Specimen Brush (PSB): Quantitative cultures
  • Specimen Processing
    • Gram Stain: To provide initial findings and aid in guiding therapy
    • Culture: Performed on appropriate media. Selective media is often needed
    • Quantitative and Semi-quantitative Reporting: Valuable
  • Incubation: At 35-37°C with or without a CO2-enriched environment
  • Colony Morphology and Identification
    • Colony morphology is observed
    • Identification using standard laboratory techniques, biochemical testing
    • Commercial systems are used
  • Antimicrobial Susceptibility Testing: Performed on all significant isolates
  • Prompt Communication: Timely reporting of results

Major Pathogens: Colony Morphology, Gram Stain, and Identification

Acinetobacter baumannii Complex

  • Source: ETA, BAL
  • Colony Morphology
    • BAP: Gram-negative, oxidase-negative, non-fermentative
    • Appearance: Can vary, but often appear as plump, opaque, smooth, slightly raised, and gray-white to tan
  • Gram Stain: Gram-negative coccobacilli or short rods
  • Identification
    • Oxidase: Negative
    • Glucose Fermentation: Negative
    • Motility: Non-motile
    • Commercial Identification Systems or MALDI-TOF: Used for species-level identification. A. baumannii is often multi-drug resistant

Enterobacteriaceae (e.g., Klebsiella pneumoniae, Enterobacter spp., Serratia marcescens)

  • Source: ETA, BAL
  • Colony Morphology
    • BAP: Large, gray colonies, can be mucoid
    • MacConkey Agar: Pink, lactose-fermenting colonies
  • Gram Stain: Gram-negative rods
  • Identification
    • Oxidase: Negative
    • Lactose Fermentation: Variable
    • IMViC Tests: (Indole, Methyl Red, Voges-Proskauer, Citrate)
    • Commercial Identification Systems: Carbapenem-resistant organisms are important

Pseudomonas aeruginosa

  • Source: ETA, BAL
  • Colony Morphology
    • BAP: Large, flat, spreading colonies
    • Pigment Production: Often produces a blue-green pigment (pyocyanin) and/or a yellow pigment (pyoverdine)
    • Appearance: Can have a distinct fruity odor (grape-like)
  • Gram Stain: Gram-negative rods
  • Identification
    • Oxidase: Positive
    • Glucose Fermentation: Oxidative (uses glucose but does not ferment it)
    • Motility: Motile
    • Commercial Identification Systems

Stenotrophomonas maltophilia

  • Source: ETA, BAL
  • Colony Morphology
    • BAP: Gram-negative, oxidase-positive, non-fermentative
    • Appearance: Often lavender-green colonies
  • Gram Stain: Gram-negative rods
  • Identification
    • Oxidase: Positive
    • Glucose Fermentation: Negative
    • Motility: Motile
    • Lysine Decarboxylase: Positive
    • Commercial Identification Systems

Staphylococcus aureus

  • Source: ETA, BAL, blood, pleural fluid
  • Colony Morphology
    • BAP: Medium to large, circular, opaque, smooth, golden-yellow or cream-colored colonies
    • Hemolysis: Usually beta-hemolytic
  • Gram Stain: Gram-positive cocci in clusters
  • Identification
    • Catalase: Positive
    • Coagulase: Positive

Additional Considerations

  • Antibiotic Resistance: These organisms often exhibit resistance to multiple antibiotics
  • Reporting
    • Quantitative or semi-quantitative reporting is often used. Report the amount of the organism
    • Antimicrobial susceptibility testing results
  • Quality Control
    • Specimen quality
    • Sterility of equipment
  • Prompt Communication
    • Alert the clinicians immediately

Key Terms

  • Healthcare-Associated Pneumonia (HCAP): Pneumonia acquired in a healthcare setting
  • Ventilator-Associated Pneumonia (VAP): Pneumonia that develops in patients on mechanical ventilation
  • Multidrug-Resistant Organisms (MDROs): Microorganisms resistant to multiple antibiotics
  • Endotracheal Aspirate (ETA): Secretions collected through an endotracheal tube
  • Bronchoalveolar Lavage (BAL): Fluid obtained from the lungs using a bronchoscope
  • BAP (Blood Agar Plate): A general-purpose culture medium
  • Gram Stain: Staining technique to differentiate bacteria
  • Oxidase Test: Biochemical test to detect cytochrome c oxidase
  • Glucose Fermentation: The ability of a bacterium to break down glucose with the production of acid and/or gas
  • Motility: The ability of a bacterium to move independently
  • Lysine Decarboxylase: An enzyme that converts lysine to cadaverine
  • Commercial Identification System: System that uses pre-packaged biochemical tests for identifying microorganisms
  • MALDI-TOF Mass Spectrometry: A proteomic-based technique that provides rapid and accurate bacterial identification based on protein profiles
  • Antimicrobial Susceptibility Testing: Laboratory tests performed to determine the susceptibility of a bacterial isolate to various antibiotics
  • Quantitative Culture: Cultures where the bacterial counts are reported
  • Semi-quantitative Culture: Cultures where the bacterial growth is estimated (e.g., few, moderate, many)
  • Colony Morphology: The visual characteristics of bacterial colonies on culture media (e.g., size, shape, color, texture)
  • Pathogen: An organism that causes disease
  • Biofilm: A community of microorganisms that adhere to a surface and are encased in a matrix of extracellular substances