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Microbiology

Cerebrospinal Fluid

This section covers the essential laboratory procedures involved in the diagnosis and management of cerebrospinal fluid (CSF) infections, particularly those related to acute meningitis and shunt infections. It emphasizes the integrated approach needed for accurate interpretation and timely intervention

Specimen Collection and Initial Processing

  • Specimen Source
    • Lumbar Puncture (LP): CSF collection from the lumbar cistern for suspected meningitis or other neurological conditions
    • Shunt/Reservoir: Collection from implanted devices for patients with hydrocephalus or ventriculitis
  • Aseptic Technique: Critical to prevent contamination
  • Volume: Adequate volume (1-3 mL per tube) for all necessary tests
  • Transport and Storage: Prompt transport at room temperature; store refrigerated if processing is delayed
  • Tube Order: Microbiology prioritized (typically tube 2) to minimize contamination

Colony Morphology and Identification of Major Pathogens

  • Core Competency: Recognizing and identifying key bacterial and fungal pathogens from CSF cultures is essential
  • Key Procedures
    • Gram Stain: Immediate assessment to identify bacteria and provide rapid presumptive identification
    • Culture
      • Media: BAP (Blood Agar), CHOC (Chocolate Agar), with consideration for selective media as needed
      • Incubation: 35-37°C, 5% CO2 atmosphere, incubate for at least 48-72 hours or longer
      • Examination: Colony morphology, Gram stain, etc. are examined
      • Identification: Follow standard laboratory procedures
  • Major Pathogens and Characteristics
    • Streptococcus pneumoniae: Alpha-hemolytic, Gram-positive lancet-shaped diplococci; optochin susceptible
    • Haemophilus influenzae: Gram-negative coccobacilli, requires X and V factors
    • Neisseria meningitidis: Gram-negative diplococci, oxidase positive, utilizes glucose and maltose
    • Escherichia coli: Gram-negative rods, lactose-fermenting, oxidase-negative
    • Listeria monocytogenes: Gram-positive rods/coccobacilli, beta-hemolytic, catalase-positive, tumbling motility
    • Enterobacteriaceae (e.g., Klebsiella, Proteus): Gram-negative rods, variable lactose fermentation, oxidase-negative
    • Staphylococcus aureus: Gram-positive cocci in clusters, beta-hemolytic, coagulase-positive
    • Beta-hemolytic streptococci: Beta-hemolytic, Gram-positive cocci in chains
    • Other Considerations: Knowledge of morphology and colony appearance on different media

Common Agents of Shunt Infections

  • Focus: Infections of CSF shunts
  • Key Agents
    • Coagulase-Negative Staphylococci (CoNS): Most common, especially S. epidermidis, biofilm formation
    • Staphylococcus aureus: More acute infections
    • Corynebacterium spp. (Diphtheroids): Often C. jeikeium, opportunistic pathogens
    • Propionibacterium spp./Cutibacterium spp. (e.g., P. acnes): Anaerobic
  • Analytic Considerations
    • Anaerobic cultures for suspected Propionibacterium
    • Extended incubation times
    • Antimicrobial susceptibility testing (AST) and biofilm formation
    • Prompt communication of results
  • Other Agents: Enterobacteriaceae, Pseudomonas aeruginosa, Enterococcus spp.

Correlation with Other Laboratory Results

  • Integrated Interpretation: Linking culture results with other CSF parameters and clinical presentation is essential for accuracy
  • Key Correlations
    • Bacterial Meningitis: Elevated WBC (neutrophils), low glucose, high protein, positive Gram stain
    • Viral Meningitis: Elevated WBC (lymphocytes), normal glucose, normal or slightly elevated protein, negative Gram stain
    • Fungal/Tuberculous Meningitis: Elevated WBC (lymphocytes), low glucose, high protein, variable Gram stain/acid-fast stain, cultures essential
    • Shunt Infections: Variable WBC, low glucose, high protein, and the presence of organisms
    • Traumatic Tap vs. Subarachnoid Hemorrhage: RBC counts, xanthochromia

Antigen Detection and Molecular Methods

  • Rapid and Sensitive: These methods offer rapid diagnostics and complement traditional culture
  • Antigen Detection
    • Principle: Detects specific bacterial or fungal antigens in CSF (e.g., latex agglutination, EIAs)
    • Benefits: Rapid results, high specificity
    • Targeted Pathogens: S. pneumoniae, H. influenzae, N. meningitidis, Cryptococcus neoformans, Group B Streptococcus
  • Molecular Methods
    • PCR/qPCR: Amplifies specific DNA or RNA sequences
    • RT-PCR: For RNA viruses
    • Multiplex PCR: Simultaneous detection of multiple pathogens
    • NGS (Next Generation Sequencing): Advanced identification method
    • Benefits: High sensitivity and specificity, rapid results, can detect non-culturable organisms, can provide quantitative results
    • Considerations: Cost, training, and potential for detecting non-viable organisms

Organism Pathogenicity

  • Understanding Disease: Pathogenicity explains how microorganisms cause disease
  • Key Components
    • Etiology: The cause of the disease
    • Transmission: How the organism spreads
    • Virulence Mechanisms: The tools the organism uses to cause disease
  • Examples (brief)
    • S. pneumoniae: Capsule, adhesins, pneumolysin
    • H. influenzae: Capsule, endotoxin, adhesins
    • N. meningitidis: Capsule, endotoxin, adhesins, outer membrane proteins
    • E. coli: Adhesins, capsule, endotoxin
    • Brucella spp.: Intracellular survival
  • Implications of Pathogenicity: Guiding diagnosis, determining treatment, and informing prevention strategies
  • Transmission: How a pathogen spreads