Identifying and Classifying Microorganisms

Principles of Taxonomy

  • Taxonomy is the science of classifying organisms into groups, or taxa, based on phenotypic and genotypic relatedness. It encompasses:
    • Identification: Characterizing organisms for grouping.
    • Classification: Arranging organisms into similar groups.
    • Nomenclature: Assigning universal names.
  • The species is the basic taxonomic unit:
    • Eukaryotes are groups capable of producing fertile offspring.
    • Prokaryotes are groups of closely related isolates or strains (genetic variants). Prokaryotes have no Kingdom designation.
  • Phylogeny represents evolutionary relatedness. The Three-domain system, established by Carl Woese et al., is based on rRNA sequence analysis.

Phenotypic Identification Methods

  • Microscopic Morphology: Size, shape, and staining (Gram stain, acid-fast, endospore, capsule) provide initial diagnostic information.
  • Culture Characteristics: Colony appearance and pigments help identify microbes.
    • Serratia marcescens produces a red pigment at 22∘C22^{\circ}\text{C}.
    • Pseudomonas aeruginosa produces a green pigment and a fruity odor.
    • Differential media like MacConkey agar identify E. coli by lactose fermentation (pink colonies).
  • Metabolic Capabilities: Biochemical tests like Catalase, sugar fermentation, and Urease production provide conclusive identification via pH indicators.
    • Dichotomous keys (flowcharts) utilize successive morphological or biochemical questions to differentiate species.
  • Serology: Uses commercially available antibodies to detect specific antigens (proteins/polysaccharides) in the cell wall, capsule, flagella, or pili, such as in Streptococcus pyogenes.

Genotypic Identification Methods

  • Genotyping: Identifies unique nucleotide sequences, allowing the identification of organisms that cannot be grown in culture.
  • Nucleic Acid Amplification Tests (NAATs): PCR and other techniques amplify sequences to detect small numbers of organisms.
    • Used for slow-growing pathogens like Mycobacterium tuberculosis (3−43-4 hours vs. up to 66 weeks for traditional culture).
  • Sequencing Ribosomal RNA Genes: Focuses on 16S16S (18S18S in eukaryotes) rDNA because it has little genetic variation. Sequences are compared against databases to find the nearest relative.

Characterizing Strain Differences

  • Serological Typing: Distinguishes strains based on antigens. Example: E. coli O157:H7 (O = lipopolysaccharide antigen; H = flagella antigen).
  • Molecular Typing: DNA fingerprinting uses restriction enzymes to create Restriction Fragment Length Polymorphisms (RFLPs) to trace outbreaks among phenotypically identical strains.
  • Antibiograms: Patterns of clearing (susceptibility) in response to antibiotic-impregnated discs distinguish different microbial strains.