Identifying and Classifying Microorganisms

The classification of bacteria has evolved significantly from the late 19th century to the present, moving from physical observations to genetic sequencing.
1870s: Ferdinand Cohn was the first to classify bacteria based on their shape.
1908: Sigurd Orla-Jensen pioneered the classification of bacteria based on their physiological properties.
1930s: Albert Kluyver and C. B. van Niel proposed classification systems based on evolutionary relationships.
1970: Roger Stanier determined relationships by comparing physical traits and nucleotide sequences.
Late 1970s: Carl Woese revolutionized the field by dividing prokaryotes into two major groups based on ribosomal RNA (rRNA) sequences.
Current Three-Domain System: The work of Carl Woese led to the currently accepted system consisting of three domains: Bacteria, Archaea, and Eukarya.

Taxonomy: The science of characterizing and naming organisms to arrange them into hierarchical groups known as taxa.
Taxonomy consists of three separate but deeply interrelated areas:
- Identification: The process of characterizing an organism in order to determine the group to which it belongs.
- Classification: The process of arranging organisms into similar or related groups.
- Nomenclature: The formal system of assigning names to organisms.

Multiple strategies are employed to identify microorganisms, often prioritizing practical clinical utility over evolutionary history:
- Microscopic examination: Investigating morphology and size.
- Culture characteristics: Observing how organisms grow in a laboratory setting.
- Biochemical tests: Analyzing metabolic capabilities.
- Nucleic acid analysis: Utilizing molecular techniques for identification.
- Patient symptoms: Used specifically for identifying pathogens in a clinical context.
In clinical settings, identification is often more critical than determining evolutionary relationships. Morphology and size are typically the most helpful initial clues for clinical identification.

Microscopic Morphology: This is a crucial initial step in identification.
- It quickly determines the size, shape, and staining characteristics of an organism.
- In some cases, microscopic morphology is sufficient to diagnose eukaryotic infections.
Gram Stain: This technique distinguishes between Gram-positive and Gram-negative bacteria.
- The result of a Gram stain can be sufficient to initiate appropriate medical therapy.
Special Stains: Specific stains are used to identify unique organisms.
- Acid-fast stain: Specifically helps identify Mycobacterium tuberculosis.

Observing the growth of organisms in culture provides vital clues to their identity:
- Serratia marcescens: Colonies typically appear red when grown at $22\,^{\circ}\text{C}$ .
- Pseudomonas aeruginosa: Often produces a distinct green pigment and a characteristic fruity odor.
Differential Media: Helps identify specific organisms based on their biological reactions to the media.
- Streptococcus pyogenes (the cause of strep throat): Yields $\beta$ -hemolytic colonies on blood agar and is catalase-negative.
- Escherichia coli (common cause of urinary tract infections): Ferments lactose to form pink colonies on MacConkey agar.

Microorganisms can be identified through a hierarchical series of metabolic tests:
Gram-Positive Cocci Path:
- Catalase Test: Distinguishes between different groups.
  - Catalase-Negative: Enterococcus sp.
  - Catalase-Positive: Leads to the Coagulase Test.
    - Coagulase-Positive: Staphylococcus aureus.
    - Coagulase-Negative: Staphylococcus saprophyticus.
Gram-Negative Rod Path:
- Oxidase Test:
  - Oxidase-Positive: Pseudomonas aeruginosa.
  - Oxidase-Negative: Leads to Lactose Fermentation testing.
    - Lactose Fermenter (Positive): E. coli or other coliforms.
    - Non-Lactose Fermenter (Negative): Proteus sp.
Commercial Kits: These allow for rapid identification via a battery of biochemical tests. They require an incubation period, after which the pattern of results is scored and processed by a computer to identify the organism.

Serological testing utilizes antibodies to detect specific molecules that serve as identifying markers.
Identifying markers often include proteins and polysaccharides of prokaryotic cells.
The most useful markers for identification include surface structures:
- Cell wall
- Capsule
- Flagella
- Pili
Certain species, such as those within the genus Streptococcus, contain unique carbohydrates in their cell walls that facilitate identification.

Differentiating between strains of the same species is vital for foodborne illness investigations, forensic investigations (including bioterrorism and biocrimes), and diagnosing specific diseases.
Biochemical Typing: Groups organisms with characteristic patterns into a biovar or biotype. This method can be used to trace specific strains such as Vibrio cholerae El Tor.
Serological Typing: Groups organisms with characteristic antigens into a serovar or serotype.
- E. coli strains are distinguished by the antigenic type of their flagella, capsules, and lipopolysaccharide molecules.
- Example - E. coli O157:H7: The "O" antigen refers to the lipopolysaccharide molecule, and the "H" antigen refers to the flagella.