Classificationof Prokaryotes

Extreme halophiles

Hot springs, home of thermophiles

Bacterial Taxonomy

“Taxonomy is the science that studies organisms in order to arrange them into groups; those organisms with similar properties are grouped together and separated from those that are different.”

Classification--the process of arranging organisms into similar or related groups, primarily to provide easy identification and study

Identification--the process of characterizing organisms. Practical use of classification scheme

Nomenclature--the system of assigning names to organisms.”

Identification

to identify fecally contaminated water.

to identify organisms associated with food spoilage or contamination.

to identify pathogens in the clinical microbiology laboratory.

It is important to keep in mind that the appropriate tests to employ will depend on the organisms likely present plus time, skills of the tester, and budgetary concerns.

In the clinic, patient symptoms help to define what organisms of interest are likely present.

Classification

Phenotype-based

Genotype-based

Identification vs. Classification

Classification of Bacteria (Linnaean)Classification of Bacteria (Linnaean)

*Kingdom Phylum/Division Class Order Family *Genus ( 1st name) *Species ( 2nd name

identifier)

King Philip Came Over For Good Spaghetti Escherichia coli

*Kingdom Prokaryotae Phylum/Division Gracilicutes Class Scotobacteria Order Eubacteriales Family

Enterobacteriaceae *Genus Escherichia *Species coli

Criterria for Classification

Growth on Media Non selective media Selective media Differential media Bacterial Microscopy Gram stain Biochemical Tests Oxidase, catalase

Criterria for Classification

Immunologic tests. Antibody antigen tests Genetic instability nucleic acid hybridization DNA sequence analysis

Classification Systems

Keys Numerical Taxonomy (Analytical profile

Index) API Phylogenetic Classification (GC content,

DNA sequencing) Ribosomal RNA( most conserved throughout

evolution)

Phylogenetic Tree

16S ribosomal RNA Line lengths are based on genetic

distances, i.e., longer lines reflect greater differences.

Identification

Phenotypic Characteristics

Microscopy: Gram stain, acid-fast stain, cell morphology, cell arrangements.

Colony morphology: size, color, border shape, etc. Growth on Differential media: blood agar, MacConkey

agar, etc. Growth on Selective/Differential media: MacConkey

agar, EMB agar, etc. Biochemical tests: “Most biochemical tests rely on a

pH indicator or chemical reaction that results in a color change [or gas production] when a [specific] compound is degraded.”

Catalase Test

Durham Tube: Gas Production

Urease Test

Commercial Biochemical Test Kits

Dichotomous Key

Typing of Strains

Biovar, Biotype (syn)

Serovar, Serotype (syn)

Typing via RFLP analysis

Ribotype (RFLP of rRNA) Also Phage Type

Restriction Fragment LengthPolymorphism (RFLP) Analysis

Genomic DNA is isolated, cut using specific restriction

enzymes, run out on a gel, and then Southern Blotted with specific

DNA probes.

Can you tell me which lanes contain

bacteria that are clonally related?

1 2 3 4 5

Southern Blotting RFLP generation.

Gels separate DNA by size.

This is a post-probe detection

blot.

Phage TypingPhages are viruses of bacteria.

The clear spots are due

to bacteria killing (lysis).

This is a “Soft-Agar Overlay.”

“Phage” = “Bacteriophage”

AntibiogramThe disks are impregnated with specific antibiotics.

The streaks make up a bacterial “lawn”.

These two strains have different antibiotic-resistance patterns.

Numerical Taxonomy

Numerical Taxonomy is a method of classification that pools phenotypic characteristics.

The key to numerical taxonomy is that no one trait (e.g., ability to ferment glucose) is elevated in importance above any other trait (e.g., Gram staining characteristics).

Instead, all traits are weighted equally.

Classification in numerical taxonomy is expressed in terms of a Similarity Coefficient found between two compared strains.

Greater similarity coefficient, closer relatedness is inferred. Greater than ~70% and inference is that two compared bacteria are of the same species.

G-C Content (DNA Base Ratio)

Helix has higher stability with

greater fraction G-C vs. A-T pairing.

Higher melting points therefore correspond to

greater G-C content

DNA-DNA Hybridization

Similarity is in terms of nucleotide sequence

between the two compared organisms.