Microbiology a human perspective Chapter 10 Identification and classification of prokaryotes

Biochemical Tests: Catalase Test

-Bacteria that produce catalase break down hydrogen peroxide to release oxygen gas causing bubbling Also done on plates

Characterizing Strain Differences: Phage Typing

-Certain strains of a given species are susceptible to various bacteriophages a.k.a phage -Virus that infects bacteria kills bacteria and has been used as anti-bacterial agents -Phage typing identifies organisms by phage that infect them -Phage type has been largely replaced by molecular methods

Using Phenotype to Identify Prokaryotes: Metabolic Capabilities

-Identification relies heavily on analysis of metabolic capabilities -Culture characteristics *Colony morphology can give clues to identity *-Green pigment of Pseudomonas aeruginosa *-b-hemolytic colonies of Streptococcus pyogenes (beta="complete" hemolysis)

Characterizing Strain Differences: Antibiograms

-Identify organisms based on antibiotic susceptibility -Disc impregnated with antimicrobial placed on inoculated plate *-Clear indicates microbial susceptibility *-Different strains will have different susceptibility pattern

Using Phenotype to Identify Prokaryotes: Microscopic Morphology

-Important initial step in identification *Can be used to determine size, shape and staining characteristics -Size and shape can readily be determined microscopically -Gram stain differentiate Gram (+) from Gram (-) *-Narrows possible identities of organism -Special Stains *-Acid Fast Stain

Using Phenotype to Identify Prokaryotes: MACCONKEY AGAR

-identification of lactose fermenting, Gram-negative enteric pathogens (differential) -inhibiting growth of Gram-positive organisms (selective). -Fermentation of lactose turns the medium red/pink. (acidic environment from fermenting lactose).

Describe the information provided by: Staining reactions

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Explain how rRNA sequences can be used in identification and classification

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Explain the basis of nucleic acid hybridization and the kind of information it provides.

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Define Domain

A collection of similar kingdoms. The domain is a relatively new taxonomic category that reflects the characteristics of the cells that make up the organism.

Define Genus

A collection of similar species

Metabolic Capabilities: Biochemical Tests: Dichotomous Key

A flowchart of tests that give either a positive or negative result

Define Species

A group of closely related isolates or strains. Note that members of a species are not all identical; individual strains may vary in minor properties. The difficulty for the taxonomist is to decide how different two isolates must be in order to be classified as separate species rather than strains of the same species.

Using Genotype to Identify Prokaryotes: PCR: Sequencing Ribosomal RNA genes

AdvantageThere is little genetic variation in rRNA -16S rRNA is "gold standard" for identifying unknown bacteria and determining evolutionary relationships -Identification of organism that can't be grown in culture

Define Strain

An isolate; subgroup within a species

Metabolic Capabilities: Biochemical Tests: Breath Test

Assays for the presence of Urease, used to detect Helicobacter pylori

Describe the information provided by: biochemical reactions

Biochemical tests are most commonly used to identify various species of bacteria, but in some cases they can be used to distinguish different strains. A group of strains that have a characteristic biochemical pattern is called a biovar or a biotype.

Urease Test

Breakdown of urea by urease enzyme releases ammonia and CO2 leads to alkaline environment within tube as indicated by pink color

Describe the information provided by: DNA base ratios

Determining the G+C content offers a crude comparison of genomes. Organisms with identical G+C contents can be entirely unrelated, however.

Describe the basis for the identification of prokaryotes using: Genotypic characteristics

Differences in DNA sequences can be used to determine the point in time at which two organisms diverged from a common ancestor.

Sugar Fermentation

Fermentation of sugars results in acid production causing pH indicator to turn yellow (inverted tube w/n larger tube traps any gas produced)

Describe the information provided by: Genomic typing

Molecular methods such as pulsed-field gel electrophoresis can be used to detect restriction fragment length polymorphisms (RFLPs).

Describe the information provided by: Phage Typing

Pg 244 table 10.6 Strains of a given species sometimes differ in their susceptibility to various types of bacteriophage.

Describe the information provided by: serology

Proteins and carbohydrates that vary among strains can be used to differentiate strains. A group of strains that have a characteristic serological type is called a serovar or a serotype.

Describe the information provided by: Morphological studies

Size, shape, and staining characteristics such as Gram stain can give suggestive information as to the identity of the organism. Further testing, however, is needed to confirm the identification.

Describe the information provided by: DNA hybridization

The extent of nucleotide sequence similarity between two isolates can be determined by measuring how completely single strands of their DNA hybridize to one another.

Classification

The process of arranging organisms into similar or related groups (taxa), primarily to provide easy identification and study.

Identification

The process of characterizing an isolate in order to determine the group (taxon) to which it belongs

Taxonomy

The science that studies organisms in order to arrange them into groups (taxa); involves three interrelated areas--identification,classification, and nomenclature.

Nomenclature

The system of assigning names to organisms.

Explain why organisms are arranged in taxonomic groups

They are arranged in a hierarchical order, with the species being the basic unit. The species designation gives a formal taxonomic status to a group of related isolates or strains, which, in turn, permits their identification. Without classification, scientists and others would not be able to communicate about organisms with any degree of accuracy.

Describe the basis for the identification of prokaryotes using: Phenotypic characteristics

Traditionally, relatedness of different bacteria has been decided by comparing properties such as ability to degrade lactose and the presence of flagella. These characteristics, however, do not necessarily reflect the evolutionary relatedness of organisms.

Describe the information provided by: Numerical taxonomy

Uses a quantitive approach to phenotypic classification, comparing a battery of characteristics.

Describe the information provided by: Sequencing rRNA genes

pg 242 The nucleotide sequence of ribosomal RNA (rRNA) can be used to identify prokaryotes, particularly those difficult or currently impossible to grow in culture. The prokaryote 70S ribosome, which plays an indispensable role in protein syntesis, is composed of protein and three different rRNAs (5S, 16S, and 23S). Because of their highly constrained and essential function, the nucleotide sequence changes that can occur in the rRNA's, ;yet still allow the ribosome to operate, are limited.