Download Chapter 12 Microbial Evolution and Systematics

Chapter9 Microbial taxonomy
Three separate but interrelated parts:
1.
Classification: the arrangement of organisms
into groups or taxa based on mutual similarity
or evolutionary relatedness.
2. Nomenclature: the branch of taxonomy
concerned with the assignment of names to
taxonomic groups in agreement with
published rules.
3. Identification: the practical side of taxonomy,
the process of determining that a particular
isolate belongs to a recognized taxon.
Taxonomy is important for
several reasons
1.
2.
3.
4.
It allows us to organize huge amounts of
knowledge about organisms
Allows us to make predictions and frame
hypotheses for further research based on
knowledge of similar organisms.
It places microorganisms in meaningful, useful
groups with precise names so that
microbiologists can work with them and
communicate efficiently.
Identification of microorganisms accurately
Taxomomic ranks
Strain: one single isolate or line
 Type: sub-set of species
 Species: related strains
 Genus: related species
 Family: related genera
 Order; class; phylum; domain

Related concepts
A species: a collection of strains that have a similar
G+C composotion and 70% or greater similarity
as judged by DNA hybridization experiments.
A biovars: variant procaryotic strains characterized
by biochemical or physiological differences.
Morphovars: differ morphologically
Serovars: have distinctive antigenic properties
Type strain: it is usually one of the first strains
studied and often is more fully characterized than
other strains
Classification systems
Phenetic classification: one that groups organisms together
based on the mutual similarity of their phenotypic
characteristics.
Comparing as many attributes as possible.
Numerical taxonomy: computers may be used to analyze
data for the production of phenetic classification.
Information about the properties of organisms is
converted into a form suitable for numerical analysis and
then compared by means of a computer.
Phylogenetic classification: based on evolutionary
relationships rather than general resemblance.
defficult because of the lack of a good fossil record.
Comparision of genetic material and gene products
Major characteristics used in
taxonomy
Morphological characteristics
Physiological and metabolic
characteristics
Ecological characteristics
Genetic analysis
Molecular characteristics
Morphological characteristics
Cell shape
Cell size
Cilia and flagella
Cellular inclusions
Color
Mechanism of motility
Endospore shape and location
Spore morphology and location
Colonial morphology
Ultrastructural characteristics
Staining behavior
Physiological, metabolic and
Ecological characteristics
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Carbon and nitrogen
sources
Cell wall
constituents
Energy sources
Fermentation
products
Luminescence
Motility
Osmotic tolerance
Storage inclusions
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General nutritional type
Growth temperature
optimum and range
Mechanisms of energy
conversion
pH optimum and growth
range
Photosynthetic pigments
Salt requirements and
tolerance
Secondary metabolites
formed
Sensitivity to metabolic
inhibitors and antibiotics
Genetic analysis
The study of transformation and conjugation in
bacteria is sometimes taxonomically useful.
Plasmid-borne traits can cause errors in
bacterial taxonomy if care is not taken.
Transformation can occur between different
procaryotic species but only rarely between
genera.
E.coli can undergo conjugation with the genera
Salmonella and Shigella but not with Proteus
and Enterobacter
Molecular analysis
Historical
• guanine (G)+ cytosine (C)
(% GC)
Now
• Hybridization
• Gene characterization
– sequencing
– other
DNA-DNA hybridization
Strain 1
Heat
+
0% Homology
Strain 2
100% Homology
DNA-DNA hybridization
 Groups
bacterial strains into
species
 Below species level
• little or no relatedness
16S rRNA Sequencing
 similarity above species level
 allows relatedness comparisons of all
bacteria
 closely related bacterial species may
be identical
 development of clinical tests based on
sequence
Ribosomal RNAs as Evolutionary Chronometers
 Reasons:
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Ancient molecules
Functionally constant
Universally distributed
Moderately well conserved in sequence
across broad phylogenetic distances
The 16S rRNA or 18S rRNA Technique
The 16S rRNA or 18S rRNA Technique
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In Prokaryotes:
• 5S rRNA is too small, contains limited info
• 23S rRNA is too large, too difficult to manage
• 16S rRNA has the right size for studies
In Eukaryotes:
• 18S rRNA is used for phylogenetic measurements
The 16S rRNA or
18S rRNA Technique
The 16S rRNA or 18S rRNA Technique
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Ribosomal Database Project (RDP):
• http://www.cme.msu.edu/RDP
Compare your sequences with the database to
find out the organisms you identify
Phylogenetic
Trees from DNA
Sequences
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Distance-Matrix
Method for
generating the trees
Evolutionary
Distance (ED)
Computer compare
the sequence
differences and
build the
phylogenetic tree
based on corrected
ED
Phylogenetic Trees from DNA Sequences
Signature Sequences: unique to certain group of organisms
Applications: Phylogenetic Probes
“official” taxonomy and
nomenclature
Taxonomy: Bergey’s manual of systematic
bacteriology or Bergey’s manual of
determinative bacteriology
in some sence official
 Nomenclature: all new names are validly
published to gain standing in the
nomenclature, either by being published in
papers in the international Journal of
Systematic Bacteriology or, if published
elsewhere, by being announced in the
Validation Lists

Bergey’s mamual of systematic
bacteriology
The first edition: phenetic
Procaryotic groups are divided into four
volumes:
(1)gram-negative bacteria of general, medical,
or industrial importance.
(2)gram-positive bacteria other than
actinomycetes
(3)gram-negative bacteria with distinctive
properties, cyanobacteria, and archaea
(4)actinomycetes
Bergey’s mamual of systematic
bacteriology
The second edition: phylogenetic
Five volumes:
Volume 1: the archaea, and the deeply branching and
phototrophic bacteria
Volume 2: the Proteobacteria
Volume 3: the low G+C gram-positive bacteria
Volume 4: the high G+C gram-positive bacteria
Volume 5: the Planctomycetes, Spirochaetes,
Fibrobacteres, Bacteroidetes, and Fusobacteria
Table19.9 p441
Table19.8 p436
Identification
in the diagnostic laboratory
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Aids treatment
Susceptibility – antibiotic selection
Based on taxonomy
Simple, low cost, rapid
Steps in isolation and identification
• Step 1. Streaking culture plates
– colonies on incubation (e.g 24 hr)
– size, texture, color, hemolysis
– oxygen requirement
Blood Agar Plate
Isolation and identification
Step
2. Colonies Gram
stained
• cells observed
microscopically
Gram negative
Gram positive
Heat/Dry
Crystal violet stain
Iodine Fix
Alcohol de-stain
Safranin stain
Step 3. Isolated bacteria are speciated
 Generally
using physiological tests
Typical Culture
Laboratory Bench
Step 4.
Antibiotic susceptibility testing
Susceptible
Not susceptible
Bacterial
lawn
No
growth
Growth
Antibiotic disk
Rapid diagnosis without culture
• WHEN AND WHY?
• grow poorly
– Isolation slow
may not be clinically useful
• can not be cultured
– isolation impossible
Rapid “Strep” Test
Streptococcal
antigenic extract
Antibody
Latex beads
Bacterial DNA sequences amplified
directly from human body fluids
• Polymerase chain reaction (PCR)
• Great success in rapid diagnosis
of tuberculosis.
Microscopy
• spinal fluids
(meningitis脑膜炎)
• sputum (tuberculosis)
• sensitivity poor
Serologic identification
• antibody response to the infecting agent
• several weeks after an infection has occurred
Questions
1.
2.
3.
4.
5.
6.
7.
Why is Ribosomal RNAs used as Evolutionary
Chronometers
Why is 16S rRNA employed to study phylogenetics
rather than the smaller 5S rRNR and the large 23S
rRNA in Prokaryotics?
How to sequence 16S rRNA from inside the cells?
How to identify a prokaryotic or eukaryotic organism
based on 16S or 18S rRNA?
How to build Phylogenetic Trees from DNA Sequences?
Why it is said the archaea closer to eukarya than
bacteria is?
What is signature sequence and how can it be used?