CH14 장

Brock
Biology of
Microorganis
ms
Chapter 14
Twelfth Edition
Madigan / Martinko
Dunlap / Clark
Microbial Evolution and Systematics

Lectures by Buchan & LeCleir
Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings
I. Early Earth and the Origin and
Diversification of Life
 14.1 Formation and Early History of Earth

 14.2 Origin of Cellular Life
 14.3 Microbial Diversification: Consequences for Earth’s
Biosphere
 14.4 Endosymbiotic Origin of Eukaryotes

14.1 Formation and Early History of
Earth
 The Earth is ~ 4.5 billion years old
 First evidence for microbial life can be found in

rocks ~ 3.86 billion years old (southwestern Green
land)

Ancient Microbial Life
3.45 billion-year-old rocks, South Africa

Figure 14.1

14.1 Formation and Early History of
Earth
 Stromatolites
 Fossilized microbial mats consisting of layers of filamentous
prokaryotes and trapped sediment
 Found in rocks 3.5 billion years old or younger
 Comparisons of ancient and modern stromatolites provide
evidence that
 Anoxygenic phototrophic filamentous bacteria formed ancient
stromatolites (relatives of the green nonsulfur bacterium
Chloroflexus)
 Oxygenic phototrophic cyanobacteria dominate modern
stromatolites

Ancient and Modern Stromatolites
3.5 billion yrs old 1.6 billion yrs old
Figure 14.2

More Recent Fossil Bacteria and
Eukaryotes
1 billion yrs old rocks
prokaryotes
eukaryotic cells
Figure 14.3

14.2 Origin of Cellular Life
 Early Earth was anoxic and much hotter than

present day (over 100 oC)
 First biochemical compounds were made by abiotic
systems that set the stage for the origin of life

 Surface Origin Hypothesis

 Contends that the first membrane-enclosed, self-
replicating cells arose out of primordial soup rich in
organic and inorganic compounds in ponds on Earth’s
surface
 Dramatic temperature fluctuations and mixing from
meteor impacts, dust clouds, and storms argue against
this hypothesis

 Subsurface Origin Hypothesis

 States that life originated at hydrothermal springs on
ocean floor
 Conditions would have been more stable
 Steady and abundant supply of energy (e.g., H2 and H2S)

may have been available at these sites

Submarine Mound Formed at Ocean
Hydrothermal Spring
Cooler, more oxidized, more

acidic ocean water
Hot, reduced, alkaline

hydrothermal fluid Figure 14.4

 Prebiotic chemistry of early Earth set stage for self-

replicating systems
 First self-replicating systems may have been RNA-
based (RNA world theory)
 RNA can bind small molecules (e.g., ATP, other
nucleotides)
 RNA has catalytic activity; may have catalyzed its own
synthesis

A Model for the Origin of Cellular Life
Last Universal Common Ancestor
Figure 14.5

 DNA, a more stable molecule, eventually became

the genetic repository
 Three-part systems (DNA, RNA, and protein)
evolved and became universal among cells

 Other Important Steps in Emergence of Cellular Life

 Build up of lipids
 Synthesis of phospholipid membrane vesicles that enclosed the
cell’s biochemical and replication machinery
 May have been similar to montmorillonite clay vesicles

Lipid Vesicles Made in the Laboratory
from Myristic Acid
vesicle
RNAs
Vesicles formed on Montmorillonite clay particles

Figure 14.6

 Last Universal Common Ancestor (LUCA)

 Population of early cells from which cellular life may
have diverged into ancestors of modern day Bacteria
and Archaea

 As early Earth was anoxic, energy-generating

metabolism of primitive cells was exclusively
 Anaerobic and likely chemolithotrophic
(autotrophic)
 Obtained carbon from CO2
 Obtained energy from H2; likely generated by H2S

reacting with FeS or UV light

Major Landmarks in Biological Evolution
Figure 14.7

A Possible Energy-Generating Scheme
for Primitive Cells
Figure 14.8

 Early forms of chemolithotrophic metabolism would

have supported production of large amounts of organic
compounds
 Organic material provided abundant, diverse, and
continually renewed source of reduced organic carbon,
stimulating evolution of various chemoorganotrophic
metabolisms

14.3 Microbial Diversification
 Molecular evidence suggests ancestors of Bacteria

and Archaea diverged ~ 4 billion years ago
 As lineages diverged, distinct metabolisms developed
 Development of oxygenic photosynthesis dramatically

changed course of evolution

 ~ 2.7 billion years ago, cyanobacterial lineages developed a

photosystem that could use H2O instead of H2S, generating
O2
 By 2.4 billion years ago, O2 concentrations raised to 1 part

per million; initiation of the Great Oxidation Event
 O2 could not accumulate until it reacted with abundant
reduced materials in the oceans (i.e., FeS, FeS2)

 Banded iron formations: laminated sedimentary rocks;
prominent feature in geological record

Banded Iron Formations
Iron oxides
Figure 14.9

 Development of oxic atmosphere led to evolution of

new metabolic pathways that yielded more energy
than anaerobic metabolisms

 Oxygen also spurred evolution of organelle-

containing eukaryotic microorganisms
 Oldest eukaryotic microfossils ~ 2 billion years old
 Fossils of multicellular and more complex eukaryotes

are found in rocks 1.9 to 1.4 billion years old

 Consequence of O2 for the evolution of life

 Formation of ozone layer that provides a barrier against
UV radiation
 Without this ozone shield, life would only have continued
beneath ocean surface and in protected terrestrial
environments

14.4 Endosymbiotic Origin of
Eukaryotes
 Endosymbiosis
 Well-supported hypothesis for origin of eukaryotic cells
 Contends that mitochondria and chloroplasts arose

from symbiotic association of prokaryotes within
another type of cell

Eukaryotes
 Two hypotheses exist to explain the formation of

the eukaryotic cell
1) Eukaryotes began as nucleus-bearing lineage that
later acquired mitochondria and chloroplasts by
endosymbiosis

Models for the Origin of the Eukaryotic
Cell
Figure 14.10a

Eukaryotes
 Two hypotheses exist to explain the formation of

the eukaryotic cell (cont’d)
2) Eukaryotic cell arose from intracellular association
between O2-consuming bacterium (the symbiont), which
gave rise to mitochondria and an archaean host

Models for the Origin of the Eukaryotic
Cell
Figure 14.10b

Eukaryotes
 Both hypotheses suggest eukaryotic cell is chimeric
 This is supported by several features

 Eukaryotes have similar lipids and energy metabolisms
to Bacteria
 Eukaryotes have transcription and translational
machinery most similar to Archaea

Major Features Grouping Bacteria or
Archaea with Eukarya
Table 14.1

II. Microbial Evolution
 14.5 The Evolutionary Process
 14.6 Evolutionary Analysis: Theoretical Aspects
 14.7 Evolutionary Analysis: Analytical Methods
 14.8 Microbial Phylogeny
 14.9 Applications of SSU rRNA Phylogenetic Methods

14.5 The Evolutionary Process
 Mutations
 Changes in the nucleotide sequence of an organism’s
genome
 Occur because of errors in the fidelity of replication, UV
radiation, and other factors
 Adaptative mutations improve fitness of an organism,
increasing its survival
 Other genetic changes include gene duplication,

horizontal gene transfer, and gene loss

14.6 Evolutionary Analysis: Theoretical
Aspects
 Phylogeny
 Evolutionary history of a group of organisms
 Inferred indirectly from nucleotide sequence data
 Molecular clocks (chronometers)

 Certain genes and proteins that are measures of evolutionary
change
 Major assumptions of this approach are that nucleotide changes

occur at a constant rate, are generally neutral, and random

Aspects
 The most widely used molecular clocks are small

subunit ribosomal RNA (SSU rRNA) genes
 Found in all domains of life
 16S rRNA in prokaryotes and 18S rRNA in eukaryotes
 Functionally constant
 Sufficiently conserved (change slowly)
 Sufficient length

Ribosomal RNA
16S rRNA
from E. coli
Figure 14.11

Aspects
 Carl Woese
 Pioneered the use of SSU rRNA for phylogenetic
studies in 1970s
 Established the presence of three domains of life:
 Bacteria, Archaea, and Eukarya
 Provided a unified phylogenetic framework for Bacteria

Aspects
 The Ribosomal Database Project (RDP)
 A large collection of rRNA sequences
 Currently contains > 409,000 sequences
 Provides a variety of analytical programs

14.7 Evolutionary Analysis: Analytical
Methods
 Comparative rRNA sequencing is a routine

procedure that involves
 Amplification of the gene encoding SSU rRNA
 Sequencing of the amplified gene
 Analysis of sequence in reference to other sequences

PCR-Amplification of the 16S rRNA Gene
Figure 14.12

General PCR Protocol

Methods
 The first step in sequence analysis involves

aligning the sequence of interest with sequences
from homologous (orthologous) genes from other
strains or species

Alignment of DNA Sequences
Figure 14.13

Methods
 BLAST (Basic Local Alignment Search Tool)

 Web-based tool of the National Institutes of Health
 Aligns query sequences with those in GenBank

database
 Helpful in identifying gene sequences

Methods
 Phylogenetic Tree
 Graphic illustration of the relationships among
sequences
 Composed of nodes and branches
 Branches define the order of descent and ancestry of
the nodes
 Branch length represents the number of changes that
have occurred along that branch

Phylogenetic Trees: Unrooted (a) and
Rooted (b-d) Forms
Figure 14.14

Methods
 Evolutionary analysis uses character-state methods

(cladistics) for tree reconstruction
 Cladistic methods
 Define phylogenetic relationships by examining changes in
nucleotides at individual positions in the sequence
 Use those characters that are phylogenetically informative
and define monophyletic groups (a group which contains all
the descendants of a common ancestor; a clade)

Identification of Phylogenetically
Informative Sites
Dots: neutral sites.

Arrows: phylogenetically informative sites.
Figure 14.15

Methods
 Common cladistic methods

 Parsimony
 Maximum likelihood
 Bayesian analysis

14.8 Microbial Phylogeny
 The universal phylogenetic tree based on SSU rRNA genes is a

genealogy of all life on Earth
Animation: Generating Phylogenetic Trees

Universal Phylogenetic Tree as
Determined by rRNA Genes
Figure 14.16

 Domain Bacteria
 Contains at least 80 major evolutionary groups (phyla)
 Many groups defined from environmental sequences
alone
 i.e., no cultured representatives
 Many groups are phenotypically diverse

 i.e., physiology and phylogeny not necessarily linked

 Eukaryotic organelles originated within Bacteria

 Mitochondria arose from Proteobacteria
 Chloroplasts arose from the cyanobacterial phylum

 Domain Archaea consists of two major groups

 Crenarchaeota
 Euryarchaeota

 Each of the three domains of life can be

characterized by various phenotypic properties

Major Features Distinguishing
Prokaryotes from Eukarya

Major Features Distinguishing
Prokaryotes from Eukarya

14.9 Applications of SSU rRNA
Phylogenetic Methods
 Signature Sequences
 Short oligonucleotides unique to certain groups of organisms
 Often used to design specific nucleic acid probes
 Probes
 Can be general or specific
 Can be labeled with fluorescent tags and hybridized to rRNA
in ribosomes within cells
 FISH: fluorescent in situ hybridization
 Circumvent need to cultivate organism(s)

Fluorescently Labeled rRNA Probes:
Phylogenetic Stains
Stained with universal Stained with a

rRNA probe eukaryotic rRNA probe
Figure 14.17

 PCR can be used to amplify SSU rRNA genes from

members of a microbial community
 Genes can be sorted out, sequenced, and analyzed
 Such approaches have revealed key features of microbial

community structure and microbial interactions

 Ribotyping
 Method of identifying microbes from analysis of DNA
fragments generated from restriction enzyme digestion
of genes encoding SSU rRNA
 Highly specific and rapid
 Used in bacterial identification in clinical diagnostics

and microbial analyses of food, water, and beverage

Ribotyping
Figure 14.18

III. Microbial Systematics
 14.10 Phenotypic Analysis

 14.11 Genotypic Analysis
 14.12 Phylogenetic Analysis
 14.13 The Species Concept in Microbiology
 14.14 Classification and Nomenclature

14.10 Phenotypic Analysis
 Taxonomy
 The science of identification, classification, and nomenclature
 Systematics
 The study of the diversity of organisms and their relationships
 Links phylogeny with taxonomy

 Bacterial taxonomy incorporates multiple methods

for identification and description of new species
 The polyphasic approach to taxonomy uses three
methods
1) Phenotypic analysis
2) Genotypic analysis
3) Phylogenetic analysis

 Phenotypic analysis examines the morphological,

metabolic, physiological, and chemical characters
of the cell

Some Phenotypic Characteristics of
Taxonomic Value
Table 14.3

Some Phenotypic Characteristics of
Taxonomic Value
Table 14.3

 Fatty Acid Analyses (FAME: fatty acid methyl ester)

 Relies on variation in type and proportion of fatty acids
present in membrane lipids for specific prokaryotic
groups
 Requires rigid standardization because FAME profiles
can vary as a function of temperature, growth phase,
and growth medium

Fatty Acid Methyl Ester (FAME) Analysis
Figure 14.19a

Fatty Acid Methyl Ester (FAME) Analysis
Figure 14.19b

14.11 Genotypic Analysis
 Several methods of genotypic analysis are available and used

 DNA-DNA hybridization
 DNA profiling
 Multilocus Sequence Typing (MLST)
 GC Ratio

Some Genotypic Methods Used in
Bacterial Taxonomy

 Genomes of two organisms are hybridized to examine
proportion of similarities in their gene sequences

Genomic Hybridization as a Taxonomic
Tool
Figure 14.20a

Tool
Figure 14.20b

Tool
Figure 14.20c

 Provides rough index of similarity between two
organisms
 Useful complement to SSU rRNA gene sequencing
 Useful for differentiating very similar organisms
 Hybridization values 70% or higher suggest strains

belong to the same species
 Values of at least 25% suggest same genus

Relationship Between SSU rRNA and
DNA Hybridization
97
95
25
Figure 14.21

 DNA Profiling
 Several methods can be used to generate DNA
fragment patterns for analysis of genotypic similarity
among strains, including
 Ribotyping: focuses on a single gene
 Repetitive extragenic palindromic PCR (rep-PCR)

and Amplified fragment length polymorphism
(AFLP): focus on many genes located randomly
throughout genome

DNA Fingerprinting with rep-PCR
Figure 14.22

 Multilocus Sequence Typing (MLST)

 Method in which several different “housekeeping
genes” from an organism are sequenced (~450-bp)
 Has sufficient resolving power to distinguish between
very closely related strains

Multilocus Sequence Typing

 GC Ratios
 Percentage of guanine plus cytosine in an organism’s
genomic DNA
 Vary between 20 and 80% among Bacteria and
Archaea
 Generally accepted that if GC ratios of two strains differ
by ~ 5% they are unlikely to be closely related

14.12 Phylogenetic Analysis
 16S rRNA gene sequences are useful in taxonomy;

serve as “gold standard” for the identification and
description of new species
 Proposed that a bacterium should be considered a new
species if its 16S rRNA gene sequence differs by more
than 3% from any named strain, and a new genus if it
differs by more than 5%

 The lack of divergence of the 16S rRNA gene limits its

effectiveness in discriminating between bacteria at the
species level, thus, a multi-gene approach can be used
 Multi-gene sequence analysis is similar to MLST, but
uses complete sequences and comparisons are made
using cladistic methods

 Whole-genome sequence analyses are becoming

more common
 Genome structure; size and number of chromosomes,
GC ratio, etc.
 Gene content
 Gene order

14.13 The Species Concept in
Microbiology
 No universally accepted concept of species for
prokaryotes
 Current definition of prokaryotic species
 Collection of strains sharing a high degree of similarity
in several independent traits
 Most important traits include 70% or greater DNA-DNA
hybridization and 97% or greater 16S rRNA gene
sequence identity

Taxonomic Hierarchy for
Allochromatium warmingii

Microbiology
 Biological species concept not meaningful for
prokaryotes as they are haploid and do not undergo
sexual reproduction
 Genealogical species concept is an alternative
 Prokaryotic species is a group of strains that based on
DNA sequences of multiple genes cluster closely with
others phylogenetically and are distinct from other
groups of strains

Multi-Gene Phylogenetic Analysis
16S rRNA genes

gyrB genes
luxABFE genes
Figure 14.24

Microbiology
 Ecotype
 Population of cells that share a particular resource
 Different ecotypes can coexist in a habitat
 Bacterial speciation may occur from a combination

of repeated periodic selection for a favorable trait
within an ecotype and lateral gene flow

A Model for Bacterial Speciation
Figure 14.25

Microbiology
 This model is based solely on the assumption of

vertical gene flow
 New genetic capabilities can also arise by horizontal
gene transfer; the extent among bacteria is variable

Microbiology
 No firm estimate on the number of prokaryotic

species
 Nearly 7,000 species of Bacteria and Archaea are
presently known

14.14 Classification and Nomenclature
 Classification
 Organization of organisms into progressively more
inclusive groups on the basis of either phenotypic
similarity or evolutionary relationship

 Prokaryotes are given descriptive genus names and

species epithets following the binomial system of
nomenclature used throughout biology
 Assignment of names for species and higher groups of
prokaryotes is regulated by the Bacteriological Code
- The International Code of Nomenclature of Bacteria

 Major references in bacterial diversity

 Bergey’s Manual of Systematic Bacteriology (Springer)
 The Prokaryotes (Springer)

 Formal recognition of a new prokaryotic species

requires
 Deposition of a sample of the organism in two culture
collections
 Official publication of the new species name and description
in the International Journal of Systematic and Evolutionary
Microbiology (IJSEM)
 The International Committee on Systematics of

Prokaryotes (ICSP) is responsible for overseeing
nomenclature and taxonomy of Bacteria and Archaea

Some National Microbial Culture
Collections
KCCM Korean Culture Center of Microorganisms Seoul, Korea http://www.kccm.or.kr

KACC Korean Agricultural Culture Collection Suwon, Korea http://kacc.rda.go.kr
Table 14.6

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Brock

Microbial Evolution and Systematics

 14.1 Formation and Early History of Earth

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 The Earth is ~ 4.5 billion years old

 First evidence for microbial life can be found in

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

3.45 billion-year-old rocks, South Africa

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

3.5 billion yrs old 1.6 billion yrs old

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Early Earth was anoxic and much hotter than

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Surface Origin Hypothesis

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Subsurface Origin Hypothesis

 Steady and abundant supply of energy (e.g., H2 and H2S)

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

Cooler, more oxidized, more

Hot, reduced, alkaline

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Prebiotic chemistry of early Earth set stage for self-

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

Last Universal Common Ancestor

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 DNA, a more stable molecule, eventually became

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Other Important Steps in Emergence of Cellular Life

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

Vesicles formed on Montmorillonite clay particles

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Last Universal Common Ancestor (LUCA)

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 As early Earth was anoxic, energy-generating

 Obtained energy from H2; likely generated by H2S

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Early forms of chemolithotrophic metabolism would

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Molecular evidence suggests ancestors of Bacteria

 Development of oxygenic photosynthesis dramatically

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 ~ 2.7 billion years ago, cyanobacterial lineages developed a

 By 2.4 billion years ago, O2 concentrations raised to 1 part

 O2 could not accumulate until it reacted with abundant

reduced materials in the oceans (i.e., FeS, FeS2)

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Development of oxic atmosphere led to evolution of

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Oxygen also spurred evolution of organelle-

 Fossils of multicellular and more complex eukaryotes

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Consequence of O2 for the evolution of life

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Contends that mitochondria and chloroplasts arose

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings

 Two hypotheses exist to explain the formation of

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings