classification of microorganisms -...

PowerPoint® Lecture

Presentations prepared by

Bradley W. Christian,

McLennan Community

College

C H A P T E R

© 2016 Pearson Education, Ltd.

Classification of

Microorganisms

10


The Study of Phylogenetic Relationships

• Taxonomy is the science of classifying organisms

• Shows degree of similarity among organisms

• Systematics, or phylogeny, is the study of the

evolutionary history of organisms


The Three Domains

• Developed by Woese in 1978; based on

sequences of nucleotides in rRNA

• Eukarya

• Animals, plants, fungi

• Bacteria

• Archaea

• Methanogens

• Extreme halophiles

• Hyperthermophiles


Figure 10.1 Three-Domain System.


Table 10.1 Some Characteristics of Archaea, Bacteria, and Eukarya


Table 10.2 Prokaryotic Cells and Eukaryotic Organelles Compared


The Three Domains

• Eukaryotes originated from infoldings of

prokaryotic plasma membranes

• Endosymbiotic bacteria developed into organelles


Figure 10.2 A model of the origin of eukaryotes.

Early cell Bacteria

Chloroplast

Archaea Mitochondrion

DNA

Eukarya


A Phylogenetic Tree

• Grouping organisms according to common

properties

• Fossils

• Genomes

• Groups of organisms evolved from a common

ancestor

• Each species retains some characteristics of

its ancestor


Scientific Nomenclature

• Common names vary with languages and

geography

• Binomial nomenclature is used worldwide to

consistently and accurately name organisms

• Genus

• Specific epithet (species)


Table 1.1 Making Scientific Names Familiar


Figure 10.5 The taxonomic hierarchy.


Classification of Prokaryotes

• Prokaryotic species: a population of cells with

similar characteristics

• Culture: bacteria grown in laboratory media

• Clone: population of cells derived from a single parent

cell

• Strain: genetically different cells within a clone


Figure 10.6 Phylogenetic relationships of prokaryotes.


Classification of Eukaryotes

• Protista: a catchall kingdom for a variety of

organisms; autotrophic and heterotrophic

• Grouped into clades based on rRNA

• Fungi: chemoheterotrophic; unicellular or

multicellular; cell walls of chitin; develop from

spores or hyphal fragments

• Plantae: multicellular; cellulose cell walls; undergo

photosynthesis

• Animalia: multicellular; no cell walls;

chemoheterotrophic


Classification of Viruses

• Not a part of any domain; not composed of cells;

require a host cell

• Viral species: population of viruses with similar

characteristics that occupies a particular

ecological niche


Methods of Classifying and Identifying

Microorganisms• Classification: placing organisms in groups of

related species

• Lists of characteristics of known organisms

• Identification: matching characteristics of an

"unknown" organism to lists of known organisms

• Clinical lab identification


Methods of Classifying and Identifying

Microorganisms• Morphological characteristics: useful for identifying

eukaryotes; tell little about phylogenetic

relationships

• Differential staining: Gram staining, acid-fast

staining; not useful for bacteria without cell walls

• Biochemical tests: determine presence of bacterial

enzymes


Figure 10.8 The use of metabolic characteristics to identify selected genera of enteric bacteria.


Applications of Microbiology 10.1


Biochemical Tests

• Rapid identification methods perform several

biochemical tests simultaneously

• Results of each test are assigned a number


One tube containing media for 15 biochemical tests

is inoculated with an unknown enteric bacterium.

After incubation, the tube is observed for results.

The value for each positive test is circled, and

the numbers from each group of tests are

added to give the code number.

Comparing the resultant code number with a

computerized listing shows that the organism in

the tube is Citrobacter freundii.

Glu

co

se

Gas

Lysin

e

Orn

ith

ine

H2S

Ind

ole

Ad

on

ito

l

Lacto

se

Ara

bin

ose

So

rbit

ol

V–

P

Du

lcit

ol

Ph

en

yla

lan

ine

Ure

ase

Cit

rate

Code Number Microorganism Atypical Test Results

62352

62353

Citrobacter freundii

Citrobacter freundii

Citrate

None

Figure 10.9 One type of rapid identification method for bacteria: EnteroPluri test from BD Diagnostics.


Serology

• The science that studies serum and immune

responses in serum

• Microorganisms are antigenic—they stimulate the

body to form antibodies in the serum

• In an antiserum, a solution of antibodies is tested

against an unknown bacterium


Serology

• In the slide agglutination test, bacteria

agglutinate when mixed with antibodies produced

in response to the bacteria

• Serological testing can differentiate between

species and strains within species


Figure 10.10 A slide agglutination test.


Serology

• Enzyme-linked immunosorbent assay (ELISA)

• Known antibodies and an unknown type of bacterium

are added to a well; a reaction identifies the bacteria

• Western blotting

• Identifies antibodies in a patient's serum; confirms HIV

infection


Figure 10.11 An ELISA test.


Figure 18.14a The ELISA method.


If Lyme disease is suspected in a patient:

Electrophoresis is used to separate Borrelia

burgdorferi proteins. Proteins move at

different rates based on their charge and size

when the gel is exposed to an electric current.

The bands are transferred to a nitrocellulose

filter by blotting. Each band consists of many

molecules of a particular protein (antigen). The

bands are not visible at this point.

The proteins (antigens) are positioned on the filterexactly as they were on the gel. The filter is thenwashed with patient's serum followed by antihumanantibodies tagged with an enzyme. The patientantibodies that combine with their specific antigenare visible (shown here in red) when the enzyme'ssubstrate is added.

The test is read. If the tagged antibodies stick tothe filter, evidence of the presence of themicroorganism in question—in this case, B.burgdorferi—has been found in the patient'sserum.

Lysed

bacteria

Larger

Polyacrylamide

gel

Proteins

Smaller

Paper towels

Salt

solution

Gel

Sponge

Nitrocellulose

filter

Figure 10.12 The Western blot.


Phage Typing

• Test for determining which phages a bacterium is

susceptible to

• On a plate, clearings called plaques appear where

phages infect and lyse bacterial cells


Figure 10.13 Phage typing of a strain of Salmonella enterica.


Fatty Acid Profiles

• FAME: Fatty acid methyl esters provide profiles

that are constant for a particular species


Flow Cytometry

• Uses differences in electrical conductivity between

species or fluorescence


Figure 18.12 The fluorescence-activated cell sorter (FACS).

Fluorescently

labeled cells

Laser beam

Laser

Fluorescence

detector

Electrically

charged

metal plates

Collection

tubes

The separated cells

fall into different

collection tubes.

As cells drop between

electrically charged

plates, the cells with

a positive charge

move closer to the

negative plate.

Electrode gives

positive charge to

identified cells.

Detector of

scattered light

Laser beam strikes

each droplet.

Cell mixture leaves

nozzle in droplets.

A mixture of cells is

treated to label cells

that have certain

antigens with

fluorescent-antibody

markers.

ElectrodeFluorescence detector

identifies fluorescent

cells by fluorescent

light emitted by cell.


DNA Base Composition

• DNA base composition

• Guanine + cytosine %

• Two organisms that are closely related have similar

amounts of various bases


DNA Fingerprinting

• DNA fingerprint

• Electrophoresis of restriction enzyme digests of an

organism's DNA

• Comparing fragments from different organisms provides

information on genetic similarities and differences


Figure 10.14 DNA fingerprints.


Nucleic Acid Amplification Tests (NAATs)

• Use of PCR to amplify DNA of an unknown

microorganism that cannot be cultured


Nucleic Acid Hybridization

• Nucleic acid hybridization measures the ability

of DNA strands from one organism to hybridize

with DNA strands of another organism

• Greater degree of hybridization, greater degree of

relatedness


Figure 10.15 DNA-DNA hybridization.


Nucleic Acid Hybridization

• Southern blotting uses nucleic acid hybridization

to identify unknown microorganisms using DNA

probes


Plasmid

SalmonellaDNAfragment

Unknown bacteria

are collected

on a filter.A Salmonella DNA

fragment is cloned in

E. coli.

The cells are lysed,

and the DNA

is released.

Cloned DNA fragments are markedwith fluorescent dye and separatedinto single strands, formingDNA probes. The DNA is separated into

single strands.

DNA probes are added

to the DNA from the

unknown bacteria.

Fluorescent probe

Salmonella DNA

DNA from

other bacteria

DNA probes hybridize withSalmonella DNA from sample.Then excess probe is washedoff. Fluorescence indicatespresence of Salmonella.

Figure 10.16 A DNA probe used to identify bacteria.


DNA Chips

• A DNA chip (also known as a microarray)

contains DNA probes and detects pathogens by

hybridization between the probe and DNA in the

sample

• Detected by fluorescence


Figure 10.17a-b DNA chip.


Figure 10.17c-d DNA chip.


DNA Chips

• Ribotyping

• rRNA sequencing

• Fluorescent in situ hybridization (FISH)

• Fluorescent DNA or RNA probes stain the

microorganisms being targeted

• Determines the identity, abundance, and relative activity

of microorganisms in an environment


Figure 10.18 FISH, or fluorescent in situ hybridization.


Putting Classification Methods Together

• Dichotomous keys

• Identification keys based on successive questions

• Cladograms

• Maps that show evolutionary relationships among

organisms; based on rRNA sequences


Figure 10.19 Building a cladogram.


Dichotomous Keys: Overview

PLAY Animation: Dichotomous Keys: Overview

dichotomous_keys.html


Dichotomous Keys: Sample with Flowchart

PLAY Animation: Dichotomous Keys: Sample

with Flowchart

dichotomous_sample.html


Dichotomous Keys: Practice

PLAY Animation: Dichotomous Keys: Practice

dichotomous_practice.html

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