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Figure 2.11b
Cytoplasmic
membrane
Endoplasmic
reticulum
Ribosomes
Nucleus
Nucleolus
Nuclear
membrane
Golgi
complex
Cytoplasm
Mitochondrion
Chloroplast
Eukaryote
© 2012 Pearson Education, Inc.
Figure 2.11a
Cytoplasm
Nucleoid
Ribosomes
Plasmid
Cell wall
Prokaryote
© 2012 Pearson Education, Inc.
Cytoplasmic
membrane
Bacterial Structures
Microscopic Techniques: Dyes and
Staining
•Simple stains
•Basic dyes with positive charge stick to
cells
•Acid dyes provide background stain
•Differential stains
Gram stain - separates
bacteria into two categories
based on type of cell wall
Acid Fast Stain
Gram-positive
Gram-negative
Gram-positive Cell Wall
Thick layer of peptidoglycan
Teichoic acids
Gram-negative Cell Wall
Thin layer of peptidoglycan
Outer membrane – and cytoplasmic membrane
lipopolysaccharide
(LPS)
Morphology of
Prokaryotic Cells:
Cell Groupings
Flagella - motility
E. coli O157:H7
Rotate like a propeller
Proton motive force used for energy
Presence/arrangement can be used as an
identifying marker
Cell Wall
Provides rigidity to the cell
(prevents it from bursting)
Cell Wall
•Peptidoglycan - rigid molecule; unique
to bacteria
•Alternating subunits of NAG and NAM
form glycan chains
•Glycan chains are connected to each
other via peptide chains on NAM molecules
Cell Wall
Cell Wall
Gram-negative
Thin layer of peptidoglycan
Outer membrane - additional
membrane barrier; porins permit passage
lipopolysaccharide (LPS)
- ex. E. coli O157:H7
endotoxin
- recognized by innate immune system
Cytoplasmic membrane
•Defines the boundary of the cell
•Semi-permeable; excludes all
but water, gases, and some
small hydrophobic molecules
•Transport proteins function as
selective gates (selectively permeable)
•Control entrance/expulsion of
antimicrobial drugs
•Receptors provide a sensor system
•Phospholipid bilayer, embedded with proteins
Cytoplasmic membrane
Electron transport chain - Series of proteins that eject protons from the cell,
creating an electrochemical gradient
Proton motive force is used to fuel:
•Synthesis of ATP (the cell’s energy currency)
•Rotation of flagella (motility)
•One form of transport
If a function of the cell membrane is
transport…..
• How is material transported in/out of the cell?
– Passive transport
• No ATP
• Along concentration gradient
– Active transport
• Requires ATP
• Against concentration gradient
Active Transport
Internal Structures:
Endospores
Enzymes bind substrate and generate a
product, enzyme is unchanged
Some enzymes require a cofactor to bind
substrate
Competitive Inhibition
Non-competitive Inhibition
6.1. Principles of Metabolism
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• Can separate metabolism
into two parts
CATABOLISM
ANABOLISM
Energy source
(glucose)
– Catabolism
Cell structures
(cell wall, membrane,
ribosomes, surface
structures)
• Processes that degrade
compounds to release
energy
• Cells capture to make ATP
Energy
Macromolecules
(proteins, nucleic acids,
polysaccharides, lipids)
Energy
Subunits
(amino acids,
nucleotides, sugars,
fatty acids)
– Anabolism
• Biosynthetic processes
• Assemble subunits of
macromolecules
• Use ATP to drive reactions
– Processes intimately linked
Energy
Precursor
metabolites
Waste products
Nutrients
(acids, carbon
dioxide)
(source of nitrogen,
sulfur, etc.)
Catabolic processes harvest
the energy released during the
breakdown of compounds and
use it to make ATP. The
processes also produce
precursor metabolites used in
biosynthesis.
Anabolic processes (biosynthesis)
synthesize and assemble subunits
of macromolecules that make up
the cell structures. The processes
use the ATP and precursor
metabolites produced in
catabolism.
Components of Metabolic
Pathways
• Role of the Chemical Energy Source and
the Terminal Electron Acceptor
• Some atoms, molecules more
electronegative than others
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Terminal
electron
acceptors
– Greater affinity for electrons
– Energy released when
electrons move from low
affinity molecule to high
affinity molecule
H2
• (E.g., glucose to O2)
Relative tendency to give up electrons
H2S
S0
Organic
carbon
compounds
Energy
release
Organic
carbon
compounds
CO2
SO4
FeOOH
Fe2+
NH4+
NO2– ( to form NH4+)
NO3– ( to form NH4+)
Mn2+
MnO2
Relative tendency to give up electrons
Energy
sources
NO3– ( to form NH2)
O2
(a) Energy is released when electrons are moved from an energy source with a
low affinity for electrons to a terminal electron acceptor with a higher affinity.
ATP is made in catabolic reactions and
used in anabolic reactions
•
6.3. The Central Metabolic
Pathways
Transition Step
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GLUCOSE
– CO2 is removed
from pyruvate
– Electrons reduce
NAD+ to
NADH + H+
– 2-carbon acetyl
group joined to
coenzyme A to form
acetyl-CoA
– Takes place in
mitochondria in
eukaryotes
2
Pentose phosphate
pathway
Starts the oxidation of glucose
Yields
Glycolysis
Oxidizes glucose to pyruvate
1
~
~
+
Reducing
power
ATP
by substrate-level
phosphorylation
Pyruvate
Yields
CO2
Reducing
power
Biosynthesis
Pyruvate
3a
Pyruvate
NAD+
Acids, alcohols, and gases
CoA
Transition step
CO2
Yields
Transition step:
CO2 is removed, a redox reaction generates
NADH, and coenzyme A is added.
Fermentation
Reduces pyruvate
or a derivative
5
CO2
Reducing
power
AcetylCoA
AcetylCoA
NADH + H+
x2
CO2
CoA
CO2
3b
TCA cycle
Incorporates an acetyl
group and releases CO2
(TCA cycles twice)
Acetyl-CoA
1 The acetyl group is transferred
to oxaloacetate to start a new
round of the cycle.
Yields
~
~
+
Reducing
power
ATP
by substrate-level
phosphorylation
CoA
Respiration
Uses the electron transport
chain to convert reducing
power to proton motive force
4
Yields
~
~
ATP
by oxidative
phosphorylation
NADH + H+
2 A chemical
rearrangement occurs.
Oxaloacetate
Citrate
A redox reaction
generates NADH.
8
NAD+
Isocitrate
NAD+
3
Malate
Water is added.
7
A redox reaction
generates NADH
and CO2 is
removed.
NADH + H+
H 2O
CO2
Fumarate
-ketoglutarate
NAD+
4
FADH2
6
CoA
A redox reaction
generates FADH2-
NADH + H+
FAD
5 The energy released
during CoA removal is
harvested to produce ATP.
CoA
Succinyl-CoA
Succinate
CoA
~ ~
ATP
~ + Pi
ADP
CO2
A redox reaction
generates NADH,
CO2 is removed,
and coenzyme A
is added.
The Electron Transport Chain—Generating Proton
Motive Force
• Calculating theoretical maximum yields
– In prokaryotes:
•
•
•
•
Glycolysis: 2 NADH 6 ATP
Transition step: 2 NADH  6 ATP
TCA Cycle: 6 NADH  18 ATP; 2 FADH2  4 ATP
Total maximum oxidative phosphorylation yield = 34
ATP
– Slightly less in eukaryotic cells
• NADH from glycolysis in cytoplasm transported
across mitochondrial membrane to enter electron
transport chain
– Requires ~1 ATP per NADH generated
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
GLUCOSE
2
Pentose phosphate
pathway
Starts the oxidation of glucose
Glycolysis
Oxidizes glucose to pyruvate
1
Yields
~
~
+
Reducing
power
ATP
by substrate-level
phosphorylation
Yields
Reducing
power
Biosynthesis
5
Acids, alcohols, and gases
Pyruvate
Pyruvate
3a
Fermentation
Reduces pyruvate
or a derivative
Transition step
CO2
CO2
Yields
Reducing
power
AcetylCoA
AcetylCoA
X2
CO2
CO2
3b
TCA cycle
Incorporates an acetyl
group and releases CO2
(TCA cycles twice)
Yields
~
ATP
by substrate-level
phosphorylation
~
+
Reducing
power
4
Respiration
Uses the electron transport
chain to convert reducing
power to proton motive force
Yields
~
ATP
by oxidative
phosphorylation
~
Fermentation
• The incomplete breakdown of glucose with an
organic compound serving as the final electron
acceptor
• Only pathway operating is glycolysis
6.5. Fermentation
• Fermentation end products varied; helpful in
identification, commercially useful
– Ethanol
– Butyric acid
– Propionic acid
• 2,3-Butanediol
• Mixed acids
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Pyruvate
Fermentation
pathway
Microorganisms
End products
Lactic acid
Ethanol
Butyric acid
Propionic acid
Mixed acids
2,3-Butanediol
Streptococcus
Lactobacillus
Saccharomyces
Clostridium
Propionibacterium
E. coli
Enterobacter
Lactic acid
Ethanol
CO2
Butyric acid
Butanol
Acetone
Isopropanol
CO2
H2
Propionic acid
Acetic acid
CO2
Acetic acid
Lactic acid
Succinic acid
Ethanol
CO2
H2
CO2
H2
(yogurt, dairy, pickle), b (wine, beer), (acetone): © Brian Moeskau/McGraw- Hill; (cheese): © Photodisc/McGraw-Hill; (Voges-Proskauer Test), (Methyl-Red Test): © The McGraw-Hill Companies, Inc./Auburn University Photographic Services
The Electron Transport Chain—Generating Proton
Motive Force
• Components of an Electron Transport Chain
– Most carriers grouped into large protein complexes
• Serve as proton pumps
• Three general groups are notable
– Quinones
• Lipid-soluble molecules
• Move freely, can transfer electrons between complexes
– Cytochromes
• Contain heme, molecule with iron atom at center
• Several types
– Flavoproteins
• Proteins to which a flavin is attached
• FAD, other flavins synthesized from riboflavin
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Fig. 6.20
Prokaryotic cell
Cytoplasmic
membrane
Electron Transport Chain
NADH dehydrogenase
Uses of Proton Motive Force
Ubiquinol
veoxidase
force
rive:
H+ (2 or 4)
H+ (0 or 4)
Ubiquinone
Path of
electrons
ATP synthase
(ATP synthesis)
10
Active transport
(one mechanism)
H+
Rotation of a flagella
H+
H+
Proton motive force
is used to drive:
Transported
molecule
Outside of
cytoplasmic
membrane
2 e– –
Cytoplasm
Succinate
dehydrogenase
NADH
+
NAD+
2 H+
1/
H2O
2
O2
Terminal
electron acceptor
H+
3 ATP
+ 3 Pi
3 ADP
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Glycolysis
Pentose phosphate
pathway
Glucose 6-phosphate
Fructose 6-phosphate
Lipopolysaccharide
(polysaccharide)
Ribose 5-phosphate
Erythrose 5-phosphate
Nucleotides
amino acids
(histidine)
Amino acids
(phenylalanine,
tryptophan,
tyrosine)
Peptidoglycan
Dihydroxyacetone
phosphate
Lipids
(glycerol
component)
3-phosphoglycerate
Amino acids
(cysteine,
glycine, serine)
Anabolic
Pathways—
Synthesizing
Subunits from
Precursor
Molecules
Phosphoenolpyruvate
Amino acids
(phenylalanine,
tryptophan, tyrosine)
Pyruvate
Pyruvate
Acetyl-CoA
Acetyl-CoA
Amino acids
(alanine,
leucine, valine)
Lipids
(fatty acids)
Oxaloacetate
Amino acids
(aspartate, asparagine,
isoleucine, lysine,
methionine, threonine)
X2
- ketoglutarate
TCA cycle
Amino acids
(arginine, glutamate,
glutamine, proline)
• Role of Electron Carriers
– Energy harvested in stepwise process
• Electrons transferred to electron carriers, which
represent reducing power (easily transfer electrons to
molecules)
– Raise energy level of recipient molecule
• NAD+/NADH, NADP+/NADPH, and FAD/FADH2
Microbial Growth
• Growth= an increase in the number of cells,
not an increase in size
• Generation=growth by binary fission
• Generation time=time it takes for a cell to
divide and the population to double
4.2. Prokaryotic Growth in Nature
• Microorganisms historically studied in laboratory
• But dynamic, complex conditions in nature have
effects on microbial growth, behavior
– Cells sense changes, adjust to surroundings
– Synthesize compounds useful for growth
– Can live singly
• Most live in polysaccharideencased communities
• Termed biofilms
• Cause slipperiness of rocks
in stream bed, slimy “gunk”
in sink drains, scum in toilet
bowls, dental plaque
Biofilms
• Formation of biofilm
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Planktonic bacteria
move to the surface
and adhere.
Bacteria multiply
and produce
extracellular polymeric
substances (EPS).
Other bacteria may
attach to the EPS
and grow.
Cells communicate and create
channels in the EPS that allow
nutrients and waste products
to pass.
Some cells detach
and then move to
other surfaces to
create additional
biofilms.
Bacteria divide
by binary fission
Calculating cell number
over time
t=time; 0=cell number at
start; n= number of
divisions based on
generation time
Nt=N0 x 2n
The Growth Curve
• Growth curve characterized by five stages
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Stationary
phase
Number of cells (logrithmic scale)
1010
Death
phase
108
106
Log or
exponential
phase
Phase of
prolonged decline
104
102
Lag
phase
100
Time (hr)
(days/months/years)
Primary and Secondary metabolites
Some factors that influence growth in
foods…temperature
• Remember that some microbes grow well at
cooler temperature, others more slowly
Some of the factors that influence growth in
foods… Water Availability (aw)
Food
(aw)
Microbe
Minumum
(aw)
Fresh meat
0.99
Spoilage
microbes
0.91
Hot dog
0.92
Pseudomonas
0.97
Ham
0.91
Staphylococcus
aureus
0.86
Dried fruit
0.72-0.8
Yeasts
0.81
Molds
0.80
Some factors that influence growth in
foods….pH
Foods
pH of food
Microbe
Minimum pH
of microbe
beef
5.5
Most spoilage
microbes
4.0
milk
6.3
molds
1.5
spinach
5.5
yeast
2.5
apples
3.0
E. coli
4.0
Microbes in food production
• Lactic Acid bacteria
• Yeasts…Saccharomyces cerevisiae
• Molds
Overview of Digestion
Fig. 24.1
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Oral cavity containing
tongue and teeth
Dental caries
Periodontal disease
Parotid salivary gland
Mumps
Stomach
Gastritis
Gastric ulcer
Gallbladder
Function
Oral cavity
Obtains and
processes food
Salivary
glands
Secrete saliva
Esophagus
Transports food to
stomach
Stomach
Stores food; mechanical
digestion; breaks down
some proteins
Pancreas
Secretes digestive
enzymes
Liver
Produces bile to assist
in fat digestion
Salivary glands
Esophagus
Esophagitis
Liver
Hepatitis
Organ
Pancreas
Pancreatitis
Small intestine
Enteritis
Duodenal ulcer
Appendix
Appendicitis
Large intestine
Dysentery
Colitis
Rectum
Anus
Food
molecules
Gallbladder Stores bile until
needed
Small
intestine
Site of most digestion
and absorption
of nutrients
Large
intestine
Absorbs some water
and minerals;
prepares waste
Villus
Epithelial cells
Microvilli
Capillaries
Lymphatic vessel
Smooth
muscle
Nerve fibers
Upper digestive tract
Lower digestive tract
How do organisms cause food
poisoning?
• Food borne intoxication: bacteria grow
within the food and produce toxins, the toxins
are what lead to food poisoning symptoms
• Examples: Clostridium botulinum
Staphylococcus aureus
Mechanisms of pathogenesis
• Attachment: pili or adhesins
• Toxin production: two kinds of toxins
1)increase secretion of water and
electrolytes
2)cause cell death
• Alterations in host cells
• Cell invasion
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Features of intestinal infections
1. Attachment—pili and adhesins
2. Toxin production
1. Enterotoxins cause release
of electrolytes
2. Cytotoxins cause cell
death
3. Alteration of epithelial cells
1. Attaching and effacing
2. Type III secretion systems
4. Cell invasion
Toxin production
Cl–
Na+
H2O
Cytotoxins cause
cell death. Toxins
absorbed into the
bloodstream result
in systemic effects.
Enterotoxins
increase secretion
of water and
electrolytes.
Alterations in the host cells
Pedestal
Inject effector
proteins
Attachment and effacing
(A/E) lesions formed after
bacterium injects various effector
proteins. One protein functions
as a receptor for the bacterium.
Another induces rearrangement
of actin filaments, resulting in the
formation of a pedestal under
the bacterium.
Cell invasion
Bacterium is engulfed and
multiplies within host cell.
An effector protein injected by
the bacterium induces the
engulfment by causing
rearrangement of host cell actin.
Type III Secretion System
Membrane ruffling
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Shigella enters
via M cells
A-B toxin on
chromosome. bad.
Causes HUS, hemolytic
uremic syndrome.
1
Epithelial
cell
M cell
Intestinal lumen
Macrophage
Shigella cells
2
Dead
macrophage
Bacteria make it into blood vessels, and
3
kill the endothelial cells.
A-B toxin:
B-binds cell,
A goes into cell—causes cell death.
4
M cells take up Shigella
cells and transport them
across the epithelium. They
multiply in the macrophages
that ingest them, leading to
death of that host cell.
Shigella cells attach to the
base of the epithelial cells
and induce those cells to
take them in. From there,
they escape the endosome
and multiply in the
cytoplasm.
Shigella cells cause the
host cell actin to polymerize.
This forms an “actin tail”
that propels a bacterium
within the host cell, sometimes with enough force
to move it into a
neighboring cell.
Neutrophils
Infected epithelial cells die
and slough off. An intense
inflammatory response
leads to bleeding and
abscess formation.
Courtesy of Philippe J. Sansonette, M.D., Professeur Institut Pasteur
Shiga-toxin E. coli (STEC)
• Obtain from the consumption of animal
products
• Attacks the colon, produce A/E lesions
• Produces Shiga toxins
• O157:H7 causes bloody diarrhea which may
lead to hemolytic uremic syndrome
Diarrhea causing E. coli
• Classified according to virulence
– Entertoxigenic E. coli (ETEC)
– Enterpathogenic E. coli (EPEC)
– Shiga toxin-producing E. coli (STEC)
– Enteroinvasive E. coli (EIEC)
– Enteroaggregative E. coli (EAEC)
– Diffusely adhering E. coli (DAEC)
Vibrio cholerae
• Causative agent of cholera
• General Characteristics: Curved gram
negative rod, facultative anaerobe, single polar
flagella, pili
• Can exist in saltwater for extended periods of
time, halotolerant
• Different serotypes based on O antigen, O1 is
current serotype circulating, O antigen is part
of the lipopolysaccharide.
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Cholera
Fig. 24.1
Vibrio cholerae
. a marine bacterium, halotolerant,
and tolerant to alkaline conditions.
Gr- curved rod.
Oral cavity containing
tongue and teeth
Dental caries
Periodontal disease
Parotid salivary gland
Mumps
NOT acid tolerant—high infective
dose
Stomach
Gastritis
Gastric ulcer
Gallbladder
Function
Oral cavity
Obtains and
processes food
Salivary
glands
Secrete saliva
Esophagus
Transports food to
stomach
Stomach
Stores food; mechanical
digestion; breaks down
some proteins
Pancreas
Secretes digestive
enzymes
Liver
Produces bile to assist
in fat digestion
Salivary glands
Esophagus
Esophagitis
Liver
Hepatitis
Organ
Pancreas
Pancreatitis
Small intestine
Enteritis
Duodenal ulcer
There have been 7 pandemics of
cholera
Appendix
Appendicitis
Large intestine
Dysentery
Colitis
Rectum
Anus
Food
molecules
Colonizes the small intestine, binds
to cells via pili and an A-B toxin
causes electrolytes to leave the cells.
Gallbladder Stores bile until
needed
Small
intestine
Site of most digestion
and absorption
of nutrients
Large
intestine
Absorbs some water
and minerals;
prepares waste
Villus
Epithelial cells
Microvilli
Capillaries
Lymphatic vessel
Smooth
muscle
Nerve fibers
Upper digestive tract
Lower digestive tract
A-B toxin
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
B binds to cell
A entersV. cholerae
bacterium
1
The A-B toxin’s B subunit attaches
to receptors on cell membrane;
the A subunit enters the cell.
B
2
The A subunit locks a G protein
in the “active” mode, turning on
adenylate cyclase.
4
A
OFF
10 µm
Plasma membrane
of intestinal cell
A
G protein
ON
cAMP activates ion transport
channels in the membrane
causing Cl– and other electrolytes
to pour out of the cell.
ATP
Cl–
3
Adenylate cyclase causes the
conversion of ATP to cAMP.
Adenylate
cyclase
K+
Na+
●
HCO3–
H2O
cAMP
5
Water follows electrolytes
out of the cell by osmosis.
© VeronikaBurmeister/Visuals Unlimited
Infections of the GI tract that are not
from food spoilage
• Helicobacter pylori can colonize the stomach.
• Gr-, curved rod with polar, sheathed flagella
• Many people have H. pylori with no symptoms
(1 in 5). However, some strains can cause
stomach cancer and 90% of stomach cancer
patients have H. pylori in their stomach.
How does H. pylori colonize the
stomach? It’s sterile, right?
• High Acid
– H. pylori survives acids by using urease to
transform urea into ammonia, creating an alkaline
pocket
• Thick Mucus layer protects stomach surface
cells (epithelium)
– H. pylori cells have sheathed flagella that allow
them to swim down into the thick mucus (less
acidic under the mucus).
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
•
Clostridium difficile
Fig. 24.1
Gr+, rod, spore forming, obligate Parotid
anaerobe.
salivary gland
Mumps
Oral cavity
Produces
containingcytotoxins
tongue and teeth
•DentalMember
of gut microbiome, in low
numbers,
caries
Salivary glands
Organ
Function
Oral cavity
Obtains and
processes food
Salivary
glands
Secrete saliva
Esophagus
Transports food to
stomach
Stomach
Stores food; mechanical
digestion; breaks down
some proteins
Pancreas
Secretes digestive
enzymes
Liver
Produces bile to assist
in fat digestion
Periodontal disease
--It most commonly occurs in patients in
Esophagus
hospitals on antibiotic therapy.
Esophagitis
•
Can also be acquired
Difficult
to kill with disinfectants (spores)
Liver
Hepatitis
Stomach
Gastritis
Gastric ulcer
Gallbladder
Pancreas
•
Mild to severe symptoms includingPancreatitis
colitis
Small(inflammation
intestine
of the colon).
Enteritis
Large intestine
ulcer
•Duodenal
Treatment:
Often stopping the antibiotics,
if
Dysentery
Appendix
possible, alleviates the problem. Colitis
Appendicitis
Rectum
Gallbladder Stores bile until
needed
Small
intestine
Site of most digestion
and absorption
of nutrients
Large
intestine
Absorbs some water
and minerals;
prepares waste
Anus
Food
molecules
Villus
Epithelial cells
Microvilli
Capillaries
Lymphatic vessel
Smooth
muscle
Nerve fibers
Upper digestive tract
Lower digestive tract
Control of Microbial Growth
A few terms
• Bacteriostatic: inhibits bacterial growth
• Bactericidal: something capable of killing bacteria
• Antiseptic: an agent that is used to inhibit/kill
bacterial growth on skin and mucus membranes
• Disinfectant: an agent that is used to inhibit/kill
bacterial growth on inanimate objects
What parts of a bacterial cell are
sensitive to physical treatments and
chemicals?
• Plasma membrane
• DNA and proteins
Are all microbes equally sensitive?
What level of microbial control or elimination is needed ?
Appropriate procedures depend on 4 main features:
1. Type and number of microbes (are there spores? What kind? Are there pathogens? )
Bacterial endospores
Mycobacterium, Pseudomonas sp.
Protozoan cysts and oocysts (Giardia lambia and Cryptosporidium parvum).
Naked viruses lack a lipid envelope, this makes them more resistant.
HIV—with lipid envelop—is very sensitive.
2. Environmental conditions
3. Risk of infection
4. Composition of the item to be treated.
Physical Methods
Moist Heat
Dry Heat
Filtration
Radiation
electromagnetic
ionizing
High Pressure
Chemical Control
Alcohol
Aldehydes
Biguanides
Ethyline Oxide Gas
Metals
Ozone
Peroxygens
Phenolics
Quaternarey
Ammonium
Compounds
Gene transfer in bacteria
• There are three types of gene transfer
1. Transformation—DNA enters and is
incorporated (cell imports it)
2. Conjugation—DNA moves from one bacterial
cell to another during cell to cell contact
3. Transduction—a virus injects DNA into the
bacterial cell
All types of gene transfer
• Involve unidirectional transfer of
information (donor-->recipient)
• Require the integration of newly acquired
DNA “homologous recombination”
• Increases genetic diversity
Terms to remember
• Replicon – DNA containing an origin of
replication (O.R.) This allows the DNA to be copied.
If no
O.R. then it must be incorporated into the chromosome for
replication.
• Homologous recombination – DNA has parts that
match the DNA in the chromosome, and bind to it to be
incorporated into the chromosome.
• Competent– A bacterial cell that can take up DNA from the
environment is termed ‘competent’
• Naked DNA—outside of the cell, or virus
Conjugation
• Transfer of genes
between 2 bacterial
cells
• Gram negative cells
use a sex pilus
• F(+) cells have F
plasmid, F(-) lack F
plasmid
Conjugation between (F+) and F() cells
First, the F pilus binds to specific receptor on
the F- cell.
Pilus retracts (gets shorter) and brings the
cells closer.
The F plasmid requires an Origin of
Transfer—we will refer to this as the OoT!!
Without the OoT, DNA can not be moved
using the F pilus.
The OoT is nicked to open the DNA and one
single strand of the plasmid moves through
the pilus to the other cell.
Then the complement of each of the ssDNA
plasmids is made, and voila! 2 F+ cells.
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Transformation
There are structures
bacterial cells use to bind
and import DNA.
Gene conferring StrS
1
Recipient chromosome
Gene
Conferring StrR
Double-stranded DNA binds to the surface of a competent cell.
2
Single strand enters the cell; the other strand is degraded.
These include the type 4
pili aparatus, which are
also used for making pili.
Some bacteria take in any
DNA, while others stick
to specific DNA
sequences and take in
only DNA they want.
3
The strand integrates into the recipient cell’s genome by
homologous recombination.
4
Streptomycin-sensitive
daughter cell
Streptomycin-resistant
daughter cell
After replicating the DNA, the cell divides.
5
Non-transformed cells (StrS) die on streptomycin-containing medium,
whereas transformed cells (StrR) can multiply.
Transduction
• Transfer of genes from a phage to bacterial cell
• Generalized transduction: occurs with lytic or
lysogenic phage (section 8.7)
• Specialized transduction: occurs with lysogenic
phage (section 13.3)
Plasmids-types
• Can be broad host range
• Or specific to particular species
• Some can be maintained within the same cell as
others (Arranged in groups by compatibility)
• Some can not, and are not compatible
• High copy number vs. low copy number
• Conjugative plasmids, carry genes needed for
conjugation
• Mobilizable plasmids have an OoT, but not the
conjugation genes. If conjugative and mobilizable
are together, they can both be moved to a new cell.
Resistance Plasmids (R plasmids)
Transposons…way to move
genes between organisms
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Insertion sequence
Mobile element
Transposase gene
Inverted repeat
5′
3′
Inverted repeat
3′
T C G A T G…
A G C T A C…
5′
5′
3′
…C A T C G A
....G T A G C T
3′
5′
Composite transposon
Mobile element
Insertion sequence
Antibiotic-resistance gene
Insertion sequence
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Vancomycinresistance gene
(encoded on a
transposon
on a plasmid)
• How did this S.
aureus become
Vancomycin
resistant S. aureus
(VRSA)?
Plasmid
Staphylococcus aureus
sensitive to vancomycin
Enterococcus faecalis
resistant to vancomycin
Enterococcus faecalis
plasmid transferred
by conjugation
Transposon
jumps from
one plasmid
to another.
Plasmid from
Enterococcus
faecalis is
destroyed.
Vancomycin-resistant
Staphylococcus aureus