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220 Cards in this Set

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Antony von Leeuwenhoek
1st person to observe & describe microorganism accurately (bacteria & protozoa)
made simple lenses (50-300x)
Robert Hooke
1st person to publish the depiction of a microorganism (fruiting structure of molds "Hairy Mould" colony)
Francesco Redi
proved biogenesis of large organisms
generation of maggots
Luis Pasteur
proved biogenesis of microorganisms through "swan-neck" flasks
Robert Koch & his postulates
1. suspected pathogenic organism should be present in all cases of the disease & absent from healthy animals
2. suspected organism should be grown in pure culture (only one)
3. inject pure culture-> healthy host experimental animals develop same signs & symptoms
4. organism should be re-isolated & shown to be same as original
Established relationship between Bacillus anthracis & anthrax
Developed pure culture methods (grown on solid media, isolation of pure culture)
Basic Properties of Cells
1. Metabolism
2. Reproduction (growth)
3. Differentiation
4. Communication
5. Movement
6. Evolution
Prokaryotes
single cells
lack nucleus
70S ribosomes
1-10 micrometer
5x10^3 cells on earth
Bacteria
simple shapes: round, rod, spiral
cell walls: peptidoglycan
Archaea
cell wall: more complex
found in extreme environments: high T, P, salt
Eukaryotes
single-celled or multicellular
have true nucleus
80S ribosome
10-100 micrometers
Protozoa
single-celled
no cell walls
motile
aquatic environments
part of food chain
some are pathogens
Algae
single-celled or multicellular
have cell walls
photosynthetic
in soils, lakes, oceans
some produce toxins
Fungi
single-celled or multicellular
have cell walls
no photosynthetic pigments
nature's "recyclers"
some are pathogens
Virus
acellular
made of nucleic acid & protein
obligate intracellular parasites
Carl Woese
used rRNA sequencing to compare organisms
led to discovery of Archaea
Monomorphic
single shape
Pleomorphic
variable shape
Hopanoid
embedded membrane protein
sterol-like
stabilize membrane
Mesosome
invaginations of plasma membrane
-cell wall formation in cell division
-chromosome replication & distribution
-secretory processes
artifacts of chemical fixation process
Internal Membrane System
mesosomes
complex infoldings in photosynthetic bacteria & prokaryotes with high respiratory activity
-aggregates of spherical vesicles, flattened vesicles, tubular membranes
Inclusion Bodies
Made of organic or inorganic
Nutrient & energy storage
Poly-B-hydroxybutyrate
Inclusion body for phosphate storage (Biodegradable plastics)
Magnetosome
Inclusion body, iron containing orientation in magnetic field
Nucleoid
irregularly shaped region
single circle of double-stranded DNA
Plasmid
extra-chromosomal DNA
Small, circular, mini-chromosomes
Extra genetic info not required for cell growth & may provide selective advantage
Transfer between bacteria via conjugation
Gram Staining
1. smear Crystal Violet for 1 min (all purple)
2. iodine solution for 3 min (all purple)
3. decolorize with alcohol ~20 sec (G+ purple, G- colorless)
4. counterstain with safranin 1-2 min (G+ purple, G- pink)
Capsule
thick, structured difficult to wash
made of polysaccharide &/or protein
adherence, resistance to desiccation, resistance to phagocytosis, improve motility
Slime Layer
thin, slimy, easy to wash off
made of polysaccharide &/or protein
adherence, resistance to desiccation, resistance to phagocytosis, improve motility
Fimbrae
only in G-
used for attachment (adhesion protein)-> invasion nutrient uptake
not for motility
short, hair-like structures
Pili
type of fimbrae for DNA transfer
Pattern of Bacterial Movement
"Runs" CCW rotation of flagella filament
"Stop/Tumbles" CW
Endospores
"resting/dormant state" structure within bacterial cell
DNA & proteins
survival of adverse conditions
can "reactivate" in favorable environment
i.e. Bacillus, Clostridium
types: central, terminal, subterminal
Sporulation
spore formed under stress
Stages in Spore Transformation
Reactivation
-prepare spores for germination
Germination (Spore into vegetative)
-spore swelling
-rupture of absorption of spore coat
-loss of resistance
-increased metabolic activity
Outgrowth
-emergence of vegetative cell
Requirements for Microbial Growth
Temperature
pH
Osmotic pressure
Oxygen concentration
Macroelements
Trace elements
Organic growth factors
Low Temperature Growth Microbes
Membranes with high levels of unsaturated fatty acids; semifluid
Psychrophiles
-growth at 0-20C (<15)
-polar habitats
Psychotrophs (facultative)
-growth at 0-7C (20-30C)
-food spoilage
Stalk
Caulobacter
Plasma membrane
Nutrient absorption & surface attachment
Hypha
Streptomyces
Plasma membrane, ribosomes, DNA
Increase metabolism
High Temperature Growth Microbes
Protein Structure - more H bonds, prolines & chaperones
DNA - histone-like proteins
Membrane - more saturated, branched, high MW lipids & ether linkages
Thermophiles
-prefer 55-65C or higher
-hot water lines, compost piles
Hyperthermophiles
-prefer 80-100C
-geothermal areas of ocean floor
Streptomyces thermoautotrophicus
Moderate Temperature Growth Microbes
Mesophiles
-prefer 20-45C
-human pathogens
pH preference Microbes
Neutrophile (5.5-8)
Acidophile (<5.5)
Alkalophile (>8.5)
internal pH near neutrality through proton/ion exchange
acid-shock proteins (chaperones)
change pH by: -acidic/basic waste products
most media have buffers to prevent growth inhibition
Osmotic Pressure Preference MIcrobes
Isotonic
Hypotonic (lower solute outside cell) = osmotic lysis
Hypertonic (higher solute out) = plasmolysis
Osmotolerant (high solute out)
i.e. Staphylococcus aureus (3M salt on skin)
Extreme halophiles (intra K/Na high) in marine environments like Great Salt Lake, Utah
Oxygen Requirement Microbes
Obligate aerobe (requires O2)
Facultative anaerobe (prefers O2)
Microaerophile (requires O2 in reduced concentrations 2-10%)
Aerotolerant anaerobe (doesn't require but unharmed by O2)
Obligate anaerobe (can't use/detoxify O2 & killed by it)
Macroelements
Carbon (heterotroph/autotroph)
Oxygen, Hydrogen (carbs, lipids, proteins, nucleic acids, organic compounds, H2O)
Nitrogen (AAs, nucleotides, carbs, lipids from protein, ammonium, nitrate, nitrite; N2 fixation from atmosphere)
Sulfur
Phosphorus
Organic Growth Factors
Amino Acids (protein synthesis)
Purines & Pyrimidines (nucleic acid synthesis)
Vitamins (enzyme cofactors)
-cell can't synthesize this because of lack of synthetic enzymes, so supplied by environment
Passive Diffusion
doesn't require energy
dependent on size of gradient
H2O, O2, CO2, small uncharged molecules move
large molecules, ions, polar substances aren't able to move
Facilitated Diffusion
not energy dependent
from high to low concentration
size of concentration gradient impacts rate of uptake
facilitated by permeases
transports glycerol, sugars, & AAs
more in eukaryotic cells
conformational change of carrier after binding an external molecule-> release of molecule on cell interior
Binary fission
process of cell division in bacteria
Measurement of Bacterial Growth
Direct Counts with Calibrated Slide Chamber
-easy, inexpensive, quick
-eukaryotes & prokaryotes
-can't distinguish living from dead
Plate Count Methods
-Spread Plate
-Membrane Filtration
Indirect/Turbidity
-more cells in test tube-> more cell mass-> more light scattered-> less light detected
Total Cell Count
turbidity/microscope count
doesn't distinguish living from dead
Viable Cell Count
plate count or membrane filtration
only live cells grow
Culture Media
Chemically defined media (precise composition known)
Complex media (unknown)
Contains:
Peptones (protein hydrolysates prepared by partial digestion of protein)
Extracts (aqueous, beef/yeast)
Agar (Sulfated polysaccharide used to solidify liquid media)
Selective Media
inhibit certain microbes
"Brilliant green agar"
G+ bacteria inhibits
Differential Media
"Blood agar" hemolysis
Selective & Differential Media
"MacConkey agar"
inhibits G+ & detects lactose fermentation
Enrichment Media
isolation of specific organisms
nitrogen fixing bacteria isolate in media without nitrogen (Siderophore in Azobacter Vinelandii)
Sterilization
all life forms (bacterial endospores & viruses) are either destroyed/removed from object/habitat
chemicals, heat, radiation
Disinfection
killing, inhibition, removal of microorganisms that may cause disease on inanimate objects
-may not kill endospores
ethanol, cationic detergents
Sanitization
microbes reduced to levels "considered safe by public health standards"
Antisepsis
reduction of total microbial population on living tissue (prevention)
Germicide
kills microbes
-not endospores
Bacteriostasis
stops bacterial growth
must be continually present
D value/ Decimal Reduction Time
time required to kill 90% at specific temperature
Factors affecting Antimicrobials
1. Population size
2. Population Composition
-tough to kill bacterial endospores & Mycobacterium tuberculosis but easy to kill "vegetative" growing cells
3. Duration of exposure
4. Concentration
5. Temperature (higher, enhances antimicrobial activity)
6. Local environment
-pH, viscosity, organic matter (inhibits chemical agents)
-cause leakage on membrane-> proteins denature/inhibit-> nucleic acids damage/break
Methods of Microbial Growth Control
Physical control methods
-Heat
-Low Temperature
-Radiation
-Filtration
Chemical Control Methods
-Alcohols
-Aldehydes
-Detergents
Heat as Method of Microbial Growth Control
(+) widely-used methods, inexpensive & easy to apply
(-) may not sterilize, some damaged by heat
Dry Heat
-Hot oven (170C for 2 hours) for glassware, glass pipettes
-Direct flaming for bunsen (loops) & incineration of trash & contaminents
Moist Heat
-Boiling (10 min-vegetative) sanitize but don't kill spores
-Autoclave (saturated steam under pressure) for endospores, media, and surgical instruments
-Pasteurization for brief 60C like wine
Thermal Death Time
minimum time needed to kill organisms in suspension of specific temperature
Z value
increase in temperature required to reduce D to 1/10
Filtration
heat-sensitive agents
antibiotics, vitamins, AAs
filter air using surgical masks, cotton plugs on culture vessels, high-efficiency particulate air filters (HEPA) for laminar flow biological safety cabinets
Low Temperature for Microbial Growth Control
Refrigeration (4C) or Freezing (-20C)
stops production due to lack of liquid water
some killed by ice crystal disruption of cell membranes
bacteriostatic
Radiation for Microbial Growth Control
Non-iodizing radiation
-UV light wavelength 260nm
-"germicidal lamps" damages DNA & kills germs
-surface sterilization
(-)poor penetration of glass, water
Ionizing radiation
-X-rays, gamma rays
-kill spores
-for lab & medical supplies
-generates electrons, OH, H but disrupts DNA & protein
i.e. Clostridium botulinum & Deinococcus radiodurnas (500x resistant)
Desiccation
drying-bacteriostatic
freeze-dried foods
Osmotic Pressure
High concentration of sugars/salts
Food preservation like jams & salted meats
Ideal Chemical Agent
active against wide range of microbes
non-corrosive & nontoxic
soluble in water, long shelf-life
non-staining, pleasant odor
cost-effective
Phenol & Derivatives
denature protein & disrupt membrane
Alcohol
Ethanol, Isopropanol
denatures proteins & dissolves lipids
doesn't kill spores
Halogens
Iodine
antiseptic/oxidize cell constituents & iodinates proteins
Heavy Metals
Cu
algicide in water but toxic
Detergents
Quaternary ammonium compounds
cleaning & sanitization in hospitals & labs
doesn't kill G-
Aldehydes
Formaldehyde
sterilize surgical instruments
combine with & inactivate nucleic acid & proteins but allergical reaction
Sterilizing Gases
Ethylene oxide
sterilization of heat-sensitive medical equipment
kills bacteria, mold & fungi
Evaluation of Chemical Control Methods
use-dilution tests: rate at which selected bacteria are destroyed 95%
i.e. Staphylococcus, Salmonella, Pseudomonas
Phenol Coefficient: dilutions of test agent compared to dilution of phenol
Mn
SOD, oxygen-evolving complex of photosystem II
Zn
SOD, alcohol dehydrogenase
Cu
SOD, cytochrome oxidase
Co
Vitamin B12, nitrile hydratase
Mo
Nitrogenase, nitrate reductase
Ni
CO Dehydrogenase, Hydrogenase
Major Nutritional Types of Microorganisms
1. Carbon (Heterotroph/Autotroph-CO2)
2. Energy (Chemotroph: organic-glucose, inorganic-H2S; Phototroph)
3. Electron (Organotroph-glucose; Lithotroph-Fe2+, H2)
Chemoorganotrophic heterotrophy
most bacteria, fungi, protozoa, most pathogenic microbes
Photolithotrophic Autotrophy
Cyanobacteria, Algae
capable of "oxygenic photosynthesis"
Chemolithotrophic Autotrophy
nitrifying bacteria, iron-oxidizing bacteria, sulfur-oxidizing bacteria
Photoorganotrophic Heterotrophy
purple & green nonsulfur bacteria
capable of "anoxygenic photosynthesis"
in lakes, streams, soil
Glucose Catabolism Pathways & Endproducts
Glycolysis//Pyruvate
Pentose phosphate//3-7Carbon sugars
Entner-Doudoroff//Pyruvate
Substrate-level Phosphorylation
synthesis of ATP from ADP by phosphorylation coupled with exergonic breakdown of a high energy organic substrate molecule
Summary of Glycolysis
Glucose + 2ADP + 2Pi + 2NAD -> 2 pyruvate + 2ATP + 2NADH + 2H
Fermentation
In absence of aerobic/anaerobic respiration
Regenerates NAD, oxidizes NADH to NAD
if NAD not regenerated, glycolysis stops
uses organic molecules as electron acceptor (endogenous, usually pyruvate or derivative)
forms alcohols/organic acids
homolactic fermenters- yogurt, sauerkraut, pickles
alcoholic fermentation- alcoholic beverages, bread
ATP yield from Aerobic oxidation of glucose by eukaryotic cells
Glycolytic Pathway: substrate-level phosphorylation (2 ATP)
oxidative phosphorylation with 2NADH (6 ATP)
2 Pyruvate to 2 Acetyl-coA: oxidative phosphorylation with 2 NADH (6 ATP)
TAC: substrate-level phosphorylation (GTP) (2 ATP)
oxidative phosphorylation with 6 NADH (18 ATP)
oxidative phosphorylation with 2 FADH2 (4 ATP)
TOTAL: 38 ATP
Pasteur Effect
facultative microbes have both fermentation & aerobic respiration
with O2, consumes less glucose
without O2, consumes more glucose
Biosynthesis
Turnover-> continual degradation & re-synthesis of cellular constituents
1. Macromolecules synthesized from limited number of monomers->saves genetic storage capacity, biosynthetic raw material & Energy
2. Many enzymes used for both catabolism & anabolism-> saves material & Energy
3. Catabolic & anabolic pathways not identical despite shared enzymes-> permits independent regulation
Synthesis of Sugars & Polysaccharides by Heterotrophs
Gluconeogenesis- synthesize glucose & fructose from noncarbohydrate precursors
most other sugars made from them
functional reversal of glycolysis
7 enzymes shared
4 enzymes unique to gluconeogenesis
Level of Organization of Molecules to Cells
Inorganic molecules (CO2, NH3, H20, PO4)
Monomers (Nucleotides, AAs, sugars, fatty acids)
Macromolecules (Nucleic acids, proteins, polysaccharides, lipids)
Supramolecular systems (membranes, enzyme complexes)
Organelles (Nuclei, mitochondria, ribosomes, flagella)
Cells (Bacteria, algae, fungi, protozoa)
Catabolism of Chemoorganotrophic Heterotrophy
1. Larger nutrients broken down
Polymers-> Monomers
Proteins-> AAs
Carbs-> Simple Sugars
Lipids-> Fatty acids & glycerol
(no energy release)
2. Monomer units-> acetyl coA
synthesis of ATP, NADH &/or FADH2
3. complete oxidation of molecules to CO2, ATP, NADH & FADH2
TCA cycle
electron transport/oxidative phosphorylation
Conversion of pyruvate to acetylaldehyde
Pyruvate Decarboxylase
Pyruvate-> Acetylaldehyde + CO2
ethanol producing yeast & bacteria
Lactic Acid Fermentation
Lactate Dehydrogenase
Pyruvate-> Lactate (NADH->NAD)
homolactic fermentation
Lactic acid bacteria
yogurt & in tooth decay
Formic acid fermentation
useful in identification in members of Enterobacteria
1. Mixed acid fermentation
forms formic acid (CO2 + H2), ethanol, acetic acid, lactic acid, suscinic acid (E.coli)
2. 2,3 Butanediol fermentation
forms 2,3 butanediol, ethanol (Enterobacter)
Methyl Red Test
Voges-Proskauer Test
use fermentation endproducts to identify bacteria
measures acidity of fermentation endproducts
mixed acid-> acidic
2,3 butanediol-> neutral
positive methyl red-> acid-> red
negative methyl red-> neutral-> yellow
Strickland Reaction
1AA oxidized
2nd AA= electron acceptor
i.e. Clostridium (anaerobe prefer high-protein environment)
oxidizes alanine; produces NADH + ATP
reduces glycine; regenerates NAD
Tricarboxylic Acid Cycle (Citric Acid Cycle, Kreb's Cycle)
completes oxidation & degradation of glucose & others into CO2
produces ATP via substrate phosphorylation
produces reduced coenzymes (NADH, FADH)
common in aerobic bacteria, free-living protozoa, most algae & fungi
amphibolic provides carbon skeletons for biosynthesis
catabolic: acetyl coA from lipids & proteins
anabolic: TCA intermediates for synthesis
3 stages based on size of intermediates (6, 5, 4) which are separated by 2 decarboxylation reactions (2 CO2)
3 NADH & 1 GTP produced by substrate-level phosphorylation
1 FADH2 generated by 1 acetyl-coA & substrate-level phosphorylation
Electron Transport & Oxidative Phosphorylation
only 4 ATP synthesized directly from oxidation of glucose to CO2
most ATP made when NADH & FADH2 oxidized in electron transport chain
-electron carriers operate together to transfer electrons from NADH & FADH2 to terminal electron acceptor
-electron flow from carriers with more negative E0 to positive
-in mitochondria (euk) plasma membrane (pro)
Electron Transport Chain in Eukaryotes
4 complexes of carriers (transport electron to O2)
coenzyme Q & cytochrome C connect complexes
pro- branched, different cytochromes, utilize electron acceptors (nitrate)
ATP synthesized by protons pumped across membrane
-proton motive force (gradient of protons & membrane potential due to unequal distribution of charge)
-ATP synthase (protons into mitochondrial matrix)
released energy to make ATP by oxidative phosphorylation (3 ATP per NADH, 2ATP per FADH2)
Chemolithotrophs
Hydrogen oxidizing bacteria (H2->2H+2e; Ni/Fe Hydrogenase)
-alcaligenes, Streptomyces thermoautotrophicus
-roistonia eutropha: 2 Hydrogenases
Nitrifying bacteria: oxidize ammonia to nitrite/nitrate (nitrification)
-NH4+1 1/2 O2-> NO2+H2O+2H nitrosomonas
-NO2+1/2 O2-> NO3 nitrobacter
-nitrite oxidoreductase
-Ammonia-oxidizing bacteria: Hydrosylamine oxidoreductase, ammonia, monoxygenase
CO oxidizing bacteria (CO->CO2)
-Aerobic (Mo in active site of CO-Dehydrogenase, MOlybdopterin) i.e. Streptomyces thermoautotrophicus, Oligotropha carboxidovorans
-Anaerobic (Ni in active site of CO-Dehydrogenase) i.e. Carboxydothermus hydrogenoformans
Iron Oxidation by Acidithiobacillus ferroxidans
Carbon Assimilation (Carbon fixation)
CO2 incorporated via Calvin-Benson cycle
energy source- light (photoautotroph)/inorganic chemicals (chemolithotroph)
product: organic carbon needed by heterotrophs
Nitrogen Fixation
enzyme: nitrogenase
very high Energy requirement
N2 + 8H + 8e + 16 ATP -> 2NH3 + H2 + 16ADP + 16Pi
widespread among microorganisms
-Rhizobium - symbiont with legumes
-many free-living bacteria (Azotobacter)
-Cyanobacteria
nitrogenase reaction is anaerobic inhibited by O2
Anaplerotic Reactions
replenish TCA cycle intermediates -> allow TCA to function during periods of active biosynthesis
1. Anaplerotic CO2 fixation
2. Glyoxylate Cycle
Peptidogylcan Synthesis
1. sugars synthesized in cytoplasm linked to carrier uridine diphosphate
2. UDP-NAM (N-acetyl muramic acid) linked to AA (no ribosomes/tRNA required, ATP needed)
3. NAM-peptide from UDP to bactoprenol phosphate
4. NAG (N-acetyl glucosamine) from UDP to NAM-peptide-> forms peptidoglycan repeat unit
5. peptidoglycan repeat unit transported across membrane by bactoprenol
6. peptidoglycan unit added onto growing peptidoglycan chain
7. bactoprenol carrier returns to inside of membrane
8. transpeptidation-crosslinks formed in peptidoglycan
Inhibited by some antibiotics->weak cell wall, bacteria lysis
penicillin inhibits transpeptidation
Measurement of Virulence
ID50 - infectious dose for 50% of population
LD50 - lethal dose for 50%
Pathogenesis of Bacterial Disease
process of disease causation
1. Attachment of microbe
2. Invasion through host
3. Evasion of host defenses
4. Mechanisms of damage to host tissues
Contact & Entry into Host
source of organism: disease reservoirs
modes of transmission: contact, vehicles (food/H2O), vectors
portals of entry: respiratory, gastrointestinal, urogenital, skin
attachment- adherence structure (fimbrae, capsules, adhesins)
Invasion & Growth in Host
passive invasion (cuts, scratches, insect, phagocyte engulfs)
active invasion (production of lytic enzymes, attack on cell surface)
growth in or on host cells (host may supply heme groups/ATP)
Evasion of Host Defenses
survival of bacteria
evasion of complement system: G-bacteria-lengthen "O" chains on LPS
resistance to phagocytosis: capsules (Streptococcus pneumoniae, Neisseria Haemophilus)
Damage to Host Tissues
Direct cell damage via host cell invasion & lysis
Toxin-mediated damage
-endotoxin (G-) produced as bacteria divide or die
-exotoxin (G+ & some G-) produced by living bacteria
Exotoxin
source: G+ & some G- living bacteria
chemistry: protein
effect on host: specific
heat stability: unstable at 60-80C
toxicity: highly in small amounts
immunology: can vaccinate
fever: usually none
genetics: reside on plasmid
disease: tetanus, botulism, diphtheria
Endotoxin
source: G- bacteria dying
chemistry: Lipid A portion of LPS
effect on host: fever, weakness, inflammation
heat stability: heat-stable
toxicity: moderate (high lethal)
immunology: no vaccines
fever: induces via IL-1 release
genetics: encoded by chromosomal genes
disease: Meningocococal meningitis, typhoid fever, G-
Neurotoxins
Block or activate specific neurons
May be ingested pre-formed
-Clostridium botulinum (Botox blocks nt release-> "flaccid paralysis")
May be formed in body
-Clostridium tetani
Membrane-Disrupting Toxins
Phospholipases (exotoxin)
-Clostridium perfringins alpha toxin (unstable host cell membrane)
Endotoxin in Bacterial Diseases
indirect effects on host
LPS enters monocytes & macrophages
Host cells release cytokines-> fever
Vaccines
Active immunization
-vaccine administered & induces protective immunity
Passive immunization
-artificially acquired
-can be produced by injecting preformed antibodies
Whole-Organism Vaccines
-inactivated/attenuated
Purified macromolecule vaccines
-capsules, toxoids, surface antigens
Recombinant vector vaccines
-attenuated recombinant nonpathogenic microorganism
-genes encode antigen from pathogen
DNA vaccines (not approved yet)
Therapeutic index
ratio toxic dose to therapeutic dose
higher ratio, more effective agent
Categories of Effectiveness
Narrow spectrum drugs
-affects one or a few types of microbes
-Penicillin G kills G+
Broad Spectrum drugs
-affects many types of microbes
-Tetracycline kills G+, G-, intracellular bacteria
Categories of Antimicrobial Agents
"Natural" synthesized by microbes
-Fungi-Penicillum; Cephalosporium
Semi-synthetic- isolated from microbes, modified in lab
Synthetic- directly synthesized in lab
Mode of Action of Antimicrobial Agents
cell wall synthesis
protein synthesis
nucleic acid synthesis
specific enzymes
Cell Wall Synthesis Inhibitors
Penicillin
-B-lactam antibiotic inhibits transpeptidation step of peptidoglycan synthesis
-bacteriocidal
Penicillin G ("natural" penicillin)
-effective against G+, some G-
-must be injected, destroyed by stomach acid
Resistance to Penicillin
some bacteria produce penicillinase (B-lactamase)
penicillin converted to penicilloic acid
Protein Synthesis Inhibition
Tetracycline binds 30S subunit, inhibits aminoacyl tRNA binding
-broad spectrum affects both G+ & G- & intracellular bacteria
-bacteriostatic
Aminoglycosides: Streptomycin
-binds 30S subunit; cause misreading of mRNA
-bactericidal
-toxic side effects: deafness & allergic response
Macrolide : Erythromycin
-bind 50S subunit
-broad spectrum
-bacteriostatic
Inhibition of Nucleic Acid Synthesis
Quinolines: Nalidixic acid
Fluoroquinolones: Ciprofloxacin
-inhibit DNA gyrase; inhibits DNA replication
-broad-spectrum
Inhibition of Specific Enzymes
Sulfonamide inhibits folic acid synthesis; inhibits purine & pyrimidine synthesis
-competes with -aminobenzoic acid during folic acid synthesis (decline of folic acid in cell, inhibits DNA/RNA synthesis)
Botulism
causative agent: Clostridium botulinum
mode of transmission: ingestion of toxin in food
signs/symptoms: flaccid paralysis, death due to respiratory/cardiac failure
virulence factors: neurotoxin-blocks nerve transmission
treatment: antisera
prevention: improved food preservation foods
Staphylococcal "Food Poisoning"
causative agent: Staphylococcus aureus
source: person preparing food
foods: ham, cream pies, puddings, any foods cooked & left at R.T.
incubation hour: 1-8 hours
signs & symptoms: nausea, vomit, abdominal cramps, diarrhea
treatment: oral or IV rehydration, recovery in 24-48 hours
prevention: handwashing by food preparer & refrigeration of foods
Helicobacter Peptic Ulcer Disease
causative agent: Helicobacter pylori
signs & symptoms: erosion & ulceration of stomach lining
treatment: antibiotics & pepto-bismol
-ammonia that's produced is toxic to epithelial cells of stomach & protease, catalase, phospholipase
-damages epithelial cells
Diphtheria
causative agent: Corynebacterium diphtheriae
mode of transmission: airborne & respiratory droplets
signs & symptoms: cold-like fever & sore throat; Pseudomembrane in throat
virulence factors: exotoxin (encoded by lysogenized phage); toxin inactivates EF-2, blocks protein synthesis
treatment: antibiotics & anti-toxin antisera
prevention: DPT vaccine
Bacterial Pneumonia
predisposing factors:
-prior viral infection-cold/influenza
-exposure to pollution
-smoking
-poor nutrition, alcoholism, drug use
-other health problems- heart & lungs
Streptococcal Pneumonia
causative agents: Streptococcus pneumoniae, G+ coccus; forms capsule
signs & symptoms: fever, chest pain & cough
virulence: capsule pneumolysin (damages blood vessels in lungs)
treatment: antibiotics
vaccine: Streptococcal pneumoniae vaccine; penetration into lower respiratory tract-> Streptococci trapped by mucus & removed by ciliary action -> macrophages phagocytosis-> ciliated epithelium damaged by viruses, toxins, smoke, chemicals-> fluid accumulates-> lowered activity of macrophages-> growth on damaged ciliated epithelium-> growth in fluids & alveoli-> fluid accumulation
Anthrax/Woolsorters' Disease
causative agent: Bacillus anthracis
mode of transmission: cuts in skin (anthrax); inhalation (Woolsorter's disease)
signs/symptoms: cutaneous anthrax-skin pustule; pulmonary anthrax-Woolsorter's disease-Pneumonia (100% lethal)
treatment: antibiotics (if early)
prevention: vaccine
Tetanus
causative agent: Clostridium tetani
mode of transmission: puncture wound
signs/symptoms: muscle spasms, "lock-jaw", death from suffocation
virulence factors: neurotoxin: tetanospasmin; blocks inhibitory neurons->overactivity muscles
treatment: passive immunization with human anti-tetanospasmin immunoglobulin; vaccine booster
prevention: DPT vaccine
Booster dose
reexposure to immunizing antigen
higher immunity against antigen back to protective levels after it decreases or after specified time
Bubonic Plague
causative agent: Yersina pestis
mode: vector: rat flea
reservoir: squirrels, rats, prairie dogs
signs & symptoms:swollen & blackened lymph nodes (buboes), fever
Taxis
arrangement/order
homos
Law
nemein
to distribute/govern
classification
arrangement of organisms into groups/taxa based on similarities/evolutionary relatedness
nomenclature
assignment of names to taxonomic groups in agreement with published rules
How old is the planet?
4.6 billion years
When did the first prokaryotic cells occur?
3.5-3.8 billion years ago
found in stromatolites & sedimentary rocks
stromatolites
layered rocks formed by incorporation of mineral sediments into microbial mats (anaerobic)
When did oxygen production being and who did it?
Cyanobacteria (oxygenic phototrophs) 2.5-3 billion years ago
When did eukaryotes arise?
from prokaryotes 1.4 billion years ago
Evolution of eukaryotes
1. nuclei, mitochondria & chloroplasts arose by invagination of plasma membranes to form double membrane structures containing genetic material & capable of further development & specialization
2. Endosymbiotic hypothesis
-fusion of ancient bacteria & archaea
-chloroplasts arose from free-living phototrophic bacterium that entered symbiotic relationships with primitive eukaryotes
-mitochondria arose similarly
Taxonomic Ranks
Domain
Phylum
Class
Order
Family
Genus
Species
Higher organisms
groups of interbreeding natural populations reproductively isolated from other groups
Prokaryotes/species
asexual collection of strains sharing stable properties
Phenotypically distinguishable
Genetically similar
-similar GC composition
-70% or > DNA sequence similarity
-same sequence of core housekeeping genes (genes coding for products that are required by all cells & expressed continually)
Population dropped from single organism/pure culture isolate
Population distinguishable from at least some other population within particular taxonomic category
Strains within species might differ slightly
Natural Classification Systems
according to shared characteristics
most desired since reflects biological nature of organisms
1st used by Linnaeus
1. Phenetic Classification
-reveal evolutionary relationships but not dependent on phylogenetic analysis
2. Numerical Taxonomy (multistep process)
-phenetic classification systems
-code info about properties or organisms (1=has trait, 0=don't)>50 characters
-construct similarity matrix
-construct dendograms
3. Phylogenetic Classification System
-evolutionary relationships
-"molecular chronometers"-use of protein/nucleic acid sequence as measure of change over time (16S ribosomal RNA, proteins: histones, cytochrome C)
Biovars
biochemical physiological strain variant
Morphovar
morphological variant
Serovar
antigenic variant
Type Str
usually one of first strains of species studied
often most fully characterized
not necessarily most representative member of species
Binomial system of nomenclature
Carl von Linne (Carolus Linnaeus)
-genus name-italicized & capitalized
-species name-italicized but not cap
-can be abbreviated after first use
"Classical Characteristics"
Morphological: cell shape, size, Gram stain
Metabolic: O2 required, fermentation end products
Ecological: T required, habitat
Genetic: chromosome exchange via conjugation/transduction
"Molecular Characteristics"
Protein comparison: AA sequence
Nucleic acid base composition: GC content
Nucleic acid hybridization
Nucleic acid sequencing
Comparison of proteins
AA sequence of proteins with same function
-electrophoretic mobility
-determination of immunological cross-reactivity
-enzymatic properties
Nucleic acid hybridization
measure of sequence homology
bind nonradioactive DNA to nitrocellulose filter-> incubate filter with radioactive ssDNA-> measure amount of radioactive DNA attached to filter-> retained radioactivity can be used to determine homology
Usually comparison of rRNA genes
-critical organelle in all microbes
-perform same functional role in all ribosomes (structural elements)
-structure changes slowly with time (provide stable sequence for comparison)
Increasingly, comparison of entire genomes
Microbial Chronometers
Nucleic acids/proteins used as "docks" to measure amount of evolutionary change over time
Assumptions:
-sequences gradually change over time
-changes are selectively neutral & relatively random
-amount of change increases linearly with time
Problems:
-rate of sequence change can vary over time
-different molecules & different parts of molecules can change at different rates
Creating phylogenetic trees from molecular data
1. align sequences
2. determine number of positions that are different
3. express difference-> evolutionary distance
4. use measure of difference to create tree
Microbial Genetics
info about evolutionary & horizontal gene transfer
Future: improved diagnostic methods, new vaccines/disease treatments, better understanding of microbial diversity, discovery of novel enzymes for industrial uses
Escherichia coli genomic map & genome
genome size: 4.6 x 10^6 bp
gene number: 4288
58% no known function
10% energy & metabolic pathways
12-14% transport protein genes
8% replication, transcription, translation
Single chromosome
Length: 1.4 mm
Semiconservative DNA replication
each synthesized copy contains one new & one old strand
Bidirectional DNA replication
two replication forks move around the DNA forming intermediates
Mutations
change of single bases, additions or deletions
Recombination
whole chromosomes/regions of chromosomes transferred between cells
Point Mutation
due to mistakes by DNA polymerase which occurs rarely; corrected by "proofreading"
due to alteration of bases after DNA synthesis caused by chemicals or radiation
Silent Mutation
no effect
Direct Selection of Mutant
treat cells with mutagen-> add agent to kill "wildtype"-> only mutants will grow (penicillin-resistance)
Indirect: Replica Plating
expose cells to mutagen-> compare growth on complete media (lacking essential components) (lysine)-> rapid screening method
Ames Test
uses mutant Salmonella strain (his auxotroph unable to synthesize histidine)-> mix Salmonella his- with test chemical-> plate on histidine-deficient media-> only revertant his- cells will grow
mutagenesis-> carcinogenesis
Missense mutation
changes AA
Neutral mutation
change to equivalent AA
nonsense mutation
creates stop codon
Insertion/Deletion
caused by intercalating agents (insert themselves between stacked bases of helix, ethidium bromide)
causes distortion of DNA
causes "frame shift" mutations
Morphological mutants
change in cell wall, colony morphology
Lethal mutant
cell death
Conditional mutant
mutation expressed in certain conditions
Biochemical mutants
Auxotroph (can't synthesize certain compound)
Prototroph (wildtype)
Microbial Recombination
Process in which >1 nucleic acid molecules are rearrange & combined to produce a new nucleic sequence
Occurs following horizontal gene transfer
One-directional
Donor-> recipient
Vertical Gene Transfer
genes transferred from parent to offspring
Horizontal Gene Transfer
transfer of genes from one mature, independent organism to another
exogenote
DNA transferred to recipient
endogenote
genome of recipient
merozygote
recipient cell that is temporary diploid as result of transfer process
conjugation
DNA transfer between bacteria requires direct contact via sex pilus
Can transfer plasmids, episomes, partial or whole chromosomes
Plasmids
cDNA-<30 genes
exist independently of chromosome
Have replication origins
episome
plasmid that can integrate into chromosome
conjugative plasmids
contain genes for pili & transfer copies of themselves
Fertility factors
conjugative plasmids contain genetic info for sex pili formation
Episomes
Resistance factors
have genes for antibiotic resistance
Encode enzymes capable of destroying or modifying antibiotics
Usually conjugative plasmids & may contain transposons
Col plasmids
encode bacteriocins
-bacteriocidal=proteins that destroy other bacteria
Virulence plasmids
encode toxin genes-> make host more pathogenic
Metabolic plasmids
encode genes for enzymes
some allow degradation of unusual carbon sources
Transposable Elements
segments of DNA that move from one chromosomal location to another
allow movement of genes between chromosome & plasmids
allows antibiotic resistance genes to move from chromosome->plasmid-> other cells
tra operon
include genes for pili on F plasmid
most info required for plasmid transfer
insertion sequences
assist plasmid integration into host cell chromosome->allows existence as episome/outside bacterial chromosome
Bacterial conjugation
transfer of DNA by direct cell to cell
1946-Lederburg & Tatum mixed 2 auxotrophic strains-> incubated culture for several hours in nutrition media->plated on minimal media->growth
U-tube experiment
Bernard Davis
demonstrated that direct cell to cell necessary
after incubation in nutrient medium, bacteria plated on minimal medium-> no prototrophs
F+ x F- mating
cell with fertility factor plasmid transfers plasmid to F- cell
F+ = recipient cell
Hfr x F- mating
Hfr (high frequency of recombination) cell has F factor plasmid integrated into chromosome (exhibits high efficiency of chromosomal gene transfer in comparison with F+ cells)
Transfers part/all of chromosome to F-
F- cell remains F- but may recombine with donated DNA
DNA transfer begins when integrated F factor nicked at site of transfer origin since only part of F factor transferred at start, F- doesn't become F+ usually
Hfr Mapping
used to map relative location of bacterial genes based on observation that chromosome transfer occurs at constant rate
Interrupted mating experiment "Blender"
-Hfr x F- mating interrupted at various intervals
-order & timing of gene transfer determined
-break conjugation bridge to study sequence of gene entry into recipient
Finding E. Coli Genetic Map
gene locations expressed in minutes, reflecting time transferred
Larger size-> maps from >1 Hfr required-> F plasmid integrated at different locations
Multiple maps superimposed-> 0 time=threonine locus
F' Conjugation
F factor in Hfr leaves, may pick up portion of chromosomal DNA-> F-
F' plasmid transferred to F- which then becomes F' cell
Transformation
uptake of "naked" DNA
first observed by Griffith 1928
recipient must be "competent" for DNA uptake
-dependent on growth cycle
-required expression of proteins needed for transformation
smooth/rough DNA experiment
Generalized Transduction
during lytic life cycle of bacteriophage virus assembly may incorporate host genes into progeny virus
Specialized Transduction
specific bacterial genes transferred
occurs via bacteriophages with lysogenic life cycles