Growth and metabolism Flashcards
Limits on bacterial growth
Exhausted nutrients Accumulation of toxic byproducts Antibiotics from neighboring Immune function Environmental (ex physical barriers)
Generation time
Time to double population
Range from <30 min - 20 hours
Only relevant during exponential phase
Major nutrient requirements
Carbon - both oxidation and cell structures Nitrogen - protein and DNA Phos - phospholipids and DNA Sulfur - proteins Iron - enzyme cofactor (metabolism)
Carbohydrate sources
Provides energy (ATP) and C for growth
Use depends on specific degradative enzymes
- can be diagnostic
Amino acids as nutrients
Sources of C, N, S
Proteins too big -> secrete proteases -> absorb acids and small peptides
Some require specific acids (no enzymes to create)
Some can use ammonia (NH4) and nitrate
Nucleic acids as nutrients
Sources of C, P, N
Secrete nucleases -> absorb DNA and RNA
Phospholipids as nutrients
Provide C, N, P
Secrete phospholipases
-> host cell lysis due to degradation
- tied to Fe metabolism (lysed cells release)
Iron metabolism
Necessary for enzymes (oxidative) Key site of host-pathogen interaction - host factors bind/sequester iron - bacteria makes - phospholipase - lyses cells to release Fe - siderophores/chelators to extract
Growth factors
Not metabolized for energy Used to make essential metabolites Diagnostic - E coli - only requires glucose, NH4 and ions - others "fastidious"
Temperature requirements
Most pathogens grow best at 37C (body core)
Most env’t grow best at 22-30C (room temp)
Exceptions
- Mycobacterium leprae -> growth in colder body areas (hands vs core)
- Legionella - grows at high temp (45C)
- Listeria - grows at low temp (4C = refrigerator -> food poisoning)
Physical requirements for growth
Temp (separate slide)
pH - usu 6-8 with 7.4 optimal (body)
- Clostridium botulinum - grows less in acid
Osmotic pressure
- cell wall helps withstand pressure changes
- high salt or sucrose inhibits growth (pickles, jam)
Growth in lab
Peptone broth + glucose
- peptone = meat + pepsin = protein and aminos
Blood agar - supports almost all pathogens
Agar - non-digested sugar - solidifies
- need solid medium to isolate colonies
Measurement of bacteria
Optical density - proportional to concentration
- spectrophotometer or naked eye turbidity
- limit is 10’6/mL
Metabolic activity (CO2 or ATP production)
- sensitive for bacteremia
Direct count - chamber in microscope
- insensitive (10’5/mL) -> any visible bacteria indicates infection
- includes both living and dead cells
Plate count - most common, accurate
- after serial dilution, 24h incubation
- yields colony forming units (CFUs)
Growth curves
Usu log scale number/mass vs time
Lag phase - metabolic activity, mass and size inc but no division
- adaptation to new medium -> new RNA, proteins
Exponential phase - binary fission
- constant generation time (bacteria, medium, temp, etc)
- “balanced growth” all components double in same time (mass, DNA)
Stationary phase - steady state of death and growth
- can last days-years
- usu 10’9/mL for pathogens in test tube
Death phase - death>growth, can last years
- unusual morphology
Glycolysis
aka Embden-Meyerhof pathway
Glucose + 2 ATP + NAD -> pyruvate + 4 ATP + 1 NADH
ATP from substrate-level phosphorylation (direct)
TCA cycle
aka tricarboxylic acid cycle, Krebs
Produces “reducing equivalents” aka NADH -> ETC
Glyoxylate shunt: fatty acit -> acetate -> AcCoA -> NADH
Eletron transport chain
Sim to mitochondria
Cytochromes in inner membrane
NADH -> electron acceptor -> proton gradient ->
ATP via oxidative phos
Electron acceptors:
aerobic: O2 -> H2O
anaerobic: not O2, ex NO3 -> N2
Fermentation overview
Glycolysis but no TCA and ETC (no enzymes, no O2)
Pyruvate + NADH -> NAD+ + reduced fermentation product
ATP - much less than respiratory chain (substrate level only)
Fermentation product excreted (most carbon)
Lactic acid fermentation
Lowers pH - inhibits other microbes
Ex:
Lactobacilli in intestine, vagina -> inhibit Candida
- also in yogurt, cheese, etc
Strep pyogenes (strep throat)
Butyric acid fermentation
Clostridium!!
- also produce butenol-acetone, acetic acid, CO2, H2, EtOH…
Distinctive odor
Protective against some pathogens (E coli)
Propionic acid fermentation
Lactic acid -> propionic acid + acetic acid + CO2
Corynebacteria, Propionibacteria, Bifidobacteria
Swiss cheese
Propionibacteria -> deep follicles -> acne
Mixed acid fermentation
Enterobacteriae (Gram (-) bacilli)
Pyruvate -> AcCoA + formic acid
- formic acid -> H2 + CO2
- AcCoA -> EtOH + acetic + succinic (“mixed acid”)
Ex:
- Salmonella -> H2 gas (gas gangrene)
- Shigella - lacks enzyme - formic acid builds up
- E coli
Butanediol fermentation
Mixed acid fermentation -> acetoin -> 2,3 butanediol
(less acidic product)
Non-fecal enteric bacteria (Klebsiella, Enterobacter)
vs acidic fecal (E coli, Salmonella, Shigella)
Ethanol fermentation
Pyruvic acid + NADH -alcohol dehydrogenase> EtOH + CO2
Saccharomyces cerevisiae -> beer, wine
(CO2 -> gas in slight acidity)
Candida albicans -> fungal infections (no gas due to alkalinity)
Strickland reaction
Clostridia in wounds
Paired amino acid fermentation (one donor, one acceptor)
-> decarboxylated acids
-> odor, volatile
Obligate aerobes
Require O2, all ATP via O2 respiration
(no fermentation enzymes)
Mycobacterium tuberculosis -> upper lobes of lung
Pseudomonas aeruginosa - not technically correct definition, no fermentation but can have NO3 respiration
Facultative anaerobes
Both respiration and fermentation
Grow fastest with O2 (more ATP from fermentation)
Many pathogens
E coli
Staphylococcus
Obligate anaerobes
Only fermentation
Oxygen toxic - require reducing conditions
- flavin enzymes -> H2O2 -> superoxide (O2-), ROS -> damage
- lack catalase (for H2O2) and superoxide dismutase
Ex Bacteroides fragilis (intestine, abcesses)
Aerotolerant anaerobes
Lack respiratory system
Can survive O2 exposure but prefer anaerobic
Lactic acid bacteria
- Lactobacilli
- Streptococcus
Clostridium (tetani, botulinum)
Microaerophiles
Aerotolerant only at low concentrations (ex 5% vs 20% atm)
Campylobacter jejuni
Anaerobic tissues
Mouth, GI
Surface tissue if other bacteria or host is using up O2
Culture if reducing medium (ex cysteine)
Glucose fermentation
Usu GI microbes
(E coli, Klebsiella, Salmonella, Shigella, Citrobacter)
vs lactose - requires additional enzyme to cleave
only E coli and Klebsiella, Citrobacter slow
Pseudomonas doesn’t use either…
