Bacteria Flashcards
what are the different bacterial morphologies?
- cocci
- rods
- curved
- spiral
- exotic - star-shaped cells
what is the cocci morphology of bacteria?
- simplest, round-shaped cells
- cell machinery at septum for elongation
- some form pairs = diplococci
- some form chains = streptococci
- division planes can be parallel or perpendicular
- form complex organisations like tetrads, sarcinae (multiple perpendicular divisions
- can form microcolonies (piles of cells)
what is the rod morphology of bacteria?
- elongated cells
- bacilli = 1 rod
- diplobacilli = 2 rods
- multiple chains of rods = streptobacilli
what is the name given to a bacteria with a mixture of round cocci and rods?
coccobacilli
what is the curved morphology of bacteria?
- governed by cytoskeleton proteins to create curved shape
- asymmetrical growth is maintained
what is the spiral morphology of bacteria?
spirillum:
- cell shape adapted to organism lifestyle
- helps bacteria move as a corkscrew and penetrate into mucus of epithelial cells
how can bacterial morphologies change during the cell cycle?
- in aquatic environment it forms swarmer cells - has flagella so can swim
- during life cycle, forms a stalk appendage so bacteria can adhere to surfaces
- as stalk grows, they divide asymmetricaly
- produce mobile form (swim) and immobile form (adhere)
what is the advantage of bacteria being small?
large SA:V ratio
- increases nutrient exchange and growth rate
- higher intracellular nutrient concentration
- rapid evolution due to high selection rate of mutations and faster divisions
why do bacteria appear to be different colours?
- they themselves aren’t coloured
- they produce pigments as colonies
- pigments fulfil roles in bacterial life cycles
- Prodigiosin = immunosuppressant
- staphyloaxanthin and violacein = antioxidant, detoxify ROS
- pyocyanin = cytotoxicity, neutrophil apoptosis, proinflammatory
why do bacteria have odours?
- they form biproducts from metabolism which aren’t necessity for life, but do produce odour
- contribution to human odours via degragation of aprocrine products
- conversion of leucine to isovaleric acid by Staph
- production of propanoic acid by propionibacteria - decarboxylation of amino acids to form polyamines
- putrescine, spermidine, cadaverine have role is ROS and signalling
what is the gram staining process?
- colony sample spread onto glass slide and stained with crystal violet, which is +ve charge and penetrates cell envelope
- iodine solution, -ve charge, penetrates cell envelope, and allows crystal violet to form large complexes
- crystal violet is either stuck in envelope or washed off
- counter stain safranin is applied and stains cells that had crystal violet washed off
what is gram positive in the gram stain?
- These bacteria retain methyl violet in their thick peptidoglycan cell walls
- contain no outer membrane
- Examples of Gram-positive bacteria are
Staphylococcus and Streptococcus
what is gram negative in the gram stain?
- These bacteria have only thin peptidoglycan cell walls
- contain an outer membrane
-Gram negative bacteria therefore appear pink-red. - Examples of Gram-negative
bacteria are Escherichia, Pseudomonas and Neisseria
what bacteria does the gram stain not work on?
Mycobacteria
- has a unique cell envelope composition where the lipids interfere with the staining process
what are S-layers?
- found in gram +ve and -ve
- non-covalently bound to cell surface- - facultative (don’t exist in most model organisms)
- proteinaceous crystalline arrays - self assembly products
- 20% of bacterial production
- not all bacteria have them
- function unknown
what are capsules?
- found in gram +ve and -ve
- made of polysaccharide
- some made of amino acids (poly-gamma-D-glutamate)
- covalently bound to peptidoglycan (gram +ve) or outer membrane (gram -ve)
- resistannt to phagocytes and bacteriophages
- keep environment hydrated
what are exopolysaccharides?
- homo- or heteropolysaccharides
- non-covalently bound to cell surface
- important for biofilm formation
- form aggregates to protect fom environment
- enables formation of colonies
- economic importance: xanthan gum
what are the key components of the outer membrane in gram negative bacteria?
- phospholipid bilayer
- innerface = phospholipids
- outerface = lipopolysaccharide, contains hydrophobic region
- variable-O-antigen polysaccharide determines antigen on cell surface
- contains porins for solute transport
- lipoproteins are covalently linked to peptidoglycan
- LPS endotoxin is a potent activator of the immune system: can trigger inflammation
what is peptidoglycan?
- made of murein
- maintains cell shape and forms exoskeleton
- acts as scaffold to display proteins
- protective role
- acts as a sieve to regulate dynamic exchanges with environment
- elastic 3D network
- present in almost all bacteria
- resistant to osmotic stress
what is the composition of peptidoglycan?
- made of murein
- glycan chains alternating N-acetylglucosamine G and N-acetylmuramic acid
- substituted via short peptides (L and D amino acids)
how is peptidoglycan assembled?
- transpeptidation reaction with enzymes which are covalently bound to the stem
what are the key components of the cytoplasmic membrane?
- phospholipid bilayer
- unsaturated fatty acids modulate membrane fluidity and permeability due to kinks in hydrocarbon chain
- amphipathic molecules for compartmentalisation
- hopanoids modulate membrane fluidity and permeability
- protein transporters make membrane selectively permeable for specific polar molecules
what is the bacterial chromosome made up of?
- dsDNA
- singular, circular chromosome
- 0.5-14.8mbp
- organised as a nucleoid: histone-like proteins enable supercoiling
what is the bacterial plasmid made up of?
- dsDNA
- variable copy number
- 2-600kbp
- self-transferable by horizontal transfer
- conjugation: physical contact between bacteria to transfer plasmid DNA
- carry resistance genes
what is the gene structure of bacteria?
- no introns
- continuous coding sequence via open reading frame (ORF)
- operons: one promoter, several ORFs
- genes are smaller than eukaryotic genes
how is gene transcription initiated?
- RNAP scans DNA forming a loose complex
- sigma factor binds to shine-dalgarno sequence upstream of start codon (-35 and -10)
- DNA is unwound to form an open complex
- transcription begins and sigma factor is released
what is rho-independent transcription termination?
- requires palindromic GC-rich region upstream of an AT-rich sequence
- once the GC-rich region has been transcribed, it forms a hairpin
- hairpin causes dissociation of RNAP, helped by the AT-rich sequence (few H-bonds)
what is rho-dependent transcription termination?
- rho proteins recognise and bind to 72 GC-rich residues
- RNA-dependent ATPase wraps the downstream RNA around itself
- once it reaches the polymerase, rho unwinds the RNA-DNA heteroduplex and releases RNAP
how is the genetic machinery different between prokaryotes and eukaryotes?
- transcription site is cytoplasm for bacteria (nucleus for eukaryotes)
- 1 RNAP in bacteria, 3 RNAP eukaryotes
- termination involves palindromic GC-rich region (eukaryotes require AAUAAA)
- mRNA is modified in eukaryotes, but not in bacteria
how large are the ribosomes in bacteria compared to eukaryotes?
70S in bacteria
- 50S + 30S
80S in eukaryotes
- 60S + 40S
what are the differences in translation between bacteria and eukaryotes?
- bacterial 70S ribosomes interact with mRNA productively in presence of tRNA
- 30S subunit recognises SD sequence
- transcription and translation are coupled in bacteria
- eukaryotic 80s ribosomes bind mRNA efficiently in absence of tRNA
- 40S subunit is guided by 5’ cap on mRNA
- compartmentalised in eukarytoes
- translation inhibited by cycloheximide
what does bacterial growth require?
- temperature
- pH
- osmotic pressure
- nutrients
- oxygen
what are the cardinal temperatures for bacteria?
Minimum: below this, no growth ca occur as the membranes gel and transport processes are too slow to maintain metabolism
Optimum: an increased enzymatic activity so that reactions occur at the maximal possible rate
Maximum: after this point, proteins become denatured, cytoplasm collapses and cell lysis occurs, causing metabolic reactions to halt
name 4 bacteria and their optimal temperatures:
- P. vacuolata = 4C
- psychrophile - E. coli = 37C
- mesophile - T. aquaticus = 70C
- thermophile - P. fumarii = 106C
- extreme thermophile
how are psychrophiles adapted to cold temperatures?
- increased membrane fluidity
- more unsaturated and polyunsaturated methyl-branched fatty acids
- limit membrane cohesion - production of anti-freeze proteins
- AFPs bind to ice crystals to inhibit their growth by covering water-accessible surfaces of ice - production of cryoprotectants
- trehalose and exopolysaccharides lowers freezing temp of water to preserve fluids - production of cold-adapted enzymes
- more a-helices and less weak bonds to combat low enthalpy
how are thermophiles adapted to high temperatures?
- genome protection
- by DNA-binding proteins stabilise DNA
- reverse DNA genes form supercoils - harder to pull apart
- high GC% - difficult to denature - modify membrane composition
- stable ether-linked phosolipids
- single lipid layer: glycerol tetraethers - thermostable proteins
- more ionic and hydrophobic interactions to form stronger bonds - thermostable chaperonins
- maintain protein folding
- thermosome in Pyrodictium abyssi
how are acidophile metabolisms adapted to their environment?
- increased membrane impermeability
- use protons as currency
- ATP depends on oxidative phosphorylation and needs H+ for this
- use H+ to import/export Na+
- H+ powers motility: when pumped in, H+ triggers conformational change to move flagella
- H+ secretion systems via antiporters
- DNA/protein repar mechanisms
- reverse membrane potential and increased osmolarity (K+ ions)
how are alkaliphile metabolisms adapted to their environment?
- Na+ as currency
- use Na+ to make ATP
- used for motility, imports/exports etc
- high affinity transporters for Na+
how are bacteria adapted to osmotic stress?
- regulate water movements by passive diffusion and aquaporins
- produce compatible solutes like proline, glutamic acid and betaine
- release solutes via mechano-sensitive channels
- halophiles accumulate osmolites in their cytoplasm
what are halophiles?
- require high salt conc to grow
- S-layer is stabilised by Na+ ions, and K+ must be >4M in the cell
what are non-halophiles?
cannot be exposed to high salt conc
what are halotolerant bacteria?
can deal with high osmotic pressure by high salt conc, but to an extent
what are extreme halophiles?
- usually archaea
- need high salt conc
what nutrients do bacteria require?
- nitrogen
- sulphur
- phosphorus
- vitamins
- K+, Ca2+, Mg2+ (cofactors)
- trace elements (Fe, Cu, Zn)
what nutrients do bacteria require?
- nitrogen
- sulphur
- phosphorus
- vitamins
- K+, Ca2+, Mg2+ (cofactors)
- trace elements (Fe, Cu, Zn)
what do bacterial metabolisms need?
- energy source
- source of electrons
- carbon source
bacteria can switch from one metabolism to another depending on the available resources
what are the toxic forms of Reactive Oxygen Species (ROS)?
- superoxide
- hydrogen peroxide
- hydroxyl radical (most toxic)
what is the detoxification reaction in ROS?
02 + 4e- + 4H+ = H20
what enzymes do bacteria produce to detoxify ROS?
- catalase/peroxidase: convert H2O2 to H20
- superoxide dismutase + catalase converts O2 to H2O2 then H2O
- superoxide reductase + catalase coverts O2 to H2O2 then H2O
what are the different oxygen requirements for different bacteria?
- obligate aerobes: catalase + superoxide dismutase
- use exclusively O2 for respiration
- P. aeruginosa - facultative aerobes: catalase + superoxide dismutase
- can use O2 for respiration
- E. coli - microaerophiles
- require O2 for respiration
- C. jejuni - anaerobes aerotolerant: superoxide dismutase
- do not use O2 for respiration
- S. mutans - obligate anaerobes = C. difficile
what are the 3 direct measurements of bacterial growth?
- Microscopic counts
- flow cytometry
- viable counting
what are microscopic counts for bacterial growth?
- manually count no. bacteria per unit on glass slide
ads: cheap
disads: time-consuming, must be accurate
what is flow cytometry?
- machine pumps cell suspension through capillary
- suspension is hit by a laser and measures scattered light by the bacteria present
ads: can distinguish live/dead bacteria, distinguish specific species using antibodies, high sensitivity
disads: expensive equipment
what is viable counting?
- serial dilutions of inoculum is spread on agar
- colonies are counted using cell counters
ads: distinguishes different species, identifies live bacteria
disads: time consuming, assumes one colony comes from one cell
what are the 3 indirect measurements of bacterial growth?
- optical density
- dry weight
- metabolic diversity
what is optical density?
Optical density
- expose cell suspension to light through prism with given wavelength
- measure unscattered light, which is proportional to no. cells in suspension
disads: requires high cell densities, doesn’t distinguish live/dead cells, OD values vary depending on organisms, doesn’t work with molds/filamentous bacteria
what is the dry weight measurement of bacterial growth?
- measures cell weight after freeze-drying
disads: time-consuming, requires expensive equipment
what is the metabolic activity measurement of bacterial growth?
- measure production of specific metabolites
ads: can measure activity of a specific group of bacteria
disads: complicated to do
what is the bacterial growth curve?
- exponential growth of bacteria
- new generation every 20-30 mins
what are the 4 phases of the bacterial growth curve?
- lag phase
- metabolism starts, but no division - log phase
- exponential increase in population
- logarithmic growth due to binary fission - stationary phase
- microbial deaths balance production of new cells
- fermentation releases acidic compounds which lower pH of the medium, so can no longer sustain growth - death phase: decrease in population
what is the thermal death point?
- min temp at which all organisms are killed in 10 mins in a particular liquid
what is the thermal death time?
- min time required to kill all bacteria in a particular liquid at a give temperature
what are the 3 methods microbes can be killed using heat?
- moist heat (boiling/autoclave): 15 mins at 121C under pressure to kill spores
- dry heat (oven): direct flaming, incineration >150C for 2 hrs
- pasteurisation (mild heat)
- HTST: 72C for 15secs, kills 99.9% viable microorganisms in milk
- UHT: 140C, 2-5secs
how can irradiation kill microbes?
- UV, x-rays and gamma rays have short enough wavelengths to kill bacteria
- the shorter the wavelength, the more energy provided (indirect proportional)
ionising radiations:
- food industry, medical/lab equip
- DNA destruction via ROS and breaking double strand
non-ionising radiation:
- surface decontamination
- DNA damage
how can filtration kill microbes?
- sterilise gases or liquids that can be damaged by heat
- porosity of filters (1mm to 0.01um) can be chosen for specific microbes
- nucleopore filter: membrane with holes that filters microbes
- membrane filter: mesh of continuous polymers
- depth filters: fibres stuck on top of one another
what are bacteriostatic antimicrobial agents?
- doesn’t kill bacteria, only stops growth
- maintain no. cells
- maintain viable cell count
what are bactericidal antimicrobial agents?
- stop growth
- trigger cell death as viable cell count decreases
what are bacteriolytic antimicrobial agents?
- cause cell lysis
what are sterilants?
- eliminate all forms of microorganisms, including spores, on objects
- e.g. ethylene oxide: gas so is effective
- most powerful antimicrobial compound we can use
what are disinfectants?
- kill microorganisms, but not endospores, on objects
- e.g. alcohol (60-85% in water)
what are antiseptics and germicides?
- inhibit growth
- kill microorganisms on tissues
- e.g. handwash
how can antimicrobial activity be measured?
- disc diffusion technique
- Minimum Inhibitory Concentration (MIC): growth inhibition measurement
- Minimum Bactericidal Concentration (MBC): cell death measurement
what is the disc diffusion technique to measure antimicrobial activity?
- inoculate plate with a liquid culture sample
- discs containing antimicrobial agents are placed on surface
- incubate for 24-48hrs
- test organism shows susceptibility to some agents, indicated by zones of inhibition
what is the MIC growth inhibition measurement of antimicrobial activity?
- MIC is the lowest concentration of a drug inhibiting the visible growth of a test organism after overnight incubation
what is the MBC cell death measurement of antimicrobial activity?
- MBC is the lowest conc of a drug killing 99.99% of a test organism after overnight incubation
- test no. viable cells after being in contact wit antimicrobial agent
what are phenolic compounds?
- aromatic derivatives
- low anaesthetic at low conc, antibacterial at high conc
- disrupts cytoplasmic membrane and denatures proteins
how are alcohols used in antimicrobial control?
- denatures proteins and disrupts cytoplasmic membranes
- lipid solvent
- active conc between 60-85%
how are aldehydes used in antimicrobial control?
- alkylating agents containing aldehyde groups
- formalin prevents bacterial growth
- modify proteins and DNA, leading to cell death
what are quaternary ammonium compounds?
- interact with phospholipids of the cytoplasmic membrane
- cationic detergents
- long chains with charge tail allows disruption of membrane
what are halogen-releasing agents?
- chlorine-releasing agents
- bleach ionises to form Na+ and hypochlorite ion OCl-, in equilibrium with hypochlorous acid (HOCl)
- form chlorinated bases in DNA and oxidation of proteins - iodine-releasing agents
- iodine/iodophores
- target DNA and proteins
what are the two major therapeutic strategies against pandemics?
- antibiotics
2. vaccinations
who described germ theory?
Louis Pasteur 1860
what is Koch’s Postulates?
- in 1890s, Koch discovered the causal relationship between a microbe and a disease
- used germ theory
- microbe must be found in all organisms suffering the disease, but not in healthy organisms
- microbe can be isolated from diseased organism and grown in culture
- cultured microbe should cause disease when applied to healthy organism
- microbe is reisolated from the inoculated host and identified as being identical to the original microbe
who discovered penicillin?
Alexander Fleming in 1928, produced by a fungus mold
what are the major classes of antibiotics?
- Cell wall inhibitors
- beta-lactams
- glycopeptides - protein synthesis inhibitors
- aminoglycosides
- cyclines
- MLS
- oxazolidinones - DNA metabolism inhibitors
- quinolones
what causes antibiotic resistance?
- antibiotic overuse and misuse in human therapeutics
- farming: animals can be fed with antibiotics at subtherapeutic doses (4x human consumption)
- agriculture: treatment of diseases in plants
- aquaculture
- pets
what are the key properties of an ideal antibiotic?
- It must display selective toxicity towards the target bacteria
- must inhibit an essential process or inhibit virulence - it must be stable in the host and active at low concentration
- It must be cheap
what are beta-lactams?
- penicillin antibiotic that inhibits peptidoglycan polymerisation
- they target D, D-transpeptidase penicillin binding proteins (PBP) in bacteria
- they form covalent bonds with amino acid in position 4, to form a crosslink from C-terminal to acceptor group
- they are structural analogs of D-Ala-D-Ala C-terminal residues in peptide stem
- they are used by PBP as substrates and inactivate these enzymes irreversibly
what are the 4 models of antibiotic resistance by bacteria?
- drug inactivation: beta-lactamases
- target modification: low affinity
PBPs/overexpression - efflux/impermeability: efflux systems in gram -ve bacteria
- bypass: alternative pathway
what are the 4 methods in which are bacteria resistant to beta-lactams?
- inactivation by beta-lactamases
- mutation of the target enzyme PBP
- secretion of the antibiotic by gram negative bacteria
- modification of the synthetic pathway targeted by beta-lactams
how do bacteria inhibit beta-lactams via beta-lactamases?
beta-lactamases can hydrolyse the antibiotic, causing it to become inactive as the structure no longer resembles D-Ala-D-Ala:
- nucleophilic attack by catalytic serine
- covalent complex penicillin-beta-lactamase
- penicillin hydrolysis
beta-lactamases evolved from the same family as PBPs
how does mutation in the target enzyme PBP make bacteria resistant to beta-lactams?
- used by gram-positive bacteria
Two methods:
1. They form low affinity PBPs so that there reduced affinity of beta-lactams for PBP
- Overexpress PBP targetted by beta-lactams
how does mutation in the target enzyme PBP make bacteria resistant to beta-lactams?
- used by gram-positive bacteria
Two methods:
1. They form low affinity PBPs so that there reduced affinity of beta-lactams for PBP
- Overexpress PBP targetted by beta-lactams
how do gram negative bacteria secrete the beta-lactam in antibiotic resistance?
e. g. P. aeruginosa
1. overproduction of the MexAB-OprM system
- carbapenem resistance
- Overproduction of the MexEF-OprN system
- imipenem resistance
how do bacteria modify the synthetic pathway that is targeted by beta-lactams in antibiotic resistance?
- they remodel how peptidoglycan is synthesised
- L, D transpeptidases cause different type of crosslinking which is beta-lactam insensitive
- they form a 3,3 bond rather than a 3,4 bond
