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Microbiology
Week 1 - Microbiology and landmarks
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Microbiology - study of micro-organisms too small to see with the naked eye - Prokaryotic - no
true membrane bound nucleus ( bacteria, archaea) Prokaryotes generally single chromosome,
no membrane, although still have genetic material, have no distinct nucleus
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Viruses - acellular
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Microbes dominate diversity
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Multiply very rapidly, E
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Bacteria – peptidoglycan wall - Mostly very small 1-6 μm in size, unicellular, Diverse range of
nutrition, Found in diverse range of environments, mostly harmless, many beneficial, others
cause serious disease
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Many found in extreme environments, Now know they are everywhere,
first non-extreme archaeon isolated in 2004
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Mycelium with spores, Yeast single cell budding
fungi, Degrade organic matter, absorptive heterotrophs
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Chitin wall prevents phagocytosis, Produce hyphae and mycelium with
specialised structures for spore production
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Protists - very diverse group, don't belong to animal, plant or fungi, size range from 1-150um
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Smallest microbe (< 0
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Require a cell to multiply, infect most living cells
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History of microbes 3
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8 billion years ago - Appearance of first recognisable cells, Cyanobacteria, Produce
oxygen during photosynthesis - 6H20 + 6CO2 ® C6H1206 + 602 (sunlight and chlorophyll)
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Development of eukaryotes
300 million years ago - Appearance of multicellular organism- evolve in presence of microbes
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Post-Renaissance - Jenner invents vaccination, 1798
Modern era - Spanish flu, 1918, Pandemic kills >20 million worldwide, Discovery of penicillin,
1929, Winning the battle against bacterial disease, Smallpox eradicated, 1979, AIDS (HIV)
identified, 1980’s
Microbes still essential to balanced ecosystem - all nutrient cycles need microbes - degrade
organic matter, recycle carbon, fix nitrogen (rhizobium), recycling nitrogen
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Infects 20% of
World’s population, Causes 3 million deaths every year
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Ebola
virus
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Malaria Plasmodium spp
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No known Archaeal diseases
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Cell morphology - First step in classical identification of bacteria: Gram positive (purple) – rods
or cocci, Gram negative (pink) – rods, cocci, curved
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Grouping Diplococci - 2 cocci together
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Grape like clusters - cocci in several planes
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Grouping dependant on division, see picture above
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Neisseria meningitidis (Gram –ve cocci)
Human only host, colonises throat and infects blood and brain, Meningitis
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Yoghurt production, Nonpathogenic
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Non-staining - Treponema pallidum (G-ve)
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Mycobacterium tuberculosis
(G+ve) - Acid fast stain, tuberculosis
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Structure of prokaryotic cell:
Motility from flagella - move towards sources of nutrients - chemoattractants, and away from
toxic agents - chemorepellants, moves counterclockwise - swims one direction, move
clockwise, tumble

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Fimbriae - attachment - Hair-like fibres on cell surface, Allow bacteria to adhere to solid
surface, Environment, biofilm production, Recognise a specific molecule on cell surface, often
carbohydrate, Disease; tissue specificty, host specificity, First step in invasion of host cell
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Generally carbohydrate, can be polypeptide
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Regular crystalline like layer of protein on the surface, eg hexagonal array, Protects
from bacteriophage and other agents, sieve, Interest in nanotechnology
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1,400 mm long within E
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Approximately 4 x 106 bp in E
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Plasmids - dsDNA molecules
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May integrate into genome,
Generally not required for growth and division; may confer critical phenotype
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Cytosol - Ribosomes (70S cf eukaryotic 80S)
densely packed in cytosol, mRNA – rapidly degraded (factory for protein synthesis)
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Specialised structures (exclusive to specific bacteria); Gas vacuoles –
buoyancy, Magnetosome - orientation
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Has phospholipid bilayer with embedded
proteins
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Structure of Eukaryotic cell - contains mitochondrion, centriole, vesicles, cytosol, rER/sER,
ribosomes, plasma membrane, flagellum, golgi, chromatin, nucleus with nuclear envelope and
microtubules
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:
Gram positive envelope - 1 membrane, thick peptidoglycan, teichoic acids embedded in
peptidoglycan
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Peptidoglycan structure - single molecule, disaccaride backbone, crosslinked via pentapepide,
strength, gives cell shape, unique to bacteria - target for host defense and antibiotics
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Endospores - Very resistant to heat, dessication, radiation, chemicals - bacillus and clostridium
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IV Cortex laid
down between the two membranes, Ca dipicolinic acid accumulated
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Sporogenesis - Activation – heating, Germination – spore swelling rupture,
absorption of spore coat, loss resistance, increase in metabolic activity, Outgrowth
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Nutrients needed for - biosynthesis and energy - source of reducing power for hydrogen or
electrons - results in growth
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H, O, also in many
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Macronutrients - 95% of any cell consists of only a few major elements: Carbon, Hydrogen,
Nitrogen, Oxygen, Phosphorous, Sulphur
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Micronutrients - needed in trace amounts or growth stops - manganese, zinc, cobalt, nickel,
copper
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Virtually any organic compound
can be used by some bacteria (HETEROTROPHY)
Used as: backbone for all cellular components, energy source, supplies electrons CHO +
-
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Carbon heterotrophs - glucose often
preferred C-source - variety of C-sources in vast - Many common soil bacteria can utilise
unusual compounds – pesticides, rubber, antibiotics
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Carbon can be fixed chemically - CO2 + H2S ® CH2O + S
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Nitrogen - Required for protein, nucleic acid, some lipids, carbohydrate - Organic molecules,
ammonia, nitrate (NO3-), which becomes reduced to ammonia (NH4+) (assimilatory nitrate
reduction), Nitrogen gas via nitrogen fixation (N2 to NH3) eg Rhizobium - nitrogenase
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Sulphur - generally absorbed as sulphate and reduced
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Carbon - Autotrophs - CO2 main or only source of carbon
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Energy source - Phototrophs - make energy using
light
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Electron source - lithotrophs (rock eaters)reduced inorganic compounds
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g
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Culture of bacteria - defined synthetic media - Known contents - chemcially defined,
concentration of each constituent known, specific source of nutrients and ions
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Uses - selectivity - specific carbon sources, limits other nutrients
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Complex media - Unknown contents - undefined composition - Include digested
proteins (casein), Extracts of complex substances (Beef extract, Yeast extract)
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Additives for enriched media
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Differential media - Distinguish between organisms,
Can allow tentative identification
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Prokaryotes can live almost anywhere - Most survive in “normal” conditions of
water, pH and temperature, Extremophiles - High temperature, salinity, pressure, pH, anoxia
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Water activity measure of availability - aw = relative humdity/100, limits growth
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pH tolerance - measure of H+ concentration - Acidophils from pH 0-5
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5 to 11
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Neutrophiles from pH 5
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All maintain an internal pH of 5
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Temperature - microbes dont regulate temperature, temperature increases speed up
reactions - increasing temp demages cells as it denatures enzymes and disrupts membranes
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Psychrophilic - cold-loving (-2 to 20°C)
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Hyperthermophilic - Very high temp (80-113°C)
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Radiation - As wavelength decreases, energy increases, Generally bacteria are susceptible to
ionising radiation eg gamma allow some can survive
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Have recombination repair systems to deal
with this and repair – balance)
Bacteria growth in culture - The growth of microorganisms is greatly affected by the chemical
and physical nature of their surroundings
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Requirements can include specific nutrients, e
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Glucose, Certain amino acids
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Substances can be included in media to specifically prevent growth of certain type(s) of
bacteria
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Oxygen - Obligate aerobe – Oxygen essential for growth, Facultative anaerobe – Grows best
with but also without oxygen, Aerotolerant anaerobe – Grows equally well with or without
oxygen, Obligate anaerobe – Cannot grow in the presence of oxygen, Microaerophile –
Requires low level of oxygen
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Temperature - Psychrophile – optimum ≤15ºC, Psychrotroph – optimum of 20 to 30ºC but can
grow as 0ºC, Mesophile – optimum 20 to 45ºC, Thermophile – optimum 45 to 65ºC,
Hyperthermophile – anything up to 113ºC
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Solid culture - agar plates, used for general maintenance of cultures, used for separating mixed
cultures, small scale growth of bacteria
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Growth and division of a bacterial cell - cells grow and then divide by binary fission, generation
time can be <20min, continous process as long as environmental conditions permits,
population increases exponentially
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Innoculated cells often depeleted in energy and
essential cellular components (e
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ATP, ribosomes, enzyme cofactors), Need to prepare for
growth
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Balanced
growth - All cellular components in balance, In up-shift or down-shift balance is disturbed,
Growth decreases until balance resumed
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g
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Death phase - Number of viable cells slowly declines, Can be logarithmic, Dependent upon
conditions, Death rate can slow as numbers decrease, Survival of resistant cells
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Only a few generations are possible
before nutrients are depleted
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Safe - can be properly sterilized
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Complete
conversion of substrate is possible
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Much idle time: Sterilization, growth of inoculum, cleaning after the fermentation
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Continuous culture – a culture of microorganisms in a medium which is maintained under
constant environmental conditions with a constant nutrient supply so that it can grow over an
extended period of time
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Maintenance of steady state conditions Media fed into the chemostat at a steady rate, Biomass and spent media removed from
chemostat at the same steady rate
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Advantages - Works all the time: low labour cost, good utilization of reactor
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Automation may be very appealing
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Disadvantages - Often
disappointing: promised continuous production for months fails due to infection or
spontaneous mutation of microorganisms to non producing strain
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Week 4 - Bacterial pathogen
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Pathogen bacteria - bacteria capable of inducing damage
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Bubonic (transmitted via flea bite - zoonosis), pneumonic (direct respiratory)
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Diphtheria - Corynebacterium diphtheriae
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Spread by airborne route
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Scarlet fever: Streptococus pyogenes - scarlet fever late 19th early 20th century – red rash, fatal,
puerperal fever and death post child birth, streptococcal gangrene, necrotising fasciitis,
airborne and foodborne transmission – spread rapidly within families, could loose ¾ of one
family
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Whooping cough: Bordetella pertussis - Highly contagious infection of respiratory system Dry
cough, severe bouts, whoop
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Vaccine effective, but short lived protection, last few years
reappearance of disease
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Legionnaire’s disease – Legionella pneumophila, (air
conditioning and conference centre)
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Newly evolved bacteria - 'killer E
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g
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Through misues bioterrorism - plague, anthrax, botulinum toxin
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Many free- living bactaria and facultative pathogens
acquire new genes that help them survive in a new environment
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Pathogen - organism capable of inducing damage
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Virulence of a bacterium depends on - infectivity, inasiveness and
pathogenic potential
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Identifying causative pathogen - Koch's postulates - Organism must be present in all cases
(absent in healthy individual) Organism must be isolated from host and grown as pure culture
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Microorganism must be isolated from experimentally infected, diseased host
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Stages of pathogenesis:
Reservoir and transmission - Human respiratory system – airborne, inanimate objects, hands
etc
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Humans; soil – direct
contact; sexually transmitted; wound puncture eg rusty nail
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Mode of transmission often dictates site of infection
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Oral – GI tract, generally local eg most Salmonella
but may become systemic eg typhoid
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Adhere to and colonise host - pili (fimbriae) - long thin structures extending from cell surface,
terminal adhesins, bind specifically to host cell carbohydrate
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Evasion of host immune system - Capsule - Carbohydrate, anti-phagocytic, May mimic host
molecules, Enzymes - destroy components of immune system, Modify surface molecules to
mimic host antigens, Invade host cells, hide from immune system
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Growth and multiplication in select environment - adapt to environment - temp, pH, redox
potential, osmolarity, nutrients, iron, differential gene expression
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g
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Induce
inappropriate immune response, results in disease
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Week 5 - Phylogeny and Important microbes
Phylogeny
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Taxonomy words, done for cataloguing, predictions, grouping organisms to allow work with
similar organisms, accurate identifications, 3 parts:
Classification - arrangement based on evolutionary relatedness
Nomenclature - assignment of taxonomic groups
Identification - defining where a particular isolate belongs
Earliest microbial ancestor anaerobic
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Prokaryotes divided into 2 domains - bacteria and archaea
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Bacteria - prokaryote cells, no nucleus, bacterial ribosomal RNA, diacyl gylcerol disesters are
main membrane lipids
Archaea - Archaeal ribosomal RNA, isoprenoid glycerol diether in membrane
Eukaryotes - Contain nucleus, eukaryotic rRNA, gylcerol fatty acyl diester membrane lipids
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May get divisions within species, strains exhibit some distinct
different to other strains within a species
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Phenetic classification - arranges groups based on mutual similarity of phenotypic
characteristics
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e
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Classification system 2 - phylogenetic classification - based on evolutionary relationships
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Phylogenetic classification - molecular chronometers - nucleic acid - DNA/RNA and protein
change over time, Uses housekeeping essential genes, Assumes there is an evolutionary clock,
Gradual change over time without alteration of function Assumes changes are selectively
neutral, occur randomly and increase linearly over time, Highly different sequences indicate
divergence long ago
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Indicators of phylogeny - can use full gene/protein sequence or fragments, ribosomal RNA,
DNA, Protein
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 Assessing similarity - make alignments of sequences, using computers determine level of
diversity, make phylogenetic tree - similar organisms cluster tighter together, then get larger
groupings
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Can be
useful in assigning the specific class of bacteria
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TB - infection mainly in the lungs but can be
found in other organs
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Detected by X-ray - tubercles seen as opaque
patches, or mantoux test - 0
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Cells
slightly curved or straight rods
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o Bacillus - used for industrial scale enzymes - amylases, proteases, lipases
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Gram
positive rods which are 2-3um long and 0
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Heterotroph: primarily aerobic but
weakly fermentative, mesophile, motile with lateral flagella, forms heat and diesiccation
resistant endospores situated centrally
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o Streptomyces - fungi-like bacteria, live in soil, produces geosmin, used for wide range of
antibiotics, no nuclear membrane
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Apical spores - spores produce germ tube that grow by apical growth
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o Escherichia - in gut, phylum - proteobacteria, gram negative rod, 50% G+C content DNA,
optimum growth at 37degrees, facultative anaerobes, make vitamin K2, strengthens bones, ecoli - used in most biology labs, used to make proteins, used as model organism, can cause
salmonella
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Causes S
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 Archaea carious cell wall types but no peptidogylcan, can be gram positive or negative,
resistant to b-lactam antibiotics, very abundant and diverse
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2
phylogenetic groups - crenarchaeota - most sulphur metabolising thermophiles, Euryarchaeota
- methanogens, thermophiles, s-dependant thermophiles
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Autotrophic H2 and CO2 only substrates for growth and methane production
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 Eukarya - Saccharomyces cerevisiae - 'budding yeast' - life cycle in picture

Week 6 - Controlling Microbes/Prevention of bacterial Disease
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Aseptic techniques - Sterile solutions, plastics, glassware, environment
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Appropriate protective clothing
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In industry - hospitals prevent spread of infection, food industry - prevebt food spoilage and disease, pharmaceutical
industry - avoid contamination of medical supplies
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g
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Biological safety class 2 – may cause disease, treatable and unlikely to
spread to others (e
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Salmonella)
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Biological safety class 4 –
serious disease high risk to workers and public (frequently viruses - Ebola), needs protective
clothing
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Cell death - Micro-organisms not killed instantly - population death usually occurs
exponentially, tail end of slightly more resistant cells
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Decimal reduction time - time required to kill 90% of microorganisms or spores in a
sample at a specific temperature
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Conditions influencing effectiveness of antimicrobial agent activity - population size - larger
popn longer required, population composition - microorganisms differ markedly in their
sensitivity to antimicrobial agents, spores, concentration of antimicrobial agent - although
relationship is not linear, duration of exposure - temperature, higher temperatures usually
increase amount of killing, local environment (many factors) e
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pH, micro-organism more
readily killed at low pH - viscosity and concentration of organic matter e
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Heat - moist heat (autoclaves)- most effective, effective against all types of microorganisms,
hot water penetrates cells, degrades nucleic acids, denatures proteins, and disrupts
membranes
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Moist heat - used to kill endospores efficiently
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Steam enters top chamber and
forces air out through bottom vent
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Pasteurization controlled heating at temperatures, below boiling, preserves food
without destroying flavour/texture, reduces total microbial population (monitor
Mycobacterium tuberculosis and Coxiella burnetii – very heat resistant bacteria), flash
pasteurization (high temperature short-time – HTST), 72°C for 15 seconds then rapid cooling,
Now routine milk treatment, ultrahigh-temperature (UHT) sterilization, 140 to 150°C for 1 to 3
seconds, UHT milk, cream etc
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g Listeria – food poisoning) Freezing some microorganisms killed by ice crystal disruption of cell membranes, NOT a method of
killing -80°C used for storage of bacteria (addition of glycerol prevents formation of ice
crystals)
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Ionizing radiation (gamma
radiation – cobalt 60) - penetrates deep into objects, destroys bacterial endospores, used for
sterilization and pasteurization of antibiotics, hormones, sutures, plastic disposable supplies,
and food
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Disinfection - killing, inhibition of vegetative cells -pathogenic organisms, generally on

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inanimate objects, disinfectants - agents, usually chemical, used for disinfection
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Antiseptics - chemical agents that
kill or inhibit growth of microorganisms when applied to tissue
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Lister 1867, surgery, commonly used as laboratory and
hospital disinfectants, denatures proteins, disrupts cell membranes, tuberculocidal, effective
in presence of organic material, and long lasting, disagreeable odor, skin irritation, Triclosan
used in hand sanitizers, bactericidal
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Alcohols - bactericidal, fungicidal, not sporicidal
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70% more effective than 95% ethanol
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g
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Chlorine - disinfects water supplies, used in food industry
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Detergents (quaternary
ammonium compounds) – solubilise membranes, denature proteins, no effect on spores
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Prevention of infection:
Dirty drinking water and poor sanitation - leads to diarrhoeal disease - The cause of this
epidemic was dilapidated and broken sanitation and water infrastructure, much of which is
still in the same state as a year ago, so the coming rainy season is likely to facilitate the spread
of the disease
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Clean water drastically reduces the incidence of water-borne diseases such as
typhoid fever, diarrhoea etc
Decrease spread - e
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Plague doctor - wore masks with slits for nose which filtered through
aromatic herb, stick to avoid contact with flea infected clothing, cloth sealed to mask
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Avoid infection and disease - consider reservoir and
mode of transmission and then remove
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Bactericidal antibiotics – kill bacteria, Bacteriostatic antibiotics –
stop growth of bacteria,
E
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Peniciilin G - action on cell wall - Inhibits cell wall transpeptidases, Inhibits cross-linking
and peptidoglycan synthesis, Loss of integrity of cell wall, Distortion of wall, weak spots, Rapid
cell lysis, works by transpeptidase reaction - Transpeptidase attacks peptide bond in D-ala-Dala substrate, Release of terminal D-ala
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Penicillin looks like D-ala-Dala substrate, Transpeptidase binds b-lactam ring instead of D-alaD-ala
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5>10 min)
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All bacteria have peptidoglycan - access can be a problem - Gram -ve bacteria resistant to
Penicillin G (benzyl penicillin), outer membrane excludes many antibiotics, hydrated molecules
must pass through porins (~ 700 D cut off) ® modified penicillins eg ampicillin is more effective
against G-ve bacteria
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Resistant to B-lactamase, enhance stability in acid, taken orally
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Protein synthesis; 30S and 50S
ribosomal subunits (streptomycin - aminoglycosides, tetracyclines, chloramphenicol)
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Characteristics of useful antibiotics - selective toxicity - ability of drug to kill or inhibit
pathogen while damaging host as little as possible
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Toxic dose - drug level at which drug becomes too toxic for patient (i
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,
produces side effects) Therapeutic index - ratio of toxic dose to therapeutic dose
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Problems with bacterial resistance to antibiotics - resistance mechanisms -Alteration
in target protein (Methicillin resistant S
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Antibiotic altering enzyme, Efflux pump (tetracycline resistance)
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Immunisation - protection against disease - Passive immunisation –(inject infected patient
with protective antibody ) - tetanus, Active immunisation/ vaccination (induce a protective
immune response with appropriate antigen, prophylactic) – most vaccines
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Attenuated bacteria (lost virulence
attributes), repeated subculture, often at raised growth temp - undefined mutations, resulting
in reduced virulence eg BCG vaccine (Mycobacterium bovis subcultured) defined mutations
are now required for new vaccines
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Week 7 - Industrial applications for microbes/Fungi
How we exploit microbes:
 Microbes in food:
o Mushrooms - naturally occurring fungi
o Fusarium venenatum is a microfungus with a high protein content
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Grown
in continually oxygenated water in large, sterile fermentation tanks
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Used to make marmite
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 Fermentation - conversion of carbohydrates into alcohols or acids
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o Used in plant products - soy beans to soy sauce, olives, coffee
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Cerevisiae fermentation in
bread making produces CO2, Ethanol and imparts flavour
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Also relies on
growth on bacteria to provide flavour - propionbacterium
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 Secondary metabolism - microorganisms can alter metabolism when placed under stressful
conditions - usually growth/nutrient limitation, can be particular phases of growth cycle,
thought to be an attempt to promote survival/eliminate competition
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o Overproduction by regulatory mutants - mutants lack ability to control build up of biochemical
intermediates
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 Biopolymers - microbially produced polymers, usually polysaccharides, used in food industry to
modify texture, used in pharmaceutical industry as gelling agents
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 Biocontrol - uses of bacteria or their products as bioinsecticides
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Is a gram-positive, spore forming soil bacterium
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Doesn't accumulate in soil or non-target species, readily lost
from environment
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Recombinant products - genes from one organism can be inserted into different organisms
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g
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coli, human insulin produces E
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Bioremediation - Use of bacteria, fungi or enzymes to return an environment altered by
contaminant to its original state
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Bioaugmentation - microorganisms that can help clean up a particular contaminant are added
to contaminated soil or water
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Intrinsic bioremediation - occurs naturally in contaminated soil or water
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Benefits of bioremediation - cost effective, uses natural processes, treat widely dispersed
contaminants, minimised environmental disturbance, rapidly mineralise contaminants and
eliminate need for disposal
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Sewage treatment - microorganisms metabolise solid waste, microbes oxidise the nutrients
and in the process energy, in the form of heat and chemicals are produced
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Fungi:
 Fungi or the Eumycota are a kingdom of eukaryotic organisms, Were once classed as primitive
plants, Lack chlorophyll, Now are considered as unique organisms that differ from animals and
plants in their nutritional mode, growth and reproduction
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Acquire nutrients by
absorption, Fungi digest food outside the body by secreting hydrolytic enzymes into their food
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Very important bio-degraders
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The partner
supplies food to the fungus, The fungus provides the partner with a protective environment
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 Structure - fungal body is called a thallus, can be a range of sizes
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Fungal cells can be divided into yeasts and moulds
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Nucleus – Fungi usually have haploid genome,
Plasma membrane – ergosterol; chitin and glucan synthase proteins
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Cytoskeleton – microtubules, microfilaments, intermediate filament
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Yeast cells are egg shaped and contain
most of the eukaryotic cell organelles, Larger than bacteria - around 3um, No flagella
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Lots of
hyphae intertwine to form mycelium
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Septa act as structural support
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High density of vesicles in hyphal tip, Vesicles produced from Golgi apparatus
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Spitzenkorper - a vesicle rich body that determines direction
of hyphal growth
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Fungal reproduction - see picture
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Spores important for dispersal
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e
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This subsequently
undergoes meiosis to yield spores
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8 subdivisions now recognised:
Chytridiomycota - Simplest of the fungi, Unicellular or mycelial thalli
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amphibians chytridiomycosis, Asexual and sexual reproduction
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g
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Some of industrial importance, Foods (e
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sufu, tempeh), antiseptics and
other drugs, meat tenderizers, and food colouring
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Examples of ascomycetes - Blue/green food spoilage moulds-Plant disease: Powdery mildews;
Dutch elm disease; Ergot - Truffles
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g S
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Human
pathogens
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, asci) containing
haploid ascospores, Many exists in filamentous or yeast forms
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Basidiomycota - club fungi, during sexual reproduction form basidia - most common are
mushroms, jelly fungim birds nest and bracket - fungal foray
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These undergo mitosis to
eventually form the basidiospores which are released from the basidium into the
environment
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Glomeromycota - 230 described species, only asexual reproduction, no sexual stage, Mainly
arbuscular mycorrhizal fungi, form intracellular associations within roots of almost all
herbaceous plants and tropical trees, Colonise plant roots, increase their absorption capacity
and protect roots
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Microsporidia - Often considered protists, obligate intracellular parasites that infect insects,
fish, and humans, Infected humans often immunocompromised, Symptoms include diarrhoea,
pneumonia, encephalitis and nephritis
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Advantages of fungi - food and drink, Very important biodegraders, Antibiotics, anticholesterol statins, cyclosporin (immune suppression), ergot alkaloids (migraines)Biopesticides
(e
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nematodes and insects), Increase crop productivity, Bioremediation, Biofuels –
mycodiesel, ethanol, research - Model eukaryotic organisms
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 Virus - microscopic organism consisting mainly of nucleic acid in a protein coat, multiplying
only in living cells and often causing disease
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 Smallpox - spread through Europe and to Americans, killed many, variolation - scratching scab
material from someone with smallpox into skin o inoculated people
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 1911 - Rous sarcoma virus - responsible for classic first cell transmission of solid tumour, in
chickens
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 1935 - Tobacco mosaic virus
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1952 - hershey0chase experiment, showed that DNA and not protein of T2bacteriophage
entered the bacterial cell
...

 1995 - Multiple drug treatment for AIDS -protease inhibitors and reverse transcriptase
...

Virus structure:
 Protein coat on the outside, membrane envelope including free DNA and RNA
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Study of viruses:
 Indirect - effect on host
 Direct - property of virus itself
o Plaque assays - make serial dilutions of virus
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After
attachment, overlay cells with semi solid media to restrict diffusion of virus particles
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o Rate zonal centrifugation -mixture of biomolecules including whole viruses, spun in a
centrifuge, fractionate gradient produced - heavier particles at bottom
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o Denaturation of TMV - Helix to disc to monomers
...

o PCR - polymerase chain reaction- heat DNA to mel strands, cool to allow primers to anneal to
target sequences, incubate to allow polymerase to extend primers, hear DNA to melt stands
again, cool to allow primers to anneal to target sequences and extend again
...

o Gel electrophoresis - add mixture of nucleic acids onto gel matrix and apply voltage, molecules
move through gel towards the anode and are separated by the gel matrix based on their size
...

o Protein expression systems - vector carrying foreign gene, new proteins produced and gene
expressed
...
Expression of
virus proteins can identify function and lead to virus assembly
...

By morphology - viruses in many shapes and sizes, viruses which look very similar can cause
very different disease and may be genetically very different
...

By serology - ability of antibodies to recognise the virus, viruses are very different in
antigenicity due to difference in their genetic sequence
...
Viruses which are closely
related may not be recognised by the same antibodies
...


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o

o

o

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By genetic material alone - whether single stranded, double stranded or circular DNA, or if
RNA + or -ve, segmented, or double stranded segmented
...
Also, not all RNA is message sense or used for
translation
...
Translation - mRNA’s get translated into proteins by the
ribosome
...
g
...
Encapsidation - These new copies of the viral genome get put into
virus particles and make mature virions
...
ssDNA viruses include organisms
infecting: bacteria, Mammals, Birds and plants
...
Replication requires
the conversion of the ssDNA into dsDNA first, Then replication follows the same rules as the
central dogma
...
Genome is
composed of two strands of complementary RNA annealed together as with the dsDNA
viruses
...
g
...
Genomes cannot function as mRNA
for translation
...
This family of
viruses use an enzyme not found in any host cells known as RNA-dependant RNA polymerase
(RdRP) for transcription and replication
...


Ss positive sense RNA Viruses - Viral genome consists of ssRNA - the same polarity as mRNA
...
Some genomes can be
translated directly by the ribosome into proteins, Majority of viruses replication in the
cytoplasm of the infected cell, Use virus encoded RNA dependant RNA polymerases (RdRp) to
replicate their genome
...
The viral proteins then work with specific cellular proteins to copy

o

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o

the viral genome into an dsRNA copy of itself
...
Genome consists of single stranded
RNA molecules of opposite polarity to mRNA – therefore they cannot be directly translated
into protein, Before translation can occur the RNA must be copied into positive sense
...
e
...
g
...
The enzyme Reverse Transcriptase or ‘RT’ is unique to viruses and has the
ability to copy RNA back into DNA, reversing the flow of genetic information in the Central
Dogma, Virus also carries 2 copies of the RT enzyme with it to allow reverse transcription to
occur as soon as it enters the host cell
...
Retroviruses have the ability to integrate their genome into the
host chromosomal DNA – then every time the cell divides each progeny cell gets a copy of the
viral genome, Our DNA genome is approximately 8% retroviral DNA
...
Unlike the Retroviruses, this
occurs inside the virus particle on maturation
...


Week 10 - Vaccines and Drugs




Treating virus infections - Virus replication requires living cells so the eradication of viruses is
obviously difficult –the virus, not the cell, must be targeted
...
Cellular response - kills infected cells but leaves the problem of viruscell infection
...
In most cases response to
internal proteins has little effect on humoral immunity to infection
...
A successful vaccine works by blocking uptake of the virus so must
generate Abs that bind to the receptor binding protein
...
Killed –inactivated virus or parts of the virus that do not infect
but still generate an antibody response
...

How a vaccine works - Neutralization of the virus may result from (in order of importance)
...
Aggregation of
virus by polyvalent antibody
...

3 major sites of viral replication - Mucosal surfaces of respiratory tract and GI tract
...
Infection at mucosal surfaces
followed by spread systemically via blood and/or neurones to target organs:picorna; measles;
mumps; HSV; varicella; hepatitis A and B
...

Antiviral drugs - In reality most antiviral drugs target viral enzymes
...
Drug resistance
- High mutation rate and large progeny numbers make viral evolution in response to selective
pressure very fast
...
Relative
fitness of drug resistant virus vs wild type virus in vivo can influence whether drug resistant
viruses proliferate
...




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