microbial growth Flashcards
outline the process of the streak plate technique
1) flame inoculation loop
2) dip into bacterial suspension
3) carefully wipe the loop over the plate covered in solid growth medium
4) either lift the loop every time you change direction or use a single touch spread where it is not lifted
5) the idea is to have colonies spread out so you can see individual colonies
what are the different types of culture medium
1) solid = contains agar
2) liquid = doesnt contain agar
3) semisolid= contains soft agar
all mediums contain water which is essential for growth
what is a minimal medium
a medium with minimal nutrients which are essential for growth
what is a complex/undefined/basal medium
a medium with water, a carbon source, salts and a source of amino acids of which the exact composition is unknown
what is a transport media
a medium for transport which doesnt promote growth as is lacking in carbon and nitrogen
what is a defined media
a medium where proportions of components are known such as amino acids
what is a differential media
a medium which is used to distinguish one microbe from another
what is a selective media
a meidum which is used to grow selective microbes
outline binary fission
1 ) a cell synthesises its constituents such as proteins until it reaches a point where it has doubled its mass
2) the cell divides into two cells
3) the cycle starts again
define binary fission
an organism duplicates its genetic material, dividing into two parts, with each new organism receiving one copy of DNA
outline how microbial cells divide
1) prokaryotes make a protein (FtsZ) which forms a constricting ring in the middle of the cell = the Z ring
2) the assembly of the Z ring requires polyermerization of FtsZ in short filaments and the association with the membrane
what mechanisms does the Z ring use to find the middle of the cell
1) Min system
2) nucleoid occlusion
what is the min system
- consists of three proteins (minC, minD and minE) which move around the cell interior to prevent polyerization of FtsZ at the poles of the cell
what is the role of MinD, C and E
1) MinD polyermizes at one pole of the cell and binds to MinC
2) MinC acts as an FtsZ inhibitor preventing FtsZ polyermization (this can only occur when C is bound to D)
3) E forms a ring close to the pole preventing CD from polyermizing further into the middle
4) D dissolves in one pole and polyermizes in the opposite pole forming another CD complex
5) E forms another ring close to that pole
what is the main role of the min system
to protetc the poles of the cell from FtsZ polyermization therefore from the contricting ring which seperates cells
what is nucleoid occulusion
1) as chromosomes replicate in the middle of the cell
2) newly replicated regions migrate to the poles so the amount of DNA in the middle decreases
3) the NO protein inhibits unwanted cell division where the nucleoid is localised
4) only when newly replicated chromosomes are about to spily the FtsZ protein find the space to form the Z ring which triggers cell division
what are the phases of microbial growth
1) lag phase
2) exponential phase
3) stationary phase
what is the lag phase
the time that the cell needs to adapt to the new growth conditions and resume growth
what is the exponential phase
once growth resumes at a steady rate the cells growth and divide exponentially
what is the stationary phase
the total number of cell remains the same but the ration of viable to dead cells change
why can microbes not grow continuously
1) run out of food
2) produce toxic by-products
3) produce grwoth inhibition products
what is balanced microbial growth and how is it maintained
- when cells do not exceed a certain cell density
- to maintain a balanced growth dilute cultures after a certain number of generations
what is used in a lab to maintain balanced growth
a chemostat
- a bioreactor to which fresh medium is continuously added
- fresh medium from a reservoir is fed to a culture at a constant rate
- the volume of culture remains constant by removing excess overflow
- rate of addition of fresh medium determines the growth rate in the culture
how can microbial growth be measured
1) turbidity
2) colony count
3) cell viability assay
4) direct counting
5) flow cytometer
how can turbidity of a culture be measured
- measure the cell density using a spectrophotometer which passes light through the culture
- the absorbance or optical density is measred
bacterial cells scatter the light reducing the light detected
how does colony count work to work out microbial density
make serial dilutions of cultures to avoid having too many cells
spread a potion of each dilution onto agar ensuring single cells are seperated
incubate at a set temp and time
identify dilution playte with 25-250 colonies
count number of colonies
what is a cell viability assay
1) luminescence based assay for the determination of the viability of cells
2) uses quantification of ATP as an indicator of metabolically active cells
3) ATP changes luciferin to oxyluciferin and light
4) the luminescent signal i sproportional to ATP produced
how can you measure cell number with a microscope
- use a counting chamber (hemocytometer)
- add cell suspension and view under microscope
how can you count cells with a flow cytometer
- uses a laser beam to simultaneously count and analze many physical and chemcial characteristics of individual cells
- sample focuses such that one cell at a time passes through the laser beam
- eahc cell interrupts the light path scattering the light giving info about size and shape of cell
what must happpen to cells before entering the flow cytometer
labelled to collect into about genetic or biochemical composition
what affect does temp have on growth
high
- proteins denature and chemical reactions slow or stop
low
- protein activity slows down but doesnt stop
how do microbes cope with high temps
thermophiles have large amounts of chaperone proteins ( heat shock proteinc which stabalise new proteins or help refold damaged ones)
how do microbes cope with high pressures
1) cell envelipe= extent of permability to water means they can balance pressure
- more permable membranes
- negative DNA supercoil so DNA more stable
what stresses does high osmotic pressure put on cells
1) decreases water for cells
2) reduces cell turgor by decreasing differences in osmotic pressure
what do cells do to maintain osmotic pressure
1) accumulate molar conc of pottasium and chloride
2) accumulatin of organic osmotic solutes
how do cells cope with extreme PH
pump protons in and out cells maintaining constant internal PH
