microbial growth

Question 1

outline the process of the streak plate technique

Answer 1

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

Question 2

what are the different types of culture medium

Answer 2

1) solid = contains agar
2) liquid = doesnt contain agar
3) semisolid= contains soft agar
all mediums contain water which is essential for growth

Question 3

what is a minimal medium

Answer 3

a medium with minimal nutrients which are essential for growth

Question 4

what is a complex/undefined/basal medium

Answer 4

a medium with water, a carbon source, salts and a source of amino acids of which the exact composition is unknown

Question 5

what is a transport media

Answer 5

a medium for transport which doesnt promote growth as is lacking in carbon and nitrogen

Question 6

what is a defined media

Answer 6

a medium where proportions of components are known such as amino acids

Question 7

what is a differential media

Answer 7

a medium which is used to distinguish one microbe from another

Question 8

what is a selective media

Answer 8

a meidum which is used to grow selective microbes

Question 9

outline binary fission

Answer 9

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

Question 10

define binary fission

Answer 10

an organism duplicates its genetic material, dividing into two parts, with each new organism receiving one copy of DNA

Question 11

outline how microbial cells divide

Answer 11

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

Question 12

what mechanisms does the Z ring use to find the middle of the cell

Answer 12

1) Min system
2) nucleoid occlusion

Question 13

what is the min system

Answer 13

• 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

Question 14

what is the role of MinD, C and E

Answer 14

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

Question 15

what is the main role of the min system

Answer 15

to protetc the poles of the cell from FtsZ polyermization therefore from the contricting ring which seperates cells

Question 16

what is nucleoid occulusion

Answer 16

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

Question 17

what are the phases of microbial growth

Answer 17

1) lag phase
2) exponential phase
3) stationary phase

Question 18

what is the lag phase

Answer 18

the time that the cell needs to adapt to the new growth conditions and resume growth

Question 19

what is the exponential phase

Answer 19

once growth resumes at a steady rate the cells growth and divide exponentially

Question 20

what is the stationary phase

Answer 20

the total number of cell remains the same but the ration of viable to dead cells change

Question 21

why can microbes not grow continuously

Answer 21

1) run out of food
2) produce toxic by-products
3) produce grwoth inhibition products

Question 22

what is balanced microbial growth and how is it maintained

Answer 22

• when cells do not exceed a certain cell density
• to maintain a balanced growth dilute cultures after a certain number of generations

Question 23

what is used in a lab to maintain balanced growth

Answer 23

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

Question 24

how can microbial growth be measured

Answer 24

1) turbidity
2) colony count
3) cell viability assay
4) direct counting
5) flow cytometer

Question 25

how can turbidity of a culture be measured

Answer 25

• 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

Question 26

how does colony count work to work out microbial density

Answer 26

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

Question 27

what is a cell viability assay

Answer 27

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

Question 28

how can you measure cell number with a microscope

Answer 28

• use a counting chamber (hemocytometer)
• add cell suspension and view under microscope

Question 29

how can you count cells with a flow cytometer

Answer 29

• 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

Question 30

what must happpen to cells before entering the flow cytometer

Answer 30

labelled to collect into about genetic or biochemical composition

Question 31

what affect does temp have on growth

Answer 31

high
- proteins denature and chemical reactions slow or stop
low
- protein activity slows down but doesnt stop

Question 32

how do microbes cope with high temps

Answer 32

thermophiles have large amounts of chaperone proteins ( heat shock proteinc which stabalise new proteins or help refold damaged ones)

Question 33

how do microbes cope with high pressures

Answer 33

1) cell envelipe= extent of permability to water means they can balance pressure
- more permable membranes
- negative DNA supercoil so DNA more stable

Question 34

what stresses does high osmotic pressure put on cells

Answer 34

1) decreases water for cells
2) reduces cell turgor by decreasing differences in osmotic pressure

Question 35

what do cells do to maintain osmotic pressure

Answer 35

1) accumulate molar conc of pottasium and chloride
2) accumulatin of organic osmotic solutes

Question 36

how do cells cope with extreme PH

Answer 36

pump protons in and out cells maintaining constant internal PH