L29 - Bacterial Growth And Replication Flashcards
Where do bacteria form biofilms?
On any surface where there is moisture
What do bacteria attach to? What happens afterwards?
- bacteria attach to surface
- grow and become enveloped in extracellular matrix (ECM) (polysacch, proteins and DNA)
What are the stages of biofilm formation?
- inital attachment
- irreversible attachment
- maturation 1
- maturation 2
- dispersal
What happens during initial attachment?
Individual bacteria attach weakly to a surface
What happens during irreversible attachment?
Attachment becomes irreversible using fimbrae and pilli
What do bacteria do after attachment?
Multiply, attract other microbes to attach
What happens during maturation 1?
- bacteria secrete a sticky, protective ECM
- contiue to join and multiply
What happens during maturation 2?
Biofilm grows in size and structure form large 3D colony
What happens during dispersal?
Sections of the biofilm break off - cells can go and colonise new areas
What do biofilms protect against?
- phagocytosis
- antibiotics
- disinfectants
How are biofilms a huge problem in healthcare?
Growth on medical devices
What are examples of biofilm in healthcare?
- dental plaque
- urinary catheter
- heart valve - endocarditis, enterococcus sp
- lung tissue - cf, pseudomonas aeruginosa
How do nutrients affect growth of biofilm?
Nutrients required for cellular biosynthesis and energy generation
- macroelements - C, H, O, N, water, S, P, K, Ca, Mg, Fe
- trace elements - Mn, Co, Mo, Ni, Cu vits, gf
How does iron affect growth of biofilm?
- used for energy generation
- iron in body not available
- withing mammalian cells (90% stored in ferritin/haem group, 8% stored elseqhere)
- outside mammalian cell (1-2% attached to transporters
- non complexed iron exists as Fe3+, insoluble
What is the bacterial iron transport system?
Siderophores
What are siderophores?
- low MW compounds with high affinity for iron
- produced and exported from some bacteria when iron low
- bind iron and allow uptake into the cell
- -remove iron complexed with transferrin, uptake
How does oxygen affect growth of biofilm?
- aerobe (with O) - microaerophile - can grow in low conc
- anaerobe (w/o O) - obligate anaerobe - cannot grow in O, faculative anaerobe - can grow in O if available
How does temperature affect growth of biofilm?
- psychrophiles - -40C to 20C, optimum <15C // lysteria monocytogene
- thermophiles - 45 to 100C // thermophilus aquaticus
- mesophiles - 20 to 40C, organisms of med and pharm importance
How does pH affact growth of biofilm?
Most organisms or medical importance are neutrophiles - pH6.5-7.5
What are the different ways of growing bacterial culture?
- suspension
- colony
What are bacterial cultures in suspension? Why?
- Bacteria grown in complex liquid media as batch culture
- to determine growth rate/ effect of antimicrobial agents
What are bacterial cultures in colonies ilke? Why?
- bacteria grown on complex media solidified with agar
- to obtain pure culture/ viable count/ assess diversity/ aid identification
What is the bacterial replication?
Binary fission - one bacteria cell grows and divides to 2 identical daughter cells
How does binary fission happen?
- cell elongage ~ double
- cell copies chromosome
- septum beings to form
- 2 copies of chromosome pulled apart
- septum formation continues until 2 cells formed
How do bacteria multiply?
Doubling as fast conditions will allow - exponential growth
What is generation time?
Time taken for bacteria to divide, double
What is rate of cell division determined by?
- time required for DNA replication
- conditions
What are the differnent phases?
- exponential phase
- stationary phase
- death phase
What is exponential phase?
- the cells are behaving in a constant predictable
- generation time is constant
- ideal phse to use the bacteria for research
What is stationary phase?
- population running out of resources
- no increase/decrease in cell numbers
- some are dividing, some are dying = unpredictably
What is death phase?
- decline in cell numbers
- some are persister - dont die - viable but non culturable cells
Where can generation time be read from?
Exponential phase of growth curve
What is generation time (g) during exponential growth?
NT = N0 x 2^n
N0 - cells initially present
NT - cells at T
n - number of generations
How do you calculate generation time?
Generation time = time/no of gens
How do you do a direct measurement of bacterial number - viable count?
- dilute sample of bacteria
- spread on agar plate
- incubate overnight @ 37C
- count colonies
- VC expressed as Colony forming units/ml CFU/ml
- best way to measure no. actively dividing cells
- exc clumps/chains of cells
- conditions must be suitable for growth
- requires overnight culture
How do you do an indirect measurement of bacterial number - optical censity (OD)?
- OD increases with increasing cell no over time
- cell no directly related to OD
- read off cell no from standard curve of OD vs cell number
- cells must be in exponential phase for OD to represent no dividing cells
