Culturing microorganisms Flashcards
Why are microorganisms cultured?
Generate biomass of microorganisms
To manufacture compounds the microbes synthesise
Achieve max yield
What are primary metabolites?
Produced in processes essential for normal microbial functioning
What are secondary metabolites?
Produced in non-essential processes
What are bioreactors?
Used in large scale, commercial production of microbial cultures
Large fermentation tanks optimised for microbial growth.
What are the components of a bioreactor?
Metal/plastic tank = inouts/outputs for liquids + gases
**Paddles for mixing culture **-> ensure even distribution of food, oxygen + temp throughout
Probes = monitor pH, temp + dissolving gases
Ports = adding ingredients + removing products
Sterilisation system -> steam injection
Nutrient medium –> for microbial growth –> liquid (broth) or solid (agar)
How nutrient availability regulated inside a bioreactor?
Fresh medium circulated by paddles
Pop size increases –> nutrient demand exceed nutrient supply –> constant supply ensures microbes = nutrience they need
How is temperature regulated in a bioreactor?
Heating/cooling water jacket surrounds vessel
Too low = bacterial enzymes not work
Too high = bacterial enzymes denature
How is pH regulated in bioreactor?
Monitored by pH probe –> automatically adjust by adding acids/bases
Build up of CO2 = reduce pH –> inhibit enzyme activity –
> keep optimal pH allows microbial enzymes to function efficiently
How are oxygen levels regulated in bioreactor?
Sterile air pumped in
Pop size increases –> O2 demand exceed O2 supply –> aerobic respiring microbes require O2
How is contamination/waste regulated in bioreactor?
Steam sterilisation bvetween batches + removal of waste products
Unwanted microbial contamination creates competition from other microbes + build up of toxic waste = kill culture
How can contamination be prevented?
Aseptic techniques-> ensures nutirent medium = sterile + high yield of desired product made
What is batch fermentation?
Microbes grow in fixed volume in individual batches until nutrients depletes + waste accumulates
Empty + clean vessel b4 starting new batch
What is continuous fermentation?
Involves continuously supplying fresh nutirents + removing culture broth
Maintain growth of culture indefinintely
What is the microbial growth curves in batch cultures?
Closed system = growth pattern over time.
Lag phase
Log phase (exponential)
Stationary phase
Death phase
Describe the growth patterns:
Lag phase: Slow initial growth -> adapt to environment + produce essential enzymes
Log phase: rapid doubling of cell numbers occurs under ideal conditions –> growth rate at max
Stationary phase: growth rate plateaus -> nutrirents diminsih + waste accumulates –> cell growth = cell death
Death phase: Cell death rate exceeds cell growth rate due to resource limitation + build up of toxins
How to culture microbes in the lab?
- Sterlise all equipment b4 use (innoculating loop in bunsen burner)
- Dip sterilised wire in innoculating loop into a starter culture –> broth containing bacterial suspension
- Transfer microbes to Petri dish -> contain sterile nutrient medium (lightly zig zag loop across agar)
- Close plate + lightly tape –> not completely sealed (prevent growth of anaerobic microbes)
- Label plates with relevant info (type, date, conditions)
- Incubate plates upside down under required conditions
- Repeats steps 1-6 for control agar dish with no bacteria
- Assess microbial growth observing colony formation on agar.
What are the factors that may affect microbial growth?
Temperature: incubate duplicate plates at diff temperatures
pH: add buffer solutions to agar to maintain different ph levels
Nutrient availability: Prepare agar with varying nutrient concentrations
Antomicrobial substances: Add diff antimicrobial compounds to agar plates
How can you investigate factors which affect growth of microorganisms?
set up colonies in diff conditions of temp
set up serial dilutions of nutrients/pH at set temp
How to do serial dilution?
Number of bacteria/ml = number of colonies x dilution of sample
Work backwards
