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1.19.1.14 - Growth of bacteria - Coggle Diagram
1.19.1.14 - Growth of bacteria
Growth of bacteria
Growth is by binary fission
Colonies form on plates as piles of bacteria
Some bacteria are very motile they do not form colonies as they spread too thinly
Bacteria have generation times also known as doubling times= the length of time required for a single bacterial cell to divide into 2 daughter cells.
If bacteria double every 20-30 mins, a colony can form in about 12-24 hours, but it can also take months for slower bacteria to form colonies
Growing bacteria
Sterile media
Aseptic techniques
Flaming loops
Measuring bacterial growth
Direct counting by microsope- but cannot distinguish from non-viable cells
Colny counting - serial dilutions spread on surface of plate, mix bacteria with agar and pour plate, miles-misra method (serial dilutions on a spot dish), membrane filtration
Absorbance in liquid culture- measurement of absorbance of liquid culture at 600nm
Miles and Misra method
The sample is diluted in log steps - e.g a 10 fold, 100 fold, 100 fold dilution etc.
A set volume is plated
As the dilution gets lower you get to the point where there are few enough bacteria that when the liquid dries into the media they stay separate and then form separate colonies
You count the colonies from 50 microlitres of the 10^-4 dilution and then calculate up to how many would be in 1 ml.
The numbers are expressed at colony-forming units per ml. They are a viable count of the bacteria in the original sample.
Absorbance
increase in optical density over time
Generation - time for one full round of division
Growth rate - how much doubling per hour
Can measure optical density
Phases of growth in liquid culture
Lag phase- bacteria adjust to ne medium and start to metabolise andgrow
Exponential phase - bacteria grow at their best this is the phase where growth rate is determined
Stationary phase- bacteria reaches a limit on resources in the middle and can not exceed a maximum population ( growth rate= death rate)
Decline/death phase - Medium is exhausted of a component needed for growth0 new bacteria can not grow and bacteria are still dying,
Studies:
Physiology studies are carried out in liquid media as this allows the measurement of growth-phases and rates.
Comparison of log vs non-Log representation. Log illustrates the exponential nature of bacterial growth and allows the distinction of phases.
Bacterial respiration
A terminal electron acceptor is a compound that receives or accepts an electron at the end of the respiratory electron transport
Humans terminal electron acceptor is oxygen hence we need oxygen to live
Bacteria adapted to a range of terminal electron acceptors and hence can colonise a lot of ecological niches
Some examples of electron acceptors (the reduced product) are H20 and CO2 for oxygen, and NO2-, N2O or N2 for nitrate (NO3-)
What are the respiration types:
Aerobe - require oxygen to grow
Microaerophile - can cope with low amounts of oxygen
Facultative anaerobe - can grow with and without oxygen
Anaerobe - cannot grow in presence of oxygen
This is clinically relevant because - a deep wound will be colonised by different bacteria based on oxygen gradient
Impact of pH
pH standard for most media is buffered at pH7
When bacteria grow, they may produce acidic metabolites that over time will inhibit their growth - some media are buffered to compensate for that.
Most pathogens are neutrophile, even bacteria that survive in stomach. That bacterium balances the pH in the close environment
The trye acidophiles and alkaliphiles are found in extreme environments are not pathogenic - those environments and are not pathogenic - those environments existed before the first higher organisms developed.
Impact of temperature
The bacteria that infect of colonise animals are mesophilic
Bacteria adapted to some animals have sightly different optima
For example birds average temperature is 42 degrees and Campylobacter which is commonly found in birds grows best at 42 degrees. It has co-evolved for this environment. ---> Flexibility within a range
That said it can also grow ok mammalian body temperatures and cause disease.
Storage in fridge slows bacteria growth but doesn't stop it completely. Milk in a fridge spoils slowly but the organisms in the milk can still grow very slowly. If you had the milk at 37 degrees, it would spoil very quickly . Allows pathogens to survive and replicate outside/between hosts.
What is the impact osmolarity (water potential on growth
The membrane of bacteria is semi-permeable
Bacteria need to balance their water potential to excessive movement of water in out or of the cell
Bacteria accumulate or lose solutes to balance the water potential across the membrane
Bacteria prefer to maintain a slight positive pressure an inflow of water which is resisted by the cell wall.
Nutrition
Bacteria acquire nutrition from the immediate environment- that environment may be a wound or intestine etc.
Bacteria can be:
Fastidious - require specific supplements
Non-fastidious - grow from basic chemicals
Most veterinary relevant organisms pathogens are chemoheterotrophs - they use organic chemicals as sources of energy
Clinical relevance
Part of the host defence is to limit access to certain nutrients - reduced bacterial growth. Bacteria can compete for compounds so one organism can potentially block colonisation/infection by another
Types of growth media
Minimal media: A basic salts based growth media often with a single carbon source
Nutrient media - A medium with all the basic requirements for growth but without specific supplements.
Enriched media: nutrient media with additional organism specific supplements
Selective media: A nutrient medium with the basic requirements for growth and added supplements that allows one bacterial type to grow more than another
Indicator medium- A medium with indicators which react to specific bacteria
Selective indicator media: A combination of selective and indicator media
Transport medium: A medium designed to protect the organisms in the sample
Once there is a pure culture:
Once cultured, tests can be done on the bacterial isolates:
speciation through: molecular tests, biochemical tests
antibiotic sensitivity
Typing to determine epidemiology of infections
For long term epidemiology studies, we need to preserve bacteria. This is done by:
Sub culturing (labour intensive and bacteria can change)
Freeze drying (Lyophilisation
Freezing with cryoprotection (DMS or Glycerol)
How to get rid of unwanted growth of bacteria:
Sterilisation: refers to a process that eliminates or kills all forms of life. Main methods to do this are heat, chemical and irradiations
Washing: Will not sterilise unless you can use something that kills the microorganisms. But can remove and reduce risk of contamination or spread of infection
Disinfection: Do not assume all disinfectants kill all pathogens. Disinfectants may not be compatible with all uses.
Disinfection:
There are a range of different chemicals used to disinfect.
They damage in a general manner by reacting with different targets such as proteins or nucleic acids - most effects are to denature cell components- can damage or affect cells. Prions are very tolerant as already denatured so not fully effective or need extreme compounds
Some are washes that can be used on patients
Others are caustic or toxic
Selection must account for a lot of factors: Non tainting for food industry, effectiveness in environment they will be used in, effectiveness against the organism you want to target.