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Why do we use mircoscopes?
microbes are too small to see with the naked eye
- low = 10x
- high dry= 40x
- immersion = 100x
ocular x objective
example 10x40= 400x high dry
How to use a microscope? (tips for success)
what objective you should start with?
- adjust light intensity for clarity
- dont turn the light all the way
- reduce intensity to moderate
- clean them
What is a we mount ? And why we use this technique? And Advantages?
- wet mount a living suspension of the organism
- organisma are alive
- see motility
- cellular arrangements
- see the size and shap
what is hay infusion?
hay infusion is a mixture of lake and and grass or hay. the mixture has O2 pumped into it facilitates the growth of microbes in the lake.
Wet mount Vs smear prep?
- wet mount
- see motility, cell arrangement, organism is alive
- Smear prep
- dead organism but allow to see a specific structure
- microorganism food
- contain all chemicals compounds necesary to support growth
- different organisms have different requirements
Types of media and what each grows?
- two forms
- broth liquid
- agar solid
- complex =rich media
- for isolation of
- tryptic soy agar
- brain heart infusion
- potato dextose agar
Chemically defined media?
- exact chemicals and amount is known
- used to isolate specific groups
Living tissue media?
- made of living organism
- growing host specific pahogen unless live on bacteria
How to calculate amount of media powder given amount of water?
Definition of Sterilization?
the complete destruction of viable organisms in a given environment to prevent contamination.
Type of sterilization?
- media and metal objects
- steam sterilization
Type of sterilization?
type of sterilization?
- heat chemicals, antibiotics
- gasous- anything that will melt
- plastic, petri dishes
What is streak plating and its purpose?
- a petri plate containing some type of sterile nutrient agar( tsa,BHI,
- purpose to seperate the bacteria in mixed culture so the individual cells can be isolated
when should you flamethe inocluating loop?
Between the streaking of each sector
What is a dillution plating and whats its purpose?
- a method used to estimate #s of bacteria in a sample
- purpose is to
How to perform/calculate several dilutions?
colony forming units
A disease or two caused by old milk?
pour plate method?
second method of obtaining isolated colonies by dilution
Shape/form of colonies?
Margin of colonies?
- diplo= pairs
- tetras= groups of four
- strep= chains of three or more
- facultative- grow without O2
- Aerobic- requires O2 to grow
- Anerobic- O2 is toxic ,requires no oxygen environment to live
- microaerophilic- O2 is toxic at atmosphere concentrations only small amount
What is Smear Prep?
a procedure used to prepare a bacterial sample for staining in which the bacteria is heat fixed to a slide
What is it used for?
staining and increasing contrast
Disadvantages of smear prep?
motility no longer be studued ditorts the cells apperance
- if liquid broth place a loopfull of broth directly on to the slide
- if isolated first drop of water on to slide and which a small amount of bacteria is dispersed using a loop
Why is the gram stain useful?
What are the 4 steps of the gram staining procedure?
what are the difference in the outer protection between gram + and gram -?
What do these cells liik like in the end?
Why is a capsul useful for bacteria?
- act as a barrier between the cells
- reserve cabroydrate
- aid the cell in adhering
what is a capsule?
a viscous substance which accumulates and coats the cell and becomes the outermost layer made up of polyssacharides(few Proteins)
What is a negative stain?
steaming the background leaving the capsule unstained since it cant be stained.
What is Schaeffer-fulton?
stain capable to distiquish between spores and vegetative cells found primarily in the clostridium and bacillus genera
What is an endospore?
its a dorment form of the bacterium that allows it to servive lean envrionmental conditions./ spores form within the cell( vegetative cell)
What bacterium form endospore?
- the clostridum
- bacillus genera
Why do we steam?
- its a primary stain in malachite green is forced into the spore coat.
- To distiquish between the spore and the vegetative cell
What is an acid fact bacterium?
- stain important for differential for identifying bacteria in the gendo mycobacterium
- large amounts of lipids and waxes in their cell wall makes them inpermeable and resistant
- alos known as the acid fast stain
- allows to identify bacteria with large amounts od lipids and waxes in their cell walls
- most commoly in the genus mycobcterium
- will appear red-non blue
Why special measures to dye them?
due to that they are impermeable and resistant to many chemicals
What special measures?
what bacteria are acid fast?
What is a selective Media?
- has a selection agent or agents added to it prohibits the growth of some organisms
- allows the organism of interact to grow while inhibits the growth of others.
What is differential Media?
- growth of organisms not inhibited
- contains substances that allow different groups grow differently than others when grown on the same media
Mannitol salt agar(MSA)
- selects gram + bacteria growth
- highest level of salts
- mannitol and phenol red
- when fermented turn yellow
- non mannitol fermenters remain pink
- crystal violet bile salts
- inhibits growth of gram+
- contains lactos and neutral red ph indicator
- turn to red colonies
- non lactos colonies do not change
Kliglers Iron Agar (KIA)
- determinator of varbohydrate fermentation patterns and gas production
- determines gram - rod Enterobactericeae
KIA agar slant
inoculated with a needle( not Loop) stab then streak slant
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