Lab1 Flashcards
List the 4 basic microbiology techniques learnt in lab 1
Gram stainmicroscope usestreak platelawn plate
bacteria and fungi can grow in the lab if what conditions are provided for?
Suitable nutrientsenvironment favourable for growth
How have the fomulation of many types of culture media formed?
Species of bacteria and fungi differ in their requirements for suitable nutrients and favourable growth environment
what are the two main kinds of culture media?
- liquid2. solidified with agar
what is agar?
gelling agent derived from seaweed which melts at 98-100degreesC
What conditions must the culture medium be in prior to use?
It must be sterilised and remain sterile prior to use
How are most culture media sterilised?
by autoclaving
what is autoclaving?
Uses steam under pressure to generate sufficient moist heat to kill both vegetative and heat or chemical resistant forms of microorganisms
give examples of heat or chemical resistant microorganisms
bacterial endospores, protozoan cysts
What is the pressure commonly used for autoclaving?
15 pounds per square inch for 15 mins.this increases temperature of steam to 121 degrees C
What happens to inoculated material used in the lab before disposal?
they are autoclaved before disposal
what is the innoculum
microorganisms of interest (that you want to transfer onto the sterile medium)
What is innoculation
the transfer of microorganisms of interest onto the sterile medium
What kind of technique must be used to innoculate?
aseptic technique
why must aseptic technique be used for innoculation
prevents contamination by extraneous microorganisms
where may extraneous microorganisms be?
in the air, on lab equipment or surfaces or on hands of laboratory workers
Give another reason why aseptic technique is used
also helps to prevent contamination of the lab worker by the microorganims being handled
When does growth/division occur?
when a freshly inoculated culture is incubated under favourable conditionsthis is 37degreesC for most human pathogens
How is growth of bacteria indicated by in liquid media?
cloudiness (turbidity)
how is growth of bacteria indicated by in solid media?
bacteria and fungi generally for colonies
The appearance of _______ is important in identification
colonies (colour, shape etc)
Description of colonies is what kind of appearance?
Macroscopic appearance
what is microscopic appearance?
term used to describe the appearance of cells
give the importance of a gram stain
it is used to quickly begin the process of identifying the microorganism
why are bacteria stained?
They are almost transparentstaining increases their visibility under the light microscope
What must be prepared before bacteria can be stained?
a heat fixed smear of the bacteria
What does staining reveal?
The overall shape of the bacterial cells (cocci, bacilli, vibrios, sprirlla)the arrangement of cells (single cells, chains, pairs, clusters, tetrads)
What characteristics are used in the identification of bacteria?
Characteristics of cellular morphology: size, shape and arrangement
How do yeast cells differ from bacterial cells?
By size, yeast cells are eukaryotic, usually bigger
briefly describe how to prepare a heat fixed smear
- sterilise loop, and run slide over flame a few times to sterilise slide2. place loopful of sterile saline on centre of slide3. sterilise loop4. For solid media: remove bacterial colony and emulsify in saline using circular motion. 4. For broth culture: [dont need saline] invert broth a few times, take 2/3 loopfuls of broth onto slide.5. spread drop on the slide to size of 10c coin6. sterilise the loop 7. allow smear to air dry, then heat fix smear by running through flame a few times8. check slides are labelled
what is the purpose of heatfixing the smear?
to fix bacteria to slideto kill microorganisms without altering their structure too muchto help microorganism adhere to slide better and take up the slide
why is it necessary to flame sterilise inoculating loop both before and after the preparation of smear?
To eliminate contamination of slide preparations nad inoculum plate so that bacteria stained and seen under microscope is the bacteria of interest
Why is gram stain used extensively in bacteriology
it is valuable.it allows the differentiation of bacteria into two large groups based on differences in cel lwall composition
What are the two types of gram stained bacteria
Gram positivegram negative
What colour are gram positive organisms?
blue-violet throughout the procedure
What colour are gram negative organisms?
red, form acceptance of the counterstain, safranin. The violet iodine complex is lost when washed with acetone or another solvent
Why are yeast cells not gram positive cells even though they retain the crystal violet-iodine complex?
Yeast cells are eukaryotes and lack the bacterial cell wall morphology
Name the 4 staining solutions used in gram staining
Crystal violetGram’s IodineAcetone/alcohol mixSafranin
Why is the stain with acetone the cruicial step in gram staining?
If the cell is over treated with acetone (i.e. destainer is left for too ong,) then even gram positive organisms will appear negativeIf cell is not treated well enough with destainer, then gram negative organisms will appear positive.Aim for 5 seconds
Why is it necessry to stain bacterial cells?
Bacteria are almost transparent. Staining increases their visibility under the microscope
What causes the specific arrangement and pattern distribution of bacterial cells?
Uhm… i actually dunno. LOL, the type of bacteria they are? ugh.
Why do gram positive and gram negative bacteria stain differently?
Although both gram + and gram - cell walls contain peptidoglycan, the gram + cell wall is thickerthe gram - cell wall has additional lipopolysaccharides
Why is a four phase streak used?
To streak bacteria onto an agar plate in order to obtain well-separated ‘single’ coloniesfrom these colonies, pure cultures of the different bacteria can be isolated
What is the value of isolating a pure culture/?
Clinical specimens frequently contain mixed populations of bacteriaonce pure cultures are obtained, the organism implicated in an infection can be identifiedtreatment can then be modified if necessary
The wire loop is heated four times during this procedure. What is the significance of each flaming
To eliminate bacterial growth from other colonies… ?
When is an agar plate lawn used?
To test when bacteria are able to grow in the presence of anti-microbial agents
how does the agar plate lawn work
anti-microbial agent is placed in the centre of a freshly spread lawn of bacteria spread on an agar plateas bacteria grow overnight, they will cover the plate as usual, unless the agent inhibtis the growth. if growth is inhibtied we will see a zone of inhibitiontherefore it is crucial to obtain full coverage of the agar plate
What is colony morphology used for?
an aid in identification as the colony morphology varies according to species
What are the different kind of shape forms?
round, spindle shaped, irregular, pin point, filamentous
What arethe different types of elevation?
convex, raised, dome shaped, flat, umbonate
What are the different types of margins?
filamentous, lobed, erose, entire
What other categories of colony morphology are there?
colour, texture, odour
