3.4 Microbiology Flashcards
What are the three shapes of bacteria called and what do they look like?
Bacillus - rod shaped
Coccus - spherical
Spiral - corkscrew shaped
What is a gram stain? (basic definition)
A method of staining the cell walls of bacteria as an aid to their identification.
What colour is gram positive and gram negative bacteria after staining?
Gram positive - stain purple after the crystal violet stain is added.
Gram negative - stain red after counterstain safranin is added.
Why is it that gram positive bacteria stains the colour it stains?
The gram positive bacteria contains a thick layer of peptidoglycan in their cell wall that retains the crystal violet stain and does not get rinsed away by alcohol or stained red by the counterstain safranin.
Why is it that gram negative bacteria stains the colour it stains?
Gram negative bacteria stains red as it has a thin layer of peptidoglycan and then has extra layers of protection from different layers. This means it does not retain the crystal violet stain very well and the purple colouring is washed away by the alcohol. But the counterstain safranin can stain gram negative bacteria so the bacteria is ultimately red.
What bacteria is susceptible to lysozymes and penicillin and why?
Gram positive is susceptible to lysozymes due to the thick layer of peptidoglycan and the lack of outer lipopolysaccharide layer in the cell wall. Bacteria constantly make and break chemical links in their cell walls and lysozyme hydrolyses the bonds holding peptidoglycan molecules together.
Which bacteria are not affected by lysozymes and penicillin and why? And thus how to do you control/kill them?
Gram negative bacteria is immune due to it’s more chemically complex cell wall containing a thinner layer of peptidoglycan and many lipopolysaccharides. To control them you need a different class of antibiotics that interfere with the cells ability to make proteins.
what are the bacteria usually placed in for culturing?
Petri dishes usually or a flask sometimes
What nutrients are needed and how does the bacteria get it in a petri dish?
Labs use a nutrient media to provide nutrients, and this provides -
- A carbon and an energy source, usually glucose
- Nitrogen for amino acid synthesis
What are the conditions needed for micro-organisms to grow?
- Nutrients
- Growth factors including vitamins
- temperature (25-45)
- pH (7.4)
- Oxygen
What does the term obligate aerobes mean?
the micro organism requires oxygen for metabolism
What is the term used to describe micro organisms that grow best in the presence of oxygen but can survive in its absence?
facultative anaerobes
What does obligate anaerobes mean?
cannot grow in the presence of oxygen
What is the difference between defined and undefined culture media?
A defined medium contains only known ingredients and an undefined medium contains components that are not all known.
What does the term selective medium mean?
It means the medium only allows certain bacteria to grow e.g only allows gram negative to grow or something like that
What are two potential problems that must be avoided when working with bacteria?
- contamination of cultures from the environment
- contamination of the environment by the culture
What does the term aseptic technique mean?
Laboratory practice that maintains sterility in apparatus and prevents contamination of the equipment and the environment.
How do you prevent contamination of pure cultures and apparatus by bacteria from the environment?
- Sterilise all apparatus and media before use to prevent initial contamination
- Handle cultures carefully, flaming the necks of culture vessels before opening and closing and use equipment such as sterile loops to prevent subsequent contamination
How do you prevent contamination to the environment by the bacteria being used in the experiments?
- Sterilise the work surface before and after the experiment using a disinfectant
- Use the correct handling techniques to prevent the contamination of personnel and the immediate environment by the organisms being cultured.
What steps must be taken during the process of inoculation? (include steps to prevent contamination)
- Hold the culture bottle in one hand; remove the cap with the little finger of the other hand. Do not place the cap down onto the work surface
- Flame the mouth of the bottle for two or three seconds
- Pass the inoculating loop through a flame until its red hot, and allow it to cool in the air.
- Lift the lid of the petri dish just enough to allow entry of the inoculating loop.
- Secure the petri dish lid with two pieces of adhesive tape. Do not seal all the way round as this could create anaerobic conditions and, potentially, encourage the growth of pathogenic micro organisms.
- incubate at around 25 degrees. Cultures should NOT be cultured at 37 degrees as this is an ideal temp for pathogens.
- Do not open petri dishes after incubation
What is a pathogen?
An organism that causes disease in its host.
What is an autoclave and what is it used for?
It is a preferred method of sterilisation in a laboratory. It is a sealed container in which glass and metal equipment is heated at 121 degrees in steam under pressure for 15 minutes after the required pressure has been reached.
What is done to disposable materials after use during inoculation?
Placed in plastic autoclavable bags and autoclaved and then placed in a dustbin.
What is the difference between total count and viable count?
-A total count uses methods that count all the cells present but cannot distinguish between live and dead cells.
-A viable count only counts the cells that are capable of reproducing (forming colonies) and are therefore alive.
