Microbiology Flashcards
Draw and label the structure of a prokaryotic cell
Flagellum Pilus Circular DNA Plasma Membrane 70s Ribosome Peptidoglycan Cell Wall Slime Capsule Plasmid
What is the function of the plasma membrane in a prokaryote?
- barrier between the environment and the cytoplasm
- controls entry and exit of substances into and out of the cell
What is the function of the peptidoglycan cell wall in a prokaryote?
- prevents osmotic lysis
- rigidity and cell structure
What is the function of the slime capsule in a prokaryote?
- protect against other cell attack
- prevents cell drying out
What is the function of pili in a prokaryote?
Transferring plasmids (genetic information) by conjugation
What is the function of the flagellum in a prokaryote?
For mobility
What is the function of the plasmid in a prokaryote?
- contains extra bacterial genes, e.g. bacterial resistance
- can be exchanged between bacteria during conjugation allowing spread of resistance
What are the 3 bacterial shapes?
- Bacillus (Rod)
- Coccus (sphere)
- Spirillum (Spiral)
What are prokaryotes?
- no membrane bound nucleus
- no membrane bound organelles
- small ribosomes (70s)
- cell wall made up of peptidoglycan
- small cells, 0.5-5um
- reproduce by binary fission
What is the structure of gram positive bacteria?
- cell wall has a thick layer of peptidoglycan and a plasma membrane
What happens to gram positive bacterium in the Gram stain?
- peptidoglycan holds onto crystal violet dye
- dye is not washed out by ethanol and therefore appears purple in the Gram stain
What is the structure of Gram negative bacterium?
- thick outer layer of lipopolysaccharides with a thin layer of peptidoglycan
- plasma membrane beneath
What happens to Gram negative bacterium in the Gram stain?
- crystal violet dye is removed when rinsed with ethanol (dissolves the stained lipopolysaccharide layer)
- peptidoglycan is stained with the counter-stain and therefore appears pink in the gram stain test
What antibiotics interfere with Gram positive bacterium?
- Penicillin
- prevents bonds interlinking peptidoglycan forming
- when the bacteria divide, the cell walls are weak and collapse
- water uptake by osmosis bursts the cell
What antibiotics interfere with Gram negative bacterium?
- Require antibiotics that interfere with the cell’s metabolism/protein synthesis
- e.g. vancomycin
- penicillin is not effective as the outer layer protects the peptidoglycan
Why are animal cells not damaged by penicillin?
Animal cells do not have a cell wall, not damaged by penicillin
Definition of a Gram Stain
A method of staining the cell walls of bacteria as an aid to their identification
What is the method for the Gram-stain test?
- create a flame-fixed emulsion of bacterial samples on a slide
- flood with crystal violet, leave for 1 min then rinse of excess with sterile distilled water
- Add lugol’s iodine solution, leave for 1 min then rinse
- Flood with decolouriser (ethanol) for 30 seconds until run-off is clear
- Counter-stain with Safranin, leave for 1 min then rinse
- Gently blot dry slide
- Observe slide under oil immersion on a microscope
Why do you add alcohol/ethanol?
Alcohol removes unbound stain and lipopolysaccharide layer
Conditions necessary for growth and bacteria: temperature
- 25-45’c as metabolism is regulated by enzymes.
- mamallian pathogens optimum is 37’c
Conditions necessary for growth and bacteria: pH
- pH 7.4
- slightly alkaline conditions
- fungi prefer neutral to acidic conditions
Conditions necessary for growth and bacteria: oxygen
- some require oxygen
- some require anaerobic conditions
Conditions necessary for growth and bacteria: nutrients
- need a carbon and energy source ususally glucose
- need nitrogen source in organic or inorganic form for amino acid synthesis
- need growth factors; minerals and vitamins
What is the aseptic technique when handling bottles and caps?
Hold bottle cap in little finger and flame the bottle neck beofre and after extracting bacteria
Why is sterilisation important?
To stop contamination of environment from culture and contamination of culture from environment
Define obligate aerobes
require oxygen to survive
Define obligate anaerobes
require anaerobic conditions to survive, oxygen is toxic
Define facultative anaerobes
grow better in the presence of oxygen but can survive in it’s absence
What is the method for serial dilution?
- 9cm3 of sterile water is placed in a series of 6 sterile tubes
- using a sterile pipette add 1cm3 of culture into first tube
- transfer 1cm3 from the first tube into the second
- repeat this for all six tubes
- transfer 1 cm3 of eavh sample onto a sterile petri dish for each of the six tube onto agar plate
- seal plates with tape and incubate at 25’c
How to estimate population size from a serial population?
- let the bacteria grow
- select a dish containing 20-200 colonies that are distinct and sperate
- count the number of colonies
- multiply by the appropriate dilution factor
What is turbidity?
- indirect method
- cloudiness of culture is measured as bacteria numbers increase
- uses a colourimeter
- then population is read off calibration culture
Why is important in serial dilution to count between 20-200 colonies?
- over 200 is too greater number to count and make sure there is no overlap
- under 20 as there is a greater risk of accumulated error if two bacteria are in one colony
Why is important in serial dilution to count between 20-200 colonies?
- over 200 is too greater number to count and make sure there is no overlap
- under 20 as there is a greater risk of accumulated error if two bacteria are in one colony
What aseptic techniques are used?
- steriles all equipment by autoclaving at 121’c for 15 minutes
What can obligate anaerobes not do in the presence of oxygen?
Metabolise
