Fundamentals of Microbiology Flashcards
What is Microbiology?
- Study of micro organisms (micro meaning small and biology meaning living things)
- e.g. bacteria, virus, fungi
Why is Microbiology important?
- Medicine
- Food and Drink production
- Agriculture
- Pharmaceutical Industry
- Genetic Engineering
How did they make pencillin?
- Penicillium mold naturally produces the antibiotic penicillium
- Scientists grew penicillium mold in fermentation tanks
- Then separated the penicillium product from the mold
- Penicillin was purified to use as an antibiotic
What are the main differences between prokaryotic and eukaryotic cells?
- Nucleus and membrane bound organelles : present in Eukaryotes only
- Cell size: Eukaryotes (10-100um), Prokaryotes (less than 5um)
- Organism type: Eukaryotes are multicellular, prokaryotes are unicellular
- Ribosomes: Eukaryotes have large 80S, Prokaryotes are small 70S
What is the classification system?
- Domain
- Kingdom
- Phylum
- Class
- Order
- Family
- Genus
- Species
How are organisms named using the classification system?
- Using genus (generic) and species (specific)
- Using capital letter only for genus and in itailics
- Escherichia coli
How does antibiotic resistance spread
- Animals get antibiotics and develop resistant bacteria in the gut
- Drug resistant bacteria can remain on meat ad spread to humans
- Fertilizer containing animal faeces on crops
- Humans get antibiotics and spread to community
What are the different prokaryotic cell shapes?
- Round
- Rod
- Curved Rod
- Short Rod
- Spiral
- Long, helical
What are the different prokaryotic cell shape arrangements?
- Single (coccus)
- Pair of coccus (diplococcus)
- 4 cells arranged into a square (tetrad)
- Chain of cocci (streptococcus)
- Cluster of cocci (staphylococcus)
- Single rod (bacillus)
- Chain of rods (Streptobacillus)
What is the cell wall made of in prokaryotes?
- Peptidoglycan (murein)
How do you distinguish between gram positive and gram negative bacteria?
- Their ability to retain a dye. Expose to dye (methyl violet), wash and wash with a decolourising agent (absolute alcohol)
- Gram negative can’t retain the dye (go pink), gram positive can (go violet) as they have a thicker cell wall
What are the 2 major roles of bacteria cell wall?
- To protect the cell against osmotic rupture particularly in diluted media and mechanical damage
- To assign bacterial shapes and their subsequent division into gram positive or negative micro organisms and antigenic attributes
What is the cell wall like of gram negative bacteria?
- Have a thinner cell wall, less sturdy
- Made more complex by layer of lipoprotein, polysaccharide and phospholipid known as the outer membrane. (Between membrane and cell wall surface)
What is the cell wall like of gram positive bacteria?
- Simple in structure compared
- Comprising of several layers of peptidoglycan connected to each other by cross linkages to form rigid scaffold
- Contain acidic polysaccharides = teichoic acids. Phosphate groups impart an overall negative charge
How do bacteria grow?
- Binary fission
- Replication of DNA, membrane pinches inwards, separation of DNA and ribosomes, cell wall forms around 2 new daughter cells
What are the 4 stages of the bacterial growth curve?
- Lag phase
- Exponential growth phase
- Stationary phase
- Death phase
What is the lag phase?
- Number of cells doesn’t change
- Metabolic adjustment (trying to adapt to environment).
- Cells grow larger and are metabolically active
What is the log or exponential phase?
- Actively dividing by binary fission and numbers increase exponentially
- The medium undergoes continuous change, as nutrients are consumed and waste excreted.
- Steep increase
What is the stationary phase?
- Waste products accumulate and nutrients gradually used up. Gradual depletion of oxygen
- The number of new cells created is equal to the number of cells dying
What is the death phase?
- The number of dying cells exceed the number of dividing cells due to accumulation of toxic products and exhaustion
- May be reversed by replenishment
- Steady decrease
What are the factors that affect microbial growth?
- Temperature
- pH
- Oxygen
- Osmotic pressure
- Light
- Media used
How does temperature affect microbial growth?
- Bacteria grow between 20-40 degrees (mesophiles)
- Low temperature = minimum growth, reduced enzyme activity
- Optimum temperature = optimum growth
- Too high = decline due to thermal desaturation. Enzyme and proteins denatures. Asymmetrical curve
How does pH affect microbial growth?
- Most micro organisms grow best around pH 7 (neutrality)
- Fungi prefer acidic conditions.
- Acidophiles have optimum growth rate in acidic conditions and alkaliphile have optimum growth rate in alkaline conditions
How does oxygen affect microbial growth?
- In a static culture, micro organisms occupy different regions of the medium reflecting on their oxygen usage
- Obligate aerobes : Grow near the surface
- Obligate anaerobes : Occupy zones of no oxygen
- Facultative anaerobes : adjust metabolism to oxygen (mostly at top but some are lower)
- Aerotolerant anaerobes : Don’t use oxygen and aren’t inhibited by it
- Microaerophiles: Have specific oxygen requirement, can only grow in narrow range of oxygen tensions
How does osmotic pressure affect microbial growth?
- Isotonic solution: No net movement of water
- Hypertonic solution: (Solution is more conc) Water moves out and cell shrinks
- Hypotonic solution: (Solution is more dilute) Water moves in and cell counteracts pressure to prevent swelling and lysis
How does light affect microbial growth?
- Phototrophic organisms require light for photosynthesis
- In labs : correct wavelength is used and doesn’t act as a heat source
- Fluorescent light produces little heat and not correct wavelengths in excess of 750nm for purple and green photosynthetic bacteria
How does the media used affect microbial growth?
- Chemically defined media: contain only chemically known components
- Selective media: Favour growth of some, inhibit others
- Enriching media: Contains added essential nutrients a specific organism needs
- Differential media: Help distinguish bacteria by the colour of colonies or change in medium
What are the elements required for growth and metabolism?
- Carbon, hydrogen, oxygen, nitrogen
- Sulfur, phosphorous
- Iron, calcium, potassium, sodium, magnesium, manganese
What is the main composition of bacteria?
- 70% Water
- 1% inorganic ions
- Carbohydrates, Amino acids, Lipids, Nucleotides and their precursors
- 7% Nucleic acids
- 15% Proteins
What is the laboratory cultivation media?
- Media are either produced by liquids or solidified with agar(extract of seaweed and sets to form a firm gel)
What are the types of bacterial growth media?
- Nutrient agar
- MacConkey medium: peptone, lactose, sodium taurocholate, agar
- Blood agar: Nutrient agar, 5% sheep or human blood
- Chocolate agar: Heated blood agar
What are the methods for counting bacteria?
- Direct microscopic
- Plate count (pour and spread)
- The most probable number
- Indirect method (turbidity)
What is the Direct Microscopic method for counting bacterial cells?
- By using a Petroff Hausser Chamber (a grid with 25 large squares)
- Bacterial suspension is added, microscopic count. Bacteria in large squares are counted and averaged
How can a fluorescent dye be used in bacterial counting?
- Penetrate cell membranes and detect live and dead cells. Some dyes cant detect dead cells. Dead cells can be stained read and live cells are stained green
What is a coulter counter?
- Rapid and accurate within a range of concentrations
- Does not differentiate between live and dead cells
- Is an electronic device that counts cells. It measures the change in resistance in an electrolyte solution when a cell enters through a small opening in the container wall. It counts the number of cells going through the opening
What is the Plate count method for counting the number of cells?
a) Pour plate method
- Serial dilution. Sample poured onto sterile plate, allowed to solidify and incubated
b) Spread plate method
- Spread sample evenly, plate incubated until bacterial colonies grow on the surface of the medium
What is the Most Probable method for counting cells?
- Statistical procedure to determine the number of bacterial cells in a sample.
- Sets of 5 lactose broth tubes are inoculated with 3 different volumes of pond water (10ml, 1ml, 0.1ml)
- Bacterial growth is assessed with change in the colour red -> yellow
What is the Indirect (turbity) method for counting cells?
- As the number of bacteria in suspension increase, the turbidity also increases and causes less light to hit the detector
- A spectrophotometer is used to measure the turbidity of the cell suspension
How do you test for the most efficient antimicrobial?
- tube dilution assay - determines me minimum inhibitory concentration of antibiotic
- disc diffusion method
How do you carry out the tube dilution assay?
- Organism is incubated with serially diluted antibiotic (he lowest conc capable of preventing microbial growth is the MIC)
What is the disc diffusion method?
- Bacterium to be tested is spread on an agar plate then paper discs soaked in appropriate antibiotic is placed on the surface
- Incubate and measure of the zone inhibition (clear ring) measured in mm
Also known as me Kirby- Bauer disk diffusion test
What are the different sterilisation methods?
- Physical methods
- chemical methods
What are the main forms of physical sterilization?
- Heat sterilisation 100 degrees for 10 mins
- Filtration
- irradiation
- sound waves
- pressure
- sunlight
What is the moist heat sterilisation method?
- Performed by steam under pressure (autoclaves)
- autoclaves remove air from the chamber and replace with pure saturated steam
What is the irradiation method?
- Nonionizing radiation : Ultraviolet
- Ionizing radiation : X rays, gamma rays, high energy electron beams
- UV radiation causes formation of thymine dimers in DNA leading to lethal mutations in microbes
What is the membrane filtration method?
- Used to sterilise heat labile substances
