Topic 6.1- Bacteria and disease Flashcards
Why are aseptic techniques important when culturing microorganisms?
to produce uncontaminated culture so results are reliable + repeatable
List the basic aseptic techniques
- wipe surfaces with antibacterial wipes/cleaner
- set up Bunsen burner nearby. Convention currents prevent microbes from entering culture
- flame inoculating loop + neck of bottles before use
- minimise time that vessels containing bacteria are open
- sterilise all equipment e.g. using an autoclave
- wear protective clothing
Outline how to culture microorganisms
- Transfer bacteria to agar plate using sterile inoculating loop or pipette
- Tape lid on at 2 ends then invert the dish and incubate. In school labs ensure that dish isn’t airtight + do not incubate above 25 degrees to avoid growth of pathogen
Explain the difference between a streak plate and a spread plate
- streak plate: aims to obtain single colonies by rotating the plate to build layers of the culture on at least 3 separate streaks
- spread plate: aims to distribute microorganisms evenly with a sterile spreader
Describe the 3 types of nutrient medium
-liquid broth
-solid agar
(usually contain nitrogen, carbon, + minerals, often enriched with protein from extract of yeast, blood or meat)
-selective mediums (contain highly specific nutrient balance, only certain microorganisms grow)
Give the advantages of using a broth medium
- can provide anoxic + oxic conditions depending on depth, which helps to identify microbes/ determine their optimum conditions
- can grow a very large quantity of bacteria
Give the advantage of using agar as the medium
can obtain single, discrete pure colony for study
Name the 4 phases of the bacterial growth curve
- lag phase
- log phase
- stationary phase
- death phase
What happens during the lag phase?
microorganisms need to adjust to the environment before reproducing so population size only increases slowly
What happens during the log phase?
after every round of division population size doubles (exponential growth)
What happens during the stationary phase?
reproduction rate= death rate, so population size stabilises at its max
What happens during the death phase?
microorganisms die due to build up of toxic waste products + lack of nutrients
Name 3 methods used to estimate the growth of a bacterial culture
- cell count
- turbidity measurement (type of colorimetry to measure opacity)
- dilution plating
Explain how to conduct a cell count
- dilute broth sample with equal volume of trypan blue to stain dead cells blue
- use a calibrated haemocytometer with volume 0.1mm3. Count the cells in each of the sets of squares and calculate the mean
- number of bacterial cells = counted number x10^4 per cm3
Suggest advantages and disadvantages of using a cell count
advantage: only counts living cells
disadvantages:
- slow
- expensive equipment
- large margin for human error
Explain how to conduct a turbidity measurement
- use colorimetry. measure absorbance or % transmission of samples with known microorganism count
- plot calibration curve: absorbance/ % transmission (y-axis) number of microorganisms (x-axis)
- record absorbance/ % transmission of unknown sample
Suggest advantages and disadvantages of using a turbidity measurement
advantages:
-quick
-can be conducted in the field
disadvantages:
-counts both living and dead cells
-requires calibration curve from known samples
-assumes equal density of cells across culture
Explain how to conduct dilution plating
- grow a colony from a single microorganism
- perform serial dilution with distilled to see single colonies
- prepare a lawn plate and count colonies
- number of cells= number of colonies x dilution factor
What are endotoxins?
lipopolysaccharides that are an integral part of the outer layer of the cell wall of Gram-negative player
What are exotoxins?
soluble proteins that are produced and released into the body by bacteria as they metabolise and reproduce in the cells of their host
Define lipopolysaccharides
large molecules containing a lipid element and a polysaccharide element
What is tuberculosis?
a lung disease caused by Mycobacterium tuberculosis and M.bovis
What’s the primary infection of tuberculosis?
the initial stage of tuberculosis when M.tuberculosis has been inhaled into the lungs, invaded the cells of the lungs and multiplies slowly, showing no obvious symptoms
What is a tubercle?
the result of a healthy immune response to an infection by M.tuberculosis. a localised inflammatory response forms a mass of tissue containing dead bacteria and macrophages
What adaptation does Mycobacterium tuberculosis have?
it can avoid the immune system, allowing some bacteria to survive the primary infection stage. they produce a thick, waxy outer layer which protects them from macrophage enzymes- this means they can remain deep in the tubercles in the lungs (dormant or growing slowly)
What makes dormant or slowly growing tuberculosis bacteria active?
when the person becomes malnourished, weakened or their immune system doesn’t work well. these bacteria can then cause active tuberculosis
Define an antibiotic
a drug that either destroys microorganisms or prevents them from growing and reproducing
What is selective toxicity?
a substance is toxic against some types of cells or organism but not others
What is penicillin?
the first antibiotic discovered. affects the formation of bacterial cell walls and is bactericidal
What are bactericidal antibiotics?
kill bacteria
What is meant by bacteriostatic antibiotics?
inhibit the growth of bacteria
What’s tetracycline?
a bacteriostatic antibiotic that inhibits proteins
Define an antibiotic-resistant organism
not affected by an antibiotic even if it was effective in the past
What factors determine the effectiveness of an antimicrobial drug?
- the concentration of the drug in the area of the body infected (affected by how easily the drug can affect the tissue and how quickly it’s excreted)
- the local pH
- whether the pathogen or the host tissue destroy the antibiotic
- the susceptibility of the pathogen to the particular antibiotic used
How are bacteria harmful?
- produce exotoxins
- endotoxins are surface trigger immune response
- invade and destroy host tissues
How does Salmonella spp. cause disease?
- Gram negative bacterium with polysaccharide endotoxins on outer membrane, triggers release of cytokines
- acute inflammation results in diarrhoea
- releases toxins into the host when bacterium dies
How does Staphylococcus spp. cause disease?
secretes exotoxins (soluble proteins)
- barrel shaped proteins embed in host cell membrane so contents leak
- protease toxins
- superantigens trigger 20% of T cells so can cause toxic shock
How does Mycobacterium tuberculosis cause disease?
- triggers inflammatory response by infecting phagocytes in the lungs
- infected phagocytes are sealed in waxy-coated tubercles so bacteria remain dormant. first infection is symptomless
- if another factor weakens immune system, bacteria become active and destroy lung tissue
How does penicillin work?
beta-lactam bactericidal antibiotic prevents formation of the peptidoglycan cross-links in cell wall, causing osmotic lysis
How does tetracycline work?
bacteriostatic antibiotic, prevents protein synthesis by binding to small subunit of ribosome so tRNA cannot attach. therefore inhibits growth and division
(bacteriostatic antibiotics may also inhibit nucleic acid formation)
What causes antibiotic resistance?
- random genetic mutation, often plasmid e.g. antigen change shape
- these bacteria have selective advantage in the presence of antibiotics
- reproduce and pass on allele for resistance to offspring
- directional selection causes resistant strain
How do hospitals minimise the spread of antibiotic resistant bacteria?
- screening and quarantining of sick patients
- hygiene code of practice e.g. alcohol-based wipes, gels
- antibiotics prescribed only when necessary and course completed to minimise selection pressure
