Topic 6: Microbiology and Pathogens Flashcards
What is a microbial culture?
A method of multiplying microbial organisms by letting them reproduce in a predetermined culture medium under controlled lab conditions
What factors must be controlled when making a microbial culture?
Nutrient levels
Oxygen
pH
Temperature
What are the main problems when culturing microorganisms?
1 - Harmless microorganisms might mutate to dangerous ones
2 - Pathogens enter and grow - can cause disease
3 - Contamination with unwanted microorganisms will ruin the investigation
What precautions should be taken when culturing microorganisms?
Use sterile equipment
Do not remove culture from lab
Dispose of culture safely
What does sterile mean?
Being sterile is to be free from living microorganisms and their spores
How do you dispose of microorganism cultures safely?
Seal in a plastic bag and sterilise at 120 degrees for 15 minutes under high pressure and then throw it away
Outline the method to culture microorganisms
- Choose a microorganism to culture
- Obtain a culture of the microorganism
- Select and make up the nutrient medium
- Innoculate the medium
What are the different types of nutrient medium?
Liquid broth, nutrient agar and selective medium
What is nutrient agar?
A solid nutrient elly extracted from seaweed used in Petri dishes
How do you make nutrient agar?
Pour nutrient broth and molten agar into a Petri dish and let it set at 50C
What is the advantage of agar
Sets at 50C but melts at 90C so you can keep it at a high temperature
What is a selective medium?
A growth medium with a specific combination of nutrients so only a certain type of microorganism grows on it
Define innoculation
The process by which microorganisms are transferred into a culture medium under sterile conditions
Explain how to innoculate a liquid nutrient medium
Innoculating loop scrapes bacteria from solid culture to liquid nutrient broth to form innoculating broth.
Flask stoppered with cotton wool
Incubate at suitable temp
Regularly mix
Why do you stopper a flask of liquid culture when culturing it?
To prevent contamination by microorganisms in the air
Why do you mix a flask of liquid culture regularly when culture?
To ensure the broth is aerated bc microorganisms need oxygen
Explain how to innoculate a solid nutrient medium
Sterilise the innoculating loop in a Bunsen Dip into bacteria suspension Streak across agar surface Replace lid, clse tape and label Turn upside down
What does aseptic mean?
Sterile - free from contamination from harmful bacteria
Why are aseptic techniques used when culturing bacteria?
To ensure the procedure is safe and to prevent the contamination of the culture.
Give examples of aseptic technique
Using sterile equipment
Using flamed equipment
Replacing the lid of the petri dish as soon as possible
What is a pure culture?
A culture containing only one type of microorganism
How can a pure culture be made?
Making conditions aerobic/anaerobic depending on organism
Making selective medium specific to it
Reinnoculate a plate with the microorganism on it (identified by using an indicator)
What information do we need to measure the growth of a culture?
The number of cells at different times
Name some methods used to measure the growth of cultures
Cell count
Dilution plating
Turbidimetry
Size/area
What is a cell count?
A method of measuring the growth of cultures of bacteria or single selled fungi
What equipment is needed for a cell count?
Haemocytometer
Microscope
What is a haemocytometer?
A thick glass microscope slife with a rectangular chamber that holds liquid. It has perpendicular lines
What is trypan blue?
A dye that stains dead cells blue and leaves living cells white
Why do we dilute cells with trypan blue before putting them into a haemocytometer?
To stain them so counting is easier
To separate the cells to make them easier to count
What is the rule for cells on lines when counting cells in a haemocytometer?
Count them if they’re on the top or right line
Don’t count them if they’re on the bottom or left
How is a haemocytometer used?
Put under a microscope and viewed under a low power so the cells can be seen
Why is it important to not move the haemocytometer when counting?
Will mve the cells so counting becomes incorrect
Air bubble may be introduced
What is turbidity?
The cloudiness of a liquid caused by a large number of individual particles in the solution
What is turbidimetry?
A method of measuring the concentration of a substance by measuring the amount of light that passes through it. A specialised form of colorimetry
What are calibration curves?
Graphs of known concentrations against their absorbances
How are calibration curves made?
Control culture is made and samples taken at regular intervals
Put into colorimeter and turbidity is measured
Do cell count of the same sample
Repeat over time
Plot graph of turbidity against cell count and compare unknown turbidities to get cell count directly
How is turbidity measured?
The absorbance of light of a sample is measured by a colorimeter
What is dilution plating?
A method used to obtain a culture plate with a countable number of bacterial colonies
What assumption is made during dilution plating?
One colony comes from one bacterial cell
Define total viable cell count
A measure of the number of cells that are alive in a given volume of culture.
Why is dilution plating needed?
Because directly counting the number of colonies from a concentrated solution is difficult bc there are so many so they clump together. By dilution you can get a smaller number of cells
What calculation can be done to get the total viable cell count?
number of colonies x dilution factor
How can you check the accuracy of dilution plating?
Doing a cell count with a haemocytometer
What do you do if there are two or more plates with enough colonies to count in dilution plating?
Count the number of colonies in each plate then calculate a mean
How can you find the mass of microorganisms/fungi in a culture?
Sample of broth centrifuged/filtered to remove bacteria from liquid
Material dried until mass stabilises
Measure the dry mass overtime
Increase in mass = increase in growth
By what process do bacteria reproduce?
Binary fission
Define generation time
The time between divisions of a bacteria
What kind of scale is used for bacterial growth and why?
A logarithmic scale because the numbers are huge bc bacteria reproduce hella fast
What equation can be used to find the number of bacteria in a population?
Nt = No x 2^kt Nt - no of organisms at time t No - number of organisms at the start k - exponentional growth constant t - time
What equation can be used to find the exponential growth constant?
k=(log10Nt - log10No)/(tlog10(2))
How can the growth of a bacterial population be shown
A graph with axis time (t) against log10Nt (y)
Name the phases in the bacterial growth curve
Lag phase
Log phase
Stationary phase
Death phase
Explain the lag phase of the bacterial growth phase
Line is flat because bacteria are adjusting to their environment so growth is slow
Enzymes and genes still activating
Explain the log phase of the bacterial growth phase
Bacteria growing really fast
Growth time depends on number of nutrients and amount of space
Explain the stationary phase of the bacterial growth phase
Growth rate is zero - new cells forming = cells dying
Nutrients being used up
Waste products starting to build up
Explain the death phase of the bacterial growth phase
No. of cells dying exceeds the no of new cells forming
Nutrients have run out
Waste products have built up too high
Why doesn’t the exponential growth of bacteria continue forever?
Nutrient exhaustion - nutrient levels can’t support growth
Build up of waste products - levels can become toxic and inhibits growth (eg CO2 lowers pH)
How can bacteria cause infection in the body?
Destroying host tissue
Releasing toxins
How does TB cause infection?
Destroys host tissue
Name ways bacteria can enter the body
Wounds
Natural openings
Name the ways in which bacteria can be transferred
Droplet infection
Vector (intermediate carrier)
Direct contact
Touching the surface someone infected has touched
Why do bacteria produce toxins?
By products of their metabolism
Name the different types of toxins
Endotoxins
Exotoxins
What are exotoxins?
Soluble proteins that are produced and released into the body outside of the bacterial cell
What effect do exotoxins have on the body and why?
Widespread effect because they can travel through body fluids so they have a range of effects
What are endotoxins?
Lipopolysaccharides in the membrance of Gram negative bacteria that may be released from the bacteria if it breaks down
What effects do endotoxins have on the body and why?
Local to the site of infection because the toxins are in the bacteria for the majority of the time
What effect does salmonella have on the body?
Inflammation in the gut lining
What are antibiotics?
Drugs that destroy microorganisms or prevent them from reproducing
What kind of illnesses are antibiotics used for?
Bacterial not viral
What is selective toxicity?
Damage is done to the pathogen not the host cell
Name the first antibiotic
Penicillin
How can antibiotics be classified?
Bacteriocidal or bacteriostatic
Define bacteriostatic antibiotics
Completely inhibit the growth of microorganisms
Name a bacteriostatic antibiotic and its mechanism
Tetracyline - interferes with protein synthesis in small ribosomes in bacteria
Define bacteriocidal antibiotics
Antibiotics that kill bacteria
Name a bacteriocidal antibiotic and its mechanism
Penicillin breaks open bacteria cell walls
Define broad-spectrum antibiotics
Destroys a wide range of harmful pathogens
Define narrow-spectrum antibiotics
Targets one or two pathogens
What does the effectiveness of a drug depend on?
Drug concentration
pH
Susceptibility of pathogen (is it antibiotic resistant)
Whether pathogen can destroy the antibiotic
What is antibiotic resistance?
When bacteria mutate so drugs are no longer effective on them
Why are mutations common in bacteria?
Because bacteria reproduce rapidly
How does natural selection lead to antibiotic resistance?
Allele which causes resistance allows bacteria to survive. When bacteria reproduce, the allele is passed on quickly through other generations. Allele frequency increases.
What is horizontal transmission?
When bacteria can acquire resistance from other species through the transmission of their plasmids
How can we control the spread of antibiotic resistance?
- Prescribe antibiotics sparingly
- Complete the course of antibiotics
-Good hygiene measures:
Alcohol gels in hospitals
Staff wearing clean clothing
Isolating ill patients
Screening patients for infection before they enter the hospital
Monitoring levels of healthcare acquired infections
Why are viral infections specific to particular tissues?
Host cells have antigens and only a few viruses have complementary receptors
What is influenza
A virus which infects the tissues of the breathing system - especially the lungs
What are the modes of transmission of influenza?
Droplet infection Direct contact with: -animal droppings -virus filled mucus -surfaces contaminated with the virus
What are the modes of infection of influenza
Ciliated epithelial cells of the lungs are infected. Viral RNA takes over the cell, reproduces and cell releases viruses and die
What are the pathogenic effects of influenza?
Headaches, runny nose, coughing, vomiting, fever etc
How is influenxa treated?
No cure but antivirals used to relieve symptoms
Vaccine used as a preventative measure
What is stem rust fungus
A fungus which affects cereal crops
What are the modes of transmission of stem rust infection
Wind carries pores
Infected plant fragments in the soil
What are the modes of infection of stem rust infection
Spores germinate in water on the plant, the fungus enters the plant through stomata and enzymes digest plant cells and nutrients are absorbed
What are the pathogenic effects of stem rust fungus
Nutrients lost from plant
Stem weakened
Blisters on leaf surface
Plant loses control over water loss
How can stem rust be controlled?
Dont use nitrate rich fertilisers
Use earlier maturing plants
Bigger spaces between plants
Fungicides
What is malaria?
Fever caused by a parasite called Plasmodium spp.
What is the mode of transmission of malaria?
Mosquito vectors transfers parasite to humans
Mouthpiece pierces the skin and the saliva has anticoagulants so blood doesnt clot while it gets its blood meal.
What is the mode of infection of malaria?
Parasites travel to the liver –> RBC –> reproduce asexually then burst out of RBC —> eventually transmitted back to the mosquito taking the blood meal
What are the pathogenic effects of malaria?
Shaking, fever, sweatin
Muscle pains
Liver damage
Anaemia
Define an endemic disease
A disease that is constantly present in an area or country
Given an example of an endemic disease
Malaria
Why is it difficult to treat endemic diseases?
Widespread
Hard to get people to cooperate
No. of hosts
Expensive
Name two ways we can control malaria
Prevent bites
Control number of mosquitos
How can we prevent mosquito bites
Insect repellents
Mosquito nets with insecticides
Screens on door
Clothing to cover skin
How can we control the number of mosquitos
Avoid standing water/sewage
Introduce predators
Pesticides
Water treatment to kill larvae
Name some of the ethical issues with controlling diseases
People unsure about safety of vaccines
Difficult to obtain consent where medical education is little
Money can be spent better elsewhere
Name some of the social issues with controlling diseases
People need to change habits
Change social attitudes - difficult
Name some of the economic issues with controlling diseases
Treatment, control and prevention is expensive and countries are normally poor
Difficult to allocate resources
What is the role of the scientific community in controlling diseases
Prevention: getting insecticide filled nets
Treatment: using drugs in combination is more effective
What are antigens?
Any substance that stimulates an immune response from the body
What substances can be antigens?
Proteins, glycoproteins, carbohydrates, toxins and even whole organisms
What is a non-specific response?
The body’s defence against pathogens which is triggered by any pathogen
What are the three main types of non-specific response?
Inflammation
Fever
Phagocytosis
Explain the stages leading to inflammation
Tissue damage causes mast cells and basophils to release histamines
Blood vessels dilate - local heat and redness to reduce effectiveness of pathogens
Leaky capillaries cause leucocytes to be released and engulf pathogens and antibodies to disable them
What are histamines?
Chemicals which cause blood vessels to dilate and cause capillaries to leak
Explain why you get a fever while ill
Hypothalamus increases body temperature:
- reduce the effectiveness of pathogens
- immune system works better at a higher temperature
What is a phagocyte?
A general term for an WBC that engulfs and digests pathogens any other foreign material in the blood
Name the 2 types of phagocytes
Neutrophils and macrophages
Name some characteristics of neutrophils
Granulocyte
Can only ingest a few pathogens before it dies
Cannot renew lysosyme stores
Name some characteristics of macrophages
Agranulocyte
Made from monocytes from the tissue
Can renew their lysosyme stores
Explain how phagocytes work
Engulfs pathogen
Phagosome surrounds pathogen and fuses with lysosom
Enzymes break down pathogen
Cytokines released
What do cytokines do?
Signal for other WBCs
Stimulate immune response
What are opsonins?
Chemicals which bind to pathogens and label them so WBCs can detect them more easily
What is the immune response?
Body’s specific response to invasion by pathogens
Name the four characteristics of the immune system
Distinguish self from non-self
Diverse
Specific
Immunological memory
What are the two main types of WBC in the immune system?
Lymphocyte and macrophage
What are the two types of lymphocytes in the immune system?
B lymphocytes
T lymphocytes
Name the types of B lymphocytes
B effectors
Plasma cells
B memory
Name the types of T lymphocytes
T killer
T helper
T memory
Where do B cells get made and develop?
Bone marrow then develop in the lymph gland
Where do T cells get made and develop?
Bone marrow then develop in the thymus gland
What do plasma cells do?
Produce antibodies
What do T killer cells do?
Produce chemicals to kill infected cells
What do T helper cells do?
Activate plasma cells and B cell APCs
What are the body’s two specific responses to infection?
The humoral response and the cell mediated response
When is the humoral response activated?
When the antigens of the pathogens are outside of body cells
Name the two main stages of the humoral response
T helper activation
B effector stage
Summarise the stages of the T helper activation stage
Macrophage engulfs pathogen
Surrounds with phagosome and lysosome
Separates antigen and binds it to MHC
Becomes APC
T cell with complementary receptor binds to antigen
T cell divides and replicates into activated T memory and T helpers
Summarise the stages of the effector stage
B cells act as macrophages and become APCs
T helper binds to B cells and releases cytokines
B cells divide into B effector and B memory
B effector –> plasma cell —> antibodies
What is clonal selection?
The selection of cells that carry the right antibody for a specific antigen
What are antibodies?
Glycoproteins that are released into circulation and bind to the antigen of pathogens
How can antibodies cause pathogen deaths?
1) Agglutination - pathogens clump then get engulfed
2) Opsonisation - acts as an opsonin and signals for phagocytes
3) Neutralisation - neutralises toxins
Name adaptations of plasma cells
Hella endoplasmic reticulum
Ribosomes
When is the cell mediated response used?
When the pathogen enters the body cells
Summarise the stages of the cell mediated response
Body cells acts as an APC
T killer w complementary receptor binds to APC
T helper releases cytokines so T killer rapidly divides
Release enzymes which make pores in membrane so water +ions enter membrane and cause cell to burst
Pathogens destroyed
T memory cells made
What is the difference between the response of T memory cells and B memory cells
T memory cells release T killer cells
B memory cells produce antibodies
What is the primary immune response?
When the body has come into contact with a pathogen and its antigen for the first time. Involves the production of antibodies and memory cells
What is the secondary immune response?
When the body comes into contact with a pathogen again and memory cells are used
What is natural active immunity?
When the body produces its own antibodies against a pathogen and its antigen encountered naturally
What is natural passive immunity?
When antibodies are made by the mother and passed onto the baby via breastmilk or placenta
What is immunisation?
The process of protecting people from infection by giving them passive or active artificial immunity
What is a vaccination?
The procedure of giving someone dead or inactive pathogen to stimulate an immune response
What is passive artificial immunity?
When the antibodies from one individual are extracted and injected into another
What is active artificial immunity?
When the body produces its own antibodies and memory cells to an antigen from a vaccination
How is the pathogen in a vaccination made non-infectious?
Toxin detoxified
Inactive/dead pathogen
Attentuated pathogens - viable but modified so they dont cause disease
What is the only disease to be eradicated?
Smallpox
Why is it not possible to totally eradicate many diseases?
They have multiple hosts - animals, soils and waters
What is the elimination of a disease?
Where it is no longer seen in humans but exists in the environment or animal so vaccinations still needed
What is meant by a controlled disease?
When a disease is still occurring but not to the extent that it is a health problem
Define herd immunity?
When a large proportion of a population is immune to a pathogen (usually by vaccination) which lowers the risk of infection
What are the pros of vaccination?
Person is protected against infection - dont die
Herd immunity - those who cant get immunised are still safe
Costs of treating disease minimised
What are the cons of vaccination?
Some children cant be vaccinated bc of allergies or extreme immune response
Name a bacteria which produces exotoxins
Staphylococcus aureus
Name a bacteria which produces endotoxins
Salmonella
