1
Q
What did Robert Hooke do in 1664?
A
- Invented microscope
- Describes microscopic structure of blue molds, using 30x magnification microscope
- First to use the word ‘cell’ saw that all living things have the same structure
- Book Micrographia very influential due to excellent artwork
2
Q
What did Leeuwenhoek do in 1684?
A
- Develops powerful microscopes (300x mag.), discovers “wee animalcules” in pond water
- First evidence of bacteria and protists
- He is considered the ‘father of microbiology’
3
Q
What did Needham and Spallanzani do in the 1700s?
A
John Needham
-There is a “life force present in all matter that causes spontaneous generation
-Boiled hay in water to kill everything, left it to exposed air and microbial growth occurred
Lazzaro Spallanzani
-Repeated experiment but sealed the flask
-Microbes did not re-appear
4
Q
What did Pasteur do in 1861?
A
- Vaccination
- Fermentation
- Pasteurization
- Disproved the theory of spontaneous generation (non-living objects can give rise to living organisms)
- Swan neck flask experiment:
- Boiled hay in swan neck flasks
- Broke neck of one of the flasks: bacterial growth in that one
- In the one with no broken neck which allowed air to pass, no bacteria growth
5
Q
What did Koch do in 1876?
A
-Staining methods for microscopy
-use of solid growth media (agar)
-Identified bacteria which caused:
-Tuberculosis
-Cholera
-Anthrax
-Discovered spores and explained survival of anthrax in soil
-Germ theory: important in disproving spontaneous generation
-Koch’s postulates:
An organism that causes a disease must;
1. Be found in all cases of the disease
2. Be isolated from the diseased host in pure culture
3. Produce same disease in experimentally-infected host
4. Be re-isolated from the experimentally-infected host
6
Q
What did Fleming do in 1928?
A
Found mold growing on a petri dish killed the bacteria around it
- Mold juice killed many bacteria, including agents of pneumonia
- Penicillin notatum
7
Q
What did Florey and Chain do in 1935-1945?
A
- Purified penicillin and developed mass production
- First really effective antibiotic. Discovery helped the Allies to win WWII
- Still dependent on fungus to make penicillin as it is hard to make
8
Q
How many trillion microbial cells does the human body have?
A
40
9
Q
What is normal flora?
A
o Found at specific sites
o Specialised for that site
o Mostly bacteria
o Acquired at birth, from diet, from the environment
10
Q
Where does microflora initially come from, and how does it develop?
A
- Concept of normal flora is flexible and differs between individuals, locations and times
- At the beginning, microflora comes from the mother
- It then develops based on diet, medicines, environment, people interacted with
- Broadscale similarities but individual differences
11
Q
What are positives of having natural microflora?
A
‘Prime’ the immune system
Provide nutritional benefits
Compete with pathogens
12
Q
What are negatives of having natural microflora?
A
Can cause disease if moved to the wrong location
Staphylococcus from nose to open wound can cause wound infection
Can cause disease even in normal habitat
Streptococcus can cause tooth decay
13
Q
What is a pathogen?
A
Disease-causing microorganism
14
Q
What are obligate pathogens?
A
pathogens that are always harmful
o Viral infection usually damages or destroys the host cell
15
Q
What are opportunistic pathogens, and what factors do they depend on?
A
only cause disease under certain conditions
o Numbers- abnormally high cell density
o Location- get into the wrong place
o Host health- immune system compromised
o Virulence factors- gain antibiotic resistance
Bacteria can horizontally share genes and evolve extremely easily
16
Q
What is the concept of the One Health concept?
A
- Healthy people Healthy environments Healthy animals
- Activities in homes and farms influence disease as a whole
- Consider animals and the environment when we are trying to manage human diseases
- Many human diseases originate in animals
- Use of antimicrobials in agriculture impacts on human pathogens
- Disease is influenced by urbanization, globalization, climate change, pollution
17
Q
What are major infectious disease problems due to?
A
- Emerging infections due to new pathogens
- New problems with old pathogens
18
Q
Describe what tuberculosis is?
- Bacterial cause
- How it is spread
- What it does
- Symptoms
- How many are affected
- Treatment
A
o Caused by bacterium Mycobacterium tuberculosis
o Spread person-person by airborne droplets (coughing)
o Infects lungs cough, chest pain, weight loss, death
o Disease can be ‘latent’ for many years (no symptoms)
o Symptoms are non-specific: difficult to diagnose
o One-third of world’s population is infected
o New strains of TB are resistant to antibiotic treatment
Drug resistant TB is difficult to treat and treatment is expensive
19
Q
Describe the role of microbes in the production in the food production chain step
A
Good-
- Rumen microbes help cows digest grass
- Symbiotic fungi enhance plant growth
- Soil microbes recycle wastes into nutrients
Bad-
- Pathogens infect animals and plants
- Irrigation water can be contamined with human pathogens
- Post harvest contamination can ruin food crops
20
Q
Describe the role of microbes in the processing in the food production chain step
A
Good-
Many foods require fermentation
Bad-
Factories can be contaminated by microbes
21
Q
Describe the role of microbes in the distribution in the food production chain step
A
Good-
Some microbes are important food sources
Bad-
Refrigeration needed to stop microbial growth
22
Q
Describe the role of microbes in the restaurant preperation in the food production chain step
A
Poor hygiene –> food poisoning
23
Q
Describe the role of microbes in the restaurant consumer in the food production chain step
A
Good-
Good gut microflora beneficial to health
Bad-
Poor gut microbiota causes poor health
24
Q
Describe the role of microbes in the retail in the food production chain step
A
-Poor hygiene –> food poisoning
25
Describe the role of microbes in the home consumers in the food production chain step
Good-
Good gut microflora beneficial to health
Bad-
Poor gut micriobiota causes poor health
26
What is the process of food production?
```
Production
Processing
Distribution
Restaurant
Restaurant preparation
Restaurant consumer
Retail
Home preparation
Home consumers
```
27
How do microbes maintain soil health?
o Fixing nitrogen from nitrogen gas to ammonia, which is extremely energy costly
o Breaking down organic wastes into inorganic nutrients
o Suppressing pathogenic microbes (animal+ plant pathogens)
o Breaking down toxic molecules like pesticides and pollutants
28
Why are microbes useful in animals? Use a specific example
o Cellulose: sugar polymer, abundant in plants, carbon-rich but difficult to digest
o Rumen microbes break down cellulose sugars organic acids, CO2, CH4
Rumen in cow
• Anaerobic environment
o Anaerobes killed by oxygen
• Not all living things need air
• Mutualistic relationship between cow and microflora
o Organic acids and dead microbial cells then digested by animals
o Methane is a potent gas --> cows farting contributes to climate change
29
Why are microbes useful in plants? Give two specific examples
• Microbes promote plant growth via mutualism
o Mutualism- ecological interaction where both partners benefit
o Mycorrhizal fungi (in most plants) enhance water + inorganic nutrient uptake, receive sugars from plant
o Rhizobium bacteria (in legume roots) fix nitrogen, receive sugars in return
30
What are the main plant pathogens?
• Just like humans and animals, plants are subject to disease caused by microbes
o Fungi and viruses are the main problems
o Crop pathogens cause global losses of about 30% of total yield
31
Talk about the Tobacco mosaic virus
Tobacco mosaic virus is a very simple entity (only 3 genes) but it can infect many crops
Remains infectious long after the virus has left the plant
32
What are the main animal pathogens?
• Pathogens can be viruses, bacteria, fungi or protists
33
Why are pathogens infecting farm animals a problem?
• Pathogens infecting animals inflict suffering, death, and massive economic losses (about 20% of total production)
34
Why was the foot and mouth disease so devastating, and how was it introduced?
o Devastating viral disease infecting many farm animals
o Infects cows, pigs, sheep and goats but not humans
o Resulted in huge economic losses
o Cause of UK outbreak was that pigs were fed waste products including meat illegally imported from infected animals
35
What is zoonosis and why is it a problem?
Human infection arising from animals
o Human pathogen may be normal flora for the animal (e.g. salmonella in chickens)
o Animal is a vector for disease (e.g. ticks and lyme disease)
Target shaped rash
High fever
Fatigued
36
What is fermentation?
Microbial transformation of foods
| • Anaerobic metabolism of sugars --> alcohols, acids, CO2 due to the action of fungi and/or bacteria
37
Why is fermentation useful?
• Many products:
o Beer
Barley (source of sugars to support fermentation)+
Hops (natural preservative and provides bitterness)+
Water+
Yeast
• Saccharomyces (brewer’s yeast)
• Yeast is a unicellular fungi whilst mold is multicellular filamous fungi
=Beer
o Bread
o Cheese
38
Why does spoilage happen and how much food is lost to spoilage?
• Spoilage is due to the growth of fungi and bacteria, or due to enzymes these microbes secrete
-Approx 20% of all food is lost to spoilage
39
How is spoilage prevented?
• Spoilage is prevented by refrigeration, preservatives and fermentation
o Refrigeration double edged sword some microbes grow better at colder temperature
o Some preservatives such as sugar and salt are not great for human health
o Fermentation- curtails activity of pathogens by making good microbes ferment the food and reduce nutritional content for bad microbes
40
What does spoilage do?
• Spoilage makes food taste bad and reduces nutritional content
41
How does food poisoning occur?
• Food poisoning can occur from spoiled or fresh food- different sets of microbes are responsible for spoilage vs poisoning
42
What is a food-borne infection?
• Food-borne infection- microbes grow in gut
| o Salmonella enterica
43
What is food-borne intoxication?
• Food-borne intoxication- microbes make toxins in food
| o Clostridium botulinum
44
What can lead to food poisoning?
o Origins of food- determines types and number of microbes
o Storage and preparation- refrigeration, raw/cooked
Cooking doesn’t destroy all toxins but will destroy live pathogens
o Human factors- hygiene
45
What does maintaining a safe food supply depend on?
```
• Maintaining a safe food supply depends on managing microbes at every stage in the production chain
o Soil
o Plants
o Animals
o People
o Factory
o Kitchen
```
46
How many bacteria do we have in our gut biome?
40 trillion
47
What does our gut microbiome depend on?
o High fibre diet- high bacteriodetes
o High protein and fat diet- high firmicutes
• Gut microbiome depends on mother’s microbiome
o Baby inoculated by mother and maintains microflora for a while after birth
o Caesura is bad for this microflora inoculation
48
What is a healthy microbiome important for?
```
o Proper food digestion
o Nutrient absorption
o Resistance to pathogens
o Immune functioning
o Mental health limited evidence
```
49
What is a bad microbiome linked to?
o Allergies
o Irritable bowel disease
o Type 2 diabetes
o Cancer
o Obesity
Fat mice have a distinct gut microbiome
Transplanting fat mouse microbes into a germ-free mouse makes it fat too
50
What is the vast majority of all biodiversity?
Microbes
| Specifically archaea and bacteria
51
What is a barrier to studying all microbes?
• Most microbes cannot be isolated or cultured
| o They cannot be studied nor streaked on agar plates
52
What is biogeochemistry?
Biogeochemistry- Biological processes that impact on chemistry at a global scale. Most of these reactions are done by microbes
53
What are the 4 laws of ecology?
* Everything is connected
* Everything must go somewhere
* Nature knows best
* There is no such thing as a free lunch
54
What do autotrophs use as their carbon source?
carbon dioxide
55
How do autotrophs get their food source?
Fix CO2 into sugars
56
What do autotrophs use as an energy source?
• May use light as energy source (photoautotrophs) or may use chemical energy sources (chemoautotrophs)
57
Are autotrophs good for climate change?
Yes-
| • Autotrophs convert inorganic C to organic C, act as sinks for CO2 and hence act to limit climate change
58
Why is too much algae bad?
• Algae perform 50% of global photosynthesis
o But algae in large quantities such as where eutrophication from fertilizer run off occurs (where there is an excess of nitrate and phosphorus) is bad, as when the algae die the heterotrophs break them down and degrade them into CO2 and methane
59
What are methanogens?
Archaea
60
What do methanogens do?
o Consume CO2 and H2 and produces
| o Are chemoautotrophs- CO2 is the carbon source, and H2 is the energy source
61
Are methanogens good or bad for global warming?
o Impact on climate change- acts as sinks for CO2 (good) but act as sources of CH4, so overall are very bad
Methane has 20 times the warming potential of CO2
62
Under what conditions do methanogens die?
o Methanogens are anaerobic and are killed by oxygen
63
How do heterotrophs get their carbon sources and energy?
needs to eat other organisms, or other organic carbon sources; these also supply energy
64
Are heterotrophs good for climate change?
• Heterotrophs are sources of carbon dioxide --> bad for climate change
65
What are methanotrophs?
Bacteria
66
How do methanotrophs get their food?
o Consume methane and produce CO2
| o Are heterotrophs- methane acts as both their carbon and energy source
67
What are methanotrophs impact on climate change?
o Impact on climate change- acts as sinks for methane but acts as sources of CO2- overall good
68
Why are methanotrophs useful for removing pollutants?
o Methanotrophs are useful for removing methane, and they also attack other pollutants like trichloroethene (TCE)
Contains enzyme called MMO- Methane Monooxygenase Enzyme which has a broad substrate range
• Hence also works to clean up TCE
MMO takes molecular oxygen to methane and uses it to convert methane to a formaldehyde
Feed it methane so that is attacks pollution
• Accident metabolism
• Cleans up groundwater
69
What are decomposers?
o A key group of heterotrophs; recycle dead cells back to carbon dioxide
70
How do protists get their food?
```
o Protists (protozoa) are often predators of other microbes; these include ciliates, flagellates, amoebae
o Not all protists are predators, some are ‘detritivores’ (scavengers) and somre are photosynthetic
```
71
What s an autotroph?
An organism that is able to form nutritional organic substances from simple inorganic substances such as carbon dioxide
72
What is a heterotroph?
• Heterotroph-An organism deriving its nutritional requirements from complex organic substances
73
What is bioremediation?
The cleanup of pollution by microbes
74
What microbes are good for bioremediation?
o Hydrocarbon-degrading bacteria (including methanotrophs)
75
Describe the coral-algal symbiosis
o Corals are primitive animals (Phylum Cnidaria) which depend on symbiotic microscopic algae to supply them with food
o The coral get nutritional benefits from this mutualistic relationship through nutritional sugars, while algae get a safe place to live and get protection from environments
76
How do algae get their food?
o The algae are photoautotrophs, and convert CO2 and light to sugars
77
How do the coral get their food?
o Coral are heterotrophs, and convert sugars to CO2
78
What are lichen?
o Lichens are primary producers in some terrestrial habitats, especially in dry environments
o Not plants
o Symbiosis mutualistic relationship between the heterotrophic fungus and the autotrophic algae
o Two way nutrient symbiosis
o Algae are encroached to the surface and fungi feeds inorganic nutrients to algae
79
Why is biotechnology important?
```
• Food
o Fermentation
o Enzymes modify food
• Drink
o Synthetic yeast for wine making
• Medicine
o Tweaking microbes so they work better
• Agriculture
o GMO food engineered to resist herbicides
• Fuel
o Algae can make ethanol as end products enhance yield of molecules
• Cleaning
o Enzymes genetically engineered
```
80
Describe traditional biotechnology
• Biotechnology has been done by humans for more than 9000 years
o Fermentation to preserve foods or make alcohol
• Early fermentations used mixed cultures of naturally-occuring bacteria and fungi
• Earliest firm evidence for alcoholic fermentation also comes from around 7000BC
o Analysis of jars from China revealed traces of alcohol and the plants used for fermentation
81
What is the difference between cellular biotechnology and molecular biotechnology?
Cellular biotechnology
- Need some biology skills (especially microbiology)
- Don’t need an understanding of DNA, RNA and proteins
- When using a whole cell for your purposes
Molecular biotechnology
- Need high level biology skills (especially microbiology
- Need understanding of DNA, RNA and proteins
- When modifying and using DNA for your purposes
82
What are viruses used for in biotechnology?
Vectors to carry genes into new hosts
| Source of enzymes- e.g. T4 ligase that joins DNA together
83
What are archaea used for in biotechnology?
Source of thermostable polymerase enzymes for copying DNA sequences
Enzymes used for PCR
Live in extreme environments
84
What are bacteria used for in biotechnology?
Hosts for cloning DNA
| Hosts for expressing proteins
85
What are algae used for in biotechnology?
Conversion of carbon dioxide and light into biofuels such as ethanol and hydrogen gas
86
What are fungi used for in biotechnology?
Moulds- antibiotic synthesis e.g. penicillin
Yeast- excellent cloning and overexpression hosts
Very easy to quickly grow
87
What are the advantages of E.Coli bacteria over yeast Saccharomyces?
Fastest growth
Very easy to extract or add plasmid DNA
Lots of plasmids for it
Very easy to extract protein from
88
What are the advantages of yeast Saccharomyces over E.Coli bacteria?
Better for cloning and expressing eukaryote genes
Generally recognised as safe (GRAS)
Due to its increased similarity to eukaryotic genes, less likely to not function in other eukaryotic organisms
This is due to:
-Codon usage
-different preferences for codon usage, so between eukaryotic organisms, codon preference tends to remain the same so less likely codons will not be translated
-Post translation modification --> will be more likely to do same transformation to protein than eukaryote due to the fact that it is a eukaryote
-Better in the food industry
89
What is a plasmid?
circular DNA elements found in microbes; replicate independently of the chromosome(s)
90
What are plasmids used for?
Plasmids are the most commonly used vector for delivery of foreign DNA into a target host cell
91
What type of transfer is used to transfer plasmids between bacteria?
Lateral gene transfers
92
What are key features of the plasmid used for biotech and why are they useful?
```
o Selectable marker
Enables us to cells to take up plasmid
Amp
• Ampicillin resistance gene
Can be any selector gene
o Cloning site
Add foreign genes here
Restriction enzymes recognition
• Has to be specific sequence for recognition to cut
o Replication functions
Ensures persistence in host
Ori
• Allows replication to occur properly, so that the two daughter cells have the plasmid
```
93
How do you force a cell to take up a plasmid?
Electric/heat shock
94
What does cloning mean?
to make copies of a biological entity
o Creating identical organisms
o Making copies of a bit of DNA by adding it into a plasmid, then replicating the plasmid
95
What does DNA cloning mean?
Molecular biology technique that makes many identical copies of a piece of DNA
96
What does recombinant DNA mean?
DNA that has been formed artificially by combining constituents from different organisms
97
What does a genetically modified organism mean?
GMOs are organisms that have had their characteristics changed through the modification of their DNA.
98
What are the enzymes needed to make a GMO?
o Thermostable polymerase-Copying DNA
o Restriction enzyme-Cutting DNA
o T4 ligase-Joining DNA
99
How do you make a GMO?
1. Extract DNA from organism of interest
2. Copy the DNA
3. Digest the DNA into pieces
4. Digest the plasmid
5. Ligate the DNA and plasmid together
a. In the ligation mixture
i. Some plasmids that are recombinant (contain bits of foreign DNA)
ii. The starting plasmid (non-recombinant)
iii. Non-ligated bits of foreign DNA
6. Add the mix into cloning host (transformation)
7. Select plasmid- containing cells
a. E.g. plate on antibiotic agar
8. Screen to find gene of interest
a. Sequence-based screen
b. Phenotypic screen
9. Final product is a GMO
100
What are the risks of GMOs?
a. GMO risks
i. Depends on what foreign genes were added
ii. Antibiotic resistance genes may be transferred into pathogens
iii. Commercial risks- legal constraints and public perceptions
101
What are some examples of GMOs?
i. Golden rice- vitamin A
1. More nutritious
ii. Antibacterial genes in pathogens
1. Bad
102
What is a vaccine?
a primary defense against infectious disease: these save approx. 3 million lives every year
103
What do vaccines protect against?
Diseases that have no cure and which can only be fought through prevention e.g. rabies
104
How do vaccines work and what are they made of?
• Vaccines work by training the immune system to recognize antigens associated with an invader- they may consist of
o Live attenuated microbes
o Killed microbes
o Antigens (proteins) produced in a GMO host
105
How are vaccines made?
1. Isolate antigen gene (HBsAg for hepatitis B virus)
2. Selectively copy DNA through thermostable polymerase
3. Cloning antigen gene
a. Yeast plasmid and HBsAG are digested and ligated to form a recombinant plasmid
4. The recombinant yeast plasmid and the yeast cloning host are transformed so that the yeast is a GMO
5. The gene is expressed and the protein purified
• Yeast is most commonly used for vaccines
BIOL1997
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