Biology of Microorganisms

Question 1

Microorganisms can be ___________. They can form _________ complexes (like phyloplankton) that are composed of a group of _________ cells. _____________ organisms consist of different cells with different functions.

Answer 1

unicellular, multicell, identical, multicellular

Question 2

What was the first organisms on Earth?

Answer 2

Microorganisms

Question 3

Microorganisms are everywhere!

How many in the human body? How many human cells?

How many microbial cells in a gram of soil?

How much of earth’s biomass is microbial?

Answer 3

4x10¹³ (human cells: 3x10¹³)

1 billion

10-20%

Question 4

Which domains of the tree of life do microbes exist on?

Which kingdoms?

Answer 4

All three: bacteria, archaea, and eukarya

Six: archaea, bacteria, protists, fungi, plantae, animalia

Question 5

Fungi

What task do they do in nature?

How many species? Give an example.

Answer 5

Decomposers: break down matter for plants/animals to reuse

100,000+ (Saccharomyces Cerevisiae - yeast)

Question 6

Plants (Algae)

What ties together these microbes?

What different forms are they found in? [4]

Give an example

Answer 6

That they are photosynthetic (using chlorophyll to capture light)

Single cells, filaments, colonies, diatoms

Chorella

Question 7

What is a lichen?

What do the separate bits do?

Answer 7

Symbiosis between algae and fungi

Algal part fixes carbon dioxide into organic carbon (nutrient for fungus), while fungus forms protective coating to guard it from external conditions

Question 8

Protozoa

What are they?

Give an example

What is one of the big differences between protozoa and bacteria?

Answer 8

Single celled predators and parasites (eukaryotes)

Amoeba

Protozoa are 1000 times larger (and also are eukaryotes, with organelles, as opposed to bacteria)

Question 9

Protozoa

Where are they often found in animals? What are they called?

What features do they have? [3]

Give examples

Answer 9

In the gut (called rumen protozoa)

Highly specialized, move rapidly (often ciliated), often engulf and degrade material (eg: plant stuff)

Entodinium, epidinium, diplodinium

Question 10

Archaea

What separates them from prokaryotes?

Where are they often found? What do they get called?

Where are they found in humans?

Answer 10

Different cell walls (no peptidoglycan) and membranes, eukaryote-like biochemical processes, and different rRNA and RNA polymerases (3 vs bacteria’s 1)

Extreme environments (salty, high temp, pH extremes, etc) = extremophiles

In the gut (methanogens = making methane)

Question 11

Bacteria

How many species?

How many RNA polymerases?

Which one is a model specimen?

Answer 11

5000+ (more all the time)

1

Escherichia coli

Question 12

What is the last type of microbe? [Not protozoa, fungi, algae, archaea, or bacteria]

How many of them are there per mL of seawater?

Answer 12

Viruses

50 million per mL

Question 13

In terms of microbial diversity, what are we talking about when we refer to inter-individual variation?

Answer 13

The degree to which each person’s microbiome has unique aspects (eg: 6 people had forearm swabs, 182 species identified, 98 were potentially new to science, and only 4 were common to all tested)

Question 14

What tools can we use to classify microbes? [5]

Answer 14

Cell ultrastructure (eukaryote/prokaryote), overall cell morphology, motility, physiological diversity (growth characteristics), and DNA sequencing

Question 15

What’s the difference between eukaryotic and prokaryotic/archaea microbes?

Answer 15

Eukaryotes have a nucleus and other internal membranes (with organelles)

Question 16

Fungal morphology: what is the difference between coenocytic hypha and septate hypha?

Why would they produce spores?

Answer 16

Spores are very hardy, and can survive where the hyphae would not (contains genetic info of fungus/bacteria, and can grow again when conditions are right)

Question 17

What’s the typical size of the prokaryote E coli? The Eukaryote yeast?

What is generally true about size in regards to prokaryotes and eukaryotes?

What’s the smallest microbe to be found? Largest?

Answer 17

1-3uM, 10uM

Prokaryotes are (usually) smaller

0.5uM (archaea nanoarchaeum equitans), 750uM (bacteria thiomargarita namibiensis)

Question 18

What are the most common shapes for bacteria?

Other shapes?

What is the word for a bacteria that changes shape? Stays the same?

Answer 18

cocci (grapes) and bacilli (rods)

square, comma-shape (vibrio), spiral

Pleomorphic, monomorphic

Question 19

What is the name of the stain to identity bacterial cell wall morphology?

Colours?

Why do they have the colours that they have?

Answer 19

Gram stain

Red/pink -, blue/purple +

Gram negative cells have an outer membrane (beyond the cell wall) - this stops the stain getting to the peptidoglycan

Question 20

Bacteria nutrition: what does heterotrophic mean?

What do you call bacteria that require inorganic carbon sources (like carbon dioxide)?

Answer 20

Requires an organic food source

Autotrophic

Question 21

DNA sequencing of microorganisms: which gene sequences are often used? Why?

Which organism does this fail to detect? Why?

Answer 21

Ribosomal RNA gene sequences (16S rRNA) - they all have it, it’s essential for growth, there are several copies per cell. In addition, there are eight conserved regions (used for primer design), and nine variable regions (to compare with)

Often fails to detect archaea (different conserved regions)

Question 22

What is binary fission?

Are bacteria haploid or diploid?

Answer 22

Process by which microbes (often prokaryotes) grow, replicate DNA, and divides into two cells

Haploid (just one set of genes)

Question 23

What is vertical gene transfer?

What is horizontal gene transfer? What types are there?

Answer 23

Process by which daughter cells receive exactly the same chromosomal genetic material

Horizontal: exchange of genetic material from one cell to another that is not its offspring (transformation = picking up naked DNA, conjugation = connecting via pilus, transduction = via bacteriophage)

Question 24

What shape are prokaryotic chromosomes?

Where does DNA replication start? How does this work?

What are the smaller extrachromosomal circles of DNA called? What is usually encoded here?

Answer 24

Circular

Origin of replication (replication fork then goes bidirectionally)

Plasmids (nonessential but useful functions)

Question 25

What is transformation? What state must the cell be to undertake it?

Describe Fred Griffith’s and Oswald Avery’s experiments on streptococcus pneumoniae

Answer 25

Uptake of free DNA (the cell must become ‘competent’ - the cell membrane is slightly porous, allowing DNA to get in)

Fred noticed that S form (with capsule) killed mice, whereas R (rough form - no capsule) did not. When he mixed heat-killed S form with alive R form, the mice died. Oz did the latter, but also included various components to degrade cellular material (lipase, RNase, protease, DNase) - only DNase prevented the killing, and so he proved that the transforming substance was DNA

Question 26

What is conjugation?

What types of bacteria usually undertake this process?

What is usually transferred?

What was Lederberg and Tatum’s experiment to show this?

[NB: bacillus anthraxis (capsule = pathogenicity)]

Answer 26

Transfer of DNA by a protein structure called a pilus [F/fertility plasmid, allowing for conjugation, can also be passed]

Related species

Plasmids and transposable elements

Two related bacteria, each with opposing metabolic weaknesses - neither grows on a minimal medium alone, but if they are mixed before plating you do get a few colonies (so genetic information is transferred between bacteria)

Question 27

Transduction - what is it?

How does it occur?

The causative organisms can be _______ or _________. What does this mean?

Answer 27

Transfer of DNA between bacteria via bacteriophages

When bacteriophages are being packaged up, part of the actual bacteria genome, or a plasmid, is packaged instead of the viral genome (when temperate phage excises itself from genome). The phage darts off anyway and injects this into another bacteria.

Virulent (always lytic cycle), temperate (lysogenic, but can enter lytic cycle)

[Example: corynebacterium = causative agent of diptheria. Toxin gene is transmitted by phage]

Question 28

What is the name for the amount of time it takes for a bacterial cell population to increase in size 2x?

How long for E coli (faster) and mycobacterium leprae (slower)

What environmental factors can contribute? [4]

Answer 28

Doubling time/generation time

20 minutes, 20 days

Nutritional availability, temperature, pH, osmolarity

Question 29

E coli doubling time is 20 minutes, but the time it takes to replicate DNA is 40 minutes - how does this work?

Answer 29

Fast growing bacteria have multiple cycles of DNA replication in progress (multiple replication forks at once)

Question 30

_____ is an increase in the number of cells in the population

_____ ____ is change in cell number per unit time

__________ ____ is time required for cell number to double

Answer 30

Growth

Growth rate

Generation time

Question 31

What are the five typical phases of prokaryote growth cycle?

Answer 31

[NB: Absorbance on Y axis]

Question 32

Five phases of prokaryote growth cycle (lag, exponential, deceleration, stationary, death): what is happening in each phase?

Answer 32

Lag: adjustment of cell metabolism to conditions

Exponential: cell growth at max. rate possible in medium

Deceleration: limiting supply of nutrient(s) or more toxins

Stationary: no more growth and some dying cells

Death: decline of viable cells due to nutrient deficit or toxicity

Question 33

What type of graph paper do you plot the growth rate of prokaryotes during exponential phase? What does this allow for?

Using this graph, how would you calculate the maximum growth rate?

Answer 33

Semilog - allows for easy calculation of doubling time

u = ln(cells_t2-cells_t1)/(t₂-t₁)

NB: t2-t1 = generation time (h or min)

Question 34

What is aseptic technique? How do you achieve this?

How would you deal with anaerobic bacteria?

Answer 34

Exclusion of contaminants - culture media sterilized, and all materials contacting media must be kept sterile

Either using a carbon dioxide hook to get the air out of tubes (as it is more dense), or using an anaerobic cabinet

Question 35

What type of plates (3) and cultures (3) are there?

Answer 35

Question 36

What methods are there for measuring growth? [NB: not growth rate]

Answer 36

Cell number (cell counts, viable counts)

Cell molecules (total dry mass, protein, DNA, etc)

Light scattering (optical density)

Question 37

Measuring cell growth: how do you do a viable count?

What about microscopy count?

Answer 37

Culture will have a specific optical density (eg: E Coli OD650 = 1.6 = 2x10^9 cells/mL). So then do a dilution series, and then place the appropriate dilutions on an agar plate. Each viable cell will form one colony (forming unit). Then just count the number of cells in one quarter of the plate, multiply by four, and then calculate based on the dilution factor…

Use a microscope with a grid square. Count the number per grid, and then multiply by a factor (determined by the volume of the grid) = cells/mL

Question 38

Measuring cell growth: how does light scattering work?

What are the potential issues? WHat can be done?

Answer 38

Light comes in, hits off cells, and then goes to a detector - the more cellular matter you have, the more light scattering you will have, and the higher optical density you will have

Cell density might be too high or too low. For low, let it grow longer. For too high, use dilutions… or use a different method

Question 39

Measuring cell growth: what’s the only method for measuring live cell numbers?

What is useful for measuring growth rates?

What is FISH?

Answer 39

Viable count

Light scattering

Fluroscent in situ hybridization - count specific bacterium in a sample of many other bacteria [uses DNA probe, so doesn’t count any burst cells]

Question 40

What is the difference between a closed and open system?

Disadvantages of batch/closed? Advantages of open/chemostat?

Answer 40

Batch: no addition after innoculation, growth rate varies, medium composition varies, difficult to change growth rate

Chemostat: continuous addition of medium, constant volume, consistent environment, can vary growth rate

Question 41

Growth rate is affected by pH. Most bacteria are __________.

What does this mean for food preservation?

Answer 41

Neutrophilic (like neutral conditions)

Weak acids get used as preservatives, as this will combat most forms (neutral and basic) of bacterial infection

Question 42

Growth rate is affected by temperature. Most bacteria are __________. What are other categories?

Answer 42

Mesophiles (avg 39)

Psychrophile (avg 4), thermophile (avg 60), hyperthermophile (avg 88)

Question 43

Growth rate affected by osmolarity. What categories are there?

Answer 43

Nonhalophiles (very low salt tolerance - e coli), halotolerant (low-medium - staph aureus), halophiles (medium-high - vibrio fischerii), extreme halophiles (high-silly - halobacterium salanarum)

Question 44

Why is it important to understand the nutrition of microbes? [3]

Answer 44

Environmental cycling of major elements (N, S, C) dependent upon microorganism activity

Industrial applications: microorganism ability to derive energy from chemical oxidations

Biotechnological applications: ability to grow on different media is important (eg: oil spill remediation)

Question 45

Which two functions do nutrients serve to microbes?

Answer 45

Used as building blocks for construction of cell components (amino acids <> proteins, sugars <> polysaccharides, nucleotides <> DNA/RNA, fatty acids, metal ions [for cofactors, osmoregulators, and structural stability])

Supply energy required to drive cellular processes (polymerization of monomers, synthesis of monomer units from simple nutrients, maintenance of osmotic balance, degradation to create ATP)

Question 46

What is anabolism and catabolism?

Answer 46

Anabolism: biochemical processes involved in synthesis of cell constituents from simpler molecules (requires energy)

Catabolism: biochemical reactions leading to the breakdown of complex to simple molecules and the production of energy

Question 47

How many phosphate-phosphate high energy bonds does ATP have? What is the main reason for them being high energy?

Answer 47

Two (repulsion of negative charges)

Question 48

Basic composition of medium for growth of microorganisms: what is essential? What might need to be added? [3/5]

Answer 48

Essential: water, carbon source, nitrogen source

Added: sulphur source, phosphorus source, inorganic salts in macro amounts (Na, K, Mg), trace inorganic salts (Cu, Mn, Co, Zn, Ni), source of energy (light, oxidation of organic compounds)

Question 49

Carbon Source: what is an organism called if it uses carbon dioxide? What if it can be organic molecules?

What are most bacteria?

Answer 49

Autotroph, Heterotroph

Heterotrophs

Question 50

Nitrogen source

Typical bacterium is what % nitrogen? Where in cell is it present?

Where do most bacteria obtain nitrogen from?

Where do others (eg: cyanobacteria) get it from? What specific cell does the cyanobacteria use to do this?

Answer 50

12% (in proteins, nucleic acids, and cell wall)

As ammonium ions (NH₄^-) or nitrate (NO₃^-)

They fix atmospheric nitrogen to ammonia in heterocysts

Question 51

Phosphorus source: where is it obtained? What is it used for?

Sulphur source: where obtained? What is it used for?

Answer 51

Phosphate ions (PO₄^3-) - synth: nucleic acids and phospholipids

Sulphate (SO₄^2-) or sulphide (S^2-) - structural role in 2 amino acids and some vitamins

Question 52

Other macronutrients: what is magnesium used for? What about calcium?

What about micronutrients?

What are ‘growth factors’ in a nutrient sense?

Answer 52

Mg: stabilizes ribosomes, cell membranes, and needed for enzyme activty, Ca: stabilizes cell wall & endospore heat stability

Usually required by enzymes

Low molecular weight organic compounds (vitamins, amino acids, purines, and pyrimidines)

Question 53

Culture media can be ______ or _____. What is each one best for?

Answer 53

Liquid: good for growing large quantities of microbes, Solid: needed for growing some microbes, and very few can degrade agar (as it is not a carbon source)

[NB: Agar is seaweed extract. Usually added to liquid media at 1.5% w/v. Solid@40 degrees centigrade]

Question 54

What is complex media? When is it often used?

What is defined media?

Answer 54

Complex: composition of media not completely known (inexpensive, lots of shit grows on it, can contain lots of different waste products like animal blood, inexpensive organic materials, etc). Often used to culture organisms from clinical samples.

Defined: composed of known quantities of chemically pure organic and/or inorganic compounds

Question 55

What are the classifications for organisms by source of carbon? [2]

Classifications for organisms by source of energy? [2]

Answer 55

Autotrophs (bulk of carbon from carbon dioxide), heterotrophs (bulk of carbon from organic sources)

Phototrophs (use light energy), chemotrophs (use energy released by redox reactions)

Question 56

Cyanobacteria derive energy from light and most of their carbon from carbon dioxide. Which nutritional category are they?

Purple non-sulphur bacteria derive energy from light and bulk of carbon from organic sources. They are…?

Nitrosomonas europaea derive energy from redox reactions and carbon from carbon dioxide. They are…?

Most bacteria, all fungi, and all protozoa derive energy from redox reactions and carbon from organic sources. They are…?

Answer 56

Photoautotrophs

Photoheterotrophs

Chemoautotrophs [NB: doesn’t have to be carbon - nitrogen for this one, fe2+ to 3+ for acidithiobacillus]

Chemoheterotrophs

Question 57

How could acidothiobacillus thiooxidans be used for bioleaching? What’s the downside?

Answer 57

Uses inorganic sources to make energy. Pyrite (Iron and sulphur) is often present in mine waste, so it can be used to separate valuable materials… but does generate sulphuric acid and iron III (rust) = shit for environment

Question 58

What is a Winogradsky Column?

Answer 58

Take pond water, add carbon [sawdust, sodium bicarbonate, shredded newspaper, etc], add sulfur [ground eggshell and egg yolk], shake, and leave for 3-5 months = end up with very defined growth bands (specific microbes grow where it is best full of interdependent communities) [also works the same in gastro-intestinal tract: aerobic -> anaerobic, increasing bacterial density, etc]

Question 59

Type of energy/nutrition of gut bacteria? Aerobic or anaerobic?

How much energy do humans take from high sugar crap? Where is it absorbed from? What about healthier/fibrous stuff? What does this mean for gut bacteria?

Answer 59

Anaerobic Chemoheterotrophs

>70% (stomach and small intestine), 5-10%. There’s a reason why most bacteria live in the lower parts of the tract - and you can starve them if you don’t eat enough healthy stuff

Question 60

What do bacteria use to move? What protein are they composed of? Where is the “motor” found?

Because it’s a protein that extrudes out of the cell, they are ______________

Answer 60

Flagella (composed of flagellin) - complex protein motor is embedded in plasma membrane

Immunoreactive

Question 61

Flagella arrangements:

Size (length and width)?

Types? [4]

Answer 61

5-20um long, 0.01-0.02um wide

Monotrichous (one on one end), amphitrichous (one on each end), lophotrichous (tuft on one end), peritrichous (all around cell)

Question 62

Flagellum has how many sections? What are they?

What shape is the flagellum? What type of rotation produces forwards movement? And what type produces tumbling?

How many genes required for synthesis?

Answer 62

3: long filament (made up of flagellin), hook, basal structure (converts chemical energy into rotary motion)

Helical (corkscrew) - anticlockwise, clockwise

>40

Question 63

What do microorganisms respond to in terms of motility? What is this called?

Where are the receptors for this process found?

Movement in peritrichous (flagellae all around) bacteria is made up of two types of action: ____ and _______

Answer 63

Chemicals in environment (chemotaxis) - towards attractants, and away from repellents

Chemoreceptors in the cell membrane

Runs and tumbles

Question 64

Bacterial chemotaxis usually involves runs and tumbles (in terms of the type of movement). What does this look like?

How is it different when there is no attractant or repellent?

Answer 64

No chemotaxis = random looking movement hither and thither

Question 65

How does motility increase bacterical pathogenicity? How do we know this is the case?

Answer 65

Helps them move towards the things they need to attach to, and can also help with the attachment itself (and we know because not-motile bacteria [such as mutant vibrio cholerae] are less virulent than their flagella-having wild types)

Question 66

How can flagella be used in diagnostic microbiology? [2]

Answer 66

Because they are immunoreactive - to help identify strains, cell surface structures like this (that elicit an immune response) can be used. For example: E coli O157:H7 has distinct lipopolysaccharide antigens (O) and flagella antigens (H)

Can also help identify bacteria by doing a motility test (stick pin w/bacteria into semi-solid agar - motile bacteria will spread away from the line of inoculation) [eg: pneumonia - klebsiella pneumoniae non-motile, pseudomonas aeruginosa motile]

Question 67

In detecting motility, you can do a line inoculation in semi-solid agar. What indirect method could you use?

Answer 67

Stain for flagella (extra width, fluorescence, etc) and phase contrast light microscopy, or use electron microscopy

Question 68

What do the following refer to:

Aerotaxis

Phototaxis

Magnetotaxis

Magnetosomes

Answer 68

A: moving to/away from oxygen

P: moving to/away from light

M: orientation/movement along magnetic force lines

M2: crystals of magnetite (Fe₃O₄) that allow for magnetotaxis

Question 69

What can drive motility in eukaryotes?

What is the structure of a eukaryotic flagella? Length/width? How do cilia differ?

Which group of protozoa are cilia restricted to?

Answer 69

Flagella and cilia

Nine peripheral mictrotubule pairs (composed of tubulin [flagellin in bacteria]) around central pair - 40um x 0.15-0.3um (diff length)

Ciliates (many rumen bacteria)

Question 70

Cell movement: cilia vs flagella…

Length?

Number?

Type of movement?

Speed of movement?

Seen in?

Answer 70

short vs long

100s vs 1-8

complicated 3D vs wave-like/undulating

Fast vs slow

eukaryotes vs eukaryotes/prokaryotes

Question 71

How many groups can protozoa (eukaryotes) be grouped into on the basis of motility? Names?

Answer 71

Four: flagellates (move by flagella), amoebas (cytoplasmic streaming - propelled by movement of cytoplasm into “pseudopodia”/”hands”), ciliates (move by cilia - paramecium, vorticella, etc), and sporozoans

Question 72

How do bacteria communicate with each other?

What are the molecules bacteria release called? What responds?

What is quorum sensing? Can you think of an example (and the specific autoinducer)?

Answer 72

Emitting and reacting to chemical signals

Autoinducers (bacteria of the same species)

It is when enough of a cell population is present that autoinducer molecules accumulate, allowing each cell to sense how many other cells there are, and to coordinate activity. [Example: vibrio fischerii - luminescent bacteria whose glow is triggered by QS of autoinducer homoserine lactone]

Question 73

Vibrio fischerii bioluminescence (autoinducer serine lactone): concentration in normal seawater? In light organs of fish?

Higher concentration of this bacteria activates the enzyme _________ and oxidises the organic compound ________

Answer 73

10² cells/mL, > 10¹⁰ cells/mL

luciferase, luciferin(s)

Question 74

Biofilms (on teeth): how is this another example of quorum sensing?

What about bacterial toxins?

Answer 74

Encasement of microbial cells in matrix polysaccharide (plaque) occurs as the concentration of autoinducer signalling molecules increases (attached monolayer -> microcolony -> mature biofilm)

Some (eg: salmonella) wait until cell density is sufficient befoire releasing the toxin that sickens their host (makes it less likely the immune system will destroy the pathogen as its population is increasing)