MGY277 Final Exam Notes
Unit 1 – Perspectives
Unit 2 – The Bacteria
• Define the terms “magnification”, “resolution”, “contrast”, and “refraction” and apply them when looking at an image from
a microscope.
- Magnification: the increase in the apparent size of the object compared to the size of the actual object
- Refraction: light rays change direction due to change in medium (refractive index – measure of speed of light as it passe
through a medium)
- Resolution: minimum distance at which two points can be distinguished as individuals
- Contrast: the ability to see objects against the background
• Recite the steps of a Gram stain and why the stain can differentiate between different types of bacteria.
1. Crystal violet (primary stain) – bacteria stain purple
2. Iodine (mordant) – bacteria remain purple, cells less soluble now
3. Alcohol (decolourizer) – Gram-positive remain purple, gram-negative become colourless
4. Safranin (counterstain) – gram-positive cells remain purple, gram-negative cells appear pink
• Recall the different types of staining and how they are usefully applied.
- Gram stain: differentiate negative from positive gram cells
- Acid-fast stain: detect small group of organisms that don’t readily take up stain, i.e. Mycobacterium genus
- Capsule stain: capsules stain poorly, negative stain used, or India ink for suspension of carbon particles
- Endospore stain: Bacillus, Clostridium form dormant endospore, resist gram stain; use heat to facilitate uptake o
primary dye – often malachite green – by endospore, counterstain Safranin used to visualize other cells
- Flagella stain where flagella is for prokaryotic motility
• Know what the basic components of the Gram-positive and Gram negative cell envelope are
- Gram-positive: thick peptidoglycan, teichoic acid stick out above peptidoglycan, no outer membrane – hence no porin, n
LPS, sensitive to lysozyme
- Gram-negative: thin peptidoglycan, no teichoic acid, outer membrane – w/ porin, LPS, not sensitive to lysozyme
• Describe the outer membrane of the Gram-negative bacteria
- Outer membrane is unique lipid bilayer embedded with protein; porins and lipoprotein within outer membrane with lipoprotei
attaching to peptidoglycan
• Describe the structure of peptidoglycan: cell wall made of peptidoglycan, subunits N-acetylmuramic acid (NAM) & N
acetylglucosamine (NAG) form glycan chains; glycan chains are linked by tetrapeptide chains (4 AA strings)
- NAG found in mammals, NAG is a derivative of NAM, unique to bacteria
- Wall peptides are short and have unique “D” AA’s (D found in bacteria, L in proteins)
• Describe the basics of how bacteria obtain nutrients
- Trans-membrane highly specific proteins (transport systems) allows for movement of nutrients, small molecules, waste &
other compounds
- Facilitated diffusion: passive transport, not useful in low-nutrient environments & rarely used by prokaryotes
- Active transport: movement against [gradient], requires energy – Proton motive force & ATP (ABC transporter), commonl
used by bacteria
- Group translocation: transported molecule chemically altered upon entry via phosphorylation (i.e. glucose, where energy use
during transport is regained when sugar is broken down)
• Describe how bacteria sense their environment
- Membrane-spanning sensor kinase (SK), transfers a phosphate from ATP to response regulator (RR) – usually found as homo
dimer – RR is transcription factor which turns genes on or off in response
- RR controls response output – RR usually DNA binding protein changing affinity for DNA once phosphorylated
o I.e. Salmonella sense acid to recognize if within host, or sense O2 to regulate genes needed for anaerobic metabolism
• Describe a “typical” bacteria genome
- E. Coli strain CFT073 genome has about 5 million BPs, about 5000 genes
o Other E. coli strains have slight differences in genes they contain, due to frequent gene exchange amongst bacteria
- Contains anywhere from <500 to >8000 genes but only subset expressed at any given time
- Bacteria don’t have histones, introns or exons
• Describe bacterial gene regulation (i.e. how genes are turned on and off)
- Mechanism to control transcription: DNA-binding proteins & alternative sigma factors
- Alternative sigma factors: Standard sigma factor is component of RNA polymerase that recognizes specific promoters fo
genes expressed during routine growth conditions
- Alternative sigma factors can replace the standard factor & recognize different sets of promoters to control expression o
specific groups of genes (i.e. heat shock)
, MGY277 Final Exam Notes
Unit 3 – The Virus
• Describe the physical and genomic features of viruses.
- Viewed as DNA or RNA within protective coating, not alive outside of a cell (not metabolically active or replicating)
- Obligate intracellular parasites – reproduction reliant on intracellular resources
- Can be eukaryotic or prokaryotic (bacteria infected by bacteriophage – don’t infect humans)
- Difficult to study in lab: can’t be grown in pure culture, require live host, EM (expensive) to see
- Small size: 10 – 10000 x smaller than cells they infect (smallest is 17 nm diameter needs 2 genes to replicate), largest i
mimivirus – 2x larger than mycoplasma bacterium
- Virion: nucleic acid + protein coat (capsid), where nucleocapsid comprised of capsid containing genome
- Viruses have spikes for attachment to receptor sites & aid in entry to host cell
- Naked virus: Nucleoplasmid with capsomere subunits & spikes
- Enveloped virus: nucleoplasmid in matrix protein (immediate role during infection), surrounded by envelope w/spikes (oute
bilayer)
- Icosahedral: 20 flat triangles, efficient design using least energy to assemble
- Helical: spiral staircase like helix arrangement, can be short & rigid or long & filamentous
- Complex: intricate structures, i.e. phage w/ icosahedral head and long helical tail w/spikes & tail fibers
- Genomic features: DNA or RNA; linear or circular; double or single-stranded; always segmented; genome sizes vary, DNA
virus typically larger than RNA – large RNA are not stable
• Understand how viruses are classified and grouped, both formally and informally.
- ICVT 2009 report: > 6000 viruses, 2288 species, 348 genera, 87 families & 6 orders
- Formal classification based on: genome structure (ss/ds, RNA or DNA), hosts infected (bacteria, archea, animal, plant o
insect), viral shape & structure (enveloped or naked, icosahedral/helical, dimensions of capsid), disease symptoms
- Families end in –viridae, some names indicate appearance (coronae – crown like appearance), others named for geographi
area first isolated
- Genus ends in –virus (Enterovirus or Herpesvirus)
- Species name often name of disease, i.e. Poliovirus causes poliomyelitis
- Viruses commonly referred to only by species name, in contrast to virus nomenclature of genus, species
- Informal: Epstein-Barr for herpesvirus (Michael Epstein & Evon Barr discovered herpesvirus)
o Based on shared route of infection: Enteric viruses through fecal-oral route, Respiratory viruses through respirator
route, Zoonotic viruses animal to human transmission, Arboviruses spread by arthropods such as ticks & mosquitos
Arboviruses can infect widely different species, i.e. West Nile encephalitis, yellow fever, dengue fever
- Viral Taxonomy: Nucleic Acid (ds DNA, ss DNA, ds RNA or ss RNA), ss DNA & ds RNA are only naked, revers
transcribing viruses all enveloped
o Nucleic Acid > Outer covering > Family > Members (family have viruses w/ similar structural features, infect simila
host & cause similar diseases)
• 5-step infection cycle for enveloped and non-enveloped viruses.
1. Attachment: > 1 spikes binds specific receptor on plasma – specificity of spike to receptors accounts for resistance
2. Penetration & uncoating: Membrane fusion of enveloped virus, receptor-mediated endocytosis of enveloped + naked virus
a. Membrane fusion leads to nucleocapsid entering cytoplasm & envelope remaining with plasma membrane
b. Receptor-mediated endocytosis: enveloped virus enters intact with enveloped virus staying within endosoma
membrane. Naked virus cannot fuse with endosome of host membrane, must damage endosome & releas
nucleocapsid into cytoplasm
o Virion must localize to site of replication, may be nucleus or cytoplasm
o Uncoating of NA & disassembly of virus may occur simultaneously upon entry, or upon final destination
3. Synthesis of viral proteins & genome replication: Requires viral gene expression + replication
a. DNA viruses: usually replicate in nucleus, requires host machinery for gene expression & DNA synthesis, encode
own DNA polymerase (ss – or ss + to ds ± DNA to ss + RNA (mRNA) to protein)
b. RNA viruses: usually replicate in cytoplasm, require RNA polymerase replicase – enzyme catalyzing replication o
mRNA/+ ss RNA from RNA template (ss – RNA or ds ± DNA)
Replicase lacks proofreading ability – accounts for genetic mutations where pre-existing immunity ineffective
as seen in seasonal influenza (new vaccine needed due to mutations)
Segmented RNA viruses undergo re-assortment to alter host specificity, non-human to human transmission
c. Reverse transcribing viruses: encode reverse transcriptase which makes DNA from RNA; retroviruses have ss + RNA
genome, single ds ± DNA complementary strand synthesized => integrated into host genome (can’t eliminate)
4. Assembly: protein capsid must be formed; genome & enzymes packaged within capsid, occurs in nucleus or organelles, man
assemble near plasma membrane close to site of release
5. Release: enveloped virus released via budding (integrates into host plasma membrane upon budding & becomes coate
w/matrix proteins), naked virus released when host cell dies (apoptosis initiated by virus or causing lysis of cell during release
Unit 1 – Perspectives
Unit 2 – The Bacteria
• Define the terms “magnification”, “resolution”, “contrast”, and “refraction” and apply them when looking at an image from
a microscope.
- Magnification: the increase in the apparent size of the object compared to the size of the actual object
- Refraction: light rays change direction due to change in medium (refractive index – measure of speed of light as it passe
through a medium)
- Resolution: minimum distance at which two points can be distinguished as individuals
- Contrast: the ability to see objects against the background
• Recite the steps of a Gram stain and why the stain can differentiate between different types of bacteria.
1. Crystal violet (primary stain) – bacteria stain purple
2. Iodine (mordant) – bacteria remain purple, cells less soluble now
3. Alcohol (decolourizer) – Gram-positive remain purple, gram-negative become colourless
4. Safranin (counterstain) – gram-positive cells remain purple, gram-negative cells appear pink
• Recall the different types of staining and how they are usefully applied.
- Gram stain: differentiate negative from positive gram cells
- Acid-fast stain: detect small group of organisms that don’t readily take up stain, i.e. Mycobacterium genus
- Capsule stain: capsules stain poorly, negative stain used, or India ink for suspension of carbon particles
- Endospore stain: Bacillus, Clostridium form dormant endospore, resist gram stain; use heat to facilitate uptake o
primary dye – often malachite green – by endospore, counterstain Safranin used to visualize other cells
- Flagella stain where flagella is for prokaryotic motility
• Know what the basic components of the Gram-positive and Gram negative cell envelope are
- Gram-positive: thick peptidoglycan, teichoic acid stick out above peptidoglycan, no outer membrane – hence no porin, n
LPS, sensitive to lysozyme
- Gram-negative: thin peptidoglycan, no teichoic acid, outer membrane – w/ porin, LPS, not sensitive to lysozyme
• Describe the outer membrane of the Gram-negative bacteria
- Outer membrane is unique lipid bilayer embedded with protein; porins and lipoprotein within outer membrane with lipoprotei
attaching to peptidoglycan
• Describe the structure of peptidoglycan: cell wall made of peptidoglycan, subunits N-acetylmuramic acid (NAM) & N
acetylglucosamine (NAG) form glycan chains; glycan chains are linked by tetrapeptide chains (4 AA strings)
- NAG found in mammals, NAG is a derivative of NAM, unique to bacteria
- Wall peptides are short and have unique “D” AA’s (D found in bacteria, L in proteins)
• Describe the basics of how bacteria obtain nutrients
- Trans-membrane highly specific proteins (transport systems) allows for movement of nutrients, small molecules, waste &
other compounds
- Facilitated diffusion: passive transport, not useful in low-nutrient environments & rarely used by prokaryotes
- Active transport: movement against [gradient], requires energy – Proton motive force & ATP (ABC transporter), commonl
used by bacteria
- Group translocation: transported molecule chemically altered upon entry via phosphorylation (i.e. glucose, where energy use
during transport is regained when sugar is broken down)
• Describe how bacteria sense their environment
- Membrane-spanning sensor kinase (SK), transfers a phosphate from ATP to response regulator (RR) – usually found as homo
dimer – RR is transcription factor which turns genes on or off in response
- RR controls response output – RR usually DNA binding protein changing affinity for DNA once phosphorylated
o I.e. Salmonella sense acid to recognize if within host, or sense O2 to regulate genes needed for anaerobic metabolism
• Describe a “typical” bacteria genome
- E. Coli strain CFT073 genome has about 5 million BPs, about 5000 genes
o Other E. coli strains have slight differences in genes they contain, due to frequent gene exchange amongst bacteria
- Contains anywhere from <500 to >8000 genes but only subset expressed at any given time
- Bacteria don’t have histones, introns or exons
• Describe bacterial gene regulation (i.e. how genes are turned on and off)
- Mechanism to control transcription: DNA-binding proteins & alternative sigma factors
- Alternative sigma factors: Standard sigma factor is component of RNA polymerase that recognizes specific promoters fo
genes expressed during routine growth conditions
- Alternative sigma factors can replace the standard factor & recognize different sets of promoters to control expression o
specific groups of genes (i.e. heat shock)
, MGY277 Final Exam Notes
Unit 3 – The Virus
• Describe the physical and genomic features of viruses.
- Viewed as DNA or RNA within protective coating, not alive outside of a cell (not metabolically active or replicating)
- Obligate intracellular parasites – reproduction reliant on intracellular resources
- Can be eukaryotic or prokaryotic (bacteria infected by bacteriophage – don’t infect humans)
- Difficult to study in lab: can’t be grown in pure culture, require live host, EM (expensive) to see
- Small size: 10 – 10000 x smaller than cells they infect (smallest is 17 nm diameter needs 2 genes to replicate), largest i
mimivirus – 2x larger than mycoplasma bacterium
- Virion: nucleic acid + protein coat (capsid), where nucleocapsid comprised of capsid containing genome
- Viruses have spikes for attachment to receptor sites & aid in entry to host cell
- Naked virus: Nucleoplasmid with capsomere subunits & spikes
- Enveloped virus: nucleoplasmid in matrix protein (immediate role during infection), surrounded by envelope w/spikes (oute
bilayer)
- Icosahedral: 20 flat triangles, efficient design using least energy to assemble
- Helical: spiral staircase like helix arrangement, can be short & rigid or long & filamentous
- Complex: intricate structures, i.e. phage w/ icosahedral head and long helical tail w/spikes & tail fibers
- Genomic features: DNA or RNA; linear or circular; double or single-stranded; always segmented; genome sizes vary, DNA
virus typically larger than RNA – large RNA are not stable
• Understand how viruses are classified and grouped, both formally and informally.
- ICVT 2009 report: > 6000 viruses, 2288 species, 348 genera, 87 families & 6 orders
- Formal classification based on: genome structure (ss/ds, RNA or DNA), hosts infected (bacteria, archea, animal, plant o
insect), viral shape & structure (enveloped or naked, icosahedral/helical, dimensions of capsid), disease symptoms
- Families end in –viridae, some names indicate appearance (coronae – crown like appearance), others named for geographi
area first isolated
- Genus ends in –virus (Enterovirus or Herpesvirus)
- Species name often name of disease, i.e. Poliovirus causes poliomyelitis
- Viruses commonly referred to only by species name, in contrast to virus nomenclature of genus, species
- Informal: Epstein-Barr for herpesvirus (Michael Epstein & Evon Barr discovered herpesvirus)
o Based on shared route of infection: Enteric viruses through fecal-oral route, Respiratory viruses through respirator
route, Zoonotic viruses animal to human transmission, Arboviruses spread by arthropods such as ticks & mosquitos
Arboviruses can infect widely different species, i.e. West Nile encephalitis, yellow fever, dengue fever
- Viral Taxonomy: Nucleic Acid (ds DNA, ss DNA, ds RNA or ss RNA), ss DNA & ds RNA are only naked, revers
transcribing viruses all enveloped
o Nucleic Acid > Outer covering > Family > Members (family have viruses w/ similar structural features, infect simila
host & cause similar diseases)
• 5-step infection cycle for enveloped and non-enveloped viruses.
1. Attachment: > 1 spikes binds specific receptor on plasma – specificity of spike to receptors accounts for resistance
2. Penetration & uncoating: Membrane fusion of enveloped virus, receptor-mediated endocytosis of enveloped + naked virus
a. Membrane fusion leads to nucleocapsid entering cytoplasm & envelope remaining with plasma membrane
b. Receptor-mediated endocytosis: enveloped virus enters intact with enveloped virus staying within endosoma
membrane. Naked virus cannot fuse with endosome of host membrane, must damage endosome & releas
nucleocapsid into cytoplasm
o Virion must localize to site of replication, may be nucleus or cytoplasm
o Uncoating of NA & disassembly of virus may occur simultaneously upon entry, or upon final destination
3. Synthesis of viral proteins & genome replication: Requires viral gene expression + replication
a. DNA viruses: usually replicate in nucleus, requires host machinery for gene expression & DNA synthesis, encode
own DNA polymerase (ss – or ss + to ds ± DNA to ss + RNA (mRNA) to protein)
b. RNA viruses: usually replicate in cytoplasm, require RNA polymerase replicase – enzyme catalyzing replication o
mRNA/+ ss RNA from RNA template (ss – RNA or ds ± DNA)
Replicase lacks proofreading ability – accounts for genetic mutations where pre-existing immunity ineffective
as seen in seasonal influenza (new vaccine needed due to mutations)
Segmented RNA viruses undergo re-assortment to alter host specificity, non-human to human transmission
c. Reverse transcribing viruses: encode reverse transcriptase which makes DNA from RNA; retroviruses have ss + RNA
genome, single ds ± DNA complementary strand synthesized => integrated into host genome (can’t eliminate)
4. Assembly: protein capsid must be formed; genome & enzymes packaged within capsid, occurs in nucleus or organelles, man
assemble near plasma membrane close to site of release
5. Release: enveloped virus released via budding (integrates into host plasma membrane upon budding & becomes coate
w/matrix proteins), naked virus released when host cell dies (apoptosis initiated by virus or causing lysis of cell during release
