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90 Cards in this Set

  • Front
  • Back
What are the main differences between bacterial and eukaryotic cells?
1. Organization
2. Reproduction
3. DNA association
4. Presence of lysosomes?
5. Ribosome?
6. Peptidoglycan?
1. Bacteria have no internal membranes or organelles whereas Eukaryotes have a nucleus and complex internal membranes.
2. Bacteria reproduce by binary fission, Euk via mitotis.
3. Bacteria-no histones or nuclear membrane with one circular chromosome whereas Eukaryotes have more than one chromosome and it's linear.
4. Bacteria-no lysozomes because they are membrane-bound.
5. Bacteria: 70S
Eukaryotes: 80S
6. Bacteria have a cell wall with peptidoglycan, Euk's don't
1. What is the composition of the bacterial cell wall?

2. What is it's purpose?

3. Where and what are the cross-linked peptides?
1. Peptidoglycan-
It is common to both Gram + and Gram - but thicker in Gram + and is on the inner layer of the cell wall.
Comprises of long chains of (sugars) NAM and NAG cross-linked by peptide side-chains and cross-bridges.

2. Purpose: gives rigidity and shape to the organism. It protects against osmotic pressure.
The peptidoglycan wall is the site of action of penicillins, cephalosporins and is degraded by lysozyme.

3. Transpeptidases form the cross-linked peptides on the NAM sugars
What forms endotoxin in bacterial cell walls?
LPS is the endotoxin so it's only specific to Gram - cells. It is the integral component of the outer membrane.
The O antigen is the outer portion of LPS with multiple repeating subunits (good for use in diagnosis) and the Lipid A (fatty acid) is the core component of which is toxic in the LPS.
A core polysaccharide separates the O antigen from the Lipid A fatty acid.
Where do you find the periplasmic space?
Only in Gram - bacteria because it is between the outer membrane and the cytoplasmic membrane.
Which type (Gram + or Gram -) has thick Murein?
Gram +
The cell wall is 80% of it's dry weight.
(Whereas Gram- it's only 10-20%)
How does transfer of genetic material occur in bacteria (3)?
1. Conjugation
2. Transduction
3. Transformation
What are the steps involved in a gram stain? (5)
1. Fix cells on slide
2. Crystal Violet
(b/c it passes through all cell walls and makes all cells appear blue)
3. Iodine solution
(forms complexes with crystal violet to make an insoluble dye)
4. Wash/dehydrate with mild solvent (ethanol) to wash out CV-I complex
-B/c thin walls of Gram - allows for CV-I complex to leave, it appears clear
-Thick walls of Gram+ retains blue dye
5. Add Safranin stain
-stains gram - cells red/pink
What acids are present in bacterial inner membranes (gram +)?
1. Teichoic acid (1st thing you hit on surface of bacteria). It's a surface antigen.
2. Lipoteichoic acid-goes all the way to the lipid bilayer
What ribosome does bacteria have?
70S
The subunits are 50S and 30S
What is the chemical composition and function of a capsule?
The capsule of bacteria is made of polysaccharide and it protects against phagocytosis. It isn't present in all bacteria. It also mediates adhesion to human tissues for colonization.
What is the chemical composition and function of a spore?
Bacteria can sporulate when nutrients like Carbon and Nitrogen are low. They get a thick, keratin-like coat (Dipicolinic acid) that make them resistant to heat, dehydration, radiation and chemicals.

Not all bacteria have.
What is the chemical composition and function of Pilus or fimbria?
Composed of Glycoprotein.
Two types:
1. mediates attachment to cell surfaces
2. Sex pilus mediates attachment of bacteria during conjugation.
What is the chemical composition and function of flagella?
Protein ( have a basal body with a central rod, an L, P, S, M ring and a hook/filament coming out of it. It is a 360 degree Hook/Rod motor)
Motility
-There are receptors that push them away or push towards a source
What are the 4 main phases of bacterial cell growth?
1. Lag phase: upon initial entry into environment. You don't see rapid growth right away because of a period of adaptation with vigorous metabolic activity.

2. Exponential growth (Log): until environment becomes limiting

3. Stationary phase: when bacteria reach maximum cell yield b/c of depletion of sources

4. Death/Decline phase:
unfavorable conditions
What are Bacteria considered as far as their nutritional requirements?
Heterotrophs:
-utilize complex organic substances
1. Obligate aerobes
2. Facultative anaerobes
3. Obligate anaerobes
4. microaerophiles
What enzymes do strict anaerobes (obligate) lack? What energy source do they use?
Superoxide dismutase and catalase

They ferment or use nitrate, sulphate, etc. for electron acceptor.
What internal factors limit growth in bacteria (2)?
1. Metabolites (end-products)
2. Catabolites
What external factors limit growth in bacteria? (5)
1. Adherance
2. Food
3. pH
4. Temperature (mesophiles affect us)
5. Oxygen
What is the main enzyme that mediates DNA replication?
DNA-dependent DNA polymerase
There are 3 stop codons and 1 start codon, what are they?
Start: AUG (Met)

Stop: UAA, UAG (Tyr) and UGA (Trp)

The genetic code is unambiguous, degenerate, and universal
What enzyme covalently attaches an amino acid to a tRNA molecule?
aminoacyl-tRNA synthetase
There are 2 sites on the ribosome, one that is growing and one that the new incoming tRNA gets added to. What are they?
The 30S subunit is associated with the mRNA and the 50S subunit is associated with the peptidyl-t-RNA site (one growing) and the Aminoacyl-tRNA site (incoming one) .
What is transformation referring to as a source of genetic exchange in bacteria?
Transformation: gene transfer by uptake of exogenous bacterial DNA(naked DNA) by another bacterium in the vicinity.
-It is not mediated by a plasmid or phage
-Active process (specialized machinery)
-Horizontal transfer
-small amount of DNA
-Competent
-many times when dying bacteria release bacteria
-
What happens if you don't get re-combination of new donor DNA?
It's an unsuccessful transfer, no uptake is really occurring.
What is transduction gene transfer?
Transduction is a process of DNA transfer by means of a bacterial virus (bacteriophage).
-mediated by bacterial virus
-error in viral life-cyle
-Two types exist:
1. Generalized: any fragment of bacterial chromosome can be transferred
2. Specialized: only genes next to insertion site are transferred

-can get 1-2% of the genes this way-horizontal transfer
What are the 4 most common toxins due to a prophage?
1. Cholera
2. Botulism
3. Diptheria
4. Pyrogenic toxin: Scarlet fever erythrogenic factor
What are plasmids?
Plasmids are: extrachromosomal, double-stranded, circular DNA molecules.
-Replicon: self-replicating abilities
-Can be multi-copy
-Encode toxins, resistance to antibiotics, heavy metals
-Are: 1. Conjugative (transmissible)-sex pilus (F+)
or
2. Non-mobile: (non-transmissible), these are smaller than the transmissible ones; F-
-Can be identified in almost all strains of bacteria tested
-
What is conjugation gene transfer?
Conjugation involves transfer of a plasmid gene. It is the mating of 2 bacteria, during which DNA is transferred from the donor to the recipient cell.
-It has the highest potential for largest amount of genetic info transfer; horizontally.
-Need physical contact between cells involving a sex pilus and is controlled by the F plasmid that carries the genes for the proteins required for mating.
-Rolling-circle method replication
What is the clinical relevance of plasmids?
They can be antibiotic-resistant: those carrying the R plasmid)
-Or they can be resistant to heavy metals, exotoxin production and pili formation
What are transposons?
Transposons (jumping genes) are pieces of DNA that move readily from one site to another, either within or between the DNAs of bacteria, plasmids and bacteriophages.
-They can jump from:
1. chromosome-->plasmid
2. plasmid-->plasmid
3. plasmid-->chromosome
-Unlike plasmids or bacteriophages, transposons cannot replicate independently of of recipient DNA.
-can be conservative or duplicative in mechanism
-can lead to mutations
What is a composite transposon and why is it significant?
Composite transposons are flanked by two separate IS elements which may or may not be exact replicas. Instead of each IS element moving separately, the entire length of DNA spanning from one IS element to the other is transposed as one complete unit. Composite transposons will also often carry one or more genes conferring antibiotic resistance.
What is an IS element?
An IS element is a short DNA sequence that acts as a simple transposable element. Insertion sequences have two major characteristics: they are small relative to other transposable elements (generally around 700 to 2500 bp in length) and only code for proteins implicated in the transposition activity (they are thus different from transposons, which also carry accessory genes such as antibiotic resistance genes). These proteins are usually the transposase which catalyses the enzymatic reaction allowing the IS to move, and also one regulatory protein which either stimulates or inhibits the transposition activity. The coding region in an insertion sequence is usually flanked by inverted repeats.
What is a mutation?
What are the three types?
A mutation is a change in the base sequence of DNA, as a consequence of which different A.As are incorporated into a protein-resulting in an altered phenotype.
-It is a heritable, sudden change
-It refers to both the change and the process by which it occurs.
*3 types:
1. Base substitution
2. Insertion (type of frameshift)
3. Frameshift mutations
What is the result of a base substitution (mutation)?
1. You can get a missense mutation where a different amino acid is inserted. This results in an A.A. change or a silent change.
or
2. You can get a nonsense mutation where the base substitution generates a STOP codon and this destroys the protein function. (UAA, UGA, UAG)
What is a frame shift mutation?
It occurs when 1 or more base pairs are added or deleted, which shifts the reading frame on the ribosome and results in the incorporation of the wrong amino acid downstream from the mutation and results in an inactive protein.
What is an insertion mutation?
Insertion of additional pieces of DNA (eg: transposons) or an additional base can cause profound changes in the reading frames of the DNA and in adjacent genes.
Mutations can be spontaneous and/or induced, how?
1. Spontaneous: mainly by errors in replication ( 1/10 million base pairs)
You can see the tautomeric shift where the normal amino form is converted to the imino form with rare A-C base pairing.
or
2. Induced by the environment-
this is at a higher frequency and usually depends on exposure to a mutagen (chemicals, radiation, virus)
What type of mutation does UV light cause?
It's an induced mutation that can cause thymine-thymine dimers (cross-linking)
What is the difference in transition and transversion induced mutations?
1. Transition mutation-it is the most common and it is replacing a purine with a purine (A, G) or a pyrimidine with a pyrimidine (C, T).

2. Transversion-not as common, replace a purine with a pyrimidine or a pyrimidine with a purine
What is the lytic cycle?
The lytic cycle is one of the two cycles of viral reproduction, the other being the lysogenic cycle. These cycles should not, however, be seen as separate, but rather as somewhat interchangeable. The lytic cycle is typically considered the main method of viral replication, since it results in the destruction of the infected cell.
What is the lysogeny cycle?
Lysogeny in prokaryotes is characterized by integration of the bacteriophage nucleic acid into the host bacterium's genome. The newly integrated genetic material, called a prophage can be transmitted to daughter cells at each subsequent cell division, and a later event (such as UV radiation) can release it, causing proliferation of new phages via the lytic cycle.
What is gene cloning?
It is the artificial incorporation of one or more genes into the genome of a new host cell by various genetic recombination techniques.
-cut and paste
-amplify genes of interest
What is the sequence of cloning?
1. Candidate DNA is extracted from source, purified and cut or cleaved into small fragments with restriction enzyme (BamHI)-"sticky ends".
2. Cleave plasmid (vector) with BamHI
3. mix cut (sticky ends) genomic and plasmid DNA
4. Covalently seal recombinant DNAs with LIGASE.
5. Transform into bacteria
6. Select amp resistant/tet sensitive colonies
Why use PCR (polymerase chain reaction) for gene cloning techniques?
What are the three things you need for a PCR? What enzyme do you use and why?
Because you can exponentially amplify DNA in a short time.
-It is sample-limiting however because the gene is present in low copy number but is great for rapid clinical diagnosis.

*3 things needed:
1. Taq DNA polymerase
2. Primers (that flank gene) with dNTP
3. Template DNA
-You use Taq DNA polymerase. This enzyme is very heat stable.
1. What temperature do you denature the protein?
2. Anneal the primer?
3. Extend/DNA synthesis?
4. How many cycles?
1. 94 degrees C-denature protein
2. 55 degrees C-anneal primer
3. 72 degree C-syntesize
4. 25-30 times
What are the methods used in a southern blot?
1. Gel
2. Probe
3. What you are after
In a southern blot-
1. Gel=DNA
2. Probe=oligonucleotide (large), use to find DNA of complementary sequence
3. Trying to see if a gene is present/organism exists
What are the methods used in a northern blot?
1. Gel
2. Probe
3. What you are after
Northern Blot:

1. Gel=mRNA
2. Probe=oligonucleotide
3. Looking to see if a gene is expressed.
What are the methods used in a western blot?
1. Gel
2. Probe
3. What you are after
Western blot:

1. Gel=SDS, protein
2. Probe=antibody
3. Looking to see if a protein is produced.

The detergent SDS gel unfolds proteins and binds uniformly so they get a net negative charge.
What is a an array hybridization?
It is a type of northern blot that looks at the expression of hundreds of genes at once. (gene expression)
-Oligonucleotides to each gene are arranged on the blot
-Probe with cDNA (mRNA)
What type of blot/technique would you use to determine the proteomics of p. gingivalis?
Do an array hybridization (northern blot) to the: conditioned epithelial culture medium or a control culture medium.
What do stem cells and cancer cells use to maintain telomere length and keep replicating?
Telomerase
How many doublings do normal human cells do to limit replication while telomeres keep getting shorter?
60-70 doublings
Tumors maintain sustained angiogenesis or they die after how big?
1-2 cm
-lots of cytokines are involved and the tumor vessels are abnormal
There are three things that tumor cells must do in order to invade other tissue, what are they?
1. Loosen contacts between cells
2. Degrade EC matrix
3. Migrate from original site

-some show tropism (migrate to certain locations)
-some lodge in nearest capillary bed
What is Hereditary Nonpolyposis Colon Cancer Syndrome?
It is a hereditary DNA repair defect
-Failure of mismatch repair (no spellchecker)
-inherit 1 mutation, acquire the other
-familial colon cancers
What is Xeroderma Pigmentosum?
It is a hereditary DNA repair defect.
-failure of nucleotide excision repair system
-small sun exposure leads to skin cancers
What are balanced translocations? How do you find them? What tumors do you most commonly find them associated with?
-common chromosomal change seen in a karyotype

2 kinds:
1. Either put proto-oncogene next to a promotor
2. Or create a fusion gene that makes a bad, growth-promoting product

-Most common in hematopoietic tumors
-Example: Ph chromosome
How does chronic myelogenous leukemia come about?
Via balance translocations in chromosomes.
-Affiliated with chromosomes 9 and 22
What do deletions of chromosomes affect?
-all or part of a chromosome
-Usually: deletion of a tumor-suppressor gene
-most commonly associated with solid tumors
-Example: del 13q14 in retinoblastoma
What are the 2 types of chemical carcinogenic agents?
1. Direct-acting agents:
-carcinogenic as-is
-most are chemotherapy drugs
-cause secondary malignancies (leukemia)

2. Indirect-acting agents:
-requires conversion to become carcinogenic
-Examples: hydrocarbons (tobacco), aflatoxin B (nuts, aspergillus-infected grains), nitrites (food preservatives)
What is the mechanism of chemical (direct and indirect-acting) agents?
The highly reactive electrophile groups bind to DNA and target RAS and p53.
What does ionizing radiation effect as a carcinogenic agent?
-causes chromosome breakage and translocations
-Examples: miners, atomic bomb, head/neck
What effects does UV light produce as a carcinogenic agent?
-Pyrimidine dimer formation
-repair pathways become overwhelmed if too much exposure
-squamous cell carcinoma, melanoma
What are grading and staging used for?
-Used for malignant tumors
-Useful for determining treatment and prognosis
What does grading neoplasms tell you?
Grading is used for:
-telling you how nasty the tumor looks
-pathologic evaluation of tumors under microscope
-mitoses, pleomorphism, necrosis, other variables
-somewhat useful but not like staging
What does staging a neoplasm tell you?
Tells you:
-how far the tumor has spread
-clinical evaluation of the patient using imaging and surgery
-TNM system (Tumor, Nodes, Metastases)
-Staging is clinical whereas grading is microscopic
-very useful
What is the TNM system and where is it used?
Tumor
Nodes
Metastases

-Used to determine stage that a tumor is in
What are the cardinal signs of inflammation?
1. Redness
2. Swelling
3. Heat
4. Pain
5. Loss of function
What is the time course of inflammation? What are the cell types involved?
1. Acute
2. Chronic
1. Acute inflammation: less than 48 hours; PMN's
2. Chronic inflammation: over 48 hours ; Mononuclear cells (macrophages, lymphocytes, plasma cells)
What are the key physiologic events involved in acute inflammation? (3)
1. Changes in vascular flow and caliber (hemodynamics)
2. Changes in vascular permeability (vascular leakage)
3. Leukocyte exudation
The 1st step involved in acute inflammation is changes in vascular flow and caliber (hemodynamic changes), what 4 characteristics do you see in order?
1. Vasoconstriction (to avoid blood loss and it's transient and inconstant)
2. Vasodilatation (helps cells get to where they need to be-1st seen in arterioles,then capillaries)
3. Slowing of circulation (outpouring of albumin-rich fluid into extravascular tissues so there is a high concentration of RBC's in the blood vessels and of high viscosity)
4. Leukocyte margination (PMN's become orientated at the periphery of vessels and start to stick)
How does vascular permeability change in acute inflammation?
This is change in vascular leakage and in inflammation from arteriole to venule, the intravascular hydrostatic pressure increases and the colloid osmotic pressure decreases so that you get edema.
-Normally both intravascular hydrostatic pressure and colloid osmotic pressure are equal
Leukocyte exudation is divided into 4 steps, what are they? (Acute inflammation-3rd step)
1. Margination, rolling, adhesion
2. Diapedesis (transmigration across the endothelium)
3. Migration toward chemotactic stimulus
4. Phagocytosis
Phagocytosis has 3 distinct steps, what are they?
1. Recognition and attachment
2. Engulment
3. Killing or degradation
(either oxygen dependent or independent)
-Them most important is the oxygen-dependent: Myeloperoxidase dependent
What are the chemical mediators of inflammation?
1. Vasoactive amines (histamine and serotonin)
2. Arachidonic acid metabolites
(via cyclooxygenase and lipoxygenase)
3. platelet activating factor (PAF)
4. Cytokines
5. Nitric oxide
6. Lysosomal constituents of leukocytes
7. Oxygen-derived free radicals
Cyclooxygenase has 2 pathways, what are they? What mediates it?
It is an arachidonic acid metabolite pathway and is a chemical mediator of inflammation.
-Pathways:
1. Prostaglandins: vasodialator
-causes pain, fever, protects stomach lining, maintains platelet function
2. Thromboxane: vasoconstrictor
What are the 4 outcomes of acute inflammation?
1. Abscess formation
2. Progress to a chronic inflammation
3. Resolution
4. Repair
Chronic inflammation arises in various organs in 1 of 3 ways, what are they?
1. Following acute inflammation
2. After repeated bouts of acute inflammation
3. Without prior acute inflammation (Tb, Rheumatoid arthritis)
What are the cells and mediators involved in chronic inflammation?
-macrophages, plasma cells, lymphocytes
-proliferation of fibroblasts and small blood vessels
-increased connective tissue
-tissue destruction
The RES system is the mononuclear phagocyte system and it consists of what?
1. Bone marrow (Monoblast/ stem cell)
2. Peripheral blood (Monocyte)
3. Tissue (Macrophage-->differentiation or activation)

-macrophage origin
Macrophages share 4 things in common with each other, what are they?
1. Mobility
2. Phagocytosis (and pinocytosis)
3. Ability to become activated (T cells, lymphokines
4. Ability to secrete large quantities of chemical mediators
Macrophages have 4 major functions, what are they?
1. Produce toxic, biologically active substances such as oxygen metabolites
2. Cause influx of other cells such as other macrophages and lymphocytes
3. Cause fibroblast proliferation and collagen deposition
4. Phagocytosis
What are the 3 ways that macrophages accumulate?
1. Continued recruitment from circulation (secondary to chemotactic factors)
2. Division
3. Prolonged survival
What is the predominant cell type at 48 hours?
Macrophage
What is a chronic granulomatous inflammation?
A type of pattern of chronic inflammation defined by presence of granulomas (small collections of modified epithelioid histiocytes/macrophages and Langhan's giant cells) usually surrounded by a rim of lymphocytes.
There are 2 factors necessary for granuloma formation, what are they?
1. Presence of indigestible organism or particlles
2. Cell-mediated immunity (T cells)
Granuloma occur in response to various diseases, name some.
-Foreign body
-Tb
-Fungal infections
-Sarcoidosis
-Schitosomiasis
-Leprosy
What are the 3 outcomes of chronic inflammation?
1. Resolution/regeneration/restitution of normal struction
2. Repair/organization/healing by C.T. scar/fibrosis
3. Continue indefinitely (Rheumatoid arthritis)
What are the factors necessary for resolution of inflammation?
1. Removal of offending agent
2. Regenerative ability if cells have been destroyed (Labile and stable cells)
3. Intact stromal framework
What are the components necessary for repair of inflammatory response?
1. Angiogenesis or neovascularization of new blood vessels
2. Migration and proliferation of fibroblasts
3. Deposition of extracellular matrix
4. Remodeling or maturation and organization of the fibrous tissue