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60 Cards in this Set
- Front
- Back
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1. When the physician collects a specimen what will the diagnostic laboratory do?
Three things... Why can testing be more difficult in dentistry? What happens to the results? |
1. Direct stain
2. Culture to purify and obtain sufficient organisms for further tests 3. Diagnostic tests -metabolic -PCR -sequencing -antibody detection tests Flora of the oral cavity contaminates the specimen being tested Results will be returned to the physician to initiate treatment |
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2. By what three methods is laboratory identification of pathogenic bacteria done by?
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1. DNA sequence
-16S rRNA -specific genetic markers (by PCR detection) 2. Antibody Reactivity (specific antigens recognized) 3. Biochemical tests -initially divided into two groups by Gram stain -further structural/biochemical classification |
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3. What makes ups the cell wall of bacteria and is special to cell walls?
What is the structure of this? (two parts) |
Peptidoglycan
1. Alternating residues of NAG and NAM 2. Peptide chain |
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4. What makes up the peptide chain?
What is special about the fourth AA? |
Three variant AA w/ a terminal D-alanine
D-Ala can be cross linked to the 3rd AA of a neighboring chain **cross linking gives rigidity and adds strength to the peptidoglycan |
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5. How is the crosslink between the 4th D-Ala and the 3rd AA?
How does it differ between gram positive and gram negative bacteria? |
Can be direct or via a bridge
Gram-Positive -insert bridge between D-Ala and 3rd AA -add extra AA in between Gram-Negative -direct peptide bond -"intra-peptide" bridge |
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6. How are gram positive bacteria structurally?
Two characteristics.... |
1. Only a single (inner or cytoplasmic) membrane
2. Thick peptidoglycan layer |
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7. How are gram negative bacteria structurally?
Three characteristics... |
1. Have an outer membrane
2. Thin peptidoglycan layer 3. Have an inner (cytoplasmic) membrane) |
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8. What are the steps of the Gram stain?
Steps 1 - 2 |
1. Staining w/ crystal violet
**both gram pos. and neg. appear purple under oil-immersion light microscopy 2. Treatment w/ iodine aggregates the crystal violet **iodine makes crystal violet less diffusable |
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9. What is the third step of the Gram stain?
Why is this step important? What happens as a result? (two things) |
Washing w/ ethanol collapses peptidoglycan and extracts lipids from the outer membrane
When peptidoglycan layer collapses nothing can get in or out 1. Gram positive retains purple stain 2. Gram negative has thin peptidoglycan layer so have breakage and stain leaks out |
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10. What role does the bacterial wall have on shape?
What are the basic bacterial shapes? (seven shapes) |
Bacterial wall (peptidoglycan) confers shape
1. Cocci (round) 2. Bacillus (rod) 3. Vibrio (U shaped) 4. Fusiform (tapered ends) 5. Spirochete) 6. Filamentous (branching) 7. Pleomorphic (no consistent shape) |
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11. What are some basic bacterial arrangements?
Five arrangements... How does morphology depend on age of culture? |
1. Diplo
2. Tetrad 3. Sarcinae 4. Strepto (chain) 5. Staphylo (grape like shape) For some species gram stain morphology can vary depending on age of culture **Rhodococcus go through a cocci-rod morphology switch depending on culture time |
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12. What happens to most bacteria w/o a cell wall?
What can some bacteria do though w/o a cell wall? |
They will lyse (burst)
**cell membrane can't w/stand pressure w/o cell wall Undergo a programed change in their cell membranes to allow them to survive w/o a cell wall **can still divide and cause disease then |
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13. What is CWD?
What are L-forms? Which bacteria have no cell walls? |
Cell Wall Defective Forms
-changes or partial loss of the cell wall L-forms have complete loss of of the cell wall Mycoplasma |
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14. What is teichoic acid (TA)?
What is TA covalently linked to? What do teichoic acids covalently attached to the cell wall do? What can TA act as a barrier to? |
Acidic polysaccharide glycerol or ribitol joined by phosphate groups and A can be linked to it
Covalently linked to muramic acid Confer a negative charge Act as a barrier to penetration of negatively charged molecules |
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15. What does the amount of teichoic acid depend upon?
(two things) Which bacteria is teichoic acid primarily associated w/? Besides teichoic acid what else can be covalently linked to the cell wall? |
1. Bacterial species
2. Amount of peptidoglycan Gram-positive bacteria Proteins by sortases |
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16. What is the first step of cell wall biosynthesis?
Where does this step occur? |
NAM is synthesized from NAG
**synthesis of peptidogylcan subunit Cytoplasmic step |
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17. What is the second step of cell wall biosynthesis that occurs in the cytoplasm?
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A pentapeptide chain is attached to NAM
**usually D-Ala **D-Ala is derived from L-Ala by a racemase |
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18. What is the third step of cell wall biosynthesis?
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Undecaprenyl carrier transfers the NAG-NAM-pentapeptide subunit to the outer leaflet of the cytoplasmic (inner) membrane
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19. What is the fourth step of cell wall biosynthesis?
What carries out the addition of new subunits? How are the new subunits added? |
Autolysins cleave hole in the cell wall to allow subunit to be put into existing cell wall
Penicillin Binding Proteins (PBP) Added by transglycosylation carried out by high molecular weight binding proteins |
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20. What occurs during the fifth step of cell wall biosynthesis?
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Transpeptidation reactions
1. High molecular weight PBPs crosslink D-Ala to the 3rd AA of an adjacent subunit w/ or w/o an intramolecular bridge 2. Low molecular weight PBPs cleave off the D-Ala (5th AA) w/o crosslinking |
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21. What does Phosphomycin/Fosfomycin inhibit?
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It is a PEP analog that inhibits the synthesis of NAM from NAG
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22. What is D-cycloserine?
What does it do? |
Analog of D-Ala
Inhibits the racemase reaction and addition of D-Ala to the AA1-AA2-AA3 chain |
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23. What does Vancomycin (glycopeptide antibiotic) bind to?
What does it do? |
Binds to D-Ala -- D-Ala chain
Sterically hinders PBPs from carrying out transpeptidation and transglycosylation reactions **very large molecule so covers D-Ala -- D-Ala |
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24. What are β-lactams structurally related do?
What do they do? What is basic structure look like? |
Related to D-Ala
Tightly bind PBPs inhibiting their transglycosylation and transpeptidation activity Square w/ N at one corner and a C=O at one corner |
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25. What do cell wall synthesis inhibitory antibiotics require?
What do growing cells produce? In the absence of new subunits what happens? |
Require growing cells for activity
Growing cells need to increase their cell wall so they produce autolysins to cleave the wall prior to inserting new peptidoglycan Autolysins will eventually destroy the cell wall ***resulting cells are osmotically fragile |
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26. What are three ways to become resistant to cell wall inhibitory antibiotics?
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1. Enter a quiescent state
2. Convert to L-forms 3. Mycoplasma -- lacks cell wall |
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27. What are bacterial membranes primarily made of?
(two things) What do bacterial membranes do not contain? What is the exception? |
1. Phosphotidylethanolamine
2. Phosphotidyglycerol Do not contain sterols Exception is Mycoplasma which incorporates sterols from their environment into their membranes |
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28. What is the membrane structure of gram positive?
Five things... |
1. Inner leaflet - phosphotidylethanolamine
2. Outer leaflet - phosphotidylglycerol 3. Lipoproteins 4. Lipoteichoic acid 5. Membrane proteins |
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29. What are both leaflets of the cytoplasmic membrane in gram negative made of?
What is the inner leaflet of the outer membrane predominantly made of? What is the outer leaflet of the outer membrane composed of |
Made of a mix of phosphotidylglycerol and phosphotidylethanolamine
Phosphotidylethanolamine LPS (lipopolysaccharide) **LPS is an endotoxin that causes endotoxic shock **endotoxic shock is only in gram negative bacteria |
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30. What is LPS composed of?
Three components... |
1. Lipid A
**causes endotoxic shock 2. Core glycolipid 3. O-specific oligosaccharide subunit compose of repeating sugar subunits |
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31. What do the outer membranes of Gram negative bacteria contain for transport?
What are these? |
Porins
Water filled channels through which molecules pass through **can be either non-specific (water-filled) or specific **usually size regulated |
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32. What are the different types of transporters located in the cytoplasmic membrane of Gram negative and positive bacteria?
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Do no require energy
1. Free diffusion 2. Facilitated diffusion Require energy 1. Active transport 2. Group translocation |
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33. How is facilitated diffusion?
What is uniport? What is symport? What is antiport? |
Down a concentration gradient using a carrier
Single molecule in or out (uniport) Two molecules moving in same direction (symport) **both molecules must be present in high concentration Two molecules moving in opposite directions **uses proton gradient often |
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34. What is required in active transport?
What happens in group translocation? What is an example? |
Either ATP or proton motive force (PMF)
Group translocation results in the phosphorylation of the transported substrate Example: use in transport of sugar and phosphorylate sugar as brough in |
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35. How does the outer membrane of Gram negative bacteria affect antibiotics?
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Acts as an additional barrier for antibiotics b/c they are not negative enough to enter porins
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36. What are the classes of drugs that can affect membranes?
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1. Those which disorganize membrane structure leading to disintegration of the cell
2. Produce a pore through the membrane allowing diffusion in and out through the pore 3. Inhibit membrane bound transporters 4. Produce changes in permeability towards specific ions 5. Inhibit synthesis of essential membrane components |
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37. Are most antibiotics specific for bacterial membranes?
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No b/c there are few differences between eukaryotic and prokaryotic membranes
**therefore they cannot be used therapeutically |
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38. How are bacterial membranes different?
Three ways.... |
1. Higher protein content
2. Phosphatidylcholine is rare 3. Lack sterols **except mycoplasma |
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39. What is polymyxin composed of?
What does it do? Can it affect dormant (non-replicating) cells? Where is it predominantly used? |
Composed of a FA tail and cyclic peptide head group
Disrupts bacterial membrane causing release of cytoplasmic components Will destroy dormant cells On skin |
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40. What is the binding of polymyxin due to?
What does it induce at high concentrations? What are polymyxins more effective against? Why? |
Due to FA tail and cyclic peptide head group
Induces autolytic mechanisms More effective against Gram negative bacteria B/c they have a higher affinity for LPS and phosphotidylethanolamine |
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41. What are cationic peptides?
What are examples? What is special about cationic peptides? Are they FDA approved? |
Class of antibiotics that self-assemble to form pores in membranes
**punch hole in cytoplasmic membrane so it leaks Examples: -antimicrobial peptides (AMPs) -nisin (food preservative) Can target proteins found only in bacterial membranes Not approved by FDA for human treatment |
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42. What is a glycocalyx?
What are the three types of layers? |
Polymer layer outside the bacterial cell composed of polysaccharides or polypeptides
1. Capsule 2. Slime layer (matrix) 3. Fibrils |
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43. What is a capsule?
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Discrete layers associated w/ individual cells that is made up of different things
(polypeptides and polysaccharides) |
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44. What are the four functions of capsules?
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1. Mediate adherence
2. Protect from phagocytosis 3. Protect from antibiotics 4. Protect from desiccation **capsulated can live on surface b/c protect from drying |
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45. What is a slime layer (matrix)?
What are fibrils? |
Secreted layer (by bacteria) which embeds the cells
**biofilm --> i.e. caries Thin tangled layer of polysaccharides |
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46. What are the two functions of a matrix?
What is the function of fibrils? |
1. Carbohydrate reservoir
2. Matrix for biofilm formation Fibrils function in signal transduction **cell to cell communication |
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47. What are fimbriae?
What are the made of? What are they involved in? How are fimbriae different from flagella? |
Short flagella-like structures
Made of protein Involved in attachment (adherence) Not made of flagellin or involved in motility |
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48. How do pili compare to fimbriae?
What are pili involved in? |
Longer and thinner than fimbriae
Involved in.... 1. Adherence 2. Protection 3. Conjugation 4. DNA transfer |
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49. What are flagella composed of?
What does flagella use to power bacterial motility? How does the flagella attach to the bacteria? |
Composed of flagellin (protein) in long chains
Uses the proton motive force to rotate flagella to propel the bacteria Hook and basal body attach the flagella to the bacteria |
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50. What are the three arrangements of flagella?
What are flagella most important for? (three things) |
1. Monotrichous (polar)
2. Lopotrichous 3. Peritrichous 1. Chemotaxis 2. Eukaryotic cell invasion 3. Biofilm formation |
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51. What are gliding and twitching?
What is it due to |
They are alternate forms of motility that do no depend on flagella
Type IV pillus |
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52. How is motility detected?
How can the presence of flagella be detected? (two ways) |
Detected by direct observation (microscope) and/or motility medium
1. Dyes -dyes add extra width to structure and absorb light making them visible 2. Antibody stains -antibodies recognize flagellin so possible to detect the flagella |
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53. What is motility medium?
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Semisolid medium that will hold non-mobile bacteria in place but motile microbes can swim through it
Presence of turbidity throughout the tube is a positive test for the presence of motility |
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54. How does DNA fit in the bacterial cell?
What does DNA gyrases do? Why is DNA gyrase necessary? |
DNA is supercoiled and compacted
DNA gyrases bind to DNA and catalyze strand cleavage and exchange to affect DNA supercoiling Needed to get DNA accessible for transcription and replication |
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55. What do quinolones and floroquinolines bind to?
What do they therefore inhibit? What does novobiocin and ciprofloxacin inhibit? |
Bind to DNA gyrase:ATP comples
Block transcription and DNA replication Inhibit DNA gyrase activity |
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56. How is PCR used as a diagnostic indicators?
What are some advantages of PCR? |
Many pathogens have genes unique to only the single species so the ability of PCR to amplify that gene can by used to indicate that the specific pathogen is present
It's rapid and can detect pathogen w/o purifying it |
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57. What are the three steps of one PCR cycle?
What does PCR require? |
1. Denaturation
2. Annealing 3. Extension Requires generation of a template and the design of specific primers |
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58. What are the primers in PCR used with and why?
How is the amplified gene detected? (two ways) |
Primers are used w/ Taq polymerase to amplify the gene from the template
1. Agarose gel electrophoresis and ethidium bromide staining or 2. Incorporation by increase in fluorescence during the PCR reaction (automated) |
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59. How can a species be identified using PCR?
Is a purified sample necessary for PCR? |
Species can be identified by the presence of an amplified gene present only in that species
PCR can be don on purified bacteria or a mixture of bacteria from a clinical specimen |
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60. What is 16S rRNA sequencing used for?
What is 16S rRNA? |
Used to identify a specific species
**by sequencing regions of the 16S rRNA it's possible to determine the species of bacteria present in a sample **has dental purpose b/c can see if bacteria is there even if you can't culture it A region of the genome that has highly conserved to highly variable regions |
