Biol 120 Final (new material)

  1. Mutant Isolation
    (Selectable vs Nonselectable)
    • Selectable mutation- progeny outgrows parent strain
    • Nonselectable mutation- ex. auxotroph: lost ability to synthesize substance required for growth and metabolism (must add that substance to culture)
  2. Replica Plating
    • what you screen for is what you get
    • -you can cause many mutations at once, but you screen for the one that you want
    • -Master plate- best media for microorganisms
    • -Velveteen- grow then stamp plate over. This plate has selectable media. By keeping orientation the same, can figure out which colonies are mutants
  3. Mutation Hot Spots
    certain genome sequences experience a lot of mutations
  4. Rate of mutation in RNA genome vs DNA genome
    Rate is 1000x higher in RNA
  5. Tautomers
    variation in chemical structure of bases
  6. Point Mutation
    • Usually 1-2 bp substitution
    • -can be synonymous mutation due to redundancy in codons
  7. Mis-sense mutation
    change one codon > faulty protein
  8. Nonsense Mutation
    change one codon > incomplete protein (due to stop codon)
  9. Silent Mutation
    • change one codon > normal protein
    • (usually 3rd nucleotide of codon causes this)
  10. Transition vs Transversion
    Transition: replace purine w/ purine or pyramidine w/ pyramidine

    Transversion: replace purine w/ pyramidine or pyramidine w/ purine
  11. Insertion or Deletions
    • make sequence longer or shorter
    • if not in multiples of 3, will change the reading frame of the codons
  12. Inversion
    • occurs when fragment of DNA flips in orientation
  13. Frameshift Mutations
    Change reading frame (insertion: +1, deletion: -1)
  14. Suppressor Mutation
    restoration of normal gene function by adding/deleting bp near original mutation
  15. Revertant Strain
    Strain in which wildtype phenotype lost in mutant is restored
  16. Same Site Revertant
    2 mutations occur: 1 cancels the other
  17. Second Site Revertant
    • 1. mutation somewhere else in the gene
    • 2. mutation in other gene can restore enzyme function (ex. a produced coenzyme mutates and now fits mutated enzyme)
    • 3. mutation in other gene can result in another metabolic pathway
  18. Jimmy was trying to isolate a histidine auxotroph. He replica plated his culture on a complete medium and on a minimal medium that lacked only the histidine growth factor. How would he identify his auxotroph?

    A. it would grow on the complete medium, but not on the minimal
  19. The original DNA coding strand lookls like this: 5'-ATG-3' (codes for f-Met). The mutated DNA looks like this: 5'-AAG-3' (Lys). What kind of mutation is this?

    C. transversion
  20. Induced mutations: Mutagenic Treatment (3 methods)
    • 1. Chemical
    • 2. Physical
    • 3. Biological
  21. Chemical Mutagens
    Base Analogs- chemicals that look like the molecules they substitue

    Alkylating Agents- cause change to DNA by alkylating it (ex. Ethidium bromide)
  22. What class of mutation is most common?

    A. neutral
  23. Radiation as a Mutagen
    a. Non-ionizing: UV light is absorbed by nucleotide bases to form pyramidine dimers (2 adjacent pyramidine bases become covalently bonded)

    b. Ionizing: short wavelength such as Xray or gamma rays
  24. Biological Mutagens
    • a. Transposon mutagenesis
    • b. Mutations that arise from mutation repairs
  25. Transposable Elements
    "jumping genes"
    • 1. Transposase- recognizes, cuts, and ligates DNA
    • 2. Short inverted terminal repeats at end of DNA (10-1000 bp long)
  26. 2 Types of Transposable Elements
    • 1. Duplicated when inserted at target sequence
    • 2. Not duplicated at target sequence (can be used to cause mutations if you know the target sequence)
  27. Ames Test
    Way to test for carcinogens: Use an auxitroph that requires histidine. Do Kirby Baur test using possible carcinogen on disc. If revertants grow, the substance causes mutations.
  28. Biological Mutagens
    • 1. Mechanism to exise single nucleotide may make a mistake
    • 2. Allelic Exchange- based on genetic recombination. Usually a large chain of DNA. Replace HisG w/ tagged DNA (antibiotic). *Don't have to be equal sized pieces of DNA. Since bacteria is haploid, only one copy of gene needs to be replaced.
  29. Bacteriophage Mutations
    • sometimes DNA can integrate into host DNA.
    • 1. attB sequence in bacteria genome
    • 2. attP sequence in virus genome
    • By using integrase, lysogenic phages can incorporate an entire plasmid into bacteria genome
  30. In early Earth, reducing compounds came from:
    volcanoes, solar radiation, heat
  31. Liquid Soup Theory
    Mimicked atmosphere and found that certain biological molecules could be produced (such as CoA)
  32. Thermal Vent Theory
    formation of first cell was on a substrate porous enough to collect needed molecules (mimicked semipermeable membrane)
  33. Insertion sequences and transposons both must have ______ for transposition to take place

    D. transposase
  34. Where do we find the evidence for early microbial life?

    A. stromatolites
  35. Evidence for Microbial Life
    • 1. microfossils- decayed cells filled with calcium carbonate or silica
    • 2. stromatolites- 3.4 billion years old
    • 3. biosignatures- organic molecules found in sedementary rock. could only be formed by microbes
    • 4. Isotope ratios- ratio of certain isotopes is altered by microbes. ex. microbes fix 12CO2 more readily than 13CO2
  36. Early earth was a reducing environment. Chemicals present:
    CH4, NH3, H2, CO2
  37. Amphiphilic molecules as first barrier for biological molecules
    First have a bilayer vessicle then a protocell
  38. Metabolist Model of Morowitz & Wachterhauser
    • CO2 based metabolism originated through self sustaining reactions
    • -catalyzed by metal sulfides
  39. RNA World Theory
    RNA is the first molecule to serve as genetic material. Also served as a catalyst (ex. splicing of introns & exons is catalyzed by RNA)
  40. The first cell was likely:

    D. all of the above
  41. Cyanobacteria and early earth
    Oxygen was not present in the atmosphere until cyanobacteria. Took 1.6 billion years to go from 0% to 20% atmospheric oxygen.
  42. Endosymbiosis
    • Mitochondria is like coxiella (very small, similar genome).
    • Chloroplasts likely came from cyanobacteria
  43. Species definition for microbes
    • share many properties and differ significantly from other groups/strains
    • 1. 16S ribosome sequence must differ by more than 3% from other organisms
    • 2. 70% or greater genomic sequence variability
  44. Strain
    • a population of microbes descended from a single individual or pure culture
    • May be 99.9% similar to test strain
  45. Biovars
    • Biovars- strains that are biochemically or physiologically different
    • Morphovars- Strains that vary in morphology
    • Serovars- Strains that vary in antigenic properties
  46. Classical Taxonomy
    based on colony morphology, cell shape, etc.
  47. Molecular Classification
    • 1. G&C content. Higher concentration=higher melting temperature. Extract DNA and do melting curves.
    • 2. Nucleic acid hybridization: greater percent hybridization = closer related species
    • 3. Nucleic acid sequences of 16S ribosome
  48. Bergey's Manual of Systematic Bacteriology
    1920s. There are now 5 volumes.
  49. International Journal of Systematic and Evolutionary Microbiology
    Help name novel species
  50. Phylogeny
    Monophyletic Groups
    • Clades- groups of related organisms
    • Monophyletic Groups- Group of species that share a common ancestor that is not shared by others
  51. Internal Nodes on a phylogenetic tree represent...
  52. The first primitive organism

    B. used RNA both as a catalyst and a unit of heredity
  53. Which of the following gas was not present on early earth?

    C. oxygen
  54. Beneficial Interactions between humans and microbes
    • 1. Normal flora
    • 2. Food via digestion (vitamin K)
  55. Parasytic Interactions
    Harm us by overstimulating immune system or killing our cells
  56. Pathogenicity
    • Pathogenicity- ability of microbe to cause disease or inflict damage
    • Infectivity- Qualitative; how easily can cause disease
    • Virulence- Quantitative; # microbes that elicit disease
    • Infection- Growth of organism in host
    • Disease- damage to host
  57. Primary Pathogen vs Opportunistic Pathogen
    Primary Pathogen- breach defense of healthy host

    Opportunistic Pathogen- cause disease in a compromised host
  58. The LD50 of Salmonella typhimunum is given as 5*105 cells. This means that 5*105 cells

    B. will kill 50% of mice injected with this dose
  59. Examples of virulence
    • S. pnemoniae is highly virulent (100 organisms needed)
    • S. typhimurium is moderately virulent (10,000s of bacteria)
  60. Attenuation
    A microbe that is not able to cause disease.
  61. Normal flora determined at birth
    • breast fed= lactobacillus
    • bottle fed= enterobacteria to digest proteins
  62. Gastrointestinal Tract
    • 1011 bacteria / gram of feces
    • Synthesize vitamins (K, B12)
    • Steroid metabolism
    • Acetic acid production
    • Glycosidase reactions
    • Methane production
  63. Rodents not exposed to bacteria
    required 30% more food than those with bacteria
  64. Normal Flora & Urogenital Tract
    • Bladder & kidneys are sterile
    • Urethra can be colonized by facultatively aerobic gram negative rods & cocci
    • Vagina can be colonized by different bacteria based on hormonal factors
  65. Microbial causes of chronic disease
    • Chlamydophila pnemoniae: artherosclerosis (walking pnemonia)
    • Diabetes (type 2) linked to virus
    • Crohn's disease- mycobacteria
    • **Microbes trigger chronic disease by triggering immune response
  66. Host factors in infection
    • 1. age
    • 2. stress
    • 3. diet
    • 4. genetics
  67. Nosocomial Infections
    associated with hospitals. Many sick people and bacteria with antibiotic resistance
  68. Steps of Harful Interactions w/ Microbes
    • 1. Exposed
    • 2. Adherence
    • 3. Invasion
    • 4. Colonization & growth
    • 5. Result
  69. Exposure
    microbe must be able to penetrate barriers (skin, mucous membrane, etc.)
  70. Adherence
    • Receptor mediated (specific)
    • ex. N. gonorrhea in urogenital epitherlia has CD66 which binds to opa
    • -capsule, fimbrae, or pilli may be involved (bind to host cell glycoproteins)
  71. Invasion
    • penetrate epithelium to initiate pahtogenesis
    • *if gets in blood or lymph, can get in any other part of body
  72. Proteins for invasion by microbe
    • ex. Hyaluronidase (breaks down hyaluron in connective tissue)
    • ex. Colleginase (breaks down colligen)
    • *allows deeper tissue invasion
  73. S. aureus: function of coagulase and kinase
    forms clot around microbe so that antibodies can't detect it
  74. Colonization & Growth
    must have appropriate environmental conditions & nutrients
  75. Trace Elements In Human Body
    • Iron is difficult for microbes to obtain.
    • Transferrins & lactoferrins keep iron in host body.
    • Siderophores are used by microbes to take in iron
  76. Bacteriemia vs Septicemia
    • Bacteremia- presence of transient bacteria in blood
    • Septicemia- blood poisoning. Presense of pathogens in blood
  77. Virulence Factors
    genes involved in pathogenesis, aid in establishment and maintenence of disease
  78. Invasiveness
    • ability of organism to grow in host tissue in large number that inhibits host function
    • ex. S. pneumoniae: hemolysis inhibits host lung function
  79. Exotoxins
    proteins that cause damage (at site or travel to other parts of body)
  80. AB Toxins (exotoxins)
    • A=active part (inserted into cell to cause damage)
    • B=binding part
  81. Cholera exotoxins
    • AB toxin disrupts electrolyte balance to cause diarrhea and frothing at mouth
    • -toxin affects adenyl cyclase so that ATP -> cAMP causing Cl- to move to lumen and Na+ movement is blocked. Water moves into intestinal lumen and does not get taken back in.
  82. Diptheria exotoxins
    • Disrupts protein synthesis by ADP riboslyation of elongation factor 2
    • -toxin binds to epithelial layer of intestinal tract. Without protein synthesis, cells begin dying.
  83. Virulence refers to:

    B. the degree or intensity of pathogenicity
  84. Organisms that are ubiquitously found in nature, but that cause disease only in individuals whose defenses are somehow debilitated are called ____ pathogens

    C. opportunistic
  85. Where would you expect to find the lowest level of microbes?

    D. the kidney
  86. Which of the following does NOT describe one of Koch's postulates:

    E. characterize the gram staining characteristic of the isolated agent
  87. Which of the following is not an adhesion factor for pathogens

    A. flagella
  88. Endotoxins
    • bind to macrophages and B cells
    • -LPS releases TNF-alpha, interferon, & cytokines
    • -symptoms: fever, activation of clotting factors, clots in arteries, low blood pressure, shock, death
  89. Bacteria in Blood: endotoxins & antibiotics
    If antibiotics kill bacteria in blood, could make things worse. Bacteria can release more endotoxins.
  90. Endotoxins vs exotoxins
    Need a lot of endotoxins for disease (micrograms of it) but only need a little bit of exotoxins (picograms)
  91. Endotoxins vs Exotoxins
    • Exotoxins: proteins can be destroyed with heat
    • -have specific action on cells or tissue
    • - highly toxic, often fatal
    • - highly immunogenic (stimulate antitoxin production)
    • - no fever

    • Endotoxins: LPS complex (gram negative) heat stable
    • - general infection; causes fever, diarrhea, vomitting
    • - weakly toxic, rarely fatal
    • - poorly immunogenic
    • - pyrogenic (causes fever)
  92. Immunology of Host
    • 1. Nonspecific Passive: physical / anatomical defenses (ex. salty skin)
    • 2. Nonspecific Adaptive: immune system
  93. Lymphatic System
    • complex organs and circulatory system
    • Lymph: WBCs, mainly lymphocytes
    • Lymph Nodes: Swollen when sick. Meeting grounds for immune system
    • Spleen: organ where immune system cells confront invader
  94. Lymphocytes
    T Cells, B Cells, Natural Killer Cells
  95. Leukocytes
    Lymphocytes, monocytes, dendritic cells, macrophages, PMNs (neutrophils, basophils, eosinophils)
  96. Phagocytes:
    • Polymorphonuclear leukocytes (PMNs)
    • Macrophages
    • Monocytes
  97. Specific Immunity
    • Recognize and destroy an individual pathogen or their products
    • Phagocyte presents part of the digested pathogen to lymphocytes which triggers immune response
  98. Phagocytes and lysosomes
    • Phagocyte engulfs bacteria via endocytosis to form "phagosome" which fuses with lysosome to become "phagolysosome" where the bacteria is killed.
    • *Very acidic environment
  99. 3 ways bacteria can escape death if engulfed
    • 1. Some bacteria can break out of phagosome w/ hemolysis enzyme and live in cytoplasm (ex. Listeria monocytogenes)
    • 2. Some bacteria can secrete proteins in cytoplasm that interfere with phagolysosome production (ex. Legionella pneumophila in air conditioning vents)
    • 3. Some can thrive in the phagolysosome (acidic environment) (ex. Coxiella burnetti)
  100. Pattern Recognition Molecules (PRMs)
    PRMs are on macrophages. They recognize bacteria (not specific bacteria, but specific TYPES of bacteria)
  101. Cytokines
    released from leukocytes. Cause redness, swelling, pain, and heat
  102. Antigen
    • molecule capable of interfering with specific components of the immune system. Host will:
    • a. destroy antige directly
    • b. produce antibodies to target antigen
  103. Adaptive Immune Response
    • requires more time and exposure to agent
    • a. cell mediated (T Cells)
    • b. humoral (B Cells -> antibodies)
  104. Antibodies
    • circulate in blood. Can also be inside cells if that's where pathogen is.
    • *Not effective against pathogens that live within cells???
  105. T Cells & B Cells live in lymphoid organs and await antigetns that activate them
    • macrophage presents antigen which activates T Cells
    • T Cell activates B Cells which release antibodies
  106. Self Reactive Immune Cells
    self reactive cells are destroyed during development of the immune response. Immune cells are not able to react with self antigens
  107. Long Living Memory Cells
    • B Cells and T Cells can become long living memory cells. Will remember specific antigen and respond quickly if exposed again.
    • *basis for vaccine production
  108. Humoral Immunity
    Antibody mediated immunity. Effective against viruses & bacteria in blood/lymph & toxins
  109. Antibody binding sites
    Most have 2 binding sites, both for same antigen. If binds to both, would get clumping effect
  110. Immunoglobulin (IgG)
    • major circulating antibody in blood
    • when exposed to antigen:
    • IgM is first to appear (can bind 10 antigen, 5 to each arm)
    • IgA is present in secretions (breast milk)
    • IgD secred in blood/lymph & immature B Cell surfaces
    • IgE bind to mast cells, release histamine (allergy symptoms)
  111. Types of T Cells
    • Cytotoxic T Cell- combat viruses and bacteria that infect cells (apoptosis=cell death), recognize antigen and release granules to kill pathogen
    • Helper T Cell- trigger macrophage to seek invader. Secrete cytokines.
    • Regulatory T Cell- oversee this system
  112. Superantigens
    • causes massive immune response
    • ex. S. aureus can cause toxic shock syndrome
    • Don't bind to normal receptor area which means not very specific. Can bind to any T Cell.
  113. Vaccines
    • 1st encounter: mostly IgM (slow due to delay)
    • 2nd encounter: mostly IgG and a LOT of it (very fast)
  114. Active Immunity
    1. Natural
    2. Artificial
    • 1. Natural: ex. chicken pox
    • 2. Artificial: vaccination
  115. Passive Immunity
    1. Natural
    2. Artificial
    • 1. Natural: maternal antibodies
    • 2. Artificial: Injection of antiserum from someone who has previously formed antibodies
Card Set
Biol 120 Final (new material)