Microbiology Module 15

  1. B-Lactam Antibiotics (Slide 2)
    The "Cillins" all contain B-lactam structure that inhibit transpeptidase (analogues of D-Ala)

    Different side chains confer different properties, ranges

    Resistence often develops when B-lactamase (penicillinase) gene is acquired by pathogen

    Cephalosporins, also have B-lactam structures
  2. Vancomycin (Slide 3)
    MRSA: Methicilin resistant staphorius. Can use vancomycin

    Large MW antibiotic effective against G+ organisms (Not orally bioavailable)

    Binds to D-Ala--D-Ala residues and blocks transpeptidase as well as other steps in cell wall synthesis

    Resistance due to changing final D-Ala to other AA or similar structure

    VRE (Vancomycin Resistant Enterococcus) is known to transfer resistance horizontally.
  3. Protein Synthesis Inhibitors (Slide 4)
    Aminoglycosides (strepto- and gyntomycin) contain cyclohexane and aminosugars

    Binds to 30S ribosomal subunit

    Resistance widespread due to enzymatic modification of the antibiotic and mutations in ribosome targets
  4. More Protein Synthesis Inhibitors (Slide 5)
    Erythromyocin- Macrolide (Large molecule)

    Binds to 23S rRNA

    Resistance due to change in 23S rRNA sequence or macrolide-digesting enzyme


    Chloramphenicol Acetyl Transferase is common gene on Tn's confers
  5. Metabolic Antagonists (Slide 6)
    Antibiotics that target important biosynthetic pathway in the cell. THF acid pathway required in order to make all bldg blocs for DNA and RNA synthesis

    Bacteria use this pathway to make nucleotides from scrath

    Sulfonamides (Sulfamethoxazole) are PABA analogues that prevent folic acid production

    Resistance is due to mutations in synthetase or by acquiring THF transport genes
  6. Metabolic Antagonists Cont. (Slide 7)
    Trimethroprim (antibiotic) inhibits DHFR and resistance due to mutations in DHFR/THF transport acquisition

    Frequently 2 antibiotics, each inhibiting a diff. step in a common pathway are used together (i.e.: Bactrim or Septra-both containing sulfamethoxazole and trimethroprim)
  7. Topoisomerase Inhibitors (Slide 8)
    Antibiotics chemically classified as quinolones derived from nalidixic acid. Kill cells and prevent cell division by inhibiting Type II and IV topoisomerases

    Resistance due to mutations in topoisomerases, yet not widespread
  8. Antibiotic Resistance Mechanisms (Slide 9)
    Alter permeability or entry pathway for antibiotic looking to get into the cell

    Chemically modify or degrade antibiotic. Are usually found in nature and need little human help

    Mutation in target. Can kill off microbes in infected patients and those that survive are those with mutations

    Efflux pump confers multidrug resistance

    Many resistance-conferring genes are found on transposons and R-plasmids, so horizontal transfer is common
  9. Antifungals (Slide 10)
    Many antifungals are toxic to host because fungi are eukaryotic and fungal targets often structurally resemble host components

    Chitin production in cell wall is obvious potential target, currently being studied

    Ergosteroal synthesis/function is common target of many anti-fungals (Azoles, amphotericin, nystatin)

    Microtubule is also a target (Griseofulvin)
  10. Antiviral Drugs (Slide 11)
    Most antiviral drugs target replication, many are nucleoside analogues

    Some antivirals have specific targets (HIV protease inhibitor ritonavir, neurominidate inhibitor tamiful, influenza virus uncoating inhibitor amantidine)

    Resistance to these drugs is common, because viral replication is often error-prone
  11. Making Monoclonal Antibodies (Slide 13)
    • Mouse:
    • -Inject mouse with antigen
    • -Harvest spleen containing B-Cells

    • Cells:
    • -Fuse spleen cells to myeloma cells (cancerous cell derived intially from B-cell)
    • -Isolate each fused cell, grow and screen

    • Protein:
    • - Grow up high production hybridomas
    • -Isolate antibodies (IgMs and IgGs)
  12. Immunofluorescence Microscopy (Slide 14)
    Uses antibodies and monoclonal-Antibodies covalently attached to fluorescent tags

    Direct: Labeled antibody binds directly to antigen on specimen

    Indirect: unlabeled antibody binds to specimen and then a labeled AB binds to first AB, not directly to specimen (Can be used to look for Abs in serum --> exposure to antigens)
  13. ELISA (Enzyme-linked immunosorbent assay) (Slide 15)
    Direct ELISA (sandwich) uses microtiter plates pre-coated w/bound, unlabelled ABs to trap Ag. A 2nd Tagged (enzyme-linked) AB to the Ag is added and the amount of 2nd AB is determined

    Often the 2 ABs are different monoclonals, w/diff. binding specificities.
  14. ELISA (Enzyme-linked immunosorbent away) (Slide16)
    Indirect: Used to look for serum ABs to a particular substance (Exposure): Microtiter plates are pre-coated w/Ag, serum containing primary ABs is added, then tagged (secondary) AB binds to 1st AB, if it is present

    Amount of secondary AB is quantified
  15. Agglutination Tests (Slide 17)
    Used as field test (Crime scene) to check blood sample.

    Quick procedure depending on abilities of ABs to from large immune complexes w/ Ags. Complexes so large that they're able to precipitate from solution

    Needs several dilutions of ABs and/or antigens, since complexes only form at specific ration of Ab:Ag

    Typically IgMs are used, since secreted IgM is pentavalent: 5IgMs are linked together.
Card Set
Microbiology Module 15
Module 15