Microbiology Module 15

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

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.

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

Erythromyocin- Macrolide (Large molecule)

Binds to 23S rRNA

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

Chloramphenicol

Chloramphenicol Acetyl Transferase is common gene on Tn's confers

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

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)

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

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

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)

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

• 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)

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)

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.

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

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.