12 Antimicrobial Therapy
1. Chemotherapy term coined by Paul Ehrlich (father of chemotherapy) - He discovered a drug treatment for syphilis; he also developed the guiding principle of chemotherapy, which is selective toxicity (the drug should be toxic to the infecting microbe, but relatively harmless to the hosts cells). Now the term chemotherapeutic agent describes any chemical substance used in medical practice.
2. Antimicrobial agent drug used to treat disease caused by microbes.
3. Antibiotic chemical substance produced by a microorganism that has the capacity to inhibit the growth of bacteria and even destroy them. One of the first antibiotics was penicillin, discovered by Alexander Fleming in while he was carrying out experiments on Staphylococcus (1929). Some of his plates became contaminated with mold spores. As he examined them, Fleming noticed that the Staphylococcus colonies were dissolving as they neared the area where the mold was growing. He reasoned that the mold was secreting something that killed the bacteria. The mold was found to be a member of the genus Penicillium, so he named the bacteria destroying substance penicillin. Ten years later Florey and Chain had purified enough penicillin to begin experiments involving the treatment humans. It was enormously useful in the latter part of World War II.
(Antifungal and antiviral agents will be discussed later in the semester as treatment for specific diseases)
A. Selective Toxicity drug harms the microbe without causing significant damage to the host. When searching for ways to treat disease, scientists look for differences between the human (or animal) host and the pathogen. Ex. Penicillin interferes with cell wall synthesis. Animal cells have no cell walls, so penicillin is not toxic to animals.
B. Spectrum of Activity the range of different microbes against which an antimicrobial agent acts. Example: Broad spectrum: G(+) and G(-) bacteria vs. Narrow spectrum: G(-) only;
C. Modes of Action (How Do the Drugs Work?)
1. Inhibition of Cell Wall Synthesis - attach to enzymes that cross-link peptidoglycans.
Penicillins Bactericidal. Natural penicillins are extracted from cultures of the mold Penicillium notatum. Structure: contain beta lactam rings. The discovery of resistant bacterial strains led to the development of semisynthetic penicillins (resistant bacteria have beta lactamase enzymes that can break down beta lactam rings). The first of the semisynthetic penicillins was methicillin (not broken down by beta lactamase enzymes). Other familiar semisynthetics are ampicillin and amoxicillin. Allergy is rare in children but occurs in 1-5% of adults. Also used prophylactically (to prevent infection). Penicillins are not as effective against G(-) due to the presence of the outer membrane.
Cephalosporins Derived from several species of the fungus Cephalosporium. Have limited antimicrobial action. Their discovery led to the development of a large number of bactericidal, semisynthetic derivatives of cephalosporin. Structure: contain beta lactam rings. Frequently used cephalosporins include cephalexin (Keflex) and ceftriaxone (these 2 are 3rd generation cephalosporins). Have a fairly wide spectrum of activity, rarely cause serious side effects, and can be used prophylactically. Many people that are allergic to penicillin may also be sensitive to the cephalosporins.
Others Carbapenems new group of extremely broad spectrum antibiotics that have a 2-part structure (Ex. Primaxin consists of a beta lactam antibiotic and cilastatin sodium, a compound that prevents degradation of the drug in the kidneys); Bacitracin derived from the bacterium Bacillus (highly toxic, so only used topically - used on lesions and wounds of skin and mucous membranes); Vancomycin produced by Streptomyces (used to treat infections caused by methicillin-resistant staphylococci and enterococci).
2. Disruption of Cell Membrane Function
Antibiotics such as polymyxins act as detergents and distort cell membranes. Polymyxins are obtained from Bacillus polymyxa and are especially effective against G(-) bacteria such as Pseudomonas that have phospholipids in their outer membrane (along with the lipopolysaccharides).
3. Inhibition of Protein Synthesis
Protein synthesis requires the DNA code, RNA (mRNA, tRNA, and rRNA). The difference between bacterial and animal ribosomes allows antimicrobial agents to attack bacterial cells without damaging animal cells. Ex. streptomycin, erythromycin, chloramphenicol (These antibiotics can affect mitochondria. They have their own ribosomes that are similar to bacterial ribosomes.)
Aminoglycosides obtained from various species of Streptomyces and Micromnospora. The first was streptomycin (1940s); many bacteria are now resistant to it. This antibiotic can damage kidneys and the inner ear nerves, so is used only in special situations and usually in combination with other drugs. These drugs have a great ability to act synergistically with other drugs. Gentamycin is a mainstay for the treatment of Pseudomonas (if resistant, then use polymyxins).
Tetracyclines several are obtained from Streptomyces. Semisynthetic tetracyclines have been developed. All are bacteriostatic. They have the widest spectrum of activity of any antibiotic. Unfortunately, because of this they destroy both the pathogenic bacteria and the normal flora. Can cause a variety of mild to severe toxic effects (kidney and liver damage, light sensitivity, interferes with effectiveness of birth control pills, staining of teeth, abnormal bone development in fetus). Used to treat Lyme disease.
Chloramphenicol originally obtained from Streptomyces, but is now fully synthesized in the lab. It is bacteriostatic and has a broad spectrum of activity. Because of its toxic effects on bone marrow, it is the drug of last choice in the U.S. Be careful - it is sometimes sold without prescription outside the U.S.
Macrolides Erythromycin, a commonly used macrolide, is produced by Streptomyces. It is bacteriostatic and is valuable in treating infections caused by penicillin-resistant organisms or in patients allergic to penicillin. Considered one of the least toxic of commonly used antibiotics.
4. Inhibition of Nucleic Acid Synthesis
Target enzymes involved in nucleic acid synthesis (ex. DNA replication, transcription).
Rifamycin specifically targets the enzyme involved in the transcription process (mRNA synthesis); produced by Streptomyces and only used in the U.S. for treating tuberculosis; has high drug interaction. Ex. Rifampicin.
Quinolones new group of broad spectrum antibiotics; targets enzyme that unwinds DNA prior to replication; especially effective against travelers diarrhea and UTIs. Ex. Nalidixic acid used against G(-)s.
5. Interference of Metabolism
Antimicrobial compounds can function in 2 ways: 1.) by competitively inhibiting enzymes and 2.) by being erroneously incorporated into important molecules such as nucleic acids. The actions of these compounds are sometimes called molecular mimicry because they mimic the normal molecule, preventing a reaction from occurring or causing it to go awry.
a. Competitive Inhibition - Remember our discussion on enzymes, their active sites, and their substrate? In competitive inhibition an antimicrobial compound binds to an enzymes active site, so that the enzyme cannot react with its true substrate. To bind to the active site, the antimicrobial compound must be similar in structure to the true substrate.
Ex. The drug sulfanilamide is very similar to the compound para-aminobenzoic acid (PABA). Sulfanilamide competitively inhibits an enzyme that acts on PABA. Many bacteria require PABA in order to make folic acid, which they use in synthesizing nucleic acids and other compounds. When sulfanilamide is bound to the enzyme, a bacterium cannot make folic acid. Animals obtain folic acid from their diets (they dont have the enzymes to make it themselves), so their metabolism is not disturbed by these competitive inhibitors.
b. Nucleic Acid Incorporation Antimicrobial compounds such as vidarabine and idoxuridine are erroneously incorporated into nucleic acids. These molecules are very similar in structure to the nitrogenous bases. When incorporated into a nucleic acid, they garble the information that it encodes because they cannot form the correct base pairs during replication and transcription. These compounds can harm the host cells as well as the microorganisms (animal cells use the same nitrogenous bases to make nucleotides). These agents are most useful in treating viral infections, because viruses incorporate these fakes more rapidly that do cells and are more severely damaged.
A. Toxicity Some antimicrobials do exert toxic effects on the patients receiving them. These effects are discussed later in connection with specific drugs.
B. Allergy An allergy is a condition in which the bodys immune system responds to a foreign substance, usually a protein. For Ex., breakdown products of penicillins combine with proteins in body fluids to form a molecule that the body treats as a foreign substance.
C. Disruption of Normal Microflora Antimicrobials, especially broad-spectrum antibiotics, mat exert their adverse effects not only on pathogens but also on the normal or indigenous microflora (the microorganisms that normally inhabit the skin and the digestive, respiratory, and urogenital tracts and keep numbers of unwanted nonnative microorganisms in check). When these native populations are reduced, other organisms not susceptible to the antimicrobial agent invade and multiply rapidly (called superinfections) Ex. Oral ampicillin and long-term use of penicillin can often leads to destruction of normal microflora in the gut and in turn, growth of the yeast Candida. Cephalosporins, tetracyclines, and chloramphenicol often lead to oral and vaginal yeast infections. Live-culture yogurts or acidophilus in the form of a pill (both contain lactobacilli) can be given to counteract this effect (basically you are reestablishing the normal microflora).
1. Spontaneous Mutations - Most bacteria acquire antibiotic resistance by spontaneous mutations in their genetic material. Bacteria reproduce so rapidly that billions of cells can be produced in a short time; among them there will always be a few mutants. If a mutant happens to be resistant to a drug, that mutant and its progeny will survive, whereas the nonresistant cells will die. After a few generations, most of the survivors will be resistant to the drug. Understand that antibiotics do not induce mutations, but they do create environments that that favor the survival of mutant resistant organisms (see section E below).
2. R Plasmids Resistant genes usually found on R plasmids can transferred from one bacterium to another by conjugation through pili, transduction (using a viral vector), or transformation (Remember Chapter 8?)
3. L forms - Some species of bacteria can lose their cell wall and swell into irregularly shaped cells called L forms. L forms can arise spontaneously and can persist and divide repeatedly. They can spontaneously revert to normal-walled cells. In the L form, bacteria are resistant to antibiotics that effect cell wall formation (ex. penicillin).
2. Two antibiotics may be administered simultaneously so they can exert an additive effect (synergism). Ex. penicillin is often added to another antibiotic (the penicillin damages the cell wall, allowing better penetration by the other antibiotic). In the following example, another antibiotic is added to penicillin to increase its effectiveness. Augmentin = penicillin (amoxicillin) and clavulanic acid. Clavulanic acid binds to beta lactamases and prevents them from inactivating the amoxicillin.
3. Antibiotics can be restricted to essential uses only (ex. not for colds, etc.). In addition, the use of antibiotics in animal feeds could be banned.
1. There are many different kinds of infectious agents in confined area.
A. The Disk Diffusion Method (Kirby-Bauer method) A bacteria is uniformly spread over an agar plate. Filter paper disks are saturated with the drug and placed on the agar surface. Clear areas called zones of inhibition appear on the agar as round disks where the drugs inhibit the bacteria. Important to realize the results obtained in vitro (in the lab) often differ from those obtained in vivo (in a living organism). Metabolic processes in a living organism may inactivate or inhibit a drug. Well do this procedure in lab.
B. The Dilution Method A constant quantity of microbial inoculum is introduced into a series of broth cultures containing decreasing concentrations of a drug. After incubation, the tubes are examined and the lowest concentration of the drug that prevents visible growth (indicated by turbidity) is noted. Advantage to using this method over the disk method: Samples from tubes that show no growth can be used to inoculate broth that contains no drug to see if the drug was bactericidal or bacteriostatic.
C. Serum Killing Power Obtain patients blood sample while the patient is receiving an antibiotic. Bacteria are added to the patients serum (blood plasma minus clotting proteins). Growth (turbidity) after incubation indicates that the antibiotic is ineffective.
D. Automated Methods Bacteria are added to wells in trays to which a variety of antimicrobial agents have been added. The trays are inserted into a machine that measures microbial growth. Advantages: efficient, fast, inexpensive, and allows physicians to prescribe an appropriate antibiotic early in an infection rather than prescribing a broad-spectrum antibiotic while awaiting lab results.
A. Solubility in body fluids
B. Selective toxicity
C. Toxicity not easily altered (no food or drug interactions)
E. Stability (should be degraded and excreted by the body slowly)
F. Resistance by microorganisms not easily acquired
G. Long shelf life.
H. Reasonable cost
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