BMS 212 Week 6
BMS 212 Week 6 BMS 212
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This 9 page Class Notes was uploaded by Brandon Czowski on Thursday March 17, 2016. The Class Notes belongs to BMS 212 at Grand Valley State University taught by Dr. Leonard in Winter 2016. Since its upload, it has received 27 views. For similar materials see Microbiology in Biomedical Sciences at Grand Valley State University.
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Date Created: 03/17/16
Chapter 9 Sterilization: process of removing/destroying all viable microorganisms (includes endospores/viruses) • Used on inanimate objects, physical process usually involves heat Sanitation: technique used to remove microorganisms and debris from inanimate surface to a safe level in public health settings (hospitals) Disinfection: destruction of vegetative cells on inanimate surface (home setting) • Concentrated—harsh on living organisms, certain amount of expose time needed Antisepsis: chemical removing microorganisms from living tissue • Reduces number of microbes, less concentrated than disinfectants Degermination: cleansing technique on animate objects by mechanical means (scrubbing) Germicide: chemical that kills on animate and inanimate objects (catch-all) Death: permeant loss of reproductive capability -cide/-cidal: to kill -stasis/-static: prevents growing Mechanisms of antimicrobial agents • Alters cell walls and membranes o Weakening cell walls increase chance of cell death from osmotic pressure o Loss of membrane integrity causes cellular contents to leave membrane o Enveloped viruses lose ability to attach to a host • Damages proteins and nucleic acids o 3D shape of protein crucial to function o Mutations alter nucleic acids Factors influencing efficiency of agents • Treatment site • Exposure time • Concentration • Actively growing • Temperature • pH • Organic matter—antimicrobial chemicals don’t work well against Physical Methods of Control Heat: permanently denatures proteins, destroys membranes, and cells walls while disrupting nature of nucleic acids Determining organisms tolerance to heat: 1. Thermal death point: lowest temperature that kills an organism in a given time (10 minutes) 2. Thermal death time: shortest time that kills an organism at given temperature Types of heat: • Moist Heat o Boiling: disinfection & sanitation—kills vegetative cells within 10 minutes o Pasteurization: sanitation—heats enough to kill pathogens o Steam under pressure “Autoclave”: 121 degrees Celsius at 15 psi for 15 minutes (additional 15 psi on top of atmospheric 15, total 30 psi) § Only heat resistant material, not good for oils/powders § Vial can test if medium is viable • Breaking column and yellow means NOT sterile • Breaking column and red means medium sterile § Autoclave tape turns white to black indicators meaning autoclaved • Dry Heat: dehydration and oxidation of proteins and cell components o Requires: higher temps, longer times, (160 degrees Celsius for 2 hours) o Able to use on powders and oils o Burning/incineration: oxidizes all organic matter, used on heat-stable instruments, waste can be disposed Refrigeration: 4 degrees Celsius slows bacterial growth (except psychrophiles and Listeria monocytogenes) Freezing: storage of cells at -80 degrees Celsius Desiccation: drying that inhibits growth of most pathogens from the lack of liquid water Lyophilization: combining freezing with liquid nitrogen and drying using a vacuum to remove water; stores cells and food indefinitely Filtration: sterilizes heat-sensitive material (antibiotics, vaccines, enzymes) Osmotic pressure: high solute solutions dehydrate cells Radiation • Ionizing: rapid, no heat with high penetration ability to break DNA, oxidize bonds and create hydroxyl radicals o Electron beams (cathode ray machine): low penetration but rapid use with spices, meats, and medical supplies o Gamma Rays: slowly emitted from radioactive elements that penetrate better but slowly accepted to use with food o X-rays: travel farthest but lower energy takes much longer • Non-ionizing: wavelengths greater than 1 nm don’t have the energy to displace electrons but can change 3D structure of proteins and nucleic acids o UV light: can create pyrimidine dimers in DNA, used for treating liquids and lab equipment since low penetrating power (drawbacks: low power, harmful to humans) Chemical Methods of Control Factors of effectiveness: concentration, exposure time, nature of microorganism, pH, temp Good chemicals: low side effects, solubility, useful high dilutions, broad spectrum, effective in organic contaminants Low Level Disinfectants & Antiseptics • Soaps & detergents 2 o Surfactants: reduce surface tension of liquids o Disrupt cell membranes and walls § Quaternary ammonias: NOT effective against endospores, naked viruses, or mycobacteria (Pseudomonas is resistant) • Heavy Metals: binds functional groups (disulfide bonds –SH) causing inactivation/denaturing of proteins o 1% silver nitrate drops: given to all newborns in eyes to protect from mothers with gonorrhea o Copper sulfate: inhibits chlorophyll to prevent algae blooms o Copper based hand gels o Advantages: small amounts, target all cells (except endospores) o Disadvantages: high toxicity internally, allergies, resistance Intermediate to Low Disinfectants & Antiseptics • Phenols: disrupts cell walls, membranes, denatures proteins (especially useful for inserting into membranes) o –cidal: kills organisms o Broad spectrum in low concentrations o Maintains activity and works with organic loads o J. Lister used phenol is OR to control infections o Disadvantages: toxicity and irritation, resistance in some organisms Intermediate Level • Halogens: -cidal (30 minutes for vegetative and hours for endospores), acts as strong oxidizer and cuts sulfhydryl bonds within proteins o Chlorine: strong, works against organic material(best to remove first), sensitive to light (loses effectiveness) § Elemental—used in sewage, chloramine—chlorine and ammonia in liquid forms isn’t toxic, hypochlorites, chlorine oxide (gas)—disinfects large areas o Iodine: less susceptible to organic material, kills classes from high to low based on concentration § Disadvantage: irritating, staining, minor allergies § Tincture: iodine dissolved in alcohol o Idophor: complex of iodine and carrier molecule (protein) as organic compound § Less irritating § Medicine: betadine § Dairy: wipes utters with betadine • Alcohols: dissolve cell membranes and denatures proteins o Ethanol and isopropanol—concentration determines disinfectant or antiseptic o Most effective in 70-95% rather than 100 b/c water allows for activity o Non-irritating (except eyes and dries skin), inexpensive o Disadvantage: inhalation hazard, effects nervous system 3 High level Disinfectants & Antiseptics • Hydrogen Peroxide: denatures proteins and breaks membranes by forming hydroxyl radicals; closest to “perfect disinfectant/antiseptic” o Leaves not residue and doesn’t damage surface o Photosensitive o 3% common as antiseptic or low disinfectant o 30-35% liquid/vapor is sporicidal • Aldehydes: cross links DNA and proteins in cells o Glutaraldehyde (liquid) o Formaldehyde (gas) § Formalin (formaldehyde dissolved in water) o Retains potency in organic material, non-corrosive (treat lab and medical material) o High toxicity (carcinogenic) and unstable at increased pH & temperature • Gaseous Agents: cross links nucleic acids and proteins o Ethylene oxide, propylene oxide o Sterilized inanimate objects: large objects that are bulky/heat-sensitive and cannot be autoclaved, flush when finished to remove residue o Hazardous Chapter 10 Chemotherapeutic: any drug used in treatment of a disease Antimicrobial agent: used to treat infection regardless of source (all inclusive) Antibiotic: antimicrobial agent produced by an organism that targets bacteria (biosynthesized) Semisynthetic: antibiotic that has been modified in a lab Synthetic: completely synthesized in a lab Concepts of Antimicrobial therapy selective toxicity: antimicrobial agent must be more toxic to pathogen rather than host— possible by targets pathways/structures of infectious organisms that the host lack • Safety/side effects o Allergies (hives or rashes) o Toxicity (measured with therapeutic index)—can disrupt normal flora o Disruption ofNormal flora is good bacteria present in our bodies, antibiotics kill this normal flora lowering our diversity and results in less effective immune system • Spectrum of activity o Broad vs. narrow: general use of targeting single infection o –cidal vs. –static: kill vs. preventing growth • Efficacy: how well the antibiotic kills a certain bacterium Mechanisms of Action • Inhibition of cell wall synthesis—vancomycin, penicillins, cephalosporins • Inhibition of protein synthesis 4 o 30S—tetracycline, streptomycin o 50S--erythromycin • Disrupt cytoplasmic membrane--polymiixins • Inhibiting general metabolic pathway—sulfaoamides & trimehoprim • Inhibition of DNA/RNA synthesis o DNA gyrase—quinolones o RNA polymerase--rifampin • Inhibiting pathogens attachment/recognition of host Effectiveness • Concentration: drugs with high therapeutic index are safer that those with lower o Therapeutic index = toxic dose (most a patient can handle)/ therapeutic dose (minimum to stop growth) • Administration: certain concentration must be maintained to be effective o Oral: simplest, concentrations in body lowest o Intramuscular: absorbed by blood vessels near muscles, large peak at first, dwindles fast o Continuous intravenous: high concentrations achieved quickly then filtered quickly by kidneys, unless continuous quick & maintained • Resistance o Intrinsic: naturally resistant o Acquired: mutation or gene transfer cause organism to be resistant • Measuring Effectiveness: o Kirby-Bauer Test (Diffusion Susceptibility) test Efficacy : measure the zone of inhibition to determine whether the organism is susceptible, intermediate, or resistant to an antibiotic o Minimum inhibitory Concentration Test (MIC): determines lowest concentration of the antibiotic that inhibits growth Discovery of Antibiotics • Paul Ehrlich—Salvarsan; treat syphilis, focused on treating rather than prevention • Gerard Domagk—1930’s found sulfa drugs (synthesized) • Alexander Fleming (1929): Penicillin published in article describing appearance • Florey & Chain (1939): found Fleming’s article and isolated to create penicillin Origins • Bacteria o Bacillus—in soil o Streptomyces—prokaryotic but produces endospores like eukaryotes • Fungi o Penicillium o Cephalosporium (Acremonium) Mechanisms of Action 5 • Cell wall inhibitors o Beta-lactams cell wall inhibitors: Inhibit cell wall synthesis by inhibiting peptidoglycan formation by binding to transpeptidases enzyme that cross link NAM sub-units § Mechanism: inhibit peptidoglycan formation irreversibly binding transpeptidases enzymes that cross link NAM subunits § Growing cells are susceptible § Penicillin, Cephalosporins, carbapenems with functional protein rings (beta-lactams) § Beta-lactamases(produced by bacteria): breaks ring in penicillin to create inactive structure § Penicillin: Narrow Spectrum • Penicillin G: effective against Gram+, low cost, effective concentration quickly with good penetration and low side effects; resistant from beta-lactamase produced by bacteria, required injection—acid liable, absorbed poorly by small intestine • Penicillin V: more stable in acidic conditions than G, able to be taken orally but poor absorption • Methicillin (semisynthetic derivative of penicillin): resisted action of beta-lactamases in bacteria, some side effects, acid liable (needed injection); MRSA (Staph. Aureus resistant) § Penicillin Broad Spectrum • Ampicillin: stable in acid, better absorption; allergies and lots of resistance • Amoxicillin: acid stable, good absorption in less frequent doses; resistance poses issue • Augmentin: clavulonic acid (beta-lactamase inhibitor) + amoxicillin § Hypersensitivity to Penicillins • Rashes, hives, diarrhea; anaphylactic shock, death § Cephalopsorins: produced by Acremonium cephalosporium (Cephalexin, Keflex) • Mechanism: bind transpeptidases leading to cell lysing • Structure: greater resistance to beta-lactamases • Toxicity: dependent on generation • Administration: some need injection, some oral • Generations o 1 : narrow spectrum (G+ only) o Higher generations—3-5 broad spectrum from ability to th blood-brain barrier (5 gen. used for MRSA) § Carapenems: KPC (Klebsiella pneumonia—carabapenem resistant) • Resistant against most beta-lactamases 6 • Mechanism: target cell wall • Broad spectrum—only G- • Administration: never orally o Non-beta lactam cell wall inhibitors: § Narrow spectrum • Vancomycin o Mechanism: physically obstructing formation of peptide brides to NAM units o Toxicity: issues of kidney & ear o Administration: injection—poorly absorbed o VRE (vancomycin resistant enterococci); treats MRSA • Bacitracin o Mechanism: prevent synthesis of mycolic acid layer of complex cell walls o Toxicity: cidal & some side effects o Synthetic o Mycobacterium genus with extra mycolic layer (leprosy, tuberculosis) • Protein synthesis Inhibitors: generally broad spectrum, Streptomyces produces many of these drugs o Aminoglycosides § Mechanism: bind to 30S ribosomal subunit causing a change in shape resulting in codons being read improperly (incorrect amino acid seq.) § Highly toxic, broad spectrum § Ex. Kanamycin, gentamicin, streptomycin, neomycin • Streptomycin: natural product of Streptomyces that acts by changing shape of 30S subunit o Toxic to ears o Uses: bubonic plague, TB, tulermia o Broad spectrum—G- • Gentamicin: produced by genus Micromonospora that bind also to 30S subunit o Toxic to kidneys and ears o Poor absorption in small intestines o Uses like streptomycin, especially against Pseudomonas o Tetracyclines § Mechanism: target and binds to tRNA docking site (A-site) on 30S subunit prohibiting protein synthesis § Broad spectrum, inexpensive semisynthetic § Administration: orally § Uses: acne, prostate infection, intracellular parasites, agriculture (food) 7 § Toxicity/Side effects: stained teeth, interference with birth control § “Static” because it is physically blocking the A-site, preventing binding o Macrolides: alternative for allergies with penicillin § Mechanism: target 50S subunit and prevents ribosome from moving, stopping synthesis—“static” § Naturally produced—G+ § Administration: orally § Toxicity: allergies and GI issues • DNA/RNA Inhibitors o Rifampin § Mechanism: blocks RNA polymerase halting transcription § Semisynthetic that’s easily absorbed § Cidal § Uses: TB & leprosy § Side effects: red/orange bodily fluids o Quinolones (nalidixic acid) & Fluoroquinolones (ciprofloxacin) § Mechanism: target DNA gyrase (specifically towards prokaryotic DNA) and inhibits enzyme from coiling/uncoiling DNA, incorrect synthesis § Cidal § Broad spectrum, stable in acid § Administration: oral, readily absorbed across intestine—able to cross blood-brain barrier § Uses: UTI’s, STD’s, respiratory infection, anthrax § Side effects: tendon disruption in old adults, seizures in infants • Cell Membrane Inhibitors o Polymyxin: produced by Bacillus polymyxa effective against susceptible cells § Mechanism: destroys bacterial membrane by inserting themselves to Gram-negative (especially affective against Pseudomonas) § Cidal § Toxicity: human kidneys, usually used topically § One of antibiotics in triple antibiotic ointment (including bacitracin & neomycin) • Metabolic Pathway Inhibition o Sulfonamides & Trimethoprim (Bactrim or SXT): target folic acid biosynthesis— synergist effect § Mechanism(sulfonamides): structural analogs of PABA that that compete for the active site for the enzyme that produces dihydrofolic acid, inhibiting synthesis of folic acid § Mechanism(trimethoprim): structural analog that competes for binding site on enzyme responsible for converting dihydrofolic acid to THF § Synthetic—developed by Gerald Domagk (1932) 8 § Broad Spectrum: effective against G+/- and some protozoa & fungi, resistance is widespread § Toxicity: allergies highly common § Uses: UTI’s, acne, protozoan infection • Preventing Virus Attachment o Attachment antagonist: analogs of attachment/receptor proteins on cell that block binding sites for viruses cannot attach/enter host cell—deterring infection § Arildone & pleconaril are agonists of receptor on poliovirus and some colds 9
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