Control of Micro-Organisms Physical Chemical Antibiotics and Chemotherapeutics Physical Control of Microorganisms GOAL = STERILIZATION: removal of all life forms, inc. bacterial spores, viruses, etc. 2004 by Jones and Bartlett Publishers 1
Physical Control with Heat Heat causes lethal changes in cells: 1. Proteins denature 2. Cell membrane disrupted 3. Cellular water evaporates Thermal death Thermal death time: Time needed to kill a population of cells at a given temperature (E. coli 55 degrees Celsius 2 hours until they are dead) Thermal death point: minimum temperature needed to kill a population 2
Direct Flame Most rapid method to sterilize Lab: loop and bunsen burner Incinerate cattle and goats if they have anthrax Hospital red bags - biohazard The hot-air oven Dry heat no steam Good for precision glassware that is damaged by water Limited uses because they take a long time to cool down 3
Moist Heat The Autoclave Uses steam and high pressure to kill microbes Hot Water molecules penetrate microbes better than Hot air Achieve sterilization quickly: 45 minutes complete Low temperature: 121 celsius PV = nrt 4
Pasteurization DOES NOT STERILIZE Kills human pathogens Kills food spoilage organisms Temp ~63 degrees Celsius Who survives? Thermophiles, but these don t cause human disease Milk, Yogurt, Cheese, OJ, etc. Hot oil Good to use on sharp instruments, such as scalpels and dental probes Oil doesn t rust or corrode metals like water does, sharps stay sharp Problem: inconvenient, difficult to remove oil after using it 5
Filtration Used to purify liquids and air Traps all microbes larger than the pore size of the filter (0.2 micrometers) HEPA filters used in isolation rooms (TB) or for Burn wards Made of cellulose, crushed glass or diatomaceous earth Viruses can pass through Ultraviolet light Damages microbes DNA Used on food products (cereals, vegetables, fruits) Can also be used to kill microbes on surfaces and in the air (UV lamps) 6
Ultrasound High frequency sound waves propagated in liquids cause cold boiling or bubbling; when bubbles burst, shock wave is created that kills microbes!! Examples: Sonic toothbrushes; lithotripsy of kidney stones Chemical Control of Microbes 7
History Levels of control Sterilize remove all life forms Disinfect destroy all pathogens Sanitize reduce microbial populations to a safe level (government defined) Degerm remove organisms from a surface (e.g. hand washing) 8
Antiseptics and disinfectants Antiseptics: used on living organisms Disinfectants: used on inanimate objects Evaluation of Chemical Agents: The PC test (Phenol coefficient) 9
Phenol Denatures Proteins Caustic to Skin Now chemically modified Examples: Sucrets throat lozenges; Cresols used on telephone poles Halogens Group VII want one electron to be happy, steal it from microbes Chlorine public swimming pools Fluorine toothpaste, drinking water Iodine used on wounds and to clean entry site before catheter insertion 10
Heavy metals Mercury (HgCl 2 ) Copper (CuSO 4 ) Silver (AgNO 3 ) Work by cross-linking proteins together Mercury used on skin wounds Copper part of Bordeaux mixture Silver eyes of newborns; silvidine used on burns Alcohols Ethanol or Isopropanol ( rubbing alcohol) At least 63% to be useful clinically Bacteriocidal vs. Bacteriostatic Alcohol dissolve lipids (re: cell membranes) and denature proteins 11
Aldehydes Cross-links proteins together Used for embalming (preserving dead bodies) formalin Glutaraldehyde used on endoscopes before they enter ; kills spores on them (sterilization) Gases used in chemical control: Ethylene Oxide vs. Chlorine Dioxide Cross-links proteins Kills Spores Causes cancer, used on products only Cheap Cross-links proteins Kills spores Non-toxic to humans Expensive $$$$$$$ 12
Detergents Soaps / Shampoos They are positively charged, solubilize negatively charged cell membranes of microbes Dyes Bind to DNA, interfere with replication Gentian violet used in mouth to kill fungi (oral thrush) Also used on wounds (Acridine Orange) 13
Acids Lower the ph; most microbes are neutralphiles Examples: acetic acid (vinegar) used in pickling Benzoic acid: used on acne Antibiotics and Chemotherapeutic Agents Antibiotics: made by microbes, kill other microbes Natural products Treat infection Chemotherapeutics: made by humans in the lab; Synthetic products Also used to treat diseases/infections 14
Paul Ehrlich the magic bullet Salvarsan synthetic compound made from arsenic and phenol Kills syphilis-causing bacteria (spirochete) Sulfonamides = competitive inhibitors Domagk saves daughter from a blood infection Made from a dye How does it work? It binds to the enzymes bacteria use to make folic acid without folic acid, they perish. 15
Sulphamethoxazole Acyclovir Used to treat Herpes infections such as chicken pox, shingles, genital herpes, or cold sores /fever blisters How does it work? It resembles the G in DNA Virus tries to incorporate the fake G into its DNA, so the drug stops replication 16
Other Modern Chemotherapeutics Quinine/chloroquine/primaquine Malaria Isoniazid TB (tuberculosis) Metronidazole Amoebiasis / Giardia / Trichomonas Nalidixic acid + Fluoroquinolones (e.g. Cipro) Urinary Tract Infections / Gonorrhea / Chlamydia Antibiotics 17
Sites of Activity for Various Antibiotics Alexander Fleming - Penicillin Natural product of the mold Penicillium Stops bacteria from making cell wall peptidoglycan Allergic?? Take Cephalosporins, also a mold product 18
Bacterial Cell Wall Gram-Negative Cell Wall Peptidoglycan Penicillins 19
Cephalosporins Coltura originale di Cefalosporium del 1949 Aminoglycosides Antibiotic that binds to bacterial ribosomes and stops protein synthesis These were isolated from bacteria (Streptomyces) Include Streptomycin, Kanamycin, Gentamycin, etc. 20
Chloramphenicol Also binds to ribosomes and stops translation of proteins Used to treat infections of the nervous system (meningitis) Side effects include red blood cell deficiencies Tetracyclines Translation blocker General use, wide-spectrum antibiotic effective against a wide range of microbes Side effects: yellow staining of teeth or bone/kidney problems 21
Polymyxin B Affect the permeability of the cell membrane pokes holes Isolated from bacteria Used to treat gram negative infections Rifampicin Blocks Bacterial transcription Binds to RNA polymerase of bacteria Used to treat TB/Leprosy/MRSA 22
Antibiotic Resistance Mechanisms Hundreds of thousands of multidrug resistant infections in the USA each year (mostly in hospitals + nursing homes) 1. Reduce permeability to antibiotic 2. Actively pump out the antibiotic 3. Target modification: mutation changes the ribosome/cell wall enzymes so that the antibiotic doesn t bind there 4. Destroy the antibiotic (create enzyme that can degrade it) Penicillinases = β-lactamases An example of an enzyme made by some bacteria function destroy penicillin and related anti-biotics 23
The Expansion of Drug Resistance Antibiotic Susceptibility Assays Disk Diffusion Assay MIC assay 24
Antibiotics: misuse and abuse Antiviral agents 25
Antifungal agents Drugs for Parasitic Infection {QUININE PLANT} 26