Physical and Chemical Control of Microbes Physical Agents heat or radiation Chemical Agents disinfectants or antiseptics Important Terms 1. Sterilization process of killing all viable microbes 2. Bactericide kills bacteria (except endospores) 3. Germicide kills pathogenic microorganisms 4. Disinfection destroy vegatative pathogens, but not bacterial endospores 5. Sanitization mechanical removal of microbes How antimicrobial agents work I. Cell wall - prevent production or digest cell wall (penicillins) - cell becomes fragile and easily lysed - detergents and alcohols disrupt cell walls (esp. gram neg) II. Cell membrane - Detergents (surfactants) have hydrophobic and hydrophilic regions - mess up interactions that hold cell membrane together - harmful chemicals into cell, important ions out of cell III. Protein or nucleic acid synthesis - Chloramphenicol binds to ribosomes and prevents peptide bond formation - Some agents bind to DNA preventing transcription - Radiation (UV, X-rays) may cause mutations that inactivate DNA IV. Alter protein function - Denature proteins (cook an egg) to alter protein function - Moist heat or alcohols can mess up protein shape and function 1
Using heat to control microbes Moist Heat 1. Autoclave use high heat and pressure (sterilization) 2. Pasteurization high temp, short time (not sterile) kills 97-99% vegatative bacteria, but doesn t kill endospores 3. Boiling disinfection, kills most bacteria, but not endospore forming Dry Heat 1. Incineration using high temperatures to kill bacteria - used when we flame inoculating loops - ensures total destruction of of microbes 2. Dry Oven heat 150 to 180 C for 2-4 hours (kills endospores) - dry heat use limited to substances that withstand high temps. Use of Radiation 1. Ionizing radiation ejects electrons from orbits around nuclei - Gamma rays or X-rays are examples - often referred to as cold sterilization 2. Nonionizing radiation raises e- energy levels, changes bonds - UV radiation breaks bonds between nitrogen bases of DNA - may cause abnormal bonds in DNA which mess up replication, transcription, translation) 2
Some types of Germicides Halogens (F, Br, I, Cl) used in 1/3 of antimicrobial chemicals Alcohols hydrocarbons with OH group(s), may disrupt cell membranes or denature proteins Hydrogen Peroxide (H 2 O 2 ) produces free radicals that are toxic to cells (cells don t produce enough catalase to neutralize all the H 2 O 2 ) Detergents and Soaps may disrupt cell membranes or assist in removal of microbes from surfaces Antimicrobics Antimicrobial drugs Antibiotics produced by microorganisms to destroy other microbes (adaptation resulting from natural selection) Synthetics derived in labs from dyes and organic compounds Narrow spectrum work against only a few microbes ex. Bacitracin works against some gram positives Broad spectrum work against a wide variety, for ex. Tetracyclines work against many gram positive and gram negative bacteria 3
Primary sites of action of antibiotics 1. Block protein synthesis - 50S (chloramphenicol, erythromycin) - 30S (aminoglycosides, tetracyclines, streptomycin) 2. Cell membrane - polymyxins 3. Cell wall (block synthesis and repair) - penicillins, cephalosporins, vancomycin, bacitracin - beta-lactam enzyme (penicillin / ampicillin) weakens peptidoglycan 4. DNA (inhibit replication and transcription) - quinolones (inhibit gyrase), Rifampin (inhibit RNA polymerase) 5. Inhibit metabolism - Sulfa drugs shut down folic acid metabolism Antimicrobial Resistance How does this develop? Natural selection inheritance of beneficial mutations - variation in populations - overproduction of offspring - struggle for existence - differential survival and reproduction Resistance (R) factors transfer genes for resistance through conjugation, transformation or transduction Gene transfer may involve different species Bacterial evolution somewhat Lamarckian 4
Mechanisms of drug resistance Drug inactivation ex. Beta-latamase enzyme inactivates some penicillins and cephalosporins (transformation lab ampicillin resistance gene) MDR (multidrug resistant) have pumps that expel drugs from the cell (Staphylococcus, Streptococcus, E. coli) Change drug receptors Erythromycin resistance associated with changes in the 50S ribosome unit, some types of penicillin resistance due to changes in binding proteins of the cell wall 5