Microbial Control Lecture Notes PDF

8/21/2024...

8/21/2024 Learning Objectives  Explain different methods of microbial control Microbial Control  Determine best method of controlling microbes  Determine the suitability of materials/substances for autoclaves Peter Keyel, PhD MBIO3400 Images from Tortora et al Microbiology: An Introduction 14th edition, unless otherwise specified 1 2 1 2 Physical, Chemical, and Cells Die at a Logarithmic Rate Biological Control of Microbes  A variety of terms are used to describe antimicrobial control measures: - Sterilization: killing of all living organisms - Disinfection: killing or removal of pathogens from inanimate objects - Antisepsis: killing or removal of pathogens from the surface of living tissues - Sanitation: reducing the microbial population to safe levels 3 4 3 4 Physical Agents That Preserve Microbes Physical Agents That Kill Microbes  Cold  High temperature - Low temperatures slow growth and preserve strains - Moist heat is more effective than dry heat. - Refrigeration temperatures used for (4oC–8oC) - Boiling water (100oC) kills most cells. food preservation because it slows growth - Killing spores and thermophiles usually requires a - For long-term storage of cultures combination of high pressure and temperature. - Placing solutions in glycerol at –70oC - Steam autoclave - Lyophilization or freeze-drying - 121oC at 15 psi for 20 minutes 5 6 5 6 1  8/21/2024 The Autoclave First choice method to sterilize most equipment Uses steam heated to 121°C and 15 psi above normal pressure for 15 min Steam sterilizes much faster than hot air, so air evacuated from chamber If the steam does not touch it, it does not get sterilized Indicator tape used to indicate sterility Also used to decontaminate waste 7 8 7 8 The Autoclave: Warnings The Autoclave: Limitations Be careful when opening the chamber: hot steam escaping Cannot be used for everything Never tighten caps on bottle—pressure difference will cause explosion High temperature and pressure will destroy certain materials: Also, steam must be able to reach liquid/solid to be sterilized Ex: polystyrene, non-borosilicate glass and styrofoam Autoclave can become jammed if debris blocks parts Other materials will fail with age (other plastics, borosilicate glass) Double bag as needed and use secondary containment Secondary containment in case primary container fails Aluminum foil often used to cover glassware like beakers, flasks Not all liquids can survive autoclaving Volatile organics do NOT go in autoclave—explosion hazard Biologic substances usually destroyed (except prions) 9 Ex: many proteins, antibiotics, fetal calf serum, tissue culture medium 10 9 10 Other Heat Control Pasteurization  Boiling one approach to preserving food - Many different time and temperature combinations can be used.  Also wrecks taste/some foods - LTLT (low temperature/long time)  Pasteurization developed to preserve wine - 63oC for 30 minutes - HTST (high temperature/short time)  Uses lower temperatures to be gentler to food - 72oC for 15 seconds  Pasteurization does not sterilize - UHT (ultra high temperature)  Achieves 5-log reduction (99.999%) reduction - 135oC for at least 2 seconds  Kills probiotic bacteria so milk spoils not sours  Vitamin/mineral loss not nutritionally relevant 11 12 11 12 2  8/21/2024 Physical Agents That Kill Microbes Physical Removal  Filtration  Irradiation - Micropore filters with pore sizes of 0.2 m can - Ultraviolet light remove microbes, from solutions. - Has poor penetrating power - Samples from - Used only for surface sterilization 1 ml to several - Gamma rays, electron beams, and X-rays liters can be drawn through - Have high penetrating power a membrane filter - Used to irradiate foods and other heat-sensitive items by vacuum or can be forced through it using a syringe. 13 14 13 14 - Air can also be sterilized by filtration. Physical Removal - Laminar flow biological safety cabinets force air through HEPA filters, which remove > 99.9% of airborne particulate material 0.3 μm in size or larger. Figure 5.28 15 16 15 16 Chemical Agents: Commercial Disinfectants and Antiseptics Disinfectants/Antiseptics  These include:  A number of factors influence the efficacy of a given chemical agent, including: - Phenol - The presence of organic matter - Ethanol - The kinds of organisms present - Iodine (Wescodyne and Betadine) - Corrosiveness - Chlorine - Stability, odor, and surface tension - Ethylene oxide (a gas sterilant)  These damage proteins, lipids, and/or DNA - Are used to reduce or eliminate microbial content 17 from objects 18 17 18 3  8/21/2024 Commercial Disinfectants and Antiseptics  Alcohols kill quickly and efficiently  Ethanol and Isopropanol most commonly used  Most active 60%-90%  Do not kill spores; cannot penetrate protein-rich  Bleach kills spores  Bleach is corrosive to metal and people 19  Fresh 10% bleach or 25% for spill control 20 19 20 Chemical Agents Antibiotics  Antibiotics are chemical compounds synthesized by one microbe that kill or inhibit the growth of other microbial species.  Penicillin mimics part of the bacterial cell wall. - Prevents cell wall formation and is bactericidal  Other antibiotics target: - Protein synthesis - DNA replication - Cell membranes 21 22 21 22 Biological Control of Microbes  Biocontrol is the use of one microbe to control the growth of another. - Probiotics contain certain microbes that, when ingested, aim to restore balance to intestinal flora - Lactobacillus and Bifidobacterium - Phage therapy aims to treat infectious diseases with a virus targeted to the pathogen - A possible alternative to antibiotics in the face of rising antibiotic resistance 23 24 23 24 4