Control of Microbial Growth A few terms • Bacteriostatic: inhibits bacterial growth • Bactericidal: something capable of killing bacteria • Antiseptic: an agent that is used to inhibit/kill bacterial growth on skin and mucus membranes • Disinfectant: an agent that is used to inhibit/kill bacterial growth on inanimate objects What factors influence the success of microbial control? • What type of organism(s) are you targeting? • What type of environment are you treating? Are all microbes equally sensitive? What parts of a bacterial cell are sensitive to physical treatments and chemicals? • Plasma membrane • DNA and proteins ** Physical Methods • Heat – Dry: kills by oxidation, ex: incineration – Moist: promotes coagulation of proteins • Boiling (100oC) • Autoclave(121oC, 15lbs/sq inch) • Pasteurization – Classic-63oC for 30 minutes – HTST-72oC for 15 seconds – UHT-140oC for 3 seconds Autoclave Autoclaves work due to steam under pressure **Indicators used in autoclaving** What does a color change on the autoclave tape indicate? Why would it be necessary to also place an indicator in the center of the pack? What type of biological indicator is used for quality assurance in an autoclave monitoring program? Why? Where would the biological indicator be placed during the autoclaving procedure? What if the substance is heat sensitive? • Filtration is the best choice • Pore sizes can be either .45um or .22um Physical Methods • Low temperature: freezing does not kill bacteria, most cultures are stored at -80oC • Dessication: remove the water and bacteria can remain viable Physical Methods • Radiation – Ionizing radiation (gamma rays, x-rays) – Non-ionizing radiation (uv rays) – Microwaves Electromagnetic Spectrum Chemical Methods • Most are only able to reduce the numbers of organisms, not achieve sterility • Types of chemicals – – – – – – – Phenol and phenolics Halogens Alcohols Heavy metals Soaps Quaternary Ammonium compounds Biguanides As a general rule: if you need to achieve sterility then you should be thinking about physical, not chemical methods of microbial control. Evaluation of chemicals: disc diffusion method Chemical control: choosing the right germicidal chemical • Goal: sterility vs. disinfection; level of disinfection required dictates potency of chemical required • Toxicity: risk benefit analysis • Activity in presence of organic material: most are diminished or inactivated • Sensitivity of the material to be treated • Residue: toxic or corrosive vs residual desired antimicrobial effect • Cost and availability • Storage and stability: concentrate vs stock solution • Environmental risk: antimicrobials in the environment Chemical control • Phenolics Destroy cytoplasmic membranes, denature proteins (binds to tyrosine, alters cell membranes) • Triclosan • Hexachlorophene Target: vegetative bacteria +/- mycobacteria; not sporocidal; inadequate if viricidal activity is required; leave an active antimicrobial residue Chemical control • Halogens 1. Iodine (binds to tyrosine, alters cell membranes) • Tincture • Iodophore Target: all bacteria, fungi, most endospores, and some viruses 2. Chlorine (strong oxidizing agent) Target: all types of microbes and viruses; inactivated by organic matter Chemical Control • Alcohols – Mechanism of action is protein denaturation, can also dissolve lipids – Most common are ethanol and isopropanol Target: kill vegetative cells and fungi, do not kill endospores and some naked viruses Chemical control • Heavy metals exert “oligodynamic action” = ability of very low concentrations of metals to kill microbes: interfere with function of enzymes by binding to –SH • Silver (and formerly, copper, mercury, lead, arsenic) Chemical control • Surfactants (soaps) – Little value as an antiseptic – Acid-anionic surface-active sanitizers are important in cleaning of dairy equipment and utensils Chemical control • Quaternary Ammonium Compounds (Quats) – Group of positively charged detergents Action: react with the cell membrane → reduced surface tension, membrane damage Target: kill many vegetative bacteria and enveloped viruses, fungi; do NOT kill endospores, Mycobacteria, or naked viruses Chemical control – Biguanides (chlorhexidine) • • • • Used in antiseptic products Adheres to skin and mucus membranes, low toxicity Action: membrane disruption Target: wide range of bacteria, fungi, and some enveloped viruses; does not kill endospores; will not destroy naked viruses Which antiseptic works the best?