The Terminology of Microbial Control Sterilization-removing ALL microbial life; heat Sterilant- a sterilizing agent Commercial sterilization- killing C. botulinum endospores in food; used so food is not destroyed Disinfection- removing pathogens not including endospore Antisepsis- removing pathogens from living tissue Degerming- mechanical removal of microbes from a limited area; using an alcohol swab Sanitization- lowering microbial counts on eating utensils to a public health safety number

The Terminology of Microbial Control Biocide/germicide- killing microbes Fungicide- kills fungi Virucide- inactivates viruses Bacteriostasis- inhibiting, not killing, microbes Sepsis- microbial contamination Asepsis-the absence of significant microbial contamination Aseptic surgery techniques prevent microbial contamination of wounds

Rate of Microbial Death Microbes die at a constant rate. Each minute of treatment kills 90% of REMAINING microbes.

Effectiveness of Treatment Depends on: Number of microbes Environment (organic matter, temperature, biofilms) Time of exposure Microbial characteristics

Actions of Microbial Control Agents Alteration of membrane permeability- damages lipids or proteins. Damage to proteins- enzyme damage due denaturation Damage to nucleic acids- cell can no longer replicate or carry on normal metabolic functions

Question: Would a chemical microbial control agent that affects plasma membranes affect humans? 7-3

Physical methods of microbial control

Heat Thermal death point (TDP): lowest temperature at which all cells in a culture are killed in 10 min Thermal death time (TDT): time during which ALL cells in a culture are killed Decimal Reduction Time (DRT): Minutes to kill 90% of a population at a given temperature

Figure 7.10 A comparison of the effectiveness of various antiseptics.

Moist Heat Sterilization Moist heat denatures proteins by breaking H bonds Types: Boiling- can kill most bacterial pathogens and viruses in 10 minutes; endospores up to 20 hours Autoclave: steam under pressure; 15psi, 121C, 15 minutes; can kill endospores Used for media, instruments, dressings, IV equipment, syringes, ets. Time and temperature can change due to size Steam must contact item’s surface

Figure 7.2 An autoclave. Exhaust valve (removes steam after sterilization) Operating valve (controls steam from jacket to chamber) Steam to chamber Safety valve Pressure gauge Steam Door Steam chamber Air Perforated shelf Sediment screen Steam jacket Thermometer Automatic ejector valve (thermostatically controlled; closes on contact with pure steam when air is exhausted) Pressure regulator for steam supply To waste line Steam supply

Figure 7.3 Examples of sterilization indicators.

Pasteurization Reduces spoilage organisms and pathogens in some food and liquids without damaging taste of product Equivalent treatments 63°C for 30 min High-temperature short-time: 72°C for 15 sec Ultra-high-temperature: 140°C for <1 sec Thermoduric(heat resistant) organisms survive

Dry Heat Sterilization Kills by oxidation Dry heat Flaming Incineration Hot-air sterilization Hot-Air Autoclave Equivalent Treatments 170˚C, 2 hr 121˚C, 15 min

Filtration HEPA- removes microbes >0.3 µm Membrane filtration- removes microbes >0.22 µm

Physical Methods of Microbial Control Low temperature inhibits microbial growth (bacteriostatic) Refrigeration Deep-freezing Lyophilization High pressure denatures proteins Desiccation prevents metabolism Osmotic pressure causes plasmolysis

Radiation Ionizing radiation (X rays, gamma rays, electron beams) Damages DNA Can go through barriers Used by the food industry on meats, vegetables, and spices Also used on dental and medical supplies Nonionizing radiation (UV) Has to be in direct contact Microwaves kill by heat; not especially antimicrobial

Principles of Effective Disinfection Concentration of disinfectant Time Substance the disinfectant is being used against: Organic matter pH

Evaluating a Disinfectant Use-Dilution Test Disk Diffusion Method Metal rings dipped in test bacteria are dried Dried cultures are placed in disinfectant for 10 min at 20°C Rings are transferred to culture media to determine whether bacteria survived treatment Filter paper is soaked with a chemical Placed on an agar plate with an organism on it Look for zones of inhibition

Staphylococcus aureus Pseudomonas aeruginosa Figure 7.6 Evaluation of disinfectants by the disk-diffusion method. Zone of inhibition Chlorine Chlorine Chlorine O-phenylphenol O-phenylphenol Hexachlorophene O-phenylphenol Hexachlorophene Hexachlorophene Quat Quat Quat Staphylococcus aureus (gram-positive) Escherichia coli (gram-negative) Pseudomonas aeruginosa (gram-negative)

If you wanted to disinfect a surface contaminated by vomit and a surface contaminated by a sneeze, why would your choice of disinfectant make a difference? 7-7

Phenol and Phenolics Disrupt plasma membranes Lysol Bacteria

Bisphenols Hexachlorophene Triclosan Disrupt plasma membranes Used in hospital soaps Staphyloccocci and Streptococci Triclosan Found in soaps, toothpaste, cutting boards, plastic kitchenware Bacteria and yeast

Biguanides Chlorhexidine Disrupts plasma membranes Surgical hand scrubs and preoperative scrubs Bacteria

Halogens Iodine Chlorine Alters protein synthesis and cell membranes Bacteria, endospores, fungi, viruses Chlorine Prevents enzyme system from functioning Bleach Disinfects drinking water, swimming pools, and sewage Bacteria and viruses

Alcohols Ethanol, isopropanol Denature proteins, dissolve lipids Require water Bacteria and fungi

Heavy Metals Silver, Mercury, and Copper Denatures proteins Silver nitrate- may be used to prevent gonorrheal ophthalmia neonatorum Silver sulfadiazine- used as a topical cream on burns Copper sulfate- is an algicide

Surface-Active Agents, or Surfactants Soap- mechanical removal of microbes Degerming Acid-anionic detergents- dairy utensils and equipment; bacteria Sanitizing Quaternary ammonium compounds (cationic detergents)- fungicidal, amebicidal, virucidal Bactericidal, denature proteins, disrupt plasma membrane

Chemical Food Preservatives Organic acids Inhibit metabolism Sorbic acid, benzoic acid, and calcium propionate Control molds and bacteria in foods and cosmetics Nitrite Prevents endospore germination Put in ham, bacon, hot dogs, sausage Antibiotics Nisin and natamycin prevent spoilage of cheese

Aldehydes Inactivate proteins by cross-linking Bactericidal, tuberculocidal, virucidal, sporicidal Used on hospital instruments, endoscopes, and respiratory therapy equipment Glutaraldehyde, formaldehyde

Gaseous Sterilants Denature proteins Sporicidal Use: heat-sensitive material Ethylene oxide

Plasma Free radicals destroy microbes Used for tubular instruments used for arthroscopic and laparoscopic surgeries

Supercritical Fluids CO2 with gaseous and liquid properties Use: medical implants(bones, tendons, ligaments)

Peroxygens Hydrogen Peroxide Use: contaminated surfaces

Figure 7.11 Decreasing order of resistance of microorganisms to chemical biocides.

Table 7.7 The Effectiveness of Chemical Antimicrobials against Endospores and Mycobacteria