Micro Chapter 9 - Controlling Microbial Growth in the Environment-2.pdf

Chapter 9 Controlling Microbial Growth in the Environment  Lecture objectives Compare and contrast the different mechanisms of antimicrobial action Understand the clinical considerations when prescribing an antimicrobial Understand routes of admi...

Chapter 9 Controlling Microbial Growth in the Environment  Lecture objectives Compare and contrast the different mechanisms of antimicrobial action Understand the clinical considerations when prescribing an antimicrobial Understand routes of administration and safety/side effects Be familiar with how drugs become resistant and how this can be circumvented Most environments, including cars, are not sterile. A study analyzed 11 locations within 18 different cars to determine the number of microbial colony-forming units (CFUs) present. The center console harbored by far the most microbes (506 CFUs), possibly because that is where drinks are placed (and often spilled). Frequently touched sites also had high concentrations.  Some Food for Thought…. How clean is REALLY “clean”? We wash and vacuum our cars, but would you eat from the carpet? Silverware and dishes that other have been eaten from (i.e. restaurants) are put through a dishwasher, but how clean are they? That same cleaning procedure would not be appropriate for surgical tools… The term “clean” is relative. Because people do not normally eat from cars or carpets, these items do not require the same level of cleanliness that silverware does. Likewise, because silverware is not used for invasive surgery, these utensils do not require the same level of cleanliness as surgical equipment, which requires sterilization to prevent infection.  Some Food for Thought…. Why not play it safe and sterilize everything? Sterilizing everything we come in contact with is impractical, as well as potentially dangerous. Sterilization protocols often require time- and labor-intensive treatments that may degrade the quality of the item being treated or have toxic effects on users. We must consider the item’s intended application when choosing a cleaning method to ensure that it is “clean enough.” ALSO – sterilization would remove beneficial microbes!! Biosafety levels  Terminology of Microbial Control Inanimate items, such as doorknobs, toys, or towels, which may harbor microbes and aid in disease transmission, are called fomites. Two factors determine level of cleanliness needed and the subsequent protocol: 1. Application of the item. ▪ Example: Silverware vs. surgical items 2. Level of resistance to antimicrobial treatment by potential pathogens. ▪ Example: Washing clothing in a washing machine vs. canning foods ▪ Most bacteria that are found on things such as clothing and bedding are not harmful. Some such as Staphylococcus sp. or contagious fungi (think athlete's foot) are generally treatable. ▪ Clostridium botulinum is a dangerous pathogen that can grow in unproperly canned foods; must be done a specific high heat and pressure.  Terminology of Microbial Control For many clinical purposes, aseptic technique is necessary to prevent contamination of sterile surfaces. Aseptic technique - combination of protocols that collectively maintain sterility, or asepsis. Prevents contamination of the patient with microbes and infectious agents. Failure to practice aseptic technique may introduce microbes to the patient’s body, putting the patient at risk for sepsis. Systemic inflammatory response to an infection that results in high fever, increased heart and respiratory rates, shock, and, possibly, death. Can be due to infections in the blood or organs (bacteria, fungi, or viral) Medical procedures that carry risk of contamination must be performed in a sterile field. Free of all vegetative microbes, endospores, and viruses. Packaging and drapings, strict procedures for washing, application of sterilants. Terminology of Microbial Control  Selecting an antimicrobial agent Prefix -cides Bacteriocides, fungicides, viricides Methods of microbial control that kill the targeted microorganism Prefix –static Bacteriostatic, fungistatic Do not kill organisms but stop their growth, making their population static.  Selecting an antimicrobial agent Classifications: High-level germicides Kill all pathogens, including endospores Medically invasive equipment Intermediate-level germicides Kill fungal spores, protozoan cysts, viruses, and pathogenic bacteria Membrane surface medical equipment Low-level germicides Kill vegetative bacteria, fungi, protozoa, and some viruses Skin contact only  Selecting an antimicrobial agent Susceptibility of Microorganisms Death rates do vary, although they are generally constant for a particular agent for a single microbe Selecting an antimicrobial agent  Selecting an antimicrobial agent Waxy lipids in cell walls – protects against water-based chemicals and prevents desiccation  Selecting an antimicrobial agent Similar to endospores; helps allow survival during harsh conditions Cysts also aid in transportation – some human parasites have charged cell walls that do not allow attachment to intestines  Measuring Microbial Control Several factors contribute to the effectiveness of a disinfecting agent or microbial control protocol. The length of time of exposure - Longer exposure times kill more microbes. It takes longer to kill a high-population load than a low-population load Susceptibility of the agent to that disinfecting agent or protocol. Concentration of disinfecting agent or intensity of exposure. Higher temperatures/higher concentrations kill microbes more quickly and effectively. Environmental conditions pH – may alter chemistry and affect the antimicrobial action Conditions that limit contact between the agent and the targeted cells. Example: the presence of bodily fluids, tissue, organic debris (e.g., mud or feces), or biofilms on surfaces—increase the cleaning time or intensity of the microbial control protocol required to reach the desired level of cleanliness. May inactivate certain antimicrobials Measuring Microbial Control Decimal reduction time (DRT) Time required to kill 90% of microbes in a sample This is the D-value  Action of Antimicrobial Agents Alteration of Cell Wall Alteration of Membrane  Action of Antimicrobial Agents Damage to Proteins Damage to Nucleic Acids  Physical Methods of Microbial Control 1. Heath-related methods a) Moist heat b) Dry heat 2. Refrigeration and Freezing 3. Desiccation and Lyophilization 4. Filtration 5. Osmotic Pressure 6. Radiation a) Ionizing b) Non-ionizing  Heat-Related Methods Commonly used Denatures proteins, disrupts cell membranes and cell walls, disrupts structure of nucleic acids Thermal Death Point – lowest temperature that kills all cells in a broth in 10 minutes Decimal reduction time – Time required to kill 90% of microbes in a sample Thermal death time – Time to completely sterilize a set volume at a set temperature  Heat-Related Methods Moist heat – disinfect, sanitize, sterilize, and pasteurize Water conducts heat better than dry air Example: Hand in hot oven at 350°F Hand in boiling water 212°F  Heat-Related Methods - Boiling Kills cells of bacteria and fungi, trophozoites of protozoans, most viruses within 10 minutes At normal atmospheric pressure, boiling cannot exceed 100°C Slow vs fast boil does not have an impact Certain organisms (endospores, cysts, some viruses) can survive boiling  Heat-Related Methods - Autoclaving Uses pressure to achieve boiling temperatures above 100°C Optimal temperature of 121°C for 15 min destroys all microbes Steam must come in contact with all surfaces and liquids Autoclave tape – changes color when sterilization has occurred  Heat-Related Methods - Pasteurization Used for food products such as milk, ice cream, fruit juices Kills harmful bacteria such as E. coli NOT STERILIZATION – heat-loving microbes can survive, but do not spoil for the time that the are generally stored and are not generally pathogenic  Heat-Related Methods – Dry Heat Used for products that cannot get wet – powders, oils, some metals Denatures proteins and oxidizes metabolic and structural chemicals Higher temperatures for longer time than moist heat Incineration – ultimate means of sterilization Autoclave: 121°C for 15 min = Dry Heat: 171°C for 1 hour  Other Physical Methods – Refrigeration and Freezing Refrigeration (0-7°C) and Freezing (below 0°C) Decrease microbial metabolism, growth, and reproduction Chemical reactions occur slower at low temperatures Liquid water not available Refrigeration halts growth of most pathogens Some microbes can multiply in refrigerated foods Slow freezing more effective than quick freezing – forms ice crystals that puncture cell membranes Organisms vary in susceptibility to freezing Some can be stored at -30 to -80°C and can be reconstituted in certain media  Other Physical Methods - Desiccation and Lyphilization Desiccation (drying) inhibits growth due to removal of water Lyophilization (freeze-drying) used for long-term preservation of microbial cultures Instantly frozen in liquid nitrogen or carbon dioxide and any water is removed through sublimation (going directly from liquid to gas) Prevents formation of damaging ice crystals  Other Physical Methods - Dessication Osmotic Pressure Use of salt or sugar to remove water, inhibiting bacterial growth Used for a variety of foods: jellies, jams, pickles, salted fish, jerky  Other Physical Methods - Filtration Passage of either a liquid or gas through a sieve Typically filters made of porcelain, glass, cotton, etc. are used A high suction vacuum pulls the solution through the filter and particles are trapped, leaving a filtrate that is free of microbes and/or viruses (dependent on the pore size of the filter)  Other Physical Methods - Filtration HEPA air filters High efficiency particulate air filters Used in biological safety cabinets Use in air ducts of certain hospital areas Can be combined with chemical filtration and UV Can be purchased for homes Allergies  Other Physical Methods - Radiation Ionizing Radiation Wavelengths shorter than 1 nm Electron beams, gamma ray, some X rays X-rays do not penetrate metals well, thus the use of lead smocks during imaging procedures Ejects electrons from atoms to create ions Ions disrupt hydrogen bonding, oxidize double covalent bonds, and create hydroxyl radicals Ions denature other molecules (DNA) – mutations lead to cell death Some practical uses: sterilization of packaged lab items (Petri dishes, loops), delicate items sensitive to heat (pharmaceutical drugs, spices)  Other Physical Methods - Radiation Nonionizing Radiation Wavelengths greater than 1 nm Excites electrons, causing them to make new covalent bonds Affects 3-D structure of proteins and nucleic acids UV light causes thymine dimers in DNA UV light does not penetrate well (i.e. does not pass well through glass, plastic, etc.) so cells must be directly exposed Suitable for disinfecting air, transparent fluids, and surfaces of objects  Chemical Methods of Microbial Control Wide range of actions Disruption of cell walls, cell membranes, proteins, DNA Impacted by environmental factors pH, temperature, freshness of chemical Impact affected by length of exposure and concentration of microbes  Method Action Level of Uses Examples Activity Phenol and phenolics Denature Intermediate to Low Healthcare, Lister introduced during proteins/disrupt labs, homes surgery; household cell membrane cleaners Alcohols Denature Intermediate Healthcare, Swabbing before a blood proteins/disrupt labs, homes draw cell membrane Halogens Denature proteins Intermediate Healthcare, Iodine for surgery prep; labs, industry, Chlorine in swimming homes pools; Chloramine in water treatment Oxidizing Agents Denature proteins High Healthcare, Hydrogen peroxide on by oxidation industry, food surfaces; Ozone for water processing, treatment homes Surfactants Decrease surface Low Healthcare, Using soap to wash hands’ tension of labs, industry , Quaternary ammonia water/disrupt cell homes compounds in mouthwash membranes  Method Action Level of Activity Uses Examples Heavy Metals Denature proteins Low Healthcare, Not heavily used; Copper to labs, homes inhibit algae growth Aldehydes Denature proteins High Healthcare, Glutaraldehyde used for labs disinfection and sterilization in hospitals; Formalin used for embalming (must use caution) Gaseous Agents Denature proteins High Healthcare, For items that cannot be labs exposed to heat or chemicals; dangerous to work with Enzymes Denature proteins High (substrate Healthcare, Eliminate prions, such as specific) industry that cause mad-cow disease; used to reduce bacteria in food and wine Antimicrobials Wide range of Intermediate to Low Healthcare, Antibiotics; reduce bacteria actions labs, in cheese industry  Selecting an antimicrobial agent Ideally, agents for the control of microbes in the environment should be Readily available Inexpensive Fast-acting/easy to use Stable during storage/chemically stable Capable of controlling microbial growth while being harmless to humans, animals, and objects Non-toxic, non-allergenic Not corrosive or otherwise damaging on surfaces