Microbiology Lecture Notes PDF

Microbiology Sterilization and Disinfection. Dr.Sajjad Mohsin Sterilization generally refers to a process that eliminates or destroys all forms of microbial life, including bacteria, viruses, and spores. It can be applied in various contexts, in...

Microbiology Sterilization and Disinfection. Dr.Sajjad Mohsin Sterilization generally refers to a process that eliminates or destroys all forms of microbial life, including bacteria, viruses, and spores. It can be applied in various contexts, including: 1-Medical Settings: Sterilizing surgical instruments and equipment to prevent infections. 2-Food Industry: Ensuring food products are free of harmful microorganisms to extend shelf life and ensure safety. 3-Laboratories: Preparing sterile environments for experiments to prevent contamination. Disinfection: is the process of reducing or eliminating harmful microorganisms from surfaces, objects, or environments, but it does not necessarily kill all pathogens, especially resistant bacterial spores. Antiseptics: are substances used to prevent infection by inhibiting the growth of microorganisms on living tissues, such as skin or mucous membranes. Unlike disinfectants, which are used on inanimate surfaces, antiseptics are safe for application to the body. Common Types:  Alcohols: Isopropyl alcohol and ethanol are widely used for skin disinfection before procedures.  Iodine Compounds: Betadine (povidone-iodine) is used for surgical scrubs and wound cleaning.  Chlorhexidine: Often used in hand sanitizers and preoperative skin preparations.  Hydrogen Peroxide: Used for cleaning wounds, though it can be irritating to tissues.  Silver Sulfadiazine: Commonly applied to burns to prevent infection.  Microbiology Mechanism of Action: Antiseptics work by disrupting the cell membranes of microorganisms, denaturing proteins, or interfering with metabolic processes, leading to microbial death or inhibition. Considerations:  Concentration: The effectiveness of an antiseptic often depends on its concentration and the type of microorganism.  Skin Sensitivity: Some antiseptics can cause irritation or allergic reactions, so it's important to choose appropriate products for different skin types and conditions. Bacteriostatic: Agent that prevents or stops the growth of bacteria Bactericidal: Agent that kills bacteria Advantage of Sterilization: 1. Prevent transmission of disease. 2. Prevent contamination and growth of undesirable bacteria. 3. Prevent spoilage of material by microorganism. Methods of Sterilization: There are Two methods A. Chemical methods B. Physical methods A. Chemical methods: These methods rely on chemicals to eliminate undesired microorganisms. Chemical agents act primarily by one of the three mechanisms: 1. Disrupt the lipid structure of the bacterial cell membrane. EXAMPLE: 70% Alcohol, and Detergents (bipolar chemicals) 2. Modify the proteins of the bacteria. EXAMPLE: 70% Alcohol, chlorine (oxidizing agent), iodine, 2% tincture iodine, potassium iodide 3. Modify the bacterial DNA.  Microbiology common chemical sterilization methods: a. 70% Alcohol: Ethanol requires water to mediate its action. Therefore, 70% alcohol is widely used to clean the skin before some surgical operations. It acts mainly by disorganizing the lipid structure of the bacterial cell membrane. Also, 70% alcohol denatures protein structure of the bacteria as well. b. Hydrogen Peroxide (H2O2): This is a potent antiseptic that is used to clean wounds. H2O2 mode of action is by inhibiting bacterial enzymatic activity. However, its effectiveness is limited by the ability of some organisms to produce catalase. Catalase is an enzyme that degrades H2O2. c. Formaldehyde: is available as a 37% solution in water (formalin). Formalin mode of action is by denaturing proteins and nucleic acids. d. Ethylene Oxide: Ethylene oxide gas is used extensively in hospitals for the sterilization of heat sensitive materials such as surgical instruments and plastics. Ethylene Oxide kills by disruption of both proteins and nucleic acids. e. Acids & Alkalis: Strong acids and alkalis kill by denaturing proteins. Although most bacteria are susceptible, it is important to note that Mycobacterium tuberculosis and other mycobacteria are relatively resistant to 2% NaOH, which is used in the clinical laboratory to liquefy sputum prior to culturing the organism. Weak acids, such as benzoic, propionic, and citric acids, are frequently used as food preservatives because they are bacteriostatic. f. Crystal violet (gentian violet): which is used as a skin antiseptic. Its action is based on binding of the positively charged dye molecule to the negatively charged phosphate groups of the nucleic acids. g. Malachite green: a triphenylamine dye like crystal violet, is a component of Löwenstein-Jensen’s medium, which is used to grow M. tuberculosis. The dye inhibits the growth of unwanted organisms in the sputum during the 6-week incubation period.  Microbiology B. Physical Method The physical agents act either by heat, radiation or by removing organisms through filtration. a. HEAT: Heat energy can be applied in three ways: 1. Moist heat (either boiling or autoclaving) 2. Dry heat 3. Pasteurization. a. Heat: Heat kills by denaturing proteins and enzymatic cleavage of DNA. Moist heat sterilizes at a lower temperature than Dry heat, because water aids in the disruption of noncovalent bonds (e.g., hydrogen bonds). ➔ Sterilization by Moist Heat (autoclaving), is the most frequently used method of sterilization. As stated above, bacterial spores are resistant to boiling (100°C), they must be exposed to a higher temperature. This cannot be achieved (water boils at 100°C at atmospheric pressure). Therefore, the pressure is increased synthetically. For this purpose, an autoclave chamber is used in which steam, at a pressure of 15 lb/in2, reaches a temperature of 121°C and is held at that temperature for 15 to 20 minutes. This kills even the highly heat-resistant spores of Clostridium botulinum. ➔ Sterilization by Dry Heat, on the other hand, requires temperatures in the range of 180°C for 2 hours. This process is used primarily for glassware. ➔ Pasteurization: is used primarily for milk, consists of heating the milk to 62°C for 30 minutes followed by rapid cooling. (“Flash” pasteurization at 72°C for 15 seconds is often used.) This is sufficient to kill the vegetative cells of the milk borne pathogens (e.g., Mycobacterium bovis, Salmonella, Streptococcus, Listeria, and Brucella). b.RADIATION: The two types of radiation used to kill microorganisms are ultraviolet (UV) light and X-rays. Ultraviolet Light: The greatest antimicrobial activity of UV light occurs at 250 to 260 nm, which is the wavelength region of maximum absorption by the purine and pyrimidine bases of DNA. X-rays have higher energy and penetrating power than UV radiation and kill mainly by the production of free radicals such as hydroxyl radicals. X- rays are used in medicine for sterilization of heat-sensitive items, such as sutures and surgical gloves, and plastic items, such as syringes.  Microbiology c. FILTRATION: is the preferred method of sterilizing certain solutions (e.g., those with heat-sensitive components). In the past, solutions for intravenous use were autoclaved, but heat-resistant endotoxin in the cell walls of the dead gram-negative bacteria caused fever in recipients of the solutions.