Medical Microbiology Lab 1 - University of Fallujah PDF

Medical Microbiology Laboratory Culture media, Sterilization & Disinfections University Of Fallujah College Of Medicine Lecture : Lab. 1. Stage : 3 Lecturer : Fairooz Bahr Sultan Department: Microbiology Date: 24/9/2024, Tuesday  Medical Microbiology...

Medical Microbiology Laboratory Culture media, Sterilization & Disinfections University Of Fallujah College Of Medicine Lecture : Lab. 1. Stage : 3 Lecturer : Fairooz Bahr Sultan Department: Microbiology Date: 24/9/2024, Tuesday  Medical Microbiology Laboratory Lab 1 Culture media, Sterilization & Disinfections Medical Microbiology is a branch of medical science concerned with the prevention, B diagnosis and treatment of infectious diseases. Bacteria /Fungus /Parasites /Viruses  Most can only be seen with the microscope.  Microbiologists determine the type of microorganism causing the disease and find a drug, usually an antibiotic, to inhibit the microorganism. Microbiologists continue to study the microorganisms through research to determine new antibiotics. Pure Culture The term "Pure Culture" refers to a population or colony of cells growing B of other species. in the absence A pure culture is usually derived from a mixed culture (one containing many species) by transferring a small sample into new, sterile growth. Importance of "Pure Culture allows for the creation of a pure culture with only one type of microorganism. allow for microbiologists to study of one species without the worry of contamination from other organisms. Medium Is a mixed nutrient used to support microbial growth. B Colony: macroscopically visible collection of millions of bacteria originating from a single bacterial cell. Culture media Requirements ( in media ) for bacterial growth: Temperature Nutrients B(H2O, C-source, N-source (Peptone provides the organism with nitrogen, amino acids, and other elements essential for growth), Inorganic salts (sodium chloride maintains the osmotic balance), Growth factors). pH Types of culture media /Based on 1. Physical state: a) Solid medium b) Liquid medium c) Semi solid medium 2. purpose/ functional use. 3. Oxygen requirement.  solid medium liquid medium semi solid medium contains 2% agar no agar 0.5% agar for the isolation and for the observation of B identification of bacteria, for the proliferation bacterial motility and Colony morphology, of bacteria preservation of bacteria pigmentation, hemolysis Eg: Nutrient agar, Eg: Nutrient broth Eg: Motility medium Blood agar  Agar Agar: (simply called agar) is the most commonly used solidifying agent. Agar is composed of polysaccharides. It melts at 95°C and solidifies at 42oC. B 2. Based on the basis of purpose/ functional use A. General purpose media (Peptone water, nutrient broth and nutrient agar) all so called (basal media). B. Enriched medium (Added growth factors) ( Blood agar, Chocolate agar). added with additional nutrients such as blood, serum or egg to the basal medium. Prepared by adding 5–10% of sheep blood to the molten nutrient agar at 45°C. It is used B hemolytic property of the bacteria. to test the Used to grow bacteria that are exacting in their nutritional needs (fastidious microorganisms ). Blood agar Used to differentiate bacteria based on their hemolytic B traits (β-hemolysis, α-hemolysis and γ- hemolysis). β-hemolysis (complete lysis of RBC surrounding the colony). γ-hemolysis (discoloration in the medium). α-hemolysis (Show “green” hemolysis caused by hydrogen peroxide produced by the bacterium, oxidizing hemoglobin to green methemoglobin). Chocolate agar Used to Isolation and identification of B microorganisms. fastidious Modified blood agar which is heated to lyse the RBC. Growth factors release by lysis of RBC which are:  X factor / hemin.  V factor / coenzyme nicotinamide adenine dinucleotide (NAD). C. Differential Media Differential media: are media that distinguish between different groups of bacteria and identification of microorganisms based on their biological characteristics (Blood agar, Mannitol B salt agar and MaConkey agar) D. Selective medium Selective media: favor the growth of particular microorganisms. Bile salts or dyes like basic fuchsin and crystal violet favor the growth of gram-negative bacteria by inhibiting the growth of gram-positive bacteria without affecting gram-negative organisms. E. Transport media Transport Media used for transporting the samples. media 3. Based on Oxygen requirement a. Aerobic media MacConkey agar These mediaB are used to grow aerobic organisms. b. Anaerobic media (Robertson’s cooked meat medium, Thioglycolate medium) These media are used to grow anaerobic organisms. Morphology of Bacteria Bacteria have characteristic shapes. The common microscopic morphologies are: 1- Spherical or ovoid (cocci) 2- Rod-shaped (bacilli) 3- Comma-shaped (vibrio) 4- Spiral (spirillum and spirochete)  Arrangements of cocci Diplococci Streptococci B Tetrads Sarcinae Staphylococci Arrangements of Bacilli Diplobacilli Streptobacilli Coccobacilli Spiral Bacteria Vibrios Spirilla Spirochetes Colony Morphology Bacteria grow on solid media as colonies. A colony is defined as a visible mass of microorganisms B all originating from a single mother cell, therefore a colony constitutes a clone of bacteria all genetically alike. These four forms represent the most common colony shapes you are likely to encounter. Circular. Filamentous Rhizoid. Irregular  B Sterilization & Disinfections  Why We Need Sterilization Microorganisms B capable of causing infection are constantly present in the external environment and on the human body. Microorganisms are responsible for contamination and infection. Remove and destroy Microorganisms from materials and surfaces. Sterilization: complete killing of all forms of microorganisms, including bacterial spores. B Disinfection: killing or removing of harmful vegetative microorganisms. Pasteurization: Used heat at temperatures sufficient to inactivate harmful organism in milk. Disinfectant: chemical substance used to achieve disinfection. Antiseptic: disinfectant that can be safely used on living tissues. Methods of Sterilization 1. Physical methods: B  Heat ( dry heat & moist heat)  U.V. Light  Ionizing Radiation  Filtration 2. Chemical method: (used for heat sensitive equipments)  Ethylene Oxide  Gluteraldehyde Heat A. Dry heat: B Flaming (Burner- for sterilizing bacteriological loops, knives, blades.). Hot air oven expose items to 160 °C for 1 hour, it used for items that are lacking water such as:  Metals.  Glassware.  Oils/ Waxes /Powder. Heat B. Moist heat by using: B Autoclave Uses hot water. High pressure. Holding period 121 Cº for 15 min. Kill all forms of life. Sterilize culture media. U.V. Light Has limited sterilizing power because of poor penetration into most B materials. Generally used in irradiation of air in certain areas such as operating rooms and tuberculosis labs. Ionizing Radiation e.g. Gamma radiation: has greater energy than U.V. light, therefore more effective. Used mainly in sterilization of disposable plastic syringes, gloves, specimens containers and Petri dishes. Filtration Used for heat sensitive substances e.g. sera, antibiotics. B B