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5345 Yazan Al-mansour Microbial growth Hani Masaadeh Second 1 1-11-2015 0 Our lecture today is about controlling the growth of microorganisms The objective of this lecture: several factors that affect the growth of a microorganism (Temperature, moisture, ph..) To control the growth of microorganisms, we have In vivo and in vitro In vivo: when we apply antibiotics. In vitro: when we use chemical and physical methods to control the microorganism. The types of microorganisms: we classified microorganisms according to the 1-Tempreture 2-condition, which is called the optimal condition of some microorganisms (psychrophilic, mesophilic, thermophilic, halophilic, haloduric, alkaliphilic, acidophilic, and piezophilic). The importance of using “asptic techniques” in microbiology laboratory and hospitals is to avoid the contamination. Draw a bacterial growth curve and label its four phases. Differentiate between sterilization, disinfection, and sanitization. Sterilization: is a term referring to any process which kills all types of microorganisms. We also call it “Bactericidal” which describes the chemical agent which kills the microorganism. Disinfection: antimicrobial agents that are applied to nonliving objects to destroy microorganisms (reduce the number of microorganisms). We also call it “Bacteriostatic” which describes the chemical agent which inhibits growth of microorganisms. Sanitization: reduction of the microbial population to a safe level as determined by the public health standards. 1 The processes of pasteurization and lyophilization: Pasteurization: sterilization of milk, occurs at temperature 6 to3 centigrade for 30 mins. Lyophilization: when we make the bacterial growth from liquid or broth phase to powder phase, so we can keep it up to 5 years. This is for the purpose of (research papers, demonstration for students). Several physical methods used to inhibit the growth of microorganisms (heating). Differentiate between bactericidal and bacteriostatic agents. Bactericidal: kills all the forms of microorganism. Bacteriostatic: inhibits the growth or the multiplication of microorganisms. -The microbial growth: is the increase in the number of cells, not cell size. By binary fission (the duplication of bacterial cells as 1 cell becomes 2 and these 2 become 4 and so on). - We have some types of slow microorganisms such as mycobacterium tuberculosis, which means that the multiplication will take a long time (18-24h). Not as the common microorganism E.coli, salmonella which takes (15mins) to grow. - At some locations, we need to increase the growth of microorganisms, such as in the microbiology lab. In other locations we need to inhibit or kill the microorganism, such as in hospitals or operation rooms to get a sterile area. - Colonies ( ) مستعمرات جرثومية: groups of cells large enough to be seen without a microscope. Each colony contains more than one million bacterial cell, or a population of billions of cells. - The colonies are different from one microorganism to another. We can differentiate between them by the differential and selective media. 2 Now we have factors which are important for microbial growth. - Temperature: (mesophiles, thermophiles, secrophiles..) the classification here is according to the optimal temperature for one type of microorganism. - Moisture: if we want to prepare a media for moisture, it is done by sterile distilled water. - Osmotic pressure - PH (acidic, alkaline, neutral) - Barometric pressure - Gases (oxygen aerobic and nonaerobic bacteria) - Radiation: plays a high role in sterilization. - Chemicals -Presence of neighboring microbes. The requirements for microbial growth can be divided into two main categories: physical and chemical. - Physical requirements include temperature, pH and Osmotic pressure. - Chemical requirements include sources of carbon, nitrogen and sulfur. Now let's talk about it in details.. Physical requirements: A) Temperature - The minimum growth temperature is the lowest temperature at which the species will grow. - The optimum growth temperature is the temperature at which the species grows best. -The maximum growth temperature is the highest temperature at which Growth is possible. The bacteria can grow at maximum and minimum temperature, but the multiplication of the bacteria won’t be very 3 fast. For example, “E.coli” prefers the temperature of a human body which is around 36-37 and it’s the optimum temperature of it. In the previous figure you can see the type of bacteria which grows at different temperatures .. - Psychrophiles: refers to cold conditions (-10 _ 20). - Thermophiles: refers to Nonpathogenic bacteria, because it needs higher temperature than the body temperature (40 _70). - Mesophiles: refers to pathogenic bacteria which grows best in moderate temperature, neither too hot or too cold, like the body’s temperature (10 _50) . B) PH. - most bacteria prefers neutral pH, between 6.5 and 7.5 - Acidophiles grow in acidic conditions: One microorganism which is highly pathogenic and grows in acidic conditions is called Microbacterium tuberculosis or any acid fast bacteria. Molds and yeasts grow over a greater range of pH than bacteria, but the optimum pH is between 5 and 6 (acidic 4 environments). - Alkalinity also inhibits microbial growth but is rarely used to preserve foods. Example for alkaline: bacterium Vibrio-cholerea (pH 8.8). c) Osmotic pressure - Microorganisms obtain almost all of their nutrients in solution from the surrounding water (humid environment). - They require water for growth and 80 to 90% of them is water. - Extreme or obligate halophiles are organisms that have adapted so well to high salt concentrations that they need it for their growth (in the Dead Sea). - Facultative halophiles do not require high salt concentrations but are able to grow at salt concentrations of 2 to 15%. - Most microorganisms must be grown in a medium that is nearly all water. Chemical Requirements for the bacterial growth: When we see the composition of the media, we will find some of those that are present. Carbon Nitrogen Sulfur Phosphorus Potassium, magnesium and calcium Oxygen - Oxygen a very important factor because some bacteria can only grow in the presence of oxygen (Aerobic) and some can only grow in the absence of oxygen (Anaerobic) and others can grow in both. 5 Grows in the presence of O2, notice bacteria grows just on the surface Grows in presence or absence of O2 “ like E.coli “ Grows in the absence of O2 “clostridium perfringens “ Grows better in the absence of O2, but Resists the presence of O2 Anaerobic but can Grow in the presence of a small amount of O2 Chemical Requirements: Organic growth factors. Essential organic compounds that an organism can’t synthesize must be obtained from the environment. Vitamins, amino acids, purines and pyrimidines Most bacteria can synthesize all of their own vitamins and are not dependent on outside sources. Culture Media: is a liquid or a gel designed to support the growth of microorganisms. There are different types of media for growing different types of cells. 6 -About agar, on a semi solid media there is an agar but a little amount of it. - On a liquid media, there is no agar at all. Give nutrients Give nutrients inhibit the grow of bacteria give solidification for media for humidity fbffffrfvvfffff fffffff vvvvvv -E.coli needs carbohydrates to grow, such as glucose and lactose, carbohydrates give e.coli pigmentation (color).nj On lactose fermenter, gives a red or a pink colony. On glucose fermenter, gives a colorless colony. 7 Anaerobic Culture Methods )(طرق زراعة البكتيريا الالهوائيه -Because anaerobic bacteria might be killed due to the exposure to oxygen, a special media called the reducing media must be used. - Reducing media Contains chemicals (sodium thioglycolate) that combine O2 and deplete the oxygen in the culture medium. (The oxygen found on the surface of thioglycolate media. liquid media) (بتمنع وصول .)االكسجين للبكتيريا -Special anaerobic jars are used to grow anaerobes on Petri plates to observe individual colonies. The mechanism is that the Culture plates are placed in the jar, and oxygen is removed by the addition of a packet of chemicals which react with oxygen to form water. ”jars” .زجاجه خاصة عشان تنمو البكتيريا الالهوائيه فيها -Use of transparent anaerobic champers, equipped with air locks and filled with inert gases. Capnophiles Require High CO2: Microbes that grow better at high CO2 concentrations are called capnophiles (capno: smoke (CO2), philes: loving). Examples: (Nesseria Meningitides, Nesseria gonorrhea) CO2 culture methods: a) Candle jar, Cultures are placed in a sealed jar containing a lighted candle, which consumes O2 and produces CO2. What happened here is as following; There is a candle with flame, so when the jar was closed the flamed candle consumed O2 and produced CO2, so with the presence of CO2 and Solid media (Petri plate), after 24 hours we will have Capnophiles bacteria present on the bottom of the jar, we can also use liquid Media as in tubes. ) (أالحتراق يولد ثاني أكسيد الكربون 8 b) CO2-packet The packet consists of a bag containing a Petri plate and CO2 generator. ) (مضخة خاصه لثاني أكسيد الكربون 9 media types : We have different types of media used in the growth of microorganisms: 1-selective 2- Differential 3-Enrichment Selective Media, Selective and differential media are used to detect the presence of specific microorganisms associated with disease or poor sanitation. - Selective media are designed to suppress the growth of unwanted microbes and encourage growth of the desired microbes. (Just one type of microorganism will be suppressed in this media). ) (بتحط مواد بتمنع نمو أي بكتيريا غير الي بدها ياها Example: bismuth sulfite agar is used to isolate the typhoid bacterium (Salmonella typhi) from feces. Example: Sabourraud’s dextrose agar (pH 5.6), is used to isolate fungi that outgrows most bacteria at this pH. Differential Media, in this media we use an indicator to differentiate between the pigmentation of two microorganisms. - Make it easy to distinguish colonies of the desired organism from other colonies growing on the same plate. Example, 10 MacConkey agar: is a culture medium designed to selectively grow Gram-negative bacteria and differentiate them by lactose fermentation. *MacConkey agar color: -The lactose fermenter bacteria give: pink colonies (e.coli). -Non-lactose fermenter bacteria give: colorless colonies (Salmonella). at the end , forgive me for any mistake . . “We cannot direct the wind but we can adjust the sails.” this is a new beginning for us in second exams we can do it . . . Edited by: Salam Mustafa 11