Chapter 8. –Controlling Microbial Growth in Vitro Chapter ...

1 Chapter 8. Controlling Microbial Growth in Vitro Osmotic pressure is the pressure that is Chapter 8 Outline exerted on a cell membrane by solutions Introduction both inside and outside the cell. factors that Affect Microbial Growth Osmosis is the movement of a solvent, Encouraging the Growth of Microbes in Vitro through a permeable membrane, from a Inhibiting the Growth of Microbes in Vitro lower concentration of solutes (dissolved factors That Affect Microbial Growth substances) to a higher concentration of solutes. Availability of Nutrients When the concentration of solutes in the All living organisms require nutrients to external environment of a cell is greater sustain life. than that of solutes inside the cell, the Nutrients are energy sources. Organisms solution in which the cell is suspended is obtain energy by breaking chemical bonds.

2 Said to be hypertonic. Moisture factors That Affect Microbial Growth , cont. Water is essential for life. It is needed to Osmotic Pressure and Salinity, cont. carry out normal metabolic processes. Plasmolysis is a condition in which the cell Certain Microbial stages ( , bacterial membrane and cytoplasm of a cell shrink endospores and protozoal cysts) can away from the cell wall; occurs when survive a drying process (dessication). bacteria with rigid cell walls are placed into factors That Affect Microbial Growth , cont. a hypertonic solution. Temperature When the concentration of solutes outside Every organism has an optimum Growth a cell is less than that of solutes inside a temperature. cell, the solution in which the cell is The temperature (and pH) ranges over suspended is said to be hypotonic.

3 Which an organism grows best are largely If a bacterial cell is placed into a hypotonic determined by its enzymes. solution, it may not burst (because of the Thermophiles are microorganisms that rigid cell wall. grow best at high temperatures. factors That Affect Microbial Growth , cont. Mesophiles are microbes that grow best at Osmotic Pressure and Salinity, cont. moderate temperatures ( , 37o C). A solution is said to be isotonic when the factors That Affect Microbial Growth , cont. concentration of solutes outside a cell Temperature, cont. equals the concentration of solutes inside Psychrophiles prefer cold temperatures the cell. (like deep ocean water). Organisms that prefer to live in salty Psychrotrophs, a particular group environments are called halophilic of psychrophiles, prefer organisms.)

4 Refrigerator temperature (4oC). Barometric Pressure Psychroduric organisms prefer warm Microbes that can survive in high temperatures, but can endure very cold or atmospheric pressure (> psi) are know even freezing temperatures. as piezophiles. factors That Affect Microbial Growth , cont. Changes in Osmotic Pressure pH Encouraging the Growth of Microbes in Vitro pH refers to the acidity or alkalinity of a Culturing Bacteria in the Laboratory solution. Bacterial Growth Most microorganisms prefer a neutral or Think of bacterial Growth as an increase in the slightly alkaline Growth medium (pH - number of organisms rather than an increase in ) their size. Acidophiles prefer a pH of 2 to 5 Bacteria divide by binary fission (one cell divides to become two cells). Alkaliphiles prefer a pH > factors That Affect Microbial Growth , cont.

5 Binary fission continues until a colony is produced. Osmotic Pressure and Salinity Binary fission continues for as long as there is a sufficient supply of nutrients, water, and space. The time it takes for one cell to become two cells is Culturing Bacteria in the Laboratory called the generation time ( , E. coli = 20 Bacterial Population Growth Curve minutes). A population Growth curve for any particular Culturing Bacteria in the Laboratory species of bacterium may be determined by Culture Media growing a pure culture of the organism in a liquid Media (sing., medium) are used in microbiology medium at a constant temperature. labs to culture ( , grow) bacteria; media Samples of the culture are collected at prepared in the lab are referred to as artificial fixed intervals to determine the number of media or synthetic media.

6 Viable organisms. A chemically defined medium is one in which all A graph is prepared by plotting the ingredients are known. logarithmic number of viable organisms Culture media can be liquid or solid. (on the vertical or Y- axis) against the An enriched medium is a broth or solid containing a incubation time (on the horizontal or X- rich supply of special nutrients that promote the axis). Growth of fastidious organisms; example = Inhibiting the Growth of Microbes in Vitro chocolate agar. Definition of Terms A selective medium has added inhibitors that Sterilization is the complete destruction of all discourage Growth of certain organisms while microbes, including cells, spores, and viruses. allowing the Growth of a desired organism; Accomplished by dry heat, autoclaving example = PEA agar. (steam under pressure), gas, various Differential Medium chemicals, and certain types of radiation.

7 Culturing Bacteria in the Laboratory Disinfection is the destruction or removal of Inoculation of Culture Media pathogens from nonliving objects by physical or Culture media are inoculated with clinical chemical methods; pasteurization is an example of specimens ( , specimens collected from patients a disinfection technique. with a suspected infectious disease). Disinfectants are chemical substances that Inoculation involves adding a portion of a eliminate pathogens on inanimate objects. specimen to the medium. Antiseptics are solutions used to disinfect Inoculation is accomplished using a sterile skin and other living tissues. inoculating loop. Definition of Terms, cont. Culturing Bacteria in the Laboratory The suffix cide or cidal refers to killing.. Importance of Using Aseptic Technique Germicidal agents, biocidal agents, and Aseptic technique is practiced when it is necessary microbicidal agents are chemicals that kill to exclude microbes when inoculating culture microbes.

8 Media. Bactericidal agents are chemicals that specifically kill bacteria, but not necessarily bacterial Unwanted organisms are referred to as endospores. contaminants; the Growth medium or plate is said Sporicidal agents kill bacterial endospores. to be contaminated. Fungicidal agents kill fungi, including The sterility of the media must be maintained fungal spores. before inoculation. Algicidal agents kill algae. Avoid touching the surface of the agar! Viricidal agents destroy viruses. Culturing Bacteria in the Laboratory Definition of Terms (cont.). Incubation A microbistatic agent is a drug or chemical that After media are inoculated, they must be placed inhibits Growth and reproduction of microbes. into an incubator which will maintain the A bacteriostatic agent is one that specifically appropriate inhibits the metabolism and reproduction of atmosphere, bacteria.

9 Temperature, and Lyophilization is a process that combines moisture level; dehydration (drying) and freezing. This process is the process is known as incubation. widely used in industry to preserve foods, antibiotics, microorganisms, and other biologic Ultrasonic waves; used in hospitals and medical materials. and dental clinics to clean equipment. Sepsis refers to the presence of pathogens in blood Filters; used to separate cells/microbes from or tissues, whereas asepsis means the absence of liquids or gases. pathogens. Gaseous atmosphere; can be altered to inhibit Antisepsis is the prevention of infection. Growth . Using Physical Methods to Inhibit Microbial Using Chemical Agents to Inhibit Microbial Growth Growth Chemical disinfection refers to the use of chemical Heat agents to inhibit the Growth of pathogens, either 2 factors temperature and time - temporarily or permanently.

10 Determine the effectiveness of heat for Disinfectants are affected by: sterilization. Prior cleaning of the object or surface The thermal death point (TDP) of any The organic load ( , feces, blood, pus). species is the lowest temperature that will The bioburden; types and numbers of kill all of the organisms in a standardized microbes pure culture within a specified time. Concentration of the disinfectant Types of Heat Contact time Dry heat , oven, electrical incinerator, Physical nature of the object being or flame disinfected Moist heat boiling or use of an autoclave Temperature and pH. Dry Heat Sterilization Using Chemical Agents to Inhibit Microbial Growth , Using Physical Methods to Inhibit Microbial cont. Growth , cont. Should have a broad antimicrobial spectrum The autoclave Fast acting A large metal pressure cooker that uses Not affected by the presence of organic matter steam under pressure to completely Nontoxic to human tissues and noncorrosive destroy all Microbial life.