Module 08: Controlling Microbial Growth in Vitro Flashcards
What are the factors that affect microbial growth?
(1) Availability of Nutrients
(2) Moisture
(3) Temperature
(4) pH (acidity and alkalinity)
(5) Osmotic pressure and salinity
(6) Barometric pressure
(7) Gaseous atmosphere
All living organisms require this to sustain life. They are known as energy sources.
Nutrients
How do organisms obtain energy?
Organisms obtain energy by breaking chemical bonds.
Under moisture, this is known as an essential for life. It is needed to carry out normal metabolic processes.
Water
What are certain microbial stages that can survive the drying process (desiccation)?
(1) Bacterial Endospores
(2) Protozoal Cysts
How does temperature affect microbial growth?
Every organism has an optimum growth temperature. The temperature (and pH) ranges over which an organism grows best are largely determined by the optimum temperature and pH ranges of its enzymes.
These are organisms that grow best at high temperatures.
Thermophiles
These are microbes that grow best at moderate temperatures (eg. 37 degrees Celsius, human body temperature).
Mesophiles (found in warm soil and water like plants and animals)
These are microbes that prefer cold temperatures (like deep ocean water).
Psychrophiles
This is a particular group of psychrophiles, prefer refrigerator temperature (4 degrees Celsius) like bread molds.
Psychrotrophs
These prefer warm temperatures but can endure very cold or even freezing temperatures. They can also be preserved in freezing temperatures.
Psychroduric organisms
This refers to the acidity or alkalinity of a solution.
pH
What pH do most microorganisms prefer?
A neutral or slightly alkaline growth medium (pH 7.0 to 7,4)
These type of microorganisms prefer a pH of 2 to 5. They thrive in highly acidic environments, such created by sulfurous gases in hydrothermal vents and hot springs.
Acidophiles (eg fungi)
These type of microorganisms prefer a pH of >8.5.
Alkaliphiles (such as those in the intestine ph 9 and soda lakes along with soils laden with carbonate)
This is the pressure that is exerted on a cell membrane by solutions both inside and outside the cell.
Osmotic pressure
This is the movement of a solvent, through a permeable membrane, from lower concentration of solutes (dissolved substances) to a higher concentration of solutes.
Osmosis
When the concentration of solutions in the external environment is greater than that of the solutes inside the cell, the solution in which the cell is suspended is said to be what?
Hypertonic
This is a condition in which the cell membrane and the cytoplasm of a cell shrink away from the cell wall.
Plasmolysis (this occurs when a bacteria with a rigid cell walls are placed in a hypertonic solution - so this can inhibit microbial growth and multiplication)
When the concentration of solutes outside a cell is less than that of solutes inside a cell, the solution in which the cell is suspended is said to be what?
Hypotonic
If a bacterial cell is placed into a hypotonic solution, it may not burst. If it does burst, the cytoplasm escapes, this process is known as what?
Plasmoptysis
Why doesn’t the bacterial cell burst when placed in a hypotonic solution?
Because of the rigid cell wall (plants and bacteria)
A solution is said to be this when the concentration of the solutes outside a cell is equal to the concentration of solutes inside the cell.
Isotonic (normal turgor)
Organisms that prefer to live in salty environments are called what?
Halophilic organisms (vibrio cholerae)
Organisms that do not prefer to live in salty environments but are capable of surviving there called what?
Haloduric organisms (Staphylococcus aureus)
Microbes that can survive in high atmospheric pressure (<14.7 psi) are known as what?
Piezophiles
These type of microorganisms prefer the same atmosphere that humans do (~20%–21% O2 and 78%–79% N2, and <1% other gases).
Obligate aerobes
These type of microorganisms require reduced concentrations of oxygen (~5% O2).
Microaerophiles
These type of microorganisms are killed by the presence of oxygen.
Obligate anaerobes
These types of microorganisms require increased concentrations of CO2 (5%–10% CO2).
Capnophiles
This is known as the increase of number of organisms rather than the increase in their size.
Bacterial Growth (growth refers to multiplication and proliferation)
Bacteria divide by ____________ (one cell divides to become two cells) when they reach their optimum size.
Binary fission (to produce two daughter cells after DNA replication)
Binary fission will continue in what conditions?
(1) Binary fission continues through many generations until a colony is produced on solid culture medium.
(2) Binary fission continues for as long as there is a sufficient supply of nutrients, water, and space.
The time it take for one cell to become two cells is called what?
Generation time (eg. E coli, Staphylococcus spp., and Streptococcus spp.. = 20 minutes)
These are used in microbiology laboratories to culture or grow bacteria.
Media or medium
Media prepared in laboratories to culture bacteria are referred as what? This is because they do not occur naturally.
Artificial media or synthetic media
This type of medium is one in which all ingredients are known; this is because the medium was prepared in the laboratory by adding a certain number of grams.
Chemically defined medium (culture media can be liquid or solid).
This is a broth or solid containing a rich supply of special nutrients that promote the growth of fastidious organisms, for example, chocolate agar.
Enriched Medium
This has added inhibitors that discourage growth of certain organisms while allowing the growth of a desired organism, for example, PEA agar
Selective Medium
This medium permits the differentiation of organisms that grow on the medium, for example, MacConkey agar.
Differential Medium
What are some various categories of media that are not mutually exclusive?
(1) Blood Agar is enriched
(2) Differential Medium
This medium is a popular liquid medium in bacteriology laboratories; it supports the growth of all categories of bacteria from obligate aerobes to obligate anaerobes.
Thioglycollate (THIO) broth
Explain the nature of Thioglycollate (THIO) broth.
There is a concentration gradient of dissolved oxygen in the tube; organisms grow only in that part of the broth where the oxygen concentration meets their needs.
This is a selective and differential medium. Lactose fermenters are pink; nonlactose fermenters are white.
MacConkey agar
This is a selective and differential medium. S. aureus is able to ferment mannitol changing the color of the medium from red to yellow.
Mannitol Salt Agar
Obligate anaerobes will grow where there is?
0% oxygen
Microaerophiles will grow where there is?
5% oxygen
Obligate aerobes will grow where there is?
20% to 21% oxygen
These are inoculated with clinical specimens (i.e., specimens collected from patients with a suspected infectious disease).
Culture Media
This involves adding a portion of a specimen to the medium. This is accomplished using a sterile inoculating loop.
Inoculation
This is an approached practiced when it is necessary to exclude microbes from a particular area (e.g., when inoculating culture media).
Aseptic Techniques
Unwanted organisms are referred to as ___________; the growth medium or plate is said to be contaminated.
Contaminants
What should be done before inoculation?
(1) The sterility of the media must be maintained before inoculation.
(2) Avoid touching the surface of the agar.
This minimizes contamination and protects the laboratorian.
Inoculating media within a biologic safety cabinet
What happens when the media is inoculated?
After media are inoculated, they must be placed into an incubator which will maintain the appropriate atmosphere, temperature, and moisture level; the process is known as incubation.
This is the process by which the media will be placed into a incubator which will maintain the appropriate atmosphere, temperature, and moisture level.
Incubation
What are the three (3) types of incubators?
(1)A CO2 incubator (contains 5%–10% CO2)
(2) A non-CO2 incubator (contains room air)
(3) An anaerobic incubator (the atmosphere is devoid of oxygen)
This incubator contains 5%–10% CO2.
CO2 incubator (It is important to keep in mind that a CO2 incubator contains oxygen (about 15%–20%) in addition to CO2. Thus, a CO2 incubator is not an anaerobic incubator.)
This incubator contains room air; thus it contains about 20% to 21% of oxygen.
A non-CO2 incubator
This incubator contains an atmosphere that is devoid of oxygen.
An anaerobic incubator
How do you determine the bacterial population counts?
Microbiologists sometimes need to know how many bacteria are present in a particular liquid at a given time (e.g., to determine bacterial contamination of drinking water).
One can determine either the total number of bacterial cells or the number of viable (living) cells.
This can be used to determine growth by measuring the turbidity of the medium.
Spectrophotometer (As bacteria increase in number, the liquid
becomes turbid (cloudy) and less light passes through.)
This is used to determine the number of viable bacteria in a liquid sample by making serial dilutions of the liquid and inoculating onto nutrient agar; after overnight incubation, the number of colonies is counted.
Viable plate count
This for any particular species of bacterium may be determined by growing a pure culture of the organism in a liquid medium at a constant temperature.
Population growth curve
How do you make a population growth curve?
(1) Samples of the culture are collected at fixed intervals to determine the number of viable organisms.
(2) A graph is prepared by plotting the logarithmic number of viable organisms (on the vertical or y- axis) against the incubation time (on the horizontal or x-axis).
These are microbes that can survive and multiply only within living cells.
Obligate intracellular pathogens or also known as host cells
Obligate intracellular pathogens include?
Obligate intracellular pathogens include viruses and two groups of Gram-negative bacteria like rickettsia and chlamydia’s.
How do you culture obligate intracellular pathogens?
Culturing these organisms in the laboratory is a challenge; they must be grown in embryonated chicken eggs, laboratory animals, or cell cultures.
How is fungi cultured?
Fungi (including yeasts, molds, and dimorphic fungi) grow on and in a variety of solid and liquid culture media (there is no single medium that is best for all medically important fungi).
Give an example of culture media for fungi?
(1) Brain heart infusion (BHI) agar, BHI with blood, and
(2) Sabouraud dextrose agar (SDA); SDA is selective for fungi because of its low pH.
What are the media used to recover fungi?
Media to recover fungi often contains antibiotics to inhibit the growth of bacteria that may also be present in the specimen
(Caution must be exercised when culturing fungi—some are highly infectious!)
What are some examples of protozoa that can be cultured as some of them can only be cultured by research and reference laboratories.
(1) Amebae,
(2) Giardia lamblia,
(3) Leishmania spp.,
(4) Toxoplasma gondii,
(5) Trichomonas vaginalis, and
(6) Trypanosoma cruzi.
Due to the severity of diseases that protozoa cause, it is of greatest importance to culture amebae:
(1) Acanthamoeba spp.,
(2) Balamuthia spp., and
(3) Naegleria fowleri.
This is known as the complete destruction of all microbes, including cells, spores, and viruses.
Sterilization
Sterilization is accomplished by what?
Dry heat, autoclaving steam under pressure), ethylene oxide gas, and various liquid chemicals (such as formaldehyde) are the principal sterilizing agents in health care facilities.
This is known as the destruction or removal of pathogens from nonliving objects by physical or chemical methods; pasteurization is an example of a disinfection technique.
Disinfection
These are chemical substances that eliminate pathogens on inanimate objects.
Disinfectants
These are solutions used to disinfect skin and other living tissues.
Antiseptics
The suffix -cide or -cidal refers to what?
Killing
These are chemicals that kill microbes?
(1) Germicidal agents,
(2) Biocidal agents, and
(3) Microbicidal agents
These are chemical agents that specifically kill bacteria, but not necessarily bacterial endospores.
Bactericidal agents
These are chemical agents that kill bacterial endospores.
Sporicidal agents
These are chemical agents that kill fungi, including fungal spores.
Fungicidal agents
These are chemical agents that kill algae.
Algacidal agents
These are chemical agents that destroy viruses.
Viricidal agents
This is a drug or chemical that inhibits growth and reproduction of microbes.
Microbistatic agent
This is one that specifically inhibits the metabolism and reproduction of bacteria.
Bacteriostatic agent
This is a process that combines dehydration (drying) and freezing. This process is widely used in industry to preserve foods, antibiotics, microorganisms, and other biologic materials.
Lyophilization
This refers to the presence of pathogens in blood or tissues, whereas asepsis means the absence of pathogens.
Sepsis
This is known as the prevention of an infection.
Antisepsis
What are the two (2) factors that determine the effectiveness of heat for sterilization?
(1) Temperature
(2) Time
(Also, the higher the temperature, the shorter the time required to kill the organisms)
This of any species is the lowest temperature that will kill all of the organisms in a standardized pure culture within a specified time.
Thermal Death Point (TDP)
What are the two (2) types of heat?
(1) Dry heat (e.g., oven, electrical incinerator, or flame)
(2) Moist heat (boiling or use of an autoclave)
This is a large metal pressure cooker that uses steam under pressure to completely destroy all microbial life.
Autoclave (Increased pressure raises the temperature above the temperature of boiling water (above 100C) and forces steam into materials being sterilized.)
Explain the mechanism of the autoclave.
Autoclaving at a pressure of 15 psi at 121.5 degrees Celsius for 20 minutes destroys vegetative microorganisms, bacterial endospores, and viruses.
These are used to quality control measure to ensure proper autoclaving.
Control measure to ensure proper autoclaving.
Under this approach to inhibit microbial growth, most microorganisms are not killed, but their metabolic activities are slowed.
Cold
Under this approach to inhibit microbial growth, many dried microorganisms remain viable, but they cannot reproduce.
Desiccation
Under this approach to inhibit microbial growth, an ultra-violet (UV) lamp is useful for reducing the number of microbes in the air.
Radiation
Under this approach to inhibit microbial growth, these are used in hospitals and medical and dental clinics to clean equipment.
Ultrasonic waves (160 degrees for 2 hours)
Under this approach to inhibit microbial growth, these are used to separate cells/microbes from liquids or gases.
Filters
Under this approach to inhibit microbial growth, these can be altered to inhibit growth.
Gaseous atmosphere
This refers to the use of chemical agents to inhibit the growth of pathogens, either temporarily or permanently.
Chemical disinfection
Disinfectants are often affected by what?
(1) Prior cleaning of the object or surface
(2) The organic load (e.g., feces, blood, pus)
(3) The bioburden (types and numbers of microbes)
(4) Concentration of the disinfectant
(5) Contact time
(6) Physical nature of the object being disinfected
(7) Temperature and pH
What are the factors that determine the effectiveness of antimicrobial procedures?
(1) Temperature
(2) Time
(3) Concentration
(4) Presence of proteins in feces, blood, pus and vomitus
(5) Type of microbes, number o microbes and the presence of spores.
What are the characteristics of an ideal chemical antimicrobial agent?
(1) Should have a broad antimicrobial spectrum
(2) Fast acting
(3) Not affected by the presence of organic matter
(4) Nontoxic to human tissues and noncorrosive
(5) Should leave a residual antimicrobial film on surface
(6) Soluble in water and easy to apply
(7) Inexpensive and easy to prepare
(8) Stable as both a concentrate and a working solution
(9) Odorless
This may be used safely on human tissues and can reduce the number of organisms on the surface of the skin; do not penetrate pores and hair follicles.
Antiseptics
These are used by healthcare personnel to remove organisms lodged in pores or folds of the skin.
Antiseptic soaps and scrubbing
What are some pathogens that contaminate foods in our kitchens?
(1) Salmonella
(2) Campylobacter
What are some steps that we should do to inhibit the growth of pathogens in our kitchens?
(1) Problems arise when handling foods before cooking.
(2) Remain aware of pathogens when preparing foods.
(3) Wash hands frequently.
(4) Thoroughly clean plates and countertops that have had poultry or meat on them with hot soapy water.
(5) The use of antibacterial kitchen sprays is controversial.
40% of the antibiotics manufactured in the United States are used in what?
Animal feed; microorganisms resistant to these antibiotics survive!
These are transmitted in animal feces and in food products.
Drug-resistant organisms
Where are drug-resistant organisms usually found?
Use of antimicrobial agents is widespread in toys, cutting boards, in hand soaps, and many other household products; resistant microorganisms survive!
How much nutrients is entailed among organisms?
Two dozens of approximately 92-naturally occurring elements are essential for life
How much water do cells have?
70% to 95% of water to carry out their normal metabolic processes
These are microbial stages that can survive the complete drying process (desiccation). The organisms contained within the spores and cysts are in a dormant or resting state; if they are placed in a moist, nutrient-rich environment, they will grow and reproduce normally.
Bacterial endospores and protozoan cysts
Microorganisms that grow best at high temperatures are called what?
Thermophiles
(Thermophiles can be found in hot springs, compost pits, and silage as well as in and near hydrothermal vents at the bottom of the ocean)
Organisms that favor temperatures above 100°C are referred to as ______________________.
Hyperthermophiles (or extreme thermophiles).
The highest temperature at which a bacterium has been found living is about what? And what was it?
Pylorus fumarii (113 degrees Celsius)
These are organisms that prefer cold temperatures. They are seen in high altitudes, algae (often pink) and can be seen living on snow.
Psychrophiles
Psychrophiles can thrive at what temperatures?
These microbes thrive in an environment that is −13°C, has 20% of salinity and contains high concentrations of ammonia and sulfur.
This is the bacterium that causes cholera and is the only human pathogen that grows well above pH 8.
Vibrio Cholerae
In a hypotonic solution, the cell is a human cell, such as a red blood cell (erythrocyte), the loss of water causes the cell to shrink; this shrinkage is called ____________.
Crenation
In a hypotonic solution, the bacteria will then?
Die because of the loss of water and dessication
This is the formation of acanthocytes can be indicative of a number of hematologic disease processes.
Acanthocytosis
If the cell is a human cell, such as an erythrocyte, the increased water within the cell causes the cell to swell. If sufficient water enters, the cell will burst (lyse).In the case of erythrocytes, this bursting is called ____________.
Hemolysis
Piezophiles can survive up until what PSI? It is commonly seen among those that thrive under the ocean.
14.7 PSI
This is a mound or pile of bacteria containing millions of cells
Bacterial Colony
Under generation time, how long does Pseudomonas and Clostridium divide?
10 minutes
Under generation time, how long does Mycobacterium tuberculosis divide?
18 to 24 hours (Bacteria with short generation times are referred to as rapid growers, whereas those with long generation times as slow growers)
These are known to have complex nutritional requirements. Often, special mixtures of vitamins and amino acids must be added to the medium to culture these.
Fastidious organisms
What are the things that cannot grow on a medium?
(1) Obligate intracellular organisms such as viruses, rickettsia’s and chlamydia’s.
(2) Treponema pallidum (causes syphilis)
(3) Mycobacterium leprae (causes leprosy)
This is one in which the exact contents are not known. This contain ground-up or digested extracts from animal organs (e.g., hearts, livers, and brains), fish, yeasts, and plants, which provide the necessary nutrients, vitamins, and minerals.
Complex medium
These are contained in tubes and are thus often referred to as tubed media.
Liquid medium or broth
These are prepared by adding agar to liquid media and then pouring the media into tubes or Petri dishes, where the media solidifies
Solid medium
This is a complex polysaccharide that is obtained from a red marine alga; it is used as a solidifying agent, much like gelatin is used as a solidifying agent in the kitchen.
Agar
Blood agar is composed of what?
5% of nutrient agar and sheep red blood cells
Chocolate agar is composed of what?
5% of nutrient agar and hemoglobin (which makes it more enriched because hemoglobin is more readily accessible.
This is used to culture important, fastidious, bacterial pathogens, such as Neisseria gonorrhoeae and Haemophilus influenzae, which will not grow on blood agar.
Chocolate agar (Martin Lewis agar)
This inhibits the growth of Gram-positive bacteria and thus is selective for Gram-negative bacteria.
MacConkey agar
This agar inhibit the growth of Gram-negative bacteria and are thus selective for Gram-positive bacteria
Phenylethyl alcohol (PEA) agar and Colistin–nalidixic acid
(CNA) agar
This involves adding a portion of the specimen on the medium,
inoculation
Inoculation of a solid or plated medium involves the use of a sterile inoculating loop to apply a portion of the specimen to the surface of the medium; a process commonly referred to as _________.
Streaking
This is practiced in the microbiology laboratory to prevent infection of individuals and contamination of the work environment, clinical specimens, and cultures.
Aseptic Technique
Aseptic technique is practiced to prevent:
(a) microbiology professionals from becoming infected,
(b) contamination of their work environment, and
(c) contamination of clinical specimens, cultures, and subcultures.
How is the viable plate count measured?
To determine the concentration of bacteria in the original sample, the number of colonies must be multiplied by the dilution factor(s).
The first phase of the growth curve is the )___________, during which the bacteria absorb nutrients, synthesize enzymes, and prepare for cell division
Lag Phase
In this, the bacteria multiply so rapidly that the number of organisms doubles with each generation time (i.e., the number of bacteria increases exponentially). Growth rate is the greatest during at this phase. This phase appears as a steeply sloped straight line
Log Phase
Because the nutrients in the liquid medium are used up and the concentration of toxic waste products from the metabolizing bacteria build up, the rate of division slows, such that the number of bacteria that are dividing equals the number of bacteria that are dying. The result is the _____________. It is during this phase that the culture is at its greatest population density.
Stationary Phase
As overcrowding occurs, the concentration of toxic waste products continues to increase and the nutrient supply decreases. The microorganisms then die at a rapid rate; this is the _____________
death phase or decline phase.
Many industrial and research procedures depend on the maintenance of an essential species of microorganism. These are continuously cultured in a controlled environment called a ______________. Are cultured where E coli cells grow
Chemostat
This is a method disinfecting liquids. This is used today to eliminate pathogens from milk and most other beverages. It should be remembered that pasteurization is not a sterilization procedure because not all microbes are destroyed.
Pasteurization
Chemicals used to disinfect inanimate objects, such as
bedside equipment and operating rooms, are called ____________.
Disinfectants (cannot be used on tissues)
This is the reduction of microbial
populations to levels considered safe by public health standards, such as those applied to restaurants
Sanitization
This in a thermostatically controlled oven provides effective sterilization of metals, glassware, some powders, oils, and waxes. These items must be baked at 160°C to 165°C for 2 hours or at 170°C to 180°C for 1 hour.
Dry heat
This is an effective means of
destroying contaminated disposable materials. This must never be overloaded with moist or protein laden materials.
Incineration
The vegetative forms of most pathogens are quite easily destroyed by boiling for _________.
30 minutes
This conducted without the use of a pressure cooker does not destroy the endospores of bacteria— notably the anaerobe,
Home Canning (>60 degrees or 30 to 60 degrees)
This causes ice crystals to form within cells and may rupture the cell membranes and cell walls of some bacteria; hence, slow freezing should not be used as a way to preserve or store bacteria
Slow freezing
This using liquid nitrogen, is a good way to preserve foods, biologic specimens, and bacterial cultures
Rapid freezing
