Microbiology Lab pra
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| OF Medium | Semisolid Agar deep with high carbs and low peptones. Peptones support growth of bacteria that don't use carbs.
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| Description of OF Medium | 2 tubes used: 1 open to the air and 1 sealed to keep air out. Contains bromthymol blue, & it turns yellow in presence of Acids, indicating catabolism of the carb. Alkaline due to use of peptone not the carb turn dark blue due to ammonia production.
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| What has occurred if the carb is metabolized in both tubes of a OF Medium? | Fermenation
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| If an organism uses carbs in an open tube only, what will they produce in an OF Medium? | Acid, acids are produced as intermediates in respiration, and the indicator will turn yellow in the top of the open tube. Organism is called oxidative.
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| OF Test Results and Interpretations |
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| What would be found in the clear area that would not be found in the blue area of a starch agar plate after the addition of iodine? | A-amylase and glucose
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| How can you tell if an organism uses glucose aerobically? Ferments glucose? Doesn't use glucose? | Tube will be yellow throughout & leave the sealed medium green & blue. Yellow throughout or at the top of tube. Green or blue throughout
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| Aerobic organisms degrade glucose, producing carbon dioxide and water. What acid turns the indicator yellow? | Pyruvate acid
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| How can organims that don't use starch grow on a starch plate? | Low levels of peptones are in the media that supports the growth of the non-starch bacteria.
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| What is an fermentation tube? | it's used to detect acid and gas production form carbohydrates
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| What does the fermentation medium contain? | Peptone, and acid-base indicator (phenol red), and inverted tube to trap gas (Durham tube), and .5-1.0% of the desired carb.
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| What color is the tube before inoculation and after acid production? | red (neutral); acid production will turn the indicator yellow (pH of 6.8 or below).
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| Do all bacteria ferment lactose? | No
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| Why are fermentation tubes evaluated at 24 and 48 hours? | they will begin to grow oxidatively (passed 24hrs) on peptone exhausting the carbs and turn the medium red. Masking any evidence of fermentation.
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| If the medium is red for the fermentation tubes what does that indicate? | It has used up all the carbs and is growing oxidatively on peptone and it turns red because of the ammonia products produced.
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| If organism metabolized glucose aerobically, what results will occur in the fermentatin tubes? | increase acid production, turning it yellow & producing gas in the tubes.
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| If bacterium cannot ferment glucose, why not test its ability to ferment other carbohydrates? | b/c its the simplest sugar and other sugars are more complex.
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| If bacterium cannot use glucose, what result will occur in a fermentation tube? | tube will stay the same color but gas may occur.
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| What is the MRVP test? | it's used to distinguish organisms that produce large amounts of acid from glucose and organisms that produce the neutral product acetoin.
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| A positive MR test | only if an organism produces a large amount of organic acid from glucose, medium will remain red when methyl red is added, below pH 4.4
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| Negative MR Test | methyl red will turn yellow in presence of pH above 6.0
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| Positive VP Test | upper part of medium turns red in the presence of acetoin with the addition of potassion hydroxide and a-naphthol.
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| Negative VP Test | A negative will turn the upper part of medium brown due to the reaction of KOH and a-naphthol.
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| What is citrate agar used for? | Useful in identifying bacteria that can ferment citrate.
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| What does a bacteria need in order to grow on a citrate medium? | Bacteria with the enzyme citrate lyase can break down citrate to form pyruvate, which can be reduced in fermentation.
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| Positive Citrate test | bacteria use citrate and ammonium, the medium is alkalized because of ammonia produced from NH4. The indicator bromthymol blue changes to blue when medium is alkalized.
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| Negative Citrate Test |
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| What are proteins | large organic molecules that include cellular enyzmes and many structures
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| Amino Acids | Subunits that make up proteins
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| Amino acids consist of | carbon,hydrogen, oxygen, nitrogen, and sometimes sulfur.
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| Peptide bonds | bonding of amino acids forming a peptide or a larger molecule polypeptide.
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| What can bacteria do to peptides or polypeptides | bacteria can hydrolyze to release amino acids and use the amino acids as carbon and energy sources when carbs aren't available
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| Nutrient Gelatin | dissolves in warm water (50 C), solidifies (gels) when cooled below (25 C) and liquefies (sols) when heated to about 25 C.
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| What happens to the gelatin when an organism produces an exoenzyme hydrolyzes the gelatin? | It liquefies it and the gelatin doesn't solidify even when cooled below 20 C.
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| Urea is | a waste product of protein digestion in most vertebrates and is excreted in the urine.
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| Urea Agar | ids bacteria that possess urease, which liberates ammonia from the urea, which raises the pH of the medium turning the indicator fuchsia (hot pink) at pH 8.4
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| Do all bacteria produce urease? Gelatinase? | No and No bacteria would need either a particular exoenzyme (gelatinase & urease) to overcome these buffers.
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| Name the 5 types of bacteria growth | Obligate Aerobes, Facultative anaerobes, Obligate anaerobes, Aerotolerant anaerobes, and Microaerophiles
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| Obligate aerobes | Oxygen required, presence of enzymes catalase and superoxide dismutase (SOD) allows toxic forms of O2 neutralized,
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| Facultative Anaerobes | aerobic and anaerobic growth, greater growth in O2. Catalase and (SOD) enzymes present
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| Obligate Anaerobes | Only anaerobic growth, lacks enzymes to neutralize O2.
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| Aerotolerant Anaerobes | only anaerobic growth but continues in presence of O2, has one enzyme SOD, tolerates it.
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| Microaerophiles | only aerobic growth but in low O2 concentration and high CO2, they produce superoxide radicals and peroxide in lethal concentrations.
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| Majority of Bacteria are | Facultative anaerobes - capable of living with or without O2
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| 5 bacteria lacking catalase are | Streptococcus, Enterococcus, Leuconostoc, Cactobacillus,(these 4 are aerotolerant anaerobes and Clostridium.
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| Species of Clostridium | are obligate anaerobes
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| Streptococcus, Enterococcus, Leuconostoc, and Cactobacillus are | aerotolerant anaerobes and lack the cytochrome system to produce hydrogen peroxide and therefore do not need catalase.
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| Catalase test | determining the presence or absense of catalase can help ID bacteria. by adding a few drops of 3% hydrogen peroxide to a microbial colony and if catalase is present molecular O2 is released as bubbles.
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| Why does Thioglycolate broth appear red or blue? | Resazurin (dye) is added to the broth which is pink in the presence of excess oxygen an colorless when reduced. Methylene blue is also and indicator.
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| Generation or doubling time | is the time it takes for one cell to divide into 2 cells or the time required for the population of cells to double.
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| Shorter the generation time represents | the faster the growth
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| Minimum growth temperature | is the lowest temperature at which a species will grow.
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| Optimum Growth | species grows the fastest
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| Maximum growth temperature | highest temperature at which a species can grow.
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| At temperatures near the max growth temp | growth ceases, presumbably because of inactivation of enzymes.
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| Lag Phase | Little or no cell division, undergoing a period of intense metabolic activity, involving synthesis of enzymes and various molecules
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| Log Phase | Growing at fastest possible rate
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| Stationary Phase | Period of equilibrium, microbial deaths balanced production of new cells. Due to possible accumulation of waste products, exhaustion of nutrients and harmful changes in pH.
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| Death Phase | Population is decreasing at a logarithmic rate. Death rate exceeds new cells rate.
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| Phases of Growth | Lag phase, Log phase, stationary phase, and Death phase
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| What is the optimum growth temp. for human pathogens | 37 C
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| What are the types of bacteria live in relation to environment | Psychrophilic, Psychrotrophic, Mesophilic, thermophilic, hyperthermophilic
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| Psychorophilic | 0 to 20 degrees C, cold-loving microbes, found in ocean depths or certain polar regions.
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| Psychrotrophic | 20 to 30 degrees C, cold-loving, can't grow above 40 degrees, low temp food spoilage, grow well in refrigerators.
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| Mesophilic | 25 to 40 degrees C, moderate-temp loving microbes, most common, most common spoilage and disease organisms
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| Thermophilic | 45 to 65 degrees C,heat-loving microbes, temp of hot water tap, sunlight soil and hot springs. can't survive in temps below 45C
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| Hyperthermophiles | about 80 or higher degrees, heat-loving, hot springs associated with volcanic activity, sulfur usually metabolic activity
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| Dry Heat | hot-air ovens or incineration denatures enzymes, dehydrates microbes, and kills oxidation effects.
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| Moist Heat | methods include pasteurization, boiling, and autoclaving. It transfers heat energy to the microbial cell more efficiently than dry air, resulting in denaturing of enzymes.
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| Pasteuriztion | the temp is maintained at 63 degrees for 30 min or 72 degrees for 15 sec. to kill designated organisms that are pathogenic or cause spoilage
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| Boiling | 100 degrees C for 10 minutes will kill vegetative bacterial cells; however does not inactivate endospores.
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| Autoclaving | the use of steam under pressure, increase pressure raises the boiling point of water and produces steam with a higher temp. Most effective. Standard conditions are 15 psi, at 121 degrees for 15 minutes. this usually kills endospores.
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| Thermal Death Time | is the length of time required to kill all bacteria in a liquid culture at a given temp.
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| Thermal death point | is the temp required to kill all bacteria in a liquid culture in 10 minutes.
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| Decimal Reduction time | is the time, in minutes, in which 90% of a population of bacteria at a given temperature will be killed.
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| Antimicrobial Agents | are chemicals used to control the growth of microbes.
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| Disinfectants | are chemical agents used on inanimate objects to lower the level of microbes on their surface.
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| Antiseptics | are chemicals used on living tissue to decrease the number of microbes.
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| Bactericidal Agents | result in bacterial death
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| Bacteriostatic Agents | causes temporary inhibition of growth.
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| In evaluating the effectiveness of antimicrobial agents | the concentration, length of contact, and whether it's lethal are the important criteria.
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| The use-dilution test | is limited to bactericidal compounds and cannot be used to evaluate bacteriostatic compounds.
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| Zones of Inhibition | the size of inhibition of bacterial growth around a disk that is impregnated with various antimicrobial agents.
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| Minimum inhibitory Concentration, MIC of an antibiotic | is determined by testing for bacterial growth in dilutions of the antibiotic in nutrient broth.
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| Antimicrobial Drugs | antimicrobial chemicals absorbed or used internally, whether natural (antibiotics) or synthetic.
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