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Microbiology Lab pra

OF Medium Semisolid Agar deep with high carbs and low peptones. Peptones support growth of bacteria that don't use carbs.
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.
What has occurred if the carb is metabolized in both tubes of a OF Medium? Fermenation
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.
OF Test Results and Interpretations
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
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
Aerobic organisms degrade glucose, producing carbon dioxide and water. What acid turns the indicator yellow? Pyruvate acid
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.
What is an fermentation tube? it's used to detect acid and gas production form carbohydrates
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.
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).
Do all bacteria ferment lactose? No
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.
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.
If organism metabolized glucose aerobically, what results will occur in the fermentatin tubes? increase acid production, turning it yellow & producing gas in the tubes.
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.
If bacterium cannot use glucose, what result will occur in a fermentation tube? tube will stay the same color but gas may occur.
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.
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
Negative MR Test methyl red will turn yellow in presence of pH above 6.0
Positive VP Test upper part of medium turns red in the presence of acetoin with the addition of potassion hydroxide and a-naphthol.
Negative VP Test A negative will turn the upper part of medium brown due to the reaction of KOH and a-naphthol.
What is citrate agar used for? Useful in identifying bacteria that can ferment citrate.
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.
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.
Negative Citrate Test
What are proteins large organic molecules that include cellular enyzmes and many structures
Amino Acids Subunits that make up proteins
Amino acids consist of carbon,hydrogen, oxygen, nitrogen, and sometimes sulfur.
Peptide bonds bonding of amino acids forming a peptide or a larger molecule polypeptide.
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
Nutrient Gelatin dissolves in warm water (50 C), solidifies (gels) when cooled below (25 C) and liquefies (sols) when heated to about 25 C.
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.
Urea is a waste product of protein digestion in most vertebrates and is excreted in the urine.
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
Do all bacteria produce urease? Gelatinase? No and No bacteria would need either a particular exoenzyme (gelatinase & urease) to overcome these buffers.
Name the 5 types of bacteria growth Obligate Aerobes, Facultative anaerobes, Obligate anaerobes, Aerotolerant anaerobes, and Microaerophiles
Obligate aerobes Oxygen required, presence of enzymes catalase and superoxide dismutase (SOD) allows toxic forms of O2 neutralized,
Facultative Anaerobes aerobic and anaerobic growth, greater growth in O2. Catalase and (SOD) enzymes present
Obligate Anaerobes Only anaerobic growth, lacks enzymes to neutralize O2.
Aerotolerant Anaerobes only anaerobic growth but continues in presence of O2, has one enzyme SOD, tolerates it.
Microaerophiles only aerobic growth but in low O2 concentration and high CO2, they produce superoxide radicals and peroxide in lethal concentrations.
Majority of Bacteria are Facultative anaerobes - capable of living with or without O2
5 bacteria lacking catalase are Streptococcus, Enterococcus, Leuconostoc, Cactobacillus,(these 4 are aerotolerant anaerobes and Clostridium.
Species of Clostridium are obligate anaerobes
Streptococcus, Enterococcus, Leuconostoc, and Cactobacillus are aerotolerant anaerobes and lack the cytochrome system to produce hydrogen peroxide and therefore do not need catalase.
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.
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.
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.
Shorter the generation time represents the faster the growth
Minimum growth temperature is the lowest temperature at which a species will grow.
Optimum Growth species grows the fastest
Maximum growth temperature highest temperature at which a species can grow.
At temperatures near the max growth temp growth ceases, presumbably because of inactivation of enzymes.
Lag Phase Little or no cell division, undergoing a period of intense metabolic activity, involving synthesis of enzymes and various molecules
Log Phase Growing at fastest possible rate
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.
Death Phase Population is decreasing at a logarithmic rate. Death rate exceeds new cells rate.
Phases of Growth Lag phase, Log phase, stationary phase, and Death phase
What is the optimum growth temp. for human pathogens 37 C
What are the types of bacteria live in relation to environment Psychrophilic, Psychrotrophic, Mesophilic, thermophilic, hyperthermophilic
Psychorophilic 0 to 20 degrees C, cold-loving microbes, found in ocean depths or certain polar regions.
Psychrotrophic 20 to 30 degrees C, cold-loving, can't grow above 40 degrees, low temp food spoilage, grow well in refrigerators.
Mesophilic 25 to 40 degrees C, moderate-temp loving microbes, most common, most common spoilage and disease organisms
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
Hyperthermophiles about 80 or higher degrees, heat-loving, hot springs associated with volcanic activity, sulfur usually metabolic activity
Dry Heat hot-air ovens or incineration denatures enzymes, dehydrates microbes, and kills oxidation effects.
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.
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
Boiling 100 degrees C for 10 minutes will kill vegetative bacterial cells; however does not inactivate endospores.
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.
Thermal Death Time is the length of time required to kill all bacteria in a liquid culture at a given temp.
Thermal death point is the temp required to kill all bacteria in a liquid culture in 10 minutes.
Decimal Reduction time is the time, in minutes, in which 90% of a population of bacteria at a given temperature will be killed.
Antimicrobial Agents are chemicals used to control the growth of microbes.
Disinfectants are chemical agents used on inanimate objects to lower the level of microbes on their surface.
Antiseptics are chemicals used on living tissue to decrease the number of microbes.
Bactericidal Agents result in bacterial death
Bacteriostatic Agents causes temporary inhibition of growth.
In evaluating the effectiveness of antimicrobial agents the concentration, length of contact, and whether it's lethal are the important criteria.
The use-dilution test is limited to bactericidal compounds and cannot be used to evaluate bacteriostatic compounds.
Zones of Inhibition the size of inhibition of bacterial growth around a disk that is impregnated with various antimicrobial agents.
Minimum inhibitory Concentration, MIC of an antibiotic is determined by testing for bacterial growth in dilutions of the antibiotic in nutrient broth.
Antimicrobial Drugs antimicrobial chemicals absorbed or used internally, whether natural (antibiotics) or synthetic.
Created by: Twixy565 on 2010-10-19

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