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Starks's Microbiology- Lab Exam 2 Study Guide

These study guides are meant to help prepare students in Dr. John Star...
Course

General Microbiology (BIOL 3300)

19 Documents
Students shared 19 documents in this course
Academic year: 2016/2017
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Julianne Cardon
Utah State University
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Microbiology Lab Exam 2 Review Topics: Exercise 37 and 38: When is it appropriate to perform different stains? (Endospore, Acid Fast, etc.) Endospore: rods Acid Fast: forming rod Fluid Thioglycolate Its use and how to interpret results Determines oxygen tolerance for a Aerotolerance Gelatin stabs: What it is and what it is testing for. NOT FOR MOTILITY! Proteases Production: Enzymes that degrade protein can be detected the liquefaction of of the stab Growth Appearance in nutrient broth and gelatin inoculations Surface: Growth on top of bacteria Subsurface: Below the surface, turbidity or granular Sediment: Sediment in the bottom of tube Exercise 39: Catabolism: Breakdown of nutrients Anabolism: Biosynthesis of build up of cellular compounds Types of Enzymes: Exoenzymes: Enzymes that are outside the cell wall which degrade large macromolecules Coeznymes: Assist in catalytic reactions Oxidation vs Fermentation Oxidation (respiration): Organic molecules are completely degraded to carbon dioxide and water. ATP is generated a proton oxidation phosphorylation. Fermentation: The partial breakdown of organic components. ATP generated substrate level phosphorylation in low amounts glucose media Fermentation Test Microbiology Lab Exam 2 Review Topics: Stab tubs with inoculating needle and deliver about 1mL of sterile mineral oil one of the tube. You will compare the results of 1 anaerobic and 1 aerobic test. Anaerobic Aerobic Interpretation Yellow Yellow Oxidative and fermentative metabolism Green Yellow Oxidative Metabolism Green Green Sugar not metabolized (nonsaccharolytic) Specific Sugar Fermentation: Fermentation Test Inaccurate sugar tops with a loopful of bacteria, tubes initially red. Yellow Tubes: Indicate fermentation of sugar lowering pH Gas Production: gas or more is gas production of H2 and CO2 Red tubes: Indicate that organism is like Oxidative (no fermentation) The two tests that can be performed with media. Fermentation Test. Methyl Red Test (Mixed Acid Fermentation): Add drops of methyl red indicator to inoculated MR tubes Red: Indicates the organism has carried out mixed fermentation Unchanged: Organism does not create acids from glucose Test Fermentation): If methyl test fails add Barrits reagent and let stand for 30 minutes Bacteria produced from glucose Citrate Slants (Inoculation and interpretation): Fermentation Test. Slant are stabbed with culture. Culture uses citrate and ammonium salts Blue: pH has become alkaline, indicating the utilization of citrate (release of ammonia) Green: pH has not changed no utilization of citrate Oxidase Test (Media needed, procedure for test and results): Oxidative Test for cytochrome oxidase in the electron transport chain. Enzyme is detected artificial electron acceptor putting it on a swab of bacteria Purple Reaction: carries out aerobic respiration Yellow: may be aerobic but use cytochrome oxidase Catalase Test (Procedure and interpretation of results): Oxidative Test. Detects for catalases which indicate a bacteria uses aerobic respiration. Adding hydrogen peroxide test for the presence of catalase Bubbling: catalase is present No bubbling: Anaerobic catalase is not present Nitrate Test (The WHOLE procedure and interpretation of results): Oxidative Test 1. Check for gas production as a of nitrate reduction Microbiology Lab Exam 2 Review Topics: Alkaline red acidic butt: Glucose only fermentation acid on bottom ammonia on top Yellow slant: Fermented both glucose and lactose acid produced Red Glucose and lactose not fermented Cysteine Degradation (H2S formation): Testing for Cysteine Desulfurase. Ferrous salts in slant will react with hydrogen sulfide degraded from cystine. Insoluble Black Precipitate: Hydrogen sulfide production, and glucose has been fermented (yellow butt) SIM (Sulfur reduction, Indole production, and Motility): Inoculated stabbing Indole Test: Add agent and if slant turns bright red indole is being produced Hydrogen Sulfide test: If cysteine is being degraded H2S will react with ferrous salts to produce a black precipitate. (Cysteine Desulfurase) Motility Test: Is seem growth out from the line of inoculation or turbidity. Litmus Milk: Test for lactose fermentation Pink: Fermentation of lactose or dextrose, caused acid production Purple: Protease digestion of Casein (like skim milk streak plate). Causes release of ammonia and increases pH. Blue: Digestion of milk proteins Clotting: Production of Rennin, converts casein to paracasein. Acid production Yellow (reduction): Bacteria reducing the litmus dye, using it as an electron acceptor. This indicates redox reactions Exercise 44: Inoculation, processing of test kit, and interpretation of EnteroPluri tube results 1. Glucose Yellow Gas overlay moved 2. Presence of Lysine Purple (alkaline) 3. Presence of Ornithine Purple (alkaline) 4. Hydrogen Sulfide Black Indole after test 5. Fermentation of Yellow (acidic) 6. Fermentation of Lactose Yellow (acidic) 7. Fermentation of Arabinose Yellow (acidic) 8. Fermentation of Sorbitol Yellow (acidic) 9. VG test Red color after adding Barrits reagent 10. Fermentation of Dulcitol Yellow Phenylalanine Deaminase: 11. Production of Urease Purple (Alkaline) 12. Utilization of Citrate Blue (Alkaline) Exercise 70: Blood Agar: Different types of hemolysis and how to differentiate them. Microbiology Lab Exam 2 Review Topics: Hemolysis: Clear zone around the colonies, means complete lysis of red blood cells Hemolysis: Green color around cells, partially of hemoglobin Hemolysis: No effect on red medium means no hemolysis of blood B Tests for fluorescent pigments Fluorescent under uv light: Produces fluorescent pigments. Additional Topics Aseptic Technique: Disinfect work area, heat loops and needles till red hot, flame mouth of tubes, petri plate lids always diagonal. Isolation on a Streak Plate Gram Stain: Primary stain: Crystal Violet Mordant: forms insoluble complex with gram positive cells Decolorizer: Alcohol or acetone removes purple stain from gram cells Counterstain: Safranin is applied, causes cells to be pink Spore Stain: 1. Cover stain with piece of paper and saturate with malachite green, steam for 5 minutes 2. Rinse for 30 seconds after cooling 3. Counterstain with Safranin for 20 seconds Rinse with water briefly Acid Fast Stain: Motility Determination

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Starks's Microbiology- Lab Exam 2 Study Guide

Course: General Microbiology (BIOL 3300)

19 Documents
Students shared 19 documents in this course

University: Utah State University

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Microbiology Lab Exam 2 Review Topics:
Exercise 37 and 38:
When is it appropriate to perform different stains? (Endospore, Acid Fast, etc.)
- Endospore: Gram-positive rods
- Acid Fast: Gram-positive, non-spore forming rod
Fluid Thioglycolate Medium- Its use and
how to interpret results
Determines oxygen tolerance for a
organism- Aerotolerance
Gelatin stabs: What it is and what it is testing for. NOT FOR MOTILITY!
- Proteases Production: Enzymes that degrade protein can be detected by the
liquefaction of of the stab
Growth Appearance in nutrient broth and gelatin inoculations
- Surface: Growth on top of bacteria
- Subsurface: Below the surface, turbidity or granular
- Sediment: Sediment in the bottom of tube
Exercise 39:
Catabolism: Breakdown of nutrients
Anabolism: Biosynthesis of build up of cellular compounds
Types of Enzymes:
- Exoenzymes: Enzymes that are outside the cell wall which degrade large
macromolecules
- Coeznymes: Assist in catalytic reactions
Oxidation vs Fermentation
- Oxidation (respiration): Organic molecules are completely degraded to carbon
dioxide and water. ATP is generated by a proton gradient/ oxidation
phosphorylation.
- Fermentation: The partial breakdown of organic components. ATP generated by
substrate level phosphorylation in low amounts
Oxidative/Fermentative glucose media (O/F): Fermentation Test

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