Microbiology Lab Midterm #2 Review PDF

- Temperature - Psychrophiles: Grow well at 0 degree Celsius; optimum: 15 degrees or lower - Psychrotrophs: Facultative psychrophiles; grow at 0 -- 7 degrees Celsius - Mesophiles: Growth optima 25 -- 45 degrees Celsius. Majority of bacterial species are...

- Temperature - Psychrophiles: Grow well at 0 degree Celsius; optimum: 15 degrees or lower - Psychrotrophs: Facultative psychrophiles; grow at 0 -- 7 degrees Celsius - Mesophiles: Growth optima 25 -- 45 degrees Celsius. Majority of bacterial species are in group 25 -- 45 degree Celsius - Thermophiles: Grow at 55 degrees or higher - Hyperthermophiles: Grow at 90 degrees or above - Thermoduric: Bacteria that can survive boiling even though they cannot grow - Usually, they are spore forming - pH - Affects prokaryotic growth by affecting activity of enzymes; especially involved in biosynthesis - Acidophiles thrive under highly acidic conditions (usually at pH 2 or below) - Neutrophiles are neutrophilic organisms that thrive in a neutral pH environment between 6.5 and 7.5. Most bacteria are neutrophiles and they produce metabolic acids. When growing them, buffers are added to the media. Buffers produce and neutralize acids produced by bacteria - Examples: Peptones, Phosphate Salts - Alkaliphiles are capable of survival in alkaline (pH roughly 8.5 -- 11) environments, growing optimally around a pH of 10 - Salt tolerance - Non halophile: grows at less than 1% salt concentration - Halotolerant: grows at salt concentration from 0 -- 10% - Halophile: microbe that can tolerate high salt (NaCl) concentrations (halo means salt) - Moderate halophile: live at 4.7 -- 20% salt - Extreme halophile: tolerate 20 -- 30% salt concentration https://www.scienceabc.com/wp-content/uploads/2019/07/salt-graph.jpg - Different types of solutions - Isotonic: Water concentration is equal inside and outside the cell; thus, rates of diffusion are equal in both directions - Hypotonic: Net diffusion of water is into the cell; this swells the protoplast and pushes it tightly against the wall. Wall usually prevents cell from bursting - Hypertonic: Water diffuses out of the cell and shrinks the protoplast away from the cell wall; process is known as plasmolysis - Antibiotic resistency - The smaller diameter or no zone = bacteria is resistant to the antibiotic tested - This is established through the usage of the Kirby-Bauer Method - The antibiotics are impregnated onto paper disks then placed on agar plates - Minimum Inhibitory Concentration - Is the lowest concentration of an antibiotic that inhibits a visible growth of a microorganism after overnight incubation - Antiseptic - Are antimicrobial agents that are applied to living tissues or body surfaces to destroy or inhibit vegetative pathogens - They can be used on human or animal body, skin, mycoses, and wounds - Disinfectant - Are antimicrobial agents that are applied to the surface of non-living objects to destroy microorganisms - Disinfection does not necessarily kill all microorganisms, especially resistant bacterial spores; it is less effective than sterilization, which is an extreme physical and/or chemical process that kills all types of life - Disinfectants work by destroying the cell wall of microbes or interfering with their metabolism - Microbicidal - Kills microorganisms - Microbiocidal efficiency of a chemical is determined with respect to phenol, thus termed Phenol Coefficient - Phenol Coefficient - Highest dilution of antimicrobial of interest (divided by) highest dilution of phenol with same characteristics - Incubation with highest dilution of the antimicrobial of interest that kills all organisms after 10 minutes not 5 minutes - If coefficient is greater than 1, then it is more efficient than phenol - Microbiostatic - Inhibit microbial growth - Efficiency of a chemical -- determined by ability to deter microbial growth - Gelatin test - Checks if bacteria contains enzymes gelatinase - Test for pathogenicity of bacteria, as gelatinase is correlated to the breakdown of tissue collagen - Positive test: liquified - Negative test: solid  - Catalase test - Checks if bacteria contains catalase - If oxygen is reduced, the production of superoxide radicals (like hydrogen peroxide) can occur - Catalase can destroy these radicals - Positive test: bubbles - Negative test: no bubbles A close-up of a test tube Description automatically generated - Coagulase test - Checks if bacteria contains coagulase - Bacteria can form a clot around themselves for protection if coagulase is present - Positive test: clotting - Negative test: no clotting  - DNase test - Checks if bacteria contains DNase - DNase can degrade a host's DNA and shows pathogenicity of bacteria - Positive test: clear zone around colony - Negative test: no clear zone around colony A petri dish with a number Description automatically generated - Oxidase test - Checks if bacteria contains oxidase enzyme - Oxidase is important for energy production as it is used in ETC (electron transport chain) - Positive test: purple color - Negative test: light pink  - Urease test - Checks if bacteria contains urease enzyme - Urea can be broken down by urease to isolate nitrogen for use - If urease breaks down urea, ammonia will be formed - Ammonia is basic and will cause an upward shift in pH - A shift in pH causes phenol red indicator to create color change - Positive test: pink/purple color - Negative test: yellow/green color Urease - Cysteine test - Cysteine is an amino acid that contains sulfur - Checks if bacteria contains enzyme cysteine desulfurase (means removal of sulfur) - If cysteine desulfurase is present, sulfur will be removed and H2S gas is produced - Positive test: black precipitate - Negative test: no precipitate  - Phenylalanine Deamination test - Phenylalanine is an amino acid - Checks if bacteria contains phenylalanine deaminase (removal of amino group) - If the amino group is removed, then phenyl pyruvic acid is produced - We can use phenyl pyruvic acid as an indicator of bacteria containing phenylalanine deaminase - By adding ferric chloride, it will react with the pyruvic acid, if no pyruvic acid is present, then there will be no reaction - Positive test: green - Negative test: yellow - Nitrate Reduction test - Checks if bacteria contain nitrate reductase (reduction of nitrate to nitrite) - If bacteria can reduce nitrate, then nitrite ions will be present in media - We can check for these ions by adding sulfanilic acid and dimethyl naphthylamine. If there is a red color change, then nitrite is present (this means bacteria was able to perform the reduction) - What if there is no color change? What does this mean? - Nitrate was never reduced, or nitrate was reduced but the nitrite ions were reduced to ammonia via another enzyme - If there is no color change, it does not always mean that there is no nitrate reductase but rather the nitrite ions may have been reduced further by another enzyme - How can we tell if nitrate reductase is present without a color change after adding our 2 reagents above? - The usage of zinc - Zinc will reduce nitrate to nitrite - If nitrate is present in the media, then zinc will reduce the nitrate to nitrite and a red color change will occur; this means nitrate reductase was never present in the bacteria - If zinc is added and there is no color change, then this means that the bacteria had reduced nitrate already, but the nitrite ions were reduced further - Lysine and Ornithine test - Decarboxylation of lysine: cadaverine - Decarboxylation of ornithine: putrescine - Positive test: purple **Lysine Test Ornithine Test** - Tryptophan Deaminase test (TDA) - Checks if bacteria can deaminate tryptophan to make indole pyruvic acid - Positive test: red/brown A test tube with different colored liquids Description automatically generated - What type of bacteria does the API 20 system identify? - Enterobacteriaceae (gram negative)