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lab final
labs 21-31
| Question | Answer |
|---|---|
| coliforms | organisms that exist as part of the normal flora of humans and animals; capable of fermenting lactose w ensuing production of gas |
| noncoliform | enteric bacteria that lack the ability of coliforms to ferment certain carbohydrates |
| tests for gram neg rods | carbohydrate fermentation tests, urease test, IMViC series |
| nonfermenter gram neg | inhabitants of soil/water, nonsporing bacili or coccobacilli, |
| p aeruginosa isolated from | drains, sinks, faucetst, water from cut flowers, cleaning solutions, meds, and disinfectant soap solutions; spread patient-patient on hands of hospital perosnnel and direct contact |
| gram neg cocci examples | n. gonorrhoeae, n. meningitidis, and m. catarrhalis |
| neisseria speices show what morphology | diplococcus w adjacent flattened sides |
| tests for gram neg cocci | oxidase test, carbohydrate fermentation test, nitrate test |
| gram pos cocci morphology | grape like clusters |
| most common gram pos cocci | s. epidermidis on skin of humans (opportunistic pathogen) |
| s. aureus can cause | boils, impetigo, pneumonia, meningitis, food poisoning, and toxic shock syndrome |
| s. aureus spread thru | skin-skin contact, respiratory droplets and fomites |
| streptococci | gram pos cocci in chain; facultatively anaerobic and catalse not produced |
| s. aureus can be isolated by | streaking organisms on blood agar plate and observing pattern of hemolysis; coagulase tetst, dnase test, and gelatinase test |
| some streptococci are nonpathogenic, others cause | strep throat, neonatal meningitis, endocarditis, and gangrene |
| tests for streptococci | hemolysis on blood agar and CAMP test |
| gram pos rods examples | bacillis and clostridium; produce endospores |
| endospores | allow species to survive harsh environmental conditions |
| clostridial species are | obligate anaerobes |
| bacilllus species are | aerobic |
| tests for gram pos rods | hemolysis on blood agar, catalase and nitrate tests |
| in eukaryotes etc is located in | mitochondria inner membrane |
| in prokaryotes etc is located in | cytoplasmic membrane |
| cytochrome c oxidase in aerobes | accepts electrons from etc and transfers these electrons to molecular oxygen |
| some aerobes produce alternate oxidases that perform same activity as cytochrome c oxidase | this means not all aerobes are oxidase positive |
| to test for cytochrome c oxidase presence | do oxidase test |
| on oxidase strip | N,N-dimethyl-p-phenylenediamine and a-napthol are imprinted |
| let oxidase strip sit about 30 seconds | then color of inoculated area is observed |
| positive oxidase test appears | blue-purple in color |
| positive oxidase test means | possesses cytochrome c oxidase, bc enzyme oxidizes and couples reagents on strip to produce indophenol blue |
| negative oxidase test appears | no change/pink in color (no possession of cytochrome c oxidase) |
| oxidase test helps differentiate | gram neg rod/cocci bacteria in enterobacteriaceae genus |
| obligate anaerobes | undergo either fermentation or anaerobic respiration; oxygen not required for either process; lack both superoxide dismutase and catalase/peroxidase |
| facultative anaerobes | undergo aerobic respiration, anaerobic respiration, depending on conditions; can utilize oxygen so contain superoxide dismutase and catalase/peroxidase to detoxify by-products of metabolism |
| selective media | grow only one particular group of microbes |
| differential media | allow differentiation bw organisms based on metabolic capabilities |
| MacConkey agar | selective for gram negative organisms; contain crystal violet and bile salts; basis of lactose fermentation |
| neutral red indicatior of MacConkey agar | contrasts lactose-fermenting from lactose-nonfermenting organisms |
| if organism ferments lactose, causes | acidic by products to be released, causing a drop in pH; allowing colony to absorb neutral red indicator; precipitation of bile salts, so hazy precipitate around them |
| organism ferments lactose appears | pink/brick red |
| if organisms doesnt ferment lactose appears | translucent |
| to utilize glucose, it must possess a glucose transporter which | transports sugar from outside cell into cytoplasm |
| disaccharides like lactose and sucrose must first be | hydrolyzed to monosaccharide components before being transported into cell |
| B-galactosidase hydrolyzes | lactose into galactose and glucose |
| galactose is converted into intermediate compound | glucose 6-phosphate in glycolysis; then transported into cell |
| sucrase hydrolyzes | sucrose into glucose and fructose |
| fructose is converted into intermediate compound | glyceraldehyde 3-phosphate and dihydroxyaceton phosphate found in glycolysis; then transported into cell |
| glycolysis precedes | aerobic and anaerobic metabolic pathways |
| respiration | complete oxidation of organic compound to carbon dioxide and water |
| aerobic respiration | oxygen only molecule that serves as final electron acceptor |
| anaerobic respiration | organic/inorganic molecule other than oxygen serves as final electron acceptor |
| Triple sugar iron (TSI) agar differentiates | organisms based on abilities to ferment glucose, surcrose, or lactose; and to liberate hydrogen sulfide gaas from sodium thiosulfate |
| TSI contains | 1% solution of lactose/sucrose and 0.1% solution of glucose; as well as sodium thiosulfate and ferrous sulfate; detects coliforms |
| phenol red in TSI detects | changes in pH due to production of acidic or alkaline end-products |
| if organism ferments glucose only, | both slant and butt of medium will become yellow (acidic end products produced) |
| slant will become alkaline due to | oxidation of acids by oxygen bc glucose concentration is low |
| acidic rxn maintained in | butt due to low oxygen concentration |
| if slant/butt are both yellow | organism ferments both lactose/sucrose |
| if no change in color in slant/butt | organism cannot ferment any carbohydrates |
| cracks or splits in TSI show that | carbon dioxide or hydrogen gases were produced as result of fermentation of carbs |
| sodium thiosulfate and ferrous sulfate allow for detection of | colorless hydrogen sulfide gas (H2S) |
| hydrogen sulfide production w peptones | peptones are degrades to amino acids (cysteine) and are reduced to pyruvate, ammonia and hydrogen sulfide gas by cysteine desulfurase |
| peptone | short chains of amino acids which serve as major carbon and energy sources for microbes |
| hydrogen sulfide production w ferrous ammonium sulfate/sodium thiosulfate | sulfate serves as final electron acceptor in anaerobic respiration and thiosulfate reductase reduce sulfate w production of hydrogen sulfide gas |
| ferrous sulfide when combines ferrous sulfate and hydrogen sulfide gas appears | black deposits w/in TSI medium |
| TSI agar used to identify | gram pos rods that are catalase neg |
| tryptophanase | hydrolyzes amino acid tryptophan into indole, pyruvic acid, and ammonia |
| pyruvic acid | precursor molecule in fermentation or respiration |
| ammonia | helps to fulfill nutritional requirements for organism |
| indole is a | waste product!!!!!! |
| indole test indirectly detects | presence of tryptophanase by measuring end-product of rxn involving enzyme |
| indole test done by | stabbing inoculated sulfide-indole-motility agar deep (SIM agar deep); then incubated; add kovac's reagent to detect tryptophan |
| sulfide-indole-motility (SIM) agar deep contains | casein peptones rich in amino acid tryptophan |
| kovac's reagent is composed of | 4- dimethylaminobenzaldehyde, butanol, and hydrochloric acid |
| if indole present (positive) | extracted into reagent layer by acidified butanol and reacts w 4-dimethylaminobenzaldehyde to form cherry red color |
| if indole not present (negative) | no color change appears |
| if organisms have cysteine desulfurase or thiosulfate reductase | hydrogen sulfide gas will be produced, when combined w ferrous ammonium sulfate, it produces ferrous sulfide (blackening in SIM) |
| if bacteria are motile in SIM | growth will not be restricted to line of stab inoculation and will be evident throughout tube |
| two fermentation patterns | mixed-acid fermentation and 2,3-butanediol fermentation |
| mixed-acid fermentation | concentrations of lactic, acetic, formic, and succinic acids are formed following glycolytic conversion of glucose to pyruvate; ethanol, CO2, and hydrogen gas are formed too, lower pH to ~4 |
| 2,3-butanediol fermentation | acids formed, but neutral products 2,3-butanediol, ethanol, carbon dioxide, and hydrogen gas predominate; pH ~6 |
| methyl red-Voges Proskauer (MRVP) test differentiates organisms by detecting | fermentative end-products following growth in buffered peptone-glucose broth |
| methyl red test | pH indicator methyl red is added to determine if acids lowered pH of medium; at pH 4=methyl red indicator turns medium red |
| Voges Proskauer test detects presence of | acetoin, which is intermediate in production of 2,3-butanediol; |
| acetoin is identified when addition of | 40% potassium hydroxide and alcoholic a-napthol solution |
| if acetoin is present | it is oxidized to diacetyl compound that imparts rose color to medium |
| Simmon's citrate agar tests for | ability of organisms to utilize citrate as sole source of carbon; also contains ammonium as sole source of nitrogen |
| if citrate is utilized, | organisms must have enzyme citrate permease, which facilitates transport of citrate into cell; then citrase oxidizes the substrate |
| during oxidation of citrate, | carbon dioxide, pyruvic acid, and oxaloacetic acid are produced |
| pyruvic acid and oxaloacetic acid are used as precursor metabolites, while | carbon dioxide combines w water and excess sodium to form soidum carbonate (alkaline end product) which raises pH of medium |
| to detect change in pH, from citrate oxidation | bromthymol blue is included in the medium; |
| bromthymol blue is green | at neutral pH |
| bromthymol blue is bleu | if pH becomes more basic |
| if organism cannot use citrate as sole carbon source, | no growth is evident on simmon's citrate agar |
| urea | waste product of protein degradation, mostly excreted in urine; some use as source of nitrogen |
| urease | hydrolyzes carbon-nitrogen bond in urea, resulting in carbon dioxide, water, ammonia |
| accumulation of ammonia allows for detection of | organisms that produce urease |
| urea agar contains | urea, peptones, dextrose; phenol red (pH indicator); pH 6.8 when supplied/salmon orange at neutral pH |
| urea causes the alkaline ammonia to accumulate | raising the pH in the medium (>8.4 the medium will be fuchsia) (positive result) |
| if urea not degraded, accumulation of ammona will | keep pH of medium neutral (negative result) |
| at acidic pH, phenol red causes urea medium to | change to yellow; but still negative bc urea not degraded |
| biogeochemical cycles | recycling of chemical elements, C, N, S |
| 4 steps in nitrogen cycle | nitrogen fixation, ammonification, nitrification, denitrification |
| nitrogen is needed for | synthesis of proteins and nucleic acids; most abundant gas in atmosphere but only few organisms can use it in gas form, most use it in ammonia (NH3) form |
| nitrogen fixation | how organisms possess enzyme nitrogenase convert atmospheric nitrogen into ammonia; completed by free-living nitrogen-fixing bacteria or symbiotic nitrogen-fixing bacteria |
| free-living nitrogen-fixing bacteria | found in high concentrations where soil and plant roots make contact; protect or sequester nitrogenase in anaerobic heterocysts |
| symbiotic nitrogen-fixing bacteria | infect roots of leguminous plants (soybeans, beans, peas, alfalfa, clover, peanuts) and form root nodules |
| root nodules | growths on plant roots that harbor bacteria |
| how nitrogen fixation works | bactiera fix nitrogen for the plant, while plant provides anaerobic conditions under nitrogenase functions and provident nutrients from plant sap |
| ammonia becomes available to other organisms when | organisms or host plants die/or are eaten |
| ammonification | amino groups are removed and converted into ammonia, which is liberated |
| in dry soils, | gaseous ammonia disappears into atmosphere |
| in moist soils, | ammonia is converted into ammonium ions that are used by bacteria and plants for amino acid, purine, pyrimidine synthesis |
| nitrification | nitrogen in ammonia oxidized to produce nitrate via 2 step process; ammonia first oxidized to nitrite ion and then nitrites oxidized to yield nitrate ions |
| nitrification requires molecular oxygen so | rxn occurs only in oxygenated water/soils |
| denitrification | nitrate ion is converted via multiple steps to more reduced forms of nitrogen (nitrite ion, nitric oxide, nitrous oxide, nitrogen gas) |
| first step in denitrification | reduction of nitrates to nitrites, requires nitrate reductase |
| denitrification results in | removal of nitrates from soil and interes w plant growth; accumulation of nitric oxide, that destroyes ozone layer |
| nitrate broth contains | potassium nitrate as source of nitrate, durham tubes |
| durham tubes | trap any nitrogen gas produced during denitrification |
| after incubation of nitrate broth, add | nitrate reagent a (sulfanilic acid) and nitrate reagent b (a-napthylamine) |
| if nitrites present (positive) | form a diazonium salt to produce red precipitate, if no color shows add zinc dust |
| if after adding zinc, no color change (positive) | nitrites were reduced by microbe in denitrification to yield nitric oxide, nitrous oxide, or nitrogen gas |
| red color after adding zinc (neg) | unreduced nitrates still present in tube and reduced to nitrites by zinc |
| catalase present in | all cells of plants, animals, aerobic bacteria; neutralizes >40,000,000 molecules of hydrogen peroxide per sec |
| catalase is located in eukaryotes | in lysosomes and peroxisomes |
| lysosomes | aid in digestion of macromolecules |
| peroxisomes | rid cell of toxic substances |
| catalase located in prokaryotes | in periplasmic space bw cell wall and cytoplasmic membrane |
| superoxide dismutase | catalyzes conversion of free radical superoxide to molecular oxygen and hydrogen peroxide |
| peroxide anion produced | toxic to cells and must be neutralized |
| catalase | converts peroxide anion to water and oxygen |
| presence of catalase is detected by | adding drop of substrate H2O2 to cells and looking for bubbling (pos) |
| if bubbling doesnt occur | organism is either anerobe or breaks down peroxide anion w peroxidase |
| Colombia CNA agar with 5% sheep blood | selective for gram pos organisms |
| Colombia CNA contains | colistin (c) and nalidixic acid (na) |
| colistin | disrupts cytoplasmic membrane of gram neg bacteria |
| nalidixic acid | prevents dna replication in susceptible gram neg organisms |
| 5% sheep blood | provides hemin and vitamin k to aid in cultivation of fastidious microbes amd differentiate organisms based on hemolysis |
| B-hemolysis | clear zones around colony, with little or no rbcs present |
| a-hemolysis | partial/incomplete hemolysis of rbc and rbc appear discolored |
| y-hemolysis | no change is observed w medium and no lysing of rbc occurs |
| mannitol salt agar contains | 7.5% sodium chloride, selective for halophiles; phenol red |
| halophiles | salt loving organisms |
| if organism ferments mannitol | acids lower pH of medium, which turn medium yellow |
| if organism does not ferment mannitol | medium stays translucent |
| fibrinogen used for | embryogenesis, tissue repair, blood clotting, cell migration/adhesion |
| insoluble fibrinogen | serves as linker in connective tissue that surrounds and supports cells in mammalian tissues |
| soluble fibrinogen | hepatocytes (liver cells) and found in plasma |
| plasma | clear, yellowish fluid portion of blood, lymph, intramuscular fluid that contains wbc and soluble clotting factors |
| prothrombin | vitamin k dependent proenzyme |
| coagulase | extracellular protein involved in formation of clots; produced by staphylococci and yersinia pestis |
| coagulase secreted by bacterial cells | binds to prothrombin to form staphylothrombin which activates thrombin (converts fibrinogen to fibrin) |
| coagulase test | inoculate organism into tube of citrated rabbit plasma; look fir fibrin clot formation; differentiate gram pos cocci |
| citrate rabbit plasma contain | citrate, EDTA, and rabbit plasma (source of fibrinogen) |
| nuclease | degrades nucleic acids and foreign double stranded DNA thru the phosphodiester bonds bw adjacent nucleotides in dna |
| foreign double stranded DNA introduced to cell by | gene transfer mechanisms or viral infection |
| dna methylase | methylates and protects host dna |
| internal/extracellular nucleases | degrade long, double stranded dna molecules into smaller pieces |
| pus | accumulation of live bacteria, live wbc, cell debris of both bacteria and wbc |
| dnases degrade pus and reduce | viscosity to allow for easier dissemination of organisms to other tissues |
| dnase test | organisms spot-inoculated onto dnase test agar plate; distinguished gram pos cocci |
| dnase agar contain | soy and casein peptones, sodium chloride, polymerized dna |
| if organisms produces dnase | is it secreted outside bacterial cell and degrades dna into smaller pieces |
| detectuib if depolymerized dna | flooding agar surface w 1N HCl and observing for medium to develop clear zone surrounding growth |
| in absence of dnase, | 1N HCl reaction w polymerized dna results in cloudy precipitate in medium |
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