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Micro- Lab Prac 3
Microbiology Lab Prac 3
| Question | Answer |
|---|---|
| Staphylococci | gram positive cocci; form clusters, produce catalase; facultative anaerobes, grow in presence of bile salts, and will grow in a 6.5% NaCl solution |
| Blood agar test | used to isolate pathogens; supplies many growth factors; hemolysin enzymes which degrade hemoglobin and lyse RBCs; different bacteria produce different hemolysins |
| Three types of hemolysis | beta hemolysis, alpha hemolysis, gamma hemolysis |
| Beta hemolysis | blood agar test; results in complete clearing around the colony |
| Alpha hemolysis | blood agar test; results in partial breakdown of RBCs; greenish color due to biliverdin spilling from RBCs |
| Gamma hemolysis | blood agar test; does not produce hemolysin, so does not lyse RBCs; no change indicated |
| Blood agar streak method | streak the plate and stab to provide area of lower oxygen concentration where streptolysins can more effectively break down RBCs |
| Mannitol salt agar | selective & differential; 7.5% salt conc. selects members of staph since they can tolerate high salt levels; contains sugar mannitol and pH indicator phenol red; fermentation of mannitol will turn agar yellow; most staph pathogens are fermenters |
| MSA agar contents | 7.5% salt; mannitol (sugar), phenol red (pH indicator) |
| Phenylethyl alcohol agar (PEA) | selective; cultivates Gram + organisms; alcohol inhibits growth of gram – organisms via DNA synthesis interference; prevents proteus species from swarming—beneficial for contaminated samples; growth = gram +; no growth = gram neg |
| Catalase | enzyme that breaks hydrogen peroxide (H2O2) into H20 and O2; bubbling will occur if evolution of O2 gas is present—indicates catalase activity; staph bubbles; strep does not |
| Staph labs | blood agar, mannitol salt agar (MSA), phenylethyl alcohol agar (PEA), catalase |
| Strep labs | 6.5% salt agar, bile esculin agar, bacitractin and optochin sensitivity |
| 6.5% salt tolerance | used to differentiate group D enterococci from Group D streptococci; group D enterococci are only streptococci that will grow in 6.5% NaCl broth; tryptic soy broth used |
| Bile esculin agar | selective and differential; used to presumptively ID enterococci and group D streptococci based on ability to hydrolyze esculin; contains oxgall (bile salts) to hinhibit growth of gram + organisms; contains nutrients, esculin, ferric citrate |
| Bile esculin agar results | when organisms hydrolyzes glycoside esculin to form esculetin and dextrose, esculetin reacts with ferric citrate to produce dark brown/black phenolic iron complex; media will turn dark brown/black if organisms can hydrolyze esculin |
| Bacitractin/optochin sensitivity tests | ID whether an organisms is susceptible/resistance to bacitratctin/optochin; blood agar plate streaked (lawn), paper disks infused w/ bacitractin/optochin applied to surface; sensitivity based on zone of inhibition (more clearance = more susceptible) |
| Bacitractin test | differentiates between beta hemolytic group A strep from beta hemolytic non group A strep b/c it is not effective on bacteria that do not have cell walls |
| Optochin | chemical that inhibits pneumococci but does not affect other alpha hemolytic streptococci |
| Gram negative bacteria labs | eosin methylene blue agar, hektoen enteric agar, MacConkey agar |
| Eosin methylene blue test (EMB) | fecal coliform bacteria; selective & differential; eosin Y, methylene blue inhibit gram + organisms; also indicator dyes -- form dark purple precipitate at low pH; sucrose, lactose—fermentable carbs; fermentation decreases pH so provides differentiation. |
| EMB outcomes | vigorous fermenters of lactose/sucrose—dark purple/blackish dye complex and growth; vigorous fermentation—green metallic sheen; slow/weak fermentation—mucoid pink; normal colored/colorless—organism does not ferment lactose/sucrose, so not fecal coliform |
| Hektoen Enteric Agar (HE) | selective & differential - salmonalla vs. shigella enterics; bile salts, dye bromthymol blue, acid fuchsin, inhibit gram +; lactose, sucrose, salicin encourage growth/differentiation of enterics; sodium thiosulfate—sulfur; ferric ammonium citrate |
| HE agar outcomes | enterics that ferment one + of the carbs lactose, sucrose, salicin, produce yellow to salmon colored colonies; non-fermenters have blue green colonies; organisms that reduce sulfur to hydrogen sulfide—black colonies/blue-green colonies w/ black center |
| MacConkey Agar (MAC) | selective and differential; separate enterics based on ability to ferment lactose; bile salts, crystal violet—inhibit growth of gram + organisms; lactose—fermentable carb; neutral red—pH indicator (turns yellow if acidic) |
| MAC results | lactose fermentation—acidic environment—pink b/c neutral red turns red; bile salts may also surround growth of fermenters; non-fermenters produce normally colored/colorless colonies |
| Kirby-Bauer test | antibiotic sensitivity |
Created by:
mbtrimm
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