Microbiology of Differentiate Media Flashcards

 Selective • Select for specific types of bacteria - Chemicals inhibit one group but allow another to grow • PEA agar, CV agar, NaCl agar  Differential • Distinguish among morphologically & biochemically related groups of orgs. - specific characteristics on media • MSA, MacConkey, EMB  Enriched • Rich media for fastidious orgs. - Contains blood, serum, or yeast extract • Blood agar, chocolate agar

 Selects for G+ orgs • Inhibits most G- orgs. • Must compare growth to a control plate (EMB, MAC, BHI or TSA)  Results – • Growth compared to control = (+) • No growth or Inhibited growth = (-) NA PEA

 Differential & selective • Selects for halophiles with high [salt] – 7.5% NaCl • Differentiates based on ability to ferment mannitol  pH indicator = phenol red • Yellow = acid production = + fermentation  Results • Growth or no growth (inhibited) • A= or Ahttp

 Selective • Selects for Gram (-) – Crystal violet & bile salts inhibit G(+)  Differential • Ability to ferment lactose • pH indicator = neutral red, turns pinkish/red when pH < 6.8  Results – Compare to PEA for control • Orgs that ferment lactose  red growth (Differential) • Non-fermenters produce non-colored colonies • Inhibited/No growth = (-) (Indicates G+ organism)

 Selective & differential media • Eosin & MB inhibit G+ bacteria • Differentiates based on ability to ferment lactose &/or sucrose – dyes interact to change the color of growth • Used for isolation of fecal coliforms  Results: • Inhibited / no growth = (-) Indicates G+ organism • Good growth = indicates G- organism - Green sheen / black = vigorous fermenter (E.coli or C. freundii) - Pink/Purple = less aggressive lactose fermenter (Enterobacter or Klebsiella) - No color (creamy/white/same color as agar) = non lactose fermenter or possible sucrose fermenter

Two tubes – • One covered w/ mineral oil to promote anaerobic growth & fermentation • Mineral oil prevents oxidation  Results • Ferment & oxidize - sealed & unsealed  yellow throughout • Oxidize only - unsealed broth  yellow; sealed staying green • Slow or weak fermenters  both tubes slightly yellow in top • Non-saccharolytic  both green or blue (strictly aerobic)

Conversion of sugar to acid, alcohol & gas (CO2)  Phenyl red broth • PR –ose  Phenyl red = pH indicator • Acid = yellow • Neutral = red • Alkaline = hot pink

CHO fermentation = yellow (acidic) = A+  CO2 production trapped in Durham tube = Gas +  Hot pink = degradation of proteins/AA NH3 (alkaline) = K+ or P+

Methyl Red & Vogues Proskauer Tests  Low acid fermentation product is maintained by E.coli • E.aerogenes converts acids to non-acidic end products (acetoin & 2,3-butanediol) which raise pH to ~6

After growth place split into two tubes • To one tube add ~4-5 drops MR (+ if turns red) • To other tube add 15 drops VP-A (Barritt’s) & 5 drops VP-B – wait 60 minutes

Presumptive test - slide test • Drop DI H2O on one side & drop of peroxide on the other • Add 1 drop of culture to each spot - If bubbles = (+) for catalase  Confirming test • Add 1-5 drops H2O2 to culture growing on slant - (+) = bubbles

Use wood end of cotton applicator place on DrySlide – read at 20 seconds  Add 1-3 drops oxidase reagent to slant • (+) = purple/black

Tests for nitrate reductase  Air bubble in Durham tube indicates gas production = denitrification  If a non-fermenter test is complete = NO3  NO2  N2  If no gas or a fermenter add reagents • Add 8 drops Nitrate A & B reagent  Results • Immediate red = (+) • If colorless add zinc - Remains colorless = (+) - Turns red = (-)

Tests for citrate permease • Citrate as only C source • pH indicator = bromthymol blue - Green pH 6.9 and blue at pH > 7.6  Inoculate w/ needle (light inoculum) • (+) if media turns royal blue • Growth no color change = (+) (rare – make sure not heavy inoculum) • No growth / No color change = (-)

Mineral oil promotes fermentation  Decarboxylation of AA  alkaline products (purple) = +  If a glc fermenter but does not have decarbixylase = yellow (-)  No color change = (-)