Microbiology Lab Tests Summary USTMED ’07 Sec C AsM (
[email protected] [email protected]) TEST
HOW
TSI
MEDIUM
Fermentation of sugar
LIA
Decarboxylation of amino acid
PRESENT
TSIA (3 sugars, Fe, agar)
LIA (lysine, Fe, agar)
Glucose, lactose, sucrose
Lysine (ornithine, arginine)
PH INDICATOR
OBSERVE
Phenol red
Sugar fermentation
Bromcresol purple
H2S production Gas production Deamination (slant)
RESULTS
Lys decarboxylation (butt) MIO
Citrate utilization test Urease test
Diffuse growth, indole production, decarboxylation of amino acid
MIO media
Ornithine, indole (if tryptophanase is present
Bromcresol purple, kovac’s reagent (HCl, dimethyl aminobenzaldehyde)
Use citrate as an inorganic carbon source
Simmon’s citrate agar (SCA)
Sodium citrate
Bromthymol blue
Hydrolyze urea
(Stuart) urea broth or Christensen’s agar
urea
Phenol red
motility Indole production Ornithine decarboxylation (butt) citrate→ammonia, change in color of medium (sign of citrate utilization) Production of NH4 and CO2 (H2O)
Alkaline butt/slant – red (-) Acid butt/slant – yellow (+) Black butt Crack, bubbles, displacement Alkaline - purple Acid/red - red Alkaline – purple (+) Acid – yellow (-) Diffuse growth vs. growth along stabline (+) red ring w/ Kovac’s Alkaline – purple (+) Acid – yellow (-) Blue slant – (+) Green slant – (-) Fuschia pink (+) Salmon pink (-)
NOTES
Due to a.a. decarboxylation
Streak slant Stab butt Portal of entry is also portal of exit; stab up to the bottom Don’t shake (semisolid) Read motility and decarboxylation before Kovac’s Bacteria are called autotrophs Soil bacteria don’t have this ability Media becomes alkaline
Agar TYPE
PURPOSE
CONTENTS
INDICATOR
MacConkey Agar
g (-), LFO
Bile salts & lactose
Neutral red, crystal violet
Salmonella Shigella agar
Inhibits normal flora coliforms, differentiation of stool pathogens
Bile salts, brilliant green agar, lactose, ferric ammonium citrate
Neutral red, sodium thiosulfate
Xylose, Lysine, Deoxycholate (XLD) Agar
Inhibits gram (+), selective for E.coli (?), inhibits proteus swarming
Xylose, lysine, ferric ammonium citrate
Hektoen Enteric Agar
Inhibits normal flora of lower GI tract
Bile salts, lactose, sucrose, starch & salicin, ferric ammonium citrate
Eosin Methylene Blue Agar (EMB)
G(-), LFO
Phenol red, sodium thiosulfate
OBSERVE
Acid – pink colonies(+) LFO Alkaline – colorless colonies (-) NLFO
H2S production
FeS (+) black centered color Normal flora – pink (salmonellashigella) Pathogens - colorless Acid- yellow (+) Alkaline – red (-) Lysine Decarboxylase Black centered colonies
fermentation Lysine decarboxylase H2S production
Lactose, Eosin & Methylene blue
Bromthymol blue, sodium thiosulfate
Eosin & methylene blue
REACTION
Fermentation (produces acid) bile salts precipitation fermentation
Acid – pink/red colonies(+) LFO Alkaline – colorless colonies (-) NLFO
fermentation
Green (-) NLFO alkaline Orange/yellow (+) LFO acid
H2S production
Black centered colonies
fermentation
Purple – LFO Green metallic sheen – E.coli (Rapid) Colorless colonies - NLFO
NOTES
Brown pH 6.8-8.0 Red pH < 6.8 Bile salts also inhibits g(+) Bile salts Inhibits gram (+) bacteria; brilliant green inhibits gram (-) coliform; ferric ammonium citrate + sodium thiosulfate produces H2S Fermentation is the first reaction Red colonies Enterobacteriaceae are xylose (+) except shigella pH > 7.6 blue pH ranges 6.0-7.6 green pH < 6.0 yellow Enterobacteriaceae are xylose (+) except shigella Purple color is precipitated eosin & methylene blue absorbed when lactose fermented
