Microbiology: Culture Media and Indicators

5-10% sheep or horse; no human blood; shows hemolysis; BA + CNA

RBC broken by heating 85 C; nutrients available; fastidious bacteria; CHOC + NYC

show acid or alkaline by colour; neutral red + phenol red + bromthymol blue

detects O2; thioglycollate broth; resazurin pink in O2

inhibit some organisms; heat stable; added before autoclave; bile salts + desoxycholate + crystal violet + brilliant green + 6-7.5% salt

heat sensitive; added AFTER autoclave + cooling; aseptic; CNA + NYC

control pH; peptones + phosphates

absorb toxins; may inhibit fastidious; transport media + BCYE

water + agar + simple protein; not enriched; no selective or differential; TSA + nutrient + BHI + Columbia + Mueller Hinton

basic + extra nutrients (blood + vitamins); fastidious organisms; extra delicacies

inhibitors (antibiotics + dyes); some grow; others blocked

tell organisms apart; visible biochemical difference

liquid broth; suppresses normal flora briefly; target pathogen grows; subculture within 12-18 h

basic; susceptibility testing; non-fastidious; starch absorbs toxins; loose agar for diffusion; low thymidine + thymine; add 5% blood for fastidious

enriched + differential; 5-10% sheep blood; hemolysin makes clear zone

enriched; NOT differential; heat-lysed blood; brown; 2% hemoglobin; fastidious Haemophilus

enriched + differential broth; grows all O2 types; hemin + vitamin K; O2 gradient; 0.075% agar; resazurin pink in O2

NO; general-purpose enriched + differential; does not suppress normal flora

enriched + selective + differential; colistin + nalidixic acid; Gram + and yeast grow; hemolysis

selective + differential; bile salts + crystal violet; lactose + neutral red; LF pink; NLF colourless

LF can revert pink to colourless when lactose used up; check 4th quadrant; mucoid colonies look NLF but are LF

selective + differential; 7.5% salt; mannitol + phenol red; S. aureus yellow; other Staph pink

yellow = acid; pinked = alkaline

selective + differential; sorbitol instead of lactose; EHEC = colourless; other E. coli = pink

selective + differential; enteric pathogens; bile salts + citrate + brilliant green; lactose + neutral red; thiosulfate + ferric ammonium citrate for H2S

thiosulfate gives sulfide; reacts with ferric ions; black precipitate; black colony centre

pink = LF; clear = NLF; black centre = H2S

selective + differential; enteric pathogens; bile salts; lactose + sucrose + salicin; bromthymol blue + acid fuchsin; H2S black

fermenters = salmon/orange/yellow; non-fermenters = greenlue-green; black centre = H2S

selective + differential; Yersinia enterocolitica; cefsulodin + irgasan + novobiocin; mannitol + neutral red; bulls-eye colony

selective + enriched; Campylobacter; 10% blood; vancomycin + amphotericin B + polymyxin B + trimethoprim + cefoperazone; sodium bisulfite lowers O2

selective + enriched; Neisseria gonorrhoeae + meningitidis; lysed horse blood; vancomycin + colistin + amphotericin B + trimethoprim

selenite; tetrathionate; BHI broth; cooked meat

brilliant green; selenium salts; tetrathionate

enrichment; feces; Salmonella

enrichment; anaerobes; overgrown by normal flora

basic = TSA + MH; enriched = BA + CHOC; selective = CNA + MAC + MSA + SMAC + SS + HEK + CIN + Campy + NYC; differential = BA + CNA + MAC + MSA + SMAC + SS + HEK + CIN + thio; enrichment = selenite + tetrathionate

mycolic acid; lipid + wax cell wall; resist acid alcohol; poor Gram stain; Mycobacterium + Nocardia + Cryptosporidium + Cyclospora

beaded; or unstained ghosts

auramine O or auramine-rhodamine; binds mycolic acid; 0.5% acid-alcohol; permanganate quencher; yellow/orange glow on dark background

Kinyoun + Ziehl-Neelsen; carbolfuchsin; phenolic

cold method; more phenol; 3% acid alcohol

uses heat/steam; carbolfuchsin; modification of Kinyoun

smear + heat fix; carbolfuchsin; filter paper; steam 10 min; cool; rinse; acid alcohol 15 sec; wash; methylene blue 2-3 min

acid-fast = red; others = blue

binds nucleic acid; orange; no cell wall organisms; Mycoplasma

binds cellulose + chitin; fungal walls; screens respiratory specimens

fermentation = no O2 + lots of acid; oxidation = O2 + little acid; asaccharolytic = no acid

change below pH 6; methyl red

weak acid indicators; bromthymol blue

glucose first; then lactose; needs lac operon enzymes

oxidative or fermentative or neither; separates non-fermenting aerobic GNB

1% glucose; peptone; bromthymol blue; semi-solid agar; green = no acid; yellow = acid

2 tubes; open = aerobic; closed = 1 cm mineral oil = anaerobic; 35 C 48 h

fermenter = both yellow; oxidizer = open yellow only; non-fermenter = both green; peptone use = blue (alkaline)

lactose to glucose + galactose; needs permease + beta-galactosidase

beta-galactosidase but no permease; makes permease later after mutation

per slides = permease but no beta-galactosidase

detects beta-galactosidase; separates LLF from NLF

looks like lactose; enters without permease; colourless to yellow o-nitrophenol; any yellow = positive

glucose + lactose + sucrose fermentation; gas; H2S; Gram-negative bacilli; enteric screening

peptone; glucose 0.1%; lactose 1%; sucrose 1%; phenol red; ferrous ammonium sulfate or ferric ammonium citrate + thiosulfate

uninoculated = orange; acid = yellow; alkaline = pink

0.1% glucose used up; slant uses peptones + makes ammonia = pink; butt stays yellow; NA/A

lots of acid; not used up in 18-24 h; no reversion; A/A

slant first then butt; slant = aerobic; butt = anaerobic; record gas + H2S only if seen

bubbles; cracks; lifting of agar; CO2 + H2

thiosulfate sulfur; ferrous sulfide; black; in butt; needs glucose fermentation

yellow/yellow; glucose + lactose and/or sucrose

pink/yellow; glucose only

pink/orange; no sugar fermented; aerobe using peptones

slantutt then H2S then gas; A/A H2S gas = A/A gH2S

mixed acid pathway = MR; 2-3 butanediol pathway = VP

mixed acid; lactic + acetic acid; pH below 4.2; methyl red; RED = positive; yellow = negative

glucose to pyruvate to acetoin; alpha-naphthol FIRST + 40% KOH second; shake for oxygen; red = positive

6 drops 5% alpha-naphthol; 2 drops 40% KOH

remove carboxyl group from amino acid; makes amine + CO2

lysine to cadaverine

ornithine to putrescine

detect decarboxylase enzyme; Gram-negative bacilli

glucose; amino acid (lysine or ornithine); bromocresol purple

glucose fermented; acid yellow; acid needed to switch enzyme on; amine alkaline; yellow to purple

3 per organism; control + lysine + ornithine

glucose only; must be YELLOW; shows growth + fermentation; purple = invalid; repeat

purple; any purple after 24 h; yellow control

bright yellow

control purple = invalid; test purple + clear = repeat; not inoculated or no amino acid

yellow = acid; purple = alkaline

remove amine group; deaminase; makes ammonia

phenylalanine deaminase; Gram-negative bacilli

L-phenylalanine; NaCl; agar; no glucose

phenylpyruvic acid + 10% ferric chloride; GREEN = positive; yellow = negative

citrate as only carbon source; O2 needed; Gram-negative bacilli

sodium citrate; ammonium salts; bromthymol blue

uses citrate + ammonia; alkaline; blue

growth + blue = positive; growth only = positive; no growth + green = negative

tryptophanase; peptone supplies tryptophan; tryptophan to indole + pyruvic acid + ammonia

Kovac's or Ehrlich's; p-dimethylaminobenzaldehyde; red ring = positive; colourless = negative

tube = benzaldehyde + red; spot = cinnamaldehyde + green/turquoise; spot works in seconds to 3 min on colonies

H2S + indole + motility; Gram-negative bacilli

tryptophan; ferrous ammonium sulfate; thiosulfate; 0.5% agar semi-solid

black precipitate; ferrous sulfide

motile = growth away from stab or turbid throughout; non-motile = along stab only; read BEFORE Kovac's

nitrate reductase; anaerobic ATP

peptone; KNO3; Durham tube

nitrate to nitrite; then maybe nitrogen gas; sulfanilic acid + dimethyl-alpha-naphthylamine; red = nitrite

gas in Durham (non-fermenter) = N2 positive; red = nitrite positive; no colour = none or beyond nitrite; add zinc

red after zinc = nitrate left = NEGATIVE; no colour = reduced beyond nitrite = N2 POSITIVE; too much zinc = false positive

thick peptidoglycan; teichoic + lipoteichoic acids; no outer membrane

thin peptidoglycan; outer membrane; LPS (lipid A + core + O antigen); porins; periplasmic space

Gram primary stain; all cells purple

mordant; fixes crystal violet complex

acetone/alcohol; strips outer membrane; Gram-negative loses colour

counterstain; Gram-negative pink

Gram positive = purple; Gram negative = pinked

Gram-negative bacilli

Gram-positive bacilli; Bacillus + Clostridium

Neisseria; Gram-negative diplococci