Culture Media

Plated Culture Media:

Acronym Description Uninoculated state and Results

XLD Xylose Lysine Deoxycholate Agar1. Yellow colonies with black centre of

xylose fermenting and hydrogen sulphide producing Gram negative enteric bacilli like:

a. Citrobacter freundiib. Proteus mirabilisc. Proteus vulgaris

2. yellow colonies without black centre of xylose fermenting BUT non-hydrogen sulphide producing Gram negative bacilli like:

a. Escherichia colib. Serratia marcescensc. Klebsiella pneumoniaed. Enterobacter aerogenese. Providencia rettgeri

Highly selective and differential medium which is used to isolate and differentiate negative enteric bacilli, especially Salmo-nella and Shigella species based on xylose fermentation and lysine decarboxylation

inhibitors: deoxycholate and citratepH indicator: phenol redcarbohydrates: xylose, dextrose and su-croseH2S indicator: Ferric ammonium citrate with thiosulphateOther components: amino acid lysine

uninoculated

beta haemolytic dome-shaped colonies

pin head coloniesStaphylococcus aureus


BAP Blood Agar Platean enriched medium that can be used to iso-late both gram positive and gram negative cocci and bacilli. The enriching substance is preferably unheated 5% defibrinated sheep blood.

this is also classified as differential culture medium because it differentiates organisms based on their haemolytic patterns exhibited by the medium.

Alpha-haemolytic- if colony is surrounded by an incomplete zone of haemolysis wich is translucent and green in colour.Beta-haemolytic- if colony is surrounded by a complete zone of haemolysis which is clear and transparentGamma-haemolytic- if colony is not sur-rounded by any zone of haemolysis; there is not change in the surrounding medium

uninoculated

large gray mucoid colonies of gram nega-tive enteric bacilli

TCBS Thiosulphate Citrate Bile Salts Sucrose Agar- a highly selective medium which iso-

late and differentiate the sucrose ferment-ing from non-sucrose fermenting species of Vibrio.- inhibitors: thisulphate, citrate, bile

salts, and high pH- indicator: bromthymol bluecarbohydrate: sucrose

Uninoculated

yellow colours of sucrose fermenting Vibrio like Vibrio cholerae and Vib-rio alginolyticus


SSA Salmonella-Shigella Agara highly selective and differential medium which is used to differentiate species of Sal-monella and Shigella

inhibitors: brilliant green, bile salts, cit-ratepH indicator: neutral redhydrogen sulphide indicator: ferric am-monium citrate with sodium thiosulphatecarbohydrate: lactose uninoculated


MSA Mannitol Salt Agarboth selective and differential culture medium used to isolate and differentiate mannitol fermenting from non-mannitol fer-menting Staphylococcus

inhibitor: 7.5% NaClpH indicator: Phenol redcarbohydrate: mannitol

Pink colonies of non-mannitol fermenting Staph like:

Staphylococcus EpidermidisStaphylococcus lugdunensisStaphylococcus saprophyticus

Yellow colonies of mannitol fermenting Staph like Staphylococcus aureus

unioculated


BSA Bismuth Sulphite Agara highly selective medium for the isolation of Salmonella typhi

inhibitors: bismuth sulphite and brilliant green

carbohydrate: glucose

uninoculated

beta haemolytic dome-shaped colonies

pin head coloniesStaphylococcus aureus


CAP Chocolate Agar Platean enriched culture medium that can be used to grow both gram positive and gram negative cocci and bacilli. the enriching sub-stance is heated blood, preferably 5% defib-rinated sheep blood

uninoculated


EMB Eosin Methylene Blue Agarboth a selective and differential culture medium which is used to isolate and differ-entiate the lactose-fermenting from the non-lactose fermenting gram negative bacilli

inhibitors: eosin and methylene bluepH indicators: eosin and methylene bluecarbohydrate: lactose

Pink to purple with dark centre colonies (fish-eye) suggestive of Enterobacter aerogenes

purple with greenish metallic sheen, suggestive of Escherichia coli

uninoculated

colourless non-lactose-fermenting colonies on EMB plate.Salmonella typhi, Shigella dysenteriae, Providencia sp. Mor-ganella sp.


HEA Hektoen Enteric Agarboth selective and differential culture medium used to isolate and differentiate the lactose fermenting from non-lactose fer-menting gram begative enteric bacilli

inhibitors: bile saltspH indicator: bomthymol bluehydrogen sulphide indicator: ferric am-monium citrate with Na thiosulphatecarbohydrates: lactose, sucrose, salicin

orange colonies without black centre- lactose fermenting, non-hydrogen sul-phide producing gram negatice enteric bacilli like E. coli, K. pneumoniae and E. aerogenes

orange colonies with black centre- lac-tose fermenting, hydrogen sulphide pro-ducing gram negative enteric bacilli like Citrobacter freundii

uninoculated

MAC MacConkey Agarboth selective and differential culture medium which is used to isolate and differ-entiate the lactose fermenting from the non-lactose fermenting gram negative bacilli

inhibitor: crystal violet and bile saltspH indicator: neutral redcarbohydrate: lactose

Pink to red colonies of lactose ferment-ing gram negative enteric bacilli

yellow to colourless colonies of lactose fermenting Gram negative enteric bacilli

uninoculated


MHA RED PIGMENTED COLONIES OF Serratia marcscens in Mueller Hinton Agar

IMVC Test

Indole Test Set-up

Tryptone Broththe liquid medium is the substrate in the IN-DOLE TEST which is used to identify the gram negative enteric bacilli based on the ability of organisms to produce the enzyme tryptophase. this enzyme decomposes this tryptone broth producing indole that subse-quently forms a red coloured complex upon the addition of Kovac’s reagent or para-dimethyl amino benzaldehyde reagent.

positive: rednegative: yellow


Kovac’s reagentpara-dimethyl amino benzaldehyde reagent

Methyl Red The liquid medium is also known as peptone glucose broth and is used to identify the gram negative enteric bacilli used on the ability of the organisms to ferment glucose pyruvic acid by one of two pathways: mixed acid fermentation pathway, which is the Methyl red test and the butylene glycol path-way which is tested by the Vogues Proskauer test.

Set-up:if the organism would ferment glucose via the mixed acid fermentation pathway, more acids are produced like lactic, acetic, formic, succinic acids, which decrease the pH of the medium to 4.4 or lower, hence, upon the ad-dition of the indicator, methyl red the broth becomes red in colour. if the organism would not use this pathway, the pH of the medium increases to 5.5 or higher, ence upon the addition of the indicator, methyl red, the broth becomes yellow in colour.


Vogues Proskauer Test

if the organism would ferment glucose via the butylene glycol pathway, an intermediate product, acetyl methyl carbinol or acetoin which is neutral is converted to diacetyl upon the addition of the VP reagent. B (40% KOH with 0.3% creatine) in the presence of VP-reagent A (5% alpha-naphthol in ab-s.methyl alcohol). Diacetyl is red in colour. Negative is yellow in colour.

Coagulase Testplasma.

-differentiates pathogenic S. aureus from the non pathogenic S. epidermidis and sapro-phyticus.

slide test: plasma + specimen positive: visible clumping negative: no visible clumping

tube testplasma+ specimen positive: visibleclot or coagulation negative: no clot or coagulation

pathogenic S. aureus - positiveNon-pathogenic S. saprophyticus/epider-midis- negative

catalase test catalase test uses 3% h202. the positive re-action is indicated by bubbling or efferves-cence

positive: all Staph sp.all gram negative enteric bacilli except Shigella dysenteriae

negative: all strep sp.





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Culture Media