MacConkey EMB

Mac Conkey
Agar
Group Members
Soh Pek Qin SEQ070029
Ahmad Amzar bn Sohaimi SEQ070002
Norain Sahari
SEQ070022
Nurul Fitrah bt Mohd Ariffin
SEQ070023
Ang Yi Hui SEQ070038
Nurul Huda Ibrahim SEQ070024
Hidayah Abd Hamid SEQ070009
Function
inhibits the growth of Gram-positive
bacteria
differentiates or distinguishes
between lactose fermenting bacteria
and non-lactose fermenting bacteria
Identification of Gram-negative
enteric pathogens
Formula
Approximate Formula* Per Liter
Pancreatic Digest of Gelatin ..................................... 17.0 g
Pancreatic Digest of Casein ........................................ 1.5 g
Peptic Digest of Animal Tissue ................................... 1.5 g
Lactose ................................................................... 10.0 g
Bile Salts ................................................................... 1.5 g
Sodium Chloride ....................................................... 5.0 g
Agar ........................................................................ 13.5 g
Neutral Red ............................................................. 0.03 g
Crystal Violet .......................................................... 1.0 mg
pH 7.1 0.2
Pancreatic digest of casein

utilize by sample in culture to breakdown protein
casein in lactose
Peptic digest of animal tissue
utilize by sample in culture to breakdown peptone into

amino acid
Lactose
source of fermentable carbohydrate
Bile salt
inhibit most of gram positive bacteria
Sodium Chloride
maintain isotonic environment in the culture
Agar
medium for bacterial growth and activities
Neutral red
pH indicator
Crystal Violet
inhibit some gram positive bacteria
Peptone:
a source of fermentable protein
Direction
Suspend the powder in 1 L of purified
water MacConkey Agar 50 g;

2. Heat with frequent agitation and boil
for 1 minute to completely dissolve the
powder.
3. Autoclave at 121C for 15
minutes.NOTE: If MacConkey Agar Base
is to be usedwithin 12 hours,omit
autoclaving and gently boil medium for
5 minutes.
Add 1% carbohydrate before or after
autoclaving, depending upon heat

lability. The surface of MacConkey
agars without salt should be
thoroughly air-dried prior to
inoculation.
4. Test samples of the finished
product for performance using
stable, typical control cultures.
Results
Colour of Media Precipitation
Microorganis
m
Colour of
Colony
Escherichia coli ATCC

8739 *
red
red
Salmonella
typhimurium ATCC
14028
colourless
yellowish
Salmonella dublin
ATCC 15480
colourless
yellowish
Shigella sonnei ATCC

11060
colourless
yellowish
Proteus mirabilis
ATCC 29906
colourless
yellowish
How This Agar Function

Acting as a visual pH indicator,
By utilizing the lactose available in the medium, Lac+
bacteria such as Escherichia coli and Klebsiella will

produce acid, which lowers the pH of the agar below 6.8
and results in the appearance of red/pink colonies.
Non-Lactose fermenting bacteria such as Salmonella and
Shigella cannot utilize lactose, and will use peptone
instead. This forms ammonia, which raises the pH of the
agar, and leads to the formation of white/colorless
colonies.
Neutral red is a pH indicator that turns red at a pH below
6.8 and is colorless at any pH greater than 6.8.
MacConkey agar plates. Proteus produces colorless

colonies as shown in the left panel. E.coli produces
bright pink colonies as shown in the right panel.
Klebsiella pneumoniae ferments lactose and produces pink colonies on

MAC.Micrococcus luteus does not grow in the presence of bile salts and crystal violet
Salmonella typhimurium does not ferment lactose and

produces colorless colonies. Pseudomonas aeruginosa also
does not ferment lactose, but since it normally produces a
diffusible blue-green, water-soluble pigment, its colonies
do not appear colorless.
FIG. 29. Example of mixed fecal flora at 24 hours. One colony type of lactose fermentor
and two of non-lactose fermentors, none of which were identified as typical causes of
gastroenteritis. (Rebecca Buxton, University of Utah)
FIG. 11. MacConkey agar plate inoculated with Serratia marcescens, Escherichia coli
(showing the pink halo) and the weak lactose fermenter Enterobacter aerogenes. (Mary Allen,
Hartwick College
Eosin Methylene Blue

Agar
Function
identification and isolation of Gram-
negative enteric rods.

inhibits the growth of Gram-positive
organisms
distinguish between lactose fermenting
and non-lactose fermenting organisms
Formula
Enzymatic Digest of Gelatin...........................10 g
Lactose...........................................................10 g
Dipotassium Phosphate................................... 2 g
Eosin Y...........................................................0.4 g
Methylene Blue..........................................0.065 g
Agar................................................................15 g
Final pH: 7.1 0.2 at 25C
Enzymatic Digest of Gelatin

is the nitrogen source in EMB Agar. Lactose and Sucrose are the
fermentable carbohydrates.
Dipotassium Phosphate
buffer.
Eosin Y and Methylene Blue

dyes that combine to form a complex at an acid pH.
inhibit grampositive bacteria to a limited degree.
differentiating between lactose fermenters and lactose nonfermenters
due to the presence or absence of dye uptake in the bacterial colonies
Lactose
source of fermentable carbohydrate
Agar
solidifying agent.
Direction
1. Suspend 37.5 g of the medium in
one liter of purified water.

3. Heat with frequent agitation and
boil for one minute to completely
dissolve the medium.
4. Autoclave at 121C for 15
minutes.
Results
Colonies of lactose fermenters are blue-black
with or without a green metallic sheen. E. coli

colonies typically are dark centered and
usually have a green metallic sheen. Colonies
of non-lactose fermenting bacteria are
colorless and translucent. Refer to appropriate
references for specific results and biochemical
MICROORGANISM
RESPONSE
REACTIONS
reactions.
Enterobacter aerogenes
Growth
Blue-black colonies with black
center
Enterococcus faecalis
Partial inhibition
Colorless, pinpoint colonies
Escherichia coli
Growth
Blue-black colonies, with

green metallic sheen
Pseudomonas aeruginosa
Growth
Colorless, may have irregular

colonies
Salmonella typhimurium
growth
colorless
FIG. 1. Eosin-methylene blue (EMB) agar plate inoculated with Escherichia coli (a gramnegative coliform bacterium) showing good growthof dark blue-black colonies
with metallic green sheen indicating vigorous fermentation of lactose and acid production
which precipitates the green metallic pigment.
FIG. 2. EMB agar plate inoculated with Enterobacter aerogenes (a

gram-negative coliform bacterium) showing good growthof brown,
dark-centered, mucoid colonies indicating lactose fermentation and
acid production. (Naowarat Cheeptham, Thompson Rivers University)
FIG. 17. EMB agar plate inoculated with (A) Escherichia coli, (B) Pseudomonas
aeruginosa, (C) Klebsiella pneumoinae, and (D) Enterobacter aerogenes. All four gram
negative bacteria grew exhibiting different morphology. Escherischia coli grew with
typical lactose fermenter morphology with excessive acid production and precipitation of
green metallic pigment (colonies with green metallic sheen). Pseudomonas aeruginosa
grew exhibiting the nonfermenter morphology (pinkish colonies), both Klebsiella
pneumoniae and Enterobacter aerogenes grew with lactose fermentation and acid
production morphology (with purple dark centered mucoid colonies). (Naowarat
Cheeptham and Carolynne Fardy, Thompson Rivers University)
How This Agar Function

By utilizing the lactose available in the medium, Lac+
bacteria such as Escherichia coli and Klebsiella will
produce acid, which lowers the pH of the agar below
6.8 and results in the appearance of dark colour
colonies.
Non-Lactose fermenting bacteria such as Salmonella
and Shigella cannot utilize lactose, and will use
peptone instead. This forms ammonia, which raises the
pH of the agar, and leads to the formation of
white/colorless colonies.
Eosin Y and methelyne blue is a pH indicator that turns
dark at a pH below 6.8 and is colorless at any pH
greater than 6.8.
References
http://en.wikipedia.org/wiki/macconkey_agar
http://www.mpbio.com/product_info.php?products_id=10060
http://www.austincc.edu/microbugz/html/eosin_methylene_blue_agar.html
http://medic.med.uth.tmc.edu/path/emb.htm
http://classes.tmcc.edu/biolabs/EMB.pdf
www.austincc.edu/microbugz/html/macconkey_agar.html
www.microbelibrary.org/ASMOnly/details.asp?id=1976&Lang
www.microbelibrary.org/ASMOnly/details.asp?id=2022&Lang
medic.med.uth.tmc.edu/path/macconk.htm

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Mac Conkey

Pancreatic digest of casein

Peptic digest of animal tissue

utilize by sample in culture to breakdown peptone into

source of fermentable carbohydrate

inhibit most of gram positive bacteria

maintain isotonic environment in the culture

medium for bacterial growth and activities

inhibit some gram positive bacteria

water MacConkey Agar 50 g;

Add 1% carbohydrate before or after

autoclaving, depending upon heat

Escherichia coli ATCC

Shigella sonnei ATCC

How This Agar Function

bacteria such as Escherichia coli and Klebsiella will

MacConkey agar plates. Proteus produces colorless

Klebsiella pneumoniae ferments lactose and produces pink colonies on

Salmonella typhimurium does not ferment lactose and

Eosin Methylene Blue

negative enteric rods.

Enzymatic Digest of Gelatin

Eosin Y and Methylene Blue

due to the presence or absence of dye uptake in the bacterial colonies

source of fermentable carbohydrate

one liter of purified water.

with or without a green metallic sheen. E. coli

Colorless, pinpoint colonies

Blue-black colonies, with

Colorless, may have irregular

FIG. 2. EMB agar plate inoculated with Enterobacter aerogenes (a

How This Agar Function

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