HAL015
Harlequin™ Chromogenic Agar for Salmonella Esterase (CASE)
Description
CASE is a selective chromogenic agar for the detection of Salmonella. It utilises a dual chromogenic
system to differentiate between Salmonella and non-target organisms that grow on the agar. The
first chromogen is a target for esterase activity present in Salmonella species. Utilisation of this
chromogen results in blue/green colonies. The second chromogen is a target for ß-glucosidase
present in non-target organisms that are able to grow on the agar and also possess esterase
activity. Utilisation of the second chromogen masks the utilisation of the first resulting in black
colonies in non-Salmonella. All other organisms are either inhibited or grow colourless on the agar.
The media is able to detect non-motile Salmonella (S.pullorum and S.gallinarum) as well as
monophasic variants (1,4,[5],12:i:-). The media is also able to detect serovars that present weak
esterase activity (S.dublin), and lactose positive strains (S.arizonae).
Typical Formulation
Growth Mix
Selective Mix
Buffer
Opacity Agents
Chromogenic Mix
Agar
Per litre
10.5g
11.0g
7.0g
7.5g
1.4g
12.5g
Appearance
Powder: Free flowing, white/cream
Finished medium: Opaque, Off White Gel
pH: 7.2 ± 0.2
Hazard classification
Refer to appropriate SDS
Method for reconstitution
Disperse 49.9g of powder in 1 litre of deionised water. Allow to soak for 10 minutes and swirl to
mix. Bring rapidly to the boil with frequent agitation and temper in a water bath to 50°C. Mix well
and dispense into sterile Petri dishes. Dry agar surface prior to use.
Media must sufficiently boil or white precipitate will be seen in the agar. This is purely aesthetic,
does not affect performance and can be avoid by sufficiently boiling the media. Extended holding
time after sterilisation can cause a loss of selectivity and similar aesthetic issues.
Storage
Store at 2-8oC in the dark for up to 14 days. Any increase in storage time must be validated by the
user.
HAL015_01
Inoculation
Streak for single colonies after selective enrichment.
Incubation
Incubate at 37±1°C for 18-24 hours.
Minimum QC Organisms
Salmonella typhimurium ATCC 14028
Salmonella enteritidis ATCC 13076
Enterobacter aerogenes ATCC 13048
Pseudomonas aeruginosa ATCC 27853
Good Growth, Blue/Green Colonies
Good Growth, Blue/Green Colonies
Growth, Black Colonies
Inhibited
Limitations
Some infrequent Salmonella isolates, typically avirulent environmental organisms that express ßglucosidase activity may produce a black colony and thus a false negative. Whilst CASE either
inhibits or masks any non-target organisms that produce esterase some isolates may result in a
blue/green colour, but with irregular morphology. All isolates can be confirmed by appropriate
biochemical and serological methods.
Interpretation
Organism
Growth characteristics
Growth
Salmonella spp.
Good Growth
Blue/Green
Growth
Black
Escherichia coli
Supressed
Colourless
Shigella spp.
Supressed
Colouress
Proteus spp.
Supressed
Colourless to Brown
Enterobacter spp. Klebsiella spp
Colour
References
Cooke. V.M. et al, A Novel Chromogenic Ester Agar Medium for Detection of Salmonellae. Appl.
Environ. Microbiol. Feb 1999 p. 807-812, Vol. 65, No. 2.
HAL015_01