Lactose TTC Agar with Tergitol

Lactose TTC Agar with Tergitol^® 7

Cat. No. 1.07680.0500
(500 g)

Selective differential medium for the detection and enumeration of E. coli und coliform bacteria in water using the membrane filtration method.

The medium complies with the recommendations of the ISO 9308-1 (1988) and the AFNOR norm NF 90-414 (1985) Water quality – Detection and enumeration of E. coli and coliform bacteria – Membrane filtration method.

Mode of Action	Quality Control
Typical Composition (g/liter)	Additives/Reagents
Preparation	Picture/Image
Experimental Procedure and Evaluation	Literature

Mode of Action
Degradation of lactose to acid is indicated by the pH indicator bromothymol blue, which changes the color of the medium under the membrane to yellow. Selectivity is achieved by the use of sodium heptadecylsulfate (Tergitol^®7) and 2,3,5-Triphenyltetrazoliumchloride (TTC) to inhibit most Gram-positive bacteria. TTC is also part of the differential system. The reduction of TTC by lactose-negative bacteria produces dark red colonies. Lactose-positive E. coli and coliform bacteria reduce TTC weakly; hence their colonies are yellow-orange.

Typical Composition (g/liter)
Lactose 20.0; peptone 10.0; yeast extract 6.0; meat extract 5.0; bromothymol blue 0.05; Tergitol^®7 0.1; agar-agar 12.7.
Additive: TTC 0.025.

Preparation
Suspend 53.9 g in 1 liter of purified water, dissolve and autoclave (121°C, 15 min). Cool the medium in a water bath to 45-50°C, add 5 ml of a sterile filtrated 0.05 % aqueous solution of TTC to 100 ml basal medium. Mix homogeneously and pour the medium into Petri dishes. The agar layer should have a height of at least 5 mm.
pH: 7.2 ± 0.2 at 25°C.
The prepared medium is clear and green.
The TTC-solution and the medium is stable for 1 week when stored at +2° - +8�°C and protected from light.

Experimental Procedure
Detailed instructions on titer determination are contained in APHA: Standard Methods for the examination of Water and Wastewater (1998).
The type of membrane filter affects the performance of the medium. The best results were obtained using cellulose-nitrate filters, e.g. from Sartorius (order no. 13906-47-ACN).
After filtration the filter is transferred, under aseptical conditions, to the agar surface.
Incubation: 21 ± 3 hours at 36 ± 2°C.

Evaluation
Lactose-positive bacteria produce yellow-orange colonies and under the membrane yellow-orange halos. The count of these typical colonies is considered to be presumptive coliform bacteria count.
Confirmation of coliform and E. coli count requires further subculture of typical colonies on a non selective agar (e.g. Tryptic Soy Agar) and Tryptophan broth, respectively.
Colonies that are oxidase negative are considered to be coliform bacteria. Coliform bacteria that form indole from tryptophane at 44 ± 0.5°C within 21 ± 3 hours are considered to be E. coli.

Quality control with the membrane filtration method

Test strains	Growth	color of medium (under membrane)	Colony color	Oxidase	Indole (44°C)
Escherichia coli ATCC 25922	+	yellow	yellow-orange	-	+
Citrobacter freundii ATCC 8090	+	yellow	yellow-orange	-	-
Pseudomonas aeruginosa ATCC 27853	+	blue	red	+	-
Bacillus cereus ATCC 11778	- / poor

Additives

EMD Cat. No.	Product	Pack Size
1.05458.0500	Tryptic Soy Agar (Casein Peptone Soymeal Peptone Agar)	500 g, 5 kg
TX1510-3	2,3,5-Triphenyltetrazolium chloride	10 g
1.09293.0100	KOVACS Indole Reagent	100 ml
1.10694.0500	DEV-Tryptophan-Broth	500 g
1.11350.0001	Bactident^® Indole (dropper bottle)	1 x 30 ml
1.13300.0001	Bactident^® Oxidase	50 test strips

E.coli

Literature

CHAPMAN, G.H. 1947. A superior culture medium for the enumeration and differentiation of coliforms. J. Bact. 53: 504 T (1947).
KULP, W., MASCOLI, C., TAVSHANJIAN, O. 1953. Use of tergitol-7 triphenyl tetrazolium chloride agar as the coliform confirmatory medium in routine sanitary water analysis. Am. J. Publ. Hlth. 43: 1111-1113 (1953).
POLLARD, A.L. 1946. A useful selective bactericidal property of Tergitol-7. Science 103: 758-759.AE