CHROMOGENIC MEDIA

DEHYDRATED CULTURE

FOR MICROBIOLOGY

DEHYDRATED CULTURE

CHROMOGENIC MEDIA FOR FOODS & WATERS

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TBX CHROMOGENIC AGAR

(TRYPTONE BILE X-GLUCURONIDE) (ISO 16649-2:2001) Cat: 1151
Selective medium for the detection and Enumeration of E.Coli in foods. FORMULA IN GRAMS PER LITRE
Casein Peptone x--D-Glucuronide 20.0 0.075 Bile Salts.1.50 Bacteriological Agar.15.00

Final pH 7.2 0.2 at 25C

Preparation
Suspend 36.6grams of the medium in one liter of distilled water. Mix well. Heat with frequent agitation and boil for one minute. Dispense into appropriate containers and sterilize at 121C for 15 minutes.

Uses
TBX Agart is based on the tryptone Bile Salts Agar medium to detect and enumerate E.coli in foods, with the addition of a chromogenic agent, x--D-Glucuronide, to detect the presence of the enzyme glucuronidase, which is highly specific for E.Coli. The released chromophore in TBX Agar is insoluble and colored target colonies are easily identified. E. coli absorbs the chromogenic agent x--D-glucuronide, and the intracellular glucuronidase enzyme activity breaks the bond between the chromophore and the glucuronide. The released chromophore is colored and builds up within the cells, causing the E. coli colonies to be blue-green colored. Casein peptone provides essential nutrients for growth: nitrogen, vitamins, minerals and amino acids. Bile Salts are inhibitors to other Gram-positive organisms and suppress coliform bacteria. Bacteriological agar is the solidifying agent. The standard ISO 16649 specifies a horizontal method for the enumeration of -glucuronidase positive E. coli in products intended for human consumption or for the feeding of animals. It uses a colony-count technique at 37 and 44C on a solid medium containing a chromogenic ingredient for the enzyme -glucuronidase. Inoculate and incubate at a temperature of 37 and 44C for 18 24 hours. Note: The negative E. coli b--glucuronidase colonies are colorless, e.g. E. coli O157:H7. The high temperatures (44C) inhibit the growth of E. coli O157:H7.

Escherichia coli ATCC 25922

Bibliography
International Standard ISO 16649-2:2001 Microbiology of food animal feeding stuffs. Horizontal method for the enumeration of presumptive Escherichia coli.

MICROBIOLOGICAL TEST
The following results were obtained in the performance of the medium from type cultures after incubation at a temperature of 37C and 44C and observed after 18-24 hours

Micro-organisms 44C Escherichia coli ATCC 25922 Salmonella typhimurium ATCC 14028 Streptococcus faecilis ATCC 19433 Klebsiella pneumoniae ATCC 13883 Good

Growth 37 Good Inhibited Inhibited Inhibited

Colony Color

Blue-green Colorless --Colorless

Inhibited Inhibited Inhibited

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Dehydrated Culture Media

E.COLI-COLIFORMS CHROMOGENIC MEDIUM

Cat: 1340
Selective medium for the simultaneous detection of E. coli and other Coliforms in water and food samples. FORMULA IN GRAMS PER LITER
Sodium Chloride.. Bacteriological Peptone Tryptophan. Chromogenic Mixture Bacteriological Agar 5.00 3.00 1.00 0.36 10.00 Phosphate Buffer.4.90 Sodium Pyruvate..1.00 Sorbitol..1.00 Tergitol -7..0.10

Final pH 6.8 0.2 at 25C

Preparation
Suspend 26.4 grams of the medium in one liter of distilled water. Heat with frequent agitation and boil for one minute. AVOID OVERHEATING. DO NOT AUTOCLAVE. Dispense into Petri Dishes. Store the plates in the refrigerator, protected from the light.

Uses
E.COLI-COLIFORMS CHROMOGENIC MEDIUM is a selective medium for the detection of E.coli and other Coliforms in waters and foods. The interaction of ingredients in the medium, such as peptone, sorbitol, etc, grants a quick colony growth, including infectious Coliforms. Tergitol-7 inhibits Gram-positive bacteria. Sodium chloride maintains the osmotic balance and the Phosphate is the buffer. Bacteriological agar is the solidifying agent. The chromogenic mixture contains chromogenic substrates as Salmon-GAL and X-glucuronide. Coliform enzymes produced, such as galactosidase and glucuronidase, cleave these substrates, resulting in the different coloration of certain bacteria colonies. The -D-galactosidase cleaves Salmon-GAL substrate, and gives a salmon to red color to the coliform colonies. The -Dglucuronidase cleaves both substrates Salmon-Gal and X-glucuronide, giving a dark blue to violet color to the colonies, easily distinguishable from other coliform colonies that have a salmon to red color. The addition of tryptophan to the medium allows the performance of the Indole test for further E. coli confirmation. Inoculate and incubate at 35 2C for 18 24. hours.
Escherichia coli Coliforms

Bibliography
Alonso, J.L. Soriano, K., Amoros I., Ferrus, M.A. 1998 Cevartitatine determination of E. coli and fecal coliforms in water using a chromogenic medium.J. Environ. Sci Health 33.

MICROBIOLOGICAL TEST
The following results were obtained in the performance of the medium from type cultures after incubation at a temperature of 35C 2C and observed after 18-24 hours Micro-organisms Escherichia coli ATCC 25922 Escherichia coli ATCC 11775 Citrobacter freundii ATCC 8090 Salmonella enteritidis ATCC 13076 Streptococcus faecalis ATCC 19433 Growth Good Good Good Good Null Colony Color Blue-dark violet Blue-dark violet Salmon Colorless ------

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Dehydrated Culture Media

SALMONELLA CHROMOGENIC AGAR

Cat: 1122
For the isolation of Salmonella in clinical samples, foods and waters. FORMULA IN GRAMS PER LITER
Sodium Citrate. Casein Peptone.. Bacteriological Agar 8.50 5.00 12.80 Chromogenic Mixture5.81 Beef Extract.5.00

Final pH 7.2 0.2 at 25C

Preparation
Suspend 37. 1 grams of the medium in one liter of distilled water. Dissolve by heating agitating frequently. Boil for one minute. DO NOT OVERHEAT. DO NOT AUTOCLAVE. Pour into Petri dishes. Keep plates refrigerated at 8-14C protecting them from light ( it may present a slight precipitate). It is recommended to prepare the plates on the same day of use.

Uses
SALMONELLA CHROMOGENIC AGAR is a new selective chromogenic medium, used for the detection and presumptive identification of Salmonella species from stool samples, foods and waters. The media used, traditionally, to differentiate species of Salmonella from the rest of Enterobacteriaceae family, based on their capacity to produce hydrogen sulfide associated with its inability to ferment lactose, are not really adequate as there are more than 2000 species of Salmonella, which do not have these characteristics. Casein peptone and Beef extract are the nutrient sources of nitrogen, vitamins, amino acids and minerals; the Chromogenic mixture aids in inhibiting Gram-positive organisms and Proteus and Coliforms in conjunction with the Sodium citrate. Bacteriological agar is the solidifying agent. To identify Salmonella species, this chromogenic agent is based on the combination of two chromogenic substrates that ease quick identification. These two chromogenes are X-gal and Magenta-caprylate. X-gal is a substrate incorporated to visualize the enzyme -D-galactosidaseproducing organisms as blue colonies. Magenta colonies are a result of the hydrolysis of Magenta-caprylate by the Lactose negative Salmonella species. Thus, non-Salmonella organisms appear blue or are not stained by any of the chromogenes of the medium. Inoculate with sample and incubate at 35 2C for 18 24 hours. Salmonella enteritidis
ATCC 13076

Bibliography
Journal Clinical Microbiology, Vol. 41 n 7 p. 3229-3232. July 2003 Robert Cassar and Paul Cuschieri. J.D. Perry, Michael Furs, Jeffrey Taylor, Et. Al. Journal Clinical Microbiology, March 1999, pag. 766-768 Vol. 37. n 3 Gallioto di camillo, p. Et. Al. (J. Clinil Microbiol. March 1999.

MICROBIOLOGICAL TEST
The following results were obtained in the performance of the medium from type cultures after incubation at a temperature of 35C 2C and observed after 18-24 hours Micro-organisms Escherichia coli ATCC 25922 Salmonella enteritidis ATCC 13076 Salmonella typhi ATCC 19430 Salmonella typhimurium ATCC 14028 Proteus vulgaris ATCC 13315 Growth Partially Inhibited Good Good Good Inhibited Colony Color Blue-green Magenta Magenta Magenta Colorless

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Dehydrated Culture Media

LISTERIA CHROMOGENIC AGAR BASE (ISO 11290:2004)

Cat: 1345
Selective medium for the detection and enumeration of Listeria monocytogenes.
FORMULA IN GRAMS PER LITER
Meat Peptone Yeast Extract Sodium Chloride Glucose Magnesium glycerophosphate.. X-glucoside 18.00 10.00 5.00 2.00 1.00 0.05 Lithium Chloride..10.00 Tryptone.6.00 Disodium Phosphate....2.50 Sodium Pyruvate...2.00 Magnesium Sulphate....0.50 Bacteriological Agar13.50

Final pH 7.2 0.2 at 25C

Preparation
Suspend 35,3 grams of the medium in 500 ml of distilled water agitating frequently and boil for one minute. Sterilize at 121C for 15 minutes. Cool to 45 - 50C and aseptically 1 vial of each supplement. Mix well and distribute into Petri dishes.
Listeria Lipase C Supplement (Cat. 6031) (1 vial for 500 ml of the medium) Lipase C Substrate ...1 gram LISTERIA CHROMOGENIC SELECTIVE SUPPLEMENT (6040) Reconstitute with 5 ml of sterile v/v ethanol - distilled water. (1 vial for 500 ml of the medium) Cycloheximide . 25 mg Ceftazidime . 10 mg Nalidixic acid ..... 10 mg Polymyxin B ... 38350 IU

Caution: Listeria Chromogenic Selective Supplement contains Cycloheximide and it is very toxic if swallowed, inhaled or comes into contact with the skin. Wear gloves and eye face protection.

Uses
LISTERIA CHROMOGENIC AGAR BASE is a selective medium for the presumptive isolation and identification of Listeria monocytogenes and Listeria spp. in food and clinical samples.It is used for confirmation after using Listeria Enrichment Broth Base Fraser (Cat. 1120). Lithium chloride and the selective supplement provide the selectivity of the medium. The differential activity of the medium is due to two factors. On one hand, the presence of the chromogenic component X-glucoside, a substrate for the detection of the enzyme -glucosidase, common to all Listeria species giving the colonies their blue colour. Other organisms that possess this enzyme, for example enterococci, are inhibited by the selective agents within the medium and by the selective supplement. On the other hand, the differential activity is also obtained by the lipase substrate, upon which the specific enzyme for L. monocytogenes acts. The lipase is responsible for the opaque white halo which surrounds L.monocytogenes. The combination of both substrates allows us to differentiate the colonies of Listeria monocytogenes from the rest of Listeria spp. since, although all are blue in colour, L.monocytogenes present an opaque white halo surrounding them. It has been observed that some strains of Listeria ivanovii, mostly pathogenic to animals although some of which have caused infections in humans, also possess lipase activity.

Listeria monocytogenes ATCC 13932

Bibliography
Ottaviani, F., Ottaviani, M. and Agosti, M (1987) Quimper Froid Symposium Proceedings, P6 A.D.R.I.A Quimper (F) 16-18 June ISO 11290-1:2004 Horizontal method for the detection and enumeration of Listeria monocytogenes Part 1: Detection Method

MICROBIOLOGICAL TEST
The following results were obtained in the performance of the medium from type cultures after incubation at a temperature of 37C 1C and observed after 24 hours Micro-organisms L. monocytogenes ATCC 1911 L. monocytogenes ATCC 13932 L. innocua ATCC 33090 Streptococcus faecalis ATCC 19433 Escherichia coli ATCC 25922 Growth Good Good Good Inhibited Inhibited Colony Color Blue Blue Blue Halo + + -

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Dehydrated Culture Media

m-EI Chromogenic Agar Base

Cat:1412
Medium to detect and enumerate the enterococcus in water through the single-step membrane filtration technique FORMULA IN GRAMS PER LITRE
Peptone.. Esculine Sodium Azide. X-Glucoside.. 10.0 1.0 0.15 0.75 Yeast Extract.......30.0 Cycloheximide.................0.05 Sodium Chloride.....15.0 Bacteriological Agar...15.0

Final pH 7.1 0.2 at 25C

Preparation
Suspend 71.95 grams of the medium in one liter of distilled water. Mix well and heat with frequent agitation for approximately a minute until the medium is completely dissolved. Sterilize in autoclave at 121C for 15 minutes. Cool at 50C and pour in Petri Dishes. For a more selective medium prepare a solution of 0.24 grams of nalidixic acid in 5ml of sterile distilled water with a few drops of sodium hydroxide 0,1N (for a better dissolution) and aseptically add to one liter of the medium.

Uses
The medium was developed as a single step procedure that does not require the transfer of the membrane filter to another substrate. Observation of blue color colonies confirm the presence of enterococci. A wide range of sample volumes or dilutions can be tested by this single step membrane-filtration procedure for the detection and enumeration of enterococci in potable, fresh, estuarine, marine and shellfish-growing waters The peptone supplies nitrogen and carbon compounds. Yeast extract provides trace elements, vitamins and amino acids. Esculin is hydrolyzed by enterococci to form esculetin and dextrose. Cycloheximide inhibits most fungi, and the sodium azide inhibits gram-negative bacteria. The XGlucoside is the substrate of the glucosidase-postive enterococci and the agar is added into the medium as a solidifying agent
Enterococcus sp

Bibliography
1. U.S. Environmental Protection Agency. 1997. Method 1600: Membrane filter test method for enterococci in water. Publication EPA-821-R97-004a. Office of Water, USEPA, Washington, D.C. 2. U.S. Environmental Protection Agency. 1986. Bacteriological ambient water quality criteria: availability. Fed. Reg. 51(45):8012. 3. U.S. Environmental Protection Agency. 2000. Improved enumeration methods for the recreational water quality indicators: enterococci and Escherichia coli. Publication EPA/821/R-97/004. Office of Science and Technology, USEPA, Washington, D. 4. Levin, Fischer and Cabelli. 1975. Appl. Microbiol. 30:66. 5. U.S. Environmental Protection Agency. 2002. Method 1600: Enterococci in water by membrane filtration using membrane-enterococcus indoxyl--D-glucoside agar (mEI). Publication EPA-821- R-02-022. USEPA Office of Water, Office of Science and Technology, USEPA, Washington, DC.

MICROBIOLOGICAL TEST
The following results were obtained of the medium from type cultures added to the supplement, after incubation at a temperature of 41C 0.5C and observed after 18-24 hours

Micro-organisms Enterococcus faecium ATCC 9790 Enterococcus faecalis ATCC 19433

Growth Good Good

Colony Color Blue Blue

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Dehydrated Culture Media

ENTEROBACTER SAKAZAKII ISOLATION CHROMOGENIC AGAR (ESIA) ISO 22964

Cat: 1446
For the isolation of presumptive Enterobacter Sakazakii

FORMULA IN GRAMS PER LITRE

Pancreatic Digest of Casein Yeast Extract -X Glucose Bacteriological Agar. 7.00 3.00 0.15 15.00 Sodium Chloride.5.00 Sodium Desoxycholate0.60 Crystal Violet.0.002

Final pH 7.0 0.2 at 25C

Preparation
Suspend 30,7 grams of the dehydrated medium in one liter of distilled water. Mix well, agitate frequently and boil for one minute. Distribute in adequate containers and sterilize and autoclave at 121C for 15 minutes. Cool to 44-47C. Pour in Petri Dishes of 15 ml.

Uses
Enterobacter sakazakii is currently considered and emerging pathogen responsible for, unweaned babies, risking severe meningitis and necrotic enterocolitis that can be the cause of mortality rate between 40-80%. The pathogenicity of Enterobacter sakazakii for un-weaned babies makes it necessary to review the manufacturing process of the products specialized for babies, guaranteeing the absence of the bacteria in the final product Additional prevention measures at a hospital include the sanitary hygiene of the prepared media; reducing the time between the preparation and its administration, to impede the multiplication of micro-organisms. The ISO normative 22964 recommends this medium for the isolation go Enterobacter sakazakii the typical colony colors of green blue greenish should be confirmed in TSA Agar Cat. 1068 in which they present yellow coloring. A Biochemical confirmation is needed from the yellow pigmentation colonies. Incubate at 41C 1C and observe after 24 hours 2 hours

Bibliography

Enterobacter Sakazakii ATCC 29544

ISO normative 22964 GUILLAUME-Gentil, O., Sonnard, V. Kandahai, M.C., Mauragg, J.D. and Jootsen, H. A simple and Rapad Cultural Method for Detection of Enterobacter Sakazakii in environmental samples. Journal of Food. Protection, 68 (1), 2005, pp. 64-69

MICROBIOLOGICAL TEST
The following results were obtained in the performance of the medium from type cultures after incubation at a temperature of 41C 1C and observed after 24 2 hours Micro-organisms Growth Good Good Good Good Inhibited Inhibited Colony Color Transparent red/violet Transparent red/violet Transparent red/violet Green-blue greenish ----------------------------------------------------------

Escherichia coli ATCC 25922 Escherichia coli ATCC 8739 Enterobacter aerogenes ATCC 13048 Enterobacter sakazakii ATCC 29544 Staphylococcus aureus ATCC 25923 Streptococcus faecalis ATCC 29212

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Dehydrated Culture Media

DEHYDRATED CULTURE

CHROMOGENIC MEDIA FOR CLINICAL SAMPLES

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CANDIDA CHROMOGENIC AGAR

Cat: 1382
Differential and selective medium for the isolation and quick Identification of Candida species of clinical importance. FORMULA IN GRAMS PER LITER
Glucose Chloranphenicol. Bacteriological Agar. 20.00 0.50 15.00 Peptone10.00 Chromogenic Mixture.....0.40

Final pH 6.1 0.2 at 25C

Preparation
Suspend 45. 9 grams of the medium in one liter of distilled water. Mix well and heat with frequent agitation until complete dissolution. Distribute into adequate containers.

Uses
CANDIDA CHROMOGENIC AGAR is an alternative chromogenic formulation to the traditional media for the detection and isolation of Candida spp. The different species of Candida produce different kinds of infections. Candida albicans is the most common and is usually susceptible to the antigfungal agents azole group. However, Candida glabrata, Candida tropicalis and Candida krusei are azole-tolerant, thus the rapid identification of the different species of Candida is essential for its correct diagnosis and treatment. In this chromogenic medium , the three different species of Candida albicans, Candida tropicalis and Candida krusei can be differentiated due to the chromogenic substrates present within the medium. Candida Chromogenic Agar allows the easy and rapid identification and differentiation of all 3 species by producing easy-to-read results in one plate, since they present different colored colonies. Colonies of Candida albicans are green, those of Candida krusei are purple-pink and those of Candida tropicalis are blue.
Candida albicans ATCC 10321 Candida krusei ATCC 34135 Candida tropicalis ATCC 1369

Bibliography
Sheehan, D.J. et. al.(1999) Current and Emerging Azole Antifungal Agents Clinical Microbiology Reviews, 12 (1): 40-79 Odds, F.C. (1988) Candida and candidiosis, 2nd ed, Baillire Tindall, London, England. Ibrahim E.H. et al. (2001) The influence of inadequate antimicrobial treatment of bloodstream infections on patient outcomes in the ICU setting. Chest, 118 (1): 146-55

MICROBIOLOGICAL TEST
The following results were obtained in the performance of the medium from type cultures after incubation at a temperature of 30-37 and observed after 24, 48 and 72 hours

Micro-organisms Candida tropicalis ATCC 1369 Candida albicans ATCC 10321 Candida Krusei ATCC 34135

Growth Good Good Inhibited

Colony Color Blue Green Purple-Pink

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Dehydrated Culture Media

URINARY TRACT INFECTIONS CHROMOGENIC AGAR (UTIC)

Cat: 1424
For the presumptive detection and differentiation of organism which cause urinary tract infections.
FORMULA IN GRAMS PER LITRE
Mix peptone.... Tryptophane.... Bacteriological Agar 16.0 2.0 16.0 Growth factors.13.0 Chromogenic Substrate 0.5

Final pH 7.2 0.2 at 25C

Preparation
Suspend 47,5 grams of the medium in one litre of distilled water. Mix well and heat agitating frequently until completely dissolved. Sterlize in autoclave at 121C for 15 minutes. Cool to 45-50C and pour into Petri Dishes.

Uses
The urinary tract infection chromogenic agar (UTIC) medium includes 2 chromogenic substrates which are cleaved by enzymes produced by Enterococcus spp, Escherichia coli and coliforms. It also includes phenylalanine and tryptophane which provide a presumptive indication of the tryptophane deaminase activity; which illustrate the presence of Proteus spp., Morganella spp. and Providencia spp. This is based on the Agar CLED. One of the chromogens is metabolised by -glucosidase enzyme activity, allowing the specific detection of enterococci which form blue or turquoise colonies. The other chromogen is cleaved -galactosidase, an enzyme produced by Escherichia coli, which grow as pink colonies. (In case of unreliable colony results, do test of Indol) Cleavage of both the chromogens shows as a result dark blue purple colonies. Tryptophane in the medium acts as an indicator of the tryptophane deaminase resulting in colonies of Proteus, Morganella, and Providencia spp of brown colour. It should be noted that, as with all chromogenic media, micro-organisms with atypical enzyme patterns may give anomalous reactions. For example 45% of Enterobacter cloacae doesnt contain -glucosidase, therefore resulting in pink colonies not distinguishable from Escherichia coli. In this case, do Indol test

Bibliography
Samra Z, Heifetz M, Talmor J, Bain E and Bahar J. Evaluation of use of a new chromogenic agar in detection of urinary tract pathogens. J Clin Microbiol. 1998;36(4): 990-4.

MICROBIOLOGICAL TEST
The following results were obtained in the performance of the medium from type cultures after incubation at a temperature of 35 2C and observed 18-24 hours

Microorganisms Escherichia coli ATCC 25922 Enterobacter aerogenes ATCC 13048 Klebsiella pneumonieae ATCC 13883 Proteus miriabilis ATCC 13315 Staphylococcus aureus ATCC 25923 Enterococcu faecalis ATCC 19433

Growth Good Good Good Good Good Good

Colour Colony Rose Dark Blue Dark Blue Light Brown White Cream (natural pigmentation) Light Blue

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Dehydrated Culture Media

MRSA AGAR
Cat: 1423
For the detection of Methicilin resistant Staphylococcus aureus FORMULA IN GRAMS PER LITRE
Peptone mixture. Chromogenic Substrate. 11.00 1.90 Growth Factors.78,00 Bacteriological Agar.12.50

Final pH 7.2 0.2 at 25C

Preparation
Suspend 103.5 grams of the medium in one liter of distilled water. Mix well and heat agitating continuously until dissolved. Sterilize in autoclave at 121C for 15 minutes. Cool to 45-50C and aseptically add 4 mg per liter of Cefoxitin supplement.

Uses
Methicilin resistant Staphylococcus aureus, MRSA, are of particular interest at an international level due to its virulence and resistance to multiple antibiotics. The antimicrobial resistance is a serious threat to public health as it is now regarded as a major hospital acquired disease worldwide. The important changes observed in the epidemiological and microbiological characteristics of the infections caused by Staphylococcus aureus are the reason for the increment and prevalence of methicilin-resitant Staphylococcus aureus nosocomial (associated to hopitalized patients) and the proliferation of methicilin-resistent Staphylococcus aureus acquired by the community The MRSA continues being a serious problem in many healthcare centres; more than 50% of the Staphylococcus aureus obtained are from Intensive Care Units (ICU) and close to 40% are from hospital patients. Effective, rapid laboratory diagnosis and susceptibility testing is critical in treating, managing and preventing MRSA infections. This chromogenic media has been designed and is adequate for the screening of Staphylococcus aureus resistant to methicilin. The -glucosidase produced by Stahphlococcus aureus cleaves the chromogenic substrate and gives a blue color to the Staphylococcus aureus colony. The Cefoxitin inhibits the growth of Staphylococcus aureus sensitive to methicilin.
Staphylococcus aureus ATCC 25923

Bibliography
Hutchison, M.J., Edwards, G.F.S., Morrison, D., ,, Evaluation of chromogenic MRSA Reference Laboratory presented at the 2005 Institute of BioMedical

MICROBIOLOGICAL TEST
The following results were obtained of the medium from type cultures, with the supplement added, after incubation at a temperature of 37C 2C and observed after 18-24 hours Micro-organisms Staphylococcus aureus ATCC 25923 Staphylococcus aureus ATCC 43300 Escherichia Coli ATCC 25922 Growth Inhibited Good Inhibited Colony Color ---Blue ----

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Dehydrated Culture Media