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Tryptic Soy Agar/Trypticase™ Soy Agar (Soybean-Casein Digest Agar)

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Tryptic Soy Agar/Trypticase™ Soy Agar (Soybean-Casein Digest Agar) Tryptic Soy Agar Formula Presumptive negative nitrate reduction reaction: Lack Difco™ Tryptic Nitrate Medium of color development denotes an absence of nitrite in the Approximate Formula* Per Liter medium; this should be confirmed by addition of Nitrate C Tryptose ................................................................... 20.0 g Reagent (zinc dust). Dextrose ..................................................................... 1.0 g Disodium Phosphate .................................................. 2.0 g 2. After adding Nitrate C Reagent: Potassium Nitrate ....................................................... 1.0 g Positive nitrate reduction reaction: Lack of color development Agar ........................................................................... 1.0 g indicates that nitrate has been reduced to nitrogen gas. *Adjusted and/or supplemented as required to meet performance criteria. Negative nitrate reduction reaction: Development of a red- Directions for Preparation from violet color within 5-10 minutes indicates that unreduced Dehydrated Product nitrate is still present. 1. Suspend 25 g of the powder in 1 L of purified water. Mix thoroughly. Limitation of the Procedure 2. Heat with frequent agitation and boil for 1 minute to com- This medium is not recommended for indole testing of coliforms pletely dissolve the powder. and other enterics.1 3. Autoclave at 121°C for 15 minutes. 4. Test samples of the finished product for performance using References 1. MacFaddin. 1985. Media for isolation-cultivation-identification-maintenance of medical bacteria, stable, typical control cultures. vol. 1. Williams & Wilkins, Baltimore, Md. 2. U.S. Food and Drug Administration. 1995. Bacteriological analytical manual, 8th ed. AOAC Inter- national, Gaithersburg, Md. Procedure 3. Pezzlo. 1992. In Isenberg (ed.), Clinical microbiology procedures handbook, vol. 1. American Society for Microbiology, Washington, D.C. 1. Obtain a pure culture of the organism to be tested from a 4. Downes and Ito (ed.). 2001. Compendium of methods for the microbiological examination of foods, 4th ed. American Public Health Association, Washington, D.C. solid culture medium. Select well-isolated colonies. 5. Flowers, Andrews, Donnelly and Koenig. 1993. In Marshall (ed.), Standard methods for the exami- nation of dairy products, 16th ed. American Public Health Association, Washington, D.C. 2. Inoculate a tube of Tryptic Nitrate Medium and incubate 6. Murray, Baron, Pfaller, Tenover and Yolken. 1999. Manual of clinical microbiology, 7th ed. Ameri- aerobically or anaerobically, as appropriate, at 35 ± 2°C can Society for Microbiology, Washington, D.C. 7. Forbes, Sahm and Weissfeld. 1998. Bailey and Scott’s diagnostic microbiology, 10th ed. Mosby, for 18-24 hours. Inc., St. Louis, Mo. 3. Read tubes for growth. 4. Test for nitrate reduction using Difco™/BBL™ Nitrate A, B Availability ™ and C Reagents or equivalents per reagent instructions. Difco Tryptic Nitrate Medium Cat. No. 236710 Dehydrated – 500 g Expected Results Difco™/BBL™ Nitrate A Reagent T 1. After adding Nitrate A and B Reagents: Cat. No. 261197 Droppers – 50 × 0.5 mL Positive nitrate reduction reaction: Development of a red- Difco™/BBL™ Nitrate B Reagent violet color within 1-2 minutes indicates that nitrate has Cat. No. 261198 Droppers – 50 × 0.5 mL been reduced to nitrite. Difco™/BBL™ Nitrate C Reagent Cat. No. 261207 Vials – 50 × 1 g Tryptic Soy Agar/Trypticase™ Soy Agar (Soybean-Casein Digest Agar) Intended Use Hycheck™ hygiene contact slides are used for assessing the Tryptic Soy Agar and Trypticase™ Soy Agar conform with speci- microbiological contamination of surfaces and fluids. fications of The United States Pharmacopeia (USP). Summary and Explanation Tryptic (Trypticase) Soy Agar (TSA) is used for the isolation The nutritional composition of TSA has made it a popular and cultivation of nonfastidious and fastidious microorgan- medium for many years. It is the medium specified as isms. It is not the medium of choice for anaerobes. Soybean-Casein Digest Agar Medium in the USP for the total The 150 × 15 mm-style plates of Trypticase Soy Agar are aerobic microbial count portion of the microbial limit testing convenient for use with Taxo™ factor strips in the isolation procedures.1 The medium is used for a multitude of purposes and differentiation of Haemophilus species. including maintenance of stock cultures, plate counting, isolation of microorganisms from a variety of specimen types Sterile Pack and Isolator Pack plates are useful for monitoring and as a base for media containing blood.2-4 It is included surfaces and air in clean rooms, Isolator Systems and other in the compendia of methods for the examination of water, environmentally-controlled areas when sterility of the medium wastewater and foods,5, 6 in the Bacteriological Analytical is of importance. Manual 7 and is used for testing bacterial contaminants in cosmetics.8 577 Section III T Tryptic Soy Agar, cont. User Quality Control NOTE: Differences in the Identity Specifications and Cultural Response testing for media offered as both Difco™ and BBL™ brands may reflect differences in the development and testing of media for industrial and clinical applications, per the referenced publications. Identity Specifications Identity Specifications Difco™ Tryptic Soy Agar BBL™ Trypticase™ Soy Agar Dehydrated Appearance: Light beige, free-flowing, homoge- Dehydrated Appearance: Fine, homogeneous, free of extrane- neous. ous material. Solution: 4.0% solution, soluble in purified Solution: 4.0% solution, soluble in purified water upon boiling. Solution is light water upon boiling. Solution is light amber, slightly opalescent. to medium, yellow to tan, clear to Prepared Appearance: Plain – Light amber, slightly opalescent. slightly hazy. With 5% sheep blood – Bright red, Prepared Appearance: Plain – Light to medium, yellow to tan, opaque. clear to slightly hazy. Reaction of 4.0% With 5% sheep blood – Bright red, Solution at 25°C: pH 7.3 ± 0.2 opaque. Reaction of 4.0% Cultural Response Solution at 25°C: pH 7.3 ± 0.2 Difco™ Tryptic Soy Agar Prepare the medium per label directions, without (plain) and with 5% Cultural Response sheep blood (SB). Inoculate and incubate at 35 ± 2°C for 18-48 hours BBL™ Trypticase™ Soy Agar under 5-10% CO2. Prepare the medium per label directions, without (plain) and with 5% sheep blood (SB). Inoculate and incubate at 35 ± 2°C for 2 days INOCULUM RECOVERY RECOVERY (incubate streptococci with 3-5% CO ). ORGANISM ATCC™ CFU PLAIN WITH SB HEMOLYSIS 2 Escherichia coli 25922 102-103 Good Good Beta INOCULUM RECOVERY RECOVERY ™ Neisseria ORGANISM ATCC CFU PLAIN WITH SB HEMOLYSIS meningitidis 13090 102-103 Good Good None Candida albicans 10231 103-104 N/A Good None Staphylococcus Escherichia coli 25922 103-104 N/A Good Beta aureus 25923 102-103 Good Good Beta Listeria Streptococcus monocytogenes 19115 103-104 N/A Good Beta (+/-) pneumoniae 6305 102-103 Good Good Alpha Pseudomonas Streptococcus aeruginosa 10145 103-104 Good N/A pyogenes 19615 102-103 Good Good Beta Shigella flexneri 12022 103-104 Good N/A CAMP Test medium with 5% sheep blood – Perform using S. aureus ATCC 33862, Streptococcus Staphylococcus sp. group B ATCC 12386 (positive) and S. pyogenes ATCC 19615 (negative). aureus 25923 103-104 Good Good Beta Streptococcus 3 4 Escherichia coli Staphylococcus aureus pneumoniae 6305 10 -10 Good Good Alpha ATCC™ 25922 ATCC™ 25923 Streptococcus pyogenes 19615 103-104 Good Good Beta CAMP Test medium with 5% sheep blood – Perform using S. aureus ATCC 25923, Streptococcus sp. group B ATCC 12386 (positive) and S. pyogenes ATCC 19615 (negative). Since TSA does not contain the X and V growth factors, it can conveniently be used in determining the requirements for these growth factors by isolates of Haemophilus by the addition of X, V and XV Factor Strips to inoculated TSA plates.3 The 150 mm plate provides a larger surface area for inoculation, making the “satellite” growth around the strips easier to read. With the Sterile Pack and Isolator Pack plates, the entire double- wrapped (Sterile Pack) or triple-wrapped (Isolator Pack) product is subjected to a sterilizing dose of gamma radiation, so that the contents inside the outer package(s) are sterile.9 This allows the inner package to be aseptically removed without introducing contaminants. Since the agar medium has been sterilized after packaging, the presence of microbial growth after sampling and incubation can be relied upon to represent true recovery and Streptococcus Streptococcus not pre-existing medium contaminants. A third rolled sterile pneumoniae pyogenes ATCC™ 6305 ATCC™ 19615 bag is included as a transport device. Isolator Pack plates have been validated to protect the medium from vaporized hydrogen peroxide when used in an Isolator System. 578 Tryptic Soy Agar, cont. The Hycheck hygiene contact slide is a double-sided paddle Trypticase™ Soy Agar (150 mm plates) for Haemophilus containing two agar surfaces for immersing into fluids or The initial specimens should be inoculated onto Chocolate II sampling surfaces. There are three slides containing TSA along Agar or another suitable medium and incubated for 18-24 with another medium: D/E Neutralizing Agar; Violet Red hours in an aerobic atmosphere supplemented with carbon Bile Glucose Agar; or Rose Bengal Chloramphenicol Agar. A dioxide. Choose one or two well-isolated colonies that fourth slide contains TSA with 0.01% TTC and Rose Bengal resemble Haemophilus species and perform a Gram stain to Chloramphenicol
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