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Principle : HiDip slide is a handy tool for isolation, enumeration and adjacent to the initial test side. Return the slide to the container and close identification of specific bacteria in urine, food and water samples. These tightly. Incubate in an up right position at indicated temperature. slides can also be used as touch slides for assessing the microbiological 2. Liquids : Loosen cap and remove the HiDip slide from the container. contamination of surfaces. Hi Dip slides are designed to monitor the Check for dehydration or contamination. Dip slide into test fluid for upto microbial flora of liquids (e.g. urine, milk, water) and equipment surfaces 15-20 seconds so that agar surface becomes totally covered. (In case of in the clinical and food industries. inadequate liquid sample availability, pour sample over the surface of the slide). Allow to drain. Tab it gently to remove excess fluid from Hi Dip slide contains a double sided, hinged plastic scull containing surface. Return the slide to the container and close tightly. Incubate in an two agar surfaces. The HiDip slides containing combination of three agar upright position at indicated temperature. Label the container for sample media are prism shaped having all three agar media on separate, number, source, date and time etc. for reference. individual surfaces. The extended agar surface above the scull allows the Interpretation : One may read counts by removing slide from the touch with test surfaces. The hinged scull allows easy touch against each container. Compare growth with chart provided to determine density of test area during sampling. The surface area of scull is divided into ten 7 colonies. If the density exceeds 10 CFU or viscocity of sample is more, units of one centimeter each to allow direct counting of microbial the sample should be diluted and dilution factor is to be accounted for in density per unit area. the count. Presence of single colony (spot) on slide surface can be Hi Dip slides are available in d ifferent agar combinations of two considered as count 102 . No colony on the slide surface can be media or three media which are available in pack of 5 tubes and 10 considered as count less than 102 . tubes. Storage & Shelf-life : Store Hi Dip slides at 15o C, optimum storage o Directions temperature is 15-25 C. Avoid the sudden temperature alterations. 1. Surfaces : Loosen cap and remove HiDip slide from container taking Expiry period is given on each box. care not to touch agar surfaces. Check for dehydration or contamination. Disposal : Used HiDip slides should be handled carefully, as it contains Gently lower the slide and press agar to touch the test surface by live microorganisms. These slides can be best disposed of either by incinerating or by immersing in a suitable disinfectant solution (i.e dettol, bending the scull around the hinge line. Apply even and firm pressure phenyl etc.) overnight or by autoclaving them after losening the cap. An for 15-20 seconds. Take care not to smudge agar over the test autoclave is not essential, a domestic pressure cooker will suffice. surface. Repeat procedure using the second agar surface on an area TM

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Media used in various combinations Cultural Response : Colour of prepared Medium (control) : Light amber for HiDip Slides Cultural characteristics after 24 - 48 hours at 35 - 37O C. CLED Agar Organisms (ATCC) Growth CLED Agar is recommended for isolation, enumeration and identification Pseudomonas aeruginosa (27853) luxuriant of urinary pathogens on the basis of lactose fermentation. Incubate slides Pseudomonas maltophila (13637) inhibited O at 35-37 C for 18-24 hours to observe cultural characteristics. Staphylococcus aureus (25923) inhibited Cultural Response : Escherichia coli (25922) inhibited Colour of prepared Medium (control) : Green Bile Esculin Medium Cultural characteristics after 18 - 24 hours at 35 - 37O C. Bile Esculin Medium is in accordance with APHA for the presumptive Organisms (ATCC) Growth Colour of colony identification of Group D Streptococci. ISO Committee has also Escherichia coli (25922) luxuriant yellow, opaque, center suggested it for the isolation and identification of Yersinia enterocolitica. slightly deeper yellow Esculin hydrolyzers hydrolyze the glycoside esculin to esculetin and Klebsiella pneumoniae (13883) luxuriant yellow to whitish blue dextrose. Esculetin reacts with ferric citrate to form a black-brown Proteus vulgaris (13315) luxuriant blue complex. Incubate slides at 35-37O C for 18-24 hours to observe cultural Salmonella Typhi (6539) luxuriant bluish characteristics. Staphylococcus aureus (25923) luxuriant deep yellow Enterococcus faecalis (29212) luxuriant slight yellowish or greenish Cultural Response : Colour of prepared Medium (control) : Amber to Yellow MacConkey Agar O Cultural characteristics after 18-24 hours at 35 - 37 C. MacConkey Agar is a differential medium for the selection and recovery Organisms (ATCC) Growth Esculin hydrolysis of the Enterobacteriaceae and related enteric gram-negative bacilli. This Enterococcus faecalis (29212) luxuriant + medium is prepared in accordance with USP and contains crystal violet, Streptococcus faecium (8482) luxuriant + NaCl and bile salts. It is very selective and suppresses growth of a number Yersinia enterocolitica (27729) luxuriant + of Gram-positive bacteria including Staphylococci. Incubate slides at Escherichia coli (25922) good — 35-37O C for 18-24 hours to observe cultural characteristics. Coliforms Key :+ = blackening of the medium and Enterobacter give pink to red colonies on this medium and —= no change Escherichia coli gives pink to red colonies with bile precipitate. Cultural Response : Sabouraud Dextrose Agar Colour of prepared Medium (control) : Red w/ purplish tinge Sabouraud Dextrose Agar is used for the cultivation of fungi particularly Cultural characteristics after 18 - 24 hours at 35 - 37O C. for the fungi associated with skin infections. This medium is also in Organisms (ATCC) Growth Colour of Colony accordance with the US Pharmacopeia for microbial limit tests. Incubate slides at 30O C for 48-72 hours to observe colony characteristics. Enterobacter aerogenes (13048) luxuriant pink to red Escherichia coli (25922) luxuriant pink to red with bile Cultural Response : precipitate Colour of prepared Medium (control) : Light amber Proteus vulgaris (13315) luxuriant colourless Cultural characteristics after 48 - 72 hours at 30O C. Salmonella Enteritidis (13076) luxuriant colourless Organisms (ATCC) Growth Salmonella Typhi (6539) luxuriant colourless Aspergillus niger (16404) luxuriant Salmonella Paratyphi A luxuriant colourless Candida albicans (10231) luxuriant Salmonella Paratyphi B luxuriant colourless Trichophyton rubrum (28191)* luxuriant Shigella flexneri (12022) fair to good colourless Saccharomyces cerevisiae (9763) luxuriant Escherichia coli (25922) luxuriant Enterococcus faecalis (29212) fair to good colourless to pink Lactobacillus casei (9595) luxuriant Staphylococcus aureus (25923) inhibited — * Growth after upto 6 days Cetrimide Agar MUGMac Medium Cetrimide Agar is used as a selective medium for the isolation of MUGMac Medium is used for selective isolation and detection of Pseudomonas aeruginosa from pus, sputum, drains etc. This medium lactose fermenting coliform organisms by a fluorogenic procedure. The helps in determining the ability of organism to produce fluorescein and medium helps to detect the presence of an enzyme ß-glucuronidase pyocyanin. Cetrimide inhibits bacteria other than Pseudomonas and thereby rapidly identifying Escherichia coli in mixed clinical aeruginosa. Pseudomonas aeruginosa colonies appear pigmented blue specimens. MUG is cleaved by the enzyme to release an end product or blue green when slides are incubated at 35-37O C for 24-48 hours. 4 - methylumbelliferone which produces a visible greenish - blue fluorescence under long wave ultra-violet light. Incubate slides at 35-37O C for 18-24 hours to observe cultural characteristics. TM

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Cultural Response : Cultural Response : Colour of prepared Medium (control) : Red w/ purplish tinge Colour of prepared Medium (control) : Deep pink O Cultural characteristics after 18 - 24 hours at 35 - 37O C. Cultural characteristics after 5 days at 25-30 C. Organisms (ATCC) Growth Fluorescence Organisms (ATCC) Growth Enterobacter aerogenes (13048) luxuriant — Aspergillus niger (16404) good Escherichia coli (25922) luxuriant + Candida albicans (10231) good Escherichia coli (25922) inhibited Key : Flourescence at 366 nm Micrococcus luteus (10240) inhibited HiCrome UTI Medium HiCrome UTI Medium is a differential medium recommended for PCA Medium (w/ growth indicator) identification and confirmation of microorganisms mainly causing Plate Count Agar is recommended for the plate count of urinary tract infections. It facilitates and expedites the identification of microorganisms in foods, water and waste water. It is also recommended O some Gram-negative and some Gram-positive bacteria on the basis of by APHA, FDA and also by ISO Committee. Incubate plates at 35-37 C different contrasted colony colours produced by reactions of genus or for 18-24 hours to observe colony characteristics. Growth indicator aids species specific enzymes with two chromogenic substrates. Incubate in easier observation and enumeration of colonies. slides at 35-37O C for 24 hours to observe cultural characteristics. Further Cultural Response : confirmation of Escherichia coli can be done by performing indole test Colour of prepared Medium (control) : Light yellow using DMACA Reagent (R035) and Proteus species can be confirmed by Cultural characteristics after 18 - 24 hours at 35 - 37O C. using TDA Reagent (R036). Pick up the suspected colony using sterile Organisms (ATCC) Growth Color of colony loop or glass rod and transfer to a piece of filter paper. Add 1-2 droop of Bacillus subtilis (6633) luxuriant red to maroon reagent or directly perform test on colony. Escherichia coli (25922) luxuriant red to maroon Cultural Response : Lactobacillus casei (9595) luxuriant red to maroon Colour of prepared Medium (control) : Light amber Staphylococcus aureus (25923) luxuriant red to maroon O Cultural characteristics after 24 hours at 35-37 C. Enterococcus faecalis (29212) luxuriant red to maroon Organisms (ATCC) Growth Colour of TDA*DMACA** Streptococcus pyogenes (19615) luxuriant red to maroon colony Escherichia coli (25922) luxuriant pink — + VRBA Medium Proteus mirabilis (10975) luxuriant light brown + — VRBA medium is selective medium used for the detection and Klebsiella pneumoniae luxuriant blue to purple, —— (13883) mucoid enumeration of coliform organisms from water and food products. It is also suggested by ISO Committee for the enumeration of coliforms. Pseudomonas aeruginosa luxuriant colourless —— (27853) It is selective due to presence of the inhibitors-bile salt and crystal violet. Organisms which rapidly ferment lactose produce red Staphylococcus aureus luxuriant golden yellow —— (25923) colonies surrounded by red-purple halo. Lactose non-fermenters and Enterococcus faecalis (29212) luxuriant blue, small —— late lactose fermenters produce pale colonies. Incubate slides at 35-37O C for 18-24 hours to observe cultural characteristics. * TDA + : Brown colour develops on growth ** DMACA + : Red colour develops on growth. Cultural Response : Colour of prepared Medium (control) : Reddish purple TSA (w/ growth indicator) Cultural characteristics after 18 - 24 hours at 35 -37O C. Tryptone Soya Agar is recommended for cultivation of a wide variety of Organisms (ATCC) Growth Colour of colony organisms. This medium is in accordance with USP. Incubate slides at O Escherichia coli (25922) luxuriant pinkish red w/bile ppt 30-35 C for 18-48 hours to observe cultural characteristics. Enterobacter aerogenes (13048) luxuriant pink Cultural Response : Colour of prepared Medium (control) : Light yellow Salmonella Enteritidis (13076) luxuriant colourless O Cultural characteristics after 18 - 48 hours at 30-35 C. Staphylococcus aureus (25923) inhibited — Organism (ATCC) Growth Color of colony Candida albicans (10231) luxuriant red to maroon Sulphate API Medium Staphylococcus aureus (25923) luxuriant red to maroon Sulphate API Medium is used for detection and estimation of Streptococcus pyogenes (19615) good-luxuriant red to maroon sulphate reducing bacteria as per American Petroleum Institute Bacillus subtilis (6633) luxuriant red to maroon Recommended Practice. Sulphate reducing bacteria convert Bacteroides vulgatus (8482)* luxuriant red to maroon sulphate to sulphide which on reaction with the ferrous ion Neisseria meningitidis (13090)** luxuriant red to maroon gives black colour. Incubate slides at 30O C for 1 week to * : incubated anaerobically ** : incubated under 5 % CO2 observe cultural characteristics. Rose Bengal Medium Rose Bengal Medium is recommended for the selective isolation and enumeration of yeasts and moulds from environmental materials and food stuffs. Rose bengal is taken up by mould and yeast colonies thereby assist in enumeration. Incubate slides at 25-30O C for 5 days to observe cultural characteristics.

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Cultural Response : VJ Agar Colour of prepared Medium (control) : Light yellow VJ Agar permits early detection of coagulase positive and mannitol Cultural characteristics after 4 days upto 1 week at 22-28O C. positive colonies of Staphylococcus aureus. It is recommended for the Sulphate API Medium microbial limit test in USP. Coagulase positive Staphylococci reduce Organism (ATCC) Growth potassium tellurite to metallic free tellurium and thus produce black colonies surrounded by yellow zones. Desulfovibrio desulfuricans (29577) good to luxuriant PA Medium Cultural response PA Medium is used for the detection of coliforms bacteria in milk, food Colour of Prepared Medium (Control) : Red 0 and water from treatment plants or distribution systems. This medium Cultural characteristics after 18 -24 hou r s at 35-37 C. has been included as a tentative standard in the “Standard Methods for Organisms (ATCC) Growth Colour of colony the Examination of Water and Wastewater”, APHA. Incubate slides at Staphylococcus aureus (25923) luxuriant black with yellow halo 35-37O C for 18-24 hours to observe cultural characteristics. Staphylococcus epidermidis (12228) fair blackish Proteus mirabilis (25933) poor black Cultural Response : Escherichia coli (25922) inhibited — Colour of prepared Medium (control) : Greyish Purple Cultural characteristics after 18 - 48 hours at 35 - 37O C. Malt Extract Agar Medium Organisms (ATCC) Growth Colour of medium Malt Extract Medium is recommended for detection, isolation and Escherichia coli (25922) good-luxuriant yellow enumeration of yeasts and moulds. Typical colonies of yeasts and Enterobacter aerogenes (13048) good-luxuriant light yellow moulds can be visualized on this media. Klebsiella pneumoniae (13883) good-luxuriant yellow Cultural Response Salmonella Typhimurium (14028) good-luxuriant — Colour of prepared medium (control): Yellow Enterococcus faecalis (29212) inhibited — Cultural characteristics after upto 48-72 hours at 25-30O C.

HS2 Medium Organisms (ATCC) Growth Aspergillus niger (16404) luxuriant HS2 medium is recommended for simultaneous detection of Salmonella, Vibrio, Citrobacter species and Escherichia coli from milk, food and Candida albicans (10231) lxuriant Saccharomyces cerevesiae (9763) luxuriant water samples. Presence of growth with or without change of colour of medium indicates microbial contamination of specimen. Pseudomonas Agar medium Incubate slides at 35-37O C for 18-24 hours to observe cultural Pseudomonas Agar medium is recommended for the detection of characteristics. pyocyanin production by Pseudomonas species. Pseudomonas Culture Response : aeruginosa colonies appear pigmented blue or blue green when slides are incubated at 35 - 370 C for 18-24 hours. Colour of prepared Medium (control) : Greenish Blue Cultural characteristics after 18-24 hours at 35-37O C Cultural response Colour of Prepared Medium (Control) : Yellow Organisms (ATCC) Colour of colonies 0 Escherichia coli (25922) blue Cultural characteristics after 18 - 24 hours at 35 - 37 C. Vibrio cholerae (15748) bluish purple Organisms (ATCC) Growth Colour Salmonella Enteritidis (13076) blue w/black centre P. aeruginosa (27853) luxuriant Blue-green Salmonella Typhimurium (14028) blue w/black centre Citrobacter freundii (8090) blue w/black centre Klebsiella pneumoniae (13883) purple Three-in-one Packing HiDip Slides format CLED Agar w/BTB Cled Agar w/BTB is recommended for isolation, and differentiation of urinary pathogens on the basis of lactose fermentation. This medium has Bromo thymol blue as pH indicator which turns yellow at acidic pH due to fermentation of lactose. Cultural response : Colour of Prepared Medium (Control) : Green Cultural characteristics after 18 -24 hours at 35-370 C. Organisms (ATCC) Growth Colour of colony Escherichia coli (25922) luxuriant yellow, opaque, center slightly deeper yellow Klebsiella pneumoniae (13883) luxuriant yellow to whitish blue Proteus vulgaris (13315) luxuriant blue Salmonella Typhi (6539) luxuriant blue Staphylococcus aureus (25923) luxuriant deep yellow Enterococcus faecalis (29212) luxuriant slight yellowish or green HiMedia container Remove Remove Remove 10 10 10

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Literature Code :TL002_04/1208