BD BBL Dermatoslide. INSTRUCTIONS FOR USE READY-TO-USE DIPSLIDE MEDIA DA Rev.: July 2003
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1 INSTRUCTIONS FOR USE READY-TO-USE DIPSLIDE MEDIA DA Rev.: July 2003 BD BBL Dermatoslide INTENDED USE BBL Dermatoslide is a two-sided slide system for the detection and isolation of pathogenic fungi from superficial infections of skin, hair, and nails. PRINCIPLES AND EXPLANATION OF THE PROCEDURE Microbiological method. Fungi play a major role in infections of the skin and its adjacent organs, such as hair and nails. Typically and most frequently, mycelial fungi like Trichophyton, Epidermophyton, and Microsporum, but also others are found as infectious agents. Yeast species like Candida albicans may also be the cause of superficial infections of the skin.. In order to allow for proper identification and therapy, both selective and nonselective isolation media must be used for the detection of the agents involved. Medium 1 is a modified Dermatophyte Test Medium Agar according to Taplin for the detection of dermatophytes, especially, but not limited to, Trichophyton, Epidermophyton, and Microsporum species. In this medium, peptones supply nitrogen and are the source of alcaline products, produced by dermatophytes. 1 When peptones are metabolized to alcaline products, a change of the phenol red indicator from yellow to red will take place. Glucose is added as a nutrient and to allow acidification by fungi able to primarily use glucose. Most fungi other than dermatophytes, including yeasts and moulds (if they are able to grow on the medium), will utilize glucose. This results in acid formation and no color change of phenol red which is the ph indicator. 2,3 Cycloheximide is an inhibitor for moulds and non-pathogenic yeasts. Chloramphenicol and gentamicin are antibacterial inhibitors. A few organisms, including saprophytes, yeasts, and bacteria, are capable of growing on the medium and change the color from red to yellow, but they are easily recognized by their distinctive colonial morphology. Medium 2 is Malt agar for the detection of yeasts. This medium is used for the detection of all types of fungi, including moulds and yeasts. Malt extract contains all necessary nutrients to support growth of fungi. The low ph of the medium results in a partial to complete inhibition of contaminating bacteria but is well tolerated by fungi. 4,5 BBL Dermatoslide is used for primary isolation of fungi, especially dermatophytes, from a variety of superficial infections. It can also be used as a transport medium from the physician s office to analytical laboratories. REAGENTS BBL Dermatoslide Formula* Per Liter Purified Water Medium 1 Medium 2 Pancreatic Digest of Casein 15.0 g Malt Extract 30.0 g Papaic Digest of Soybean Meal 5.0 Agar 18.0 Sodium Chloride 5.0 ph 4.0 ± 0.2 Agar 20.0 Glucose 10.0 Phenol Red 0.2 Cycloheximide 0.5 Chloramphenicol 0.1 Gentamicin 0.1 ph 5.5 ± 0.1 *Adjusted and/or supplemented as required to meet performance criteria. DA
2 PRECAUTIONS. For professional use only. Do not use slides if they show evidence of microbial contamination, discoloration, drying, cracking or other signs of deterioration. When using BBL Dermatoslide as a transport medium from a physician s office to a diagnostic laboratory, observe local regulations for shipping clinical specimens. Consult GENERAL INSTRUCTIONS FOR USE document for aseptic handling procedures, biohazards, and disposal of used product. Wear protective gloves when collecting and handling specimens and positive slides containing infectious agents. STORAGE AND SHELF LIFE On receipt, store BBL Dermatoslide in the dark at C in the original package until just prior to use. Avoid freezing, draught, temperature fluctuations and overheating. Unopened slides from opened packages of 10 slides can be used until the indicated expiry date. Opened slides must be used immediately. USER QUALITY CONTROL Prepare suspensions matching the McFarland standard 0.5 of the strains mentioned below. For each organism, prepare a tube (wide enough to allow introduction of a BBL Dermatoslide agar carrier) with 20 to 25 ml of sterile saline and add 1 ml of the above suspension of the strain. Open a BBL Dermatoslide, remove the agar carrier, dip it briefly into the test strain suspension, and proceed as described under Test Procedure for liquid specimens. Incubate for 5 to 7 days at 35 ± 2 C. Inspect agar surfaces for growth as described under Results and Interpretation. Strains Medium 1 Medium 2 Trichophyton mentagrophytes * ATCC 9533 Candida albicans ATCC Candida glabrata ATCC 2001 Aspergillus niger ATCC Pseudomonas aeruginosa ATCC Enterococcus faecalis ATCC Staphylococcus aureus ATCC Uninoculated Yellow, clear to slightly Colorless to light amber opaque * Note that growth of dermatophytes on this medium may be weaker or may take longer to occur than on medium 1. PROCEDURE Materials Provided BBL Dermatoslide. Microbiologically controlled. Materials Not Provided Ancillary culture media, reagents, laboratory equipment and devices for collection as described. Specimen Types BBL Dermatoslide can be used for all specimens suspected to contain fungi causing infections of skin, hair, nails, etc. Collection of Specimens, Inoculation, and Test Procedure Before use, visually inspect the agar surfaces for sterility, leaving BBL Dermatoslide unopened. Label the tube with patient name, specimen number, and date of inoculation. DA
3 Prior to the inoculation, remove the slide from the plastic tube without touching the agar surfaces. Skin: Clean the affected site with 70% ethyl or isopropyl alcohol prior to removing skin scales. Always use sterile instruments for specimen collection. Remove scales from dry and peeling lesions by scraping from the inflamed edges towards the healthy skin with a scalpel. Disposable scalpels can easily hurt the skin (it is preferable to use only the back of the blade). Scrape large areas thoroughly and collect as much material as possible. With acute inflamed or vesicular lesions, the skin of the blister must be carefully removed with scissors and forceps. It should be cultured together with the content of the blister and, if possible, scales from the surrounding area. With infiltrates or granulomatous processes, collect material from the depth and from skin folds with a sharp spoon or a vaccination lancet. Collect the material on a piece of filter paper or directly on BBL Dermatoslide. If the replica technique is applied for specimen collection, carefully scrape off the surface layers of skin with a sharp instrument and then gently press the surfaces of BBL Dermatoslide against the skin site to be checked, beginning with medium 1. Hair: Pluck the roots of dull, lusterless hair with a forceps. Infected hair breaks and loosens more easily than healthy hair. If Wood s light is used, collect fluorescent hair, even if it looks healthy in daylight. If so-called black dots appear, lift the infected hair out of the bulb by using the edge of a scalpel. Do not use cut hair as a specimen. Distribute hair onto both media of the slide. Gently press hair onto the agar with the forceps. Nails: In case of subungal infection, all grossly deformed surface parts of the nail are removed carefully with scissors, nail file, or scalpel. Chips of nail are then collected from the nail bed. In case of surface infection, nail chips or small dustlike particles are scraped from the surface of the nail body. Do not inoculate BBL Dermatoslide with large nail pieces cut from extracted nails. It is best to use a nail fraise. Inoculate both sides of the slide with the specimen. Specimens collected on swabs or with forceps: Distribute the material evenly on both agar media (beginning with agar 1) with a sterile swab or sterile forceps. Gently press the larger particles onto the agar with the forceps in order to provide contact between the specimen and the agar surface. Note that swabs taken from infected areas are not the appropriate specimens for collecting dermatophytes. For further description of the collection methods and types of specimens, consult the references. 3,4,6 After specimen collection and inoculation, return the slide into the tube and tighten the screw cap carefully. Incubate BBL Dermatoslide at C for up to 4 weeks. Inspect the agar surfaces daily. For further processing or identification of the pathogens, the inoculated slide may also be sent to a special laboratory. Results After the incubation, inspect the agar surfaces visually without opening the tube. Medium 1: of dermatophytes leads to a color change of the medium underneath the colonies from yellow to red, even before the colonies are fully developed. As growth progresses, the color of the whole medium will gradually become red and a white, gray, or yellowish mycelium will become visible. The specimen is considered negative for dermatophytes if no growth is visible after the incubation on this medium. Certain saprophytic mould contaminants may also grow on medium 1. However, in this case the colonies and the mycelium usually appear before a color change of the medium occurs. Certain cycloheximide-resistant yeasts (e.g., Candida albicans) may also produce a color change of the medium from yellow to red after extended incubation. Yeasts can be differentiated from the dermatophytes because they form smooth colonies without any mycelium. DA
4 Medium 2: All fungi (including yeasts) will grow on this medium. Cycloheximide-sensitive yeasts (Candida tropicalis, Candida parapsilosis, Candida krusei, Candida [=Torulopsis] glabrata and others) usually appear as smooth colonies on medium 2 without growth on medium 1. Note that certain dermatophytes on medium 2 may need longer incubation than on medium 1 or may grow only weakly. Note that further tests, such as microscopic and/or biochemical procedures are necessary to completely identify the pathogens isolated on this dipslide. 3,4 Most of these methods are only available in diagnostic laboratories and usually cannot be performed in a physician s office. Therefore, all dipslides with doubtful or clearly positive results should be sent to an analytical laboratory performing mycological tests. After use and prior to discarding, all used tubes and other contaminated material must be autoclaved or incinerated. For details, see GENERAL INSTRUCTIONS FOR USE document. PERFORMANCE CHARACTERISTICS AND LIMITATIONS OF THE PROCEDURE BD BBL Dermatoslide is a two-sided dipslide suitable for the isolation of fungi and must only be used for the recovery of fungi from superficial infections (skin, hair and nails). 2-5 Medium 1 is a standard selective medium for dermatophytes (e.g., Trichophyton, Epidermophyton, and Microsporum species). 3,5 Additionally, cycloheximide-resistant yeasts, such as Candida albicans will grow on the medium. This medium is inhibitory to bacteria. Certain pathogenic fungi, including certain strains of Microsporum, are inhibited by cycloheximide contained in this medium. However, these strains will grow on medium 2 of this dipslide. Medium 2 is a universal isolation medium for fungi, including saprophytic species. Most bacteria which may be present as contaminants or pathogens are inhibited by the low ph of this medium. 2,6 The number of species that may grow on the media of this slide system is large and is not restricted to those mentioned in the Results and Interpretation section. BD BBL Dermatoslide is intended to be used for primary isolation and counting. For further tests, and especially if mixed cultures occur on the media of this system, subcultures onto appropriate plated media must be made to obtain pure cultures which are mandatory for complete identification and susceptibility testing. 3,6 Dipslides must not be used for performing disc susceptibility tests. REFERENCES 1. Taplin, D., et al Isolation and recognition of dermatophytes on a new medium (DTM). Arch. Dermatol. 99: MacFaddin, J. D Media for isolation-cultivation-identification- maintenance of medical bacteria, vol. 1, p Williams & Wilkins, Baltimore, MD. 3. Summerbell, R.C Trichophyton, Microsporum, Epidermophyton, and agents of superficial mycoses. In: Murray, P. R., E. J. Baron, J.H. Jorgensen, M. A. Pfaller, and R. H. Yolken (ed.). Manual of clinical microbiology, 8th ed. American Society for Microbiology, Washington, D.C. 4. Larone, D.H. 1995: Medically important fungi - a guide to identification. 3 rd edition. ASM Press, Washington. 5. Atlas, R.M Handbook of Microbiological media. CRC Press, Boca Raton, FL. 6. Sutton, D.A Specimen collection, transport, and processing: mycology. In: Murray, P. R., E. J. Baron, J.H. Jorgensen, M. A. Pfaller, and R. H. Yolken (ed.). Manual of clinical microbiology, 8 th ed. American Society for Microbiology, Washington, D.C. PACKAGING/AVAILABILITY BBL Dermatoslide Cat. No slides DA
5 FURTHER INFORMATION For further information please contact your local BD representative. BD Diagnostic Systems Tullastrasse 8 12 D Heidelberg/Germany Phone: , Fax: Reception_Germany@europe.bd.com BD Diagnostic Systems Europe Becton Dickinson France SA 11 rue Aristide Bergès Le Pont de Claix/France Tel: Fax: BD, BD logo and BBL are trademarks of Becton, Dickinson and Company. ATCC is a trademark of the American Type Culture Collection Becton, Dickinson and Company DA
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