Felis ISSN 2398-2950

Fungal culture

Synonym(s): dermatophyte

Contributor(s): Catherine Fraser, David Godfrey

Overview

  • Dermatophyte test medium (DTM)  is a Sabouraud’s dextrose agar with cyclohexamide, gentamycin, chlortetracycline which inhibit non-dermatophyte growth, and phenol red as a pH indicator.
  • It is suitable for use in diagnosis of feline dermatophytosis Dermatophytosis.
  • The principle of the test is that pathogenic fungi utilize the protein in the medium and the resulting metabolites change the pH to alkaline and the medium from yellow to red.
  • Saprophytic fungi use carbohydrates first and no color change is seen. Once the carbohydrate is depleted, saprophytes will then utilize protein and a late color change is seen. Therefore it is important to observe the DTM daily and a color change either before or in association with the early visible growth of the colonies is positive.
  • The ideal is to also culture on a plain Sabouraud dextrose agar as this enables easier identification of the pathogen species. DTM inhibits formation of conidia - the best feature to identify the species; it can also make the gross colony morphologies less distinctive.

Sampling

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Tests

Methodologies

  • Culture is best at 30°​C but room temperature is acceptable. UV light reduces growth so a dark area is best.
  • A positive result may be seen in as little as 5 days but leave the culture for at least 14 days before treating as negative. The atmosphere can be too dry as well as too humid and a pan of water in the incubator will provide this ideal humidity.
  • Identifying particular dermatophytes is not often possible with DTM as macroconidia are not produced.
  • Setting up a subculture on Sabouraud’s dextrose agar will culture colonies that sporulate and mature macroconidia can then be identified using a fungal guide.
  • The most common dermatophytosis of cats is Microsporum canis.
  • Colonies of M. canis on Sabouraud’s agar  are white cotton/wool-like with an orange-yellow undersurface.
  • Once colonies are seen, take a small square of acetate tape, eg Scotch pressure sensitive tape® (3M), holding it in forceps touch it lightly to the surface of the culture then place it on a glass slide with a drop of lactophenol cotton blue and examine under x100 magnification. The macroconidia are abundant on Sabouraud’s media, for Microsporum canis they have thick walls with a terminal knob.

Availability

  • Fungal culture can be carried out in the practice situation as long as the test will be examined daily. DTM is commercially available, eg Dermaphyt® (Kruuse). Alternatively, all commercial laboratories will offer the test but samples should be submitted in paper or false-negative culture results may be seen due to bacterial overgrowth.

Validity

Sensitivity

  • Choosing sample material carefully remains the single most important determinant of success.

Specificity

  • Visual analysis of gross cultural morphology and microscopic analysis of hyphal form and macroconidial morphology is necessary for species identification and requires experience
  • These are more readily identifiable on colonies grown on plain Sabouraud medium.
  • It may be useful to submit positive DTM tests to a commercial laboratory for confirmation and identification of the species.

Technique (intrinsic) limitations

  • Fastidious cultural requirements.
  • Overgrowth by non-pathogenic fungi or bacteria on Sabouraud's dextrose agar plates.
  • Failure to identify species morphological characteristics on DTM.

Technician (extrinsic) limitations

  • Veterinarian or laboratory technician experience required.

Result Data

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Further Reading

Publications

Refereed Papers

Other sources of information

  • Miller W H, Griffin C E & Campbell L (2013) Diagnostic methods. In: Muller & Kirk’s Small Animal Dermatology 7th edn. Elsevier Inc, Missouri,  pp 86-91.
  • van Cutsem J & Rochette F (1991) Mycoses in Domestic Animals. Janssen Research Foundation.


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