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Efficacy of in-house fluorescent stain for fungus

March 15 2017

­­Increasing numbers of immunocompromised patients means that correctly diagnosing fungal infections is becoming more and more critical. Conventional diagnostic methods may not be the most effective, especially when there are small amounts of the causal agent in a sample. In a recent study of diagnostic tests for fungal infections, Kirani and Chandrika (2017) looked at the efficacy of an in-house calcofluor white (CFW) fluorescent stain, compared to conventional CFW stain, histopathology and culture.
The new stain was composed of the whitener  Ranipal which was first used to stain teased fungal cultures and because of high non-specific fluorescence the brightener Robin blue was added. They also determined the sensitivity, specificity, negative predictive value and positive predictive value. The authors compared the results of the in-house fluorescent stain to the conventional stain with 100 cases of suspected dermtophytosis and found that the tests agreed perfectly (35 positive, 65 negative). The samples were then compared using culture, which found 30 positive samples and 70 negative samples. They also compared the results 15 tissue section smears from clinically diagnosed fungal infections using the in-house stain and standard histopathological techniques. Using the in-house stain, 12 samples showed evidence of fungal infections (Aspergillus, Mucor, Candida, Rhinosporidium seeberi and Pneumocysttis carinii) whereas conventional histopathology identified only 10 positive samples. 

Figure 2: In-house fluorescent stain of skin scrapings Figure 1: Conventional calcofluor white staining of skin scrapings
a. In-house fluorescent stain                  b. Conventional calcofluor white 

Image source:  Kirani and Chandrika (2017)                          

These results suggest that in-house fluorescent microscopy can be a valuable tool for the diagnosis of fungal infections. There are several advantages of an in-house fluorescent stain over conventional methods: ease and speed & safety of use.  In comparison, in-house stain is cheap, readily available and not harmful. In addition, small numbers of organisms in a smear can be identified more easily, so the test is more sensitive. Samples such as skin, hair, and nails require pre-treatment with KOH. Biopsy samples require deparaffinization and pretreatment with KOH before staining. 
In developing countries where conventional CFW is prohibitively expensive or not available, the in-house stain provides an excellent, low-cost alternative which has been shown to be more sensitive than routine methods.