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Sporotrichosis is a chronic, cosmopolitan granulomatous mycosis that affects humans and animals. The infection is caused by the dimorphic fungi  Sporothrix  sp. The aims of the present study were to evaluate, standardize and validate a nested PCR technique using two DNA purification kits for the extraction of DNA from formalin fixed and paraffin-embedded tissues (FFPE) for  Sporothrix  sp. detection. FFPE mycological culture pellet samples of different  Sporothrix  species ( S. chilensis, S. mexicana, S. pallida, S. globosa, S. brasiliensis and S. schenckii ) were used as positive controls and clinical FFPE tissue samples of animals positive for  Cryptococcus  sp.,  Leishmania infantum  and  Histoplasma  sp. were used as negative controls. Ten clinical FFPE skin samples from cats with sporotrichosis were used to validate the nested PCR. These samples were cut into two distinct paraffin sectioning protocols (5 and 16 μm thick). The paraffin sections were subjected to two different DNA extraction kits (chemical and thermal extractions). A nested PCR was performed on the extracted DNA to identify the genus  Sporothrix . The chemical extraction protocol with the 5 μm thick paraffin section was more effective in extracting DNA from  Sporothrix  sp. from FFPE samples and the nested PCR technique showed the highest sensitivities (100% in the positive controls and of 50% in the skin samples of cats) and specificity (100%). Therefore, the nested PCR using this protocol has great potential to be applied in  Sporothrix  sp. diagnosis in FFPE samples of cats.

Nested PCR for the Diagnosis of Feline Sporotrichosis From Formalin-Fixed and Paraffin-Embedded Samples Using Different DNA Extraction Protocols