Comparison of Modified Fite-Faraco Staining, Auramine O Staining and Real-Time Polymerase Chain Reaction with Standard Staining Methods for Detection of Mycobacterium leprae in Slit-Skin Samples
Keywords:
Slit-skin smear, Acid fast bacilli staining, Fluorescence, Leprosy, Modified Fite-Faraco, Real-time PCRAbstract
Detection of Mycobacterium leprae by modified Fite-Faraco (FF), fluorescent Auramine O (AO) method, real-time polymerase chain reaction (PCR) of Pra gene and the standard Ziehl Neelsen (ZN) staining were compared in slit-skin samples from 32 leprosy patients attending the Raj Pracha Samasai Institute. Scraped incision method was performed to collect tissue fluid and dermal tissue from the same 4 sites including two from both ear lobes and two from the active lesions. Slit-skin smears (SSS) were prepared and stained by ZN, FF and AO methods. The collected specimens were evaluated for the present of Pra gene using real-time PCR method. Fite-Faraco was modified by omitting the deparaffinization instead submerge the slide in soybean oil for 10 minutes. There was a very good correlation among all methods. Out of 32 samples, 7 showed organisms by ZN method, 9 showed organisms by FF method, 11 showed organisms by AO method and 9 showed organisms by real-time PCR method. FF and AO stains were found to be superior to ZN in detecting leprae bacilli in SSS. Moreover, sensitivity was identical for real-time PCR, FF, and AO methods for multibacillary (MB) cases suggesting that real-time PCR, FF and AO techniques on the SSS could replace ZN without any loss of sensitivity. FF procedure uses simple equipment, easy to carry out, and convenient procedure. Therefore, the modified Fite-Faraco method described here can be used for the confirmation of the clinical diagnosed case of leprosy in resource-limited hospitals.
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