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Molecular and Classical Methods Compared for Leprosy Diagnosis

By Labmedica International staff writers
Posted on 27 Sep 2018
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Image: Auramine O stained Mycobacterium leprae in FFPE tissue section under ×40 objective of light-emitting diode fluorescence microscope A: Sample with high BI; B. Sample with low BI (Photo courtesy of Armauer Hansen Research Institute).
Image: Auramine O stained Mycobacterium leprae in FFPE tissue section under ×40 objective of light-emitting diode fluorescence microscope A: Sample with high BI; B. Sample with low BI (Photo courtesy of Armauer Hansen Research Institute).
Mycobacterium leprae is the causative agent of leprosy, a chronic granulomatous infectious disease affecting the skin and peripheral nerves. Leprosy manifests in various forms based on the immunological profiles and bacterial load in patients.

The diagnosis of leprosy mainly relies on clinical examination due to the inconsistent sensitivity and poor reproducibility of the current laboratory tests. Utilization of alternative methods to the standard Ziehl Neelsen (ZN), Fite-Faraco (FF) and Haematoxylin and Eosin (H&E) staining procedures may eventually improve leprosy diagnosis.

Ethiopian and Swiss scientists working at the Armauer Hansen Research Institute (Addis Ababa, Ethiopia) enrolled a total of 141 leprosy cases comprising 136 newly diagnosed treatment naïve and five relapse leprosy patients with any form of the disease in a prospective comparative cross-sectional study at the ALERT center from January 2015 to April 2016.

The team compared the performance of the fluorescent Auramine O (AO) staining and polymerase chain reaction (PCR) with different skin samples using a combination of ZN, FF and H&E staining as the gold standard. AO, ZN, FF, H&E and PCR tests were performed on slit skin smears (SSS). DNA was extracted from punch biopsies using two different methods with or without mechanical lysis. Punch biopsies in 10% formalin were kept for 48–72 hours before tissue processing was performed overnight using an automated tissue processor ASP 300S.

The team reported that the sensitivities were 87.6%, 59.3% and 77% for H&E, ZN and FF, respectively, whereas it reached 65.5% and 77.9% for AO in SSS and tissue sections and 91.1% for PCR in tissue samples. Moreover, samples with low bacillary index, sensitivity of AO staining (61.8%) was similar to FF (60%) and lower than PCR (86.6%). Sensitivity of PCR also increased (96.8%) when mechanical lysis was used during DNA extraction compared to enzymatic treatment alone (84.6%).

The authors concluded that their results showed that for diagnostic purposes, analysis of skin section is more sensitive than SSS, especially for samples with low bacillary load. AO staining on SSS and tissue sections was not significantly better than other routine diagnostic tests but considerably more user friendly. The sensitivity of PCR was higher than current standard methods and increased when combined with more efficient DNA extraction using mechanical and chemical lysis. They recommend AO staining for the diagnosis of leprosy in lower health facilities such as health centers and district hospitals and PCR diagnosis at referral level and research centers. The study was published on September 4, 2018, in the journal PLoS Neglected Tropical Diseases.

Related Links:
Armauer Hansen Research Institute


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