Application of a polymerase chain reaction for the detection of Mycobacterium leprae in skin tissues

Abstract

The polymerase chain reaction (PCR) based on the selective amplification of a 530-bp fragment of the gene encoding the proline-rich antigen of Mycobacterium leprae was applied on sections of fixed or frozen biopsy samples from leprosy patients. A simple procedure for the extraction of DNA from M. leprae in clinical specimens that provided suitable template DNA for amplification was developed. When PCR was applied on frozen sections, positive amplification in samples from all untreated acid-fast bacillus (AFB)-positive patients and in samples from 56% of the untreated AFB-negative patients could be detected, while biopsy samples from patients with skin diseases other than leprosy were all PCR negative. With neutral Formalin-fixed biopsy samples, positive amplification in 92% of the samples from untreated AFB-positive patients and in 61% of the samples from untreated AFB-negative patients could be detected by PCR. Biopsy samples exposed to mercuric chloride or nonbuffered formaldehyde containing fixatives were not suitable for application of PCR. This PCR holds promise as a tool for studies on M. leprae infection.

Diagnosing leprosy: revisiting the role of the slit‐skin smear with critical analysis of the applicability of polymerase chain reaction in diagnosis

“…No bacilli may be present in the sample, which is likely to be true in some bi-negative cases, or bacilli may no longer be viable, either because of a strong host immune response or because they have been killed by treatment. 12 By contrast, we cannot rule out the possibility that other factors, such as the age of the specimens, may also have an effect on the outcome of PCR. Negative results among PB patient samples may be explained by their intrinsically higher ratios of human genomic DNA to M. leprae DNA, which probably inhibits M. leprae DNA amplification.…”

Detection ofMycobacterium lepraein nasal mucosa biopsies by the polymerase chain reaction

“…Recent development of the PCR technique brought an unprecedented opportunity for fast, sensitive and specific detection of M. leprae in human beings. It was described initially for this pathogen in 1989 [14,21], being shown an useful and applicable method, and ever since it has been used in the most different clinical specimens as skin [4,5], nasal swab [8][9][10], bucal swab [10], and hair bulb [9].…”

“…The accurate diagnosis of leprosy is of fundamental importance to all aspects of leprosy epidemiology, case management, and prevention of disability [3]. The recent development of polymerase chain reaction (PCR) has brought an unprecedented opportunity for sensitive, specific, and rapid detection of M. leprae in clinical specimens [4,5].…”

A Modified PCR‐Based Method for Rapid Non‐Radioactive Detection of Clinically Important Pathogens

“…At present, PCR is used in the detection of different infectious agents, e.g. S. aureus (1,11), S. pneumoniae (6,19), Mycobacterium (7,18), Chlamydia trachomatis (3), Pseudomonas (2, 10), etc. Many published protocols utilize different modifications of a direct PCR ("nested," "multiplex," and the like) and can detect 103-104 bacteria during 4-5 hr.…”