Whole-Blood Nested-PCR Amplification of M. leprae-Specific DNA for Early Diagnosis of Leprosy

Abstract

Abstract. We evaluated the sensitivity and specificity of a nested-polymerase chain reaction (PCR) method for detection of Mycobacterium leprae DNA from whole blood. Whole-blood specimens were subjected to nested-PCR amplification of M. leprae repeat DNA sequences in 49 multibacillary (MB) and 30 paucibacillary (PB) leprosy patients, 96 household contacts (HHCs), 18 tuberculosis (TB) patients, and 35 normal healthy individuals. M. leprae DNA was detected in 95.92% (47/49) of MB, 70% (21/30) of PB, and 6.25% (6/96) of HHC, but it was not detected in 18 TB or 35 normal controls. The sensitivities of the anti-bovine serum albumin (ND-O-BSA) immunoglobulin M (IgM) and antifusion protein of ML0405-ML2331 IgG for MB were 97.96% and 89.8%, and these values for PB were 70% and 53.33%. However, the ND-O-BSA enzyme-linked immunosorbent assay (ELISA) had lower specificity, with relatively high false-positive results for TB patients (16.67%) and normal healthy controls (10%). Based on these promising findings, we propose the use of nested PCR of whole-blood samples along with ELISA test for early detection of leprosy cases.

Asymptomatic Leprosy Infection among Blood Donors May Predict Disease Development and Suggests a Potential Mode of Transmission

“…1).These six new cases were subjected to other confirmatory tests and presented positivity for M. leprae DNA in slit-skin smears and skin biopsy specimens, providing strong evidence that subclinical infection progressed to disease. The seropositivity observed in our study was slightly higher than that observed in a recent study sampling healthy controls (1/35), which assessed serum samples for antibodies against a PGL-1 synthetic antigen (ND-O-BSA) (6). It is important to emphasize that the case detection rate in the Uberlandia region was 10-fold higher than that in Yunnan province, and the absolute number of new leprosy cases detected in Brazil was 30-fold higher than the number detected in China (1).…”

“…Discrepancies may also be the result of our larger sampling. The same study did not report PCR positive results among the healthy controls, although household contacts presented a PCR positivity of 6.25% (6/96) (6). Not surprising, in addition to living in the same dwelling, household contacts often share similar conditions of nutrition, social status, hygiene, and, in some cases, consanguinity.…”

“…It is important to emphasize that the case detection rate in the Uberlandia region was 10-fold higher than that in Yunnan province, and the absolute number of new leprosy cases detected in Brazil was 30-fold higher than the number detected in China (1). Additionally, our blood donor population was more representative and does not present any specific subgroups, such as health care professionals or food handlers (6). Discrepancies may also be the result of our larger sampling.…”

“…It has recently been suggested that whole-blood nested-PCR amplification could be used for early diagnosis of leprosy (6) and that the presence of M. leprae DNA in peripheral blood may be associated with bacillary migration and a high risk for disease onset (7).…”

Application of RLEP Real-Time PCR for Detection of M. leprae DNA in Paraffin-Embedded Skin Biopsy Specimens for Diagnosis of Paucibacillary Leprosy

“…13 Recently, we have found that a nested PCR method that detects M. leprae DNA in patients' whole blood can be used for rapid and early diagnosis of leprosy. 14 In this study, we evaluated the use of a real-time PCR test targeting RLEP DNA sequence for adjunct diagnosis of PB leprosy in comparison to the nested PCR test 14 and also conventional histopathology for diagnosis of leprosy.…”