Resumo

Molecular determination of the viability of M. leprae in paucibacillary patients using 16S RNA B. G. C. Sartori 1, A. J. Finardi1, E. B. Moraes1, P. S. Rosa 1, S. Madeira-Diório 1, D. C. Nascimento 1, M. Moraes 2, I. M. F. Dias-Baptista1,* 1Instituto Lauro de Souza Lima, Bauru, 2FIOCRUZ, Rio de Janeiro, Brazil Preferred Presentation Method: Poster only Introduction: The impossibility of cultivating Mycobacterium leprae in axenic culture media has hampered in vitro studies and the development of clinical trials due to the slow multiplication of the bacillus. The present study aims at assessing the viability of bacilli through the detection of 16S rRNA specific for M. leprae with the normalization of the total quantities of mycobacteria from the quantification of total M. leprae DNA using the RLEP repetitive sequence. Methods: A total of 17 skin biopsy specimens were obtained for leprosy patients diagnosed and classified according to the Ridley & Jopling criteria. TaqMan real-time PCR assays for detection of M. leprae RNA and DNA were compared with inoculation into mice footpads (Shepard’s technique). Results: The initial assessments were on therapeutic efficacy to analyze viability of bacilli in clinical samples after 0 to 12 months of MDT. A preliminary experiment was performed to detect M. leprae viability from skin biopsy MB and PB leprosy patients. The 16S rRNA/RLEP results were positive for all analyzed biopsy samples (Ct ranging from 21.9 to 36.1) including paucibacillary. The inoculation into mice footpads were positive but with Ct ranging from 29.8 to 36.9. The samples after treatment have not yet been processed. Conclusion: The differentiation between the viable and nonviable M. leprae is very important for correct prognoses of leprosy in patients on treatment, determination of drug resistance or identification of relapse. Thus, the 16S rRNA assay is an additional molecular tool for leprosy diagnosis and determination of the viability of the leprosy bacillus. Funding: FAPESP (2010/03693-9)