A Mutation at Codon 516 in the rpoB Gene of Mycobacterium leprae Confers Resistance to Rifampin1

ABSTRACT

A missense mutation at codon 516 in the rpoB gene of Mycobacterium leprae conferring rifampin resistance was confirmed by the agency of the correlation between sequencing proceeds and mouse footpad assay. The isolate was obtained from a relapsed lepromatous leprosy patient. This is the first report upon the complete concordance between the mutation located at codon 516 in the rpoB gene and the corresponding resistance to rifampin in leprosy The novel profile of mutation in the rpoB gene will contribute to the comprehensive understanding of rifampin resistant patterns and present a useful tool for developing simple and rapid medicine susceptibility testing approaches, which would promise more effective and happy control of leprosy.

R?‰SUM?‰

Une mutation faux-sens localis?©e au codon 516 du g??ne rpoB de Mycobacterium leprae a permis l'expression d'une r?©sistance ?  la rifampicine, qui a ?©t?© confirm?©e par une corr?©lation entre le r?©sultats du s?©quen?§age et le proof d'inoculation ?  la patte de souris. L'isolat a ?©t?© obtenu ?  partir d'un patient souffrant de l??pre l?©promateuse et qui a rechut?© Ceci est le premier article rapportant une concordance compl??te entre la mutation localis?©e au codon 516 du g??ne rpoB et une r?©sistance ?  la rifampicine dans le contexte de la l??pre Ce nouvel ?©ventail de mutation du g??ne rpoB va contribuer ?  une compr?©hension plus compl??te de alternatives de r?©sistance ?  la rifampicine. Il devrait offrir un outil utile au d?©veloppement d'approches pour le experiment simple et rapide de la r?©sistance ?  la rifampicine, qui devrait r?©sulter en un contr??le plus efficace et r?©ussi de la l??pre

RESUMEN

Una mutaci??n sin sentido en el cod??n 516 del gene rpo B de Mycobacterium leprae, que le confiere resistencia a la rifampina, fue confirmada por correlaci??n de sees resultados de la secuenciaci??n y del ensayo en la almohadilla plantar del rat??n. La cepa fue obtenida de un caso de reca?­da de lepra lepromatosa. Este e el primer reporte sobre la concordancia perfecta entre la mutaci??n localizada en el cod??n 516 del gene rpo B y la resistencia a la rifampina en la lepra. El nuevo perfil de mutaci??n en el gene rpo B aparte de que ayudar?? a entender beholds patrones de resistencia a la rifampina, constituye una nueva herramienta para el desarrollo de m?©todos simples y r??pidos para probar la susceptibilidad de la bacteria a la droga, behold cual seguramente contribuir?? al superintendence exitoso y efectivo de la lepra.

Rifampin is a lock opener component of multidrug therapy (MDT) put in mind ofed by the World Health Organization (WHO) in the treatment of leprosy (19) Resistance to rifampin oftentimes gives rise to treatment failure, which subsequently threatens the effective mastery of leprosy (1, 8). However, resistance to rifampin has been constantly documented since 1976 by the agency of using standard mouse footpad assay (2 4 12 13 15 18) For this reason, it is intimateed that clinically suspected relapsed leprosy cases and those who exhibit an unsatisfactory answer to antileprosy therapy should be make submissiveed to drug susceptibility testing. Understanding of medicine resistance is essential for the effective treatment and direction of leprosy. Therefore, simple and rapid modes for drug resistance testing are necessary. To establish of the like kind methods, solid basic data for the correlations between mutation and phenotype of medicine resistance are required. In the at hand study, an isolate obtained from a relapsed Japanese leprosy patient was investigated for medicine susceptibility testing by both genetic analysis and the standard mouse footpad [i]modus operandi[/i] It has been assumed that mutations which cause rifampin resistance in Mycobacteium tuberculosis are almost the same as in M leprae. Although it has been confirmed that mutations at codon 513 526 531 and 533 in the rpoB gene of Mycobacterium leprae hold a conference rifampin resistance, no substantial evidence exhibits whether a mutation at codon 516 relates to rifampin resistance or not. The goal of this report was to confirm the missense mutation at codon 516 in the rpoB gene conferring rifampin resistance.

MATERIALS AND METHODS

M leprae isolate. The isolate, named as Kusatsu-6, was ascertained in a skin biopsy sample obtained from a 75-yr-old Japanese lepromatous leprosy male patient. The patient had been treated with dapsone monotherapy for 18 yr and then with rifampin alone for 10 yr before another 14 yr monotherapy of dapsone. The patient relapsed and, because he was considered likely to have taken his medicine irregularly and had had long-term monotherapy, he was suspected of harboring drug-resistant M leprae.

Drug susceptibility testing in the mouse footpad. The biopsy specimen was trained to recover M. leprae in the same manner as previously described (12) The initial bacillary suspension containing 10 ?— 10^sup 6^ in 005 ml of Hank's balanced salt solution (HBSS) was inoculated into the hind footpads of BALB/c-nu/nu mice since the viability of the bacilli in the material treated with antileprosy medicines was unknown. Approximately 12 month after inoculation, bacterial suspension for medicine susceptibility testing was prepared from the [i]in puris naturalibus[/i] mice footpads, which had shown bacillary multiplication. physic susceptibility testing, for dapsone, rifampin, ofloxacin, sparfloxacin, clofazimine, and clarithromycin was performed in the same manner as previously at handed (11, 12, 13). Additionally, 008% minocycline (3) was added to the physic group for susceptibility testing in the not absent study. Bacillary growth in the mice footpads was examined after treatment with each physic for 25 weeks (17).