Genotyping of Mycobacterium leprae in Myanmar and possible transmission modes.

The polymorphism of TTC repeats in Mycobacterium leprae was examined using bacilli from slit skin samples of leprosy patients attending at Central Special Skin Clinic, Yangon General Hospital and nasal swabs of their contacts to elucidate the possible mode of leprosy transmission. It was found that bacilli with different TTC genotypes were distributed among same household contacts and also harbored bacilli in patients were different TTC genotype from that harbored on the nasal mucus of the healthy contacts. Genotypes of TTC repeats were found to differ between husband under treatment and his wife and also mother under treatment and her sons living in same house. This study revealed that TTC genotype of bacilli harbored by household contacts was different with the TTC genotype by index cases. These results indicate that the family members get transmission from outside the dwellings rather than from commonly supposed their MB index cases. There might have been some infectious sources to which the populace had been commonly exposed outside the dwellings.

FTA card utility for PCR detection of Mycobacterium leprae.

The suitability of the FTA® elute card for the collection of slit skin smear (SSS) samples for PCR detection of Mycobacterium leprae was evaluated. A total of 192 SSS leprosy samples, of bacillary index (BI) 1 to 5, were collected from patients attending two skin clinics in Myanmar and preserved using both FTA® elute cards and 70% ethanol tubes. To compare the efficacy of PCR detection of DNA from each BI class, PCR was performed to amplify an M. leprae-specific repetitive element. Of the 192 samples, 116 FTA® elute card and 112 70% ethanol samples were PCR positive for M. leprae DNA. When correlated with BI, area under the curve (AUC) values of the respective receiver-operating characteristic curves were similar for the FTA® elute card and ethanol collection methods (AUC=0.6). Taken together, our results indicate that the FTA® elute card, which enables the collection, transport, and archiving of clinical samples, is an attractive alternative to ethanol preservation for the detection of M. leprae DNA.

A novel method for simple detection of mutations conferring drug resistance in Mycobacterium leprae, based on a DNA microarray, and its applicability in developing countries.

A simple method to detect mutations in the genome of Mycobacterium leprae that confer resistance to key drugs for leprosy was exploited on the basis of a reverse hybridization system. A series of oligonucleotide probes corresponding to each mutation in the folP1, rpoB and gyrA genes for dapsone, rifampicin and ofloxacin resistance, respectively, were selected and fixed on a glass slide as capture probes, to develop a DNA microarray termed the leprosy drug susceptibility-DNA microarray (LDS-DA). Mutations in clinical isolates of M. leprae were successfully identified by the LDS-DA. Feasibility studies were conducted to evaluate the performance of the LDS-DA in two developing countries, Myanmar and the Philippines. The high concordance of the results obtained by this method with the results of nucleotide sequencing strongly supports the applicability of the LDS-DA as a drug susceptibility test in place of sequencing, a time-consuming and costly procedure. This is a rapid and simple method for the simultaneous susceptibility testing of three front-line drugs for leprosy, and solves the problems of previously reported methods.

Clinical analysis of multibacillary leprosy patients after 1-year fixed World Health Organization recommended multidrug therapy at Yangon General Hospital, Myanmar.

This study included 200 randomly selected multibacillary leprosy cases who had completed 1 year of fixed World Health Organization recommended multidrug therapy (WHO-MDT) without prior dapsone (DDS) monotherapy. The time interval after release from treatment varied from a few months to 8 years. All cases were clinically reviewed in 2006 by comparison with their old clinical records. Reactions, particularly reversal reactions, occurred frequently among patients who had completed MDT within the last 3 years. It was difficult to distinguish relapse cases and late reversal reactions in skin smear-negative multibacillary cases. Based on bacteriological and histological analyses, one patient was confirmed to have relapsed 1 year after release from treatment. The overall relapse rate was 0.5%. No drug resistance mutations were detected by polymerase chain reaction or dot blot hybridization. The present study indicates that it is important to follow up patients for several years after completion of MDT in order to detect possible lepra reactions and relapses.

The frequency of drug resistance mutations in Mycobacterium leprae isolates in untreated and relapsed leprosy patients from Myanmar, Indonesia and the Philippines.

INTRODUCTION: The magnitude of drug resistance in Mycobacterium leprae to dapsone, rifampicin, and ofloxacin was studied in three Southeast Asian countries with a high prevalence of leprosy. METHODS: M. leprae from the skin of leprosy patients was collected in North Maluku and North Sulawesi in Indonesia, Yangon in Myanmar, and Cebu in the Philippines. Mutations in the drug resistance determining regions in the folP1, rpoB, and gyrA genes, which have been proven to confer resistance, were analysed. In addition, samples from 51 newly diagnosed cases and 13 patients with leprosy relapse in Cebu were submitted for susceptibility testing in the mouse footpad. RESULTS: Of 252 isolates obtained from new cases, 3% were dapsone resistant and 2% were rifampicin resistant. In samples taken from patients with relapsed leprosy (n = 53), significantly more resistance mutations were detected: 15% had dapsone resistance mutations, and 8% had rifampicin resistance mutations. Two patients with relapsed leprosy had mutations for both dapsone and rifampicin resistance. No mutations conferring quinolone resistance were detected. No mutations were detected in the folP1 gene of M. leprae isolates with a low degree of resistance to dapsone. DISCUSSION: Detection of drug-resistant cases by mutation detection in the drug resistance determining region of the genome is a practical method for monitoring resistance. A comparison of the results obtained in this study with previous data obtained prior to the use of multidrug therapy (MDT), does not indicate clearly whether the magnitude of drug resistance has changed. Larger studies of resistance mutations in M. leprae isolated from patients with relapsed leprosy are needed to confirm our results. CONCLUSION: We recommend monitoring the magnitude of drug resistance globally, by testing M. leprae DNA from relapse cases and a representative sample of new cases.

Genotypic analysis of Mycobacterium leprae isolates from Japan and other Asian countries reveals a global transmission pattern of leprosy.

The genotype of single-nucleotide polymorphism type 3, CTC, at positions 14676, 164275, and 2935685, along with four copies of 6 bp repeats in the rpoT gene, was predominant for isolates originating in the Japanese mainland. Type 1, CGA, type 2, CTA, and type 3 were detected from Korea, Indonesia, and Myanmar. No isolates with four copies of 6 bp were detected from Myanmar, Okinawa, and Japanese Brazilian patients. Type 4, TTC, with three copies of 6 bp, was detected only from Japanese Brazilians. The results indicate that infection occurred in Brazil and the disease developed later in Japan.