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The ongoing transmission of leprosy in India is worrisome, and emerging drug resistance may be one of the factors responsible for the continued transmission of leprosy in India. Emerging cases of multidrug-resistant Mycobacterium leprae pose a great threat to eradication of leprosy and must be addressed with utmost priority. We report a case of multidrug-resistant M. leprae in a case of relapse where slit skin smear (SSS) was negative and histopathology was inconclusive. Drug resistance studies in leprosy are undertaken only in smear-positive relapse cases, and detection of this type of multidrug resistance in a case with negative SSS and innocuous histopathology is rather unusual and highlights the importance of undertaking drug resistance tests even in smear-negative cases of leprosy relapse. Resistance to ofloxacin (OFL) is also a cause for concern as OFL is one of the reserve drugs recommended for treatment of rifampicin-resistant strains.

Purulia is one of the high-endemic districts for leprosy in West Bengal (the eastern part of India). The annual new case detection rate (ANCDR) of leprosy in West Bengal is 6.04/100000 (DGHS 2019-20). Our earlier report provided evidence of secondary drug resistance in relapse cases of leprosy. The aim of the current study was to observe primary drug resistance patterns for dapsone, rifampicin, and ofloxacin amongst new leprosy patients from Purulia, West Bengal in order to better understand the emergence of primary resistance to these drugs.In the present study, slit-skin smear samples were collected from 145 newly diagnosed leprosy cases from The Leprosy Mission (TLM) Purulia hospital between 2017 and 2018. DNA was extracted from these samples and the Mycobacterium leprae genome was analyzed for genes associated with drug resistance by polymerase chain reaction (PCR), followed by Sanger sequencing. Wild-type strain (Thai-53) and mouse footpad-derived drug-resistant strain (Z-4) were used as reference strains.Of 145 cases, 25 cases showed mutations in genes associated with resistance to rifampicin, dapsone, and ofloxacin (as described by the World Health Organization, rpoB, folP, and gyrA, respectively) through Sanger sequencing. Of these 25 cases, 16 cases showed mutations in ofloxacin, two cases showed mutations in combinations of ofloxacin and rifampicin, four cases showed a mutation only in rifampicin, one case showed mutations in combinations of rifampicin and dapsone, and two cases showed mutations only in dapsone.Results from this study indicated the emergence of resistance to antileprosy drugs in new cases of leprosy. As ofloxacin is the alternate drug for the treatment of rifampicin-resistant cases, the emergence of new cases with resistance to ofloxacin indicates that ofloxacin-resistant M. leprae strains are actively circulating in this endemic region (i.e., Purulia, West Bengal), posing challenges for the effective treatment of rifampicin-resistant cases.

While Type 1 reaction is seen in leprosy patients, in up to 30% of patients, late reaction is uncommon and usually is seen in the first two years of release from treatment (RFT). It is believed tha...

BACKGROUND Leprosy relapse/recurrence is a serious concern particularly in a leprosy-endemic nation such as India. It is believed that bacilli persisting even after multidrug therapy can cause relapse; recently, however, drug resistance as a cause for recurrences and chronic erythema nodosum leprosum (ENL) has been speculated. AIM To study drug-resistance patterns in cases of leprosy relapse and chronic/recurrent (c/r)ENL. METHODS This cross-sectional study conducted over a period of 1 year included patients diagnosed as having leprosy relapse and those with c/rENL. Skin biopsy specimens were examined by conventional PCR for resistance testing for rifampicin, dapsone and ofloxacin, respectively targeting the rpoB, folP and gyrA genes of Mycobacterium leprae. RESULTS In total, 61 patients (25 smear-negative) were included in the study. Of these, 37 were diagnosed as having leprosy relapse and 24 as having c/rENL. Drug resistance to at least one drug was identified in 10 cases (16.4%). Rates of drug resistance were 5.4% (2 of 37) for dapsone, 10.8% (4 of 37) for rifampicin and 2.7% (1 of 37) for ofloxacin among cases of relapse, whereas it was 12.5% (3 of 24) and 8.3% (2 of 24) for dapsone and rifampicin respectively among those with c/rENL. Multidrug resistance was seen in 3.3% patients (2 of 61). CONCLUSION Drug-resistance rate among those with c/rENL was almost equalled that of relapse. Smear-negative leprosy relapse cases also had resistance to bactericidal drugs. These findings call for modifications in criteria for testing under leprosy drug-resistance surveillance and all cases of relapse and those with recalcitrant c/rENL should be tested.

Erythema multiforme (EM)-like erythema nodosum leprosum (ENL) is a rare atypical presentation, and its late appearance after the completion of multidrug therapy (MDT) is unusual. We describe the case of a lepromatous leprosy patient who after the completion of MDT presented to us with late EM-like ENL and was found to be resistant to rifampicin. We discuss the implications of this finding and the potential role of resistant bacilli in causing reactions with atypical presentations.

Introduction The most commonly noted reactions in leprosy patients are type 1 reactions and erythema nodosum leprosum, with some rare phenomenon of host response known as Lucio phenomenon or leprosy of Lucio and Latapi which is caused by Mycobacterium lepromatosis. So far, no case of M. lepromatosis has been reported from India. Materials and methods The main objective of this study was to detect any positive cases of M. lepromatosis in India with such a complication. We screened slit skin smear/biopsy samples from lepromatous leprosy (LL) patients reporting to The Leprosy Mission Community Hospitals across the country. Eighty-eight slit skin smears were collected from leprosy patients in 70% ethanol. DNA was extracted from all these samples. Polymerase chain reaction (PCR) was done for 2 genes; one set was for 16S rRNA and the other set was for coproporphyrinogen III oxidase (hemN) gene. Then, sequencing was done for all positive amplicons. Homology of the sequences was analyzed using the Basic Local Alignment Search Tool at the National Center of Biotechnology Information database. Results Among 88 isolates, we found 4 positive cases for M. lepromatosis. All 4 were LL cases with a bacteriological index ranging from 2+ to 4+. On the basis of the National Center of Biotechnology Information Basic Local Alignment Search Tool analysis, the sequenced amplicons of both genes matched with the M. lepromatosis 16S rRNA and phosphofructokinase genes but not with hemN gene of lepromatosis. This is the first report for the presence of M. lepromatosis in LL cases from India. Conclusion This new species M. lepromatosis exists beyond Mexico, Singapore and it is the cause of DLL in India also. It may cause dual infections along with M. leprae in endemic areas like India.

Despite more than three decades of multidrug therapy (MDT), leprosy remains a major public health issue in several endemic countries, including India. The emergence of drug resistance in Mycobacterium leprae (M. leprae) is a cause of concern and poses a threat to the leprosy-control program, which might ultimately dampen the achievement of the elimination program of the country. Rifampicin resistance in clinical strains of M. leprae are supposed to arise from harboring bacterial strains with mutations in the 81-bp rifampicin resistance determining region (RRDR) of the rpoB gene. However, complete dynamics of rifampicin resistance are not explained only by this mutation in leprosy strains. To understand the role of other compensatory mutations and transmission dynamics of drug-resistant leprosy, a genome-wide sequencing of 11 M. leprae strains - comprising five rifampicin-resistant strains, five sensitive strains, and one reference strain - was done in this study. We observed the presence of compensatory mutations in two rifampicin-resistant strains in rpoC and mmpL7 genes, along with rpoB, that may additionally be responsible for conferring resistance in those strains. Our findings support the role for compensatory mutation(s) in RNA polymerase gene(s), resulting in rifampicin resistance in relapsed leprosy patients.

The ongoing transmission of leprosy in India is worrisome, and emerging drug resistance may be one of the factors responsible for the continued transmission of leprosy in India. Emerging cases of multidrug-resistant Mycobacterium leprae pose a great threat to eradication of leprosy and must be addressed with utmost priority. We report a case of multidrug-resistant M. leprae in a case of relapse where slit skin smear (SSS) was negative and histopathology was inconclusive. Drug resistance studies in leprosy are undertaken only in smear-positive relapse cases, and detection of this type of multidrug resistance in a case with negative SSS and innocuous histopathology is rather unusual and highlights the importance of undertaking drug resistance tests even in smear-negative cases of leprosy relapse. Resistance to ofloxacin (OFL) is also a cause for concern as OFL is one of the reserve drugs recommended for treatment of rifampicin-resistant strains.

Early diagnosis of leprosy is important for limiting the severity of disease, which may lead to disabilities and deformities if not treated timely. Multiplex PCR employing more than one gene, specific to target DNA, is more efficient detection tool. In the present study, slit skin scrapings, blood, nasal swabs and saliva from Paucibacillary (PB) and Multibacillary (MB) cases as well as household contacts of PB cases were tested by multiplex PCR using three different gene targets namely RLEP, 16SrRNA and sodA. We found an increase in overall diagnostic positivity for M. leprae DNA detection by M-PCR as compared to individual PCR. In case of nasal swabs using M-PCR the PPV, NPV were 0.5454, 0.8333 respectively. There is remarkable increase in PPV in SSS of PB cases and nasal swabs of HHCs using M-PCR. Conclusively, our finding suggests the utility of M-PCR for early diagnosis and household contact surveillance for leprosy.

Non-tuberculous mycobacteria (NTM) are environmental mycobacteria found ubiquitously in nature. The present study was conducted to find out the presence of various species of NTM in leprosy endemic region along with Mycobacterium (M) leprae. Water and wet soil samples from the periphery of ponds used by the community were collected from districts of Purulia of West Bengal and Champa of Chhattisgarh, India. Samples were processed and decontaminated followed by culturing on Lowenstein Jensen (LJ) media. Polymerase chain reaction (PCR) was performed using 16S rRNA gene target of mycobacteria and species was confirmed by sequencing method. Indirect immune-fluorescent staining of M. leprae from soil was performed using M. leprae-PGL-1 rabbit polyclonal antibody. The phylogenetic tree was constructed by using MEGA-X software. From 380 soil samples 86 NTM were isolated, out of which 34(40%) isolates were rapid growing mycobacteria (RGM) and 52(60%) isolates were slow growing mycobacteria (SGM). Seventy-seven NTM isolates were obtained from 250 water samples, out of which 35(45%) were RGM and 42(55%) were SGM. Amongst all the RGM, we isolated M. porcinum, M. psychrotolerans, M. alsenase, M. arabiense and M. asiaticum from Indian environmental samples. M. fortuitum was the most commonly isolated species of all RGM. Out of all SGM, M. holsaticum, M. yongonense, M. seoulense, M. szulgai, M. europaeum, M. simiae and M. chimaera were isolated for the first time from Indian environment. M. intracellulare was the commonest of all isolated SGM. Presence of M. leprae was confirmed by indirect immunofluorescent microcopy and PCR method from the same environmental samples. Phylogenetic tree was showing a close association between these NTMs and M. leprae in these samples. Several NTM species of pathogenic and nonpathogenic in nature along with M. leprae were isolated from soil and pond water samples from leprosy endemic regions and these might be playing a role in causing disease and maintaining leprosy endemicity in India.

Background Leprosy is a chronic infectious disease caused by Mycobacterium leprae and mainly affects skin, peripheral nerves. Vitamin D receptor (VDR) gene polymorphism has been found to be associated with leprosy. Vitamin D has been shown to control several host immunomodulating properties through VDR gene. Vitamin D deficiency was also found to be linked to an increased risk for several infections and metabolic diseases. Objective In the present study, we investigated the association of VDR gene polymorphism, mRNA gene expression of VDR and the vitamin D levels with leprosy and its reactional states. Methodology A total of 305 leprosy patients consisting of tuberculoid (TT), borderline tuberculoid (BT), borderline lepromatous (BL), lepromatous leprosy (LL), as well as 200 healthy controls were enrolled in the study. We identified single nucleotide polymorphisms (SNPs) of VDR Taq1, Fok1 and Apa1, as well as the expression of VDR mRNA gene using PCR-based restriction fragment length polymorphism (RFLP) analysis and real-time PCR respectively. We also performed ELISA to measure vitamin D levels. Result We observed that SNP of VDR gene (Fok1 and Taq1) are associated with the leprosy disease. The allelic frequency distribution of T and t allele (p = 0.0037), F and f allele (p = 0.0024) was significantly higher in leprosy patients and healthy controls. ff genotype of Fok1 was found to be associated with leprosy patients [p = 0.0004; OR (95% CI) 3.148 (1.662–5.965)]. The recessive model of Fok1 genotype was also found to be significantly associated in leprosy patients in comparison to healthy controls [p = 0.00004; OR (95% CI) 2.85 (1.56–5.22)]. Leprosy patients are significantly associated with t-F-a haplotype. Further, VDR gene expression was found to be lower in non-reaction group compared to that of reaction group of leprosy and healthy controls. Paradoxically, we noted no difference in the levels of vitamin D between leprosy patients and healthy controls. Conclusion Blood levels of vitamin D do not play any role in clinical manifestations of any forms of leprosy. ff genotype of Fok1 and tt genotype of Taq1 was found to be associated with leprosy per se. Association of t-F-a haplotype with leprosy was found to be significant and could be used as a genetic marker to identify individuals at high risk for developing leprosy. VDR gene expression was lower in TT/BT and BL/LL groups of leprosy in comparison to that of healthy controls., Author summary Present study was carried out to find out the association of vitamin D receptor (VDR) gene polymorphism, mRNA gene expression of VDR gene and level of vitamin D with leprosy reactions and leprosy patients. Surprisingly, level of vitamin D in leprosy patients was not found to be associated with the disease and its manifestations. VDR genotypes (Fok1 and Taq1) were found to be associated with leprosy patients. t-F-a haplotype was significantly associated with leprosy patients. Gene expression of vitamin D receptor was lower in leprosy patients in comparison to healthy controls.

Despite more than three decades of multidrug therapy (MDT), leprosy remains a major public health issue in several endemic countries, including India. The emergence of drug resistance in Mycobacterium leprae (M. leprae) is a cause of concern and poses a threat to the leprosy-control program, which might ultimately dampen the achievement of the elimination program of the country. Rifampicin resistance in clinical strains of M. leprae are supposed to arise from harboring bacterial strains with mutations in the 81-bp rifampicin resistance determining region (RRDR) of the rpoB gene. However, complete dynamics of rifampicin resistance are not explained only by this mutation in leprosy strains. To understand the role of other compensatory mutations and transmission dynamics of drug-resistant leprosy, a genome-wide sequencing of 11 M. leprae strains – comprising five rifampicin-resistant strains, five sensitive strains, and one reference strain – was done in this study. We observed the presence of compensatory mutations in two rifampicin-resistant strains in rpoC and mmpL7 genes, along with rpoB, that may additionally be responsible for conferring resistance in those strains. Our findings support the role for compensatory mutation(s) in RNA polymerase gene(s), resulting in rifampicin resistance in relapsed leprosy patients.

Objectives The emergence of multidrug-resistant (MDR) organisms for any infectious disease is a public health concern. Global efforts to control leprosy by intensive chemotherapy have led to a significant decrease in the number of registered patients. Currently recommended control measures for treating leprosy with multidrug therapy (MDT) were designed to prevent the spread of dapsone-resistant Mycobacterium leprae strains. Here we report the identification of MDR M. leprae from relapse leprosy patients from endemic regions in India. Methods Resistance profiles to rifampicin, dapsone and ofloxacin of the isolated strains were confirmed by identification of mutations in genes previously shown to be associated with resistance to each drug. Between 2009–2016, slit-skin smear samples were collected from 239 relapse and 11 new leprosy cases from hospitals of The Leprosy Mission across India. DNA was extracted from the samples and was analysed by PCR targeting the rpoB, folP and gyrA genes associated with resistance to rifampicin, dapsone and ofloxacin, respectively, in M. leprae. M. leprae Thai-53 (wild-type) and Zensho-4 (MDR) were used as reference strains. Results Fifteen strains showed representative mutations in at least two resistance genes. Two strains showed mutations in all three genes responsible for drug resistance. Seven, seven and one strain, respectively, showed mutations in genes responsible for rifampicin and dapsone resistance, for dapsone and ofloxacin resistance and for rifampicin and ofloxacin resistance. Conclusion This study showed the emergence of MDR M. leprae in MDT-treated leprosy patients from endemic regions of India.