Pathogenicity and virulence of Mycobacterium leprae.

Leprosy is caused by Mycobacterium leprae (M. leprae) and M. lepromatosis, an obligate intracellular organism, and over 200,000 new cases occur every year. M. leprae parasitizes histiocytes (skin macrophages) and Schwann cells in the peripheral nerves. Although leprosy can be treated by multidrug therapy, some patients relapse or have a prolonged clinical course and/or experience leprosy reaction. These varying outcomes depend on host factors such as immune responses against bacterial components that determine a range of symptoms. To understand these host responses, knowledge of the mechanisms by which M. leprae parasitizes host cells is important. This article describes the characteristics of leprosy through bacteriology, genetics, epidemiology, immunology, animal models, routes of infection, and clinical findings. It also discusses recent diagnostic methods, treatment, and measures according to the World Health Organization (WHO), including prevention. Recently, the antibacterial activities of anti-hyperlipidaemia agents against other pathogens, such as M. tuberculosis and Staphylococcus aureus have been investigated. Our laboratory has been focused on the metabolism of lipids which constitute the cell wall of M. leprae. Our findings may be useful for the development of future treatments.

Host-Related Laboratory Parameters for Leprosy Reactions.

Leprosy reactions are acute inflammatory episodes that complicate the course of a Mycobacterium leprae infection and are the major cause of leprosy-associated pathology. Two types of leprosy reactions with relatively distinct pathogenesis and clinical features can occur: type 1 reaction, also known as reversal reaction, and type 2 reaction, also known as erythema nodosum leprosum. These acute nerve-destructive immune exacerbations often cause irreversible disabilities and deformities, especially when diagnosis is delayed. However, there is no diagnostic test to detect or predict leprosy reactions before the onset of clinical symptoms. Identification of biomarkers for leprosy reactions, which impede the development of symptoms or correlate with early-onset, will allow precise diagnosis and timely interventions to greatly improve the patients' quality of life. Here, we review the progress of research aimed at identifying biomarkers for leprosy reactions, including its correlation with not only immunity but also genetics, transcripts, and metabolites, providing an understanding of the immune dysfunction and inflammation that underly the pathogenesis of leprosy reactions. Nevertheless, no biomarkers that can reliably predict the subsequent occurrence of leprosy reactions from non-reactional patients and distinguish type I reaction from type II have yet been found.

Essential Roles of PPARs in Lipid Metabolism during Mycobacterial Infection.

The mycobacterial cell wall is composed of large amounts of lipids with varying moieties. Some mycobacteria species hijack host cells and promote lipid droplet accumulation to build the cellular environment essential for their intracellular survival. Thus, lipids are thought to be important for mycobacteria survival as well as for the invasion, parasitization, and proliferation within host cells. However, their physiological roles have not been fully elucidated. Recent studies have revealed that mycobacteria modulate the peroxisome proliferator-activated receptor (PPAR) signaling and utilize host-derived triacylglycerol (TAG) and cholesterol as both nutrient sources and evasion from the host immune system. In this review, we discuss recent findings that describe the activation of PPARs by mycobacterial infections and their role in determining the fate of bacilli by inducing lipid metabolism, anti-inflammatory function, and autophagy.

Mycobacterium leprae promotes triacylglycerol de novo synthesis through induction of GPAT3 expression in human premonocytic THP-1 cells.

Mycobacterium leprae (M. leprae) is the etiological agent of leprosy, and the skin lesions of lepromatous leprosy are filled with numerous foamy or xanthomatous histiocytes that are parasitized by M. leprae. Lipids are an important nutrient for the intracellular survival of M. leprae. In this study, we attempted to determine the intracellular lipid composition and underlying mechanisms for changes in host cell lipid metabolism induced by M. leprae infection. Using high-performance thin-layer chromatography (HPTLC), we demonstrated specific induction of triacylglycerol (TAG) production in human macrophage THP-1 cells following M. leprae infection. We then used [14C] stearic acid tracing to show incorporation of this newly synthesized host cell TAG into M. leprae. In parallel with TAG accumulation, expression of host glycerol-3-phosphate acyltransferase 3 (GPAT3), a key enzyme in de novo TAG synthesis, was significantly increased in M. leprae-infected cells. CRISPR/Cas9 genome editing of GPAT3 in THP-1 cells (GPAT3 KO) dramatically reduced accumulation of TAG following M. leprae infection, intracellular mycobacterial load, and bacteria viability. These results together suggest that M. leprae induces host GPAT3 expression to facilitate TAG accumulation within macrophages to maintain a suitable environment that is crucial for intracellular survival of these bacilli.

The function of peroxisome proliferator-activated receptors PPAR-γ and PPAR-δ in Mycobacterium leprae-induced foam cell formation in host macrophages.

Leprosy is a chronic infectious disease caused by Mycobacterium leprae (M. leprae). In lepromatous leprosy (LL), skin macrophages, harboring extensive bacterial multiplication, gain a distinctive foamy appearance due to increased intracellular lipid load. To determine the mechanism by which M. leprae modifies the lipid homeostasis in host cells, an in vitro M. leprae infection system, using human macrophage precursor THP-1 cells and M. leprae prepared from the footpads of nude mice, was employed. RNA extracted from skin smear samples of patients was used to investigate host gene expressions before and after multidrug therapy (MDT). We found that a cluster of peroxisome proliferator-activated receptor (PPAR) target genes associated with adipocyte differentiation were strongly induced in M. leprae-infected THP-1 cells, with increased intracellular lipid accumulation. PPAR-δ and PPAR-γ expressions were induced by M. leprae infection in a bacterial load-dependent manner, and their proteins underwent nuclear translocalization after infection, indicating activation of PPAR signaling in host cells. Either PPAR-δ or PPAR-γ antagonist abolished the effect of M. leprae to modify host gene expressions and inhibited intracellular lipid accumulation in host cells. M. leprae-specific gene expressions were detected in the skin smear samples both before and after MDT, whereas PPAR target gene expressions were dramatically diminished after MDT. These results suggest that M. leprae infection activates host PPAR signaling to induce an array of adipocyte differentiation-associated genes, leading to accumulation of intracellular lipids to accommodate M. leprae parasitization. Certain PPAR target genes in skin lesions may serve as biomarkers for monitoring treatment efficacy.

Quantitative evaluation of maxillary bone deformation by computed tomography in patients with leprosy.

BACKGROUND: Facial deformation as a sequela of leprosy is caused not only by a saddle nose but also by regression of the maxilla, as well documented in paleopathological observations of excavated skeletal remains of patients with leprosy. However, maxillary changes in living patients have been evaluated only by the subjective visual grading. Here, we attempted to evaluate maxillary bone deformation in patients with leprosy using three-dimensional computed tomography (3D-CT). METHODS: Three-dimensional images centered on the maxilla were reconstructed using multiplanar reconstruction methods in former patients with leprosy (n = 10) and control subjects (n = 5); the anterior-posterior length of the maxilla (MA-P) was then measured. The difference between the MA-P of the patients and those of controls was evaluated after compensating for individual skull size. These findings were also compared with those from previous paleopathological studies. FINDINGS: Three former patients with lepromatous leprosy showed marked atrophy of the maxilla at the prosthion (-8.6, -11.1 and -17.9 mm) which corresponded with the visual appearance of the maxillary deformity, and these results were consistent with paleopathological findings of excavated skeletal remains. Additionally, the precise bone defects of the maxilla could be individually calculated for accurate reconstructive surgery. INTERPRETATION: We have successfully illustrated maxillary bone deformities in living patients with leprosy. This study also confirmed the maxillary regression described in paleopathological studies.

Mycobacterium leprae genomes from naturally infected nonhuman primates.

Leprosy is caused by the bacterial pathogens Mycobacterium leprae and Mycobacterium lepromatosis. Apart from humans, animals such as nine-banded armadillos in the Americas and red squirrels in the British Isles are naturally infected with M. leprae. Natural leprosy has also been reported in certain nonhuman primates, but it is not known whether these occurrences are due to incidental infections by human M. leprae strains or by M. leprae strains specific to nonhuman primates. In this study, complete M. leprae genomes from three naturally infected nonhuman primates (a chimpanzee from Sierra Leone, a sooty mangabey from West Africa, and a cynomolgus macaque from The Philippines) were sequenced. Phylogenetic analyses showed that the cynomolgus macaque M. leprae strain is most closely related to a human M. leprae strain from New Caledonia, whereas the chimpanzee and sooty mangabey M. leprae strains belong to a human M. leprae lineage commonly found in West Africa. Additionally, samples from ring-tailed lemurs from the Bezà Mahafaly Special Reserve, Madagascar, and chimpanzees from Ngogo, Kibale National Park, Uganda, were screened using quantitative PCR assays, to assess the prevalence of M. leprae in wild nonhuman primates. However, these samples did not show evidence of M. leprae infection. Overall, this study adds genomic data for nonhuman primate M. leprae strains to the existing M. leprae literature and finds that this pathogen can be transmitted from humans to nonhuman primates as well as between nonhuman primate species. While the prevalence of natural leprosy in nonhuman primates is likely low, nevertheless, future studies should continue to explore the prevalence of leprosy-causing pathogens in the wild.

Paleopathological evidence and detection of Mycobacterium leprae DNA from archaeological skeletal remains of Nabe-kaburi (head-covered with iron pots) burials in Japan.

The Nabe-kaburi is a unique burial method, the purpose of which is shrouded in mystery. The burials were performed during the 15(th) to 18(th) centuries in eastern Japan, and involved covering the heads of the deceased with iron pots or mortars. The identification of leprosy-specific osteological lesions among some of the excavated remains has led to the suggestion that Nabe-kaburi burials were a reflection of the social stigma against certain infectious diseases, such as leprosy, tuberculosis or syphilis. However, molecular evidence for the presence of disease has been lacking. The goal of this study was to detect Mycobacterium leprae (M. leprae) DNA in archaeological human skeletal remains from Nabe-kaburi burials. The paleopathological data from three Nabe-kaburi burials were re-evaluated before small samples were taken from affected and control areas. DNA was extracted and used as a template to target the M. leprae-specific DNA using a combination of whole genome amplification, PCR analysis and DNA sequencing. M. leprae DNA fragments were detected in the two sets of skeletal remains that had also shown paleopathological evidence of leprosy. These findings provide definitive evidence that some of the Nabe-kaburi burials were performed for people affected by leprosy. Demonstration of the presence of M. leprae DNA, combined with archeological and anthropological examinations, will aid in solving the mystery of why Nabe-kaburi burials were performed in medieval Japan.

Leprosy as a model of immunity.

 Leprosy displays a spectrum of clinical manifestations, such as lepromatous and tuberculoid leprosy, and type I and II lepra reactions, which are thought to be a reflection of the host's immunological response against Mycobacterium leprae. Therefore, differential recognition of M. leprae, as well as its degraded components, and subsequent activation of cellular immunity will be an important factor for the clinical manifestation of leprosy. Although M. leprae mainly parasitizes tissue macrophages in the dermis and the Schwann cells of peripheral nerves, the presence of M. leprae in other organs, such as the liver, may also play important roles in the further modification of seesaw-like bipolar phenotypes of leprosy. Thus, leprosy is an exciting model for investigating the role of the human immune system in host defense and susceptibility to infection.

[Present leprosy situation in the world in 2012].

The epidemiological situation of leprosy is reported by the health division of each country to WHO. The reported data is collected by WHO and is immediately run on the Weekly Epidemiological Record. On this latest edition, data from the beginning of 2012 was reported. The Enhanced global strategy for further reducing the disease burden due to leprosy (plan period: 2011-2015) emphasizes reducing grade-2 disabilities among new cases. The sustained and committed efforts by the national programmes along with the continued support from national and international partners have led to a decline in the global burden of leprosy. It is important that all endemic countries continue to provide innovative solutions to address barriers to timely case detection and treatment completion, to ensure that the current declining trend is sustained.

Clofazimine modulates the expression of lipid metabolism proteins in Mycobacterium leprae-infected macrophages.

Mycobacterium leprae (M. leprae) lives and replicates within macrophages in a foamy, lipid-laden phagosome. The lipids provide essential nutrition for the mycobacteria, and M. leprae infection modulates expression of important host proteins related to lipid metabolism. Thus, M. leprae infection increases the expression of adipophilin/adipose differentiation-related protein (ADRP) and decreases hormone-sensitive lipase (HSL), facilitating the accumulation and maintenance of lipid-rich environments suitable for the intracellular survival of M. leprae. HSL levels are not detectable in skin smear specimens taken from leprosy patients, but re-appear shortly after multidrug therapy (MDT). This study examined the effect of MDT components on host lipid metabolism in vitro, and the outcome of rifampicin, dapsone and clofazimine treatment on ADRP and HSL expression in THP-1 cells. Clofazimine attenuated the mRNA and protein levels of ADRP in M. leprae-infected cells, while those of HSL were increased. Rifampicin and dapsone did not show any significant effects on ADRP and HSL expression levels. A transient increase of interferon (IFN)-ß and IFN-γ mRNA was also observed in cells infected with M. leprae and treated with clofazimine. Lipid droplets accumulated by M. leprae-infection were significantly decreased 48 h after clofazimine treatment. Such effects were not evident in cells without M. leprae infection. In clinical samples, ADRP expression was decreased and HSL expression was increased after treatment. These results suggest that clofazimine modulates lipid metabolism in M. leprae-infected macrophages by modulating the expression of ADRP and HSL. It also induces IFN production in M. leprae-infected cells. The resultant decrease in lipid accumulation, increase in lipolysis, and activation of innate immunity may be some of the key actions of clofazimine.

[Present leprosy situation in the world in 2011].

The epidemiological situation of leprosy is reported by the health division of each country to WHO. The reported data is collected by WHO and is immediately run on the Weekly Epidemiological Record. On this latest edition, data from the beginning of 2010 was reported. The Enhanced global strategy for further reducing the disease burden due to leprosy (plan period: 2011-2015) emphasizes reducing grade-2 disabilities among new cases. The burden of leprosy continues to decline globally as a result of sustained efforts carried out by national leprosy programmes along with continued support from both national and international partners. Improving the management of complications through the development of an effective referral service and increased community awareness about the disease will ensure that cases present for diagnosis at an early stage and will help reduce the disease burden further.

Essential role of hormone-sensitive lipase (HSL) in the maintenance of lipid storage in Mycobacterium leprae-infected macrophages.

Mycobacterium leprae (M. leprae), the causative agent of leprosy, parasitizes within the foamy or enlarged phagosome of macrophages where rich lipids accumulate. Although the mechanisms for lipid accumulation in the phagosome have been clarified, it is still unclear how such large amounts of lipids escape degradation. To further explore underlying mechanisms involved in lipid catabolism in M. leprae-infected host cells, we examined the expression of hormone-sensitive lipase (HSL), a key enzyme in fatty acid mobilization and lipolysis, in human macrophage THP-1 cells. We found that infection by live M. leprae significantly suppressed HSL expression levels. This suppression was not observed with dead M. leprae or latex beads. Macrophage activation by peptidoglycan (PGN), the ligand for toll-like receptor 2 (TLR2), increased HSL expression; however, live M. leprae suppressed this increase. HSL expression was abolished in the slit-skin smear specimens from patients with lepromatous and borderline leprosy. In addition, the recovery of HSL expression was observed in patients who experienced a lepra reaction, which is a cell-mediated, delayed-type hypersensitivity immune response, or in patients who were successfully treated with multi-drug therapy. These results suggest that M. leprae suppresses lipid degradation through inhibition of HSL expression, and that the monitoring of HSL mRNA levels in slit-skin smear specimens may be a useful indicator of patient prognosis.

Current status of leprosy: epidemiology, basic science and clinical perspectives.

Leprosy has affected humans for millennia and remains an important health problem worldwide, as evidenced by nearly 250 000 new cases detected every year. It is a chronic infectious disorder, caused by Mycobacterium leprae, that primarily affects the skin and peripheral nerves. Recent advances in basic science have improved our knowledge of the disease. Variation in the cellular immune response is the basis of a range of clinical manifestations. The introduction of multidrug therapy has significantly contributed to a decrease in the prevalence of the disease. However, leprosy control activities, including monitoring and prevention programs, must be maintained.

Chimpanzees used for medical research shed light on the pathoetiology of leprosy.

Leprosy is a chronic infectious disorder caused by Mycobacterium leprae, which mainly affects skin and peripheral nerves. It is classified as either paucibacillary or multibacillary based upon clinical manifestations and slit-skin smear results. It is speculated that leprosy develops after a long latency period following M. leprae infection. However, the actual time of infection and the duration of latency have never been proven in human patients. To date, four cases of spontaneous leprosy have been reported in chimpanzees who were caught in West Africa in infancy and used for medical research in the USA and Japan. One of these chimpanzees was extensively studied in Japan, and single-nucleotide polymorphism analysis for the M. leprae genome was conducted. This analysis revealed that the chimpanzee was infected with M. leprae during infancy in West Africa and the pathognomonic signs of leprosy appeared after at least 30 years of incubation. Analysis of leprosy in chimpanzees can contribute not only to medical research but also to the understanding of the pathoetiology of leprosy.

[Leprosy in a chimpanzee].

Leprosy is suspected to develop after a long period of latency following infection with Mycobacterium leprae (M. leprae) during infancy, but definitive proof has been lacking. We found a rare case of leprosy in a chimpanzee (Pan troglodytes) born in West Africa (Sierra Leone) and brought to Japan around 2 years of age. At 31, the ape started exhibiting pathognomic signs of leprosy. Pathological diagnosis, skin smear, serum anti-phenolic glycolipid-I (PGL-I) antibody, and by PCR analysis demonstrated lepromatous leprosy. Single-nucleotide polymorphism (SNP) analysis verified the West African origin of the bacilli. This occurrence suggests the possibility of leprosy being endemic among wild chimpanzees in West Africa, potentially posing a zoonotic risk.

[Present leprosy situation in the world in 2010].

The epidemiological situation of leprosy is reported by the health division of each country to WHO. The reported data is collected by WHO and is immediately run on the Weekly Epidemiological Record. On this latest edition, data from the beginning of 2010 was reported. In almost all of the highly endemic countries, control activities have been integrated within the general healthcare system. However, early case detection and prompt treatment with MDT remain the cornerstone of leprosy. In order to reduce the physical, mental and socioeconomic burden of leprosy, much remains to be done.

[Diagnosis of Hansen's disease].

The Leprosy Mailing List (LML) is an e-mailing list open to whoever interested in the field from all over the world. It acts as a forum for exchanging information related to Hansen's disease. It was established in February 2001 in Italy, and the present moderator of the LML is Dr. Salvatore Noto. He and his colleague have recently introduced an atlas for diagnosing Hansen's disease which they brought together information and photos collected through the LML. The atlas is divided into three sections, (1) Introduction, (2) Cardinal signs, and (3) Diagnosis and the clinical spectrum of leprosy, and they are all accompanied with relevant photos. This time, Dr. Noto kindly permitted us to translate the atlas into Japanese to be published in the Japanese Journal of Leprosy and posted in the Japanese Leprosy Association homepage. This article includes the translation and some of the most informative photos. For more information, please refer to the homepage where you will find all photos in the atlas.

Whole-genome expression analysis of Mycobacterium leprae and its clinical application.

The whole-genome sequence analysis of Mycobacterium leprae, which was completed in 2001, revealed the characteristics of this microbe's genomic structure. Half of the M. leprae genome consists of a limited number of protein-coding genes and the rest comprises non-coding regions and pseudogenes. We performed membrane array and tiling array analyses to analyze the gene-expression profile of the M. leprae genome and found that pseudogenes and non-coding regions were expressed similarly to coding regions at the RNA level. The RNA expressions were confirmed by real-time PCR analysis. Expression of these RNAs in clinical samples showed varying patterns among patients, thus indicating that the analysis of RNA expression patterns, including non-coding regions and pseudogenes, may be useful for understanding the pathological state, prognosis, and assessment of therapeutic progress in leprosy.