Historical review of leprosy
The leprosy bacillus (Mycobacterium leprae) was first discovered by the Norwegian physician Gerhard Armauer Hansen in 1873, and thus the bacillus was also called Hansen's bacillus. Although this was the first human bacterial pathogen to be described, it remains to be one of the least understood pathogens. This is because it was not possible to grow the bacillus in culture .
Origin of leprosy
Leprosy is a disease of great antiquity, having been recognized from Vedic times in India and from Biblical times in the Middle East, with case reports from almost over 3000 years ago (http://www.who.int/topics/leprosy/en/). It probably originated in the tropics of Asia and Africa and spread to the rest of the world. Leprosy has always been held in superstitious dread, and the person suffering from leprosy was considered ‘unclean’ and a social outcast. Even now, leprosy is endemic in many regions of the world and a public health problem in India. The first case of leprosy was reported in Rio de Janeiro, Brazil, in 1600 .
The genus Mycobacterium contains at least 54 recognized species. The majority of these are saprophytes, but some do cause disease in humans. The most important ones are M. leprae and Mycobacterium tuberculosis. Stefansky (1905) described Mycobacterium lepraemurium as the cause of leprosy in rats. Tuberculosis in humans was subsequently shown to be caused by two types of bacteria (human and bovine). Three other types of tubercle bacilli were also recognized: Mycobacterium microti from voles; Mycobacterium avium from birds; and Mycobacterium jiscium, Mycobacterium marinum, Mycobacterium ranae and others from fish, lizards, snakes, frogs and cold-blooded animals. Johnes (1985) described Mycobacterium paratuberculosis from cattle. Mycobacteria were isolated from skin ulcers for the first time in Australia (Mycobacterium ulcerans, 1948) and later in Sweden (Mycobacterium balnei, 1954), Uganda (Mycobacterium buruli, 1964) and elsewhere .
Saprophytic mycobacteria have been isolated from a number of sources including Mycobacterium butyricum from butter, Mycobacterium phlei from grass, Mycobacterium stercoris from dung and Mycobacterium smegmatis from smegma. Atypical mycobacteria occur in soil, water and other environmental sources (including photochromogens, scotochromogens, nonphotochromogens and rapid growers).
Cause of leprosy
M. leprae, the causal agent of leprosy, is found as a strongly acid-fast bacillus with rounded ends measuring 1.5–8 μm in length and 0.2–0.5 μm in diameter. They are present in different forms in vivo, like masses within the lepra cells called globi or as bundles of cigars or in a palisade arrangement. M. leprae stains poorly or not at all by Gram staining or as beaded Gram-positive bacilli (http://www.who.int/topics/leprosy/en/) . Those bacilli that stain as solid acid-fast rods with carbol fuchsin stain are believed to be viable, whereas those bacilli that stain irregularly are dead and degenerating (http://www.who.int/topics/leprosy/en/) .
Electron microscopic studies of the M. leprae cell wall structure concluded that the cell wall is composed of peptidoglycans bound to branched-chain polysaccharides, consisting of arabinogalactans, which support mycolic acids, and lipoarabinomannan. The outermost structure is the capsule that has lipids, especially phthiocerol dimycocerosate and phenolic glycolipid (PGL-1), which has a trisaccharide bound to lipids by phenol. This trisaccharide is antigenically specific for M. leprae.
On examination of the M. leprae genome, it appears that this genome is much smaller than that of M. tuberculosis and rich in inactive or deleted genes, thereby explaining the difficulty in maintaining this bacterium in axenic culture. It has 2770 genes, with a coding percentage of 49.5% and a molecular weight of 2.2 × 109 Da, with 3 268 203 base pairs (bp) and a guanine + cytosine content of 57.8% .
Epidemiology of leprosy
Leprosy has been a major public health problem in tropical countries for many decades. Nearly half a million cases are estimated to exist worldwide, mainly in the Asian and African subcontinents. Countries where leprosy continues to be a major problem include Brazil and India. Although no nonhuman sources of infection have been established, naturally occurring infection in monkeys and armadillos has been reported . Leprosy is one of the most common peripheral neuropathies worldwide, with a global prevalence of 1.4 per 10 000 population. It is most common in tropical and subtropical regions of south and south-east Asia, Africa, Latin America, eastern Pacific and western Mediterranean; 82% of all reported cases are found in Brazil, India, Indonesia, Myanmar and Nigeria, but the prevalence has decreased conspicuously owing to the introduction of multiple drug therapy (MDT) in the 1980s. India has not been able to eliminate leprosy as yet and ranks first in terms of absolute number of cases [1,4]. The report of the National Leprosy Elimination Programme in March 2012 states that there were about 0.13 million cases of leprosy in India, 9.7% of which were children. A child leprosy survey conducted in the rural and urban areas of western Maharashtra yielded a prevalence rate of 10.5 and 1.5 per 10 000, respectively. The prevalence rate of leprosy in Brazil is 1.54 cases per 10 000 inhabitants, with 33 955 new cases in 2011, 61% of which were multibacillary [1,3].
Mechanisms of leprosy transmission
Leprosy is transmitted by close and prolonged contact through inhalation of the bacilli contained in nasal secretions or through skin erosions. The main route of transmission is the nasal mucosa. Other transmission routes including blood, vertical transmission, breast milk and insect bites are also possible . The presence of specific DNA sequences of M. leprae in nasal swabs or biopsies in healthy individuals living in endemic areas suggests that the carrier plays a significant role in the transmission. M. leprae-like organisms have also been reported to be present in soil. The mode of transmission of leprosy has not been fully established, but the nose and skin are considered as the main portals of exit as well as entry .
Clinical classification of leprosy
The disease is formally classified into a range of subtypes for the purposes of correct implementation of MDT; in 1982, WHO classified leprosy as multibacillary or paucibacillary. These are the two indices, based on the observation of nasal smears or skin smears, which help in assessing the amount of infection, viability of bacilli and disease prognosis.
Leprosy patients were diagnosed and classified clinically, bacteriologically and histopathologically, according to the Ridley–Jopling scale as lepromatous, borderline lepromatous, mid-borderline, borderline tuberculoid, tuberculoid, suspicious (or) early intermediate (I). Multibacillary patients were defined as mid-borderline, borderline lepromatous or lepromatous having a bacterial index of greater than 0 whereas borderline tuberculoid, tuberculoid patients with bacterial index of 0 were defined as paucibacillary .
The bacterial index is the only objective way of monitoring the benefits of treatment, and it represents the bacterial load. The index is calculated by totalling the number of positives (+) given to each smear and dividing this number by the smears collected. A minimum of seven sites should be examined; smears from four skin lesions or nasal swab and smears from both ear lobes. The results are expressed on a logarithmic scale (http://www.who.int/topics/leprosy/en/).
1. 1+ At least 1 bacillus in every 100 fields.
2. 2+ At least 1 bacillus in every 10 fields.
3. 3+ At least 1 bacillus in every field.
4. 4+ At least 10 bacilli in every field.
5. 5+ At least 100 bacilli in every field.
6. 6+ At least 1000 bacilli in every field.
In paucibacillary leprosy, the bacterial index is less than 2, and in multibacillary leprosy, it is greater than 2.
The type of leprosy and the bacterial index are the main patient-related factors that determine the transmission of leprosy among contacts.
During the course of microscopic examination of smears, it is possible to distinguish and count the number of solid staining organisms (organism that stain completely) and irregularly staining bacilli (organisms which do not stain completely). The percentage of solid-staining bacilli in a stained smear is referred to as the morphological index. The total of the morphological index for all sites divided by the number of sites gives the average morphologic index for the patient .
Based on the clinical criteria that uses the number of skin lesions (macules, papules and nodules) and nerve damage, leprosy is classified as single-lesion paucibacillary leprosy (one skin lesion with no nerve involvement), paucibacillary leprosy (two to five skin lesions asymmetrically distributed with one nerve involvement and definite loss of sensation) and multibacillary leprosy (many skin lesions symmetrically distributed with many nerve involvement).
The bacillus is spread via respiratory droplets and contact with active skin lesion; humans are the principal reservoirs [6–11]. The organism binds lamin-2 to α-dystroglycan on Schwann cells. They tend to infect skin and peripheral nerves in cooler areas of the body such as chin, malar eminences, earlobes, knees and distal extremities . They proliferate best at 80°F (30°C). The incubation period can range from 3 to 10 years between infection and clinical manifestation of disease. Although respiratory droplets and contact with active skin lesion are believed to be the main modes of transmission, many patients have no identifiable contacts [8,9]. The two extreme ‘polar’ forms of the disease are the lepromatous and tuberculoid types.
Tuberculoid leprosy is a relatively benign, yet resistant form of the disease. Neurological manifestations are characterized by patchy cutaneous sensory loss, particularly to temperature rather than pinprick or light touch. Sensory losses are confined to areas of rash but are followed by multiple mononeuropathies, with large nerve sensory and motor involvement. The nerves affected are primarily the pressure/trauma-dependent nerves, with the ulnar nerve involved in most sural, radial and branches of the facial nerve (lagopthalmos). There can be ulnar and posterior auricular nerve thickening, ulnar nerve abscesses and painless injuries. The typical axonal damage that occurs in tuberculoid leprosy is the result of infiltrating inflammatory cells and granuloma. It is multifocal and often leads to a total obliteration of fibres. The thickened perineum contains inflammatory infiltrates and often becomes fused to the epineurium. The granulomata are made up of epitheloid histocytes multinuclear plasma cells and lymphocytes. Frequently, nerves become caseated. Old lesions show hyalinized fibrotic fascicles with only a few intact axons .
In contrast to the tuberculoid form, nerve damage in lepromatous leprosy is widespread and symmetrical, with extensive intracutaneous nerve involvement. It can resemble distal symmetric polyneuropathy, and areas of anhydrosis, alopecia and anaesthesia are not uncommon. There is a poor correlation between cutaneous lesions and anaesthesia. Sensory loss tends to occur in the coolest areas of the body (ears, dorsal surfaces of the hands and feet, dorsolateral forearms and anterolateral parts of the legs).
Early in the disease course, the palms and soles of the feet and deep tendon reflexes are spared. Later, large nerve trunks become involved, and symptoms extend to the cool surface of the face, corneal anaesthesia, with impaired closure of the eyes can lead to painless ulcers. It involves more severe cutaneous manifestations, with nodules, macules and papules, and extensive skin infiltrates occurring with or without sensory loss. There can be alopecia over all skin lesions, loss of eyebrows and lashes (madarosis) and hypertrophy of the infiltrated facial skin (leonine faces). Primary damage to the nerves is the result of direct infiltration of the organism into macrophages, Schwann cells, perineural cells, fibroblasts, endothelium and infrequently the axons. Foamy cells are the result of macrophages and Schwann cells filled with the bacillus and other debris. Early in the disease course, the overall axonal architecture is preserved; there is only a mild inflammatory reaction and an absence of lymphocytes, giant cells and granulomata, as well as segmental demyelination. Later ‘onion-skinning’ of the axon can be seen, caused by a separation of perineural layers, by infiltrating foamy macrophages. There is also fibroblast proliferation and collagen deposition. Ultimately, collagen and hyaline material replace the nerve fibres .
Symptoms and signs pertaining to involvement of the skin and nerves are most commonly encountered, including hypopigmented macules and sensory loss . At least two of the following findings have to be present for a clinical diagnosis of leprosy.
1. A characteristic patch or skin lesion with impaired sensations.
2. A thickened (or) tender cutaneous or peripheral nerve with impairment of sensations in the area supplied by it.
3. Acid-fast bacteria in the skin smear.
Reversal reaction/reactions and relapses
Reversal reactions are downgrading response, a weakening over time of cell-mediated immunity with conversion from tuberculoid type leprosy to lepromatous type. Borderline leprosy has characteristics of both tuberculoid and lepromatous types, most likely because of a downgrade or reversal reaction, with a gradual conversion to lepromatous leprosy as a result of a decline in cell-mediated immunity .
Reversal reactions are hypothesized to be brought about by disturbances in immunological balance as a result of immune reactivity to M. leprae antigens. Identical antigenic determinants of the host might also contribute to the autoimmune phenomenon. Three types of reactions are recognized.
1. Type I (reversal) reaction associated with changes in morphological index, borderline tuberculoid, mid-borderline and borderline lepromatous; patients usually suffer from these reactions; when not promptly treated it results in nerve damage and deformities.
2. Type II reaction (erythema nodosum leprosum reactions) affecting multibacillary patients.
3. The Lucio phenomenon, which is less well understood, associated with extensive skin necrosis owing to acute vasculitis and occlusion of arterioles whose endothelium is massively invaded by M. leprae cells.
Relapse in leprosy is defined as recrudescence of the disease activity after successful completion of a prescribed course of therapy. This is considered to result from renewed multiplication of a few live organisms .
Early diagnosis of subclinical or carrier state leprosy has been problematic. Although M. leprae can be detected in the nasal mucosa of people who have been exposed to leprosy, this finding predicts neither clinical disease nor infectivity . The diagnosis of other forms of leprosy is usually made on clinical grounds (i.e. the finding of anaesthetic skin lesions in the presence of thickened peripheral nerves). However, demonstration of acid-fast bacilli in slit skin smears provides laboratory confirmation of the diagnosis of multibacillary disease . Until recently, the only method for studying immunity in leprosy was a skin test for delayed hypersensitivity, the lepromin test first described by Mitsuda . Present-day serological diagnosis of leprosy relies solely on detecting immunoglobulin (Ig) M antibodies to PGL-1 using synthetic neoglycoconjugate surrogate of PGL-1. Such serology in the past relied on ELISA. In recent years, there have been major developments in the creation of kits, such as the M. leprae dipstick and a 10-min Leprosy Lateral Flow Test Kit The concordance with conventional ELISA is excellent. Prospects for antigens with serological sensitivity and specificity greater than PGL-1 are dim. PGL-1 in its various formats is probably the best serological tool for leprosy that can be developed.
IgG and IgM anti-PGL-1 antibodies and IgG anti-lipoarabinomannan-B antibodies were measured by indirect ELISA. IgM anti PGL-1 antibodies were also measured by a gelatin particle agglutination test .
Assay of interferon-γ and other cytokines
T-cell response can be easily measured using delayed type hypersensitivity skin testing. Other alternatives include the assay of interferon-γ and other cytokines in whole blood as a measure of T-cell responses and development of a simple dipstick assay for interferon-γ to complement the detection of anti-PGL-1 antibodies. Such assays can also be used to measure other cytokines such as interleukin-5, interleukin-10 or transforming growth factor beta whose production may distinguish individuals developing clinical disease from exposed individuals who will remain healthy. At present, such whole blood assays provide a suitable tool that addresses T-cell responses on a limited immunoepidemiological scale .
The advent of PCR technology has provided unparalleled sensitivity and specificity for the diagnosis of leprosy. Diagnostic PCR-based assays are highly sensitive and specific, able to detect small numbers of M. leprae in various types of clinical specimen, such as slit skin smears, skin biopsies and nasal swab specimens, as well as M. leprae in the environment. However, PCR in the leprosy context is confined to the research arena.
During the last 10 years, several PCR methods have been developed to amplify different gene sequences of M. leprae. Molecular biological methods have renewed interest in a variety of techniques such as DNA-PCR, reverse-transcription PCR, nested PCR, Nucleic Acid Oligonucleotide-Based amplification and modern quantitative amplification (real-time PCR). Many kinds of PCR amplification have been included in leprosy research: gene amplification of a 347-bp product from a bacterial genomic library , amplification of the gene that encodes a 36-kDa antigen  and PCR-based selective amplification of a 530-bp fragment of the gene encoding the proline-rich antigen of M. leprae, a specific 360-bp DNA probe encoding 80% of the 18-kDa protein gene of M. leprae in-situ hybridization  or PCR using a unique sequence of 16S ribosomal RNA . Such sophisticated PCR is difficult to implement during routine diagnosis.
PCR based on the selective amplification of a 531-bp fragment of the gene encoding the proline-rich antigen of M. leprae was applied to nasal swab specimens from a leprostus patient, occupational contacts and endemic and nonendemic control. To prevent false-positive amplification use of dUTP and uracil DNA glycosylase in all PCRs was detected by using a 531-bp modified template as an internal control . In some experiments, primers to amplify a 212-bp region of the human β-globulin gene were used as internal positive control. PCR amplification of a 531-bp fragment of the M. leprae pra gene in fresh sample was evaluated for its usefulness in the detection of leprosy bacilli .
Dapsone was the first effective chemotherapeutic agent against leprosy. Its use as monotherapy for several years led to the development of a resistant strain of leprosy bacilli. In view of this, MDT is now recommended ; this has been the standard for treatment of leprosy since 1982 (WHO study group, 1982). A multidrug regimen of several antimicrobials is used to eradicate M. leprae. The types of drugs and duration of treatment depend on the amount of pathogen present. Several newer drugs active against M. leprae have emerged, which are being evaluated to improve the treatment and reduce the duration of treatment in multibacillary leprosy. Prominent among these are quinolones (pefloxacin, ofloxacin, sparfloxacin, moxifloxacin), ansamycins (rifabutin, KRM-1648), macrolides (clarithromycin), tetracyclines (minocycline), fusidic acid and other sulfones (brodimoprim). Of these, the quinolones, minocycline and clarithromycin appear to be the front-runners in providing alternative drug treatment of multibacillary leprosy [25,26].
Appropriate understanding and accurate diagnosis of leprosy is essential due to its prevalence for millions of years globally. Leprosy is an endemic worldwide and a major public health problem in India. It can be controlled and eliminated by early case detection and chemotherapy. Leprosy research has gained importance since the development of molecular techniques such as PCR. It is apparent that PCR can be established as a diagnostic procedure for leprosy patients and subclinical cases or as a tool for drug assessment.
Conflicts of interest
There are no conflicts of interest.
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