Efficacy of single-dose ROM therapy plus low-dose convit vaccine as an adjuvant for treatment of paucibacillary leprosy patients with a single skin lesion.

The recent World Health Organization multicentric field study on the treatment of paucibacillary (PB) leprosy patients with single skin lesion (SSL) and a single dose of rifampin-ofloxacin-minocycline (ROM) brought new hope to those who are engaged in the eradication of leprosy from India. Being encouraged by the WHO report, we undertook the present hospital-based study and found that PB leprosy patients with SSL were morphologically and histopathologically heterogeneous. The histological spectrum of SSL ranged from indeterminate through tuberculoid (TT) to borderline tuberculoid (BT) leprosy, and most patients had active BT leprosy. Ninety new, untreated PB leprosy patients with SSL were included in the present study for comparative assessment of the efficacies of ROM and ROM plus Convit vaccine therapies. Children, pregnant women, lactating mothers and patients with any thickening of nerves were excluded. All patients were bacteriologically negative (skin-smear test) but lepromin reactive. The patients were divided into two groups after proper matching for morphological and histological status of SSL: a) The test group included 60 patients and the control group included 30 patients. The test group was given a single dose of ROM initially and two injections of low-dose Convit vaccine, one initially and the other at the end of 3 months. b) The control group was given only a single dose of ROM initially. Both groups were followed clinically every 2 weeks for 6 months and retested for histological, bacteriological and lepromin status at the end of 6 months. Thereafter, they were followed clinically every month for another 6 months. In the test group, the SSL resolved in 33.3%, regressed in 48.3%, and remained active in 18.3% of the patients, while the granuloma disappeared in 70% of the cases. Only one patient developed neuritis, and in another patient the disease relapsed on the eighth month. On the other hand, the SSL in the control patients resolved, regressed and remained active in 13.3%, 63.3% and 23.3% of the cases, respectively, while the granuloma disappeared in 53.3% of the cases. In the seven patients who remained active, the disease course was progressive, and two of them developed neuritis. The clinical outcome of the patients treated with ROM plus low-dose Convit vaccine was statistically superior to those treated with single-dose ROM therapy alone.

Why relapse occurs in PB leprosy patients after adequate MDT despite they are Mitsuda reactive: lessons form Convit's experiment on bacteria-clearing capacity of lepromin-induced granuloma.

It is amazing how after years of scientific research and therapeutic progress many simple and basic questions about protective immunity against Mycobacterium leprae remain unanswered. Although the World Health Organization (WHO) has recommended short-term multidrug therapy (WHO/MDT) for the treatment of paucibacillary (PB) leprosy patients, from time to time several workers from different parts of the globe have reported inadequate clinical responses in a few tuberculoid and indeterminate leprosy patients following adequate WHO/MDT despite the fact that they are Mitsuda responsive. A few borderline tuberculoid patients harbor acid-fast bacilli (AFB) in their nerves for many years even though they become clinically inactive following MDT, a fact which has been ignored by many leprosy field workers. Keeping these patients in mind, we have attempted to investigate the cause of the persistence of AFB in PB cases and have looked into the question of why Mitsuda positivity in tuberculoid and indeterminate leprosy patients, as well as in healthy contacts, is not invariably a guarantee for protectivity against the leprosy bacilli. We have: a) analyzed the histological features of lepromin-induced granulomas, b) studied the bacteria-clearing capacity of the macrophages within such granulomas, and c) studied the in vitro leukocyte migration inhibition factor released by the blood leukocytes of these subjects when M. leprae sonicates have been used as an elicitor. The results of these three tests in the three groups of subjects have been compared and led us to conclude that the bacteria-clearing capacity of the macrophages within lepromin-induced granuloma (positive CCB test) may be taken as an indicator of the capability of elimination of leprosy bacilli and protective immunity against the disease. This important macrophage function is not invariably present in all tuberculoid and indeterminate leprosy patients or in all contacts even though they are Mitsuda responsive and are able to show a positive leukocyte migration inhibition (LMI) test. It is likely but not certain that this deficit of the macrophage is genetically predetermined and persists after completion of short-term WHO/MDT. Thus, after discontinuation of treatment slow-growing, persisting M. leprae multiply within macrophages leading to relapse.

Immunotherapy of lepromin-negative borderline leprosy patients with low-dose Convit vaccine as an adjunct to multidrug therapy; a six-year follow-up study in Calcutta.

The present report, which describes management of lepromin-negative borderline leprosy patients with low-dose Convit vaccine, is an extension of our earlier study on the treatment of lepromatous leprosy patients with low-dose Convit vaccine as an adjunct to multidrug therapy (MDT). The test Group I, consisting of 50 lepromin-negative, borderline leprosy patients, were given low-dose Convit vaccine plus MDT. The control group II consisted of 25 lepromin-negative, borderline leprosy patients given BCG vaccination plus MDT and 25 lepromin-negative, borderline leprosy patients given killed Mycobacterium leprae (human) vaccine plus MDT. The control group III consisted of 50 lepromin-positive, borderline leprosy patients not given any immunostimulation but given only MDT. Depending upon the lepromin unresponsiveness, the patients were given one to four inoculations of the various antileprosy vaccines and were followed up every 3 months for 2 years for clinical, bacteriological and immunological outcome. All patients belonging to the test and control groups showed clinical cure and bacteriological negativity within 2 years. However, immunologic potentiation, assessed by lepromin testing and the leukocyte migration inhibition test (LMIT), was better in the test patients receiving low-dose Convit vaccine plus MDT than in the control patients receiving BCG vaccine plus MDT or killed M. leprae vaccine plus MDT or MDT alone. But the capacity of clearance bacteria (CCB) test from the lepromin granuloma showed poor bacterial clearance in the test patients. However, there was no relapse during 6 years of follow up. Two mid-borderline (BB) patients had severe reversal reactions with lagophthalmos and wrist drop during immunotherapy despite being given low-dose Convit vaccine.

Immunotherapy of far-advanced lepromatous leprosy patients with low-dose convit vaccine along with multidrug therapy (Calcutta trial).

This report describes a promising mode of treatment of lepromin-unresponsive, far-advanced, lepromatous (LL) leprosy patients with antileprosy vaccines as an adjunct to multidrug therapy (MDT). The Trial Groups included 50 highly bacilliferous, lepromin-negative, untreated LL patients. They were given MDT for 2 years. Of them, 30 patients were administered a mixed antileprosy vaccine containing killed Mycobacterium leprae of human origin plus M. bovis BCG. The remaining 20 patients were given M. bovis BCG. Depending on the severity of lepromin unresponsiveness, they were given one to six inoculations at 3-month intervals. Another 20 similar LL patients were taken in the Control Group. They were given only MDT for 2 years. From the start of the study, all patients belonging to the Trial and Control Groups were followed every 3 months for clinical, bacteriological and immunological outcomes. Within 2 years all 50 patients of the Trial Groups and 19 of the 20 patients of the Control Group became clinically inactive and bacteriologically negative. However, the clinical cure and the falls of the bacterial and morphological indexes were much faster in those patients receiving the mixed vaccine therapy than in those patients who were given BCG plus MDT or only MDT. The immunological improvements in the patients of the Trial and Control Groups were assessed by: a) lepromin testing at the beginning of the study and at 3-month intervals and also by b) the in vitro leukocyte migration inhibition (LMI) test at both the beginning and end of the study. As the patients were given more and more vaccinations, the incidence of lepromin conversion increased, more so in the patients receiving the mixed vaccine. Thus, 63%, 15% and 5% of the patients became lepromin positive in those patients receiving the mixed vaccine, BCG, and MDT only, respectively. Lamentably, the vaccine-induced lepromin positivity was temporary and faded away within several months. At the beginning of the study, the LMI test against specific M. leprae antigen was negative in all patients of both the Trial and Control Groups. After the end of the chemo-immunotherapy schedule, the LMI test became positive in 50% and 20% of LL patients receiving the mixed vaccine and BCG, respectively. None of the Control Group could show LMI positivity after completion of the MDT schedule. These results show that treatment of LL patients with the mixed vaccine and MDT could quickly reverse the clinical course of the disease, remove immunologic anergy in some patients, and induce a rapid decrease in the bacterial load in them.

Biometals in skin and sera of leprosy patients and their correlation to trace element contents of M. leprae and histological types of the disease; a comparative study with cutaneous tuberculosis.

The present study has provided information on the biometal contents of killed and dried Mycobacterium leprae as well as dermal granulomas induced by the invading mycobacteria in various histological types of leprosy patients. For comparison, the biometal contents of the contralateral leprosy-unaffected skin of the same patients also were measured. The study also reports changes of serum levels of the biometals in these patients which were compared with those in healthy control subjects and patients with skin tuberculosis. These data show that M. leprae is rich in zinc. During the course of the evolution of the disease there is gross alteration of the dynamics of the inflammatory cell population that infiltrates into leprosy granulomas, resulting in the alterations of trace element contents of the disease-affected skin lesions. Interestingly, the changes of the biometal contents in the granulomas of the patients with skin tuberculosis are similar to those in leprosy patients. It is postulated that the significant decrease of the contents of copper, zinc, iron, calcium and magnesium in the disease-affected skin in comparison to that of the contralateral healthy skin is a local effect, perhaps due to erosion or influx of biometal-deficient inflammatory cells into the affected skin with eventual loss of connective tissue of skin and mobilization of tissue-bound microelements into the vascular compartment. On the contrary, the changes in biometal levels in the sera of leprosy patients appear to be a general effect perhaps due to the release of interleukin-1, a product of inflammatory cells, causing hypercupremic, hypozincemic and hypoferremic responses in the hosts. Moreover, growth and multiplication of M. leprae, especially in polar lepromatous leprosy patients with a high bacillary load, demand essential biometals which may be mobilized into the bacterial bodies from the hosts. This perhaps results in the change in the homeostasis of the essential biometals in the hosts.

An eight-year field trial on antileprosy vaccines among high-risk household contacts in the Calcutta metropolis.

One-hundred-seventy-nine lepromin-negative household contacts were vaccinated with heat-killed Mycobacterium leprae, BCG, or a combination of the two. Vaccination induced lepromin positivity in 131 of these contacts. Over an 8-year follow-up period, 12 lepromin-positive contacts developed leprosy, all tuberculoid; while 2 lepromin-negative vaccinated contacts developed leprosy, both lepromatous. Overall, 7.8% of the vaccinated contacts developed the disease. Seven-hundred-fourteen household contacts were not vaccinated, and served as controls. Among the 504 who were lepromin positive, leprosy developed in 35, all tuberculoid, over the 8-year follow up. Among the 210 lepromin-negative unvaccinated contacts, 61 developed leprosy: tuberculoid in 29, borderline in 4, lepromatous in 8, and indeterminate in 20. Overall, 13.5% of the 714 unvaccinated contacts and 29.0% of the 210 unvaccinated, lepromin-negative contacts developed leprosy. Vaccination could not induce lepromin positivity in all contacts. The three vaccines were equally effective in inducing lepromin positivity. Vaccination reduced the overall incidence of leprosy from 13.5% to 7.8% among household contacts but did not reduce the incidence of lepromatous leprosy (1.2% of all the vaccinated and 1.1% of all the unvaccinated contacts).

Tumor necrosis factor: status in reactions in leprosy before and after treatment.

BACKGROUND: Tumor necrosis factor-alpha (TNF) is an important mediator of immunologic responses to chronic infections. METHOD: Sera from 25 patients with acute reactions (6 with type 1 upgrading, 8 with type 1 downgrading, and 11 with type 2 reaction) were assayed for TNF before treatment and after clinical remission of the acute episode. The results were compared with serum TNF levels in healthy controls and fresh pauci- and multibacillary leprosy patients. RESULTS: TNF levels in acute reactions were higher than in the control groups (significant only in upgrading reaction). In type 1 reaction, serum TNF concentrations fell to approximately the levels of the control patients following treatment and clinical remission. In type 2 reaction, however, levels of TNF were seen to rise further (became statistically significant) as a result of therapy induced clinical remission. CONCLUSIONS: The rise in TNF-alpha level in reactions in leprosy is significant and indicates its active role in immunopathogenesis. The corresponding decline in TNF-alpha levels seen following regression of type 1 (lepra) reactions was not observed in the case of type 2 (ENL) reaction. This probably reflects the enhancement of cellular immunity in such cases and/or an attempt by the immunologic process to overcome specific inhibitors.

Nutritional status of children of urban leprosy patients staying at preventoria based on biochemical parameters.

Measurements of nutritionally relevant biochemical and endocrine variables were made on 60 apparently healthy children (group A) whose parents suffered from leprosy and who had been separated at the age of 4 years and brought up in preventoria. Most of the measurements were also made on a comparison group of healthy children from the same poor socio-economic class (group B). In both groups the serum concentrations of cholesterol and triglycerides were well below those found in Western populations. Almost all the children in both groups were anaemic, but serum iron and ferritin levels were satisfactory. Folate and vitamin B12 levels were measured in group A only and were low in a significant proportion. Deficiency of these water-soluble vitamins may be a cause of the anaemia. Low albumin levels were found in 40% of group A children, compared with 2% in group B. The concentrations of calcium and magnesium were lower and that of phosphate higher in group A than in B. In both groups one-third of the children had low levels of serum zinc. Fifteen per cent of group A children had biochemical evidence of vitamin A deficiency, but none were deficient in vitamin E. Levels of total T3 and total T4 were below the lower limit of normal in a substantial proportion of children in both groups. Concentrations of parathyroid hormone were increased in parallel with the low values for serum calcium. Radiological studies of ossification centres in 57 group A children showed delayed maturation in 11 cases. The relevance of these findings to previous studies of the children of lepers in India is discussed.

Reconstruction of anti-leprosy drug depleted complement haemolytic activity by addition of zymosan-treated sera (a source of C142) and CratEDTA (a source of C3-C9).

This paper describes the mechanism of in vitro interaction of human serum complement system with anti-leprosy drugs (dapsone and clofazimine) and anti-lepra reaction drugs such as chloroquine. These drugs could inhibit the complement-mediated lysis of erythrocytes both via direct and alternative pathways, but only at hypertherapeutic doses. Attempts were made to restore the drug depleted complement-mediated lysis of erythrocytes by adding zymosan-treated guinea-pig sera (a source of C142) and also by adding Crat-EDTA sera (a source of C3-C9). Destroyed complement-mediated haemolytic activity by dapsone could be restored by early complement (C142) components, while complement-mediated haemolytic activity blocked by clofazimine could be regenerated by adding both late (C3-C9) and early (C142) complement component. However, chloroquine-mediated inhibition of the complement-mediated haemolysis activity could not be appreciably restored by adding both early and late complement reagents.

Delayed clearance of circulating immune complexes in mice following administration of antileprosy drugs.

In this report we describe an animal experiment which showed delayed clearance of preformed 125I-HSA-anti-HSA immune complexes (with five times excess HSA) from the circulation of mice treated with antileprosy drugs (dapsone, clofazimine, and rifampin--multidrug therapy for 7 days) in comparison with normal (untreated) mice. The results also showed delayed retention of the preformed immune complexes in the spleen and kidneys of the antileprosy-drug-treated animals. The exact mechanism of the delayed handling of preformed immune complexes in mice fed antileprosy drugs could not be ascertained. However, in light of the anticomplementary effects of clofazimine and dapsone, as reported earlier, and in light of the large accumulation of clofazimine and rifampin in macrophages, it has been postulated that in the drug-fed animals either the immune complexes could not be phagocytosed by macrophages, through the avenue of their C3b receptors, or the immune complexes could not be downgraded easily within the macrophages overloaded with clofazimine and rifampin. These results might have clinical significance and might throw some light on the prolonged persistence of circulating immune complexes in the vascular bed of lepromatous patients even after clinical remission of erythema nodosum leprosum.

In vivo effects of anti-leprosy drugs on the rat peritoneal macrophages and lymphocyte subpopulations.

The present study describes the in vivo effects of anti-leprosy drugs on rat peritoneal macrophages and T-cell homeostasis. It was observed that BCG-elicited rat peritoneal macrophages produced more H2O2 and expressed more Ia antigen on their cell surfaces compared with resident peritoneal macrophages. Furthermore, elicited macrophages isolated from rats administered multidrug therapy (MDT), consisting of dapsone, clofazimine and rifampicin in high dose (10 x MDT) released more O2-. On the contrary, there was a significant decrease in the Ia antigen expression on these macrophages. Anti-leprosy drug treatment in high dose (10 x MDT) decreased the total number of blood T-helper (W3/25+) cells and increased the total number of blood T-suppressor (OX-8+) cells which resulted in a significant decrease in a W3/25: OX-8 ratio. Electron microscopy of elicited macrophages isolated from 10 x MDT treated rats showed development of many filipodia compared with control macrophages. These data show that 10 x MDT treatment in rats for 1 month alters the homeostasis of blood T-cell subpopulations which perhaps decreases the Ia expression on macrophages. However, the increase in O2- production and the appearance of filipodia on the macrophages is due to a direct effect of drugs on the macrophages. MDT treatment for 1 month in a therapeutic dose has no effect on the above-mentioned parameters.

Anti-leprosy drugs inhibit the complement-mediated solubilization of pre-formed immune complexes in vitro.

Incubation of pre-formed immune complexes (IC) (125I-HSA--anti-HSA) with normal human serum resulted in solubilization of IC. When various anti-leprosy drugs were added to human sera, solubilization of IC was fairly explicit with clofazimine, whereas this effect was marginal with dapsone. Rifampicin hardly displayed this effect. Aspirin, chloroquine, and prednisolone, the drugs used in addition to multi-drug therapy to control reactions in leprosy, were in a position to inhibit the solubilization of 125I HSA--anti-HSA by normal serum only at a very high dose. From the current data of the inhibition of solubilization of pre-formed IC along with our earlier observations on the modulation of complement-mediated haemolysis by these drugs, it may be possible to postulate that clofazimine as well as chloroquine affect early complement components. This may in turn be responsible for preventing the deposition of C3 complement onto IC.

Effect of anti-leprosy drugs on superoxide anion production by rat peritoneal macrophage with special reference to light exposed clofazimine.

The present study describes the in vitro effect of anti-leprosy drugs on superoxide anion (O2-) production by rat resident peritoneal macrophages. Of the three drugs tested i.e. clofazimine, rifampicin and dapsone, the first was most effective in increasing O2- production in a dose dependent manner, while rifampicin had some stimulatory effect and dapsone exhibited minimal action. Furthermore, when clofazimine and dapsone were added together it was observed that the increase of O2- production by macrophages due to clofazimine was not significantly altered by the addition of dapsone. Moreover, it was found that killed Mycobacterium leprae could induce a lesser amount of O2- production in comparison to that of Staphylococcus aureus and the enhancement of O2- release due to clofazimine was stimulus dependent. This increase of O2- release after addition of clofazimine was inhibited by the addition of p-bromophenacyl bromide. Another interesting finding was that the enhancement of O2- production by clofazimine gradually decreased as clofazimine was exposed to light for days. On further investigation it was found that ultraviolet, NMR, infrared and mass spectra of the light unexposed and exposed drug were similar, but the diffusion current of the polarogram of light exposed drug was remarkably more than that observed in light unexposed drug, indicating, thereby, a possible increase in the electron accepting capacity of the light reacted molecule. As far as we know this is the first report describing the effect of light exposed clofazimine on the respiratory burst activity of macrophages.

Sexually transmitted diseases in leprosy patients in north and northeastern India. A futile search for human immunodeficiency virus antibody.

Three-hundred-eighty-four leprosy patients were clinically examined for sexually transmitted diseases (STD) in north and northeastern India, revealing a high incidence (5.2%) of STD among them. Eighteen males, one female, and one eunuch were found to have chancroid ulcer, gonococcal urethritis, lymphogranuloma inguinale, and primary chancre. Of these patients, only 100, selected randomly, could be screened serologically for STD due to Treponema pallidum, herpes simplex (type 1 and 2), Entamoeba histolytica, hepatitis-associated virus, cytomegalovirus, Chlamydia trachomatis and human immunodeficiency virus (HIV); 100 control sera were included for comparison. In addition, sera from another 133 normal subjects and another 176 lepromatous patients were also screened for HIV antibody. Thus, a total of 233 normal sera and 276 leprosy sera were tested for HIV antibody. Although our leprosy patients have shown significantly high incidences of clinical STD and also high seropositivity against T. pallidum, herpes-simplex viruses types 1 and 2, hepatitis-associated virus, and cytomegalovirus, the search for antibody against HIV was negative. Our clinical and serological data suggest promiscuity in our patient population. The absence of HIV antibody in this high-risk population, however, seems to be an enigma.

A study on nutrition, growth and development of a high-risk group of children of urban leprosy patients.

This paper describes a cross-sectional study of the physical development of a high-risk group of 182 socially deprived healthy children of leprosy patients ranging from preschool age to early teens. They were rescued at the age of 4 years from the distress of leprosy colonies where they were born, and brought up in Government Homes (Preventorium) under better environmental conditions. Of them 135 children could be followed clinically for 10 years for the development of childhood leprosy. Another 94 children of leprosy patients living with their parents were included for comparison. A group of 158 normal children of similar economic status and age group were included as controls. It was observed that, although better environment, food and training were provided in the Preventorium, so that the children could be brought into the national mainstream, nevertheless 5 children developed an indeterminate type of leprosy during the course of 10 years. This is the first report of growth and development of children of leprosy patients from the Indian sub-continent.