Quality control tests for vaccines in leprosy vaccine trial, Avadi.

All the vaccines supplied for the large scale comparative leprosy vaccine trial of ICRC bacilli, M.w, BCG plus killed M. leprae (candidate vaccines), BCG and normal saline (control arms) at CJIL Field Unit, Chennai were tested for quality control by the suppliers following the procedures laid down in the WHO protocol for killed M. leprae. Quality control for BCG was carried out at BCG vaccine laboratory as per protocol. Toxicity and sterility tests were done on all the vaccine batches/lots received. As part of the quality control, bacterial count, and protein estimation were also done. Studies showed that the bacterial content and protein concentration were comparable with the original preparations. Vaccines were free from micro-organisms, toxic materials and safe for human use. Thus the quality of all vaccine preparations was satisfactory.

Comparative leprosy vaccine trial in south India.

This report provides results from a controlled, double blind, randomized, prophylactic leprosy vaccine trial conducted in South India. Four vaccines, viz BCG, BCG+ killed M. leprae, M.w and ICRC were studied in this trial in comparison with normal saline placebo. From about 3,00,000 people, 2,16,000 were found eligible for vaccination and among them, 1,71,400 volunteered to participate in the study. Intake for the study was completed in two and a half years from January 1991. There was no instance of serious toxicity or side effects subsequent to vaccination for which premature decoding was required. All the vaccine candidates were safe for human use. Decoding was done after the completion of the second resurvey in December 1998. Results for vaccine efficacy are based on examination of more than 70% of the original "vaccinated" cohort population, in both the first and the second resurveys. It was possible to assess the overall protective efficacy of the candidate vaccines against leprosy as such. Observed incidence rates were not sufficiently high to ascertain the protective efficacy of the candidate vaccines against progressive and serious forms of leprosy. BCG+ killed M. leprae provided 64% protection (CI 50.4-73.9), ICRC provided 65.5% protection (CI 48.0-77.0), M.w gave 25.7% protection (CI 1.9-43.8) and BCG gave 34.1% protection (CI 13.5-49.8). Protection observed with the ICRC vaccine and the combination vaccine (BCG+ killed M. leprae) meets the requirement of public health utility and these vaccines deserve further consideration for their ultimate applicability in leprosy prevention.

Viability of M. leprae while undergoing laboratory procedures.

Studies were carried out to assess whether various methodological procedures adopted while conducting experiments, or, maintaining M. leprae under different conditions affected the number of organisms made available or their viability. Results of mouse foot-pad experiments showed that bacilli survived for one day at 37 degrees C, 7 days at 20 degrees to 30 degrees C and for 90 days in lyophilized conditions. Repeated daily exposure of the material preserved in refrigerator at +4 degrees C, to room temperatures showed that bacilli survived for only up to five days; whereas, with single exposure they survived up to 14 days. M. leprae were found to lose infectivity after 30 minutes of exposure to various disinfectants and ultra violet light. Centrifugation at high speed did not affect the viability of M. leprae.

Comparative assessment of viability of M.leprae by mouse foot-pad and fluorescent staining techniques.

Morphological characteristics have been used as a parameter to assess the viability of M.leprae in leprosy patients. However, with the advent of the mouse foot-pad technique, viability of M.leprae is determined by growing the bacilli in the mouse foot-pad. In recent years, a fluorescent staining technique using fluorescent diacetate-ethidium bromide (FDA-EB) has been used to assess the viability of cultivable mycobacteria as well as M.leprae. The purpose of this study was to compare the viability of M.leprae by both mouse foot-pad and fluorescent staining techniques. M.leprae strains from both untreated and treated patients as well as mouse passaged strains of M.leprae were used for the comparison. Percentage of green-stained bacilli in the inoculum was compared with that of multiplication of M.leprae in the mouse foot-pad. It was observed that there was no correlation between the estimates of viable M.leprae by fluorescent staining and by mouse foot-pad inoculation. FDA-EB staining appears to reflect only trends as absence of green staining cells had overall general correlation with loss of infectivity to mouse foot-pad but, the converse was not found to be true.

Coupling of proteins to liposomes and their role in understanding delayed type of hypersensitivity in human and mice.

Liposome-coupled lepromin was found to elicit a 3-week skin reaction in leprosy patients similar to that elicited by whole Mycobacterium leprae. The present study suggests that the presentation of antigens in a specific orientation is necessary for evoking delayed type hypersensitivity response in humans.

Evaluation of two multidrug regimen in hospitalised multibacillary cases.

Fifty three multibacillary leprosy cases were treated with two regimens of MDT L1 consisting of Rifampicin, Dapsone and Ethionamide and L2 consisting of Rifampicin, Dapsone and clofazimine. The results were compared at regular intervals and at the end of the study (24 months). Clinical inactivity, bacteriological negativity, ENL reactions, upgrading reactions were seen in L1 group in 65%, 4.54%, 50% and 41% of cases respectively while 65%, 25.8%, 30% and 45% respectively in L2 regimen group. Zero percent morphological Index was achieved in all cases in L1 regimen 90% in L2 regimen cases. No viability was found on mouse foot pad inoculation after 6 months in L1 while after 18 months in L2 cases.

Evaluation of the efficacy of candidate vaccines against M. leprae infection in mice.

Delayed-typed hypersensitivity (DTH) response and protection value of some of the candidate vaccines alone and in combination with BCG has been investigated. It was observed that both M.w. and BCG gave heightened DTH and good protection. On the other hand both M. leprae and ICRC evoked moderate DTH and gave poor protection. However on combining any of these candidate vaccines with live BCG, the lowering of DTH and poor protection was observed except in the M. leprae combination which in spite of low DTH gave better protection.

Follow up of BL/LL patients on a slightly modified WHO regimen of multidrug therapy.

56 lepromatous leprosy patients with an initial average BI of 4.45 were administered once a month 600 mg of Rifampicin, 100 mg of Clofazimine on alternate days and 100 mg of Dapsone daily. None of these patients became smear negative in 2 years, and the same regimen was continued further. Two patients have become negative in 3 years and treatment has been stopped in them. The study indicates that highly bacilliferous LL/BL patients are likely to need 3 years or more of MDT for achieving bacterial negativity.

Dapsone resistance in North India.

An attempt has been made to screen the resistant strains of M. leprae in lepromatous patients from the eight northern States of India. By using the mouse foot pad technique it was found that in a total of 69 clinically suspected patients 33 (47.8%) harboured M. leprae with some degree of dapsone resistance. A detailed epidemiological study in these parts of the country may reveal the prevalence rate.

Immunological potential of a cultivable mycobacterial strain M. habana against leprosy bacillus in mouse foot pad.

A strain of a typical mycobacteria M. habana originally afforded protection against M. tuberculosis challenge in mice, was tested for its immunological potential against leprosy bacillus in the mouse foot pad. The vaccine strain M. habana has arrested the growth of M. leprae into the mouse foot pad better than BCG (Phipps) and unvaccinated control.

Prevalence of drug resistance in Dharmapuri and A. Pallipatti areas of Tamil Nadu.

An evaluation of leprosy control project was undertaken in Dharmapuri and A. Pallipatti areas of Tamil Nadu to study the prevalence rate of drug resistance among the leprosy patients. At the end of 5 years of assessment 266 patients were still found to be bacteriologically positive among whom 25 patients were suspected to be clinically dapsone resistant. By mouse foot pad technique the drug resistant prevalence rate was found to be 1.1 per cent in these two areas.