Data-Medicos 
Dermagic/Express No. 68 
11 Agosto 1.999. 11 August 1.999. 

~ Lepra (Hansen) y vacunas ~ 
~ Leprosy (Hansen disease) and vaccine ~ 


EDITORIAL ESPANOL 
================= 
Hola Amigos de la red, DERMAGIC de nuevo con ustedes. La lepra, enfermedad bien conocida desde la antiguedad, se ha convertido en un verdadero reto para nuestros investigadores en la busqueda de una VACUNA, que proteja CONTRA la infeccion del mycobacterium Leprae. 
Muchos intentos se han hecho, y hay varios grupos trabajando en ello, en VENEZUELA el Grupo del Dr. Convit trabaja con Cepas de Mycobacterium Leprae, tambien con BCG, quiza uno de los pioneros en esta busqueda ansiosa, los resultados, alentadores. En ARGENTINA se esta trabajando con Cepas de Mycobacterium Bovis y vaccae. Otros paises (Brasil) también con EL BCG SOLO o con Mycobacterium Leprae, Pero encuentro que en la India NUEVA DELHI, se ha estado trabajando con 4 cepas, entre las que destacan Mycobacterium Habana y Mycobacterium w, este ultimo del cual segun ellos se pondra al mercado LA PRIMERA VACUNA contra la Lepra producida por Cadila Pharmaceuticals, (referencia 50), porque NO ES PATOGENO. Pero si revisamos bien TODAS las referencias, NO SON VACUNAS PROPIAMENTE DICHAS, en el sentido estricto de la PREVENCION de la infeccion, puesto que se usan en combinacion con poliquimioterapia. (MTD). Recordemos tambien que la clasica vacuna BCG (bacillus Calmette-Guerin) , que protege conta la tuberculosis, tambien protege contra la lepra.. 

Por mi parte felicito a todos estos investigadores, pero seguiremos esperando por UNA REAL VACUNA contra la LEPRA... Espero que les guste este DERMAGIC, 


Saludos a todos !!! 

Dr. Jose Lapenta R.,,, 

EDITORIAL ENGLISH 
================= 
Hello Friends of the net, DERMAGIC again with you. The leprosy, very well-known illness from the antiquity, has become a true challenge for our investigators in the search of a VACCINE that protects AGAINST the infection of the mycobacterium Leprae. 
Many intents have been made, and there are several groups working in it, in VENEZUELA the Group of the Dr. Convit begins with Strains of Mycobacterium Leprae, also the BCG, maybe one of the pioneers in this anxious search, the results, encouraging. In ARGENTINEAN are working with Strains o f Mycobacterium Bovis and vaccae. In other countries (Brazil) with THE BCG ALONE or plus Mycobaterium Leprae, But I find that in the India NEW DELHI, has been working with 4 strains, among those are the Mycobacterium Habana and Mycobacterium w, this last of which will put on to the market THE FIRST VACCINE against the Leprosy produced by Cadila Pharmaceuticals, according to them, (reference 50), because it IS NONPATHOGEN. But if we revise ALL the references well, they ARE NOT VACCINE PROPERLY, this in the strict sense of the PREVENTION of the infection, since they are used in combination with multidrugtherapy (MTD). Let us also remember that the classic VACCINE BCG (bacillus Calmette-Guerin) that protects against the tuberculosis, it also protects against the leprosy.. 

I congratulate all these investigators, but we will continue waiting for A REAL VACCINE against the LEPROSY... I hope you like this DERMAGIC, 


Greetings to ALL, !! 
Dr. Jose Lapenta R.,,, 
=================================================================== 
REFERENCIAS BIBLIOGRAFICAS / BIBLIOGRAPHICAL REFERENCES 
=================================================================== 
===================================================================== 
1.) Causative organism and host response. 
2.) The GroES antigens of Mycobacterium avium and Mycobacterium paratuberculosis. 
3.) Human T cell recognition of the Mycobacterium leprae LSR antigen: epitopes 
and HLA restriction. 
4.) Quality control tests for vaccines in leprosy vaccine trial, Avadi. 
5.) Comparative leprosy vaccine trial in south India. 
6.) Effectiveness of bacillus Calmette-Guerin (BCG) vaccination in the 
prevention of leprosy; a case-finding control study in Nagpur, India. 
7.) Effectiveness of Bacillus Calmette Guerin (BCG) vaccination in the 
prevention of childhood pulmonary tuberculosis: a case control study in 
Nagpur, India. 
8.) Tuberculin sensitivity and skin lesions in children after vaccination with 
two batches of BCG vaccine. 
9.) Leprosy vaccine: influence of dissolved oxygen levels on growth of a 
candidate strain (Mycobacterium w), and storage stability of the vaccine. 
10.) Studies of vaccination of persons in close contact with leprosy patients in 
Argentina. 
11.) 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. 
12.) BCG vaccination protects against leprosy in Venezuela: a case-control study. 
13.) Immunoprophylactic trial with combined Mycobacterium leprae/BCG vaccine 
against leprosy: preliminary results. 
14.) IgM antibodies to native phenolic glycolipid-I in contacts of leprosy 
patients in Venezuela: epidemiological observations and a prospective study 
of the risk of leprosy. 
15.) Immunological changes observed in indeterminate and lepromatous leprosy 
patients and Mitsuda-negative contacts after the inoculation of a mixture 
of Mycobacterium leprae and BCG. 
16.) Comparative study of the 48-hour response to soluble antigens obtained from 
human and armadillo leprosy material in lepromatous leprosy patients and 
normal persons, contacts of leprosy patients. 
17.) Association of HLA specificity LB-E12 (MB1, DC1, MT1) with lepromatous 
leprosy in a Venezuelan population. 
18.) Immunotherapy with a mixture of Mycobacterium leprae and BCG in different 
forms of leprosy and in Mitsuda-negative contacts. 
19.) A 35-kilodalton protein is a major target of the human immune response to 
Mycobacterium leprae. 
20.) Immunogenicity and protection studies with recombinant mycobacteria and 
vaccinia vectors coexpressing the 18-kilodalton protein of Mycobacterium 
leprae. 
21.) Mycobacterial infections: are the observed enigmas and paradoxes explained 
by immunosuppression and immunodeficiency? 
22.) Leprosy patients with lepromatous disease recognize cross-reactive T cell 
epitopes in the Mycobacterium leprae 10-kD antigen. 
23.) [BCG vaccination to Mycobacterium leprae infection in mice] 
24.) Human leukocyte antigens in tuberculosis and leprosy. 
25.) Modulation of protective and pathological immunity in mycobacterial 
infections. 
26.) IL-2 and IL-12 act in synergy to overcome antigen-specific T cell 
unresponsiveness in mycobacterial disease. 
27.) Dharmendra antigen but not integral M. leprae is an efficient inducer of 
immunostimulant cytokine production by human monocytes, and M. leprae 
lipids inhibit the cytokine production. 
28.) Inhibition of multiplication of Mycobacterium leprae in mouse foot pads by 
immunization with ribosomal fraction and culture filtrate from 
Mycobacterium bovis BCG. 
29.) Techniques for genetic engineering in mycobacteria. Alternative host 
strains, DNA-transfer systems and vectors. 
30.) Leprosy vaccine: influence of dissolved oxygen levels on growth of a 
candidate strain (Mycobacterium w), and storage stability of the vaccine. 
31.) Lymphostimulatory and delayed-type hypersensitivity responses to a 
candidate leprosy vaccine strain: Mycobacterium habana. 
32.) Randomised controlled trial of single BCG, repeated BCG, or combined BCG 
and killed Mycobacterium leprae vaccine for prevention of leprosy and 
tuberculosis in Malawi. Karonga Prevention Trial Group [see comments] 
33.) 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. 
34.) A case-control study of the effectiveness of BCG vaccine for preventing 
leprosy in Yangon, Myanmar. 
35.) Immunotherapy of far-advanced lepromatous leprosy patients with low-dose 
convit vaccine along with multidrug therapy (Calcutta trial). 
36.) Protective immunization of monkeys with BCG or BCG plus heat-killed 
Mycobacterium leprae: clinical results. 
37.) Studies of vaccination of persons in close contact with leprosy patients in 
Argentina. 
38.) Restoration of proliferative response to M. leprae antigens in lepromatous 
T cells against candidate antileprosy vaccines. 
39.) Does bacille Calmette-Gu´erin scar size have implications for protection 
against tuberculosis or leprosy? 
40.) Protective efficacy of BCG against leprosy in S~ao Paulo. 
41.) Post-vaccination sensitization with ICRC vaccine. 
Author 
42.) Sensitization and reactogenicity of two doses of candidate antileprosy 
vaccine Mycobacterium w. 
43.) Tuberculin sensitivity and skin lesions in children after vaccination with 
two batches of BCG vaccine. 
44.) Association between leprosy and HIV infection in Tanzania. 
45.) A follow-up study of multibacillary Hansen's disease patients treated with 
multidrug therapy (MDT) or MDT + immunotherapy (IMT). 
46.) Novel O-methylated terminal glucuronic acid characterizes the polar 
glycopeptidolipids of Mycobacterium habana strain TMC 5135. 
47.) Regional lymphadenitis following antileprosy vaccine BCG with killed 
Mycobacterium leprae. 
48.) A major T-cell-inducing cytosolic 23 kDa protein antigen of the vaccine 
candidate Mycobacterium habana is superoxide dismutase. 
49.) Supervised Multiple Drug Therapy Program, Venezuela 
50.) NII DEVELOPES WORLD'S FIRST ANTI-LEPROSY VACCINE 
===================================================================== 
===================================================================== 
1.) Causative organism and host response. 
===================================================================== 
Lepr Rev 1999 Mar;70(1):95-102 
Krahenbuhl JL 

Whether or not the leprosy elimination target is met in all endemic 
countries by the year 2000, the MDT programme will have greatly reduced 
worldwide prevalence. However, our workshop chairmen were asked to ignore 
the prevalence-based leprosy 'elimination' programme and focus on 
recommendations for a long term, incidence-based eradication target where 
transmission is blocked. They were asked to be concerned with basic leprosy 
research goals in the post 2000 era. The members of our workshops are 
actively productive workers, committed to their special interests. They are 
fully cognizant of the obstacles faced daily in working with leprosy and M. 
leprae, the requirement for clever experimental design even with the 
availability of the powerful tools of molecular biology which can now be 
brought to bear on some of the research obstacles. They are also aware of 
our lack of understanding about leprosy and M. leprae. How do you block 
transmission if you don't know how infection is transmitted? Can infection 
be detected, diagnosis made earlier? Is there a non-human reservoir host, a 
carrier state, an environmental source? What is the basis of M. leprae's 
predilection for nerves, the mechanisms underlying reactions? What needs to 
be targeted to treat reactions? Can a vaccine play a role? There is nothing 
startling in the workshops' recommendations. Other individuals and groups 
of experts have made the same suggestions, with slightly varying 
priorities. What one can read between the lines of these reports, is a 
sense of urgency to get as much done as soon as possible. Worldwide 
interest in leprosy will soon be diminished, not by design but as a 
consequence of the laudable success of the MDT programme. The experiment is 
still underway, but chemotherapy alone, killing bacilli in the detectable 
human host, does not appear to be the answer to blocking transmission. A 
number of goals must be addressed while there are still intact national and 
international leprosy programmes, while there are still leprosy treatment 
and research centres that can co-ordinate and facilitate the necessary 
trials for early diagnosis, early detection of reactions, evaluation of 
immunosuppressive regimens for reactions. A key recommendation is concerned 
with the means of measuring progress. A clear and explicit means of 
reporting incidence, prevalence and 'case detection' should be implemented 
to avoid a distorted picture of worldwide leprosy. These recommendations 
are non-controversial. What should be done is clear. The uncertainty is in 
determining who will do the work. Who will fund the laboratories engaged in 
this work? Look around you. There are fewer scientists attending this 
Congress but browsing the abstracts and attending our sessions and posters 
clearly revealed to me that fewer of us are doing far better work than in 
the past. Alternative sources of funding will help. Tuberculosis research 
is enticing researchers away from leprosy in the developed countries but is 
visibly sustaining leprosy research in many centres in developing 
countries. Formation of alliances was a key goal of this Congress. I asked 
my colleagues from Carville to identify in their own discipline, dedicated 
people, committed laboratories that will sustain their leprosy research 
efforts over the next 5, 10 or more years. These are the people with whom 
we wish to collaborate, form alliances, share resources and expertise, 
address the future of worldwide leprosy. 

===================================================================== 
2.) The GroES antigens of Mycobacterium avium and Mycobacterium paratuberculosis. 
===================================================================== 
Vet Microbiol 1999 Jun 1;67(1):31-5 
Cobb AJ, Frothingham R 
Veterans Affairs Medical Center, Durham, NC 27705, USA. 

The GroES antigen provokes a strong immune response in human beings with 
tuberculosis or leprosy. We cloned and sequenced the Mycobacterium avium 
and Mycobacterium paratuberculosis GroES genes. M. avium and M. 
paratuberculosis have identical GroES sequences which differ from other 
mycobacterial species. This supports the current formal designation of M. 
paratuberculosis as M. avium subsp. paratuberculosis. Immunodominant 
epitopes from Mycobacterium tuberculosis GroES are conserved in M. avium, 
but some Mycobacterium leprae epitopes are distinct. GroES is unlikely to 
be specific as a serologic or skin test reagent, but may be an appropriate 
component of a broad mycobacterial vaccine. 

===================================================================== 
3.) Human T cell recognition of the Mycobacterium leprae LSR antigen: epitopes 
and HLA restriction. 
===================================================================== 
FEMS Immunol Med Microbiol 1999 Jun;24(2):151-9 

Oftung F, Lundin KE, Meloen R, Mustafa AS 
Department of Vaccinology, National Institute of Public Health, Oslo, 
Norway. [email protected] 

We have in this work mapped epitopes and HLA molecules used in human T cell 
recognition of the Mycobacterium leprae LSR protein antigen. HLA typed 
healthy subjects immunized with heat killed M. leprae were used as donors 
to establish antigen reactive CD4+ T cell lines which were screened for 
proliferative responses against overlapping synthetic peptides covering the 
C-terminal part of the antigen sequence. By using this approach we were 
able to identify two epitope regions represented by peptide 2 (aa 29-40) 
and peptide 6 (aa 49-60), of which the former was mapped in detail by 
defining the N- and C-terminal amino acid positions necessary for T cell 
recognition of the core epitope. MHC restriction analysis showed that 
peptide 2 was presented to T cells by allogeneic cells coexpressing HLA-DR4 
and DRw53 or DR7 and DRw53. In contrast, peptide 6 was presented to T cells 
only in the context of HLA-DR5 molecules. In conclusion, the M. leprae LSR 
protein antigen can be recognized by human T cells in the context of 
multiple HLA-DR molecules, of which none are reported to be associated with 
the susceptibility to develop leprosy. The results obtained are in support 
of using the LSR antigen in subunit vaccine design. 

===================================================================== 
4.) Quality control tests for vaccines in leprosy vaccine trial, Avadi. 
===================================================================== 
Indian J Lepr 1998 Oct-Dec;70(4):389-95 

Sreevatsa, Hari M, Gupte MD 
BCG Vaccine Laboratory, Guindy, Chennai. 

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. 

===================================================================== 
5.) Comparative leprosy vaccine trial in south India. 
===================================================================== 
Indian J Lepr 1998 Oct-Dec;70(4):369-88 

Gupte MD, Vallishayee RS, Anantharaman DS, Nagaraju B, Sreevatsa, 
Balasubramanyam S, de Britto RL, Elango N, Uthayakumaran N, Mahalingam VN, 
Lourdusamy G, Ramalingam A, Kannan S, Arokiasamy J 

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. 

===================================================================== 
6.) Effectiveness of bacillus Calmette-Guerin (BCG) vaccination in the 
prevention of leprosy; a case-finding control study in Nagpur, India. 
===================================================================== 
Int J Lepr Other Mycobact Dis 1998 Sep;66(3):309-15 

Zodpey SP, Shrikhande SN, Salodkar AD, Maldhure BR, Kulkarni SW 
Clinical Epidemiology Unit, Government Medical College, Nagpur, India. 

A hospital-based, pair-matched, casecontrol study was carried out at 
Government Medical College Hospital in Nagpur in central India to estimate 
the effectiveness of BCG vaccination in the prevention of leprosy. The 
study included 314 incidence cases of leprosy [diagnosed by World Health 
Organization (WHO) criteria] below the age of 32 years. Each case was pair 
matched with one control for age, sex and socioeconomic status. Controls 
were selected from subjects attending this hospital for conditions other 
than tuberculosis and leprosy. A significant protective association between 
BCG and leprosy was observed (OR 0.29, 95% CI 0.21-0.41). The vaccine 
effectiveness (VE) was estimated to be 71% (95% CI 59-79). The BCG 
effectiveness against multibacillary and paucibacillary leprosy was 79% 
(95% CI 60-89) and 67% (95% CI 45-78), respectively. It was more effective 
during the first decade of life (VE 74%; 95% CI 38-90), among females (VE 
82%; 95% CI 64-90), and in the lower socioeconomic strata (VE 75%; 95% CI 
32-92). The prevented fraction was calculated to be 51% (95% CI 38-62). In 
conclusion, this study has identified a beneficial role of BCG vaccination 
in the prevention of leprosy in central India. 

===================================================================== 
7.) Effectiveness of Bacillus Calmette Guerin (BCG) vaccination in the 
prevention of childhood pulmonary tuberculosis: a case control study in 
Nagpur, India. 
===================================================================== 
Southeast Asian J Trop Med Public Health 1998 Jun;29(2):285-8 

Zodpey SP, Shrikhande SN, Maldhure BR, Vasudeo ND, Kulkarni SW 
Department of Preventive and Social Medicine, Government Medical College, 
Nagpur, India. 

A hospital-based, pair matched, case control study was carried out to 
estimate the effectiveness of BCG vaccination in the prevention of 
childhood pulmonary tuberculosis. The study included 126 incident cases of 
pulmonary tuberculosis (diagnosed by WHO criteria) below/equal the age of 
12 years. Each case was pair matched with one control for age, sex, 
socio-economic status. Controls were selected from subjects attending study 
hospital for conditions other than tuberculosis and leprosy. The 
significant protective association between BCG and childhood pulmonary 
tuberculosis was observed (OR = 0.39, 95% CI = 0.22, 0.68). The overall 
vaccine effectiveness was 61% (95% CI = 32%, 78%). BCG was nonsignificantly 
more effective in underfives, among males and in upper-middle socioeconomic 
strata. The overall prevented fraction was estimated to be 47.53% (95% CI = 
21.41%, 67.25%). Results of this study thus demonstrated a moderate 
effectiveness of BCG vaccination in prevention of childhood pulmonary 
tuberculosis in a Central India population. 

===================================================================== 
8.) Tuberculin sensitivity and skin lesions in children after vaccination with 
two batches of BCG vaccine. 
===================================================================== 
Indian J Lepr 1998 Jul-Sep;70(3):277-86 

Vallishayee RS, Anantharaman DS, Gupte MD 
CJIL Field Unit (ICMR), Avadi, Chennai. 

BCG is one of the vaccines used, as control arm, in an ongoing large scale 
comparative leprosy vaccine trial in South India. The objective of the 
present study was to examine, in the local population, the sensitizing 
ability, as measured by skin test reactions to tuberculin, and 
reactogenecity, in terms of skin lesions at the site of vaccination, for 
the two batches of BCG vaccine used in the above trial. The study was 
undertaken in 816 tuberculin-negative, previously not vaccinated school 
children, aged five to 14 years. Each child received one of the two batches 
of BCG vaccine or normal saline (control), by random allocation. At 12 
weeks from vaccination, character and size of local response, at the 
vaccination site, were recorded. At the same time, the children were 
retested with tuberculin and post-vaccination reactions to the test were 
measured after 72 hours. At three years after vaccination all available 
children were re-examined for the presence and size of BCG scar at the site 
of vaccination. It was found that healing of vaccination lesions was 
uneventful, with both batches of BCG. The mean size of the lesion was 
similar for the two batches, the overall mean being 6.3 mm. The mean size 
of post-vaccination tuberculin sensitivity increased with age, and it was 
14.5 mm and 15.6 mm. The sensitizing effect attributable to the vaccine was 
11 mm and 12 mm, for the two batches of BCG respectively. This study showed 
that the two batches of BCG, in a dose of 0.1 mg, used in the ongoing 
leprosy vaccine trial were acceptable in terms of vaccination lesion and 
were highly satisfactory in terms of development of hypersensitivity. 

===================================================================== 
9.) Leprosy vaccine: influence of dissolved oxygen levels on growth of a 
candidate strain (Mycobacterium w), and storage stability of the vaccine. 
===================================================================== 
Vaccine 1998 Aug;16(13):1344-8 

Mukhopadhyay A, Panda AK, Pandey AK 
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi, India. 
[email protected] 

The growth of Mycobacterium w, a candidate strain for leprosy vaccine in 
submerged culture, was inhibited by the presence of over 40% oxygen 
saturation in the medium. Intracellular levels of superoxide dismutase and 
catalase were very low in the beginning. However, under controlled 
oxygenation, these levels increased with time. The augmentations of these 
antioxidant enzymes were associated with the elevated oxygen consumption by 
the culture. By maintaining the oxygen level below 20% during 6-day 
culture, it was possible to grow Mycobacterium w in five production batches 
up to a cell density of 3.7 +/- 0.70 x 10(9) bacilli ml-1. The shelf life 
of the vaccine produced in different batches was more than 2 years, both at 
4 degrees C and at 26 degrees C. This provides a cost-effective, unit 
culture technology for the production of this candidate leprosy vaccine 
from a nonpathogenic organism, which will facilitate the widespread use of 
the vaccine. 

===================================================================== 
10.) Studies of vaccination of persons in close contact with leprosy patients in 
Argentina. 
===================================================================== 
Vaccine 1998 Jul;16(11-12):1166-71 

Bottasso O, Merlin V, Cannon L, Cannon H, Ingledew N, Keni M, Hartopp R, 
Stanford C, Stanford J 
Instituto de Inmunologia, Facultad de Ciencias Medicas, Universidad 
Nacional de Rosario, Argentina. 

A total of 670 adults living or working with leprosy patients, were 
examined for a BCG vaccination scar, and skin-tested with four new 
tuberculins. Based on the results 513 were vaccinated, 65 with Bacille de 
Calmette et Guerin (BCG) alone, 66 with BCG plus killed Mycobacterium 
vaccae and 382 with killed M. vaccae alone. Skin-testing was repeated 2-3 
years later on 344 subjects, when all three vaccines were found to have 
been highly successful in increasing responses to Tuberculin and Leprosin A 
(p < 0.0005) with increased immune recognition of common and 
species-specific antigens. Mean diameters of induration to each skin-test 
were greatest in recipients of BCG alone (p < 0.05), which suggests that 
better immuno-regulation occurs after receiving vaccines that incorporate 
M. vaccae. The results suggest 10(8) M. vaccae alone might prove a valuable 
future vaccine, which would not require selective pre-vaccination procedures. 

===================================================================== 
11.) 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. 
===================================================================== 
Int J Lepr Other Mycobact Dis 1998 Jun;66(2):182-9 

Chaudhuri S, Hajra SK, Mukherjee A, Saha B, Mazumder B, Chattapadhya D, Saha K 
Department of Leprosy, School of Tropical Medicine, Calcutta, India. 

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. 

===================================================================== 
12.) BCG vaccination protects against leprosy in Venezuela: a case-control study. 
===================================================================== 
Int J Lepr Other Mycobact Dis 1993 Jun;61(2):185-91 

Convit J, Smith PG, Zuniga M, Sampson C, Ulrich M, Plata JA, Silva J, 
Molina J, Salgado A 
Instituto de Biomedicina, Caracas, Venezuela. 

A total of 64,570 household and other close contacts of about 2000 leprosy 
cases were screened for eligibility for entry into a trial of a new leprosy 
vaccine. The screening procedure included a clinical examination for 
leprosy and for the presence of BCG and lepromin scars. Ninety-five new 
cases of leprosy were identified, and the prevalence of BCG and lepromin 
scars among them was compared with similar data from matched controls 
selected from among those with no evidence of leprosy. The difference in 
the prevalence of BCG scars in the two groups was used to estimate the 
protection against leprosy conferred by BCG vaccination. One or more BCG 
scars was associated with a protective efficacy of 56% (95% confidence 
limits 27% to 74%). There was a trend of increasing protection with four or 
more BCG scars, but this was not statistically significant. There was no 
evidence that the efficacy of BCG varied with age or according to whether 
or not the contact lived in the same household as a case. The protective 
effect was significantly higher among males, and was significantly greater 
for multibacillary than for paucibacillary leprosy. 

===================================================================== 
13.) Immunoprophylactic trial with combined Mycobacterium leprae/BCG vaccine 
against leprosy: preliminary results. 
===================================================================== 
Lancet 1992 Feb 22;339(8791):446-50 

Convit J, Sampson C, Zuniga M, Smith PG, Plata J, Silva J, Molina J, 
Pinardi ME, Bloom BR, Salgado A 
Instituto de Biomedicina, Caracas, Venezuela. 

In an attempt to find a vaccine that gives greater and more consistent 
protection against leprosy than BCG vaccine, we compared BCG with and 
without killed Mycobacterium leprae in Venezuela. Close contacts of 
prevalent leprosy cases were selected as the trial population since they 
are at greatest risk of leprosy. Since 1983, 29,113 contacts have been 
randomly allocated vaccination with BCG alone or BCG plus 6 x 10(8) 
irradiated, autoclaved M leprae purified from the tissues of infected 
armadillos. We excluded contacts with signs of leprosy at screening and a 
proportion of those whose skin-test responses to M leprae soluble antigen 
(MLSA) were 10 mm or more (positive reactions). By July, 1991, 59 
postvaccination cases of leprosy had been confirmed in 150,026 person-years 
of follow-up through annual clinical examinations of the trial population 
(31 BCG, 28 BCG/M leprae). In the subgroup for which we thought an effect 
of vaccination was most likely (onset more than a year after vaccination, 
negative MLSA skin-test response before vaccination), leprosy developed in 
11 BCG recipients and 9 BCG/M leprae recipients; there were 18% fewer cases 
(upper 95% confidence limit [CL] 70%) in the BCG/M leprae than in the BCG 
alone group. For all cases with onset more than a year after vaccination 
irrespective of MLSA reaction the relative efficacy was 0% (upper 95% CL 
54%; 15 cases in each vaccine group). Retrospective analysis of data on the 
number of BCG scars found on each contact screened suggested that BCG alone 
confers substantial protection against leprosy (vaccine efficacy 56%, 95% 
CL 27-74%) and there was a suggestion that several doses of BCG offered 
additional protection. There is no evidence in the first 5 years of 
follow-up of this trial that BCG plus M leprae offers substantially better 
protection against leprosy than does BCG alone, but the confidence interval 
on the relative efficacy estimate is wide. 

===================================================================== 
14.) IgM antibodies to native phenolic glycolipid-I in contacts of leprosy 
patients in Venezuela: epidemiological observations and a prospective study 
of the risk of leprosy. 
===================================================================== 
Int J Lepr Other Mycobact Dis 1991 Sep;59(3):405-15 

Ulrich M, Smith PG, Sampson C, Zuniga M, Centeno M, Garcia V, Manrique X, 
Salgado A, Convit J 
Instituto de Biomedicina, Caracas, Venezuela. 

In a randomized, double-blind vaccine trial in Venezuela, about 29,000 
contacts of leprosy patients have been vaccinated with either a mixture of 
heat-killed Mycobacterium leprae and BCG or BCG alone, and are being 
re-surveyed annually to detect new cases of leprosy. All contacts had a 
serum sample collected at the time of entry into the trial, and 13,020 of 
these sera have been analyzed for antibodies to phenolic glycolipid-I 
(PGL-I). Antibody levels have been related to various characteristics of 
the contacts and to their risk of developing leprosy in the following 4 
years. A strong association was found between PGL-I antibody level and the 
risk of developing leprosy, in spite of possible modification of the 
incidence rate induced by vaccination. Antibody levels were higher in 
females than in males, and declined progressively with age. Household 
contacts had higher levels than did non-household contacts, and levels were 
higher in individuals from the state in Venezuela which has the highest 
incidence of the disease. No substantial differences were found in antibody 
levels between contacts of multibacillary and paucibacillary patients, 
which may in part reflect the influence of treatment, and there was no 
clear association with the presence of BCG or lepromin scars or with 
skin-test responses to PPD and leprosy soluble antigen. The assay of 
antibodies to PGL-I seems unlikely to provide a sensitive or specific test 
for infection with M. leprae, and measuring PGL-I antibody levels as a 
screening procedure to identify those at high risk of developing leprosy is 
unlikely to be particularly useful in most leprosy control programs. Such 
assays may be useful for the epidemiological monitoring of changes in the 
intensity of infection with M. leprae in a community and for the study of 
carefully defined groups of contacts during some phases of control programs. 

===================================================================== 
15.) Immunological changes observed in indeterminate and lepromatous leprosy 
patients and Mitsuda-negative contacts after the inoculation of a mixture 
of Mycobacterium leprae and BCG. 
===================================================================== 
Clin Exp Immunol 1979 May;36(2):214-20 

Convit J, Aranzazu N, Pinardi M, Ulrich M 

This investigation was carried out to study the possibility of eliciting 
favourable immunological changes in small groups of Mitsuda-negative 
patients with indeterminate leprosy, lepromatous patients who were 
bacteriologically negative after prolonged treatment with sulphones, and in 
Mitsuda-negative contacts by means of stimulation with a mixture of 
autoclaved tissues from Mycobacterium leprae-infected armadillos and living 
BCG. A radical change was observed in the specific immunological activity 
of the indeterminate group, all of whom initially had occasional bacilli in 
cutaneous nerves in biopsies taken from hypopigmented spots, and in the 
persistently Mitsuda-negative contacts. The 48 hr and 30 day reactions to 
lepromin, the 48 hr reaction to supernatant antigen from lepromin, the test 
for bacillary clearence and in vitro lymphocyte transformation (LTT) to M. 
leprae from human and armadillo lesions all became positive. Of the 
lepromatous patients studied, only one became positive to all the criteria 
mentioned above. In the others, the 48 hr reaction to supernatant antigen, 
the LTT to antigen from a humn source, and the clearance test remained 
negative, while the Fernandez and Mitsuda reactions became positive. These 
results are discussed in terms of the possible use of this stimulation 
procedure in the prevention and immunotherapy of leprosy. 

===================================================================== 
16.) Comparative study of the 48-hour response to soluble antigens obtained from 
human and armadillo leprosy material in lepromatous leprosy patients and 
normal persons, contacts of leprosy patients. 
===================================================================== 
Int J Lepr Other Mycobact Dis 1976 Jan-Jun;44(1-2):284-6 

Convit J, Pinardi ME, Aranzazu N 
We prepared antigens by precipitating with 80% ammonium sulfate 
supernatants of human and armadillo antigen at a concentration of 160 X 
10(6) bacteria per ml. The precipitate was resuspended, dialyzed and 
filtered. The antigen obtained was inactivated with trypsin during 30 
minutes. The tests made with these antigens were negative for the 48-hour 
test in lepromatous patients and highly positive in normal persons who were 
contacts of leprosy patients. 

===================================================================== 
17.) Association of HLA specificity LB-E12 (MB1, DC1, MT1) with lepromatous 
leprosy in a Venezuelan population. 
===================================================================== 
Tissue Antigens 1984 Jul;24(1):25-9 

Ottenhoff TH, Gonzalez NM, de Vries RR, Convit J, van Rood JJ 

To investigate whether an association could be found between HLA and 
lepromatous leprosy a population study was performed in Tachira, Venezuela. 
This was done in the same endemic area in which recently both non-random 
parental HLA-haplotype and preferential segregation of the HLA specificity 
LB-E12 (MB1, DC1, MT1) was demonstrated in lepromatous leprosy patients 
from multicase families. In this study 32 lepromatous patients and 32 
healthy controls were typed for HLA-A, -B, -C, -DR and the specificities MB 
and MT. The frequency of LB-E12 (MB1, DC1, MT1) showed a significant 
increase in lepromatous leprosy patients (p = 0.04). This is the first 
report concerning HLA and leprosy which confirms in the same endemic area 
an association observed in families on the population level. 

===================================================================== 
18.) Immunotherapy with a mixture of Mycobacterium leprae and BCG in different 
forms of leprosy and in Mitsuda-negative contacts. 
===================================================================== 
Int J Lepr Other Mycobact Dis 1982 Dec;50(4):415-24 

Convit J, Aranzazu N, Ulrich M, Pinardi ME, Reyes O, Alvarado J 

A total of 529 weak or non-reactors to M. leprae, including 
Mitsuda-negative contacts and patients with leprosy, were vaccinated once 
or repeatedly, as necessary, with a mixture of 6 x 10(8) purified, 
heat-killed M. leprae and 0.01 mg to 0.2 mg of viable BCG. Clinical, 
histopathological and immunological criteria were used to evaluate the 
response of these individuals. Clinical changes, including sharper 
definition of borders and progressive flattening and regression of lesions, 
were observed in 57% of the active LL cases and 76% of the active BL cases. 
Histopathological study revealed infiltration of the lesions by mononuclear 
cells, appearance of epithelioid differentiation, and fragmentation of the 
microorganisms. Delayed-type skin tests with soluble antigen from purified 
M. leprae became positive in significant numbers of each group studied. 
These results demonstrate the efficacy of combined immunotherapy in 
low-resistance forms of leprosy and potential utility in the 
immunoprophylaxis of the disease. 

===================================================================== 
19.) A 35-kilodalton protein is a major target of the human immune response to 
Mycobacterium leprae. 
===================================================================== 
Author 
Triccas JA; Roche PW; Winter N; Feng CG; Butlin CR; Britton WJ 
Address 
Centenary Institute of Cancer Medicine and Cell Biology, Newtown, New South 
Wales, Australia. 
Source 
Infect Immun, 64(12):5171-7 1996 Dec 
Abstract 
The control of leprosy will be facilitated by the identification of major 
Mycobacterium leprae-specific antigens which mirror the immune response to 
the organism across the leprosy spectrum. We have investigated the host 
response to a 35-kDa protein of M. leprae. Recombinant 35-kDa protein 
purified from Mycobacterium smegmatis resembled the native antigen in the 
formation of multimeric complexes and binding by monoclonal antibodies and 
sera from leprosy patients. These properties were not shared by two forms 
of 35-kDa protein purified from Escherichia coli. The M. smegmatis-derived 
35-kDa protein stimulated a gamma interferon-secreting T-cell proliferative 
response in the majority of paucibacillary leprosy patients and healthy 
contacts of leprosy patients tested. Cellular responses to the protein in 
patients with multibacillary leprosy were weak or absent, consistent with 
hyporesponsiveness to M. leprae characteristic of this form of the disease. 
Almost all leprosy patients and contacts recognized the 35-kDa protein by 
either a T-cell proliferative or an immunoglobulin G antibody response, 
whereas few tuberculosis patients recognized the antigen. This specificity 
was confirmed in guinea pigs, with the 35-kDa protein eliciting strong 
delayed-type hypersensitivity in M. leprae-sensitized animals but not in 
those sensitized with Mycobacterium tuberculosis or Mycobacterium bovis 
BCG. Therefore, the M. leprae 35-kDa protein appears to be a major and 
relatively specific target of the human immune response to M. leprae and is 
a potential component of a diagnostic test to detect exposure to leprosy or 
a vaccine to combat the disease. 

===================================================================== 
20.) Immunogenicity and protection studies with recombinant mycobacteria and 
vaccinia vectors coexpressing the 18-kilodalton protein of Mycobacterium 
leprae. 
===================================================================== 
Author 
Baumgart KW; McKenzie KR; Radford AJ; Ramshaw I; Britton WJ 
Address 
Centenary Institute of Cancer Medicine and Cell Biology, University of 
Sydney, Newtown, New South Wales, Australia. 
Source 
Infect Immun, 64(6):2274-81 1996 Jun 
Abstract 
The activation of antigen-specific T lymphocytes is essential for the 
control of leprosy infection in humans and experimental animals. T cells 
recognize a variety of protein antigens from Mycobacterium leprae, 
including the 18-kDa protein, which is limited in distribution among 
mycobacteria and which is absent from Mycobacterium tuberculosis and the 
vaccine strain, Mycobacterium bovis BCG. Adjuvant preparations of 
mycobacterial protein antigens have had limited protective efficacy for 
experimental infections in animals. Since recombinant vectors may elicit 
more effective T-cell responses than adjuvant preparations, recombinant 
vaccinia virus (VV18) and M. bovis BCG (BCG18) vectors expressing the 
18-kDa protein of M. leprae were prepared. Both VV18 and BCG18 stimulated 
anti-18-kDa protein antibody and lymphocyte proliferative responses. 
Sequential immunization with VV18 followed by BCG18 induced higher levels 
of specific immunoglobulin G2a antibodies than immunoglobulin G1 
antibodies, in contrast to immunization with VV18 or BCG18 alone. The 
protective efficacy of immunization with VV18 from a challenge with BCG18 
was examined in two murine models of mycobacterial infection. After 
intravenous challenge, mice immunized with recombinant vaccinia virus 
exhibited lower initial levels of replication and earlier clearance of 
BCG18 from their spleens than mice immunized with vaccinia virus expressing 
an unrelated protein. After footpad infection in a dissemination model, 
there was earlier clearance of BCG18 from specifically immunized mice. 
However, immunization of mice with VV18 did not prevent a productive 
mycobacterial infection. 

===================================================================== 
21.) Mycobacterial infections: are the observed enigmas and paradoxes explained 
by immunosuppression and immunodeficiency? 
===================================================================== 
Author 
Maes HH; Causse JE; Maes RF 
Address 
Microbiology and Genetics Unit, University of Louvain Medical School, 
Brussels, Belgium. 
Source 
Med Hypotheses, 46(2):163-71 1996 Feb 
Abstract 

The enigmas and paradoxes observed in tuberculous patients, in Bacille 
Calmette-Gu&acute;erin-vaccinated people and in Bacille Calmette-Gu&acute;erin-treated 
cancer patients have been examined, in an attempt to explain them through 
the mechanisms of immunodeficiency and immunosuppression. A dual effect is 
postulated: an immunosuppression induced by the infecting mycobacteria that 
adds to a pre-existing or emerging state of immunodeficiency of the 
infected individual. The immunological cellular and humoral anergies 
observed at the beginning of a tuberculous therapy are usually lifted after 
the first two weeks of treatment. This restoration of immune responsiveness 
may be attributed to the destruction or to the growth inhibition of 
immunosuppressive mycobacteria. The observation that drugs cytocidal in 
vitro do not always sterilize the patients under treatment whereas 
bacteriostatic drugs do, may find an explanation in the dual 
immunosuppression induced by cytocidal drugs and mycobacteria. The fact 
that Bacille Calmette-Gu&acute;erin applied as an immunotherapy to residual 
cancer has either a favorable or an unfavorable action may be due to the 
immunosuppressive activity attached to some Bacille Calmette-Gu&acute;erin 
strains and to some cancers. The variable protective activity of Bacille 
Calmette-Gu&acute;erin vaccines may be due to the immunological status of the 
vaccinated people and the compositional differences between strains. The 
protective activity of subunit vaccines in experimental models can be 
attributed to the elimination of immunosuppressive factors present in whole 
killed mycobacteria. 

===================================================================== 
22.) Leprosy patients with lepromatous disease recognize cross-reactive T cell 
epitopes in the Mycobacterium leprae 10-kD antigen. 
===================================================================== 
Author 
Hussain R; Dockrell HM; Shahid F; Zafar S; Chiang TJ 
Address 
Department of Microbiology, The Aga Khan University, Karachi, Pakistan. 
Source 
Clin Exp Immunol, 114(2):204-9 1998 Nov 
Abstract 
T cell responses play a critical role in determining protective responses 
to leprosy. Patients with self-limiting tuberculoid leprosy show high T 
cell reactivity, while patients with disseminated lepromatous form of the 
disease show absent to low levels of T cell reactivity. Since the T cell 
reactivity of lepromatous patients to purified protein derivative (PPD), a 
highly cross-reactive antigen, is similar to that of tuberculoid patients, 
we queried if lepromatous patients could recognize cross-reactive epitopes 
in Mycobacterium leprae antigens as well. T cell responses were analysed to 
a recombinant antigen 10-kD (a heat shock cognate protein) which is 
available from both M. tuberculosis (MT) and M. leprae (ML) and displays 
90% identity in its amino acid sequence. Lymphoproliferative responses were 
assessed to ML and MT 10 kD in newly diagnosed leprosy patients 
(lepromatous, n = 23; tuberculoid, n = 65). Lepromatous patients showed 
similar, but low, lymphoproliferative responses to ML and MT 10 kD, while 
tuberculoid patients showed much higher responses to ML 10 kD. This 
suggests that the tuberculoid patients may be recognizing both 
species-specific and cross-reactive epitopes in ML 10 kD, while lepromatous 
patients may be recognizing only cross-reactive epitopes. This was further 
supported by linear regression analysis. Lepromatous patients showed a high 
concordance in T cell responses between ML and MT 10 kD (r=0.658; P<0.0006) 
not observed in tuberculoid patients (r=0.203; P>0.1). Identification of 
cross-reactive T cell epitopes in M. leprae which could induce protective 
responses should prove valuable in designing second generation 
peptide-based vaccines. 

===================================================================== 
23.) [BCG vaccination to Mycobacterium leprae infection in mice] 
===================================================================== 
Author 
Nomaguchi H; Yogi Y; Matsuoka M; Fukotomi Y; Okamura H; Nagata K; Nagai S; 
Ohara N; Yamada T 
Address 
National Institute for Leprosy Research. 
Source 
Nippon Rai Gakkai Zasshi, 65(2):106-12 1996 Jul 
Abstract 
BCG vaccine (Tokyo strain) was given in BALB/cA mice intradermally 1 or 3 
months before Mycobacterium leprae (M. leprae) challenge as modified 
Shepard's method. The vaccine dosage was 10(7-8) or 10(6). The BCG gave 
good protection in both dosages and both challenges against M. leprae 
infection. Lymphocytes proliferations of BCG-vaccinated splenocyte cultures 
in response to M. leprae lysate or BCG components (hsp65, 38 kD, 30 kD or 
12 kD protein) were tested, and potent proliferative responses were seen in 
the cultures with M. leprae lysate and hsp65. Furthermore, gamma-IFN 
productions were positive in the cultures with M. leprae lysate or hsp65, 
but negative with other antigens. The production of gamma-IFN with hsp65 
was never inhibited with polymyxin B, but inhibited with IL-10. These 
results show that BCG (Tokyo strain) is a useful vaccine for M. leprae 
infection in mice, and one of the components of BCG, hsp65, may be a 
effective antigen component for protection of M. leprae infection inducing 
Th1 type cytokine. 

===================================================================== 
24.) Human leukocyte antigens in tuberculosis and leprosy. 
===================================================================== 
Author 
Meyer CG; May J; Stark K 
Address 
Institute for Tropical Medicine, Berlin, Germany. [email protected] 
Source 
Trends Microbiol, 6(4):148-54 1998 Apr 
Abstract 
Human mycobacterial infections are characterized by a spectrum of clinical 
and immunological manifestations. Specific human leukocyte antigen (HLA) 
factors are associated with the subtypes of leprosy that develop and the 
course of tuberculosis after infection. The identification of protective 
mycobacterial antigens presented by a broad variety of HLA molecules will 
have important implications for the design of vaccines. 

===================================================================== 
25.) Modulation of protective and pathological immunity in mycobacterial 
infections. 
===================================================================== 
Author 
Ottenhoff TH; Spierings E; Nibbering PH; de Jong R 
Address 
Department of Immunohematology and Blood Bank, University Hospital, Leiden, 
The Netherlands. [email protected] 
Source 
Int Arch Allergy Immunol, 113(4):400-8 1997 Aug 
Abstract 
Mycobacterial infections represent major problems to global health care. 
Tuberculosis is feared particularly because of its high mortality rates 
whereas in leprosy the occurrence of immunopathology, particularly nerve 
damage, is a major problem since the bacillus itself is relatively 
harmless. Thus, both effective vaccination strategies as well as novel 
immunomodulating regimens are warranted for the control of morbidity and 
mortality in mycobacterial diseases. Since CD4+ Th1 cells and type-1 
cytokines play a key role both in protective immunity and immunopathology 
in mycobacterial infections, we here describe new pharmacological and 
cytokine-based strategies to regulate Th1 immunity. 

===================================================================== 
26.) IL-2 and IL-12 act in synergy to overcome antigen-specific T cell 
unresponsiveness in mycobacterial disease. 
===================================================================== 
Author 
de Jong R; Janson AA; Faber WR; Naafs B; Ottenhoff TH 
Address 
Department of Immunohematology & Bloodbank, University Hospital Leiden, The 
Netherlands. 
Source 
J Immunol, 159(2):786-93 1997 Jul 15 
Abstract 
IL-12 secretion by APC is critical for the development of protective 
Th1-type responses in mycobacterial (Mycobacterium avium and Mycobacterium 
tuberculosis) infections in mice. We have studied the role of IL-12 and 
IL-2 in the generation of Mycobacterium leprae-specific T cell responses in 
humans. Leprosy patients were defined as low/nonresponders or high 
responders based on the level of T cell proliferation in M. 
leprae-stimulated PBMC. In high responders, M. leprae-induced proliferation 
was markedly suppressed by neutralizing anti-IL-12 mAb (inhibition 55 +/- 
6%). Neutralization of IL-2 activity resulted in an inhibition of 77 +/- 
4%. Given the importance of endogenous IL-2 and IL-12 in M. leprae-induced 
responses, we investigated the ability of rIL-2 and rIL-12 to reverse T 
cell unresponsiveness in low/nonresponder patients. Interestingly, rIL-12 
and rIL-2 strongly synergized in restoring both M. leprae-specific T cell 
proliferation and IFN-gamma secretion almost completely to the level of 
responder patients. A similar synergy between rIL-2 and rIL-12 was also 
observed in high responders when suboptimal M. leprae concentrations were 
used for T cell stimulation. Our data demonstrate a crucial role for 
endogenous IL-12 and IL-2 in M. leprae-induced T cell activation. Most 
importantly, we show that rIL-2 and rIL-12 act in synergy to overcome 
Ag-specific Th1 cell unresponsiveness. These findings may be applicable to 
the design of antimicrobial and antitumor vaccines. 

===================================================================== 
27.) Dharmendra antigen but not integral M. leprae is an efficient inducer of 
immunostimulant cytokine production by human monocytes, and M. leprae 
lipids inhibit the cytokine production. 
===================================================================== 
Author 
Nakamura C; Fukutomi Y; Kashiwabara Y; Oomoto Y; Kojima M; Hayashi H; 
Onozaki K 
Address 
Department of Hygienic Chemistry, Faculty of Pharmaceutical Sciences, 
Nagoya City University, Japan. 
Source 
Int J Lepr Other Mycobact Dis, 65(1):63-72 1997 Mar 
Abstract 
Killed integral Mycobacterium leprae, Mitsuda antigen, and 
chloroform-treated M. leprae, Dharmendra antigen (Dh-Ag), have been used 
for the classification of leprosy patients based on cell-mediated immunity. 
Heat-killed M. leprae also were used as a component of the Convit vaccine. 
Human blood monocytes were stimulated with M. leprae or Dh-Ag and their 
cytokine-inducing ability was compared. Monocytes were cultured in the 
presence of fresh human serum because of the efficiency of cytokine 
induction and the phagocytosis of M. leprae have been shown to be optimal 
in the presence of fresh serum. M. leprae and Dh-Ag were equally 
phagocytosed by monocytes. Dh-Ag was more potent than M. leprae in the 
induction of immunostimulatory/proinflammatory cytokines, interleukin-1 
(IL-1), IL-6 and tumor necrosis factor (TNF). In contrast, a comparable 
level of IL-1ra, an immunosuppressive cytokine, was induced by M. leprae 
and Dh-Ag. The lipids extracted from M. leprae induced none of these 
cytokines by monocytes. Nevertheless, when monocytes were pretreated with 
the lipids followed by stimulation with Dh-Ag, productions of IL-1, IL-6 
and TNF were all inhibited in a dose-dependent manner. However, the lipids 
did not inhibit the cytokine production induced by other stimuli including 
BCG and lipopolysaccharide. Moreover the lipids did not affect the 
production of IL-1ra. These results suggest that the lipids from M. leprae 
are responsible for the poor cytokine-inducing ability of M. leprae, thus 
favoring their infection. These results also suggest that Dh-Ag rather than 
integral M. leprae may be useful as a vaccine candidate because Dh-Ag is 
able to induce a large amount of cytokines from monocytes. 

===================================================================== 
28.) Inhibition of multiplication of Mycobacterium leprae in mouse foot pads by 
immunization with ribosomal fraction and culture filtrate from 
Mycobacterium bovis BCG. 
===================================================================== 
Author 
Matsuoka M; Nomaguchi H; Yukitake H; Ohara N; Matsumoto S; Mise K; Yamada T 
Address 
National Institute for Leprosy Research, Tokyo, Japan. 
Source 
Vaccine, 15(11):1214-7 1997 Aug 
Abstract 
Immunization of mice with the ribosomal fraction from ruptured 
Mycobacterium bovis Bacillus Calmette-Gu&acute;erin (BCG) and the culture 
filtrate reduced remarkably the multiplication of Mycobacterium leprae in 
the foot pads of mice. This is the first reported case of the protective 
activity against M. leprae multiplication in mice of the BCG ribosomal 
fraction and culture filtrate. The inhibition was more evident with the 
culture filtrate than with the ribosomal fraction. When the ribosomal 
proteins separated from ribosomal RNA were injected into mice, only slight 
inhibition was observed. Ribosomal RNA alone did not inhibit at all, in 
contrast to the conclusion reported by Youmans and Youmans. 

===================================================================== 
29.) Techniques for genetic engineering in mycobacteria. Alternative host 
strains, DNA-transfer systems and vectors. 
===================================================================== 
Author 
Hermans J; de Bont JA 
Address 
Department of Food Science, Agricultural University, Wageningen, The 
Netherlands. 
Source 
Antonie Van Leeuwenhoek, 69(3):243-56 1996 Apr 
Abstract 
The study of mycobacterial genetics has experienced quick technical 
developments in the past ten years, despite a relatively slow start, caused 
by difficulties in accessing these recalcitrant species. The study of 
mycobacterial pathogenesis is important in the development of new ways of 
treating tuberculosis and leprosy, now that the emergence of 
antibiotic-resistant strains has reduced the effectiveness of current 
therapies. The tuberculosis vaccine strain M. bovis BCG might be used as a 
vector for multivalent vaccination. Also, non-pathogenic mycobacterial 
strains have many possible biotechnological applications. After giving a 
historical overview of methods and techniques, we will discuss recent 
developments in the search for alternative host strains and DNA transfer 
systems. Special attention will be given to the development of vectors and 
techniques for stabilizing foreign DNA in mycobacteria. 

===================================================================== 
30.) Leprosy vaccine: influence of dissolved oxygen levels on growth of a 
candidate strain (Mycobacterium w), and storage stability of the vaccine. 
===================================================================== 
Author 
Mukhopadhyay A; Panda AK; Pandey AK 
Address 
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi, India. 
[email protected] 
Source 
Vaccine, 16(13):1344-8 1998 Aug 
Abstract 
The growth of Mycobacterium w, a candidate strain for leprosy vaccine in 
submerged culture, was inhibited by the presence of over 40% oxygen 
saturation in the medium. Intracellular levels of superoxide dismutase and 
catalase were very low in the beginning. However, under controlled 
oxygenation, these levels increased with time. The augmentations of these 
antioxidant enzymes were associated with the elevated oxygen consumption by 
the culture. By maintaining the oxygen level below 20% during 6-day 
culture, it was possible to grow Mycobacterium w in five production batches 
up to a cell density of 3.7 +/- 0.70 x 10(9) bacilli ml-1. The shelf life 
of the vaccine produced in different batches was more than 2 years, both at 
4 degrees C and at 26 degrees C. This provides a cost-effective, unit 
culture technology for the production of this candidate leprosy vaccine 
from a nonpathogenic organism, which will facilitate the widespread use of 
the vaccine. 

===================================================================== 
31.) Lymphostimulatory and delayed-type hypersensitivity responses to a 
candidate leprosy vaccine strain: Mycobacterium habana. 
===================================================================== 
Author 
Singh NB; Gupta HP; Srivastava A; Kandpal H; Srivastava UM 
Address 
Division of Microbiology, Central Drug Research Institute, Lucknow, India. 
Source 
Lepr Rev, 68(2):125-30 1997 Jun 
Abstract 
Lymphostimulatory and delayed-type hypersensitivity (DTH) immune responses 
to a candidate antileprosy vaccine Mycobacterium habana have been 
quantified in inbred AKR mice. M. habana vaccine in three physical states, 
live, heat-killed and gamma-irradiated, was given intradermally to separate 
groups of mice and after 28 days these mice were given subcutaneous 
challenge with heat-killed M. leprae and heat-killed M. habana in the left 
hind footpad. Live BCG vaccine alone and in combination with 
gamma-irradiated M. habana were also compared similarly. A sufficient 
degree of DTH response was generated in mice by M. habana vaccine in all 
physical forms against two challenge antigens (lepromin and habanin). The 
BCG combination with M. habana did not increase the DTH response indicating 
internal adjuvanticity endowed in M. habana. The active hypersensitivity of 
immunized mice was transferable to syngeneic mice by the transfer of 
sensitized cells from the donor to the recipient mice intravenously. M. 
leprae-infected Rhesus monkey PBMC have shown comparable stimulatory 
response with M. habana (sonicate), and M. leprae (sonicate) antigens. The 
possibility of developing M. habana as a candidate antileprosy vaccine is 
discussed. 

===================================================================== 
32.) Randomised controlled trial of single BCG, repeated BCG, or combined BCG 
and killed Mycobacterium leprae vaccine for prevention of leprosy and 
tuberculosis in Malawi. Karonga Prevention Trial Group [see comments] 
===================================================================== 
Source Lancet, 348(9019):17-24 1996 Jul 6 
Abstract 
BACKGROUND: Repeat BCG vaccination is standard practice in many countries 
for prevention of tuberculosis and leprosy, but its effectiveness has not 
been evaluated. The addition of Mycobacterium leprae antigens to BCG might 
improve its effectiveness against leprosy. A double-blind, randomised, 
controlled trial to evaluate both these procedures was carried out in 
Karonga District, northern Malawi, where a single BCG vaccine administered 
by routine health services had previously been found to afford greater than 
50% protection against leprosy, but no protection against tuberculosis. 
METHODS: Between 1986 and 1989, individuals lacking a BCG scar were 
randomly assigned BCG alone (27,904) or BCG plus killed M leprae (38,251). 
Individuals with a BCG scar were randomly allocated placebo (23,307), a 
second BCG (23,456), or BCG plus killed M leprae (8102). Incident cases of 
leprosy and tuberculosis were ascertained over the subsequent 5-9 years. 
FINDINGS: 139 cases of leprosy were identified by May, 1995; 93 of these 
were diagnostically certain, definitely postvaccination cases. Among 
scar-positive individuals, a second BCG vaccination gave further protection 
against leprosy (about 50%) over a first BCG vaccination. The rate ratio 
for all diagnostically certain, definitely postvaccination cases, all ages, 
was 0.51 (95% CI 0.25-1.03, p = 0.05) for BCG versus placebo. This benefit 
was apparent in all subgroups, although the greatest effect was among 
individuals vaccinated below 15 years of age (RR = 0.40 [95% CI 0.15-1.01], 
p = 0.05). The addition of killed M leprae did not improve the protection 
afforded by a primary BCG vaccination. The rate ratio for BCG plus killed M 
leprae versus BCG alone among scar-negative individuals was 1.06 
(0.62-1.82, p = 0.82) for all ages, though 0.37 (0.11-1.24, p = 0.09) for 
individuals vaccinated below 15 years of age. 376 cases of postvaccination 
pulmonary tuberculosis and 31 of glandular tuberculosis were ascertained by 
May, 1995. The rate of diagnostically certain tuberculosis was higher among 
scar-positive individuals who had received a second BCG (1.43 [0.88-2.35], 
p = 0.15) than among those who had received placebo and there was no 
evidence that any of the trial vaccines contributed to protection against 
pulmonary tuberculosis. INTERPRETATION: In a population in which a single 
BCG vaccination affords 50% or more protection against leprosy, but none 
against tuberculosis, a second vaccination can add appreciably to the 
protection against leprosy, without providing any protection against 
tuberculosis. 

===================================================================== 
33.) 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. 
===================================================================== 
Author 
Chaudhury S; Hajra SK; Mukerjee A; Saha B; Majumdar V; Chattapadhya D; Saha K 
Address 
School of Tropical Medicine, Calcutta, India. 
Source 
Int J Lepr Other Mycobact Dis, 65(1):56-62 1997 Mar 
Abstract 
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. 

===================================================================== 
34.) A case-control study of the effectiveness of BCG vaccine for preventing 
leprosy in Yangon, Myanmar. 
===================================================================== 
Author 
Bertolli J; Pangi C; Frerichs R; Halloran ME 
Address 
Epidemiology Program Office, US Centers for Disease Control and Prevention, 
Atlanta, GA, USA. 
Source 
Int J Epidemiol, 26(4):888-96 1997 Aug 
Abstract 
BACKGROUND: Five randomized trials, a follow-up study, and six case-control 
investigations of BCG vaccine's effectiveness (VE) for preventing leprosy 
have been conducted internationally, with widely varying estimates of VE. 
Because of the difficulty of generalizing from disparate results, local 
estimates of VE are needed for health planning purposes and are currently 
particularly relevant, given the World Health Organization's (WHO) goal to 
eliminate leprosy by the year 2000. METHODS: We conducted a case-control 
study in Yangon, Myanmar. Residents of Yangon between the ages of 6 years 
and 24 years who were listed in the National Leprosy Registry as being on 
active treatment for leprosy between December 1992 and April 1993 were 
eligible to participate in the study as cases. Control subjects were 
matched to the cases on age, sex, and neighbourhood. RESULTS: One or more 
doses of BCG were associated with a VE of 66%. The results show a 
significant trend of increasing VE with increasing number of BCG doses (one 
dose, VE = 55%; two doses, VE = 68%; three doses, VE = 87%). One dose of 
BCG vaccine appeared to provide protection substantially higher than that 
found in an earlier vaccine trial in Myanmar, but consistent with results 
from case-control studies in other countries. CONCLUSIONS: These data 
suggest that BCG reduces the risk of leprosy in Myanmar, and that BCG 
vaccination of infants, along with early case-finding and treatment, should 
be considered an important part of the leprosy intervention strategy. 

===================================================================== 
35.) Immunotherapy of far-advanced lepromatous leprosy patients with low-dose 
convit vaccine along with multidrug therapy (Calcutta trial). 
===================================================================== 
Author 
Majumder V; Mukerjee A; Hajra SK; Saha B; Saha K 
Address 
School of Tropical Medicine, Calcutta, India. 
Source 
Int J Lepr Other Mycobact Dis, 64(1):26-36 1996 Mar 
Abstract 
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. 

===================================================================== 
36.) Protective immunization of monkeys with BCG or BCG plus heat-killed 
Mycobacterium leprae: clinical results. 
===================================================================== 
Author 
Gormus BJ; Baskin GB; Xu K; Bohm RP; Mack PA; Ratterree MS; Cho SN; Meyers 
WM; Walsh GP 
Address 
Department of Microbiology, Tulane Regional Primate Research Center, 
Covington, LA 70433, USA. 
Source 
Lepr Rev, 69(1):6-23 1998 Mar 
Abstract 
Rhesus and sooty mangabey monkeys (RM and SMM) were vaccinated and boosted 
with BCG or BCG + low dose (LD) or high dose (HD) heat-killed Mycobacterium 
leprae (HKML). One group was not vaccinated. Except for a group of 
controls, all monkeys were challenged with live M. leprae. All animals were 
studied longitudinally to determine antileprosy protective efficacy. BCG 
reduced the numbers of RM with histopathologically-diagnosed leprosy by 70% 
and slowed and ameliorated the appearance of symptoms. BCG + LDHKML reduced 
the number of RM with leprosy by 89% and BCG + HDHKML by 78%. BCG did not 
protect SMM from developing leprosy, but disease progress was slowed; 
disease in SMM was exacerbated by the addition of HKML to the vaccine. RM, 
as a species, are prone to paucibacillary (PB) forms of leprosy, whereas 
SMM are prone to multibacillary (MB) forms. Thus, BCG vaccination offers 
significant protection from clinical disease and slows/ameliorates the rate 
of progression/degree of disease at the PB end and appears to at least 
ameliorate symptoms at the MB end of the leprosy spectrum. BCG + HKML 
protects at the PB end and exacerbates disease progress at the MB end of 
the leprosy spectrum. 

===================================================================== 
37.) Studies of vaccination of persons in close contact with leprosy patients in 
Argentina. 
===================================================================== 
Author 
Bottasso O; Merlin V; Cannon L; Cannon H; Ingledew N; Keni M; Hartopp R; 
Stanford C; Stanford J 
Address 
Instituto de Inmunologia, Facultad de Ciencias Medicas, Universidad 
Nacional de Rosario, Argentina. 
Source 
Vaccine, 16(11-12):1166-71 1998 Jul 
Abstract 
A total of 670 adults living or working with leprosy patients, were 
examined for a BCG vaccination scar, and skin-tested with four new 
tuberculins. Based on the results 513 were vaccinated, 65 with Bacille de 
Calmette et Gu&acute;erin (BCG) alone, 66 with BCG plus killed Mycobacterium 
vaccae and 382 with killed M. vaccae alone. Skin-testing was repeated 2-3 
years later on 344 subjects, when all three vaccines were found to have 
been highly successful in increasing responses to Tuberculin and Leprosin A 
(p < 0.0005) with increased immune recognition of common and 
species-specific antigens. Mean diameters of induration to each skin-test 
were greatest in recipients of BCG alone (p < 0.05), which suggests that 
better immuno-regulation occurs after receiving vaccines that incorporate 
M. vaccae. The results suggest 10(8) M. vaccae alone might prove a valuable 
future vaccine, which would not require selective pre-vaccination procedures. 

===================================================================== 
38.) Restoration of proliferative response to M. leprae antigens in lepromatous 
T cells against candidate antileprosy vaccines. 
===================================================================== 
Author 
Mustafa AS 
Address 
Department of Microbiology, Faculty of Medicine, Kuwait University, Safat, 
Kuwait. 
Source 
Int J Lepr Other Mycobact Dis, 64(3):257-67 1996 Sep 
Abstract 
Several studies conducted in the last decade suggest that Mycobacterium 
lepraereactive T cells exist in lepromatous patients, but their number may 
be too few to yield a detectable response in cell-mediated immunity (CMI) 
assays. Immunizations with candidate antileprosy vaccines and stimulation 
of T cells with M. leprae + interleukin-2 restore the M. leprae-induced CMI 
response in lepromatous leprosy patients. These immunizations and 
stimulation may enrich the pre-existing M. leprae-responsive T cells in 
lepromatous patients and, thereby, induce a detectable CMI response to M. 
leprae antigens upon repeat testing. To verify this proposition, we carried 
out a study in a group of 10 lepromatous leprosy patients. Peripheral blood 
mononuclear cells (PBMC) obtained from these patients were anergic to M. 
leprae antigens in proliferative assays, but they responded to the antigens 
of candidate antileprosy vaccines, i.e., M. bovis BCG, M. bovis BCG + M. 
leprae, and Mycobacterium w. The enrichment of M. leprae-responsive T cells 
was performed by establishing T-cell lines from the PBMC after in vitro 
stimulation with M. leprae, M. bovis BCG, M. bovis BCG + M. leprae, and 
Mycobacterium w. When tested for their proliferative responses, 1/10, 3/10, 
6/10 and 2/10 T-cell lines established against M. leprae, M. bovis BCG, M. 
bovis BCG + M. leprae, and Mycobacterium w, respectively, responded to M. 
leprae. These results suggest that enrichment of pre-existing M. 
leprae-responsive T cells may contribute to the restoration of the T-cell 
response to M. leprae in some lepromatous patients. Four of the 10 M. 
leprae-induced T-cell lines proliferated in response to the 65 kDa, 36 kDa, 
28 kDa, and 12 kDa recombinant antigens of M. leprae, suggesting that the 
nonresponsiveness of T cells in some lepromatous patients may be overcome 
by using recombinant antigens of M. leprae. 

===================================================================== 
39.) Does bacille Calmette-Gu&acute;erin scar size have implications for protection 
against tuberculosis or leprosy? 
===================================================================== 
Author 
Sterne JA; Fine PE; P&uml;onnighaus JM; Sibanda F; Munthali M; Glynn JR 
Address 
Communicable Disease Epidemiology Unit, London School of Hygiene and 
Tropical Medicine, UK. 
Source 
Tuber Lung Dis, 77(2):117-23 1996 Apr 
Abstract 
SETTING: Total population study in Karonga District, northern Malawi, in 
which the overall vaccine efficacy of bacille Calmette-Gu&acute;erin (BCG) has 
been found to be -7% against tuberculosis and 54% against leprosy. 
OBJECTIVE: To examine the relationship between BCG scar size and protection 
against tuberculosis and leprosy. DESIGN: Cohort study in which 85,134 
individuals were screened for tuberculosis and 82,265 for leprosy between 
1979 and 1984, and followed up between 1986 and 1989. RESULTS: Of the BCG 
scar positive individuals whose scars were measured, 31/3 2471 were later 
identified with tuberculosis and 81/31 879 with leprosy. In 19,114 
individuals, of whom 17 developed tuberculosis, tuberculin induration was 
measured at first examination. Mean scar sizes increased with increasing 
tuberculin induration in all except the oldest individuals. Mean scar sizes 
were lowest in individuals aged < 10 years, highest in individuals aged 
10-29 years and intermediate in older individuals. There was some evidence 
(P = 0.08) for an increase in tuberculosis risk with increasing scar size, 
which probably reflects the known correlation between scar size and 
tuberculin status at the time of vaccination. There was no clear 
association between BCG scar size and leprosy incidence. CONCLUSIONS: We 
find no evidence that increased BCG scar size is a correlate of 
vaccine-induced protective immunity against either tuberculosis or leprosy. 

===================================================================== 
40.) Protective efficacy of BCG against leprosy in S~ao Paulo. 
===================================================================== 
Author 
Lombardi C; Pedrazzani ES; Pedrazzani JC; Filho PF; Zicker F 
Address 
Pan American Health Organization, Bras&acute;ilia, Brazil. 
Source 
Bull Pan Am Health Organ, 30(1):24-30 1996 Mar 
Abstract 
The case-control study reported here evaluated the protective effect of BCG 
vaccine against leprosy in S~ao Paulo, Brazil. Seventy-eight patients under 
age 16 who had been diagnosed as having leprosy (cases) and 385 healthy 
individuals (controls) were selected and matched by sex, age, place of 
residence, and type of exposure to leprosy (intradomiciliary or 
extradomiciliary). The cases were drawn from an active patient registry and 
from a group of new leprosy cases treated at 50 health centers in the 
cities of Bauru and Ribeir~ao Preto in the state of S~ao Paulo. In order to 
estimate the protective effect of BCG, the prevalences of BCG scars in 
cases and controls were compared. The presence of one or more scars was 
associated with an estimated protective efficacy of 90% (95% confidence 
interval: 78% to 96%). Stratified analysis by age group, sex, socioeconomic 
level, and clinical form of the disease revealed no significant differences 
in the protection provided by the vaccine. However, it seems clear that 
more data will be needed in order to accurately assess the true relevance 
of BCG for leprosy control programs. 

===================================================================== 
41.) Post-vaccination sensitization with ICRC vaccine. 
Author 
===================================================================== 
Vallishayee RS; Gupte MD; Anantharaman DS; Nagaraju B 
Address 
CJIL Field Unit (ICMR), Madras. 
Source 
Indian J Lepr, 68(2):167-74 1996 Apr-Jun 
Abstract 
ICRC vaccine is one of the candidate anti-leprosy vaccines under test in a 
large scale comparative vaccine in trial. The objectives of the present 
study was to study the sensitization potential, as measured by Rees' MLSA 
and lepromin, and reactogenicity of this vaccine preparation in the local 
population. The study included 368 'healthy' individuals aged 1-70 years. 
Each individual received either ICRC vaccine or normal saline (control) by 
random allocation. They were also tested with Rees' MLSA and lepromin-A, 12 
weeks after vaccination. Reactions to Rees' MLSA were measured after 48 
hours and those to lepromin-A after 48 hours and three weeks. Character and 
size of local response, at the vaccination site, were recorded at 3rd, 8th 
and 15th week after vaccination. The results of the study showed that 
healing of vaccination lesion was uneventful, the mean size of the lesion 
being 10.3 mm. The mean sizes of post-vaccination reactions, to Rees' MLSA 
and lepromin (both early and late reactions), were significantly higher in 
the vaccine group compared to that in the normal saline group; the 
sensitizing effect attributable to the vaccine was of the order of 3.5 mm, 
1.7 mm and 2.2 mm respectively. In conclusion, the study has demonstrated 
that ICRC vaccine was 'safe' and produced significant sensitizing effect as 
measured by post-vaccination sensitization to Rees' MLSA and lepromin, in 
the local population. 

===================================================================== 
42.) Sensitization and reactogenicity of two doses of candidate antileprosy 
vaccine Mycobacterium w. 
===================================================================== 
Author 
Gupte MD; Vallishayee RS; Anantharaman DS; Britto RL; Nagaraju B 
Address 
CJIL Field Unit (ICMR), Avadi, Madras. 
Source 
Indian J Lepr, 68(4):315-24 1996 Oct-Dec 
Abstract 

M.w vaccine is one of the antileprosy vaccines under test in an ongoing 
comparative vaccine trial in South India. The objective of the present 
study was to examine the sensitizing ability, as measured by skin test 
reactions to Rees' MLSA and lepromin, and reactogenicity of M.w vaccine in 
the local population. Two doses of M.w, 1 x 10(9) bacilli and 5 x 10(9) 
bacilli, were used, in two separate studies of 395 and 400 "healthy" 
individuals aged 1-65 years. In each study, the study subjects received 
either M.w vaccine or normal saline (control), by random allocation. The 
results showed that healing of vaccination lesions was uneventful although 
the healing process was somewhat prolonged with the higher dose. The mean 
size of lesions was 7.0 mm and 9.5 mm with the low and high doses of the 
vaccine, respectively. The results also showed that M.w vaccine in a dose 
of 1 x 10(9) bacilli, failed to induce post-vaccination sensitization as 
measured by reactions to Rees' MLSA and by Fernandez and Mitsuda reactions 
to lepromin-A. However, when the dose of the vaccine was increased to 5 x 
10(9) bacilli the mean sizes of post-vaccination reactions to Rees' MLSA 
and lepromin-A (both early and late) were significantly larger in the 
vaccine group compared to that in the control group. The sensitizing effect 
attributable to the vaccine was of the order of 1.5 mm to 1.8 mm. 

===================================================================== 
43.) Tuberculin sensitivity and skin lesions in children after vaccination with 
two batches of BCG vaccine. 
===================================================================== 
Author 
Vallishayee RS; Anantharaman DS; Gupte MD 
Address 
CJIL Field Unit (ICMR), Avadi, Chennai. 
Source 
Indian J Lepr, 70(3):277-86 1998 Jul-Sep 
Abstract 
BCG is one of the vaccines used, as control arm, in an ongoing large scale 
comparative leprosy vaccine trial in South India. The objective of the 
present study was to examine, in the local population, the sensitizing 
ability, as measured by skin test reactions to tuberculin, and 
reactogenecity, in terms of skin lesions at the site of vaccination, for 
the two batches of BCG vaccine used in the above trial. The study was 
undertaken in 816 tuberculin-negative, previously not vaccinated school 
children, aged five to 14 years. Each child received one of the two batches 
of BCG vaccine or normal saline (control), by random allocation. At 12 
weeks from vaccination, character and size of local response, at the 
vaccination site, were recorded. At the same time, the children were 
retested with tuberculin and post-vaccination reactions to the test were 
measured after 72 hours. At three years after vaccination all available 
children were re-examined for the presence and size of BCG scar at the site 
of vaccination. It was found that healing of vaccination lesions was 
uneventful, with both batches of BCG. The mean size of the lesion was 
similar for the two batches, the overall mean being 6.3 mm. The mean size 
of post-vaccination tuberculin sensitivity increased with age, and it was 
14.5 mm and 15.6 mm. The sensitizing effect attributable to the vaccine was 
11 mm and 12 mm, for the two batches of BCG respectively. This study showed 
that the two batches of BCG, in a dose of 0.1 mg, used in the ongoing 
leprosy vaccine trial were acceptable in terms of vaccination lesion and 
were highly satisfactory in terms of development of hypersensitivity. 

===================================================================== 
44.) Association between leprosy and HIV infection in Tanzania. 
===================================================================== 
Author 
van den Broek J; Chum HJ; Swai R; O'Brien RJ 
Address 
Ministry of Health, Tuberculosis and Leprosy Central Unit, Dar es Salaam, 
Tanzania. 
Source 
Int J Lepr Other Mycobact Dis, 65(2):203-10 1997 Jun 
Abstract 
SETTING: An epidemiological study of the interaction of leprosy and HIV 
infection in Tanzania. OBJECTIVE: To establish the prevalence of HIV 
infection among leprosy patients, and to measure the association of HIV and 
leprosy by comparing the HIV prevalence in leprosy patients and blood 
donors. DESIGN: Testing for HIV infection in consecutively diagnosed 
leprosy patients (new and relapsed after MDT) in all regions in Tanzania 
successively for a period of 3 to 6 months during 1991, 1992 and 1993. 
RESULTS: Out of the total estimated eligible leprosy patients, 697 patients 
(69%) entered the final analysis. The HIV prevalence among these leprosy 
patients was 12% (83/697) as compared to 6% (8960/ 158,971) in blood donors 
examined in Tanzania during the same period. There were no significant 
differences in HIV seroprevalence by age, sex, residence or type of 
disease. However, the adjusted odds ratio (OR) of the presence of a BCG 
scar was 1.9 [95% confidence interval (CI) 1.1-3.3] among HIV-positive 
leprosy cases compared to HIV-negative leprosy cases. Comparing leprosy 
cases with blood donors as controls, the logistic regression model, 
controlling for sex, age group and residence, showed the OR for HIV 
seropositivity among leprosy patients to be 2.5 (95% CI 2.0-3.2). This 
association existed in all strata, but was strongest in the 15-34-year age 
group. No difference of HIV status between multibacillary and 
paucibacillary leprosy could be shown to exist. The point estimate of the 
population attributable risk of HIV infection for leprosy was 7%. 
CONCLUSION: HIV infection is associated with leprosy and might reverse the 
epidemiological trend of the slow decline in case notification in Tanzania 
if HIV infection is increasing greatly. Previous BCG vaccination loses its 
protection against leprosy in the presence of HIV infection. A repeated 
study is recommended in order to validate these findings, whereby recording 
of the disability grading of the cases is necessary to adjust for delay in 
diagnosis. 

===================================================================== 
45.) A follow-up study of multibacillary Hansen's disease patients treated with 
multidrug therapy (MDT) or MDT + immunotherapy (IMT). 
===================================================================== 
Author 
Rada E; Ulrich M; Aranzazu N; Rodriguez V; Centeno M; Gonzalez I; Santaella 
C; Rodriguez M; Convit J 
Address 
Instituto de Biomedicina, Caracas, Venezuela. 
Source 
Int J Lepr Other Mycobact Dis, 65(3):320-7 1997 Sep 
Abstract 
Multibacillary (MB) leprosy patients treated with multidrug therapy (MDT) 
or MDT + immunotherapy (IMT) with BCG + heat-killed Mycobacterium leprae 
were tested annually for their ability to proliferate in vitro to the 
mycobacterial antigens BCG, M. leprae soluble extract, and intact M. 
leprae. IgM antibody responses to phenolic glycolipid I (PGL-I) were 
measured, as well as serum nitrite levels in patients' sera, before, during 
and after treatment. Patients who received only MDT did not present 
cellular reactivity to intact M. leprae antigens, in contrast to the 
results obtained with BCG, which elicited reactivity at time zero, that 
increased after treatment. Regarding PGL-I antibody variations in relation 
to the initial value, we observed a statistically significant marked 
decrease at the end of 2 years which continued to fall in successive 
evaluations. MB patients showed high initial serum nitrite concentrations 
which dropped drastically with treatment. This decay was apparently 
associated with the bacillary load present in these patients. The group 
submitted to IMT + MDT showed high and long-lasting T-cell responses to 
mycobacterial antigens in a significant number of initially unresponsive MB 
patients. There was a marked increase to M. leprae soluble extract and BCG, 
as well as a more variable response to whole bacilli. The antibody levels 
in this group of patients are sustained for a somewhat longer period and 
decreased more slowly during the 5-year follow up. 
Language 

===================================================================== 
46.) Novel O-methylated terminal glucuronic acid characterizes the polar 
glycopeptidolipids of Mycobacterium habana strain TMC 5135. 
===================================================================== 
Author 
Khoo KH; Chatterjee D; Dell A; Morris HR; Brennan PJ; Draper P 
Address 
Department of Microbiology, Colorado State University, Fort Collins 80523, 
USA. 
Source 
J Biol Chem, 271(21):12333-42 1996 May 24 
Abstract 
Mycobacterium "habana" strain TMC 5135, which has been proposed as a 
vaccine against both leprosy and tuberculosis, is considered to be a strain 
of serotype I of the recognized species Mycobacterium simiae. We have now 
shown that each of these strains possesses characteristic polar 
glycopeptidolipids (GPL) which are sufficiently different to allow 
unequivocal strain identification. Thin layer chromatographic analysis 
demonstrated that M. habana synthesizes a family of apolar GPLs and three 
distinct polar GPLs (pGPL-I to -III) which exhibited migration patterns 
different from those of M. simiae serotype I (pGPL-Sim). Using a 
combination of chemical, mass spectrometric, and proton-NMR analyses, the 
GPLs from M. habana were determined to be based on the same generic 
structure as those from the M. avium complex, namely N-fatty 
acyl-D-Phe-(O-saccharide)-D-allo-Thr-D-Ala-L-alaninyl-O-m onosaccharide. 
The de-O-acetylated apolar GPLs contain a 3-O-Me-6-deoxy-Tal attached to 
the allo-Thr and either a 3-O-Me-Rha or a 3,4-di-O-Me-Rha attached to the 
alaninol. In the pGPLs, oligosaccharides were found to be attached to the 
allo-Thr. The oligoglycosyl alditol reductively released from the least 
polar pGPL-I was fully characterized as L-Fucp alpha 1 in --7 with 
3-(6-O-Me)-D-Glcp beta 1 in --7 with 3-(4-O-Me)-L-Rhap alpha 1 in --7 with 
3-L-Rhap alpha 1 in --7 with 2-(3-O-Me)-6-deoxy-Tal. In pGPl-II and -III, 
the terminal Fuc residue is further 3-O-methylated and 4-O-substituted with 
an additional 2,4-di-O-Me-D-GlcA and 4-O-Me-D-GlcA, respectively. The 
corresponding oligosaccharide from pGPL-Sim was shown to be of identical 
molecular weight to pGPL-II but terminating with a 3,4-di-O-Me-GlcA. 
Enzyme-linked immunosorbent assay-based serological studies using anti-M. 
habana and anti-M. simiae sera against whole cells and purified pGPLs 
firmly established the polar GPLs as important antigens and indicated that 
the terminal epitopes L-Fuc-, 2,4-di-O-Me-D-GlcA, and 4-O-Me-D-GlcA 
uniquely present in pGPL-I, -II, and -III, respectively, confer sufficient 
specificity for the identification of M. habana as a distinct serotype of 
M. simiae. 

===================================================================== 
47.) Regional lymphadenitis following antileprosy vaccine BCG with killed 
Mycobacterium leprae. 
===================================================================== 
Author 
De Britto RL; Ramanathan VD; Gupte MD 
Address 
CJIL Field Unit (Indian Council of Medical Research, Avadi, Madras, India. 
Source 
Int J Lepr Other Mycobact Dis, 65(1):12-9 1997 Mar 
Abstract 
Phase-II and extended Phase-II studies were conducted in three different 
sets of the population in Thiruthani Taluk, Chengalpattu District, South 
India, involving BCG and killed Mycobacterium leprae (KML) combination 
vaccines to ascertain the acceptability of the vaccines. In the Phase-II 
study, 997 healthy volunteers were vaccinated on individual randomization 
with one of the vaccines arms: BCG 0.1 mg + 6 x 10(8) KML, BCG 0.1 mg + 5 x 
10(7) KML, BCG 0.1 mg + 5 x 10(6) KML, BCG, 0.1 mg or normal saline. Blood 
samples were taken and the serum was tested for antibody levels against 
phenolic glycolipid-I (PGL-I) and the 35-kDa protein of M. leprae. In this 
study, we observed regional suppurative adenitis in 6% (6 out of 100), 3% 
(3 out of 100), and 3% (3 out of 100) of the vaccinees in the BCG 0.1 mg + 
6 x 10(8) KML, BCG 0.1 mg + 5 x 10(7) KML, and BCG 0.1 mg + 5 x 10(6) KML 
vaccine arms, respectively, in the 13-70 year age group. Earlier BCG scar 
status, skin-test reactions to lepromin-A, Rees' MLSA, and serum antibody 
levels against PGL-I and the 35-kDa protein did not help to identify the 
group at risk of developing suppurative adenitis. Suppurative adenitis 
appears to have a different relationship between the age of the subject and 
the dose of the vaccine. In order to overcome the problem of regional 
suppurative adenitis and to know the mechanism involved, an extended 
Phase-II study was conducted in similar groups of the population by 
reducing the BCG and KML doses, i.e., with BCG 0.05 mg + 6 x 10(8) KML, BCG 
0.05 mg + 5 x 10(7) KML, and BCG 0.01 mg + 5 x 10(7) KML. Biopsy specimens 
were collected from lymph nodes of the suppurative adenitis cases and were 
subjected for culture and histopathological examination. The observations 
showed that regional suppurative adenitis could be reduced to 1% in the BCG 
0.05 + 6 x 10(8) KML group, 0.5% in the BCG 0.05 + 5 x 10(7) KML group, and 
0.5% in the BCG 0.01 + 5 x 10(7) KML group. This phenomenon of suppurative 
adenitis appears to be related to the total dose of mycobacterial antigens. 
Suppurative adenitis was seen by weeks 18 and 20 post-vaccination in the 
latter two lower doses; whereas it was seen by week 8 in the higher dose of 
the combination vaccines. No case of suppurative adenitis was observed in 
the BCG 0.1 mg group. Culture and histopathology ruled out the 
possibilities of progressive BCG infection and superadded infection. 
Considering the above results, BCG 0.05 mg + 6 x 10(8) KML was acceptable 
for a large-scale vaccine trial in South India. 

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48.) A major T-cell-inducing cytosolic 23 kDa protein antigen of the vaccine 
candidate Mycobacterium habana is superoxide dismutase. 
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Author 
Bisht D; Mehrotra J; Dhindsa MS; Singh NB; Sinha S 
Address 
Division of Membrane Biology, Central Drug Research Institute, Lucknow, 
India. 
Source 
Microbiology, 142 ( Pt 6)():1375-83 1996 Jun 
Abstract 
This study describes the purification and immunochemical characterization 
of a major 23 kDa cytosolic protein antigen of the vaccine candidate 
Mycobacterium habana (TMC 5135). The 23 kDa protein alone was salted out 
from the cytosol at an ammonium sulfate saturation of 80-95%. It 
represented about 1.5% of the total cytosolic protein, appeared 
glycosylated by staining with periodic acid/Schiff's reagent, and showed a 
pl of approximately 5.3. Its native molecular mass was determined as 
approximately 48 kDa, suggesting a homodimeric configuration. 
Immunoblotting with the WHO-IMMLEP/IMMTUB mAbs mc5041 and IT61 and activity 
staining after native PAGE established its identity as a mycobacterial 
superoxide dismutase (SOD) of the Fe/Mn type. The sequence of the 18 
N-terminal amino acids, which also contained the binding site for mc5041, 
showed a close resemblance, not only with the reported deduced sequences of 
Mycobacterium leprae and Mycobacterium tuberculosis Fe/MnSODs, but also 
with human MnSOD. In order to study its immunopathological relevance, the 
protein was subjected to in vivo and in vitro assays for T cell activation. 
It induced, in a dose-related manner, skin delayed hypersensitivity in 
guinea-pigs and lymphocyte proliferation in BALB/c mice primed with M. 
habana. Most significantly, it also induced lymphocyte proliferative 
responses, in a manner analogous to M. Ieprae, in human subjects comprising 
tuberculoid leprosy patients and healthy contacts. 


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49.) Supervised Multiple Drug Therapy Program, Venezuela 
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Dr Jacinto Convit Director, Institute of Biomedicine, Caracas 

A supervised multiple drug therapy program (SMDT) for the treatment of 
leprosy has been in progress in our country since 1985. It has been 
supported by the Novartis Leprosy Fund since 1991. In contrast to the WHO 
MDT regime, the SMDT program provides a single treatment regime for both 
multibacillary (MB) and paucibacillary (PB) leprosy, differing only in the 
duration of treatment (two years for MB; six months for PB). Twice a month, 
health workers visit patients at home to supervise the taking of 
medication–600 milligrams of clofazimine each visit and 600 milligrams of 
rifampicine once a month. The daily 100 milligram dose of dapsone is 
checked indirectly with sulfone-in-urine tests done at random. 

The Venezuelan program also includes health education activities, 
examination of patients’ families, and a research program in connection 
with the quest for a leprosy vaccine. Once the treatment has been 
completed, former patients are kept under surveillance over a period of two 
(for PB) or five (for MB) years for a possible relapse of the disease. 

Our leprosy program in Venezuela has brought highly gratifying results. 
More than 4,200 patients have been cured and are now under post-treatment 
surveillance; a further 3,000 are still in treatment. Although the number 
of newly detected cases has scarcely changed, averaging around 450 a year, 
the number of patients undergoing treatment has gone down distinctly. The 
program’s activities have also brought about an improved public attitude to 
the disease. Most new patients seek treatment on their own initiative, and 
the manifest improvement in the condition of those who have been treated is 
the best publicity for the program. 

To secure the success of the leprosy program we have had to reorganize the 
Public Health Dermatology Services and reinforce their infrastructure and 
central data registration system. Carrying out the program of visits at 
patients’ homes necessitated providing the health workers with 
transportation and allowances to defray travel expenses. Finally, an 
extensive health education program had to be mounted so as to ensure that 
patients come regularly for follow-up examinations after they are cured. 

Not least thanks to the backing we have received from the Novartis Leprosy 
Fund, we have been able to solve all these problems or move them closer to 
a solution. 

Our future efforts will be directed toward integrating our leprosy work 
with the control of other endemic diseases such as tuberculosis, 
leishmaniasis, and Chagas’ disease. The training programs for this are now 
under way, and some are already completed. In future, MDT as recommended by 
the WHO will be used. We also plan to develop a vaccination program in 
conjunction with the current curative program and, through further research 
projects, to improve early diagnosis. 

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50.) NII DEVELOPES WORLD'S FIRST ANTI-LEPROSY VACCINE 
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The National institute of Immunology, New Delhi has developed 
Anti-Leprosy vaccine and has conducted Phase I,II and III clinicaltrials to 
study its immunotherapeutic and immunoprophylactic effects in leprosy 
patients. The development of this vaccine was initiated during early 1980s. 
Phase II clinical trial of this vaccine was launched in December 1986 in 
two Urban Leprosy centres of Delhi namely safdarjung Hospital and Dr. Ram 
Manohar Lohia Hospital after obtaining due approval of Drugs controller 
Genral of India and the Institutional Ethics commuttee of Hospitals. 

Patients receving this vaccine as adjunct to multidrug therapy (MDT) 
have shown repid clinical improvement, bacteriological negativity and 
histopathological upgradation. This observation of hastening of healing 
induced by Mw vaccine has been consistent from 1987 till date. The vaccine 
is also free from any serious side effects and well accepted by rural as 
well as urban population. The encouraging results of clinical trials in 
Delhi Urban Leprosy Centres led the Institute to expand the trials in 
larger population in field situation in Kanpur Dehat. The vaccine was 
tested on Leprosy patients as well as their healthy households contacts. 
The data produced has been thoroughly examined by two separate Expert 
Committees constituted by the Department of Biotechnology. The statistical 
analysis of the immunotherapeutic data of Mw vaccine with MDT shows the 
improvement in clinical profiles of the leprosy patients as early as six 
months. 

The technology for manufacture of the product has been trasferred to 
M/s cadila Pharmaceuticals, Ahmedabad. Drugs controller General of India 
has provided the clearance for commercialisation of this vaccine to M/S. 
Cadila Pharmaceuticals. Cadila will soon launch the product in the market, 
witth a mechanism for post market surveillance. Looking at the problem 
globally, although leprosy is found in about 80 countries in Asia, Africa 
and Latin America, India alone contributes to about 60 per cent of the 
global pool of leprosy patients. Though the number of leprosy patients in 
the world have reduced from approximately 12 million to 6 million from 1985 
to 1995, there are difficulties in accurate estimation of disease burden 
due to ambiguity in early detection of the disease and self-healing nature 
in a large number of cases. 

This is the first anti-leprosy vaccine developed in the world. while 
its immunotherapeutic effects have been well established, its role for 
immunoprophylaxis is being examined by regulatory agencies. There are 
indicators showing that the vaccine has profound effects on healthy 
household contacts. 

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DATA-MEDICOS/DERMAGIC-EXPRESS No (68) 11/08/99 DR. JOSE LAPENTA R. 
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