Limited susceptibility of cynomolgus monkeys (Macaca fascicularis) to leprosy after experimental administration of Mycobacterium leprae.

Cynomolgus monkeys are a useful model for human tuberculosis, but susceptibility to M. leprae is unknown. A cynomolgus model of leprosy could increase understanding of pathogenesis-importantly, neuritis and nerve-damaging reactions. We administered viable Mycobacterium leprae to 24 cynomolgus monkeys by three routes, with a median follow-up period of 6 years (range = 1-19 years) involving biopsies, nasal smears, antiphenolic glycolipid-1 (PGL-1) antibody serology, and lepromin skin testing. Most developed evanescent papules at intradermal M. leprae inoculation sites that, on biopsy, showed a robust cellular immune response akin to a lepromin skin test reaction; many produced PGL-1 antibodies. At necropsy, four monkeys, without cutaneous or gross neurological signs of leprosy but with elevated PGL-1 antibodies, including three with nasal smears (+) for acid fast bacilli (AFB), showed histological features, including AFB, suggestive of leprosy at several sites. Overall, however, cynomolgus monkeys seem minimally susceptible to leprosy after experimental M. leprae administration.

Longitudinal ocular survey of 202 Filipino patients with multi-bacillary (MB) leprosy treated with 2 year WHO-multiple drug therapy.

The aim of this study was to describe the ocular conditions in multibacillary (MB) leprosy patients treated with 2 year WHO multiple drug therapy (MDT), consisting of dapsone, clofazimine and rifampin, a regimen expected to reduce ocular complications of leprosy. We conducted comprehensive eye examinations in 202 Filipino MB leprosy patients before, during, and after WHO 2 year MDT. Assessments were carried out for at least 5 years. Inflammatory "lepra" reactions occurred in 62% (reversal reaction, 52%; erythema nodosum leprosum, 10%); most were mild. Eye abnormalities consisted mostly of diminished corneal sensitivity before MDT (6%) and lagopthalmos (n = 7, 3.4%). Six of 7 lagopthalmos cases occurred in a subset of 132 patients with facial patches (5%). Visual acuity scores, intra-ocular pressures and pupil cycle times were unremarkable. Bacillary invasion, keratitis, episcleritis, iridocyclitis, ectropion, synechiae, glaucoma and cataract formation were not detected. Scleral clofazimine pigmentation was frequent, resolving in most within 3 years of treatment cessation. Facial patches at presentation may denote a higher risk for lagopthalmos. We propose the generally low rates of ocular problems reflected mild lepra reactions, due to anti-inflammatory properties of clofazimine, a relatively young cohort, and a readily accessible community-based clinic permitting earlier diagnosis and prompt treatment.

Long-term relapse risk of multibacillary leprosy after completion of 2 years of multiple drug therapy (WHO-MDT) in Cebu, Philippines.

From 1987 to 1994, we enrolled 500 subjects completing 2-year WHO multiple drug therapy (MDT) for multibacillary leprosy in a prospective relapse study. Relapse was defined as new skin lesions and an increase in the bacterial index (BI) > or = 2+ (> or = 100x) at any single slit-skin smear site. At the study end in 2006, follow-up was 6,401 subject-years, a mean of 12.8 years/subject. We observed 23 relapses, 6-16 years after MDT (mean, 10.5 years; 95% confidence interval [CI], 9.2-11.8), peaking in Years 11-12 (> 1%/year). The cumulative risk was 6.6% (95% CI, 5.0-8.2%). In a subset of 181 subjects with pre-MDT average BI > or = 4+, 11 relapses occurred (cumulative risk, 10.1%). In mouse footpad assays, Mycobacterium leprae from relapsed subjects were rifampin and clofazimine sensitive. Taken together, the data suggest relapses are related to activation of dormant organisms (persisters) not killed by MDT rather than new infection.

Clinical and histologic features of skin lesions in a cynomolgus monkey experimentally infected with mycobacterium ulcerans (Buruli ulcer) by intradermal inoculation.

Buruli ulcer, caused by Mycobacterium ulcerans, is a destructive infection that most commonly affects the skin. Animal models for Buruli ulcer include guinea pigs, rats, mice, and armadillos, but each is limited in replicating the spectrum of human disease. Here, a cynomolgus monkey was infected with two concentrations of M. ulcerans (1.0 and 2.2 x 10(8)) by intradermal inoculation, 3 months apart. All injection sites developed papules that progressed to ulcers with undermined borders within 2-4 weeks. The rate of progression and size of the ulcers were proportional to the numbers of organisms inoculated. Biopsies from ulcer edges showed ulceration, robust inflammatory cell infiltrates, granulomatous-like responses, mild edema, and extracellular acid-fast bacilli. The ulcers healed spontaneously between Weeks 8 and 12, with no signs of systemic infection. This report, the first to describe a non-human primate experimentally infected with M. ulcerans, suggests that cynomolgus monkeys are modestly susceptible and develop some of the clinical and histologic features of Buruli ulcer.

A randomized, double-blind, double-dummy, controlled dose comparison of thalidomide for treatment of erythema nodosum leprosum.

In a randomized, double-blind, double-dummy controlled study, 22 men with erythema nodosum leprosum (ENL) received six capsules containing either 100 mg (group A, n = 12) or 300 mg (group B, n = 10) of thalidomide daily for one week. A six-week, four capsules per day taper followed, in which group A received 50 mg/day of thalidomide in weeks 2 and 3, then dummy capsules in weeks 4 through 7, while group B had gradual decrements every two weeks. Both regimens caused comparable improvement in 19 patients at day 7 (group A [12 of 12] versus group B [7 of 10]; P = 0.08), but slower tapering in group B showed less re-emergence of ENL through week 7 (P = 0.02, versus group A). Most patients developed new lesions soon after stopping treatment. Slower tapering from a higher initial thalidomide dose may improve clinical ENL responses, but high recurrence rates after discontinuation indicates further assessment is needed to identify better tapering regimens.

Parallel assessment of 24 monthly doses of rifampin, ofloxacin, and minocycline versus two years of World Health Organization multi-drug therapy for multi-bacillary leprosy.

Monthly doses of rifampin, ofloxacin, and minocycline (ROM) are expected to be effective treatment for multi-bacillary leprosy. Patients with MB leprosy received ROM (n = 10) or World Health Organization multi-drug therapy (MDT) (n = 11). Treatment with ROM was given as 24 consecutive monthly observed doses of rifampin (600 mg), ofloxacin (400 mg), and minocycline (100 mg). Treatment with MDT was given as 24 consecutive monthly observed doses of rifampin (600 mg) and clofazimine (300 mg), and unobserved daily dapsone (100 mg) and clofazimine (50 mg). Twenty patients completed the 24-month regimens with > 99% compliance. Treatments with ROM and MDT were safe, tolerable, and caused similar improvements in lesions, bacterial indices, and histology. All MDT recipients developed clofazimine-induced pigmentation. Six ROM and nine MDT recipients assessed at five or more years after completion of treatment had no evidence of relapse. Twenty-four months of treatment with ROM is a safe, well-tolerated, and convenient regimen that may provide an alternate therapy to MDT for MB leprosy. Larger trials with sufficient follow-up would better define the role of ROM.

Long-term efficacy of 2 year WHO multiple drug therapy (MDT) in multibacillary (MB) leprosy patients.

Relapse rate estimates after 2 year WHO multiple drug therapy (MDT) in multi-bacillary (MB) leprosy vary. Between 1987 and 1994, 500 MB leprosy patients completing 2 year MDT were enrolled in a prospective relapse study. The majority of patients (N = 316) were treated and followed at the physician-staffed Cebu Skin Clinic (CSC), whereas others (N = 184) received therapy from government clinics and were followed by CSC technicians in the field. Relapse definition was an increased bacteriologic index (BI) and new skin lesions, supplemented with mouse footpad inoculations. Through 2002, follow-up was 5368 person-years, with a mean of 10.8 years per patient. The absolute relapse rate was 3% (15/498; 0.28/100 person-years), with a cumulative risk estimate of 3.9% at 15 yrs. For a subset of 217 patients followed for >or=12 yrs or until relapse, relapses occurred in 9% (13/142) attending the CSC, versus 3% (2/75) assessed in the field (p = 0.09). The rate for patients followed at CSC for >or=12 yrs and a pre-treatment BI >or=2.7+ was 13% (13/98). All relapses were BL or LL, with pre-treatment BI's of >or=2.7+. Relapses occurred long after completion of therapy, between 3 and 11 yrs from the midpoint of the examination without relapse to detection, or between 6 to 13 yrs to the actual year of detection, 7 occurring at >or=10 yrs. Lesion material from all relapses contained M. leprae that was rifampin and clofazimine sensitive, whereas 3 showed partial or full dapsone resistance. [Follow-up rigor and time], medical expertise, and pre-treatment bacterial load influence relapse rates after 2 yr MDT.

Antileprosy protective vaccination of rhesus monkeys with BCG or BCG plus heat-killed mycobacterium leprae: immunologic observations

Groups of rhesus monkeys were vaccinated and boosted with Mycobacterium bovis bacillus Calmette Guerin (BCG) or BCG plus low-dose (LD) or high-dose (HD) heat-killed M. leprae (HKML), or were unvaccinated. Prior to and following vaccination-boosting and subsequent M. leprae (ML) challenge, these and unvaccinated, unchallenged control monkeys were observed longitudinally for approximately 3 years. Vaccination with BCG plus HKML initially stimulated significant in vitro blood mononuclear cell blastogenic responses to lepromin, which returned to baseline post-boosting and post-live-ML-challenge, minimally reappearing significantly 2 years post-ML-challenge. Vaccination with BCG failed to stimulated positive blastogenic responses to lepromin before ML-challenge but small, marginally positive, intermittent responses were seen post-ML-challenge. Compared to the unvaccinated ML-challenged group, significant increases in the numbers of blood CD4+ and CD8+ T-cell subsets and an increased CD4+:CD8+ ratio were observed in both BCG plus HKML-vaccinated, ML-challenged groups, but not in the BCG-only-vaccinated, ML-challenged group. CD4+CD29+ and CD4+CD45RA+ subset numbers increased significantly over time in only the BCG plus LD HKML-vaccinated, ML-challenged group. Compared to unvaccinated, ML-challenged groups, vaccination with BCG or BCG plus HKML followed by ML-challenge produced lower IgM:IgG antiphenolic glycolipid-I (PGL-I) serum antibody ratios and protected rhesus monkeys from clinical leprosy, consistent with prior observations that low IgM:IgG anti-PGL-I responses correlated with resistance to and protection from leprosy.

Minocycline in Lepromatous Leprosy

Twelve patients were treated with three dose levels of minocycline for 30 days, primarily to detect the dose-related effects on Mycobacterium leprae viability, followed by another 5 months of daily minocycline for overall efficacy and persistence of clinical and antibacterial effects. Subsequently, the patients were given standard WHO/MDT chemotherapy for multibacillary leprosy. Clinical improvement was recognizable during the first month, occurring much earlier among those on minocycline 200 mg daily than those who received minocycline 100 mg daily. A similar change also was observed in one patient 11 days after three daily doses of 100 mg of minocycline. At the end of 6 months, all patients were clinically improved with a slight reduction in the average bacterial index (BI) and logarithmic index of bacilli in biopsy (LIB). The effects of minocycline on viability by mouse foot pad inoculation and palmitic acid oxidation assays were noted beginning at 10 to 14 days of daily dosing and becoming more definite after 30 days of treatment. Both tests correlated fairly well. Doses of 200 mg daily did not appear to be more efficient than minocycline 100 daily. Phenolic glycolipid-I (PGL-I) antigen determinations done on some patients during the first month remained positive and did not correlate with changes in viability results. At the end of 6 months, after 5 months of 100 mg of minocycline monotherapy, no viable organisms could be demonstrated by mouse foot pad inoculation and palmitic acid oxidation assays; assays for PGL-I antigen were all negative