A case of relapse with drug-susceptible M. leprae after multidrug therapy.

A male born in 1930 was diagnosed as smear-positive borderline leprosy in 1971, and was treated with dapsone and/or sulfamethoxypyridazine from 1972 to 1980 with clinical improvement. However, new skin lesions with smears strongly positive appeared in August 1980, and he was diagnosed as having downgraded to lepromatous (LL) leprosy, but the bacilli recovered from the skin biopsy were fully susceptible to both dapsone and rifampin by mouse foot pad technique. Between 1981 and 1983, the patient was treated with 24 months of rifampin 600 mg and dapsone 100 mg daily, supplemented with prothionamide 500 mg daily during the initial 3 months, and his skin lesions gradually improved during treatment with the combined regimen. Afterward, the patient was kept under surveillance without treatment. From 1984 to 1986, his skin smears were negative, and no bacilli could be found from a skin biopsy taken in 1985. Then in 1987, 52 months after stopping treatment, new skin lesions appeared with a high concentration of Mycobacterium leprae (2 x 10(6)/mg tissue). The drug-susceptibility test again demonstrated that the organisms were fully susceptible to both dapsone and rifampin. Apparently the relapse was due to remultiplication of drug-susceptible persisters.

Clinical trial of pefloxacin and ofloxacin in the treatment of lepromatous leprosy.

Twenty-one previously untreated lepromatous patients were randomized into two groups and treated with either 800 mg pefloxacin (PEFLO) or 400 mg ofloxacin (OFLO) once daily. The trial consisted of two parts: monotherapy from day 0 to day 56; and combined with the World Health Organization multidrug therapy (WHO/MDT) regimen for multibacillary (MB) leprosy from day 57 to day 180. Four patients were removed from the trial because the organisms recovered from their pretreatment biopsies failed to infect mice. Among the remaining 17 cases, four (23.5%) had primary resistance to dapsone but all of them were susceptible to rifampin. The initial (day 0) proportion of viable organisms, as measured by mouse foot pad inoculation, varied tremendously from patient to patient despite randomization during admission. Definite clinical improvement was noticed in virtually all patients after 22 doses of treatment with either PEFLO or OFLO. A significant fall in the morphological index (MI) occurred as early as after 8 doses of PEFLO or after 22 doses of OFLO; the bacterial load also showed a moderate degree of reduction during the period of monotherapy. Although single-dose PEFLO or OFLO displayed only a modest degree of bactericidal effect against Mycobacterium leprae, about 99.9%, or 4 logs, of organisms viable on day 0 were killed by 22 doses of either PEFLO or OFLO. No significant difference in the therapeutic effect was detected between the two regimens. During PEFLO or OFLO monotherapy, except in one patient (case no. 10), the side effects were few and mild. Case no. 10 developed a psychic disorder after 27 days of PEFLO monotherapy, presumably due to the treatment with PEFLO. All of the patients tolerated the period of combined therapy extremely well, although some asymptomatic and transient laboratory abnormalities were observed. Because both PEFLO and OFLO displayed rapid bactericidal activities in human leprosy and were well tolerated by the patients, further clinical trials and field trials in evaluating the therapeutic effects of combined regimens containing both rifampin and PEFLO or OFLO are being organized. Since this is the first clinical trial in leprosy employing nude mice, in combination with normal mice, for monitoring the therapeutic effects of antimicrobials, the advantages, limitations and appropriate timing in using nude mice are discussed.

Effectiveness of pefloxacin in the treatment of lepromatous leprosy.

As a first clinical trial of a fluoroquinolone derivative in leprosy, ten previously untreated lepromatous leprosy patients, about two fifths of them with primary dapsone resistance but all susceptible to rifampin, were treated with pefloxacin 400 mg twice daily for 6 months. Definite clinical improvement was observed in all ten patients as early as 2 months after beginning treatment, and the morphological index was also drastically decreased to the baseline during the same period. The rapid bactericidal effects, as measured by serial mouse foot-pad inoculations, were demonstrated to the extent that about 99% of the bacilli were killed during the first 2 months of treatment. However, the bacterial load, in terms of the bacterial index and the number of acid-fast bacilli per mg of tissue, of the patients was only moderately reduced. The side effects were mild, and the patients tolerated the treatment well.

Study of 39 documented relapses of multibacillary leprosy after treatment with rifampin.

Among 39 strains of Mycobacterium leprae isolated from patients with multibacillary leprosy who relapsed after treatment with rifampin (RMP), 22 strains were resistant to RMP and 17 were susceptible. All of the RMP-resistant strains were recovered from patients who had been treated with more than 50 doses of RMP, usually given as monotherapy. RMP-susceptible strains were recovered from only six patients who had received more than 50 doses of RMP, and from 11 patients who had received no more than seven doses. The median time to relapse after the beginning of RMP therapy was 9 years (range 1-12 years) among the patients harboring RMP-resistant strains of M. leprae, and the median time to relapse after discontinuation of RMP treatment was 7 years (range 1-11 years) among the patients harboring RMP-susceptible strains. These data suggest that monotherapy with more than a few doses of RMP can be responsible for the emergence of RMP-resistant strains of M. leprae, thus emphasizing the need to employ RMP only in combination with other effective drugs in the chemotherapy of multibacillary leprosy.

Activity of ofloxacin against Mycobacterium leprae in the mouse.

Mice inoculated with 4800 Mycobacterium leprae in the left hind foot pad were treated from day 62 to day 150 after infection with 50 mg or 150 mg of ofloxacin per kg body weight, 150 mg pefloxacin per kg, or 50 mg prothionamide per kg. These drugs were administered by esophageal cannula 5 days weekly with dapsone (0.01 g per 100 g diet). Multiplication of M. leprae in the treated and in untreated control mice was assessed by monthly harvests. The treatment of mice with the smaller dosage ofloxacin, with pefloxacin, prothionamide, or dapsone uniformly resulted in a delay of multiplication of 4 months, compared to the multiplication of M. leprae in the untreated controls. The delay of multiplication (4 months) being 1 month longer than the duration of drug administration (3 months), all of the treatments may be considered as bacteriopausal or moderately bactericidal. In contast with these results, treatment of mice with 150 mg ofloxacin per kg resulted in no growth of the organisms whatever as late as 18 months after inoculation, strongly suggesting that, in that dosage, ofloxacin had killed all of the M. leprae. Such a profound killing activity has been observed only with rifampin. Although the pharmacokinetic characteristics of ofloxacin are different in man from those in the mouse, the daily dosage of 150 mg ofloxacin per kg body weight in the mouse is equivalent to 400 mg per day in man which is the usual therapeutic dosage; thus, the results obtained in the mouse may be extrapolated to man. Therefore, ofloxacin appears a very promising drug for the chemotherapy of leprosy.

Primary and secondary dapsone resistance of M. leprae in Martinique, Guadeloupe, New Caledonia, Tahiti, Senegal, and Paris between 1980 and 1985.

Primary and secondary dapsone resistance were studied among lepromatous patients living in Martinique, Guadeloupe, New Caledonia, Tahiti, Senegal, and Paris. Four hundred fifteen biopsies were taken from clinically active and bacteriologically positive (bacterial index greater than 2) patients in the 6-year period of 1980-1985. Among these, 280 biopsies that contained 5 x 10(4) acid-fast bacilli per ml with a morphological index of at least 0.10 were inoculated into the mouse foot pad, and 229 harbored infective Mycobacterium leprae. Among the 129 infective M. leprae isolated from new cases, 54% had some degree of dapsone resistance, a low degree being prominent in all cases. Among the 100 infective M. leprae isolated from relapsed cases, 79% had a high or an intermediate degree of dapsone resistance. The annual incidence of secondary dapsone resistance was estimated to be about 0.55% in Guadeloupe.

Multiplication of Mycobacterium leprae in the nude mouse, and some applications of nude mice to experimental leprosy.

Two aspects of the immune deficiency of nude mice make these animals particularly useful tools for leprosy research. Nude mice are capable of supporting multiplication of M. leprae to levels approaching 10(10) per g in peripheral body tissues. In addition, nude mice may be inoculated with greater than 10(4) (in fact, with as many as 10(8) organisms per foot pad, without provoking an immune response that prevents multiplication of the organisms. Thus, the nude mouse should be particularly suitable for detecting persisting M. leprae in treated patients, and as a model of the patient for evaluating chemotherapeutic regimens.

[Resistance of Mycobacterium leprae to dapsone and rifampicin: apropos of a survey carried out in the Cape Verde region (Senegal)]. / La résistance de Mycobacterium leprae à la dapsone et à la rifampicine: à propos d'une enquête effectuée dans la région du Cap-Vert (Sénégal).

Since 1983, primary resistance of M. leprae to DDS and rifampicin has been evaluated in new cases of lepromatous leprosy observed in the Cap-Verde region in Senegal. Out of the 13 strains isolated, 10 (77%) have been found resistant to DDS, 7 at low level, 2 at intermediate level, 1 at high level; all of them have been found sensible to rifampicin. Similar results have been obtained with 57 strains isolated from patients not yet treated coming from different geographical areas, seeing that 37, i.e. 65%, were resistant to DDS, 27 at low level, 5 at intermediate level and 5 at high level; all of them were sensible to rifampicin. Level of resistance to DDS is very different in case of acquired resistance. In 69 lepromatous patients treated for more than 5 years and showing a relapse, M. leprae was 62 times, i.e. 90%, resistant to DDS, 6 times at low level, 21 at intermediate level and 35 at high level; in addition, 13 times M. leprae was resistant to rifampicin. In order to avoid and to solve problems set by resistance of M. leprae to antibiotics, strict application of polychemotherapy of leprosy is compulsory.

Activities of pefloxacin and ciprofloxacin against Mycobacterium leprae in the mouse.

Because ciprofloxacin and pefloxacin are fluoroquinolones active against many mycobacterial species, both drugs were tested against Mycobacterium leprae in the mouse foot-pad system. Preliminary pharmacokinetic studies in the mouse showed that after a single oral dose of 150 mg/kg ciprofloxacin the peak serum concentration was 3.6 micrograms/ml, and after 50 mg/kg or 150 mg/kg pefloxacin peak serum concentrations were, respectively, 9.2 micrograms/ml and 16.9 micrograms/ml, the half-lives for serum elimination being about 2 hr for both drugs. The activity of daily 50 mg/kg and 150 mg/kg ciprofloxacin and pefloxacin against M. leprae was then tested in mice infected with 5 X 10(3) M. leprae. The growth of M. leprae was not prevented in mice treated continuously with either 50 mg/kg or 150 mg/kg ciprofloxacin, indicating that this drug had no or a limited bacteriostatic effect at the dosages used. In mice treated continuously with 50 mg/kg pefloxacin, growth of M. leprae was not prevented, but at monthly harvests the number of bacilli in the foot pads remained less than those of control mice (p less than 0.05). No growth of M. leprae occurred in mice treated continuously with 150 mg/kg pefloxacin. In mice treated for only 3 months with daily 150 mg/kg pefloxacin, the growth-delay that followed the stopping of the drug was 126 days, suggesting that approximately 99% of the M. leprae were killed. The pharmacokinetics of pefloxacin being more favorable in man than in the mouse, pefloxacin appears a possible drug for the chemotherapy of leprosy.

Experimental leprosy in the armadillo and nude mice: comparative histobacteriology and ultrastructure.

In last 14 years, armadillo has proved an ideal animal model for studying experimental leprosy and mass production of Mycobacterium leprae. However recently a number of groups working with nude mice have claimed its ability as a better experimental model as far as leprosy research and production of leprosy bacilli is concerned. We therefore decided to compare experimental M. leprae infection of both armadillo and nude mice. We compared the degree of infection as well as the physiological and morphological state of proliferating bacteria by histobacteriological and electron microscopic studies. Histobacteriological studies of the infected nude mice showed that the highest number of acid-fast bacilli (AFB) was present in the leprome formed at the site of inoculation, the foot-pad, but often the morphological index (MI) was low. A bacillary diffusion was observed, however the internal organs did not show extensive lesions and contained few AFB with a still lower MI. Similarly the AFB present in the nodes were few in number, formed smaller globi than those formed in the foot-pad, and were essentially non-solid staining bacilli with a lower MI. In the armadillo on the other hand, lesions were extensive and apparent in the internal organs, a much higher AFB count was found and the MI was higher than in nude mice. Ultrastructural studies showed that a much higher proportion of M. leprae cells inside armadillo tissues existed as intact bacteria than in nude mice. Essentially damaged bacteria were found in the nodes from nude mice. The number of M. leprae bacilli in lung, liver, spleen and kidney from all the nude mice was too little to be studied under the electron microscope. Intact bacilli were observed only in mice foot-pads, which however contained two distinct populations of M. leprae; the intact and the damaged bacilli, which were arranged together in separate globi.

Hepatitis in leprosy patients treated by a daily combination of dapsone, rifampin, and a thioamide.

A 13% incidence of hepatitis was observed among 54 cases of multibacillary leprosy treated daily with the three-drug combination of dapsone, rifampin, and a thioamide (ethionamide or prothionamide). No hepatitis was observed among 109 cases of paucibacillary leprosy treated daily with the two-drug combination of dapsone and rifampin. Symptoms were jaundice in five cases and nausea plus vomiting associated with a significant increase of transaminase levels in two cases. In five cases, the symptoms appeared during the first two months of therapy and in two cases, later. Discontinuing treatment with rifampin and the thioamide but not dapsone resulted in recovery. When rifampin was resumed without the thioamide, the hepatitis did not recur. Viral etiology could be eliminated in six cases. Neither sex, age, weight nor the fact that the patient was a new case or a relapse case appeared to be a contributing factor. Hepatotoxicity caused by administration of a thioamide might have been potentiated by the concurrent administration of rifampin.

[Activity of rifampicin administered daily and intermittently on experimental tuberculosis in mice]. / Activité de la rifampicine administrée quotidiennement et d'une manière intermittente sur la tuberculose expérimentale de la souris.

Intermittent regimens with rifampicin (RMP) for chemotherapy of pulmonary tuberculosis are cheaper and can be usually supervised more easily than daily regimens. They are recommended, specially, in developing countries. The present study is designed to evaluate the reduction of RMP activity as the interval between doses is increased in experimental mouse tuberculosis. The mice are given 10 mg/kg oral doses of RMP once, twice, three times or six times a week. The RMP activity is measured as the decrease of viable counts (cfu) in the spleen of mice. RMP given once a week has no activity neither in combination with isoniazid and streptomycin on a large population of organisms nor alone on a small population. RMP given alone twice a week has an activity slightly better than RMP given once a week. RMP given alone three times a week has a weak but not worthy activity: the mean number of cfu after a three times weekly treatment is significantly lower than the mean number of cfu in control (p less than 0.001) but significantly higher than the mean number of cfu after a six times weekly treatment (p less than 0.001). These experimental results are consistent with the experimental data on the effects of RMP on the RNA polymerase. They are not favourable to the intermittent administration of RMP in the chemotherapy of tuberculosis.

[Current data on the bacteriology of leprosy]. / Données actuelles sur la bactériologie de la lèpre.

Microscopic examination and mouse foot-pad inoculation are the main tools in Mycobacterium leprae bacteriology. Microscopic examination enables to calculate bacteriological (IB) and morphological (IM) indexes. The former scores the density of acid-fast organisms in the lesions and the latter scores their viability since only solidly stained acid-fast organisms are likely to be alive. Mouse foot-pad inoculation provides the opportunity for studying the growth curve of M. leprae. Alterations of this growth curve under drug therapy enable to assess the antileprosy activity of drugs and to determine the drug sensitivity of organisms. Acquired and primary drug resistances to DDS are to date the most important events in the chemotherapy of leprosy. Researches are in progress on mice with congenitally reduced immunological capacity (nu/nu mice) and on antileprosy vaccination. Although no decisive advances have been obtained in these fields, researches in progress are promising.