Expression profile of CXCL12 chemokine during M. tuberculosis infection with different therapeutic interventions in guinea pig.

Mycobacterium indicus pranii (MIP) already established as an immune-modulator in mycobacterial infections generates immune response by acting on CXC chemokines. In the present study, the immunomodulatory effect of MIP in conjunction with chemotherapy against M.tb infection was evaluated by colony forming units (CFUs) following aerosol infection to guinea pig and by measuring CXCL12 chemokine expression using q-PCR and in situ RT-PCR. Different experimental groups included, infection (Rv), immunoprophylaxis (RvMw), chemotherapy (RvCh) and combination of immunoprophylaxis+chemotherapy (RvChMw) group and normal healthy (NH) group. In the combination of immunoprophylaxis+chemotherapy (RvChMw) group, the CFU counts reduced significantly (p<0.001) at 4th week of infection as compared to other treated groups (RvMw and RvCh group). The expression of CXCL12 was recorded in all the treated groups of animals. The study demonstrated suppressed expression of CXCL 12 in both immunoprophylaxis as well as chemotherapy groups (6th and 8th week) that become elevated in immunoprophylaxis plus chemotherapy group (10th week), at which time point no CFUs were detected in RvCh and RvChMw group. The findings indicate that the expression of CXCL12 is associated with good response to anti - tubercular treatment. Thus, prior immunization with MIP appears to show good immunomodulatory effect to release CXCL12 chemokine during infection and also correlates with enhanced effect to chemotherapy.

Predictors of unfavourable treatment outcome in patients with multidrug-resistant tuberculosis in India.

Setting: India has one of the highest global rates of multidrug-resistant tuberculosis (MDR-TB), which is associated with poor treatment outcomes. A better understanding of the risk factors for unfavourable outcomes is needed. Objectives: To describe 1) the demographic and clinical characteristics of MDR-TB patients registered in three states of India during 2009-2011, 2) treatment outcomes, and 3) factors associated with unfavourable outcomes. Design: A retrospective cohort study involving a record review of registered MDR-TB patients. Results: Of 788 patients, 68% were male, 70% were aged 15-44 years, 90% had failed previous anti-tuberculosis treatment or were retreatment smear-positive, 60% had a body mass index < 18.5 kg/m2 and 72% had additional resistance to streptomycin and/or ethambutol. The median time from sputum collection to the start of MDR-TB treatment was 128 days (IQR 103-173). Unfavourable outcomes occurred in 40% of the patients, mostly from death or loss to follow-up. Factors significantly associated with unfavourable outcomes included male sex, age ⩾ 45 years, being underweight and infection with the human immunodeficiency virus. Adverse drug reactions were reported in 24% of patients, with gastrointestinal disturbance, psychiatric morbidity and ototoxicity the most common. Conclusion: Long delays from sputum collection to treatment initiation using conventional methods, along with poor treatment outcomes, suggest the need to scale up rapid diagnostic tests and shorter regimens for MDR-TB.

Contexte : L'Inde a l'un des taux les plus élevés au monde de tuberculose multirésistante (TB-MDR), qui est associée à des résultats médiocres du traitement. Une meilleure compréhension des facteurs de risque de résultats défavorables est requise.Objectifs : Décrire : 1) les caractéristiques démographiques et cliniques des patients TB-MDR enregistrés dans trois états d'Inde de 2009 à 2011, 2) les résultats du traitement, et 3) les facteurs associés à des résultats défavorables.Schéma : Une étude de cohorte rétrospective impliquant une revue des dossiers des patients TB-MDR enregistrés.Résultats : Il y a eu 788 patients, dont 68% d'hommes, 70% âgés de 15­44 ans, 90% ayant eu un échec de leur traitement anti-tuberculose précédent ou ayant un frottis positif en retraitement, 60% ayant un index de masse corporelle < 18,5 kg/m2 et 72% ayant en plus une résistance à la streptomycine et/ou à l'éthambutol. Le délai médian entre le recueil de crachats et la mise en route du traitement de la TB-MDR a été de 128 jours (IQR 103­173). Les résultats ont été défavorables pour 40% des patients, en majorité des décès ou des pertes de vue. Les facteurs significativement associés à un résultat défavorable ont inclus le sexe masculin, l'âge ⩾ 45 ans, la maigreur et le fait d'être positif pour le virus de l'immunodéficience humaine. Des effets secondaires des médicaments ont été notés dans 24% des cas, avec des troubles gastro-intestinaux, des problèmes psychiatriques et une ototoxicité comme symptômes les plus fréquents.Conclusion : De longs délais entre le recueil de crachats et la mise en route du traitement basé sur des méthodes conventionnelles et des résultats médiocres du traitement signalent la nécessité d'intensifier la mise en œuvre des tests de diagnostic rapide et des protocoles de traitement court de la TB-MDR.

Marco de referencia: La tasa de tuberculosis multirresistente (TB-MDR) en la India es una de las tasas más altas en el mundo y se asocia con desenlaces terapéuticos desfavorables. Es preciso lograr un mejor conocimiento de los factores de riesgo que determinan la ineficacia del tratamiento.Objetivos: 1) Describir las características demográficas y clínicas de los pacientes con TB-MDR registrados en tres estados de la India del 2009 al 2011; 2) analizar los desenlaces terapéuticos; y 3) describir los factores asociados con los resultados desfavorables del tratamiento.Método: Un estudio retrospectivo de cohortes a partir del análisis de las historias clínicas de los pacientes registrados con diagnóstico de TB-MDR.Resultados: Se incluyeron en el estudio 788 pacientes; el 68% era de sexo masculino, en el 70% la edad estaba comprendida entre 15 años y 44 años, el 90% tenía antecedente de fracaso de un tratamiento antituberculoso o estaba en retratamiento con baciloscopia positiva, el índice de masa corporal era inferior a 18,5 en el 60% de los casos y el 72% presentaba resistencia adicional a estreptomicina, etambutol o ambos. La mediana del lapso entre la recogida de la muestra de esputo y el comienzo del tratamiento de la TB-MDR fue 128 días (intervalo intercuartil 103­173). Se observaron desenlaces desfavorables en 40% de los pacientes y consistieron en su mayoría en defunciones o pérdidas durante el seguimiento. Los factores que se asociaron de manera significativa con estos desenlaces fueron el sexo masculino, la edad ⩾ 45 años, el bajo peso y la serología positiva frente del virus de la inmunodeficiencia humana. Se notificaron reacciones adversas a los medicamentos en el 24% de los casos, de las cuales las más frecuentes fueron los trastornos gastrointestinales, las afecciones psiquiátricas y la ototoxicidad.Conclusión: La observación de plazos prolongados entre la recogida de las muestras de esputo y la iniciación del tratamiento cuando se utilizan los medios diagnósticos corrientes y de desenlaces terapéuticos desfavorables destaca la necesidad de ampliar la escala de aplicación de las pruebas rápidas de diagnóstico y la administración de pautas más cortas de tratamiento de la TB-MDR.

Evaluation of the diagnostic value of immunocytochemistry and in situ hybridization in the pediatric leprosy.

Leprosy is characterized by a long and variable incubation period and a chronic clinical course. Diagnosis of leprosy is essentially based on clinical features. Although the majority of cases can be diagnosed clinically yet alternative methods for diagnosis are required especially for early cases. Immunocytochemistry and in situ hybridization can be a valuable tool for diagnosis for early cases. The present study is aimed to assess the diagnostic value of immunocytochemistry and in situ hybridization in cytological specimens and to compare these techniques with Z.N. staining. This prospective study was carried out in 26 patients below 18 years of age of leprosy. Clinical examination of each patient was done and categorized according to IAL. After taking consent, three skin smears was taken, one for Z.N. staining and remaining two for immunocytochemistry and in situ hybridization respectively. Routine skin smear examination by Z.N. staining method confirmed the diagnosis in 4/26 (15.83%) and these belonged to BB, BL category. Immunocytochemistry showed positivity in 10/15 (66.6%) in BT and 72.7% in BB/BL leprosy. Immunocytochemistry improved the diagnosis by 53.85%, and the results were statistically significant (p < 0.01). In situ hybridization showed the positive results in 80% cases of BT leprosy and 90.9% cases of BB/BL leprosy. In situ hybridization improved the diagnosis by 70% in comparison to ZN staining and the results were statistically significant (p < 0.01). This study supports that immunocytochemistry and in situ hybridization enhance the diagnosis of leprosy when compared to routine skin smears stained by Z.N staining. They are important diagnostictoolsfor definitive diagnosis in early as well as established cases of leprosy.

Expression of CXCL10 (IP-10) and CXCL11 (I-TAC) chemokines during Mycobacterium tuberculosis infection and immunoprophylaxis with Mycobacterium indicus pranii (Mw) in guinea pig.

Mycobacterium indicus pranii (earlier known as Mycobacterium w) has been used as an immunmodulatory agent in leprosy and tuberculosis by mediating the release of various cytokines and chemokines. CXCL10 (IP-10) and CXCL11 (I-TAC) chemokines are involved in T-cell migration and stimulation of natural killer cells in Mycobacterium tuberculosis infection. In this study, the effect of heat killed M. indicus pranii (alone and in conjunction with chemotherapy) on disease progression was determined by colony forming units (CFUs) in guinea pig lung following their aerosol infection and the expression levels of CXCL10 and CXCL11 were studied by quantitative Reverse Transcriptase Polymerase Chain Reaction (qRT-PCR) and in situ RT-PCR. Four groups of animals included; infection only (Rv), immunoprophylaxis (RvMw), chemotherapy (RvCh) and combination of immunoprophylaxis with chemotherapy (RvChMw). In the group where immunoprophylaxis was given in combination with chemotherapy, the CFU counts reduced significantly at 4th week post-infection as compared to animals that received immunoprophylaxis or chemotherapy alone. At the same time, all groups of animals had elevated expression of CXCL 10 which was significantly high only in animals that received Mw with or without chemotherapy. Unlike to CXCL 10, study demonstrated suppressed expression CXCL 11 in both immunoprophylaxis as well as chemotherapy groups that became up-regulated in synergistic response of immunoprophylaxis and chemotherapy. Taken together, data indicates that the expression of CXCL10 and CXCL11 positively correlates with anti-tubercular treatment (at least with combination of immunoprophylaxis and chemotherapy). Therefore, prior immunization with Mw appears to be a good immunomodulator for release of chemokines and augments the effect of chemotherapy.

Clinical and histopathological evaluation of the effect of addition of immunotherapy with Mw vaccine to standard chemotherapy in borderline leprosy.

This study reports detailed analysis of clinical parameters and clearance of granuloma in borderline leprosy patients treated with immunotherapy and chemotherapy. It aims to assess the additive effect of immunotherapy (Mwvaccine) with standard MDT on clinical status of untreated borderline leprosy cases and on granuloma fraction of untreated borderline leprosy cases. Patients attending the OPD were serially recruited in two groups. A total of 150 cases in one treatment (trial) group (Mw vaccine plus MDT) and 120 cases in another treatment (control) group (MDT only) of border line leprosy have been included. After the formal written consent, detailed clinical examination, charting, smear examination of all untreated borderline patients of both groups was done, biopsies were taken from the active lesions of all patients of both groups at start of therapy and every six month thereafter till the completion of therapy. The same procedure was repeated every six months during the follow-up period. Standard MDT was given to all the patients of both groups according to type of disease. Mw vaccine 0.1 ml (0.5 x 10(9) bacilli) was injected intra-dermally at the start of therapy and every six months in addition to chemotherapy to the treatment group. The BT cases were followed up after 6 doses of MDT and 2 doses of Mw vaccine, and, the BB, BL cases were followed up after 24 doses of MDT plus 5 doses of Mw vaccine. Clinically, greater and faster improvement was observed in all the clinical parameters, faster attainment of smear negativity and two episodes of lepra reaction occurred in cases treated with combined chemotherapy and immunotherapy, as compared to controls (chemotherapy alone) wherein clinical improvement was slower in all parameters, slower attainment of smear negativity in bacillary index and seven showed the occurrence of reactions, histipathologically in addition to more rapid clearance of granuloma in immunotherapy treated group, a significant finding was an increase in the epithelioid cells population in this group. This suggests a possible immunoactivation of the macrophages especially in BB/BL immunotherapy group. Overall comparison of regression induced by chemotherapy alone with that induced by combined chemotherapy and immunotherapy shows a greater reduction in clinical parameters as well as granuloma fraction in BT cases as well as in BB/BL cases. This trial shows the potential usefulness of this approach of addition of immunotherapy to standard chemotherapy in borderline leprosy cases which leads to in faster recovery from disease reduced chances of reactions and faster granuloma clearance. Such information is expected to be useful in improving the immunotherapeutic approaches for treatinggranulomatous conditions in general and in leprosy in particular.

Histological diagnosis of early and suspicious leprosy by in situ PCR.

Leprosy is a chronic mycobacterial disease whose diagnosis is primarily based on clinico-pathological examination and supported by slit skin smears for the presence of acid fast bacilli (AFB). However, definitive diagnosis of early leprosy and those suspected to have the disease but not histologically confirmed pose major public health problems. The present study reports the utility of the in situ Polymerase Chain Reaction amplification (PCR) directed at a 530bp fragment of DNA encoding the 36kd antigen of the causative Mycobacterium leprae for the diagnosis of such patients using skin biopsies of lesions. Twenty five adult patients (aged 15-50yrs) each from the clinical categories of Early and clinically Suspect leprosy were selected for the study after obtaining permission. They had solitary lesions, which were negative for AFB on slit skin smear examination. Routine histopathology confirmed the diagnosis of leprosy in 8/25 (32%) cases in the category of Early leprosy with AFB being seen in 2 biopsies, and in 5/25(20%) cases of Suspect leprosy with AFB being seen in a solitary case. The Direct in situ PCR procedure which was performed in the histologically unconfirmed cases improved the diagnosis with positive results observed in 12/17 (70.6%) cases of Early (p=0.001) and in 12/20 (60%) cases of Suspect Leprosy (p=0.005 indicating the usefulness of the Direct in situ PCR to establish the diagnosis of leprosy in histologically doubtful cases.

Evaluation of diagnostic role of in situ PCR on slit-skin smears in pediatric leprosy.

A large proportion of early cases of leprosy in children remain AFB negative in skin smears. Such cases required additional techniques to confirm the diagnosis. In situ PCR on slit- skin smears is minimally invasive and less cumbersome as compared to skin biopsies. This study was initiated in our institute with the objective to evaluate the diagnostic value of in situ PCR on slit- skin smears in pediatric leprosy. A total of 25 cases of leprosy below 16 years of age were included in the study. After detailed history and thorough clinical examination, informed consent was obtained from the parents of children for slit- skin smears from lesion sites for AFB staining and for in situ PCR technique. Cases were clinically categorized according to IAL classification into indeterminate (I), tuberculoid tuberculoid (TT), borderline tuberculoid (BT), borderline borderline (BB), borderline lepromatous (BL) and lepromatous (LL). Most of the patients (76%) were between 9-16 years of age and 64% of the cases had history of contact with leprosy patients within the family. Skin smears were positive for AFB in only 20% of the cases. On applying in situ PCR, it was observed that 62.5% cases of I/TT/BT/BB category and 88.8% of BL/LL category gave positive signals. Overall in situ PCR confirmed the diagnosis in 72% cases while by slit smears diagnosis was confirmed in only 20% of cases. Further, out of 20 skin smear negative cases, 13 were positive by in situ PCR. Specificity of the signals of in situ PCR was established by demonstrating the absence of signals in nonleprosy dermatological conditions of vitiligo and P.alba. This study supports the potential usefulness of in situ PCR on slit- skin smears of early pediatric leprosy cases. This strategy will be especially useful in cases where skin smears are negative and in those cases where skin biopsy can not be done either because of unusual locations of lesions or because of sensitive age of the patients.

KIT ligand and bone morphogenetic protein signaling enhances human embryonic stem cell to germ-like cell differentiation.

BACKGROUND: Signaling mechanisms involved in early human germ cell development are largely unknown and believed to be similar to mouse germ cell development; however, there may be species specific differences. KIT ligand (KITL) and Bone morphogenetic protein 4 (BMP4) are necessary in mouse germ cell development and may play an important role in human germ cell development. METHODS: KITL signaling studies were conducted by differentiating human embryonic stem cells (hESCs) on KITL wild-type, hetero- or homozygous knockout feeders for 10 days, and the effects of BMP signaling was determined by differentiation in the presence of BMP4 or its antagonist, Noggin. The formation of germ-like cells was ascertained by immunocytochemistry, flow cytometry and quantitative RT-PCR for germ cell markers. RESULTS: The loss of KITL in enrichment and differentiation cultures resulted in significant down-regulation of germ cell genes and a 70.5% decrease in germ-like (DDX4+ POU5F1+) cells, indicating that KITL is involved in human germ cell development. Moreover, endogenous BMP signaling caused germ-like (DDX4+ POU5F1+) cell differentiation, and the inhibition of this pathway caused a significant decrease in germ cell gene expression and in the number of DDX4+ POU5F1+ cells. Further, we demonstrated that eliminating feeders but maintaining their secreted extracellular matrix is sufficient to sustain the increased numbers of DDX4+ POU5F1+ cells in culture. However, this resulted in decreased germ cell gene expression. CONCLUSIONS: From these studies, we establish that KITL and BMP4 germ cell signaling affects in vitro formation of hESC derived germ-like cells and we suggest that they may play an important role in normal human germ cell development.

Comparative growth pattern of multi drug resistance versus susceptible isolates of Mycobacterium tuberculosis in mice lungs.

BACKGROUND & OBJECTIVE: Rise in prevalence of multi-drug resistance (MDR) in tubercle bacilli is a serious cause of concern. As mutations with two house keeping genes rpoB and katG are associated with resistance to two important anti-tubercular drugs rifampicin and isoniazid respectively, there is a need to understand the growth kinetics of organisms with such mutated genes in experimental animals. This study was undertaken to study the growth kinetics of susceptible as well multi-drug resistance Mycobacterium tuberculosis isolates in mice. METHODS: Two MDR (having mutations in rpoB and catG) and two drug susceptible isolates of M. tuberculosis along with H37Rv were grown in mice after aerogenic infection. RESULTS: The MDR isolates grew slowly up to 3 wk though the growth was significantly different from sensitive strains. However, after 3 wk, the growth in sensitive as well MDR strains was similar, suggesting that even the mutations in the MDR strains did not have any impact on the growth kinetics. INTERPRETATION & CONCLUSION: The effect of mutations in other parts of these genes need to be studied. Retention of property of MDR strains to establish infection after aerogenic infection has epidemiological significance in terms of the transmission of MDR tuberculosis.

Immunogenicity and protective efficacy of "Mycobacterium w" against Mycobacterium tuberculosis in mice immunized with live versus heat-killed M. w by the aerosol or parenteral route.

As the disease caused by Mycobacterium tuberculosis continues to be a burden, there is a concerted effort to find new vaccines to combat this problem. One of the important vaccine strategies is whole bacterial vaccines. This approach relies on multiple antigens and built-in adjuvanticity. Other mycobacterial strains which share cross-reactive antigens with M. tuberculosis have been considered as alternatives to M. bovis for vaccine use. One such strain, "Mycobacterium w", had been evaluated for its immunomodulatory properties in leprosy. A vaccine against leprosy based on killed M. w is approved for human use, where it has resulted in clinical improvement, accelerated bacterial clearance, and increased immune responses to Mycobacterium leprae antigens. M. w shares antigens not only with M. leprae but also with M. tuberculosis, and initial studies have shown that vaccination with killed M. w induces protection against tuberculosis in Mycobacterium bovis BCG responder, as well as BCG nonresponder, strains of mice. Hence, we further studied the protective potential of M. w and the underlying immune responses in the mouse model of tuberculosis. We analyzed the protective efficacy of M. w immunization in both live and killed forms through the parenteral route and by aerosol immunization, compared with that of BCG. Our findings provide evidence that M. w has potential protective efficacy against M. tuberculosis. M. w activates macrophage activity, as well as lymphocytes. M. w immunization by both the parenteral route and aerosol administration gives higher protection than BCG given by the parenteral route in the mouse model of tuberculosis.

Two microbiological relapses in a patient with lepromatous leprosy.

A lepromatous patient treated with dapsone in the pre-MDT era to the point of smear negativity (> 6 years), relapsed 5 years after stopping treatment. He was then put on WHO-MDT for multibacillary (MB) leprosy, and was treated again; he had negative slit skin smears (3 years). He again presented with a relapse of leprosy 17 years after stopping treatment, and this time he presented with borderline leprosy in reaction.

Changing profile of disease in leprosy patients diagnosed in a tertiary care centre during years 1995-2000.

A hospital based retrospective study was carried out to determine change in the profile of disease in leprosy patients taking 1995 as baseline and compared with the profile seen in year 2000. A total of 2149 and 1703 cases were studied respectively of year 1995 and 2000. Male to female ratio slightly increased from 2.95:1 in year 1995 to 3.4:1 in year 2000. Majority of patients were of borderline type in both years. Proportion of cases with MB leprosy was nearly same in females (60.8%) and males (63.1%) in year 1995 and in year 2000 (64.8% females and 67.6% males). Proportion of highly bacillary cases has decreased over the years in females (from 20.95% in 1995 to 11.7% in year 2000, p=0.03) as well as in males (from 25% in 1995 to 15.5% in year 2000, p=0.001). Incidence of total reactions increased from 27.6% to 35.4% over the years which is significant (p<0.01). Proportion of type 1 reactions were more in reproductive age group in females in both years (p<0.05) and of type 2 reactions were significantly (p > or = 0.05) more in males in both years. Incidence of disability (both grade 1 and grade 2) was significantly more in males than in females in both years (p > or = 0.04). Grade 1 disability has significantly increased over years in females from 10.11% to 14.8%(p<0.03) as well as in males from 13.27% to 21.3%(p<0.001). Onset of reactions was associated with pregnancy/lactation in 62% of cases and with menopause in 21% of cases in 2000, which suggests strong correlation with hormonal imbalance. To conclude while leprosy incidence has declined after MDT, recognition and management of reactions in women around changes in their hormonal levels should be properly monitored for early and effective management.

PCR and in-situ hybridization for diagnosis of leprosy.

OBJECTIVE: To assess the diagnostic value of Polymerase Chain Reaction (PCR) and in situ hybridization. METHODS: This prospective study was carried out in 22 patients RESULTS: The histopathological examination confirmed the clinical diagnosis in 27.2% cases only. In situ hybridization showed a positivity of 42.8% in early (I/BT) and 46.7% in BB/BL group. In situ hybridization thus enhanced the diagnosis by 18.1%. PCR targeting 36 kDa gene of M. leprae was performed on 15 cases. In these 15 cases, histopathology confirmed the diagnosis in 4 cases (26.6%) and PCR confirmed the diagnosis in 10 cases (66.6%), thus enhancing the diagnosis by 40%. CONCLUSION: 36 kDa PCR and in situ hybridization enhance the diagnosis of leprosy when compared to routine histopathology. They are important diagnostic tools for definitive diagnosis in early and doubtful cases of leprosy.

Diagnostic value of in situ Polymerase Chain Reaction in leprosy.

OBJECTIVE: This prospective study was carried out to assess the diagnostic value of in situ Polymerase Chain Reaction in leprosy, particularly in enhancing the histopathological diagnosis. METHOD: Clinical examination of 20 patients (< 16 yr) was done and skin smear for AFB was prepared. Biopsy of lesion site was taken for histopathological examination and in situ PCR testing. RESULTS: The histopathological examination confirmed the clinical diagnosis in 45% cases only; non-specific histopathology was reported in the remaining 55% cases. In situ PCR showed a positivity of 57.1% in early/localized form of leprosy (IIBT) and 61.5% in (BB/BL) group. When compared to histopathology examination, a significant enhancement of 15% in diagnosis was seen. With in situ PCR, the diagnosis could be confirmed in 4/11 (36.3%) cases with non-specific histopathological features, (which is common in early disease) in addition to confirmation of 8/9 (88.8%) histopathologically-confirmed tissue sections. Histopathology and in situ PCR, combined together, confirmed the diagnosis in 13/20 cases (65% of total cases). CONCLUSION: Thus, in situ PCR is an important diagnostic tool especially in early and doubtful cases of leprosy.

Persister studies in leprosy patients after multi-drug treatment.

Cutaneous biopsies were collected from leprosy patients who attended the out-patient department of the Institute for treatment at different intervals, i.e., 12 months, 18 months, 24 months, 36 months, and more after beginning the multi-drug treatment therapy (M.D.T.). The patients belonged to the two drug regimens; (i) standard multibacillary (MB) M.D.T. after 12, 24, and 36 months; or (ii) standard M.D.T. + Minocycline 100 mg once a month (supervised) + Ofloxacin 400 mg once a month supervised for 12 months Biopsies were processed for mouse footpad inoculation and for estimating ATP levels by bioluminescence assay as per established methods. Viable bacilli were observed in 23.5% up to 1 year, 7.1% at 2 years, and in 3.84% at 3 years of M.D.T. by MFP and 29.4%, 10.7%, and 3.84% by ATP assay in the M.D.T. group at the same time period, respectively, but not in M.D.T. + Minocycline + Ofloxacin group after one year. The overall percentage of persisters was 5.55% by MFP and 7.14% by ATP assay up to 3 years of treatment.

Patients presenting with defined areas of sensory loss--a preliminary study.

Thirty patients presenting with circumscribed areas of clearly demonstrable hypoesthesia were chosen from amongst those attending this Institute. Their history and clinical features were recorded, lepromin test was done for reading at four weeks, and peripheral part of the hypoesthetic area was biopsied for histopathology and immunostaining. The subjects were predominantly adult males with the symptomatic sites limited to the extremities. On routine histopathological examination of the symptomatic sites, the diagnosis of leprosy, using defined criteria, could be made in six cases (20%). Immunostaining of the remaining sections showing either no pathology or a nonspecific pathology revealed the presence of mycobacterial antigen in five of the 24 cases (20.83%). Overall, leprosy could be diagnosed in 11 of the 30 cases studied (36.66%). This study shows that leprosy may be an important cause of circumscribed areas of sensory deficit.

Long-term follow up of multibacillary leprosy patients with high BI treated with WHO/MDT regimen for a fixed duration of two years.

Forty-six, newly detected, previously untreated multibacillary (MB) patients with a bacterial index (BI) of > or = 3+ who had received WHO/MDT for 2 years were followed up for a total duration of 424 person-years and a mean duration of 9.26 +/- 2.98 years per patient. The BIs of the patients continued to fall, and all of the patients, except one, reached skin-smear negativity. WHO/MDT was well accepted and well tolerated. Relapse, which was defined as an increase in the BI of 1+ or more with or without clinical evidence of activity, was observed in only one patient, giving a relapse rate of 2.2% or 0.23 per 100 person-years in patients with a BI of > or = 3+ after long-term follow up. This patient was started on a second course of WHO/MDT to which he responded favorably. WHO/MDT for a fixed duration of 2 years for MB patients as recommended by the WHO is vindicated.