Magnitude of unreported kala-azar cases in a highly endemic district of Bihar, India: A positive impact of Indian elimination programme.

BACKGROUND & OBJECTIVES: In India, kala-azar surveillance is weak and no public-private partnership exists for disease containment. Estimate of disease burden is not reliably available and still cases are going to private providers for the treatment. The present study aimed to assess the magnitude of kala-azar cases actually detected and managed at private set-up and unreported to existing health management information system. METHODS: Institution based cross-sectional prospective pilot study was conducted. List of facilities was created with the help of key informants. The information about incidence of kala-azar cases were captured on monthly basis from July 2010 to June 2011. Rapid diagnostic strip test (rk-39) or bone marrow/splenic puncture were applied as laboratory methods for the diagnosis of kala-azar. Descriptive statistics as well as chi-square test for comparison between proportions was conducted. RESULTS: Overall availability of private practitioners (PPs) was 4.59/1,00,000 population and maximum PPs (46; 93.9%) were from qualified category. The median years of medical practice was 25 yr (inter quartile-range [18, 28]). Interestingly, only a small proportion (240; 19%) of cases was managed by PPs. Amongst the PPs, only low proportion (32; 18.2%) managed >2 cases per month. The mean number of kala-azar suspects and cases identified varied significantly between different PPs' professions with p <0.048 and p <0.032, respectively. A highly significant difference (p <0.0001) was observed for kala-azar case load between qualified and unqualified practitioners. A small proportion (38; 15.8%) of kala-azar cases was not present in the public health system record. INTERPRETATION & CONCLUSION: Still sizeable proportions of cases are going to PPs and unrecorded into government surveillance system. A mechanism need to be devised to involve at least qualified PPs in order to reduce treatment delay and increase case detection in the region.

Hypertriglyceridemia: a possible diagnostic marker of disease severity in visceral leishmaniasis.

PURPOSE: Visceral leishmaniasis (VL), a protozoan disease, is 100% fatal if left untreated. Anemia is common in VL which plays a role in expression of clinically overt VL disease. Laboratory clues are scarce for strengthening clinical suspicion for severity in VL. Hypertriglyceridemia has emerged as a new concept for the diagnosis and prognosis in VL. The present study is aimed at correlating the magnitude of hypertriglyceridemia with the severity in VL. MATERIALS AND METHODS: A retrospective case-control study was conducted between January 2012 to December 2013 among 124 patients coming for treatment from VL endemic areas, who had fever of more than 15 days and did not respond to antimalarials and antibiotics. The parasitologically confirmed VL cases (n = 87) were categorized as mild/moderate (n = 60) and severe (n = 27) groups according to WHO classification for anemia and parasite burden. Serum triglycerides were assayed in VL groups along with controls (n = 37). RESULTS: Serum triglyceride level was significantly higher in VL than controls [mean values were 173.50 ± 47.67 versus 127.1 ± 53.79 mg/dl, respectively (p < 0.0001)]. Triglyceride level was significantly higher in severe than in mild/moderate group of VL [211.3 ± 50.2 mg/dl versus 134 ± 45.09 mg/dl, respectively (p < 0.0001)]. Hypertriglyceridemia (>161.7 mg/dl) was noted in all severe VL patients, compared to 31.66% of mild or moderate group (p < 0.0001). There was no significant difference between mild/moderate VL and controls. CONCLUSIONS: It is hypothesized that hypertriglyceridemia could be of additional diagnostic benefit to assess the probability and severity of VL in endemic areas.

Development of post-kala-azar dermal leishmaniasis in AmBisome treated visceral leishmaniasis: a possible challenge to elimination program in India.

We report two cases, one male (33 years) and a female (14 years), that developed Post-Kala-azar Dermal Leishmaniasis (PKDL) after successful treatment for visceral leishmaniasis (VL) or Kala-azar with AmBisome, the lipid complex of Amphotericin B. Both cases presented with hypo-pigmented macular lesions all over the body. The patients responded well to AmBisome after treatment with three courses. This first ever case report from India indicates that possibly there is no effective drug for VL until date, which can prevent post-treatment development of PKDL.

Fatal acute pancreatitis in a patient with visceral leishmaniasis during miltefosine treatment.

Pancreatitis is a known side effect of the once commonly used drug, sodium stibogluconate, for treatment of visceral leishmaniasis (VL). In India, miltefosine has recently been introduced as the first-line drug. Its side effects include loose motions, vomiting, and teratogenicity. We report here a case of a 41-year-old parasitologically confirmed male case of VL, who developed acute pancreatitis during treatment with miltefosine. On the 13 th day of treatment, he presented with abdominal pain and vomiting. The biochemical, hematological, and radiological features were suggestive of acute pancreatitis. The patient was put on conservative treatment for pancreatitis at the specialized center but succumbed to renal failure and septicaemia.

Post-kala-azar dermal leishmaniasis in a patient treated with injectable paromomycin for visceral leishmaniasis in India.

Post kala-azar dermal leishmaniasis (PKDL) is a skin manifestation that usually develops after treatment of visceral leishmaniasis (VL), a major public health problem in India. The diagnosis and management of PKDL is complex. This is the first case report from India in which PKDL occurred after paromomycin treatment for VL in an Indian patient.

Post-kala-azar dermal leishmaniasis (PKDL), HIV and pulmonary tuberculosis.

Post-kala-azar dermal leishmaniasis is usually a sequel to visceral leishmaniasis. A 25-year-old woman presented with hypopigmented maculopapular lesions all over the body for the past 4 years without any previous history of visceral leishmaniasis. She was on treatment for leprosy and pulmonary tuberculosis for the past 2 months, but did not show any improvement. Investigations confirmed that she had post-kala-azar dermal leishmaniasis associated with pulmonary tuberculosis and HIV-1 infection. She was started on treatment for the triad of diseases, and showed improvement.

Designing therapies against experimental visceral leishmaniasis by modulating the membrane fluidity of antigen-presenting cells.

The membrane fluidity of antigen-presenting cells (APCs) has a significant bearing on T-cell-stimulating ability and is dependent on the cholesterol content of the membrane. The relationship, if any, between membrane fluidity and defective cell-mediated immunity in visceral leishmaniasis has been investigated. Systemic administration of cholesterol by liposome delivery (cholesterol liposomes) in Leishmania donovani-infected hamsters was found to cure the infection. Splenic macrophages as a prototype of APCs in infected hamsters had decreased membrane cholesterol and an inability to drive T cells, which was corrected by cholesterol liposome treatment. The effect was cholesterol specific because liposomes made up of the analogue 4-cholesten-3-one provided almost no protection. Infection led to increases in interleukin-10 (IL-10), transforming growth factor beta, and IL-4 signals and concomitant decreases in gamma interferon (IFN-gamma), tumor necrosis factor alpha, and inducible NO synthase signals, which reverted upon cholesterol liposome treatment. The antileishmanial T-cell repertoire, whose expansion appeared to be associated with protection, was presumably type Th1, as shown by enhanced IFN-gamma signals and the predominance of the immunoglobulin G2 isotype. The protected group produced significantly more reactive oxygen species and NO than the infected groups, which culminated in killing of L. donovani parasites. Therefore, cholesterol liposome treatment may be yet another simple strategy to enhance the cell-mediated immune response to L. donovani infection. To our knowledge, this is the first report on the therapeutic effect of cholesterol liposomes in any form of the disease.

Clinical and laboratory comparison of different brands of amphotericin B used for the treatment of Kala-azar: an observational study.

The communication presents clinical response of cases of visceral leishmaniasis to treatment by two different brands of Amphotericin B. Fungizone was found to be slightly better than Amphotericin B, however, the difference is not statistically significant.

Wilson disease with visceral leishmaniasis: an extremely uncommon presentation.

Visceral leishmaniasis (VL), which is caused by the protozoa Leishmania donovani and transmitted by the bite of the female sand fly Phlebotomus argentipes, is common in Bihar, India. Wilson disease is an autosomal recessive disorder of copper metabolism in which copper is deposited in the brain and liver. We report a case of an extremely uncommon combination of these diseases in a patient. Treatment options for such a combination of diseases are limited and difficult.

Visceral leishmaniasis and tuberculosis in patients with HIV co-infection.

We describe here two cases, one male and one female, both age 40 years, with visceral leishmaniasis and HIV-1 co-infection. The female patient had features of Koch's abdomen. The male patient had features of tuberculous lymphadenitis and bilateral pleural effusion more marked on the right side. Both were treated with highly active antiretroviral therapy, antituberculous drugs, antibiotics, antifungal medicine (fluconazole) and miltefosine. Both patients showed marked improvement with therapy.

Reversal of T-cell unresponsiveness through serine-esterase inhibitors mediated enhanced lymphokine induced microbicidal activities in kala-azar.

After presenting processed glycoprotein of Leishmania donovani to T-cell, macrophage seeks the help of a panel of T-cells lymphokines to transform from a state that sustains intra cellular replication of parasite to an effector state for destructing parasites. But esterase and trypsin of macrophage membrane prevent T-cells to release MIF. Role of soya-bean trypsin inhibitor (STI) has been exposed in the present study with a view to alter esterase functional behaviour of macrophage for control of T-cell activation and also, if T-cells once made responsive to antigen by STI do alter macrophage response to T-cells or not. Results establish STI as potent effector molecule, which can serve as an adjuvant to candidate T-cell epitope and synthetic peptide for development of anti-Kala-azar vaccine protocol in future.

A randomized clinical trial of low dosage combination of pentamidine and allopurinol in the treatment of antimony unresponsive cases of visceral leishmaniasis.

OBJECTIVES: A randomized clinical trial of low dosage combination of pentamidine and allopurinol was carried out with objectives to assess the efficacy and toxicity as compared to full dosage of pentamidine in antimony unresponsive visceral leishmaniasis (VL) patients. METHODS: Using a randomized control clinical trial, a total of 158 antimony unresponsive patients of VL were randomly allocated into two treatment groups. Patients in one group (n=80) received half the dosage of pentamidine i.e. 2 mg/kg body weight by IM route on alternate day and allopurinol in dose of 15 mg/kg body weight in three divided dosages for 30 days; patients in the second group (n=78) received pentamidine in dose of 4 mg/kg body weight by IM route on alternate day for 15 injections in 30 days. The efficacy and safety of the two regimens were compared. RESULTS: Apparent cure i.e. clinical and pathological cure at the end of therapy, in 78 (97.5%) and 67 (86%), and ultimate cure i.e. clinical and parasitological cure at the end of follow-up of six months, in 73 (91.25%) and 58 (74.35%) patients was observed in the combination regimen and single regimen group respectively. The difference of the ultimate cure between two groups of the patients was statistically significant (p < 0.01). In single regimen group, 11 (14%) patients showed primary unresponsiveness (with no response during treatment) and nine (13%) relapse (after six months of follow-up) respectively, where as in combination regimen group, two (2.5%) patients showed primary unresponsiveness and five (6.4%) relapse respectively. By the end of the treatment, the incidence of injection-related toxicity, such as rigor and fever, was same in both groups. No hyperglycemia was observed in combination therapy probably due to reduced dose of pentamidine and three patients in single regimen developed hyperglycemia and one of them developed irreversible hyperglycemia. CONCLUSIONS: The study showed that the combination of pentamidine (half dose) and allopurinol is more effective in achieving ultimate cure with an added advantage of reduced toxicity in unresponsive cases as compared to full pentamidine dose.

Effect of immunization with lipid associated polysaccharide antigen and anti CD-2 antibodies on class II MHC expression and cellular immune response in BALB/C mice infected with Leishmania donovani.

In a bid to characterize the antigens and immunization mechanisms which may be used to produce a protective response against L. donovani, role of lipid associated polysaccharide (LPS) antigen and whole antigen was evaluated. BALB/C mice were immunized with whole or LPS antigen in combination with one of three putative adjuvents (anti CD-2 antibody/FIA/0.85% Saline). LPS antigen emulsified in anti CD-2 antibody was found to induce significant antibodies in mice on day 28 against challenge with lethal dose of L. donovani. Immunoprophylactic properties of LPS and whole antigen was investigated on day 40 through cytokine elicitation (IL-2), MIF) in culture supernatants of spleen cells, but before that MHC-II expressed on macrophage was studied. The LPS antigen in combination with anti CD-2 antibody was found to be most immuno-reactive inducing higher MHC-II expression on macrophages which was associated with substantial rise in the level of MIF and IL-2. It coincided with decline in antibody titre in 100% mice immunized with LPS antigen while Leishmania injected as whole antigen failed to induce specific macrophage and T-cell response with all the above formulations. We surmise from our data that lipid associated polysaccharide antigen linked to anti CD-2 antibody has potential for eliciting protective immunity against Leishmania.

A community-based, comparative evaluation of direct agglutination and rK39 strip tests in the early detection of subclinical Leishmania donovani infection.

In the Indian state of Bihar, the sensitivities and specificities of direct agglutination tests (DAT) and rK39 test strips for the detection of Leishmania donovani infection in humans were explored and found to be generally good (92%-100%). When 172 asymptomatic individuals [16 'case-contacts' who lived in the same households as past or current, confirmed cases of visceral leishmaniasis (VL) and 156 other subjects from neighbouring households] were tested, the same 36 (21%) individuals, including all 16 'case-contacts', were found seropositive using each type of test. When followed-up after 3 months, 18 of the individuals who had been found seropositive in the baseline survey remained seropositive, and eight (44%) of these had developed symptomatic VL, with amastigotes in their splenic aspirates. Seven (44%) of the 16 'case-contacts' but only one (5%) of the other 20 subjects found seropositive at baseline went on to develop VL within 3 months. Although the strip test appeared slightly better than DAT for predicting the development of VL in the 172 subjects, either type of test may be very useful for the early detection of asymptomatic L. donovani infection and thus the identification of those at relatively high risk of developing VL.

Milk of cow (Bos taurus), buffalo (Bubalus bubalis), and goat (Capra hircus): a better alternative than fetal bovine serum in media for primary isolation, in vitro cultivation, and maintenance of Leishmania donovani promastigotes.

Tyndalized milk of goat, cow, and buffalo was found to be a potential substitute for fetal bovine serum (FBS) in the medium for the cultivation of Leishmania donovani promastigotes. The numbers (means) of promastigotes reached 2.6 x 10(7), 2.3 x 10(7), and 2.1 x 10(7)/ml, respectively, in the medium supplemented with 10% milk of goat, cow, and buffalo, in comparison to 1.9 x 10(7)/ml in the control with 10% FBS. In primary isolation, the milk-supplemented medium showed that 22 out of 26 samples were positive for promastigotes (84.6%) and the cells were maintained successfully during the observed period of 6 months.

Visceral leishmaniasis (kala-azar)--the Bihar (India) perspective.

From a hospital-based surveillance carried out in Rajendra Memorial Research Institute of Medical Sciences, Patna, Bihar, India, the socio-economic, demographic and treatment response information of 737 patients admitted with visceral leishmaniasis (VL) during January 2001-December 2003, were analysed. The disease was two times higher in males than in females because of several factors including clothing pattern, sleeping habits and occupation. In Bihar, the second poorest state in India, poverty plays a major role in perpetuation of the disease, contributing to malnutrition, illiteracy (60%), and poor housing (82%). Further, presences of peri-domestic animal shelters around houses (63%) and vegetations (77%) facilitate breeding of sand fly vector. Clinical and laboratory characteristics were similar in the age groups <12 years and >12 years. The increasing unresponsiveness of VL patients to conventional anti-leishmanial drugs, e.g. sodium antimony gluconate (SAG) and pentamidine, has definitely posed a major therapeutic challenge in combating the disease. Amphotericin B, though costly, is highly effective. Miltefosine is a highly promising new oral drug for VL.

Evaluation of cholinesterase level in an endemic population exposed to malathion suspension formulation as a vector control measure.

The manuscript describes a study on the blood cholinesterase (ChE) level in an exposed population at different interval of time after spraying with malathion suspension (SRES) use for kala-azar vector control in an endemic area of Bihar, India. The toxicity of a 5% malathion formulation in the form of a slow release emulsified suspension (SRES) was assessed by measuring serum ChE levels in spraymen and in the exposed population. The study showed a significant decrease in ChE levels in the spraymen (p < 0.01) after one week of spraying and in exposed population one week and one month after of spraying (p < 0.01), but was still within the normal range of ChE concentration, one year after spraying, the ChE concentration in the exposed population was the same as prior to spraying (p > 0.01). On no occasion was the decrease in ChE level alarming. A parallel examination of the clinical status also showed the absence of any over toxicity or any behavioural changes in the exposed population. Hence, it may be concluded that 5% malathion slow release formulation, SRES, is a safe insecticide for use as a vector control measure in endemic areas of kala-azar in Bihar, India so long as good personal protection for spraymen is provided to minimize absorption and it can substitute the presently used traditional DDT spray.