Classical Visceral Leishmaniasis Research Articles

The Severity of Visceral Leishmaniasis Correlates with Elevated Levels of Serum IL-6, IL-27 and sCD14.

BackgroundVisceral leishmaniasis (VL) is a severe disease caused by infection with protozoa of the genus Leishmania. Classic VL is characterized by a systemic infection of phagocytic cells and an intense activation of the inflammatory response. It is unclear why 90% of infected individuals do not develop the disease while a minority develop the classical form. Furthermore, among those that develop disease, a small group progresses to more severe form that is unresponsive to treatment. The presence of inflammatory mediators in serum could theoretically help to control the infection. However, there is also a release of anti-inflammatory mediators that could interfere with the control of parasite multiplication. In this study, we took advantage of the spectrum of outcomes to test the hypothesis that the immune profile of individuals infected with Leishmania (L.) infantum is associated with the development and severity of disease.Methodology/Principal FindingsSera from patients with confirmed diagnosis of VL were evaluated for the presence of numerous molecules, and levels compared with healthy control and asymptomatic infected individuals.Conclusions/Principal FindingsAlthough differences were not observed in LPS levels, higher levels of sCD14 were detected in VL patients. Our data suggest that L. infantum may activate the inflammatory response via CD14, stimulating a generalized inflammatory response with production of several cytokines and soluble molecules, including IFN-γ, IL-27, IL-10, IL-6 and sCD14. These molecules were strongly associated with hepatosplenomegaly, neutropenia and thrombocytopenia. We also observed that IL-6 levels greater than 200 pg/ml were strongly associated with death. Together our data reinforce the close relationship of IFN-γ, IL-10, IL-6, TNF-α and IL-27 in the immune dynamics of VL and suggest the direct participation of sCD14 in the activation of the immune response against L. infantum.

Infection with Leishmania (Leishmania) infantum of 0 to 18-Month-old children living in a visceral leishmaniasis-endemic area in Brazil.

The diagnosis of Leishmania (Leishmania) infantum infection in children from birth may serve as a reference for the early identification of cases that would progress to classical visceral leishmaniasis (VL) in endemic areas. This study prospectively evaluated newborns of mothers living in the municipality of Paracatu, Minas Gerais, Brazil. The infants were followed up at 6-month intervals by clinical examination, serological tests (immunofluorescence [IIF] and enzyme-linked immunosorbent assay with rK39 [ELISA-rK39]) and polymerase chain reaction (PCR) until they had completed 18 months of age. A total of 166 pregnant women were included to evaluate the possible transfer of antibodies or even congenital transmission. Twenty-two of the women tested positive by IIF, four by ELISA-rK39, and one by PCR. Three infants of the 25 women with some positive test results were also positive in the first test (one by IIF, one by ELISA-rK39, and the third by ELISA-rK39 and PCR). One hundred and sixty infants were included in the study; of these, 43 had at least one positive sample over time. However, agreement between tests was low. Follow-up of children with a positive result in the tests studied revealed no progression to classical disease within a period of 18 months. In contrast, two children with negative IIF, PCR, and ELISA-rK39 results developed classical VL at 9 and 12 months of age. In conclusion, a positive test result was variable and sometimes temporary and agreement between tests was low. Therefore, the early diagnosis of Leishmania infection was not associated with the early identification of cases that would progress to classical VL in the endemic area studied.

Anti-Leishmania infantum IgG Antibody Avidity in Visceral Leishmaniasis

IgG avidity tests are used to discriminate acute from chronic infections. There are few reports on the IgG avidity profile of patients with visceral leishmaniasis (VL). This study investigated the anti-Leishmania IgG avidity in patients with classic VL (n = 10), patients showing clinical cure after treatment (n = 18), and asymptomatic subjects with at least one positive Leishmania test (n = 20). All subjects were from areas in Brazil where VL is endemic. Serum samples were collected from each subject on two different occasions. IgG avidity was evaluated by Western blotting. The proportion of high-avidity antibodies was higher in all samples from patients with classic VL. In contrast, low-avidity antibodies predominated in subjects with a history of VL, including 13 cases (72.2%) in the first assessment and 14 (77.8%) in the second. Fifteen (75%) of the asymptomatic subjects presented a predominance of low-avidity antibodies in the first assessment, and the frequency of high-avidity antibodies increased over time in seven subjects (35%) of this group. Antibodies against the 14- and/or 16-kDa antigen fraction were detected in the first assessment in all patients with classic VL, in 10 (55.5%) treated patients, and in 10 (50%) asymptomatic subjects. These were high-avidity antibodies in most cases. In the asymptomatic group, an increase in IgG avidity against the 14- and/or 16-kDa antigen fraction was observed in three cases (15%). The results indicate distinct responses in infected and asymptomatic subjects, probably associated with the length of time after infection. In this respect, IgG avidity tests represent a new approach to better characterize asymptomatic VL.

Comparative Study of Serologic Tests for the Diagnosis of Asymptomatic Visceral Leishmaniasis in an Endemic Area

Serologic tests have been widely used for the diagnosis of asymptomatic visceral leishmaniasis. This study evaluated five serologic tests used for the diagnosis of asymptomatic infection: enzyme-linked immunosorbent assay (ELISA) using promastigote antigen (ELISAp), ELISA using recombinant K39 (ELISA rK39), and K26 (ELISA rK26) antigens, an indirect immunofluorescence test using Leishmania (Leishmania) amazonensis promastigote antigen (IIFT), and an immunochromatographic test using rK39 antigen (TRALd). As a reference regarding the performance of the tests, patients with classic visceral leishmaniasis originating from Minas Gerais, Brazil (N = 36), were defined as the positive group and samples of healthy individuals from nonendemic areas (Argentina) (N = 127) were used as negative controls. Patients with other diseases such as cutaneous leishmaniasis (N = 53) and malaria (N = 56) were also studied to evaluate the chance of cross-reactivity in these tests. Finally, subjects from an area endemic for visceral leishmaniasis in Brazil (Porteirinha, northern Minas Gerais) (N = 1241) were screened for asymptomatic infection with Leishmania and Chagas disease. The sensitivity of the serologic tests was 50% (18/36), 66.7% (24/36), 69.4% (25/36), 83.3% (30/36), and 88.9% (32/36) for ELISAp, ELISA rK26, ELISA rK39, IIFT, and TRALd, respectively. Specificity, calculated using the truly negative group, was 96% (122/127) for TRALd, 97.6% (124/127) for ELISAp and IIFT, and 100% (127/127) for ELISA rK39 and rK26. Positivity in at least one test employing recombinant antigen was observed in 24 (45%) patients with cutaneous leishmaniasis and 47 (82.4%) with malaria. In the visceral leishmaniasis-endemic area, the positivity of the serologic tests ranged from 3.9% to 37.5%. The enzyme-linked immunosorbent assay (ELISA) tests using recombinant antigens were more frequently positive in subjects with a history of exposure to human or canine visceral leishmaniasis (ELISArK39: 14.6% [149/1017] versus 37.5% [84/224]; ELISA rK26: 12.7% [129/1017] versus 21.4% [48/224], P < 0.001 for both). Kappa agreement was low, with a maximum value of 0.449 between ELISAp and IIFT. In addition, among the 112 IIFT-positive subjects, 75 (67%) also presented positive serology for Chagas disease. In conclusion, IIFT and TRALd presented the best performance to diagnose classic cases of visceral leishmaniasis in an endemic area. Cross-reactivity of the tests with Chagas disease, cutaneous leishmaniasis, and malaria should be taken into account. However, the differences in the positivity of the tests used, together with the low agreement between results, do not permit to select the best test for the diagnosis of asymptomatic Leishmania infection.

Estudo comparativo de anticorpos IgG e IgE antileishmania como marcadores de infecção e doença em indivíduos de área endêmica de leishmaniose visceral, em São Luis, MA

Comparative study regarding IgG and IgE anti-leishmania as markers of infection and illness among residents of a visceral leishmaniasis visceral leishmaniasis endemic area, São Luis, MA in the period from May 1999 to May 2000. All the 1,016 individuals younger than 16 years old were tested for the presence of IgG by ELISA. A total of 174 (17.1%) children revealed a positive IgG test and 4 children showed symptoms of classical visceral leishmaniasis during the time of the survey: 85 IgE anti-leishmania ELISA tests with positivity of 43.5% were realized. In this sample, all 7 children with past-visceral leishmaniasis in the sample were IgG positive and 4 (57.1%) were IgE positive, even after a 7 year post treatment period. Three children with current visceral leishmaniasis were evaluated, and all of them were positive for both tests. The detection of antileishman IgE antibodies presented as a good marker for infection by Leishmania chagasi in endemic areas but not as a disease marker.

Sudanese mucosal leishmaniasis: isolation of a parasite within the Leishmania donovani complex that differs genotypically from L. donovani causing classical visceral leishmaniasis

Mucosal leishmaniasis, which is a sporadic disease in the Sudan, was shown by isoenzyme characterization and PCR to be caused by Leishmania donovani. However, it was not clear if the parasite was exactly the same strain as that causing visceral leishmaniasis (VL), or of a different strain. We utilized a new generation of molecular DNA markers, minisatellites and kinetoplast DNA, for rapid characterization of the parasite. The results show that the genotypes of some of the parasites causing VL are different from those causing mucosal leishmaniasis. The L. donovani isolates causing visceral disease, as well as post-kala-azar mucosal leishmaniasis (PKML), have been shown to possess characteristic haplotypes. However, sequencing of a portion of the cytochrome oxidase II (COII) gene indicates that the parasite that invades the oral mucosa is divergent from other parasites causing VL. It appears to possess features of a more ancestral parasite with pronounced sequence homology to L. major. This agrees with earlier studies where isolates of mucosal leishmaniasis have been shown to possess an isoenzyme profile distinct from L. donovani and a different geographical distribution, albeit often overlapping with that of L. donovani.

VISCERAL LEISHMANIASIS

Visceral leishmaniasis presents a serious problem in endemic regions that is difficult to treat or prevent. Several epidemiologic problems make the disease particularly troublesome to manage. These include the facts that classic visceral leishmaniasis is fatal if untreated and there is not reliable access to medical care in many endemic regions. When available, treatment has associated toxicity and requires the use of intravenous medications with careful monitoring for toxicity, which are complex to administer in underdeveloped nations. There is an increasing incidence of the disease in HIV-infected individuals in southern Europe, in part because of the fact that eradication of the organism from infected persons using currently available drugs appears to be difficult if not impossible. Furthermore, chronic cutaneous forms of the disease allow humans and animals to maintain the organism long-term in a bodily site that is easily accessible to the sandfly vector. More effective and less toxic treatment modalities as well as a protective vaccine are badly needed to manage this disease.

Epidemiology of the Leishmaniases

The leishmaniases are a group of zoonotic infections caused by protozoan parasites of the genus Leishmania. These infections produce a variety of different clinical diseases depending on the virulence or tropism of the parasite and differential host immune responses. Newly recognized clinical presentations, such as viscerotropic leishmaniasis in American military veterans of Operation Desert Storm, continue to challenge clinicians. Epidemics of classic visceral leishmaniasis leading to thousands of deaths are ongoing in Brazil, India, and the Sudan. Epidemics of localized cutaneous leishmaniasis are ongoing in many areas of South America, North Africa, and Central Asia. A marked increase in cases is often associated with an influx of nonimmune populations into newly cleared agricultural populations into newly cleared agricultural areas or population expansion into previously unsettled areas surrounding cities. The emergence of leishmaniasis as an important opportunistic infection in AIDS patients portends an ominous future as the HIV pandemic sweeps into the hyperendemic areas of South America, Africa, and the Indian subcontinent. Parenteral transmission via needle sharing in HIV coinfected individuals in Spain is an epidemiologically significant new mode of transmission. Finally, recent work has elucidated an enzootic transmission cycle involving L. mexicana in Texas.

Ethiopian visceral leishmaniasis patients co-infected with human immunodeficiency virus

This communication reports 7 Ethiopian visceral leishmaniasis (VL) patients co-infected with human immunodeficiency virus (HIV). The clinical and laboratory findings in 6 patients did not differ from classical VL. All patients had highly elevated anti-leishmanial antibody titres, determined by immunoglobulin G-based enzyme-linked immunosorbent assay; they most probably acquired the Leishmania infection before HIV. Amastigotes were identified in the splenic aspirates of 6 patients and in the lymph node aspirate of the 2 patients whose lymph nodes were examined. The CD4:CD8 lymphocyte ratio was depressed in those patients whose ratio was determined. Most patients showed some initial response to pentavalent antimonial therapy.

The subclinical form of experimental visceral leishmaniasis in dogs.

Pathological aspects of a subclinical form of experimental canine leishmaniasis is reported here for the first time. Fifteen mongrel dogs were used in the present study. Eight dogs were infected and seven were used as control. Four of the control dogs were inoculated with spleen cells from non-infected hamsters. The eight mongrel dogs inoculated intravenously with amastigotes forms of Leishmania chagasi evolved for periods as long as 25 months without any clinical characteristic sign of classical Visceral Leishmaniasis (VL). Most of the laboratory test results were compatible to those of the seven control animals but culture of bone marrow aspirated material and serologic testing (IIF) demonstrated or provided evidence that the animals were infected. The most important and predominant histopathological lesion in infected animals were epithelioid granulomas presented in the liver, spleen, adrenal gland and lung of some animals. Channels containing erythrocytes in some granulomas of the liver suggest that these granulomas are formed inside sinusoidal capillaries. Despite the animals were proved to be infected and presented characteristic histologic lesions, they did not present external signs of disease. The granulomatous aspect of the lesions indicates a good immunologic reactivity and suggest that a host-parasite equilibrium does exist in the dog experimental model.

Immunologic markers of clinical evolution in children recently infected with Leishmania donovani chagasi.

The study attempted to identify immunologic markers for progression of Leishmania donovani chagasi infection to disease in children in an area endemic for visceral leishmaniasis (VL). [3H]thymidine uptake of lymphocytes stimulated with L. donovani chagasi antigen from children with asymptomatic infection (25,286 +/- 11,648) and from children with self-healing subclinical infection (15,511 +/- 4681) was greater (P = .001) than that observed with lymphocytes from children who progressed to classic VL (4811 +/- 2984). The interferon-gamma (IFN-gamma) levels from asymptomatic and subclinically infected children (74 +/- 90 units/ml) were higher (P = .02) than those observed in children who progressed to VL (7 +/- 8 units/ml). Absence of lymphocyte blastogenesis and IFN-gamma production were associated with progression of infection to classic VL. In the presence of these markers, children should be closely followed to identify signs and symptoms that would permit early initiation of therapy.