A.M. El Hassan

T-cell response in human leishmaniasis

In the present communication we provide evidence for the existence of a Th1/Th2 dichotomy in the T-cell response to Leishmania antigens in human leishmaniasis. Our data suggest that the pattern of IL-4 and IFN-gamma response is polarised in these patients. Lymphocytes from individuals recovered from cutaneous leishmaniasis (CL) responded by IFN-gamma production following stimulation with Leishmania antigens whereas cells from patients recovered from visceral leishmaniasis (VL) showed a mixed pattern of IFN-gamma and IL-4 responses. The cells producing these cytokines were predominantly CD4+. Furthermore, IL-10 plays an important role in the development of post kala azar dermal leishmaniasis (PKDL) from VL. The balance between the parasitic-specific T-cell response plays an important regulatory role in determining the outcome of Leishmania infections in humans.

Genetic susceptibility to visceral leishmaniasis in The Sudan: linkage and association with IL4 and IFNGR1

Longitudinal studies in Sudan show ethnic differences in incidence and clinical phenotypes associated with Leishmania donovani. Immunologically, bias in type 1 vs type 2 cytokine responses is important. To determine whether polymorphisms at IL4/IL9 or IFNGR1 contribute to susceptibility, we examined 59 multicase families of visceral leishmaniasis (VL) with/without post Kala-azar dermal leishmaniasis (PKDL). Multipoint nonparametric analysis (Allegro) linked IL4/IL9 to VL per se (P=0.002). Transmission disequilibrium testing with robust variance estimates confirmed association in the presence of linkage between VL per se and IL4 (P=0.008) but not IL9. Stepwise logistic regression analysis showed both IL4RP2 and IL4RP1 markers contributed significantly to the association, suggesting a common disease-associated haplotype. In contrast, IFNGR1 was linked (P=0.031) and associated (P=0.007) to PKDL but not VL or VL per se. Hence, polymorphism in a type 2 cytokine gene influences underlying susceptibility to VL, whereas IFNGR1 is specifically related to susceptibility to PKDL.

Liver morphology and function in visceral leishmaniasis (Kala-azar).

AIM--To study the morphology and function of the liver in visceral leishmaniasis (Kala-azar). METHODS--Percutaneous liver biopsy specimens from 18 patients with confirmed visceral leishmaniasis were examined under light and electron microscopy before and after treatment with pentovalent antimony. The tissue was also examined for hepatitis B surface and core antigens using immunoperoxidase staining. Liver function was investigated in nine patients before and after treatment. RESULTS--Specimens before treatment showed Kupffer cells and macrophages colonised by leishmania parasites in 40% of cases. A chronic mononuclear cell infiltrate had affected the portal tracts and lobules. Ballooning degeneration of the hepatocytes, fibrosis of the terminal hepatic venules, and pericellular fibrosis were common findings. The fibrosis was related to Ito cells transforming to fibroblast-like cells. None of the patients had hepatitis B infection. All patients had biochemical evidence of liver dysfunction before treatment. Liver function improved after treatment. CONCLUSION--Visceral leishmaniasis causes morphological and functional disturbance in the liver. Focal fibrosis rather than cirrhosis occurs. The exact aetiology of hepatic damage is unclear but may have an immunological basis.

Epidemiology and clinical manifestations of Leishmania donovani infection in two villages in an endemic area in eastern Sudan

We conducted a longitudinal study in an endemic area for visceral leishmaniasis (VL) in eastern Sudan to compare the epidemiology and clinical spectrum of Leishmania donovani infection in two populations differing in ethnic background and duration of residence in the area. The study took place in two villages from April 1994 to April 1996. In Um‐Salala village, which is inhabited by members of the Masaleet tribe, half of the villagers had previous exposure to cutaneous leishmaniasis (Leishmaria major) before moving there. The population of the second village, Mushrau Koka, belong to the Hausa tribe and most were born there. The incidence of VL was 20.4/1000 person‐years in 1994/1995 and increased sharply to 38.3/1000 person‐years in 1995/1996 in Um‐Salala. A rise in the incidence of VL was also observed in Mushrau Koka but with a lower incidence, 3.3/1000 person‐years to 4.6/1000 person‐years. The incidence rate of confirmed VL reflects only a limited part of the total infection rate which includes various forms of subclinical infection. The ratio of clinical to subclinical infection in Um‐Salala was 1.2 : 1 in 1994/1995 compared with 2.6 : 1 in 1995/1996. This ratio was 1 : 11 in 1994/1995 and 1 : 2.5 in 1995/1996 in Mushrau Koka. In both villages the mean age of subclinical cases was higher, but in Mushrau Koka the mean age of subclinical cases also was higher than that of subclinical cases in Um‐Salala. The leishmanin skin test (LST) was positive in 56% of individuals in Um‐Salala and in 33% in Mushrau Koka. VL only occurred in leishmanin‐negative individuals. Post kala‐azar dermal leishmaniasis (PKDL) followed in 58% of confirmed VL patients in Um‐Salala; the low incidence of VL for Mushrau Koka did not permit to estimate a PKDL rate. The clinical manifestations resulting from exposure to L. donovani range from subclinical infection to VL and PKDL. No firm conclusion as to the difference in incidence of VL between the two villages could be reached but differences in exposure to VL and cutaneous leishmaniasis (CL) as well as other factors such as ethnic background and differences in nutritional status may play a role.

Decreased erythropoietin response in Plasmodium falciparum malaria-associated anaemia

Abstract:  One of the most serious manifestations of Plasmodium falciparum malaria is anaemia. Its established causes are increased red cell destruction and ineffective erythropoiesis. Since proinflammatory cytokines have been shown to suppress the in vitro synthesis of erythropoietin (Epo), we measured serum immunoreactive Epo in 90 Sudanese patients suffering from malaria. Even in severe cases of anaemia (blood haemoglobin <80 g/l), serum Epo levels rarely exceeded 300 U/l. For comparison, serum Epo was increased up to 12,000 U/l in a reference group of Caucasian patients with anaemia not associated with infection. Moreover, the slope of the log Epo/haemoglobin regression line was less steep in malarial anaemia. Thus, as documented for other chronic inflammatory disorders, there is a relative lack of Epo in malaria‐associated anaemia. Treatment with the antimalarial drug chloroquine may aggravate the defect in Epo production, because chloroquine inhibited Epo synthesis when tested in cell culture.

Treatment of visceral leishmaniasis with sodium stibogluconate in Sudan: management of those who do not respond

Almost all (98%) of 1593 visceral leishmaniasis (VL) patients treated with sodium stibogluconate (Pentostam; Wellcome) in Sudan between 1989 and 1995 and follow-up responded well to treatment. However, the other 33 patients, all of whom were seronegative for HIV, showed partial or no response. The two main causes of unresponsiveness were primary drug resistance (39.3%) and low drug dosages given at peripheral dispensaries (30.3%). All of those who had been sub-optimal doses were cured when adequate doses of the drug were given. A third cause was concurrent disease, particularly pulmonary tuberculosis (18%). With treatment of the concurrent disease, patients responded well to Pentostam. Eight patients who failed to respond to repeated courses of Pentostam did not benefit from pentamidine or sterol inhibitors. Three of these patients responded to liposomal amphotericin B, two responded to splenectomy in association with Pentostam therapy, and three died. Pentostam, given in adequate doses, still appears to be the drug of choice for the treatment of VL in the Sudan Liposomal amphotericin B is a suitable second-line drug.

Dichotomy of the T cell response to Leishmania antigens in patients suffering from cutaneous leishmaniasis; absence or scarcity of Th1 activity is associated with severe infections

The T cell response was studied in 25 patients suffering from cutaneous leishmaniasis caused by Leishmania major with severe (n= 10) and mild (n= 15) disease manifestations. Peripheral blood mononuclear cells (PBMC) from the patients were activated by sonicates of Leishmania promastigotes (LMP) and amastigotes (LDA), and the surface protease gp63. The proliferative responses to Leishmania antigens were lower in patients with severe disease than in patients with mild disease (P= 0·01–0·05), and such a difference was not observed in the response to purified protein derivative of tuberculin (PPD) or tetanus toxoid (TT). LMP‐induced interferon‐gamma (IFN‐γ) production was lower in patients with severe than in patients with mild disease (P < 0·05). When the IL‐4 and IFN‐γ responses of each patient were considered, two response patterns were observed in the cultures activated by the Leishmania sonicates. One response pattern was characterized by high production of IFN‐γ without production of IL‐4 (a Th1‐like pattern), the other was characterized by low IFN‐γ levels which in most cases were associated with IL‐4 production (not a Th1‐like pattern). These patterns could not be distinguished when the cells from the same donors were stimulated by TT and PPD. The percentages of patients with a Th1‐like response pattern after stimulation by LMP in patients with severe and mild disease manifestations were 30% and 80%, respectively. This difference was statistically significant (P= 0·034).

The host tissue reaction to Madurella mycetomatis: new classification

In this prospective study, three types of tissue reaction to mycetoma grains are described. Type I reaction is characterized by the adherence of neutrophils to the surface of the grain leading to its disintegration. In Type II reaction, the fragmented grain and the dead neutrophils are cleared by macrophages and multinucleated giant cells, while in Type III reaction there is a discrete well developed epithelioid granuloma with Langhanss giant cells. The three types of reaction are usually seen in different combinations in the same lesion.

Immunogenicity and safety of autoclaved Leishmania major plus BCG vaccine in healthy Sudanese volunteers

In a longitudinal study in the epidemiology of Leishmania donovani infection in an endemic focus in eastern Sudan, we observed that previous exposure or infection with Leishmania major appeared to protect against visceral leishmaniasis caused by L. donovani. We therefore conducted a study to test the safety and immunogenicity of a vaccine consisting of autoclaved L. major (ALM) plus BCG in inducing protection in vaccinated individuals. Leishmanin-negative healthy Sudanese volunteers were enrolled in the study and were divided into three groups: group (A) received ALM+BCG, group (B) received BCG alone, and group (C) received the vaccine diluent. The subjects were examined for their clinical and immunological responses before intervention, following intervention and 6-8 weeks after vaccination. Vaccinated subjects (group A) developed localized reactions at the sites of vaccine inoculation that ulcerated and healed within 4-6 weeks; 61.6% of them converted to leishmanin reactive following vaccination. Only one subject in group (C) became leishmanin-positive. A total 76.9% of the vaccinated volunteers in group (A) produced significant levels of interferon-gamma in response to L. major antigen. The vaccine produced significant cellular immune responses that may protect against natural challenge. None of the groups had systemic reactions and all the reactions observed in the vaccinated group were comparable with the BCG-vaccinated group.

Serodiagnosis of Leishmania donovani infections: assessment of enzyme-linked immunosorbent assays using recombinant L. donovani gene B protein (GBP) and a peptide sequence of L. donovani GBP.

The repetitive sequence of Leishmania major gene B protein (GBP) has previously been shown to be a useful tool in the diagnosis of cutaneous leishmaniasis (CL). Here, we have assessed enzyme-linked immunosorbent assays (ELISAs) using recombinant L. donovani GBP (rGBP) and a peptide sequence of L. donovani GBP (GBPP) in the diagnosis of L. donovani infections in Sudan. The sensitivity of the rGBP ELISA in diagnosing visceral leishmaniasis (VL) and post kala-azar dermal leishmaniasis (PKDL) was 92% and 93%, respectively. In contrast, the sensitivity of the GBPP ELISA was 55% for VL and 63% for PKDL. Plasma antibody reactivity of donors with VL and PKDL remained high for an extended period after the end of treatment. Antibody-reactivity to rGBP and GBPP was detected in 71% and 14% of plasma samples from CL patients, respectively. Plasma from healthy Sudanese donors living in an area endemic for malaria but free of leishmaniasis was negative in both assays.