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.

Serum erythropoietin concentration in anaemia of visceral leishmaniasis (kala-azar) before and during antimonial therapy.

Serum erythropoietin (Epo) concentrations and variables of red cell and iron status were studied in 27 Sudanese patients who were treated with sodium stibogluconate for visceral leishmaniasis (kala-azar). Blood haemoglobin increased from 6.4 (+/- 1.7 SD) to 9.5 (+/- 1.4) g/dl during treatment. Serum ferritin decreased concomitantly. Serum iron levels were unchanged whereas the total iron binding capacity increased slightly. The pre-treatment serum Epo concentration in relation to the blood haemoglobin concentration was not as high as expected from the one in primary haematological diseases, indicating that there is a relative lack of Epo in anaemic kala-azar patients. Serum Epo further decreased during stibogluconate therapy. The normal dependence of the serum Epo level on the blood haemoglobin concentration was lost during mid-term antimonial treatment, but it recovered thereafter. Cell culture studies with the human hepatoma cells HepG2 showed that stibogluconate (> or = 30 microg/ml) inhibited Epo gene expression. Thus, effective treatment of kala-azar with stibogluconate results in improvement of anaemia, although the drug itself may impair Epo production.

Oronasal leishmaniasis caused by a parasite with an unusual isoenzyme profile.

A 45-year-old Sudanese man from western Sudan presented with oronasal leishmaniasis of three years duration. He had no history of previous kala-azar or cutaneous leishmaniasis. The parasite isolated from the oral mucosa was characterized by isoenzymes using 12 enzymes and by polymerase chain reaction amplification of kinetoplast DNA using species-specific primers. The specific primers gave products indistinguishable from those of the Leishmania donovani complex. However, the isoenzyme profile showed a zymodeme pattern that was significantly different from the zymodemes previously reported in the Sudan and the Ethiopian region.

IL-12 enhances Th1-type responses in human Leishmania donovani infections.

IL-12 is a pluripotent cytokine that interacts with NK and T cells to play a central role in the initiation and maintenance of Th1 responses and IFN-gamma production. Because of the interactive relationship between IL-12 and IFN-gamma response to infectious organisms, a study was undertaken to examine the role of IL-12 in the immune regulation of human visceral leishmaniasis (VL). Human (Hu) VL is associated with immune dysfunction and the appearance of IL-10 mRNA, not present in healed individuals. We found that PBMC from treated VL patients produced both IL-12 p40 and IFN-gamma in response to in vitro stimulation with Leishmania donovani. The production of both IL-12 p40 and IFN-gamma were interdependent and were abrogated by the addition of exogenous Hu rIL-10. In contrast, PBMC from active VL patients did not produce IL-12 p40 or IFN-gamma in response to L. donovani lysate. Neutralizing anti-IL-10 mAb led to the enhancement of IFN-gamma production by active VL PBMC cultured with L. donovani lysate, and this enhanced IFN-gamma production was blocked by anti-IL-12 mAb. The addition of exogenous Hu rIL-12 to PBMC from active VL patients resulted in the augmentation of IFN-gamma in response to L. donovani lysate. Therefore, treatment of active VL patient PBMC with anti-IL-10 or IL-12 shifted the response toward a Th1-type response with the production of IFN-gamma. These results indicate that IL-12 may play an important role in the regulation of the cellular immune responses in Hu VL.

Neurologic changes in visceral leishmaniasis.

Neurologic changes in visceral leishmaniasis (VL) are rarely reported. From January 1992 to April 1993, 111 patients with VL were seen at Soba University Hospital in Khartoum, Sudan. Fifty-two (46%) patients had neurologic symptoms or signs; the most common symptom was a sensation of burning feet. Four patients had foot drop. Five patients had deafness and one patient had multiple cranial nerves palsies. None of our patients had vitamin deficiency or any of the other known causes of neuropathy. Nerve conduction studies in 15 patients showed evidence of axonal degeneration and demyelination, which were confirmed by histopathology and electron microscopy of nerve biopsies. There was no direct parasitic infection of the nerve and there was no neuritis. In most patients, the sensory symptoms disappeared within two weeks in most of our patients after specific anti-leishmanial treatment. Motor recovery was much slower. Audiographic studies in five patients with deafness showed it to be sensory-neural. Hearing returned to normal after treatment with sodium stibogluconate. Further studies are needed to define the etiology of the nerve pathology in patients with VL.

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.

Tinea versicolor and visceral leishmaniasis.

BACKGROUND: Visceral leishmaniasis (VL) is endemic in several areas in the Sudan. The disease is associated with depressed cellular immunity. Tinea versicolor is a normal commensal of the skin which can become pathogenic particularly in patients with depressed cell-mediated immunity. Patients with VL have a high prevalence of tinea versicolor. METHODS: One hundred and thirty patients with parasitologic confirmation of VL were screened for tinea versicolor infection. In the suspected cases the diagnosis was made by demonstrating the fungal hyphae and spores in skin scrapings. All patients were treated with sodium stibogluconate. RESULTS: Of the 130 patients with VL, 10.8% were found to have severe tinea versicolor. The fungal infection developed or became worse with the start of VL. After successful treatment of VL, the tinea lesions disappeared completely or decreased in severity. CONCLUSIONS: Depressed cell-mediated immunity that is a feature of VL is the probable underlying cause for fungal infection. Tinea infection during the course of VL is to be distinguished from lesions of post-kala-azar dermal leishmaniasis.

Kala-azar in western Upper Nile province in the southern Sudan and its spread to a nomadic tribe from the north.

Since the start in 1988 of the present epidemic of kala-azar (visceral leishmaniasis) in western Upper Nile state in southern Sudan, the epidemiology of the disease in all parts of the Sudan where kala-azar has been reported was reassessed by the Leishmaniasis Research Group in Khartoum. In this paper, the spread of the epidemic is described among a nomadic tribe originating from southern Kordofan state, who migrate every year with their cattle to the Bentiu area in western Upper Nile state where the epidemic is still raging. 200 cases from this tribe were seen in Khartoum; another 56 cases were found during a field trip to the area. In addition, the Bentiu area was visited, where 301 cases were under treatment and another 52 of 1120 individuals screened were confirmed parasitologically. 20 cases of post-kala-azar dermal leishmaniasis were found. Parasites isolated from the nomadic tribe were of the same zymodeme as parasites isolated previously from the Nuer in western Upper Nile. The epidemiological findings in each state are discussed in relation to the tribes that were affected and the ecology of the area.

Interleukin 10 production correlates with pathology in human Leishmania donovani infections.

We have found that an important Th2 cytokine, IL-10, is produced by tissues from patients acutely infected with Leishmania donovani. In all individuals tested, IL-10 mRNA production was increased in lymph nodes taken during acute disease over that observed in postacute samples. In contrast, both pre- and posttreatment lymph nodes had readily detected mRNA for IFN-gamma and IL-2. A down-regulating effect of IL-10 on leishmania-induced proliferative responses was demonstrated when Hu rIL-10 was added to cultures of PBMC from clinically cured individuals. PBMC from individuals with acute visceral leishmaniasis responded to stimulation with leishmania lysate by producing IL-10 mRNA. Simultaneously cultured PBMC collected from the same patients after successful chemotherapy produced no detectable IL-10 mRNA after leishmania antigen stimulation. Neutralizing anti-IL-10 mAb added to PBMC from patients with acute visceral leishmaniasis markedly increased the proliferative response to leishmania lysate. Finally, we observed mRNA for IL-10 and IFN-gamma concurrently in a lesion from a patient with post-kala-azar dermal leishmaniasis (PKDL). These results indicate the production of IL-10 during L. donovani infection, and suggest a role for this cytokine in the regulation of immune responsiveness during visceral leishmaniasis.

In vivo cytokine profiles in patients with kala-azar. Marked elevation of both interleukin-10 and interferon-gamma.

The immunological mechanisms underlying the susceptibility to disseminated visceral parasitism of mononuclear phagocytes in patients with kala-azar remain undefined. Resistance and susceptibility are correlated with distinct patterns of cytokine production in murine models of disseminated leishmanial disease. To assess lesional cytokine profiles in patients with kala-azar, bone marrow aspirates were analyzed using a quantitative reverse transcriptase PCR technique to amplify specific mRNA sequences of multiple Th1-, Th2-, and/or macrophage-associated cytokines. Transcript levels of IL-10 as well as IFN-gamma were significantly elevated in patients with active visceral leishmaniasis; IL-10 levels decreased markedly with resolution of disease. These findings suggest that IL-10, a potent, pleiotropic suppressor of all known microbicidal effector functions of macrophages, may contribute to the pathogenesis of kala-azar by inhibiting the cytokine-mediated activation of host macrophages that is necessary for the control of leishmanial infection.

Distinguishing post-kala-azar dermal leishmaniasis from leprosy: experience in the Sudan.

In this study 4 patients were post-kala-azar dermal leishmaniasis (PKDL), whose lesions were similar to those of lepromatous and borderline leprosy, are described. In 2 patients there was no previous history of kala-azar but they were residents of an area of known endemic kala-azar. Lack of proper clinical and laboratory assessment was behind the failure to diagnose PKDL. Consequently the patients were treated with antileprosy drugs without proof of leprosy. The 3rd and 4th patients, though suspected clinically of leprosy, were correctly diagnosed as PKDL with adequate history, clinical assessment and appropriate laboratory investigations. The salient points in distinguishing PKDL from leprosy are described and discussed.

TSH receptor antibodies in autoimmune thyroiditis.

Out of 2,322 patients attending a thyroid clinic in north west Germany over a three year period, 123 were found to have evidence of autoimmune thyroiditis (hypothyroid, latent hypothyroid or euthyroid) and 96 were available for further analysis. TSH receptor antibodies (TRAb) were detectable by receptor assay in five of these patients (and a further two who attended the clinic later) and all the TRAb positive sera showed TSH blocking activity by bioassay. All of the patients with blocking activity were hypothyroid (on treatment) and represented 15% of this group of 34 patients. This suggests that in hypothyroid patients with autoimmune thyroiditis, the prevalence of TSH receptor antibodies with blocking activity is similar in northern Europe and Japan (21% of 43 patients; (1]. In the present study, no relationship between thyroid volume as assessed by sonography and the presence or absence of blocking antibodies was apparent. As blocking antibodies were undetectable in patients at early stages of the disease (i.e., in the euthyroid or latent hypothyroid groups) it seemed unlikely that these antibodies were a major causative factor in the development of hypothyroidism.

Photoaffinity labelling of the TSH receptor on FRTL5 cells.

An investigation of the properties of TSH receptors on FRTL5 cells using affinity labelling with a 125I-labelled photoactive derivative of TSH is described. Our studies suggest that FRTL5 cells contain 2 principal types of cell surface TSH receptors. One form, probably a precursor, consists of a single polypeptide chain (Mr 120,000) with an intrachain loop of amino acids formed by a disulphide bridge. The other type of receptor consists of a water-soluble A chain (Mr 55,000) linked to an amphiphilic B chain (Mr 35,000) by a disulphide bridge. The 2 chain structure is probably derived from the single chain 120,000 protein by enzymatic cleavage of peptide sequences within the loop of amino acids formed by the intrachain disulphide bridge.

The TSH receptor: structure and interaction with autoantibodies in thyroid disease.

Studies of the TSH receptor using affinity labelling with photoactive derivatives of TSH and analysis by SDS-PAGE have shown that the receptor contains 2 subunits (A and B), linked by a disulphide bridge. Similar results are obtained with TSH receptors from human, porcine and guinea pig thyroid tissue and from guinea pig fat. Analysis of affinity labelled receptors under non-denaturing conditions suggest that subunits additional to the A and B subunits are not present. Hydrodynamic measurements indicate that the receptor A subunit has an approximately spherical structure (Stokes' radius 70 A) and when this interacts with TSH (an elongated structure with Stokes' radius 56A) a very elongated complex (Stokes' radius 104A) is formed. Isoelectric focusing studies of the TSH receptor A subunit, TSH and TSH receptor antibodies indicate that charge-charge interactions are of considerable importance in the binding of hormone and antibody to the receptor.

Characterization of TSH antagonist activity in the serum of patients with thyroid disease.

The ability of sera from patients with thyroid disease to block TSH stimulation of cyclic AMP release from isolated porcine thyroid cells has been assessed and the blocking activity characterized. TSH receptor binding activity was also measured. No blocking or receptor binding activity was detectable in patients with primary myxoedema (n = 23), Hashimoto's disease (n = 11), multinodular goitre (n = 6), or rheumatoid arthritis (n = 10). However, analysis of sera from 23 patients (out of an initial screen of 110 patients) with treated Graves' disease which did not stimulate cyclic AMP production in the bioassay showed that two of these sera contained powerful blocking and receptor binding activity. Both these patients had been treated with 131I. Analysis of the two sera by gel filtration on Sephadex G-200 indicated that blocking and TSH receptor binding activity were associated only with the IgG fraction. Digestion of the IgG with pepsin followed by reduction showed that both (Fab)2 and Fab fragments contained high levels of blocking and binding activity. Antibody divalency was not necessary therefore for TSH antagonist activity. However, our studies suggest that autoantibodies of this type with TSH antagonist activity do not occur frequently in patients from the Cardiff region with primary myxoedema, Hashimoto's or treated Graves' disease.