Ahmed M. El Hassan

Immunopathology of post kala-azar dermal leishmaniasis (PKDL): T-cell phenotypes and cytokine profile

In Sudan, post kala‐azar dermal leishmaniasis (PKDL) caused by Leishmania donovani develops in half of the patients treated for visceral leishmaniasis (kala‐azar). In most patients lesions heal spontaneously, but in others symptoms are severe and persist for years. This study examined the immunological response in lesions of PKDL patients by immunohistochemistry and compared the findings with results obtained using peripheral blood mononuclear cells (PBMCs). In all lesions, parasites or parasite antigen were present and provoked the formation of an inflammatory infiltrate consisting of a mixture of macrophages, lymphocytes, and plasma cells. In patients who had high interferon‐gamma (IFNγ) responses to Leishmania antigen in vitro, compact epithelioid granulomas were formed. The inflammatory cells were mainly CD3+ and interleukin‐10 (IL10) was the most prominent cytokine in the lesions. However, IFNγ was found in all and IL4 in most lesions, in varying amounts. PBMCs from all patients responded to Leishmania antigen by IFNγ production or proliferation. The results indicate that PKDL develops as a result of an influx of immunocompetent cells into skin, which harbours parasites. The inflammatory response to the parasites is complex. It involves several cell types and cytokines, of which some are antagonistic. It is conceivable that the balance between these cytokines determines the outcome of the disease. Copyright

The Marrara syndrome: a hypersensitivity reaction of the upper respiratory tract and buccopharyngeal mucosa to nymphs of Linguatula serrata.

The Marrara syndrome, like Halazoun in Lebanon, is a hypersensitivity reaction of the upper respiratory tract and buccopharyngeal mucosa to nymphs of Linguatula serrata. The condition follows the consumption of Marrara which consists of raw liver, lungs, trachea and rumen of goats and sheep infected with larvae of L. serrata. The adult worm is found in the nasal passages of dogs. Sheep and goats are infected by eggs from infected dogs. A survey that included 240 adult individuals in a village of endemic L. serrata infection in the Sudan showed that 20% experienced symptoms of allergic nasopharyngitis (Marrara syndrome) following the consumption of raw viscera of goats or sheep at least once in their life. In a prospective study of 24 patients who reported to hospital with the Marrara syndrome, the clinical features included itching in the throat and nose, unilateral conductive deafness, tinnitus and facial palsy. Secondary bacterial infection caused suppurative otitis media. Adult L. serrata parasites were found in the nasal passages of 56 and 47% of male and female dogs in the endemic area. Nymphs were recovered from the mesenteric lymph nodes, lungs and livers of goats in the area.

Mycetoma in the Sudan: An Update from the Mycetoma Research Centre, University of Khartoum, Sudan

This communication reports on the Mycetoma Research Centre of the University of Khartoum, Sudan experience on 6,792 patients seen during the period 1991–2014.The patients were predominately young (64% under 30 years old) males (76%). The majority (68%) were from the Sudan mycetoma belt and 28% were students. Madurella mycetomatis eumycetoma was the most common type (70%). In 66% of the patients the duration of the disease was less than five years, and 81% gave a history of sinuses discharging mostly black grains (78%). History of trauma at the mycetoma site was reported in 20%. Local pain was reported in 27% of the patients, and only 12% had a family history of mycetoma. The study showed that 57% of the patients had previous surgical excisions and recurrence, and only 4% received previous medical treatment for mycetoma. Other concomitant medical diseases were reported in 4% of the patients. The foot (76%) and hand (8%) were the most commonly affected sites. Less frequently affected sites were the leg and knee (7%), thigh (2%), buttock (2%) and arm and forearm (1%). Rare sites included the chest wall, head and neck, back, abdominal wall, perineum, oral cavity, tongue and eye. Multiple sites mycetoma was recorded in 135 (2%) of cases. At presentation, 37% of patients had massive lesions, 79% had sinuses, 8% had local hyper-hydrosis at the mycetoma lesion, 11% had regional lymphadenopathy, while 6% had dilated tortuous veins proximal to the mycetoma lesions. The diagnosis of mycetoma was established by combined imaging techniques and cytological, histopathological, serological tests and grain culture. Patients with actinomycetoma received a combination of antimicrobial agents, while eumycetoma patients received antifungal agents combined with various surgical excisions. Surgical excisions in the form of wide local excision, debridement or amputation were done in 807 patients, and of them 248 patients (30.7%) had postoperative recurrence. Different types of amputations were done in 120 patients (1.7%).

Relationship of the Sickle Cell Gene to the Ethnic and Geographic Groups Populating the Sudan

The presence of a geographical pattern in the distribution of the sickle cell gene (S gene) and its association with malaria is well documented. To study the distribution of the S gene among various ethnic and linguistic groups in the Sudan we analyzed a hospital-based sample of 189 sickle cell anemia (SCA) patients who reported to the Khartoum Teaching Hospital between June 1996 and March 2000 and 118 controls with other complaints, against their ethnic and linguistic affiliations and geographic origin. Electrophoresis for hemoglobin S and sickling tests were carried out on all patients and controls as a prerequisite for inclusion. The majority of patients (93.7%) belonged to families of single ethnic descent, indicating the high degree of within-group marriages and thus the higher risk of augmenting the gene. SCA was found to be predominant among the Afro-Asiatic-speaking groups (68.4%) including nomadic groups of Arab and non- Arab descent that migrated to the Sudan in various historical epochs. Those patients clustered in western Sudan (Kordofan and Darfur) from where 73% of all cases originate. The proportion of patients reporting from other geographic areas like the south (3.1%), which is primarily inhabited by Nilo-Saharan-speaking groups (19% of the whole sample) who populated the country in previous times, is disproportionate to their total population in the country (χ2 = 71.6; p = 0.0001). Analysis of the haplotypes associated with the S gene indicated that the most abundant haplotypes are the Cameroon, Benin, Bantu and Senegal haplotypes, respectively. No relationship was seen between haplotypes and the various hematological parameters in the sub-sample analyzed for such association. These results provide an insight into the distribution of the sickle cell gene in the Sudan, and highlight the strong link of the middle Nile Valley with West Africa through the open plateau of the Sahel and the nomadic cattle herders and also probably the relatively young age of the S gene.

Oral leishmaniasis associated with kala-azar

Mucosal leishmaniasis as an oral disease in the form of chronic periodontitis with involvement of the oral mucosa is described. Leishmania parasites were isolated from the oral lesions, lymph nodes, and bone marrow. The patient had a low-grade fever and hepatosplenomegaly that regressed along with the oral lesions after treatment with stibogluconate sodium.

Pathology of post-kala-azar dermal leishmaniasis: a light microscopical, immunohistochemical, and ultrastructural study of skin lesions and draining lymph nodes.

Background:  Whereas the clinical manifestations and treatment of post‐kala‐azar dermal leishmaniasis (PKDL) have been adequately described before, the pathology received little attention, particularly the African form of PKDL which shows some clinical differences from the disease in India. Therefore, our aim was to characterize the pathology and the immunohistopathology in PKDL lesions and correlate the histopathological findings with the clinical features of the disease.

Humoral and cellular immune responses to glucose regulated protein 78 – a novel Leishmania donovani antigen

The recently cloned glucose regulated protein 78 (GRP78) of Leishmania donovani has been suggested as a new and promising Leishmania vaccine candidate. We assessed antibody and T‐cell reactivity to GRP78 in an enzyme‐linked immunosorbent assay (ELISA) and in lymphoproliferative assays. Serological evaluation of plasma samples obtained in Sudan revealed that 89% of patients with visceral leishmaniasis (VL), 78% with post kala‐azar dermal leishmaniasis (PKDL), and 85% with cutaneous leishmaniasis (CL) had antibody reactivity to this Leishmania antigen. Plasma from healthy Sudanese individuals living in an area endemic for malaria but free of leishmaniasis and plasma from healthy Danes was negative in the assay. GRP78 antibody was detected in 10% and 5% of plasma samples from Sudanese and Ghanaian malaria patients, respectively, whereas 35% of plasma samples from otherwise healthy Sudanese individuals with a positive leishmanin skin test showed antibody reactivity to recombinant GRP78 (rGRP78). In lymphoproliferative assays, 9 of 13 isolates of peripheral blood mononuclear cells (PBMC) from individuals previously infected with L. donovani and one of three individuals previously infected with L. major showed a response to rGRP78, whereas PBMC isolates from Danish control individuals did not respond. These findings, in addition to our previous observations in experimental CL ( Jensen et al. 2001 ), confirm GRP78 as a possible vaccine antigen.

A Histopathological Exploration of the Madurella mycetomatis Grain

Although the Madurella mycetomatis grains seem to interfere with the host defense mechanisms and impede the antifungal drugs penetration, yet their histological features are not fully known and hence this study was set out to determine that. The study included 80 patients with confirmed M. mycetomatis eumycetoma. After informed written consent, surgical biopsies were obtained from the excised tissues during the patients’ surgical treatment. All sections were stained with haematoxylin and eosin, Grocott’s hexamine silver, Periodic Acid-Schiff’s, Masson-Fontana, Perl’s Prussian Blue, Von-kossa’s, Formalin Inducing Fluorescence and Schmorl’s stains. Modified bleaching technique was used. The concentrations of Zinc, Copper, Calcium, Iron, Lead, Cobalt and Nickel were determined by Atomic Absorption Spectrophotometer. The M. Mycetomatis grains appeared to consist of lipid, protein and melanin. The melanin was located on the hyphal wall as thick layers. The Zinc, Copper and Calcium concentrations in the grains were four, six, and sixteen folds higher than in normal tissue respectively, the other metals were found in the same concentrations as in normal tissue. In the grains, calcium was located in the melanin vicinity. From this study, it can be concluded that, the grains contain melanin, heavy metals, proteins, lipids and they contribute in the formation of the grain cement matrix. These elements seem to contribute in the organism pathogenicity and might impede the penetration of various anti-fungal agents.

The effect of repeated leishmanin skin testing on the immune responses to Leishmania antigen in healthy volunteers

The leishmanin skin test (LST) is used in immunogenicity studies. The effect of multiple LSTs on immune responses was assessed. None of the volunteers converted to LST positive. IFN-gamma and IL-10 levels remained unchanged. Repetition of LST does not modulate the in vivo or in vitro immune responses to Leishmania antigen.

Prevention of measles in Sudan: a prospective study on vaccination, diagnosis and epidemiology

Despite the availability of safe and effective live attenuated vaccines, measles continues to be endemic in many developing countries. Control and elimination of measles will be especially difficult in East Africa, because of its limited infrastructure and political instability. We have studied diagnostic and epidemiological aspects of measles in suburban Khartoum, Sudan. Prospective studies were carried out in a cohort of clinically diagnosed measles cases and in a cohort of newborns, which were both followed up for 2 years. The studies intended to provide a rational basis for improvement of measles vaccination strategies, and strengthen measles research infrastructure in Khartoum.