Leishmaniasis is a disease caused by the protozoan parasites of the genus Leishmania. The infection is transmitted between vertebrate hosts by the bite of blood sucking female phlebotomine sand flies (Diptera, Psychodida). It manifests mainly in three clinical forms: visceral, cutaneous,and mucocutaneous. Visceral leishmaniasis or kala-azar, the often a fatal form of the disease, is caused by species of the Leishmania donovani complex. These parasites were responsible for severe recent outbreaks in Sudan and other countries and are thought to originate in East Africa. Visceral leishmaniasis (VL) is endemic in 47 countries, with approximately 200 million people at risk of infection and an annual estimate of 500,000 cases. 0ver 90% of visceral leishmaniasis cases occur in six countries: Bangladesh, Brazil, Ethiopia, India, South Sudan and Sudan. The majority of cutaneous Leishmaniasis cases occur in Afghanistan, Algeria, Brazil, Colombia, the Islamic Republic of Iran, Pakistan, Peru, Saudi Arabia and the Syrian Arab Republic.
Diagnosis of Leishmaniasis:
These studies were done in collaboration with Infectious Diseases research institute in Seattle, USA (www.idri.org)
Diagnosis of the different forms leishmaniasis is still problematic and is based on the use of often invasive sampling techniques with low accuracy> Our lab has been involved in several studies to develop accurate, specific and less invasive diagnostic tests for leishmaniasis.
Development and comparative evaluation of two antigen detection tests for Visceral Leishmaniasis.
Development and evaluation of novel diagnostic tests for visceral leishmaniasis
[In collaboration with Foundation for Innovative New Diagnostics- FIND] (www.find.org ); (www.iend.org)
FIND and partners have been developing a LAMP test tat detects parasite DNA in blood (Notomi et al. 2000), and an ELISA assay for parasite antigens in urine. The two assays have been evaluated on samples collected from experimental animal infections, and on a limited number of human samples with very encouraging results. Promising new Leishmania antigens were detected in urine of Sudanese VL patients, identified, cloned and expressed
Evaluation of urinary antigen ELISA:
The newly developed urine ELISA test (developed by FIND and Kalon) will be evaluated using archived and fresh urine samples collected from parasitologically confirmed VL patients and controls. Urine samples will be collected from VL patients at days: 0, 7, 18 and 90, while for controls urine samples will be collected at day 0 only. The sensitivity and specificity of the ELISA test will be compared with the parasitology microscopy results.
Development of an ELISA test using antigens identified by IEND, and comparing its performance with another ELISA developed by FIND and partners
The expressed and purified Leishmania antigens will be used for immunization of rabbits for production of polyclonal sera and used for immunization of mice for production of monoclonal antibodies. The reactivity of the produced antibodies to the cloned antigens will be tested using western blots and ELISA. The reactive antibodies will be evaluated for detection of Leishmania antigens in archived and fresh urine samples from confirmed VL patients and healthy controls using Western blotting. The test will be further developed into a capture ELISA test using the produced polyclonal sera and the monoclonal antibodies. The developed ELISA test will be compared with the test developed by FIND using the same urine samples of patients and controls. The results of the two ELISA tests will be compared with the parasitology and qPCR tests done for the same sample donors.
Studies on the performance of a rapid diagnostic test (RDT) for Leishmania antigens that is under development
FIND and Standard Diagnostics (SD) have started development of an RDT for leishmania antigens in urine. IEND will provide SD with stored urine samples from VL patients and controls, to be used in development of the RDT. IEND will further test prototype RDTs coming from SD, and if successful, carry out prospective clinical studies.
Cutaneous Leishmaniasis:
between 0.7 to 1.2 million cases. CL is widely distributed, with about one-third of cases occurring in each of three epidemiological regions, the Americas, the Mediterranean basin, and western Asia from the Middle East to Central Asia. Afghanistan, Algeria, Colombia, Brazil, Iran, Syria, Ethiopia, North Sudan, Costa Rica and Peru, together account for 70 to 75% of global estimated CL incidence. Although CL is not a fatal diseases it causes disfiguring and disability beside its serious social stigma.
The main clinical forms of CL include localized or disseminated cutaneous lesions caused by L. major and L. tropica complexes in the old world and by L. braziliensis, L. mexicana, and related species in the new world. L. donovani generally causes visceral disease in the Old World, but recently cases of disseminated cutaneous leishmaniasis caused by L. donovani were reported. Clinical presentation of cutaneous Leishmaniasis ranges from single, multiple open ulcers, papules, nodules, or nodulo-ulcerative lesions, mainly on the exposed parts of the skin. In 20% of cases the parasite disseminates through the lymphatics, producing sporotrichoid-like lesions. Some lesion may heal spontaneously however other may persist for years and require medical treatment
Diagnosis of CL is often confused with Leprosy, skin tuberculosis, Buruliulcer , mycetoma, sarcoidosis and tropical ulcer. The diagnosis is based on the presence of cutaneous lesion and the confirmation is based on demonstration of leishmania parasites in samples collected from the lesion. Serology is of little value since the infection is confined to the skin.
The clinical samples and methods used in diagnosis of CL are variable with inconsistent accuracies. The commonly used sampling method is the skin scraping and the fine needle aspiration. Both are invasive, painful and require medical skills. The sensitivity of microscopic detection of leishmania amastigotes in smear ranges between 54% and 72% while molecular detection of parasite DNA in samples reaches 96%. However the molecular detection methods requires laboratory facilities, well trained lab personnel and financial resource.
Treatment of CL is quite variable, while some lesions heal spontaneously other persist and require treatment. Pentavalent antimony derivatives remain the mainstay of systemic treatment. Their efficiency is well established, but they can cause serious side effects and are toxic and injectable. Other treatments can be
Evaluation of urinary antigen ELISA:
The newly developed urine ELISA test (developed by FIND and Kalon) will be evaluated using archived and fresh urine samples collected from parasitologically confirmed VL patients and controls. Urine samples will be collected from VL patients at days: 0, 7, 18 and 90, while for controls urine samples will be collected at day 0 only. The sensitivity and specificity of the ELISA test will be compared with the parasitology microscopy results.
Development of an ELISA test using antigens identified by IEND, and comparing its performance with another ELISA developed by FIND and partners
The expressed and purified Leishmania antigens will be used for immunization of rabbits for production of polyclonal sera and used for immunization of mice for production of monoclonal antibodies. The reactivity of the produced antibodies to the cloned antigens will be tested using western blots and ELISA. The reactive antibodies will be evaluated for detection of Leishmania antigens in archived and fresh urine samples from confirmed VL patients and healthy controls using Western blotting. The test will be further developed into a capture ELISA test using the produced polyclonal sera and the monoclonal antibodies. The developed ELISA test will be compared with the test developed by FIND using the same urine samples of patients and controls. The results of the two ELISA tests will be compared with the parasitology and qPCR tests done for the same sample donors.
Studies on the performance of a rapid diagnostic test (RDT) for Leishmania antigens that is under development
FIND and Standard Diagnostics (SD) have started development of an RDT for leishmania antigens in urine. IEND will provide SD with stored urine samples from VL patients and controls, to be used in development of the RDT. IEND will further test prototype RDTs coming from SD, and if successful, carry out prospective clinical studies.
Cutaneous Leishmaniasis:
between 0.7 to 1.2 million cases. CL is widely distributed, with about one-third of cases occurring in each of three epidemiological regions, the Americas, the Mediterranean basin, and western Asia from the Middle East to Central Asia. Afghanistan, Algeria, Colombia, Brazil, Iran, Syria, Ethiopia, North Sudan, Costa Rica and Peru, together account for 70 to 75% of global estimated CL incidence. Although CL is not a fatal diseases it causes disfiguring and disability beside its serious social stigma.
The main clinical forms of CL include localized or disseminated cutaneous lesions caused by L. major and L. tropica complexes in the old world and by L. braziliensis, L. mexicana, and related species in the new world. L. donovani generally causes visceral disease in the Old World, but recently cases of disseminated cutaneous leishmaniasis caused by L. donovani were reported. Clinical presentation of cutaneous Leishmaniasis ranges from single, multiple open ulcers, papules, nodules, or nodulo-ulcerative lesions, mainly on the exposed parts of the skin. In 20% of cases the parasite disseminates through the lymphatics, producing sporotrichoid-like lesions. Some lesion may heal spontaneously however other may persist for years and require medical treatment
Diagnosis of CL is often confused with Leprosy, skin tuberculosis, Buruliulcer , mycetoma, sarcoidosis and tropical ulcer. The diagnosis is based on the presence of cutaneous lesion and the confirmation is based on demonstration of leishmania parasites in samples collected from the lesion. Serology is of little value since the infection is confined to the skin.
The clinical samples and methods used in diagnosis of CL are variable with inconsistent accuracies. The commonly used sampling method is the skin scraping and the fine needle aspiration. Both are invasive, painful and require medical skills. The sensitivity of microscopic detection of leishmania amastigotes in smear ranges between 54% and 72% while molecular detection of parasite DNA in samples reaches 96%. However the molecular detection methods requires laboratory facilities, well trained lab personnel and financial resource.
Treatment of CL is quite variable, while some lesions heal spontaneously other persist and require treatment. Pentavalent antimony derivatives remain the mainstay of systemic treatment. Their efficiency is well established, but they can cause serious side effects and are toxic and injectable. Other treatments can be utilized, such as miltefosine. Local therapy is used in uncomplicated lesions. Injections of the pentavalent antimony derivate, cryotherapy and paromomycinointmentsis are frequently used for treatment of Old World Leishmaniasis (8).
Evaluation of Point of Care tests for cutaneous leishmaniasis: Prospective evaluation of LoopampTM and CL DetectTM Rapid Test for cutaneous leishmaniasis diagnosis in Sudan
[In collaboration with Foundation for Innovative and New Diagnostics – FIND] (www.find.org )
To improve the diagnosis of cutaneous Leishmaniasis by improving the sampling methods and detection tools of parasite/ parasite products
Capacity building for drug sensitivity testing for anti-leishmanial drugs being used under national programme for visceral leishmanisis (VL) control: Monitoring of drug resistance of leishmania parasites in Sudan
[In collaboration with kalaCore- DIFD Project] (www.kalacore.org )
Objectives:
To perform drug sensitivity testing of anti-leishmanial drugs on Leishmania donovani
isolated from patients presenting with treatment failure or relapse after treatment for VL
Develop capacity of a regional referral point for drug sensitivity testing
Geographical mapping of unresponsiveness of anti-leishmanial drugs being used
under control programme using Arc View GIS
Drug discovery for Leishmaniasis:
- Immune response in Leishmaniasis:
Hepatitis/ HIV co-infection:
[In collaboration with Giessen University Germany and Witwitherland University- South Africa- Sponsored by DFG)
HBV and HIV: two pandemic human viruses with devastating effects on human health
HBV/HIV co-infection Sub-Saharan Africa:
In 2007, UNAIDS estimated 6% adult HIV prevalence in Africa versus 0.6% and 0.3% in North America and Western Europe, respectively. Sixty to 70% of the 34 million HIV carriers worldwide are in Africa. Thus, in addition to monoinfections with HIV and HBV, co-infections are widespread, between 16% to 98% of HIV-positive individuals having been exposed to or being carriers of HBV (25). No comprehensive data are available on the number of individuals dually infected with HBV and HIV in Africa because of the lack of systematic surveillance, underreporting and lack of resources. HIV infection causes immunosuppression and the establishment of acquired immunodeficiency (AIDS), leading to opportunistic infections and premature death. Due to the massive worldwide funding of HIV research and antiviral treatment in many of the African countries available, the problem of HBV (co-) infection in HIV-infected individuals has been neglected and research is now under-resourced.
Special situation for Sub-Saharan Africa:
Because of differences in the epidemiology, transmission patterns and genotypes of the viruses in Africa compared to non-African countries, observations obtained outside Africa cannot be extrapolated to Africa. Studies on HBV and HIV co-infections are a highly neglected area of research in Africa. Therefore, it is important that HBV and HIV should be studied simultaneously, especially in the African context, so that light can be shed on the interaction of the two viruses and the implications of the co-infection on a number of important areas including epidemiology, transmission routes, and the effect of co-infection of disease progression, OBI, HBV genotypes and treatment management. This project will be a natural progression of the work already initiated and will build on the knowledge base generated during the last few years.