Leishmaniasis is an infectious disease caused by different species of unicellular Leishmania parasites which are transmitted by the bite of sandflies. As first clinical symptome, a non-healing ulcerous wound of up to several centimeters diameter forms around the byte of the insect. After several months, the wound may heal spontaneously leaving a scar. If the infection was due to a less virulent species of the parasite, the patient is healed and resistant against this form for the rest of life. However, other strains of the parasite lead to mucocutaneous leishmaniasis which lead to painfull destructions of the mucosa of lipps and nose, or to visceral leishmaniasis which affects inner organs and is lethal without treatment. Cutaneous as well as visceral leishmaniasis are endemic in the Republic of Yemen, but due to limiting financial resources, the disease is neither appropriatelytly diagnosed nor treated in the population. Recorded epidemiological data exist only for some governmental hospitals, but the number of unregisterd cases may be huge. In order to shed more light in the real extent of the disease in Yemen and to contribute to future control measures, it was attempted to design new molecular procedures for the diagnosis, which are, at the one hand, highly sensitive and specific, at the other hand not too complicated and inexpensive, so that they could be performed under the limiting conditions of the country. To obtain some insight in the sitation of leishmaniasis, the data registered in governmental hospitals during the past years were collected and evaluated. In addition, samples from more than 200 patients with suspected cutaneous and visceral leishmaniasis were collected in different parts of the country. These samples were analysed first in Yemen by microscopy for cutaneous leishmaniasis and by a highly unspecific procedure called formol gel precipitation for visceral leishmaniasis Some of the samples were also analysed by a commercial ELISA. From all collected blood samples the plasma fraction and DNA from enriched leucocytes were prepared and transported to the University of Giessen for further analysis. By using the available sequence data of the Leishmania strains L. infantum (visceral leishmaniasis) and L. major (cutaneous leishmaniasis), specific PCR primers were developed for the design of new diagnostic procedures. The assays were tested by means of purifed DNA from these two strains which had been cultivated in the laboratory. Five different target sites from highly repeated sequences of the genomes were selected to develop new diagnostic PCR assays. Two assays specific for the genus Leishmania, and one assay allowing differentiation of the two strains were constucted allowing to discriminate between cutaneous and visceral leishmaniasis. Applying these tests on the patient samples collected in Yemen revealed that PCR diagnosis is more sensitive and specific than the traditional methods used. Of course, neither a comprehensive epidemiology of Yemen nor a ultimative perfect diagnostic protocol could be obtained during this work. This has to be established during the following years. However, the results reveal at the one hand that the situation of leishmaniasis in Yemen is more severe than officially stated. At the other hand, diagnostic procedures were developed which are simple and inexpensive enough to be introduced in the country. When optimized, these techniques may lead to more awareness of this disease in the country and to an improvement of the health condition of the suffering population.
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