Molecular epidemiology of human cutaneous leishmaniasis in Jericho and its vicinity in Palestine from 1994 to 2015.

Cutaneous leishmaniases (CL) are vector-borne parasitic diseases endemic in many countries of the Middle East including Palestine. Between 1994 and 2015, 2160 clinically suspected human cases of CL from the Jericho District were examined. Stained skin tissue smears and aspirates were checked by microscopy and cultured for promastigotes, respectively. For leishmanial species identification, amplification products from a PCR-ITS1 followed by RFLP analysis using Hae III. Data were analyzed using Epi Info free-software. The overall infection rate was 41.4% (895/2160), 56.3% (504/895) of the cases were male, 43.7% (391/895) female, 60.5% (514/849) children under age 14, 41.3% (259/627) of the cases were caused by Leishmaniamajor and 57.3% (359/627) by Leishmaniatropica. The case numbers peaked in 1995, 2001, 2004, and 2012. Statistically-significant clusters of cases caused by L. major were restricted to the Jericho District; those caused by L. tropica were from the districts of Jericho, Bethlehem, Nablus and Tubas. CL is seasonal and trails the sand fly season. Distribution of cases was parabolic with fewest in July. The monthly total number of cases of CL and just those caused by L. major correlated significantly with temperature, rainfall, relative humidity, evaporation, wind speed and sunshine (P<0.05, r2=0.7-0.9 and P<0.05, r2=0.5-0.8, respectively). Cases caused by L. tropica, significantly, had a single lesion compared to cases caused by L. major (P=0.0001), which, significantly, had multiple lesions (P=0.0001). This and previous studies showed that CL is present in all Palestinian districts. The surveillance of CL has increased public awareness and molecular biological methodology for leishmanial species identification is an essential addition to classical diagnosis. The overall results are discussed, correlated to climatic and environmental changes and large-scale human activities.

Population structure and geographical subdivision of the Leishmania major vector Phlebotomus papatasi as revealed by microsatellite variation.

Multi-locus microsatellite typing (MLMT) has been employed to infer the population structure of Phlebotomus papatasi (Scopoli) (Diptera: Psychodidae) sandflies and assign individuals to populations. Phlebotomus papatasi sandflies were collected from 35 sites in 15 countries. A total of 188 P. papatasi individuals were typed using five microsatellite loci, resulting in 113 different genotypes. Unique microsatellite signatures were observed for some of the populations analysed. Comparable results were obtained when the data were analysed with Bayesian model and distance-based methods. Bayesian statistic-based analyses split the dataset into two distinct genetic clusters, A and B, with further substructuring within each. Population A consisted of five subpopulations representing large numbers of alleles that were correlated with the geographical origins of the sandflies. Cluster B comprised individuals collected in the Middle East and the northern Mediterranean area. The subpopulations B1 and B2 did not, however, show any further correlation to geographical origin. The genetic differentiation between subpopulations was supported by F statistics showing statistically significant (Bonferroni-corrected P < 0.005) values of 0.221 between B2 and B1 and 0.816 between A5 and A4. Identification of the genetic structure of P. papatasi populations is important for understanding the patterns of dispersal of this species and to developing strategies for sandfly control.

Control of phlebotomine sandflies in confined spaces using diffusible repellents and insecticides.

The control of phlebotomine sandflies (Diptera: Psychodidae), the vectors of leishmaniasis, is directed mostly against adults as larvae develop in unknown or inaccessible habitats. In the current study we tested geraniol, a natural plant-derived product, as a space repellent and the synthetic pyrethroid prallethrin as a diffusible insecticide. Geraniol was dispersed in the air using diffusers with an electric fan and prallethrin was evaporated using electrically heated evaporators. Both substances were tested in inhabited bedrooms and in tents. Geraniol failed to effect significant reductions in the numbers of either Phlebotomus papatasi Scopoli in rooms or Phlebotomus sergenti Parrot in tents. In laboratory experiments, geraniol proved ineffective in preventing sandflies from feeding. By contrast, prallethrin was highly effective in reducing the number of sandflies in rooms as well as in tents. Exposure of sandflies to prallethrin in laboratory experiments caused 97% mortality rates. Both prallethrin and, to a lesser extent, geraniol reduced the number of Culex mosquitoes captured in tents. Electric liquid-vaporizers with 1.5% prallethrin are highly effective in protecting people from sandfly bites in confined spaces and may be useful in combating cutaneous leishmaniasis.

Clinical diagnosis of cutaneous leishmaniasis: a comparison study between standardized graded direct microscopy and ITS1-PCR of Giemsa-stained smears.

Parasitological diagnosis of cutaneous leishmaniasis is absolutely necessary before treatment. Direct microscopy of scrapings taken from the margins of skin lesions is the most commonly used method for clinical diagnosis of leishmaniasis. In this study to evaluate the usage of stained smears as samples for PCR and the possible advantage of PCR, we compared the sensitivity of the diagnosis of Giemsa-stained skin scrapings by standardized graded direct microscopy with that of ITS1-PCR with the material of the same area of the slide. Three 5mm x 5mm squares were marked on each of the 20 Giemsa-stained touch smears from 20 clinically diagnosed Palestinian patients. Out of the 60 squares scanned for amastigotes under 100x oil-immersion light microscopy, 45 (75%) gave usable results and 23 of these were positive for Leishmania. Fifteen (25%) squares could not be scanned microscopically, 12 because of staining that was too thick and 3 because of inadequate staining. DNA from each scanned square was extracted separately after microscopy and run through ITS1-PCR. Of the 23 microscopy-positive squares, 20 (87%) of these were positive by PCR. Of the three that were negative, one failed to extract for DNA, the second showed only one amastigote in the entire square, and the third was normally graded as +1 but was not amplified for unknown reasons. Of the 22 squares negative for microscopy, 18 (82%) were ITS1-PCR positive. Additionally, all three improperly stained squares were ITS1-PCR positive. Of the 12 darkly stained squares, 11 were positive. A negative control group of 15 German individuals from which Giemsa-stained slides containing three squares each was prepared and these slides were also microscopically scanned and tested by ITS1-PCR. Both tests were negative with both methods. Compared to microscopy (data in parenthesis), PCR showed a sensitivity of 87% (37%) and a specificity of 100% (100%). We have concluded that Giemsa-stained smears are a readily usable sampling method for PCR and that ITS1-PCR is far more sensitive than microscopy.

The recent emergence of Leishmania tropica in Jericho (A'riha) and its environs, a classical focus of L. major.

Between 1997 and 2002, 49 strains of Leishmania were isolated from the cutaneous lesions of Palestinians living in and around Jericho. A polymerase chain reaction (PCR) amplifying the ribosomal internal transcribed spacer 1 (ITS1-PCR) was applied to their cultured promastigotes and to 207 individuals' skin scrapings spotted on filter-papers, 107 of which proved positive for leishmanial DNA. Species identification was performed by restricting the ITS1-PCR amplification products from the cultured promastigotes and the amastigotes in the scrapings with the endonuclease HaeIII. Of the 49 cultures, 28 (57%) were L. major and 21 (43%) were L. tropica. Of the 107 dermal samples tested directly, 53 (49.5%) were infected with L. major, 52 (48.5%) with L. tropica and two remained unidentified. This is the first time L. tropica has been exposed in the population of the Jericho area and on such a large scale. The itinerant behaviour of some of this population precludes categorically declaring that L. tropica has recently become established in this classical focus of L. major. For this and although 88.2% of the cases of L. tropica claimed not to have travelled out of the vicinity of Jericho, local infected sand fly vectors of L. tropica must be caught, identified and, if possible, shown to harbour infections, and, if one exists, an animal reservoir host should also be exposed to endorse whether the cases caused by L. tropica were imported or autochthonous.

Epidemiology of cutaneous leishmaniasis in the endemic area of Jericho, Palestine.

This study of cutaneous leishmaniasis in Jericho city and the adjacent Aqbat-Jaber refugee camp investigated the seroprevalance of Leishmania major and the risk factors associated with acquiring the disease. Clinical and parasitology identification of cases showed children and young men were more affected, with the head most affected in children. Enzyme-linked immunosorbent assay (ELISA) was used to test sera from 190 individuals. The overall seroprevalence of cutaneous leishmaniasis was 26.3%. A case-control study of 247 individual in 37 households showed that a higher level of education of the head of the household and having children sleep under bed nets were significantly related to a lower incidence of cutaneous leishmaniasis.

Epidemiology of cutaneous leishmaniasis in the endemic area of Jericho, Palestine

This study of cutaneous leishmaniasis in Jericho city and the adjacent Aqbat-Jaber refugee camp investigated the seroprevalance of Leishmania major and the risk factors associated with acquiring the disease. Clinical and parasitology identification of cases showed children and young men were more affected, with the head most affected in children. Enzyme-linked immunosorbent assay [ELISA] was used to test sera from 190 individuals. The overall seroprevalence of cutaneous leishmaniasis was 26.3%. A case-control study of 247 individual in 37 households showed that a higher level of education of the head of the household and having children sleep under bed nets were significantly related to a lower incidence of cutaneous leishmaniasis