Predominance of Leishmania major and rare occurrence of Leishmania tropica with haplotype variability at the center of Iran

ABSTRACT Background Leishmania major is a causative agent of zoonotic cutaneous leishmaniasis in the center of Iran, Abarkouh district. Molecular characterization and precise incrimination of Leishmania species was carried out to perform controlling measurements and to design treatment programs for zoonotic cutaneous leishmaniasis. Methods All smears isolated from ulcers of suspected patients were examined under a light microscope and graded for amastigotes frequency. Extraction of DNA, PCR, RFLP and sequencing of ITS-rDNA genotype were done to increase the efficacy of Leishmania parasites identification at their species-specific level and to detect any Leishmania infections within. Results Humans were found to be infected with L. major with high infection frequency and also Leishmania tropica was identified with low occurrence for the first time as non-native species using molecular analyses. The rates of infections was considerable with microscopic observation (n= 65, 73%) out of 89 smears prepared from suspected patients. Molecular analyses showed that the density of L. major was significantly higher (n= 48, 53.93%) than L. tropica (n= 4, 4.49%) (Mann-Whitney U test: p< 0.05) and two samples (2.25%) remained ambiguous after several sequencing. L. major did not have diversity with two common haplotypes but L. tropica were found to exhibit high diversity with three novel haplotypes. Conclusion L. major was considered the causative agent of leishmaniasis in the region, but the identification of a non-native L. tropica revealed the importance of further isolation of Leishmania parasites following molecular analyses and confirmation, and also revealed the importance of further isolation of Leishmania parasites from patients of the field areas who do not have easily access to health care centers for specialized treatment strategies.

Old world leishmaniasis presenting as a nasal mass.

Mucosal leishmaniasis is a major problem in Latin America but has been rarely noticed in our region. Although there have been a few reports of mucosal involvement especially in the oral cavity from Southwest Iran, yet none have been presented from the Northeast where Leishmania tropica is the major concern. We report a patient with endonasal leishmaniasis due to L. tropica, an extremely rare entity in immunocompetent patients in our region. He presented with a mass in the left nasal vestibule causing a sense of obstruction on the same side, accompanied by occasional rhinorrhea and mild epistaxis. This case exemplifi es the need for considering leishmaniasis as a differential diagnosis for nasal obstruction in this endemic area.

An overview of a diagnostic and epidemiologic reappraisal of cutaneous leishmaniasis in Iran

Cutaneous leishmaniasis (CL) is a widespread tropical infection which has a high incidence rate in Iran. Leishmania tropica, the causative agent of anthroponotic cutaneous leishmaniasis (ACL), and Leishmania major, which causes zoonotic cutaneous leishmaniasis (ZCL), are endemic in various parts of Iran with a high incidence rate. The aim of this study was to evaluate the reappraisal of the diagnosis and epidemiology of CL in Iran, by different clinical, parasitological and molecular assays among patients suspected of CL referred to the Department of Parasitology, at the Pasteur Institute of Iran during 2006-2009. Two hundred samples from patients with ulcerative skin lesions were collected, clinical analyses were applied, data questionnaire was completed and samples were examined for CL by using both direct microscopic and culture methods. Moreover, PCR assay was applied for detection of Leishmania species in CL isolates resulting from parasitological assay. Clinical observation revealed that the majority (58 percent) of lesions was single; double lesions were observed in 22 percent of patients, and only 20 percent of CL had multiple lesions. Out of 200 patients, Leishman body was observed in 77 samples (38.5 percent) by direct smear and 40 percent by cultivation assay. Most patients (21.3 percent) had a travel history to the Isfahan province, one of the most important endemic areas of CL located in center of Iran. PCR assay by kDNA indicated 32 and 18 out of 50 isolates respectively had similar patterns with standard L. major and L. tropica. In conclusion, clinical manifestations and an appropriate diagnostic assay with a parallel molecular characterization of CL may lead to a screening evaluation of disease, prognosis, treatment and control strategies.

Epidemiology of a new focus of localized cutaneous leishmaniasis in Himachal Pradesh.

A new focus of localised cutaneous leishmaniasis has emerged along the Satluj River valley in the mountainous region of north west Himachal Pradesh. The main endemic region extends from Pooh subdivision of Kinnaur district to Kumarsain subdivision of Shimla district with adjoining Nirmand subdivision of Kullu District comprising 86 villages. The climate of the affected areas varies from temperate to subtropical. A total of 285 cases were recorded from 1988 to January, 2005. The age of these patients varied from 10 months to 75 years, with 63 children (<12Years), and a male to female ratio of 1: 0.9. The duration of disease was 15 days to 48 months with majority (85%) presenting between 1-6 months. The number of lesions varied from 1-8, and were mostly seen on exposed parts of the body. Morphologically, lesions were asymptomatic, dry, nodular or crusted nodulo-ulcerative plaques. Tissue smear positivity for amastigotes was 43%. The characterization of 14 strains of these Leishmania revealed presence of both Leishmania tropica (n=3) and Leishmania donovani (n=11). Identification of the 42 sandflies collected from the peridomestic environment of the patients, revealed Phlebotomus longiductus - 29, P. major 8, P. kandelaki 2, while 2 remained unidentified. The patients were treated with intralesional sodium stibogluconate and majority showed excellent response.

Characterization of leishmania species and L. major stains in different endemic areas of cutaneous leishmanasis Iran

Both zoonotic and anthroponotic cutaneous leishmaniasis [CL] caused by L. major and L. tropica, respectively, are endemic in different parts of Iran. This study was performed to investigate the new changes in epidemiological pattern of CL, and to identify the species of Leishmania and the strains of L. major isolated from different endemic areas of Iran. Seventy-two isolates from 245 samples collected from different endemic areas of Iran were characterized by monoclonal antibodies [mAb] specific for L. major, L. tropica, and L. infantum. Flow cytometry, isoenzyme analysis and PCR amplification were used for identification. Isoenzyme analysis was carried out by PAGE and cellulose acetate. Eight enzymes including malate dehydrogenase [MDH], malic enzyme [ME], glucose 6-phosphate dehydrogenase [G6PD], glucose phosphate isomerase [GPI], nucleoside hydrolase inosine [NHi], nucleoside hydrolase deoxy inosine [NHd], superoxide dismutase [SOD] and 6-phosphogluconate dehydrogenase [6PGD] were used for isoenzyme analysis. PCR assay was developed using specific primers against kinetoplast DNA. The isolates were compared with reference strains [RS]. Data obtained from different methods showed 45 out of 72 isolates were similar to RS of L. major and 22 similar to L. tropica, and also five samples were identified as Crithidia. Isoenzyme migration pattern of L. major isolates was indistinguishable from each other in six enzymatic systems but differences were observed in profile of two enzymes of SOD and MDH. The data indicate that L. major species are dominant in the studied endemic areas, and different strains of L. major are present in different geographic areas of Iran. Moreover, it seems that enzymatic system is more useful approach than others for characterization of Leishmania species and L. major strains

Cutaneous leishmaniasis in Multan: Species identification

To identify the Leishmania species being responsible for cutaneous leishmaniasis in Multan. Parasites were isolated from clinically and parasitologically confirmed lesions of cutaneous leishmaniasis from 30 patients by fine needle aspiration [FNA]. The bioptical materials were then cultured in Evans Tobie's medium and parasites isolated were identified by isoenzyme electrophoresis technique. Successful Leishmania isolates were obtained from 16 patients. All strains were identified by biochemical techniques as belonging to Leishmania tropica zimodeme MON7 variant PGD. The causative species was identified as Leishmania tropica

Rapid Wright's stain for the detection of imported Leishmania tropica.

Leishmania tropica (cutaneous leishmaniasis) can be detected easily, rapidly, and conveniently by the examination of a skin ulcer smear that is stained with a modified method of Wright staining of blood (ie that used for routine hematological examinations).

Cutaneous leishman iasis: iso-enzyme charcterisation of leishmania tropica

In order to identify and characterise the organisms responsible for Cutaneous Leishmaniasis, parasites were isolated from active lesions, grown in-vitro cultures and identified by iso-enzyme characterisation. Thirteen isolates from different patients were typed as L. tropica. Seven of these isolates were from Afghan refugees encamped in the suburbs of Islamabad, 3 were from patients in Multan, 1 was from a patient from Azad Jammu and Kashmir and 1 was from Besham [Swat, NWFP]. The study confirms the presence of anthroponotic Cutaneous Leishamaniasis caused by L. Tropica in Pakistan

Leishmaniasis cutánea (Botón de Oriente) / Cutaneous leishmaniasis

Se presenta un paciente de 65 años de edad, con una lesión nodular de aproximadamente 1,5 cm de diámetro, eritematosa y asintomática, ubicada en tercio superior, cara anterior de pierna derecha. Como antecedente de importancia, refiere haber viajado a Italia donde permaneció hasta 3 meses antes de la aparición de la afección. Los estudios efectuados demostraron que se trataba de una leishmaniasis cutánea (Botón de Oriente). Se analizan las formas clínicas, métodos diagnósticos y terapéuticos, y se efectúa una revisión de la literatura de esta inusual enfermedad en nuestro medio