In vitro infectivity of leishmania major isolated from patients with different clinical forms of cutaneous leishmaniasis and its association with parasite zymodems

The aim of this study was to characterize the Leishmania parasites isolated from cutaneous leishmaniasis [CL] patients in Pars Province in Iran and to compare the potential infectivity of the isolates in macrophage cell line. Moreover, attempt was made to find out the association between parasite infectivity and their zymodems. Twenty samples were taken from the skin lesion of CL patients. The samples were cultured in biphasic media followed by mass cultivation in RPMI medium. Each isolate was tested for the activity of the 5 enzymes including glucose phosphate isomerase [GPI], malate dehydrogenase [MDH], nucleoside hydrolase 1 and 2 [NH1 and NH2], and phosphoglucomutase [PGM]. The enzymatic profiles of the isolates were compared with WHO reference strains. Specific PCR [primers: LIN17 and LIN R4] and RAPD-PCR were used as complementary methods for characterization of the isolates. Isoenzyme electrophoresis showed that all of the isolates were L. major. PCR with LIN 17 and LIN R4 and RAPD-PCR with AB-07 primers further determined the isolates as L. major. Results of macrophage infectivity experiment, using J774 cell line, showed that the most virulent isolates were related to Z1 with 63% macrophage infectivity rate. A well correlation was found between the infectivity rate of the isolates and type of ulcer. Those isolates with high infectivity rate were involved in more severe, ulcerative or erythmatose lesions in CL patients. The most invasive isolates might be a good candidate for immunological studies and for vaccine development

[Characterization of leishmania parasites isolated from kala- azar patients in kohgiloyeh and boyerahmad, using semi-nested PCR]

Visceral leishmaniasis [VL] is a disease commonly known as Kala-azar caused by protozoan parasites of the genus Leishmania including L. donovani, L. infantum and L. chagasi. VL is sporadic in manyareas of Iran and is endemic in a few provinces such as Fars, Azarbayjan, Bushehr, Ardabil and Qom. VL has been reported from some areas of Kohgiloyeh and Boyerahmad and this study aimed to characterize the causative agent of VL in this region. Bone marrow sample was obtained from 6 VL patients from children department in Imam Sajad hospital in Yasuj. DNA was extracted from the obtained samples and was checked by semi-nested PCR to determine the species of the parasite. To do that, a segment of minicircle kinetoplast DNA was amplified, using LINR4 and LIN17 primers. Products of PCR were evaluated by electrophoresis, using 1.5% agarose and stained with ethidium bromide. Parasitologically examination of bone marrow smears demonstrated amastigotes form of the parasite in the samples. For mass cultivation, isolated parasites were cultured in diphasic NNN followed by RPMI 1640 media. All the samples produced a 720 bp band in PCR assay. The isolates were compared with referent strains and it was revealed that all the isolates were L. infantum. Findings of this study demonstrated that the causative agent of VL in Kohgiloyeh and Boyerahmad was L. infantum. Further study is needed to explore other aspects of VL in this region