Molecular Detection of Virulence Factor Glycoprotein (Gp63) of Leishmania spp. in Phlebotomus Sand Flies

Abstract

Introduction: Gp63 is the major surface glycoprotein of Leishmania which is prevalent in the promastigote stage of Phlebotomus sergenti. Glycoprotein 63 (gp63) or leishmanolysin is a zinc-dependent metalloprotease found on the surface of Leishmania. It was initially discovered in 1980and described biochemically and genetically as a surface antigen expressed in promastigotes of Leishmania species, having a range of substrates including casein, albumin, fibrinogen, haemoglobin, and gelatin. On the surface of amastigote, GP63 is present at a very low level. Glycoprotein 63 is a zinc-dependent metalloprotease that is active in the pH range of 7–10, which is neutral to alkaline. Materials and Methods: During the study, which was carried out in Al-Muthanna province from July 2017 to August 2018, about 2550 sand fly samples including 719 males and 1633 females were collected. Three species of sand flies were recorded according to morphological features which included Phlebotomus papatasi, Phlebotomus sergenti, and Sergentomyia sintoni. The caught specimens showed that P. papatasi was the most prevalent species. Two primers were used for the diagnosis of Leishmania spp. using a nested PCR technique, which was designed in a previous study for 1250 samples of sand flies from 25 locations in the study area. Results: Results showed that 13 samples were positive, containing DNA for the Leishmania parasite, and 12 samples were negative. Positive samples include 11 samples of L. major species and only 2 samples of L. tropica species. Additionally, the results showed that the L. major species was the dominant species in the study area. Positive samples (13) of sand flies had Leishmania parasite based on molecular diagnosis and virulence factor gene (Gp63) of Leishmania parasite was detected in sand flies using PCR method The results showed that virulence factor gene (Gp63) was detected on the surface of promastigote. This refers to the presence of Leishmania parasite in sand fly. The study is the first one which detected the presence of the virulence factors gene (GP63) of Leishmania in the body of sand flies. Conclusion: This study revealed the possibility of diagnosing leishmaniasis by virulence factor gene (Gp63) in leishmania promastigotes.