Viscerotropic Leishmaniasis; Diagnosis and species detection using the polymerase chain reaction with kinetoplast DNA-specific primers in Balb/c mice

Abstract

The study involves three groups of Balb/c mice, the first and second groups inoculated in the hind footpad (1 x 106), (3 x 106) stationary phase promastigotes of L. tropica respectively, the third group inoculated phosphate buffer saline. The aim of this study was to establish a murine model for human viscerotropic leishmaniasis, and the dissemination of L. tropica to visceral organs tissues of BALB/c mice in order to find out the ability to diagnosis the disease by polymerase chain reaction (PCR) as recent application. The samples of liver tissues were collected and DNA was extracted by Promega DNA kit purification. The polymerase chain reaction (PCR) with kDNA specific primers was used firstly to distinguish the species of parasite which isolate from patient had cutaneous lesion acquired in Iraq where only L. major and L. tropica were expected to be the causative agents. The size difference between the PCR products of L. major and L. tropica allowed differential diagnosis. The smaller product (650 bp) could be identified as derived from L. major, whereas the larger product (800 bp) was due either to L. tropica or to a member of the L. donovani complex which yielded the same size of band. The same primers were used to detect the dissemination of parasites to visceral organs via extracted DNA from liver tissues of infected animals. PCR application was very high sensitive and early diagnosis in liver tissues as a substrate for PCR reaction, all animals of group (1) which infected with (106) promastigotes were negative result at (15) days post-infection, whereas three animals from group (2) which infected with (3x106) promastigotes showed positive result at the same period. All animals except one from group (1) showed positive results at (30) days post-infection. But at the later sequences days all animals were positive.