Trypanosoma cruzi kDNA minicircle sequences integration into the vertebrate host genome: A biomarker for monitoring the efficacy of multidrug treatment of Chagas disease

Abstract

The protist Trypanosoma cruzi inserts kinetoplast DNA minicircle sequences into the host genome. Sensitive PCR with specific primer sets revealed protozoan nuclear DNA and kinetoplast DNA in agarose gels bands probed with radiolabel specific sequences for tissues of T. cruzi-infected rabbits and mice. Avian species are refractory to the infection, but chicks that hatched from T. cruzi-inoculated eggs retained the kinetoplast DNA in the embryonic germ line cells and developed parasite-free Chagas disease-like cardiomyopathy. A Target-primer TAIL-PCR with specific primer sets, Southern hybridization, cloning, and sequencing of the amplification products revealed kinetoplast minicircle sequences integration sites mainly in LINE-1 transposable elements and hitchhiking to several loci. This kinetoplast DNA biomarker was used to monitor the effect of multidrug treatment of T. cruzi-infected mice. A trypanocidal nitro heterocyclic compound in combination with an array of inhibitors of eukaryotic cell division prevented minicircle sequences transfer. Nine out of 12 inhibitors prevented the kinetoplast DNA integration into the macrophage genome. The multidrug treatment of T. cruzi-infected mice with benznidazole, azidothymidine and ofloxacin lowered the rate of minicircle sequences integrations into the mouse genome by 2.44-fold and reduced the rejection of the target heart cells.