A non-replicative antibiotic resistance-free DNA vaccine delivered by the intranasal route protects against canine leishmaniasis.

Leishmania infantum is the etiological agent of zoonotic visceral leishmaniasis (ZVL). The disease is endemic in Central and South America, Central and South East Asia, and the Mediterranean basin. Dogs are the main reservoir, with an estimated prevalence of approximately 2.5 million dogs in Southern Europe. Current treatments cause side effects, disease recurrence, and drug resistance. Therefore, the development of vaccines against canine leishmaniasis is necessary. We have generated a DNA vaccine based on the non-replicative antibiotic resistance marker-free plasmid vector pPAL that contains the encoding gene for the L. infantum activated protein kinase C receptor analog (LACK). Homologous pPAL-LACK prime-boost intranasal administration confers efficacious protection in Beagle dogs with a reduction of clinical signs and a statistically significant reduction of the parasite burden in the bone marrow of more than 90% of dogs after experimental infection with highly infective promastigotes. This DNA vaccine elicits a robust cellular immune response skewed towards the Th1 profile.

IL12 p35 and p40 subunit genes administered as pPAL plasmid constructs do not improve protection of pPAL-LACK vaccine against canine leishmaniasis.

Leishmania infantum causes zoonotic visceral leishmaniasis (ZVL) in the Mediterranean basin and South America. The parasite has been shown to co-infect HIV patients and an outbreak in central Spain was reported in the last decade. Therfore, ZVL is a public health problem, dogs being the parasite's reservoir. We have developed a DNA vaccine based on the L. infantum activated protein kinase A receptor (LACK) using different plasmid vectors and vaccinia virus strains as vehicles. Recently, we have generated an antibiotic resistance marker-free plasmid vector called pPAL. Homologous pPAL-LACK prime-boost vaccination protects Beagle dogs as well as a heterologous plasmid-virus regime. For both reasons, pPAL improves safety. IL12 was described to trigger Th1 response through IFN-γ production in infected dogs, being a good candidate for cytokine therapy in conventional treatment-unresponsive dogs. Herein, we report a complete protection study in dogs through inoculation of genes encoding for the p35 and p40 subunits which compose canine IL12 in combination with the LACK gene. A homologous plasmid-plasmid regime using independent pPAL constructs for each gene was inoculated in a 15-day interval. The infectious challenge using L. infantum promastigotes was successful. The outcome was pPAL-LACK vaccine protection suppression by IL12 administration. The important implications of this finding are discussed in the manuscript.