The conundrum of human visceral leishmaniasis in Emilia-Romagna, Italy: are wild and peridomestic animals potential reservoirs?

Leishmaniasis is a complex parasitic disease caused by intracellular protozoans of the genus Leishmania mainly transmitted by the bite of sand flies. In Italy, leishmaniasis is caused by Leishmania infantum, responsible for the human visceral and canine leishmaniases (HVL and CanL, respectively). Within Emilia-Romagna region, Italy, recent molecular studies indicated that L. infantum strains circulating in dogs and humans are different. This suggests that an animal reservoir other than dog should be evaluated in the epidemiology of HVL in Emilia-Romagna. Therefore, the main aim of this PhD project was to investigate the role of wild and peridomestic mammals as potential animal reservoirs of L. infantum in the regional zones where HVL foci are still active, also evaluating the possible role of arthropod vectors other than phlebotomine sandflies as vectors of Leishmania spp. in the sylvatic cycle of the protozoa. Overall, 206 specimens of different animal species (roe deer, rats, mice, badgers, hares, polecats, foxes, beech martens, bank voles, hedgehogs, and shrews), collected in Emilia-Romagna were screened for Leishmania with a real-time PCR, revealing a prevalence of 33% for roe deer (first report in this species). Positivity was also found in brown rats (10.6%), black rats (13.1%), mice (10%), badgers (25%), hedgehogs (80%) and bank voles (11%). To distinguish the two strains of L. infantum circulating in Emilia-Romagna, a nested PCR protocol optimized for animal tissues was developed, demonstrating that over 90% of L. infantum infections in roe deer were due to the strain isolated from humans and suggesting their possible role as reservoirs in the study area. Furthermore, the presence of Leishmania kDNA was detected in unfed larvae, nymphs and males of questing Ixodes ricinus ticks collected in regional parks of Emilia-Romagna suggesting their possible role in the transmission of L. infantum in a sylvatic or rural cycle.