Typing of Leishmania isolates from vectors and leporids of the Madrid (Spain) outbreak.

Abstract

In 2009, a large outbreak of leishmaniasis, associated with environmental changes, was declared near Madrid (Spain), in which <em>Phlebotomus perniciosus</em> was the vector, whereas the main reservoirs were hares and rabbits. Analysis of isolates from humans, vectors and leporids from the focus identified the <em>Leishmania infantum</em> ITS-Lombardi genotype. However, multilocus enzyme electrophoresis (MLEE), the reference technique for <em>Leishmania </em>typing, and sequencing of the <em>hsp70 </em>gene, a commonly used marker, were not performed. In the present study, 19 isolates from <em>P. perniciosus</em> (n=11), hares (n=5), and rabbits (n=3) from the outbreak area, all characterized as ITS-Lombardi in previous studies, were analysed by MLEE and <em>hsp70</em> sequencing. The <em>hsp70</em> results confirmed that all the analysed strains are <em>L. infantum</em>. However, by MLEE, four different zymodemes of <em>L. infantum</em> were identified based on variable mobilities of the NP₁ enzyme: MON-34 (NP₁100, n=11), MON-80 (NP₁130, n=6), MON-24 (NP₁140, n=1), and MON-331 (NP₁150 , n=1). The relative frequency of these zymodemes does not correspond to their usual occurrence in Spain. Moreover, MON-34 and MON-80 were found in <em>P. perniciosus</em>, hares, and rabbits for the first time. These findings continue to provide insights into the outbreak and call for further studies with a higher number of strains.