ORIGINAL PAPER Detection of Leishmania infantum and identification of blood meals in Phlebotomus perniciosus from a focus of human leishmaniasis in Madrid, Spain Maribel Jiménez & Estela González & Andrés Iriso & Elisa Marco & Ana Alegret & Fernando Fúster & Ricardo Molina Received: 5 February 2013 / Accepted: 15 March 2013 # Springer-Verlag Berlin Heidelberg 2013 Abstract Since 2010, the number of cases of both human visceral leishmaniasis and cutaneous leishmaniasis in south- western Madrid region (Spain) and more specifically in the town of Fuenlabrada has increased. Direct xenodiagnosis of leishmaniasis proved that hares (Lepus granatensis) from this focus are able to infect with Leishmania infantum colonized Phlebotomus perniciosus. To a better understanding of this focus of leishmaniasis, we conducted an entomological survey using CDC light traps, at the end of the seasonal transmission period of 2011 before the beginning of control measures of the disease, to study the phlebotomine sand flies species involved. Detection of Leishmania DNA in the sand flies captured was studied by kDNA-PCR and cpb-PCR. In addition, blood fed and gravid female P . perniciosus were analysed by a PCR based in vertebrate cytochrome b (cyt b) gene. Taxonomic identification of captured sand flies (n =174) as P . perniciosus (n =171) and Sergentomyia minuta (n =3) together with the analysis of blood feeding in ten sand flies that shows a high preference for hares (n =6), followed by humans (n =3), and cats ( n =1) confirm a strong association between P. perniciosus hares and humans in the focus. Moreover, 79 out of 135 (58.5 %) P . perniciosus were positive to L. infantum by PCR approaches. These data support the increase of human leishmaniasis cases in the area and the existence of an unusual sylvatic cycle alternative to the classical domestic one, where the dog is the main reservoir of L. infantum. Introduction In Spain, leishmaniasis is a zoonosis caused by Leishmania infantum and transmitted by phlebotomine sand flies belong- ing to the subgenus Larroussius (Diptera: Psychodidae, Phlebotominae); the dog being the main domestic reservoir (WHO 2010). Leishmaniasis is endemic in rural, peri-urban, and subur- ban areas of Madrid region (central Spain). Only two sand fly species Phlebotomus perniciosus and Phlebotomus ariasi out of the seven described in this region are proven vectors of the parasite L. infantum, being its main vector P . perniciosus (Conesa Gallego et al. 1999; Gálvez et al. 2010). Since 2010, a marked and unusual increase of human cases of both visceral and cutaneous leishmaniasis (VL and CL, respectively) is being observed in four municipalities of the southwestern Madrid region, where around 374 cases have been diagnosed until September 2012—of which 36.6 % are cases of visceral leishmaniasis—with an incidence rate of 21.54 cases/100,000 inhabitants. Only in the town of Fuenlabrada, 316 cases have been notified with an incidence rate of 52.99 cases/100,000 inhabitants (Suárez et al. 2012). The creation of a park adjacent to the urban area of the focus was expected to increase the hare population in this open space, and as a result of this, it has increased the population of the sand fly vectors. In fact, we have recently shown by direct M. Jiménez (*) : E. González : R. Molina (*) Unidad de Entomología Médica, Servicio de Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Ctra. Majadahonda-Pozuelo s/n, 28220 Majadahonda Madrid, Spain e-mail: mjimenez@isciii.es e-mail: rmolina@isciii.es A. Iriso : F. Fúster Subdirección General de Sanidad Ambiental y Epidemiología, Dirección General de Ordenación e Inspección, Consejería de Sanidad, Comunidad de Madrid, Madrid, Spain E. Marco : A. Alegret Concejalía de Salud, Consumo y Cooperación, Ayuntamiento de Fuenlabrada, Madrid, Spain Parasitol Res DOI 10.1007/s00436-013-3406-3