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Currently, no standardized molecular surveillance of circulating <jats:italic>Leishmania<\/jats:italic> strains is performed in European endemic areas, despite the potential public health implications of parasite biodiversity. This study aims to characterize parasite population haplogroups causing TL in two active endemic areas in southern Europe, i.e. Bologna (northern Italy) and Fuenlabrada (central Spain).<\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>Methods<\/jats:title>\n            <jats:p>In this retrospective study, we typed 87\u00a0<jats:italic>L. infantum<\/jats:italic>\u00a0samples from TL cases in the areas of Bologna and Fuenlabrada; these areas hosted the main European foci of human TL occurring in the last 15\u00a0years. Two <jats:italic>Leishmania<\/jats:italic> genomic typing targets were used: the heat shock protein 70 <jats:italic>(Hsp70<\/jats:italic>) and the cysteine peptidase b <jats:italic>(Cpb<\/jats:italic>). Simpson\u2019s index was used to calculate the discriminatory power of the used typing methods.<\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>Results<\/jats:title>\n            <jats:p>Typing results depicted the presence of a heterogeneous parasite population circulating in Bologna with two main haplogroups, i.e. <jats:italic>Hsp70<\/jats:italic>(A)_<jats:italic>Cpb<\/jats:italic>(F) (<jats:italic>n<\/jats:italic>\u2009=\u20097, 30.4%) and <jats:italic>Hsp70<\/jats:italic>(G)_<jats:italic>Cpb<\/jats:italic>(E\/F) (<jats:italic>n<\/jats:italic>\u2009=\u20097, 30.4%), differing from the reference <jats:italic>L. infantum<\/jats:italic> strain JPCM5 haplogroup and partially overlapping with <jats:italic>L. donovani<\/jats:italic> lineages. Among the samples from Fuenlabrada, <jats:italic>n<\/jats:italic>\u2009=\u200919 samples were typed by both targets, revealing a homogeneous population expressing <jats:italic>Hsp70<\/jats:italic>(A) and <jats:italic>Cpb<\/jats:italic>(E), matching the JPCM5 reference strain haplogroup. Overall, the <jats:italic>Cpb<\/jats:italic> typing method exhibited higher discrimination power as compared to the <jats:italic>Hsp70<\/jats:italic> method (Simpson\u2019s index of diversity,<jats:italic> P<\/jats:italic>-value\u2009&lt;\u20090.05).<\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>Conclusions<\/jats:title>\n            <jats:p>Our findings show differences among <jats:italic>L. infantum<\/jats:italic> populations causing TL in two southern European epidemiological foci of human leishmaniasis and support the recent discovery of <jats:italic>L. infantum\/L.donovani<\/jats:italic> hybrid strains circulating in northern Italy. These results underscore the critical need to identify the circulating <jats:italic>Leishmania<\/jats:italic> strains in endemic areas and assess their potential public health implications in active foci.<\/jats:p>\n          <\/jats:sec>\n          <jats:sec>\n            <jats:title>Graphical Abstract<\/jats:title>\n          <\/jats:sec>","DOI":"10.1186\/s40249-025-01309-5","type":"journal-article","created":{"date-parts":[[2025,5,26]],"date-time":"2025-05-26T03:12:56Z","timestamp":1748229176000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["Combination of Cpb-Hsp70 typing methods reveals genetic divergence between Leishmania infantum strains causing human tegumentary leishmaniasis in northern Italy and central Spain: a retrospective study"],"prefix":"10.1186","volume":"14","author":[{"given":"Tommaso","family":"Gritti","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Carmen","family":"Chicharro","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Eugenia","family":"Carrillo","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jose Carlos","family":"Solana","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Javier","family":"Moreno","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Elena","family":"Carra","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Margherita","family":"Ortalli","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sara","family":"Morselli","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Valeria","family":"Gaspari","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Margherita","family":"Zanazzi","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tiziana","family":"Lazzarotto","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gianluca","family":"Rugna","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0862-4937","authenticated-orcid":false,"given":"Stefania","family":"Varani","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2025,5,26]]},"reference":[{"issue":"3","key":"1309_CR1","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pntd.0004349","volume":"10","author":"M Akhoundi","year":"2016","unstructured":"Akhoundi M, Kuhls K, Cannet A, Vot\u00fdpka J, Marty P, Delaunay P, et al. 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