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Despite numerous reports of human dirofilariosis in Ukraine, epidemiological data on these filarial parasites in companion animals remain limited. The aim of the study was to conduct a molecular epidemiological survey to assess the prevalence of filarial infections in stray dogs and cats across Ukraine and evaluate factors associated with infection in dogs. In collaboration with the European Scientific Counsel Companion Animal Parasites (ESCCAP) and local non-governmental organizations (NGOs), a total of 457 blood samples (233 dogs and 224 cats) were collected between March and December 2023 from Berdychiv, Lviv, Kharkiv, Sumy and Zvenyhorodka. Molecular detection of\n                      <jats:italic>D. repens<\/jats:italic>\n                      ,\n                      <jats:italic>D. immitis<\/jats:italic>\n                      , and\n                      <jats:italic>A. reconditum<\/jats:italic>\n                      was performed using a two-step quantitative PCR (qPCR) assay with novel species-specific primers.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      The method demonstrated high sensitivity and specificity, capable of detecting DNA from a single microfilaria, with no evidence of cross-reactivity among target species. Among the canine samples, 66 (28.3%) tested positive for at least one filarial species, including cases of both mono- and co-infection. In contrast, only 8 feline samples (3.6%) were positive for\n                      <jats:italic>D. repens<\/jats:italic>\n                      or\n                      <jats:italic>D. immitis<\/jats:italic>\n                      . Statistical analysis revealed a higher prevalence among male dogs and those weighing over 10\u00a0kg, while the lowest prevalence was observed in the youngest age group (&lt;\u20093 years).\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>\n                      These findings provide the first molecular evidence of\n                      <jats:italic>Dirofilaria<\/jats:italic>\n                      spp. and\n                      <jats:italic>A. reconditum<\/jats:italic>\n                      infections in stray animals from several under-studied areas of Ukraine, highlighting the relevance of the One Health approach in mitigating the risk of zoonotic transmission.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.1186\/s12917-025-04867-w","type":"journal-article","created":{"date-parts":[[2025,7,4]],"date-time":"2025-07-04T21:13:53Z","timestamp":1751663633000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["Molecular investigation of Dirofilaria repens, Dirofilaria immitis and Acanthocheilonema reconditum in stray dogs and cats in Ukraine"],"prefix":"10.1186","volume":"21","author":[{"given":"Mateusz","family":"P\u0119kacz","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kateryna","family":"Slivinska","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alla","family":"Vyniarska","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Katarzyna","family":"Basa\u0142aj","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alicja","family":"Kalinowska","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Agnieszka","family":"Weso\u0142owska","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alicja","family":"Laskowska","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Olesia","family":"Kysterna","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andrii","family":"Klietsov","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Martina","family":"Miterp\u00e1kov\u00e1","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andrei Daniel","family":"Mihalca","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jakub","family":"Gawor","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Vitaliy","family":"Kharchenko","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anna","family":"Zawistowska-Deniziak","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2025,7,5]]},"reference":[{"key":"4867_CR1","doi-asserted-by":"publisher","first-page":"24","DOI":"10.1016\/j.vetpar.2015.04.020","volume":"213","author":"D Otranto","year":"2015","unstructured":"Otranto D, Cantacessi C, Dantas-Torres F, Brianti E, Pfeffer M, Genchi C, et al. 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J Pharmacol Pharmacother. 2010;1:94\u20139.","journal-title":"J Pharmacol Pharmacother"}],"container-title":["BMC Veterinary Research"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1186\/s12917-025-04867-w.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/article\/10.1186\/s12917-025-04867-w\/fulltext.html","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1186\/s12917-025-04867-w.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,7,4]],"date-time":"2025-07-04T21:14:02Z","timestamp":1751663642000},"score":1,"resource":{"primary":{"URL":"https:\/\/bmcvetres.biomedcentral.com\/articles\/10.1186\/s12917-025-04867-w"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,7,5]]},"references-count":47,"journal-issue":{"issue":"1","published-online":{"date-parts":[[2025,12]]}},"alternative-id":["4867"],"URL":"https:\/\/doi.org\/10.1186\/s12917-025-04867-w","relation":{},"ISSN":["1746-6148"],"issn-type":[{"value":"1746-6148","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,7,5]]},"assertion":[{"value":"3 April 2025","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"9 June 2025","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"5 July 2025","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Declarations"}},{"value":"This research was carried out in collaboration with ESCCAP and several Ukrainian NGOs, as part of a comprehensive national initiative targeting the sterilization and vaccination of stray animals, which also includes parasitological examinations. All analyses were performed on residual blood samples collected during routine checkups and diagnostic procedures. As per the Resolution on the Protection of Animals Used for Scientific or Educational Purposes (15 January 2015) [\n                      \n                      ], no ethical approval or license was required for this study. Due to the absence of formal owners, written informed consent was not obtained. However, caregivers of the animals and\/or the directors of local shelters were informed about the results of\n                      Dirofilaria\n                      \/\n                      Acanthocheilonema\n                      testing. Additionally, veterinarians were provided with comprehensive information to facilitate the implementation of appropriate treatment for infected dogs. The manuscript adheres to the recommendations in the ARRIVE guidelines [\n                      \n                      ].","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Ethics approval and consent to participate"}},{"value":"Not applicable.","order":3,"name":"Ethics","group":{"name":"EthicsHeading","label":"Consent for publication"}},{"value":"The authors declare no competing interests.","order":4,"name":"Ethics","group":{"name":"EthicsHeading","label":"Competing interests"}}],"article-number":"438"}}