Proceeding of Veterinary and Animal Science Days 2018, 6th- 8th June, Milan, Italy HAF © 2013 Vol. V, No. 1s ISSN: 2283-3927 l Bovine besnoitiosis, caused by Besnoitia besnoiti, is a (re)emerging disease in Europe (Cortes et al.,2014), including Italy (Gazzonis et al.,2014;2017). However, its economic impact is scarcely considered and generally underestimated and there are still little studied aspects concerning both the parasite and the disease. Following a natural outbreak of besnoitiosis in a dairy herd, a study was planned to characterize B.besnoiti infection in cattle through a multidisciplinary approach. Suspicious abortions and clinical cases of besnoitiosis were reported in a dairy farm (September 2017, Northern Italy) housing 216 Holstein cattle. Blood samples were collected; haematological and serological analyses (ELISA and confirmatory WB) were performed (Fernandez-Garcia et al.,2009). Histology and molecular (endpoint ITS-1 PCR (Cortes et al.,2007) and sequencing) analyses of tissues from a slaughtered cow with chronic besnoitiosis were carried out. Out of 59 ELISA-positive animals, 50 (23%) were confirmed by WB. B. besnoiti prevalence was higher in cows (41%) than in calves (12%); no heifer resulted positive. Considering haematological parameters, a significant shift in the differential leucocyte formula from lymphocyte (L%) to granulocyte (G%) was recorded in infected cows (Mean±S.D.:L=46.1±18.4,G=53.9±18.4) if compared to negative animals (Student’s T-test,p=0.012). Histology revealed a high load of B.besnoiti tissue cysts in skin, vulva, muzzle, sclera, eyelid, respiratory tract, emphasizing the possibility of parasite transmission through direct contact among animals. B.besnoiti was confirmed by PCR in other organs (heart, liver, aorta wall, tonsil) and especially in ovary, uterus and vulva, suggesting that the infection could affect cows’ fertility. Parasite DNA was also found in masseters posing an important question for food security, even if B.besnoiti is not considered zoonotic (Figure 1). The study suggests that to investigate the dynamics of bovine besnoitiosis is mandatory to associate clinical and laboratory tests, including the genetic characterization of the parasite and its eventual correlation with the disease outcome. Ethical statement: all procedures for collection of biological specimens from live animals were accomplished following good clinical practices in the respect of animal welfare according to Keywords Bovine Besnoitiosis, Serology, Haematology, Molecular Biology, Histology. CORRESPONDING AUTHOR Luca Villa luca.villa@unimi.it JOURNAL HOME PAGE riviste.unimi.it/index.php/haf Investigating on Besnoitia besnoiti (Apicomplexa, Sarcocystidae) in naturally infected dairy cattle by an integrated approach. L. Villa 1,*, A.L. Gazzonis1, S. Mazzola1, S.A. Zanzani1, C. Perlotti1, G. Sironi1, M.T. Manfredi1 1 Department of Veterinary Medicine, Università degli Studi di Milano, Via dell’Università 6, 26900 Lodi, Italy. http://creativecommons.org/licenses/by-nc-nd/3.0/deed.en Proceeding of Veterinary and Animal Science Days 2018, 6th- 8th June, Milan, Italy 70 HAF © 2013 Vol. V, No. 1s ISSN: 2283-3927 current legislation. The study was conducted with the approval of Institutional Animal Care and Use Committee of Università degli Studi di Milano (Permission OPBA_34_2017). Conflict of interest statement: the authors declare they have no conflict of interest. References Cortes, H.C., Reis, Y., Gottstein, B., Hemphill, A., Leitao, A., Muller, N., 2007. Application of conventional and real-time fluorescent ITS1 rDNA PCR for detection of Besnoitia besnoiti infections in bovine skin biopsies. Veterinary Parasitology. 146, 352-356. Cortes, H.C., Leitao, A., Gottstein, B., Hemphill, A., 2014. A review on bovine besnoitiosis: a disease with economic impact in herd health management, caused by Besnoitia besnoiti. Parasitology. 141, 1406-1417. Fernandez-Garcia, A., Alvarez-Garcia, G., Risco-Castillo, V., Aguado-Martinez, A., Marugan-Hernandez, V., Ortega- Mora, L.M., 2009. Pattern of recognition of Besnoitia besnoiti tachyzoite and bradyzoite antigens by naturally infected cattle. Veterinary Parasitology. 164, 104-110. Gazzonis, A.L., Alvarez-Garcia, G., Zanzani, S.A., Garippa, G., Rossi, L., Maggiora, M., Dini, V., Invernizzi, A., Luini, M., Tranquillo, V.M., Ortega-Mora, L.M., Manfredi, M.T., 2014. Besnoitia besnoiti among cattle in insular and northwestern Italy: endemic infection or isolated outbreaks? Parasite and Vectors. 7, 585. Gazzonis, A.L., Alvarez-Garcia, G., Maggioni, A., Zanzani, S.A., Olivieri, E., Compiani, R., Sironi, G., Ortega-Mora, L.M., Manfredi, M.T., 2017. Serological dynamics and risk factors of Besnoitia besnoiti infection in breeding bulls from an endemically infected purebred beef herd. Parasitology Research. 116, 1383-1393. Figure 1: (A) Typical skin lesions in the chronic phase of besnoitiosis in a dairy cow. (B) Histological section of skin: hyperkeratosis with lymphohistiocytic and eosinophilic flogosis. (C) Pattern of recognition of Besnoitia besnoiti tachyzoite antigens by sera from naturally infected cattle by Western Blot. (D) Agarose gel-electrophoretic analysis of amplification products from conventional Besnoitia besnoiti ITS1 rDNA PCR on tissues from a slaughtered cow with chronic besnoitiosis http://creativecommons.org/licenses/by-nc-nd/3.0/deed.en