{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,7]],"date-time":"2026-07-07T10:18:16Z","timestamp":1783419496391,"version":"3.54.6"},"reference-count":50,"publisher":"MDPI AG","issue":"7","license":[{"start":{"date-parts":[[2026,7,7]],"date-time":"2026-07-07T00:00:00Z","timestamp":1783382400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Xinjiang Uygur Autonomous Region Frontier Science and Technology Support Program for Talent","award":["XJRC-2025-KJ-KJQY-002"],"award-info":[{"award-number":["XJRC-2025-KJ-KJQY-002"]}]},{"name":"Xinjiang Uygur Autonomous Region Natural Science Foundation Youth Fund","award":["2025D01B43"],"award-info":[{"award-number":["2025D01B43"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Veterinary Sciences"],"abstract":"<jats:p>Rhodococcus equi (R. equi) primarily induces fatal pulmonary and extrapulmonary pyogenic granulomatous infections in foals, imposing substantial economic burdens on the equine industry. The emergence and spread of multidrug-resistant (MDR) R. equi have led to a therapeutic impasse in clinical settings. Although vaccination is a proven strategy against MDR pathogens, no commercial vaccine is currently available for R. equi. In this study, we employed a bioinformatics approach to systematically identify and prioritize antigenic epitopes derived from R. equi for multi-epitope vaccine design. Using ABCPred, NetMHCpan EL, and IEDB servers, 27 MHC-I and 9 MHC-II epitopes were selected from five previously validated R. equi vaccine candidates: ABC transporter, PBD2, NlpC\/P60, Esterase, and M23. These epitopes were coupled with distinct peptide linkers to construct six multi-epitope vaccine constructs, designated V1\u2013V6. The physicochemical properties, antigenicity, immunogenicity, and toxicity of the six vaccine constructs were analyzed, and the V3 and V4 constructs were ultimately selected. Using the HDOCK and Gromacs tools, the intermolecular interactions, binding affinity, and thermal stability of the V3 and V4 constructs with the equine MHC molecules EQCA-I and EQCA-II were evaluated. The results confirm that V3 and V4 exhibit strong binding affinity to EQCA-I and EQCA-II, with stable conformations following binding, indicating theoretical potential to induce humoral and cellular immunity in foals. Recombinant plasmids for V3 and V4 were constructed, and the V3 and V4 proteins were successfully prepared, confirming the feasibility of prokaryotic expression for these vaccine constructs. Immunization assays in SPF BALB\/c mice showed that the multi-epitope vaccines elicited robust antigen-specific IgG antibody responses, reflecting preliminary humoral immunogenicity. However, these murine data have translational limitations, as they cannot fully represent equine immune responses. The findings establish a crucial theoretical foundation for the advancement of vaccines targeting R. equi while offering a reference for the design of vaccines against other drug-resistant microbial pathogens.<\/jats:p>","DOI":"10.3390\/vetsci13070655","type":"journal-article","created":{"date-parts":[[2026,7,7]],"date-time":"2026-07-07T08:04:49Z","timestamp":1783411489000},"page":"655","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Designing a Multi-Epitope Vaccine Candidate Against Rhodococcus equi Based on the Bioinformatics Technique"],"prefix":"10.3390","volume":"13","author":[{"given":"Shiwen","family":"Gao","sequence":"first","affiliation":[{"name":"Xinjiang Key Laboratory of New Drug Research and Development for Herbivores, College of Veterinary Medicine, Xinjiang Agricultural University, Urumqi 830052, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Guoqing","family":"Li","sequence":"additional","affiliation":[{"name":"Xinjiang Key Laboratory of New Drug Research and Development for Herbivores, College of Veterinary Medicine, Xinjiang Agricultural University, Urumqi 830052, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiangyu","family":"Wang","sequence":"additional","affiliation":[{"name":"Xinjiang Key Laboratory of New Drug Research and Development for Herbivores, College of Veterinary Medicine, Xinjiang Agricultural University, Urumqi 830052, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Weifang","family":"Gu","sequence":"additional","affiliation":[{"name":"Xinjiang Key Laboratory of New Drug Research and Development for Herbivores, College of Veterinary Medicine, Xinjiang Agricultural University, Urumqi 830052, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dingnuoya","family":"Guo","sequence":"additional","affiliation":[{"name":"Xinjiang Key Laboratory of New Drug Research and Development for Herbivores, College of Veterinary Medicine, Xinjiang Agricultural University, Urumqi 830052, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zongping","family":"Xian","sequence":"additional","affiliation":[{"name":"Xinjiang Key Laboratory of New Drug Research and Development for Herbivores, College of Veterinary Medicine, Xinjiang Agricultural University, Urumqi 830052, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7336-2211","authenticated-orcid":false,"given":"Xuelian","family":"Ma","sequence":"additional","affiliation":[{"name":"Xinjiang Key Laboratory of New Drug Research and Development for Herbivores, College of Veterinary Medicine, Xinjiang Agricultural University, Urumqi 830052, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0004-4211-0870","authenticated-orcid":false,"given":"Jun","family":"Meng","sequence":"additional","affiliation":[{"name":"College of Animal Science, Xinjiang Agricultural University, Urumqi 830052, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8268-5044","authenticated-orcid":false,"given":"Hongqiong","family":"Zhao","sequence":"additional","affiliation":[{"name":"Xinjiang Key Laboratory of New Drug Research and Development for Herbivores, College of Veterinary Medicine, Xinjiang Agricultural University, Urumqi 830052, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0009-0994-7130","authenticated-orcid":false,"given":"Lu","family":"Liu","sequence":"additional","affiliation":[{"name":"Xinjiang Key Laboratory of New Drug Research and Development for Herbivores, College of Veterinary Medicine, Xinjiang Agricultural University, Urumqi 830052, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,7,7]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"9","DOI":"10.1016\/j.vetmic.2013.06.016","article-title":"Rhodococcus equi: The many facets of a pathogenic actinomycete","volume":"167","author":"Hapeshi","year":"2013","journal-title":"Vet. Microbiol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"2887","DOI":"10.1128\/jcm.29.12.2887-2889.1991","article-title":"Prevalence of virulent Rhodococcus equi in isolates from soil and feces of horses from horse-breeding farms with and without endemic infections","volume":"29","author":"Takai","year":"1991","journal-title":"J. Clin. Microbiol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"20","DOI":"10.1016\/j.tvjl.2011.08.014","article-title":"Rhodococcus equi pneumonia in the foal\u2014Part 1: Pathogenesis and epidemiology","volume":"192","author":"Muscatello","year":"2012","journal-title":"Vet. J."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"4908","DOI":"10.1177\/00031348211041558","article-title":"Post-Operative Wound Infection by Multiple Zoonotic Organisms","volume":"89","author":"Albert","year":"2023","journal-title":"Am. Surg."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"310","DOI":"10.1016\/j.cmi.2018.04.033","article-title":"Diagnosis and management of pulmonary infection due to Rhodococcus equi","volume":"25","author":"Lin","year":"2019","journal-title":"Clin. Microbiol. Infect."},{"key":"#cr-split#-ref_6.1","unstructured":"EFSA Panel on Animal Health and Welfare (AHAW), Nielsen, S.S., Bicout, D.J., Calistri, P., Canali, E., Drewe, J.A., Garin-Bastuji, B., Rojas, J.L.G., Gort\u00e1zar, C., and Herskin, M. (2022). Assessment of listing and categorisation of animal diseases within the framework of the Animal Health Law (Regulation"},{"key":"#cr-split#-ref_6.2","unstructured":"(EU) No 2016\/429): Antimicrobial-resistant Rhodococcus equi in horses. EFSA J., 20, e07081."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"590","DOI":"10.1111\/jvim.16002","article-title":"Opsonization but not pretreatment of equine macrophages with hyperimmune plasma nonspecifically enhances phagocytosis and intracellular killing of Rhodococcus equi","volume":"35","author":"Harvey","year":"2021","journal-title":"J. Vet. Intern. Med."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"5760","DOI":"10.1016\/j.vaccine.2005.07.050","article-title":"Virulence-associated protein-specific serum immunoglobulin G-isotype expression in young foals protected against Rhodococcus equi pneumonia by oral immunization with virulent R. equi","volume":"23","author":"Wilkie","year":"2005","journal-title":"Vaccine"},{"key":"ref_9","first-page":"444","article-title":"Protection of foals against experimental Rhodococcus equi pneumonia by oral immunization","volume":"51","author":"Prescott","year":"1987","journal-title":"Can. J. Vet. Res. Rev. Can. Rech. Vet."},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Bordin, A.I., Pillai, S.D., Brake, C., Bagley, K.B., Bourquin, J.R., Coleman, M., Oliveira, F.N., Mwangi, W., McMurray, D.N., and Love, C.C. (2014). Immunogenicity of an electron beam inactivated Rhodococcus equi vaccine in neonatal foals. PLoS ONE, 9.","DOI":"10.1371\/journal.pone.0105367"},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Rocha, J.N., Cohen, N.D., Bordin, A.I., Brake, C.N., Gigu\u00e8re, S., Coleman, M.C., Alaniz, R.C., Lawhon, S.D., Mwangi, W., and Pillai, S.D. (2016). Oral Administration of Electron-Beam Inactivated Rhodococcus equi Failed to Protect Foals against Intrabronchial Infection with Live, Virulent R. equi. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0148111"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"215","DOI":"10.1016\/j.vetimm.2004.12.006","article-title":"Assessment in mice of vapA-DNA vaccination against Rhodococcus equi infection","volume":"104","author":"Haghighi","year":"2005","journal-title":"Vet. Immunol. Immunopathol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"2457","DOI":"10.1016\/j.vaccine.2008.03.015","article-title":"Chimeric vapA\/groEL2 DNA vaccines enhance clearance of Rhodococcus equi in aerosol challenged C3H\/He mice","volume":"26","author":"Phumoonna","year":"2008","journal-title":"Vaccine"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"437","DOI":"10.1016\/j.ijmm.2004.09.011","article-title":"Immune response to vaccines based upon the VapA protein of the horse pathogen, Rhodococcus equi, in a murine model","volume":"294","author":"Vanniasinkam","year":"2005","journal-title":"Int. J. Med. Microbiol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"530","DOI":"10.1016\/j.immuni.2010.09.017","article-title":"Reverse vaccinology: Developing vaccines in the era of genomics","volume":"33","author":"Sette","year":"2010","journal-title":"Immunity"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"404","DOI":"10.1038\/nrd2224","article-title":"More than one reason to rethink the use of peptides in vaccine design","volume":"6","author":"Purcell","year":"2007","journal-title":"Nat. Rev. Drug Discov."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"1999","DOI":"10.1007\/s11095-023-03540-x","article-title":"Validation of Multi-epitope Peptides Encapsulated in PLGA Nanoparticles Against Influenza A Virus","volume":"40","author":"Heng","year":"2023","journal-title":"Pharm. Res."},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Kaushik, V., G, S.K., Gupta, L.R., Kalra, U., Shaikh, A.R., Cavallo, L., and Chawla, M. (2022). Immunoinformatics Aided Design and In-Vivo Validation of a Cross-Reactive Peptide Based Multi-Epitope Vaccine Targeting Multiple Serotypes of Dengue Virus. Front. Immunol., 13.","DOI":"10.3389\/fimmu.2022.865180"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"w20052","DOI":"10.4414\/smw.2019.20052","article-title":"Design and evaluation of a multi-epitope assembly peptide vaccine against Acinetobacter baumannii infection in mice","volume":"149","author":"Ren","year":"2019","journal-title":"Swiss Med. Wkly."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Liu, L., Cai, P., Gu, W., Duan, X., Gao, S., Ma, X., Ma, Y., Ma, S., Li, G., and Wang, X. (2024). Evaluation of vaccine candidates against Rhodococcus equi in BALB\/c mice infection model: Cellular and humoral immune responses. BMC Microbiol., 24.","DOI":"10.1186\/s12866-024-03408-z"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"e18623","DOI":"10.1016\/j.heliyon.2023.e18623","article-title":"Identification of vaccine candidates against Rhodococcus equi by combining pangenome analysis with a reverse vaccinology approach","volume":"9","author":"Liu","year":"2023","journal-title":"Heliyon"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"467","DOI":"10.1007\/s11030-021-10292-6","article-title":"ABCpred: A webserver for the discovery of acetyl- and butyryl-cholinesterase inhibitors","volume":"26","author":"Malik","year":"2022","journal-title":"Mol. Divers."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"177","DOI":"10.1007\/s00251-011-0579-8","article-title":"NetMHCcons: A consensus method for the major histocompatibility complex class I predictions","volume":"64","author":"Karosiene","year":"2012","journal-title":"Immunogenetics"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"304","DOI":"10.1007\/s00251-005-0798-y","article-title":"Automated generation and evaluation of specific MHC binding predictive tools: ARB matrix applications","volume":"57","author":"Bui","year":"2005","journal-title":"Immunogenetics"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"251","DOI":"10.1038\/s41598-021-03988-0","article-title":"Identification of vaccine and drug targets in shigella dysenteriae sd197 using reverse vaccinology approach","volume":"12","author":"Jalal","year":"2022","journal-title":"Sci. Rep."},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Ghaffari-Nazari, H., Tavakkol-Afshari, J., Jaafari, M.R., Tahaghoghi-Hajghorbani, S., Masoumi, E., and Jalali, S.A. (2015). Improving Multi-Epitope Long Peptide Vaccine Potency by Using a Strategy that Enhances CD4+ T Help in BALB\/c Mice. PLoS ONE, 10.","DOI":"10.1371\/journal.pone.0142563"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"bbaa294","DOI":"10.1093\/bib\/bbaa294","article-title":"AlgPred 2.0: An improved method for predicting allergenic proteins and mapping of IgE epitopes","volume":"22","author":"Sharma","year":"2021","journal-title":"Brief. Bioinform."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"3098","DOI":"10.1093\/bioinformatics\/btx345","article-title":"Protein-sol: A web tool for predicting protein solubility from sequence","volume":"33","author":"Hebditch","year":"2017","journal-title":"Bioinformatics"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"bbac174","DOI":"10.1093\/bib\/bbac174","article-title":"ToxinPred2: An improved method for predicting toxicity of proteins","volume":"23","author":"Sharma","year":"2022","journal-title":"Brief. Bioinform."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2278","DOI":"10.1007\/s00894-014-2278-5","article-title":"AllerTOP v.2\u2014A server for in silico prediction of allergens","volume":"20","author":"Dimitrov","year":"2014","journal-title":"J. Mol. Model."},{"key":"ref_31","first-page":"531","article-title":"Protein identification and analysis tools in the ExPASy server","volume":"112","author":"Wilkins","year":"1999","journal-title":"Methods Mol. Biol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"404","DOI":"10.1093\/bioinformatics\/16.4.404","article-title":"The PSIPRED protein structure prediction server","volume":"16","author":"McGuffin","year":"2000","journal-title":"Bioinformatics"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"5.8.1","DOI":"10.1002\/0471250953.bi0508s52","article-title":"Protein structure and function prediction using I-TASSER","volume":"52","author":"Yang","year":"2015","journal-title":"Curr. Protoc. Bioinform."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"W294","DOI":"10.1093\/nar\/gks493","article-title":"GalaxyWEB server for protein structure prediction and refinement","volume":"40","author":"Ko","year":"2012","journal-title":"Nucleic Acids Res."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"121","DOI":"10.1006\/jmbi.1996.0628","article-title":"Deviations from standard atomic volumes as a quality measure for protein crystal structures","volume":"264","author":"Pontius","year":"1996","journal-title":"J. Mol. Biol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"W407","DOI":"10.1093\/nar\/gkm290","article-title":"ProSA-web: Interactive web service for the recognition of errors in three-dimensional structures of proteins","volume":"35","author":"Wiederstein","year":"2007","journal-title":"Nucleic Acids Res."},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Ponomarenko, J., Bui, H.-H., Li, W., Fusseder, N., Bourne, P.E., Sette, A., and Peters, B. (2008). ElliPro: A new structure-based tool for the prediction of antibody epitopes. BMC Bioinform., 9.","DOI":"10.1186\/1471-2105-9-514"},{"key":"ref_38","doi-asserted-by":"crossref","unstructured":"Zhai, W., Li, X., Duan, X., Gou, C., Wang, L., and Gao, Y. (2022). Development of a microbial protease for composting swine carcasses, optimization of its production and elucidation of its catalytic hydrolysis mechanism. BMC Biotechnol., 22.","DOI":"10.1186\/s12896-022-00768-0"},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Huang, S., Zhang, C., Li, J., Dai, Z., Huang, J., Deng, F., Wang, X., Yue, X., Hu, X., and Li, Y. (2022). Designing a multi-epitope vaccine against coxsackievirus B based on immunoinformatics approaches. Front. Immunol., 13.","DOI":"10.3389\/fimmu.2022.933594"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"479","DOI":"10.1016\/j.vetimm.2011.10.011","article-title":"Development of a live, attenuated, potential vaccine strain of R. equi expressing vapA and the virR operon, and virulence assessment in the mouse","volume":"145","author":"Whitehead","year":"2012","journal-title":"Vet. Immunol. Immunopathol."},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"Trevisani, M.M., Hanna, E.S., Oliveira, A.F., Cardoso, S.A., Roque-Barreira, M.C., and Soares, S.G. (2017). Vaccination of mice with virulence-associated protein G (VapG) antigen confers partial protection against Rhodococcus equi infection through induced humoral immunity. Front. Microbiol., 8.","DOI":"10.3389\/fmicb.2017.00857"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"73","DOI":"10.1016\/j.vetmic.2004.09.006","article-title":"Foal IgG and opsonizing anti-Rhodococcus equi antibodies after immunization of pregnant mares with a protective VapA candidate vaccine","volume":"104","author":"Cauchard","year":"2004","journal-title":"Vet. Microbiol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"164","DOI":"10.1016\/j.vetimm.2013.12.003","article-title":"Assessment of the safety and immunogenicity of Rhodococcus equi-secreted proteins combined with either a liquid nanoparticle (IMS 3012) or a polymeric (PET GEL a) water-based adjuvant in adult horses and foals\u2014Identification of promising new candidate antigens","volume":"157","author":"Cauchard","year":"2014","journal-title":"Vet. Immunol. Immunopathol."},{"key":"ref_44","doi-asserted-by":"crossref","unstructured":"Chedid, C., Kokhreidze, E., Tukvadze, N., Banu, S., Uddin, M.K.M., Biswas, S., Russomando, G., Acosta, C.C.D., Arenas, R., and Ranaivomanana, P.P. (2020). Relevance of QuantiFERON-TB gold plus and heparin-binding hemagglutinin interferon-\u03b3 release assays for monitoring of pulmonary tuberculosis clearance: A multicentered study. Front. Immunol., 11.","DOI":"10.3389\/fimmu.2020.616450"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"1631","DOI":"10.1111\/febs.14067","article-title":"Ribosomal protein L7\/L12 is required for GTPase translation factors EF-G, RF3, and IF2 to bind in their GTP state to 70S ribosomes","volume":"284","author":"Carlson","year":"2017","journal-title":"FEBS J."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"23","DOI":"10.1016\/bs.mie.2020.12.004","article-title":"Design and construction of chimeric linker library with controllable flexibilities for precision protein engineering","volume":"647","author":"Huang","year":"2021","journal-title":"Methods Enzymol."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"bbaf055","DOI":"10.1093\/bib\/bbaf055","article-title":"Advances of computational methods enhance the development of multi-epitope vaccines","volume":"26","author":"Wei","year":"2024","journal-title":"Brief. Bioinform."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"e36154","DOI":"10.1016\/j.heliyon.2024.e36154","article-title":"Rational design and computational evaluation of a multi-epitope vaccine for monkeypox virus: Insights into binding stability and immunological memory","volume":"10","author":"Kaur","year":"2024","journal-title":"Heliyon"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"189","DOI":"10.1038\/nm.2285","article-title":"A multistage tuberculosis vaccine that confers efficient protection before and after exposure","volume":"17","author":"Aagaard","year":"2011","journal-title":"Nat. Med."}],"container-title":["Veterinary Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2306-7381\/13\/7\/655\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,7,7]],"date-time":"2026-07-07T10:03:10Z","timestamp":1783418590000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2306-7381\/13\/7\/655"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,7,7]]},"references-count":50,"journal-issue":{"issue":"7","published-online":{"date-parts":[[2026,7]]}},"alternative-id":["vetsci13070655"],"URL":"https:\/\/doi.org\/10.3390\/vetsci13070655","relation":{},"ISSN":["2306-7381"],"issn-type":[{"value":"2306-7381","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,7,7]]}}}