{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,25]],"date-time":"2026-03-25T12:48:41Z","timestamp":1774442921794,"version":"3.50.1"},"reference-count":61,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2021,4,6]],"date-time":"2021-04-06T00:00:00Z","timestamp":1617667200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Vet. Sci."],"abstract":"<jats:p>Consumption of contaminated poultry products is one of the main sources of human campylobacteriosis, of which <jats:italic>Campylobacter jejuni<\/jats:italic> subsp. <jats:italic>jejuni<\/jats:italic> (<jats:italic>C. jejuni<\/jats:italic>) and <jats:italic>C. coli<\/jats:italic> are responsible for ~98% of the cases. In turkeys, the ceca are an important anatomical site where <jats:italic>Campylobacter<\/jats:italic> asymptomatically colonizes. We previously demonstrated that commercial turkey poults colonized by <jats:italic>C. jejuni<\/jats:italic> showed acute changes in cytokine gene expression profiles, and histological intestinal lesions at 2 days post-inoculation (dpi). Cecal tonsils (CT) are an important part of the gastrointestinal-associated lymphoid tissue that surveil material passing in and out of the ceca, and generate immune responses against intestinal pathogens. The CT immune response toward <jats:italic>Campylobacter<\/jats:italic> remains unknown. In this study, we generated a kanamycin-resistant <jats:italic>C. coli<\/jats:italic> construct (CcK) to facilitate its enumeration from cecal contents after experimental challenge. <jats:italic>In vitro<\/jats:italic> analysis of CcK demonstrated no changes in motility when compared to the parent isolate. Poults were inoculated by oral gavage with CcK (5 \u00d7 10<jats:sup>7<\/jats:sup> colony forming units) or sterile-media (mock-colonized), and euthanized at 1 and 3 dpi. At both time points, CcK was recovered from cecal contents, but not from the mock-colonized group. As a marker of acute inflammation, serum alpha-1 acid glycoprotein was significantly elevated at 3 dpi in CcK inoculated poults compared to mock-infected samples. Significant histological lesions were detected in cecal and CT tissues of CcK colonized poults at 1 and 3 dpi, respectively. RNAseq analysis identified 250 differentially expressed genes (DEG) in CT from CcK colonized poults at 3 dpi, of which 194 were upregulated and 56 were downregulated. From the DEG, 9 significantly enriched biological pathways were identified, including platelet aggregation, response to oxidative stress and negative regulation of oxidative stress-induced intrinsic apoptotic signaling pathway. These data suggest that <jats:italic>C. coli<\/jats:italic> induced an acute inflammatory response in the intestinal tract of poults, and that platelet aggregation and oxidative stress in the CT may affect the turkey's ability to resist <jats:italic>Campylobacter<\/jats:italic> colonization. These findings will help to develop and test <jats:italic>Campylobacter<\/jats:italic> mitigation strategies to promote food safety in commercial turkeys.<\/jats:p>","DOI":"10.3389\/fvets.2021.613203","type":"journal-article","created":{"date-parts":[[2021,4,6]],"date-time":"2021-04-06T05:00:21Z","timestamp":1617685221000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":4,"title":["The Acute Host-Response of Turkeys Colonized With Campylobacter coli"],"prefix":"10.3389","volume":"8","author":[{"given":"Matthew J.","family":"Sylte","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sathesh K.","family":"Sivasankaran","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Julian","family":"Trachsel","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yuko","family":"Sato","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Zuowei","family":"Wu","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Timothy A.","family":"Johnson","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Lawrance C.","family":"Chandra","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Qijing","family":"Zhang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Torey","family":"Looft","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1965","published-online":{"date-parts":[[2021,4,6]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","first-page":"31","DOI":"10.1016\/S0168-1605(02)00232-5","article-title":"Emerging foodborne pathogens","volume":"78","author":"Tauxe","year":"2002","journal-title":"Int J Food Microbiol."},{"key":"B2","doi-asserted-by":"publisher","first-page":"6292","DOI":"10.3390\/ijerph10126292","article-title":"Foodborne Campylobacter: infections, metabolism, pathogenesis and reservoirs","volume":"10","author":"Epps","year":"2013","journal-title":"Int J Environ Res Public Health."},{"key":"B3","doi-asserted-by":"publisher","first-page":"351","DOI":"10.1051\/vetres:2005012","article-title":"Campylobacter","volume":"36","author":"Moore","year":"2005","journal-title":"Vet Res"},{"key":"B4","doi-asserted-by":"publisher","first-page":"2365","DOI":"10.1128\/AEM.54.10.2365-2370.1988","article-title":"Colonization of gastrointestinal tracts of chicks by Campylobacter jejuni","volume":"54","author":"Beery","year":"1988","journal-title":"Appl Environ Microbiol"},{"key":"B5","doi-asserted-by":"publisher","first-page":"1689","DOI":"10.3382\/ps\/pex384","article-title":"Evaluation of different Campylobacter jejuni isolates to colonize the intestinal tract of commercial turkey poults and selective media for enumeration","volume":"97","author":"Sylte","year":"2018","journal-title":"Poult Sci."},{"key":"B6","doi-asserted-by":"publisher","first-page":"6","DOI":"10.1016\/j.vetimm.2019.02.003","article-title":"Intestinal colonization and acute immune response in commercial turkeys following inoculation with Campylobacter jejuni constructs encoding antibiotic-resistance markers","volume":"210","author":"Sylte","year":"2019","journal-title":"Vet Immunol Immunopathol."},{"key":"B7","doi-asserted-by":"publisher","first-page":"114","DOI":"10.1111\/j.1574-695X.2008.00458.x","article-title":"Campylobacter colonization of the chicken induces a proinflammatory response in mucosal tissues","volume":"54","author":"Smith","year":"2008","journal-title":"FEMS Immunol Med Microbiol."},{"key":"B8","doi-asserted-by":"publisher","first-page":"191","DOI":"10.1016\/j.vetimm.2009.06.007","article-title":"Innate immune gene expression differentiates the early avian intestinal response between Salmonella and Campylobacter","volume":"132","author":"Shaughnessy","year":"2009","journal-title":"Vet Immunol Immunopathol"},{"key":"B9","doi-asserted-by":"publisher","first-page":"e11827","DOI":"10.1371\/journal.pone.0011827","article-title":"Gene expression profiling of the local cecal response of genetic chicken lines that differ in their susceptibility to Campylobacter jejuni colonization","volume":"5","author":"Li","year":"2010","journal-title":"PLoS ONE."},{"key":"B10","doi-asserted-by":"publisher","first-page":"491","DOI":"10.1111\/j.1365-2052.2010.02168.x","article-title":"Caecal transcriptome analysis of colonized and non-colonized chickens within two genetic lines that differ in caecal colonization by Campylobacter jejuni","volume":"42","author":"Li","year":"2011","journal-title":"Anim Genet"},{"key":"B11","doi-asserted-by":"publisher","first-page":"150541","DOI":"10.1098\/rsos.150541","article-title":"Cytokine responses in birds challenged with the human food-borne pathogen Campylobacter jejuni implies a Th17 response","volume":"3","author":"Reid","year":"2016","journal-title":"R Soc Open Sci."},{"key":"B12","doi-asserted-by":"publisher","first-page":"352","DOI":"10.1080\/03079457.2018.1475724","article-title":"Re-thinking the chicken-Campylobacter jejuni interaction: a review","volume":"47","author":"Awad","year":"2018","journal-title":"Avian Pathol"},{"key":"B13","doi-asserted-by":"publisher","first-page":"88","DOI":"10.1186\/s40168-018-0477-5","article-title":"The effect of the timing of exposure to Campylobacter jejuni on the gut microbiome and inflammatory responses of broiler chickens","volume":"6","author":"Connerton","year":"2018","journal-title":"Microbiome."},{"key":"B14","doi-asserted-by":"publisher","first-page":"161","DOI":"10.1113\/expphysiol.1984.sp002777","article-title":"Peristalsis and antiperistalsis in the chicken caecum are myogenic","volume":"69","author":"Hodgkiss","year":"1984","journal-title":"Q J Exp Physiol."},{"key":"B15","doi-asserted-by":"publisher","first-page":"213","DOI":"10.1637\/10950-093014-Reg.1","article-title":"Immune responses in cecal tonsils of Marek's disease virus-infected chickens","volume":"59","author":"Heidari","year":"2015","journal-title":"Avian Dis."},{"key":"B16","doi-asserted-by":"publisher","first-page":"188","DOI":"10.3382\/ps\/pey390","article-title":"Characterization of intestinal immune response to Clostridium perfringens infection in broiler chickens","volume":"98","author":"Fasina","year":"2019","journal-title":"Poult Sci."},{"key":"B17","doi-asserted-by":"publisher","first-page":"349","DOI":"10.1128\/CMR.9.3.349","article-title":"Avian gut-associated lymphoid tissues and intestinal immune responses to Eimeria parasites","volume":"9","author":"Lillehoj","year":"1996","journal-title":"Clin Microbiol Rev."},{"key":"B18","first-page":"2193","article-title":"Distribution of immunoglobulin-bearing cells in the gut-associated lymphoid tissues of the turkey: effect of oral treatment with intestinal microflora","volume":"45","author":"Naqi","year":"1984","journal-title":"Am J Vet Res."},{"key":"B19","doi-asserted-by":"publisher","first-page":"1997","DOI":"10.4049\/jimmunol.1002040","article-title":"Regulatory T cell properties of chicken CD4+CD25+ cells","volume":"186","author":"Shanmugasundaram","year":"2011","journal-title":"J Immunol."},{"key":"B20","doi-asserted-by":"publisher","first-page":"182","DOI":"10.1002\/(SICI)1097-0185(199802)250:2<182::AID-AR8>3.0.CO;2-5","article-title":"Appearance and development of lymphoid cells in the chicken (Gallus gallus) caecal tonsil","volume":"250","author":"Del Moral","year":"1998","journal-title":"Anatom Rec."},{"key":"B21","doi-asserted-by":"publisher","first-page":"e00380","DOI":"10.1128\/IAI.00380-17","article-title":"Influence of the gut microbiota composition on Campylobacter jejuni colonization in chickens","volume":"85","author":"Han","year":"2017","journal-title":"Infect Immun."},{"key":"B22","doi-asserted-by":"publisher","first-page":"67","DOI":"10.1016\/j.vetmic.2017.11.010","article-title":"Gene expression profiling of chicken cecal tonsils and ileum following oral exposure to soluble and PLGA-encapsulated CpG ODN, and lysate of Campylobacter jejuni","volume":"212","author":"Taha-Abdelaziz","year":"2017","journal-title":"Vet Microbiol"},{"key":"B23","doi-asserted-by":"publisher","first-page":"108732","DOI":"10.1016\/j.vetmic.2020.108732","article-title":"Campylobacter-derived ligands induce cytokine and chemokine expression in chicken macrophages and cecal tonsil mononuclear cells","volume":"246","author":"Taha-Abdelaziz","year":"2020","journal-title":"Vet Microbiol"},{"key":"B24","doi-asserted-by":"publisher","first-page":"52","DOI":"10.1093\/jac\/dki419","article-title":"Interaction of CmeABC and CmeDEF in conferring antimicrobial resistance and maintaining cell viability in Campylobacter jejuni","volume":"57","author":"Akiba","year":"2005","journal-title":"J Antimicrob Chemother"},{"key":"B25","doi-asserted-by":"publisher","first-page":"594","DOI":"10.1128\/JB.180.3.594-599.1998","article-title":"Identification of Campylobacter jejuni promoter sequences","volume":"180","author":"W\u00f6sten","year":"1998","journal-title":"J Bacteriol."},{"key":"B26","doi-asserted-by":"publisher","first-page":"1711","DOI":"10.4315\/0362-028X-64.11.1711","article-title":"Development of a selective differential agar for isolation and enumeration of Campylobacter spp","volume":"64","author":"Line","year":"2001","journal-title":"J Food Protect."},{"key":"B27","doi-asserted-by":"publisher","first-page":"172","DOI":"10.1186\/s12859-016-1016-7","article-title":"Growthcurver: an R package for obtaining interpretable metrics from microbial growth curves","volume":"17","author":"Sprouffske","year":"2016","journal-title":"BMC Bioinform."},{"key":"B28","doi-asserted-by":"publisher","first-page":"462","DOI":"10.1136\/jcp.35.4.462","article-title":"A selective medium for isolating Campylobacter jejuni\/coli","volume":"35","author":"Bolton","year":"1982","journal-title":"J Clin Pathol"},{"key":"B29","first-page":"p. 7.01","article-title":"Isolation of Campylobacter species from food and water","volume-title":"Bacteriological Analytical Manual","author":"Hunt","year":"1998"},{"key":"B30","doi-asserted-by":"publisher","first-page":"2","DOI":"10.1111\/j.1745-4581.2008.00111.x","article-title":"Addition of sulfamethoxazole to selective media aids in the recovery of Campylobacter spp. from broiler rinses","volume":"16","author":"Line","year":"2008","journal-title":"J Rapid Methods Automat Microbiol."},{"key":"B31","doi-asserted-by":"publisher","first-page":"648","DOI":"10.2144\/97234bm22","article-title":"Simplified agar plate method for quantifying viable bacteria","volume":"23","author":"Jett","year":"1997","journal-title":"Biotechniques."},{"key":"B32","doi-asserted-by":"publisher","first-page":"15","DOI":"10.1093\/bioinformatics\/bts635","article-title":"STAR: ultrafast universal RNA-seq aligner","volume":"29","author":"Dobin","year":"2013","journal-title":"Bioinformatics."},{"key":"B33","doi-asserted-by":"publisher","first-page":"550","DOI":"10.1186\/s13059-014-0550-8","article-title":"Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2","volume":"15","author":"Love","year":"2014","journal-title":"Genome Biol."},{"key":"B34","doi-asserted-by":"publisher","first-page":"e161","DOI":"10.1093\/nar\/gku864","article-title":"svaseq: removing batch effects and other unwanted noise from sequencing data","volume":"42","author":"Leek","year":"2014","journal-title":"Nucleic Acids Res."},{"key":"B35","doi-asserted-by":"publisher","first-page":"44","DOI":"10.1038\/nprot.2008.211","article-title":"Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources","volume":"4","author":"Huang Da","year":"2009","journal-title":"Nat Protoc."},{"key":"B36","doi-asserted-by":"publisher","first-page":"489","DOI":"10.1016\/S1369-5274(99)00005-3","article-title":"The biological cost of antibiotic resistance","volume":"2","author":"Andersson","year":"1999","journal-title":"Curr Opin Microbiol."},{"key":"B37","doi-asserted-by":"publisher","first-page":"14607","DOI":"10.1073\/pnas.241517298","article-title":"Mutation frequency and biological cost of antibiotic resistance in Helicobacter pylori","volume":"98","author":"Bjorkholm","year":"2001","journal-title":"Proc Natl Acad Sci USA"},{"key":"B38","doi-asserted-by":"publisher","first-page":"1300","DOI":"10.1128\/AAC.05516-11","article-title":"Impaired fitness and transmission of macrolide-resistant Campylobacter jejuni in its natural host","volume":"56","author":"Luangtongkum","year":"2012","journal-title":"Antimicrob Agents Chemother."},{"key":"B39","doi-asserted-by":"publisher","first-page":"11","DOI":"10.1016\/S0021-9975(96)80058-2","article-title":"Comparison of the effects of infectious bronchitis and infectious laryngotracheitis on the chicken respiratory tract","volume":"114","author":"Nakamura","year":"1996","journal-title":"J Compar Pathol."},{"key":"B40","doi-asserted-by":"publisher","first-page":"908","DOI":"10.1093\/ps\/77.6.908","article-title":"Serum levels of interleukin-6, alpha1-acid glycoprotein, and corticosterone in two-week-old chickens inoculated with Escherichia coli lipopolysaccharide","volume":"77","author":"Nakamura","year":"1998","journal-title":"Poultry Sci."},{"key":"B41","doi-asserted-by":"publisher","first-page":"152","DOI":"10.1080\/00071669889547","article-title":"Changes in plasma alpha 1-acid glycoprotein concentration and selected immune response in broiler chickens injected with Escherichia coli lipopolysaccharide","volume":"39","author":"Takahashi","year":"1998","journal-title":"Br. Poultry Sci."},{"key":"B42","doi-asserted-by":"publisher","first-page":"289","DOI":"10.2307\/1592967","article-title":"The kinetics of hemopexin and \u03b11-acid glycoprotein levels induced by injection of inflammatory agents in chickens","volume":"45","author":"Adler","year":"2001","journal-title":"Avian Dis."},{"key":"B43","doi-asserted-by":"publisher","first-page":"1295","DOI":"10.1093\/ps\/81.9.1295","article-title":"Comparison of the effects of infection with Salmonella Enteritidis, in combination with an induced molt, on serum levels of the acute phase protein, alpha1 acid glycoprotein, in hens","volume":"81","author":"Holt","year":"2002","journal-title":"Poultry Sci."},{"key":"B44","doi-asserted-by":"publisher","first-page":"3097","DOI":"10.1016\/j.vaccine.2012.02.055","article-title":"Vaccination and acute phase mediator production in chickens challenged with low pathogenic avian influenza virus; novel markers for vaccine efficacy?","volume":"30","author":"Sylte","year":"2012","journal-title":"Vaccine."},{"key":"B45","doi-asserted-by":"publisher","first-page":"47","DOI":"10.1016\/j.tvjl.2012.07.022","article-title":"Alpha(1)-acid glycoprotein modulates phagocytosis and killing of Escherichia coli by bovine polymorphonuclear leucocytes and monocytes","volume":"196","author":"Lecchi","year":"2013","journal-title":"Vet J."},{"key":"B46","doi-asserted-by":"publisher","first-page":"329","DOI":"10.1007\/s13105-015-0389-9","article-title":"Orosomucoid, an acute response protein with multiple modulating activities","volume":"71","author":"Luo","year":"2015","journal-title":"J Physiol Biochem."},{"key":"B47","doi-asserted-by":"publisher","first-page":"10","DOI":"10.1016\/j.vetimm.2017.06.006","article-title":"Widespread extrahepatic expression of acute-phase proteins in healthy chicken (Gallus gallus) tissues","volume":"190","author":"Marques","year":"2017","journal-title":"Vet Immunol Immunopathol."},{"key":"B48","doi-asserted-by":"publisher","first-page":"1997","DOI":"10.1073\/pnas.84.7.1997","article-title":"cDNA cloning and sequence of MAL, a hydrophobic protein associated with human T-cell differentiation","volume":"84","author":"Alonso","year":"1987","journal-title":"Proc Natl Acad Sci USA."},{"key":"B49","doi-asserted-by":"publisher","first-page":"1207","DOI":"10.3390\/ijms17081207","article-title":"Chicken-specific kinome array reveals that Salmonella enterica Serovar Enteritidis modulates host immune signaling pathways in the cecum to establish a persistence infection","volume":"17","author":"Kogut","year":"2016","journal-title":"Int J Mol Sci."},{"key":"B50","doi-asserted-by":"publisher","first-page":"372","DOI":"10.3389\/fimmu.2017.00372","article-title":"Immunometabolic phenotype alterations associated with the induction of disease tolerance and persistent asymptomatic infection of Salmonella in the chicken intestine","volume":"8","author":"Kogut","year":"2017","journal-title":"Front Immunol."},{"key":"B51","doi-asserted-by":"publisher","first-page":"1828","DOI":"10.3382\/ps\/pev161","article-title":"Effect of Salmonella infection on cecal tonsil regulatory T cell properties in chickens","volume":"94","author":"Shanmugasundaram","year":"2015","journal-title":"Poult Sci."},{"key":"B52","doi-asserted-by":"publisher","first-page":"e00307","DOI":"10.1128\/IAI.00307-18","article-title":"Model of persistent salmonella infection: Salmonella enterica serovar pullorum modulates the immune response of the chicken from a Th17-type response towards a Th2-type response","volume":"86","author":"Tang","year":"2018","journal-title":"Infect Immun."},{"key":"B53","doi-asserted-by":"publisher","first-page":"27","DOI":"10.1016\/j.gene.2019.02.068","article-title":"Transcriptome modulation by in ovo delivered Lactobacillus synbiotics in a range of chicken tissues","volume":"698","author":"Dunislawska","year":"2019","journal-title":"Gene."},{"key":"B54","first-page":"232835","article-title":"Pathway enrichment analysis of \u201comics\u201d data","volume-title":"bioRxiv.","author":"Reimand","year":"2017"},{"key":"B55","doi-asserted-by":"publisher","first-page":"2357","DOI":"10.4049\/jimmunol.1900899","article-title":"Platelets in inflammation and resolution","volume":"203","author":"Margraf","year":"2019","journal-title":"J Immunol."},{"key":"B56","doi-asserted-by":"publisher","first-page":"495","DOI":"10.1080\/01926230701320337","article-title":"Apoptosis: a review of programmed cell death","volume":"35","author":"Elmore","year":"2007","journal-title":"Toxicol. Pathol."},{"key":"B57","doi-asserted-by":"publisher","first-page":"65","DOI":"10.1016\/j.rvsc.2018.10.009","article-title":"Oxidative stress, apoptosis and abnormal expression of apoptotic protein and gene and cell cycle arrest in the cecal tonsil of broilers induces by dietary methionine deficiency","volume":"121","author":"Wu","year":"2018","journal-title":"Res Vet Sci."},{"key":"B58","doi-asserted-by":"publisher","first-page":"173","DOI":"10.1016\/j.resmic.2010.09.024","article-title":"MCLMAN, a new minimal medium for Campylobacter jejuni NCTC 11168","volume":"162","author":"Alazzam","year":"2011","journal-title":"Res Microbiol."},{"key":"B59","doi-asserted-by":"publisher","first-page":"144","DOI":"10.1007\/s11306-018-1438-5","article-title":"Unexpected differential metabolic responses of Campylobacter jejuni to the abundant presence of glutamate and fucose","volume":"14","author":"Van Der Hooft","year":"2018","journal-title":"Metabolomics."},{"key":"B60","doi-asserted-by":"publisher","first-page":"94","DOI":"10.1128\/JVI.01707-19","article-title":"The G Protein-coupled receptor FFAR2 promotes internalization during influenza A virus entry","author":"Wang","year":"2020","journal-title":"J Virol."},{"key":"B61","doi-asserted-by":"publisher","first-page":"1047","DOI":"10.1016\/S0006-291X(03)00488-1","article-title":"Identification of a free fatty acid receptor, FFA2R, expressed on leukocytes and activated by short-chain fatty acids","volume":"303","author":"Nilsson","year":"2003","journal-title":"Biochem Biophys Res Commun."}],"container-title":["Frontiers in Veterinary Science"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fvets.2021.613203\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2021,4,6]],"date-time":"2021-04-06T05:00:29Z","timestamp":1617685229000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fvets.2021.613203\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,4,6]]},"references-count":61,"alternative-id":["10.3389\/fvets.2021.613203"],"URL":"https:\/\/doi.org\/10.3389\/fvets.2021.613203","relation":{},"ISSN":["2297-1769"],"issn-type":[{"value":"2297-1769","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,4,6]]},"article-number":"613203"}}