{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,21]],"date-time":"2026-09-21T05:03:21Z","timestamp":1789967001374,"version":"4.0.1"},"reference-count":45,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,9,21]],"date-time":"2026-09-21T00:00:00Z","timestamp":1789948800000},"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:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>Milk fever is a serious metabolic disease affecting high-yielding dairy cows during the transition period and can result in substantial economic losses due to reduced milk production, impaired health, increased treatment costs, and culling.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>The present study aimed to identify molecular signatures associated with milk fever in Holstein dairy cows using targeted blood gene-expression profiling, genetic variation, and artificial neural network (ANN) modeling. One hundred multiparous Holstein cows were enrolled in the study, fifty of them clinically healthy and fifty diagnosed with milk fever based on clinical signs and serum calcium levels below 2.0 mmol\/L. Quantitative real-time PCR was used to measure relative mRNA expression of 13 potential calcium signaling, endocrine control and cellular metabolism-related genes.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      Single nucleotide polymorphisms (SNPs) were then detected by PCR product sequencing. ANN and hierarchical cluster analysis were used to assess the diagnostic performance of the biomarkers tested. Milk fever cows showed significant upregulation of\n                      <jats:italic>PKIB, ASIC2, PER1, NUAK1, STX1A, SRI<\/jats:italic>\n                      and\n                      <jats:italic>ITPR1<\/jats:italic>\n                      , and significant downregulation of\n                      <jats:italic>CAMK2A, ANXA6, CACNA1S, VDR, CAMK4<\/jats:italic>\n                      and\n                      <jats:italic>NESP55<\/jats:italic>\n                      (\n                      <jats:italic>p<\/jats:italic>\n                      &amp;lt; 0.05). Sequencing identified 23 SNPs in the coding region, of which 17 were synonymous and 6 non-synonymous. These variants showed significant differences in their distribution between healthy and affected cows (\n                      <jats:italic>p<\/jats:italic>\n                      &amp;lt; 0.001). The most informative biomarkers for the classification of the disease by ANN analysis were\n                      <jats:italic>NUAK1<\/jats:italic>\n                      and\n                      <jats:italic>NESP55<\/jats:italic>\n                      . Hierarchical clustering based on normalized gene-expression profiles also showed distinct clustering patterns between healthy and affected cows.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Discussion<\/jats:title>\n                    <jats:p>The identified gene-expression signatures and coding-region variants may provide candidate biomarkers and genetic markers for future studies aimed at reducing susceptibility to milk fever in dairy cattle. These results show that the combination of targeted blood gene-expression profiling, functional genetic variation, and machine-learning methods offers a powerful framework for genomic prediction, precision livestock management, and future marker-assisted breeding strategies.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.3389\/fvets.2026.1946092","type":"journal-article","created":{"date-parts":[[2026,9,21]],"date-time":"2026-09-21T04:28:40Z","timestamp":1789964920000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":0,"title":["Integrative targeted blood gene-expression profiling, coding-region genetic variants, and machine learning identify candidate biomarkers of milk fever in Holstein dairy cattle"],"prefix":"10.3389","volume":"13","author":[{"given":"Basma Mohamed","family":"Hendam","sequence":"first","affiliation":[{"name":"Department of Animal Wealth Development, Faculty of Veterinary Medicine, Mansoura University","place":["Mansoura, Egypt"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tahani M. I.","family":"Al-Hazani","sequence":"additional","affiliation":[{"name":"Biology Department, College of Science and Humanities, Prince Sattam Bin Abdulaziz University","place":["Al-Kharj, Saudi Arabia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Asmaa A.","family":"Badr","sequence":"additional","affiliation":[{"name":"Department of Animal Wealth Development, Faculty of Veterinary Medicine, Mansoura University","place":["Mansoura, Egypt"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Khairiah Mubarak","family":"Alwutayd","sequence":"additional","affiliation":[{"name":"Department of Biology, College of Science, Princess Nourah bint Abdulrahman University","place":["Riyadh, Saudi Arabia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hanan M.","family":"Alharbi","sequence":"additional","affiliation":[{"name":"Department of Biology, College of Science, Princess Nourah bint Abdulrahman University","place":["Riyadh, Saudi Arabia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ashwag","family":"Shami","sequence":"additional","affiliation":[{"name":"Department of Biology, College of Science, Princess Nourah bint Abdulrahman University","place":["Riyadh, Saudi Arabia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mariam Abdulaziz","family":"Alkhateeb","sequence":"additional","affiliation":[{"name":"Department of Biology, College of Science, Princess Nourah bint Abdulrahman University","place":["Riyadh, Saudi Arabia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nawal","family":"Al-Hoshani","sequence":"additional","affiliation":[{"name":"Department of Biology, College of Science, Princess Nourah bint Abdulrahman University","place":["Riyadh, Saudi Arabia"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ahmed M.","family":"El-Sebaey","sequence":"additional","affiliation":[{"name":"Department of Clinical Pathology, Faculty of Veterinary Medicine, Mansoura University","place":["Mansoura, Egypt"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ahmed","family":"Ateya","sequence":"additional","affiliation":[{"name":"Department of Animal Wealth Development, Faculty of Veterinary Medicine, Mansoura University","place":["Mansoura, Egypt"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2026,9,21]]},"reference":[{"key":"ref1","doi-asserted-by":"publisher","first-page":"141","DOI":"10.1007\/s10911-005-5397-x","article-title":"Adapting to the transition between gestation and lactation: differences between rat, human and dairy cow","volume":"10","author":"Horst","year":"2005","journal-title":"J Mammary Gland Biol Neoplasia"},{"key":"ref2","doi-asserted-by":"publisher","first-page":"50","DOI":"10.1016\/j.tvjl.2007.12.020","article-title":"The monitoring, prevention, and treatment of milk fever and subclinical hypocalcemia in dairy cows","volume":"176","author":"Goff","year":"2008","journal-title":"Vet J"},{"key":"ref3","doi-asserted-by":"publisher","first-page":"1530","DOI":"10.5713\/ajas.16.0310","article-title":"Genetic parameters of milk \u03b2-hydroxybutyric acid and acetone and their genetic association with milk production traits of Holstein cattle","volume":"29","author":"Lee","year":"2016","journal-title":"Asian Australas J Anim Sci"},{"key":"ref4","doi-asserted-by":"publisher","first-page":"7655","DOI":"10.3168\/jds.2019-16454","article-title":"Invited review: determination of large-scale individual dry matter intake phenotypes in dairy cattle","volume":"102","author":"Seymour","year":"2019","journal-title":"J Dairy Sci"},{"key":"ref5","first-page":"198","article-title":"Effects of negative energy balance on reproduction in dairy cows","volume":"42","author":"Knop","year":"2009","journal-title":"Lucr\u0103ri. 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