{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,20]],"date-time":"2026-05-20T01:55:05Z","timestamp":1779242105010,"version":"3.51.4"},"reference-count":49,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,2,9]],"date-time":"2026-02-09T00:00:00Z","timestamp":1770595200000},"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. Cell. Infect. Microbiol."],"abstract":"<jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>\n                      The flavivirus Duck Tembusu virus (DTMUV) exhibits high pathogenicity and transmissibility, posing a severe threat to the poultry industry in China and Southeast Asia. Although the molecular mechanisms of DTMUV pathogenesis remain unclear, its potential to disrupt intracellular calcium ion (Ca\u00b2\n                      <jats:sup>+<\/jats:sup>\n                      ) homeostasis, a known driver of viral replication, has not been investigated. Here, we investigated the role and underlying mechanism of Ca\u00b2\n                      <jats:sup>+<\/jats:sup>\n                      homeostasis in DTMUV infection.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>\n                      Fluo-4AM staining and flow cytometry of duck embryo fibroblasts (DEFs) were used to detect cytoplasmic Ca\u00b2\n                      <jats:sup>+<\/jats:sup>\n                      . To modulate Ca\u00b2\n                      <jats:sup>+<\/jats:sup>\n                      levels and AMP-activated protein kinase (AMPK) activity, we used voltage-gated calcium channel (VGCC) blockers (verapamil, diltiazem hydrochloride), a Ca\u00b2\n                      <jats:sup>+<\/jats:sup>\n                      chelator (BAPTA-AM), and an AMPK inhibitor (Compound C). Viral entry, genomic RNA replication (targeting the DTMUV NS5 gene), viral yield, and release were evaluated via qRT-PCR and plaque assays. AMPK activation was detected using western blotting with anti-phospho-AMPK\u03b1 (Thr172) and anti-AMPK\u03b1 antibodies. Statistical analyses were performed using Student\u2019s t-test or two-way analysis of variance.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      DTMUV infection significantly increased cytoplasmic Ca\u00b2\n                      <jats:sup>+<\/jats:sup>\n                      in DEFs at 6, 8, 10, and 12 hours post-infection. This elevation was suppressed by treatment with verapamil or diltiazem hydrochloride, indicating that DTMUV induces extracellular Ca\u00b2\n                      <jats:sup>+<\/jats:sup>\n                      influx via VGCCs. Functional assays showed that reducing cytoplasmic Ca\u00b2\n                      <jats:sup>+<\/jats:sup>\n                      via treatment with VGCC blockers or BAPTA-AM specifically inhibited DTMUV RNA replication, but not viral entry or release, decreasing progeny virus production. Further mechanistic analysis revealed that DTMUV infection activates the AMPK pathway in a Ca\u00b2\n                      <jats:sup>+<\/jats:sup>\n                      -dependent manner, with Compound C-mediated AMPK inhibition dose-dependently suppressing viral RNA replication and progeny yield.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>\n                      DTMUV disrupts host Ca\u00b2\n                      <jats:sup>+<\/jats:sup>\n                      homeostasis to activate AMPK, which promotes viral RNA replication. This study provides novel insights into DTMUV pathogenesis and identifies Ca\u00b2\n                      <jats:sup>+<\/jats:sup>\n                      \u2013AMPK signaling as a potential anti-DTMUV target.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.3389\/fcimb.2026.1743907","type":"journal-article","created":{"date-parts":[[2026,2,9]],"date-time":"2026-02-09T07:48:41Z","timestamp":1770623321000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":1,"title":["Disruption of cellular calcium homeostasis by duck Tembusu virus facilitates viral replication via AMPK pathway activation"],"prefix":"10.3389","volume":"16","author":[{"given":"Ziding","family":"Yu","sequence":"first","affiliation":[{"name":"Wuhu Vocational Technical University","place":["Wuhu, China"]},{"name":"Detection of Food-borne Pathogenic Microorganisms Engineering Research Center of Wuhu","place":["Wuhu, 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