{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,9]],"date-time":"2026-10-09T03:40:42Z","timestamp":1791517242639,"version":"4.3.4"},"reference-count":36,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2025,6,4]],"date-time":"2025-06-04T00:00:00Z","timestamp":1748995200000},"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>Introduction<\/jats:title>\n                    <jats:p>\n                      Yaks serve as a vital economic and ecological resource in high-altitude regions, but it faces significant health challenges from various pathogens. Among these,\n                      <jats:italic>Pasteurella multocida<\/jats:italic>\n                      and\n                      <jats:italic>Salmonella<\/jats:italic>\n                      are critical pathogens that contribute to severe diseases.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>\n                      A duplex real-time fluorescence quantitative PCR assay was developed to simultaneously detect\n                      <jats:italic>Pasteurella multocida<\/jats:italic>\n                      and\n                      <jats:italic>Salmonella<\/jats:italic>\n                      . The species-specific genes\n                      <jats:italic>kmt1<\/jats:italic>\n                      and\n                      <jats:italic>invA<\/jats:italic>\n                      were selected as target regions for primer and probe design. Following rigorous optimization, a duplex assay was established. Recombinant plasmids were constructed to serve as standards for generating standard curves. The detection thresholds were determined using SPSS statistical analysis and receiver operating characteristic curve methods. Furthermore, the assay\u2019s sensitivity, specificity, stability, and clinical applicability were evaluated.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      The established assay demonstrated high sensitivity, with detection limits of 100 and 10 copies for pMD-kmt1 and pMD-invA, respectively. No cross-reactivity was observed with six pathogens, including\n                      <jats:italic>Mycoplasma bovis<\/jats:italic>\n                      , infectious bovine rhinotracheitis virus and others. The standard curves showed strong linearity, with coefficients of determination of 0.995 and 0.998, and amplification efficiencies of 103.37% and 103.47% for pMD-kmt1 and pMD-invA, respectively. No interference was observed between high- and low-concentration templates during simultaneous detection. The intra- and inter-assay coefficients of variation ranged from 0.23% to 1.51%. Detection thresholds were determined to be cycle threshold values of 41.5 for\n                      <jats:italic>P. multocida<\/jats:italic>\n                      and 40.0 for\n                      <jats:italic>Salmonella<\/jats:italic>\n                      . Clinical evaluation was performed on 226 samples collected from yaks in seven counties of Ganzi Prefecture, Sichuan Province, China. The single infection rates of\n                      <jats:italic>P. multocida<\/jats:italic>\n                      and\n                      <jats:italic>Salmonella<\/jats:italic>\n                      were 20.35% (46\/226) and 38.50% (87\/226), respectively, while the co-infection rate was 6.19% (14\/226).\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Discussion<\/jats:title>\n                    <jats:p>\n                      This study successfully established a duplex real-time fluorescence PCR assay that enables the simultaneous detection of\n                      <jats:italic>P. multocida<\/jats:italic>\n                      and\n                      <jats:italic>Salmonella<\/jats:italic>\n                      with high sensitivity, specificity, and efficiency. The assay offers a reliable and rapid diagnostic tool that is particularly suited for clinical and epidemiological investigations in yak populations.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.3389\/fcimb.2025.1599817","type":"journal-article","created":{"date-parts":[[2025,6,4]],"date-time":"2025-06-04T01:37:09Z","timestamp":1749001029000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":5,"title":["Establishment and epidemiological investigation of a dual fluorescent qPCR assay for Pasteurella multocida and Salmonella in yaks in the Tibetan Autonomous Prefecture of Garz\u00ea, China"],"prefix":"10.3389","volume":"15","author":[{"given":"Yao","family":"Pan","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qingting","family":"Yu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qi","family":"Wang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qiang","family":"Li","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wei","family":"Tian","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lingxiang","family":"Xin","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xing","family":"Hu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Haiyue","family":"Xiao","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yuanjie","family":"Liu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Luo Rong Deng","family":"Zhu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lan","family":"Lan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Liangquan","family":"Zhu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jianping","family":"Wu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2025,6,4]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","first-page":"109586","DOI":"10.1016\/j.meatsci.2024.109586","article-title":"Understanding consumer preferences for attributes of yak meat: implications for economic growth and resource efficiency in pastoral areas","volume":"216","author":"Ai","year":"2024","journal-title":"Meat Sci."},{"key":"B2","doi-asserted-by":"publisher","DOI":"10.3390\/vaccines10020315","article-title":"Spatial, temporal, and demographic patterns in the prevalence of hemorrhagic septicemia in 41 countries in 2005-2019: A systematic analysis with special focus on the potential development of a new-generation vaccine","volume":"10","author":"Almoheer","year":"2022","journal-title":"Vaccines (Basel)"},{"key":"B3","doi-asserted-by":"publisher","first-page":"1313","DOI":"10.5455\/OVJ.2024.v14.i6.1","article-title":"Tackling salmonellosis: A comprehensive exploration of risks factors, impacts, and solutions","volume":"14","author":"Ayuti","year":"2024","journal-title":"Open Vet J."},{"key":"B4","doi-asserted-by":"publisher","first-page":"455","DOI":"10.30466\/vrf.2024.2017140.4090","article-title":"Susceptibility of Pasteurella multocida isolated from cattle in Egypt to antibiotics, silver, chitosan and curcumin nanoparticles","volume":"15","author":"Bahr","year":"2024","journal-title":"Vet Res. 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