{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,11,1]],"date-time":"2025-11-01T05:45:08Z","timestamp":1761975908486,"version":"build-2065373602"},"reference-count":23,"publisher":"MDPI AG","issue":"11","license":[{"start":{"date-parts":[[2025,10,30]],"date-time":"2025-10-30T00:00:00Z","timestamp":1761782400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100007219","name":"Shanghai Natural Science Foundation","doi-asserted-by":"crossref","award":["23ZR1476800"],"award-info":[{"award-number":["23ZR1476800"]}],"id":[{"id":"10.13039\/100007219","id-type":"DOI","asserted-by":"crossref"}]},{"name":"Major Tasks of Chinese Academy of Agricultural Sciences","award":["CAAS-ZDRW202409-2"],"award-info":[{"award-number":["CAAS-ZDRW202409-2"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>African swine fever (ASF), caused by African swine fever virus (ASFV), has inflicted severe economic losses on China\u2019s pig industry. Existing ASFV nucleic acid detection methods struggle to identify infected pigs in the pre-viremic stage, especially for recently emerged recombinant ASFV strains that exhibit delayed clinical symptoms and prolonged virus shedding, posing great challenges to ASF prevention and control. To fit the problem, this study established a TaqMan duplex quantitative polymerase chain reaction (qPCR) assay targeting the ASFV p72 gene and porcine Hp gene for early diagnosis of ASFV infection. The qPCR reaction system (20 \u03bcL) and conditions were optimized and showed high sensitivity, with detection limits of 1.42 \u00d7 101 copies\/\u03bcL for Hp and 2.23 \u00d7 101 copies\/\u03bcL for ASFV, as well as excellent specificity and reproducibility. Serum cDNA samples from pigs infected with virulent or recombinant ASFV strains were tested, and the result showed that Hp was detectable as early as 1 day post-infection (DPI), however ASFV remained undetectable until 3DPI. Then cDNA samples from cohabitation infection were tested and 80% samples were Hp-positive, although ASFV test was negative.In conclusion, this duplex qPCR assay for simultaneous detection of Hp and ASFV enables pre-viremia diagnosis of ASF, providing a valuable tool for early screening of ASFV-infected pigs.<\/jats:p>","DOI":"10.3390\/v17111444","type":"journal-article","created":{"date-parts":[[2025,10,31]],"date-time":"2025-10-31T01:23:59Z","timestamp":1761873839000},"page":"1444","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Simultaneous ASFV and Haptoglobin Detection by Duplex qPCR Enables Pre-Viremia Diagnosis of African Swine Fever"],"prefix":"10.3390","volume":"17","author":[{"given":"Yun","family":"Bao","sequence":"first","affiliation":[{"name":"College of Veterinary Medicine, Shanxi Agricultural University, Jinzhong 030801, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shimin","family":"Gao","sequence":"additional","affiliation":[{"name":"College of Veterinary Medicine, Shanxi Agricultural University, Jinzhong 030801, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shuang","family":"Li","sequence":"additional","affiliation":[{"name":"Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yijie","family":"Liu","sequence":"additional","affiliation":[{"name":"Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2291-8712","authenticated-orcid":false,"given":"Fei","family":"Gao","sequence":"additional","affiliation":[{"name":"Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Liwei","family":"Li","sequence":"additional","affiliation":[{"name":"Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Wu","family":"Tong","sequence":"additional","affiliation":[{"name":"Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7562-8941","authenticated-orcid":false,"given":"Changlong","family":"Liu","sequence":"additional","affiliation":[{"name":"Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yanjun","family":"Zhou","sequence":"additional","affiliation":[{"name":"Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yifeng","family":"Jiang","sequence":"additional","affiliation":[{"name":"Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China"},{"name":"Jiangsu Co-Innovation Center for the Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,10,30]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Shi, K., Qian, X., Shi, Y., Wei, H., Pan, Y., Long, F., Zhou, Q., Mo, S., Hu, L., and Li, Z. (2024). A triplex crystal digital PCR for the detection of genotypes I and II African swine fever virus. Front. Vet. Sci., 11.","DOI":"10.3389\/fvets.2024.1351596"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"640","DOI":"10.1126\/science.aaz1439","article-title":"Architecture of African swine fever virus and implications for viral assembly","volume":"366","author":"Wang","year":"2019","journal-title":"Science"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"438","DOI":"10.1080\/22221751.2019.1590128","article-title":"Replication and virulence in pigs of the first African swine fever virus isolated in China","volume":"8","author":"Zhao","year":"2019","journal-title":"Emerg. Microbes Infect."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"2457949","DOI":"10.1080\/21505594.2025.2457949","article-title":"Insights and progress on epidemic characteristics, pathogenesis, and preventive measures of African swine fever virus: A review","volume":"16","author":"Li","year":"2025","journal-title":"Virulence"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"2183","DOI":"10.1080\/22221751.2021.1999779","article-title":"Genotype I African swine fever viruses emerged in domestic pigs in China and caused chronic infection","volume":"10","author":"Sun","year":"2021","journal-title":"Emerg. Microbes Infect."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"752","DOI":"10.1007\/s11427-021-1904-4","article-title":"Emergence and prevalence of naturally occurring lower virulent African swine fever viruses in domestic pigs in China in 2020","volume":"64","author":"Sun","year":"2021","journal-title":"Sci. China Life Sci."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"3096","DOI":"10.1038\/s41467-023-38868-w","article-title":"Highly lethal genotype I and II recombinant African swine fever viruses detected in pigs","volume":"14","author":"Zhao","year":"2023","journal-title":"Nat. Commun."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"1603","DOI":"10.1016\/j.jia.2023.02.035","article-title":"Developing a duplex ARMS-qPCR method to differentiate genotype I and II African swine fever viruses based on their B646L genes","volume":"22","author":"Ding","year":"2023","journal-title":"J. Integr. Agric."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"228","DOI":"10.1016\/j.jia.2023.05.021","article-title":"Establishment of an indirect immunofluorescence assay for the detection of African swine fever virus antibodies","volume":"23","author":"Wang","year":"2024","journal-title":"J. Integr. Agric."},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Auer, A., Cattoli, G., Padungtod, P., Lamien, C.E., Oh, Y., Jayme, S., and Rozstalnyy, A. (2024). Challenges in the Application of African Swine Fever Vaccines in Asia. Animals, 14.","DOI":"10.3390\/ani14172473"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"66","DOI":"10.1186\/s12969-022-00721-7","article-title":"\u03b12-fraction and haptoglobin as biomarkers for disease activity in oligo- and polyarticular juvenile idiopathic arthritis","volume":"20","author":"Zeller","year":"2022","journal-title":"Pediatr. Rheumatol. Online J."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"61","DOI":"10.1016\/j.jmsacl.2022.07.001","article-title":"Haptoglobin polymorphism affects its N-glycosylation pattern in serum","volume":"25","author":"Busskamp","year":"2022","journal-title":"J. Mass. Spectrom. Adv. Clin. Lab."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"24","DOI":"10.1186\/s40813-024-00375-9","article-title":"New insights into swine dysentery: Faecal shedding, macro and microscopic lesions and biomarkers in early and acute stages of Brachyspira hyodysenteriae infection","volume":"10","author":"Carvajal","year":"2024","journal-title":"Porc. Health Manag."},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Grantz, J.M., Thirumalaikumar, V.P., Jannasch, A.H., Andolino, C., Taechachokevivat, N., Avila-Granados, L.M., and Neves, R.C. (2024). The platelet and plasma proteome and targeted lipidome in postpartum dairy cows with elevated systemic inflammation. Sci. Rep., 14.","DOI":"10.1038\/s41598-024-82553-x"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"488","DOI":"10.1111\/j.1439-0450.2006.01002.x","article-title":"Porcine acute phase protein concentrations in different diseases in field conditions","volume":"53","author":"Parra","year":"2006","journal-title":"J. Vet. Med. B Infect. Dis. Vet. Public Health"},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Miao, C., Yang, S., Shao, J., Zhou, G., Ma, Y., Wen, S., Hou, Z., Peng, D., Guo, H., and Liu, W. (2023). Identification of p72 epitopes of African swine fever virus and preliminary application. Front. Microbiol., 14.","DOI":"10.3389\/fmicb.2023.1126794"},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Duan, X., Ru, Y., Yang, W., Ren, J., Hao, R., Qin, X., Li, D., and Zheng, H. (2022). Research progress on the proteins involved in African swine fever virus infection and replication. Front. Immunol., 13.","DOI":"10.3389\/fimmu.2022.947180"},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Yang, Y., Yuan, H., Zhang, Y., Luan, J., and Wang, H. (2025). Progress in African Swine Fever Vector Vaccine Development. Int. J. Mol. Sci., 26.","DOI":"10.3390\/ijms26030921"},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Borca, M.V., Ramirez-Medina, E., Silva, E., Rai, A., Espinoza, N., Velazquez-Salinas, L., and Gladue, D.P. (2023). ASF Vaccine Candidate ASFV-G-\u2206I177L Does Not Exhibit Residual Virulence in Long-Term Clinical Studies. Pathogens, 12.","DOI":"10.3390\/pathogens12060805"},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Franco-Mart\u00ednez, L., Beer, M., Mart\u00ednez-Subiela, S., Garc\u00eda-Manzanilla, E., Blome, S., and Carrau, T. (2022). Impact of ASFV Detergent Inactivation on Biomarkers in Serum and Saliva Samples. Pathogens, 11.","DOI":"10.3390\/pathogens11070750"},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Silva, C.A., Callegari, M.A., Dias, C.P., de Souza, K.L., Romano, G.S., Hernig, L.F., Lippke, R.T., Jansen, R., Leite, F.L., and Filipe, F. (2024). Well-Being and Performance of Nursery Pigs Subjected to Different Commercial Vaccines Against Porcine Circovirus Type 2, Mycoplasma hyopneumoniae and Lawsonia intracellularis. Vaccines, 12.","DOI":"10.3390\/vaccines12111242"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"157","DOI":"10.1016\/j.vetimm.2006.04.008","article-title":"The porcine acute phase protein response to acute clinical and subclinical experimental infection with Streptococcus suis","volume":"113","author":"Sorensen","year":"2006","journal-title":"Vet. Immunol. Immunopathol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"3515","DOI":"10.1021\/acs.jproteome.4c00154","article-title":"Multiplex Assay to Determine Acute Phase Proteins in Modified Live PRRSV Vaccinated Pigs","volume":"23","author":"Tor","year":"2024","journal-title":"J. Proteome Res."}],"container-title":["Viruses"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4915\/17\/11\/1444\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,11,1]],"date-time":"2025-11-01T05:34:00Z","timestamp":1761975240000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4915\/17\/11\/1444"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,10,30]]},"references-count":23,"journal-issue":{"issue":"11","published-online":{"date-parts":[[2025,11]]}},"alternative-id":["v17111444"],"URL":"https:\/\/doi.org\/10.3390\/v17111444","relation":{},"ISSN":["1999-4915"],"issn-type":[{"type":"electronic","value":"1999-4915"}],"subject":[],"published":{"date-parts":[[2025,10,30]]}}}