{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T11:10:55Z","timestamp":1785409855308,"version":"3.56.0"},"reference-count":30,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2026,6,4]],"date-time":"2026-06-04T00:00:00Z","timestamp":1780531200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100007539","name":"Jiangsu Province Agricultural Science and Technology Independent Innovation Fund Project","doi-asserted-by":"publisher","award":["CX(22)2016"],"award-info":[{"award-number":["CX(22)2016"]}],"id":[{"id":"10.13039\/100007539","id-type":"DOI","asserted-by":"publisher"}]},{"award":["CX(22)2016"],"award-info":[{"award-number":["CX(22)2016"]}],"id":[{"id":"https:\/\/ror.org\/0335n2618","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Vaccines"],"abstract":"<jats:p>Background: Pseudorabies vaccines based on the live attenuated PRV Bartha-K61 strain remain essential for controlling pseudorabies in swine, but poor thermal stability during storage and transport limits field use. Objectives: This study aimed to develop a thermostable freeze-dried live pseudorabies vaccine through the integrated optimization of formulation and lyophilization and to preliminarily assess its humoral immunogenicity in piglets. Methods: Trehalose, mannitol, and glycine were screened as candidate lyoprotectants by single-factor experiments, followed by Box\u2013Behnken response surface optimization using viral titer retention as the response. Critical thermal parameters were determined to establish a formulation-specific lyophilization cycle. Product quality was evaluated by post-lyophilization titer, cake appearance, residual moisture, and electron microscopy, and stability was compared with a commercial freeze-dried vaccine at 2\u20138 \u00b0C, 25 \u00b0C, 37 \u00b0C, and 45 \u00b0C. Conclusions: The optimized formulation, ST005, contained 9.5% trehalose, 2.0% mannitol, and 1.5% glycine. It showed a collapse-related critical temperature of approximately \u221234.0 \u00b0C and a dried-product glass transition temperature of 69.1 \u00b0C, produced an intact cake with residual moisture below 3.0%, and preserved viral morphology after lyophilization. Titer loss remained below 1.0 log10 TCID50\/mL for 24 months at 2\u20138 \u00b0C, 9 months at 25 \u00b0C, 21 days at 37 \u00b0C, and 9 days at 45 \u00b0C, outperforming the commercial comparator. In PRV-seronegative piglets, stored ST005 induced robust gB-specific and neutralizing antibody responses after storage under refrigerated, ambient, and accelerated conditions.<\/jats:p>","DOI":"10.3390\/vaccines14060506","type":"journal-article","created":{"date-parts":[[2026,6,5]],"date-time":"2026-06-05T01:23:21Z","timestamp":1780622601000},"page":"506","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Development of a Thermostable Freeze-Dried Live Pseudorabies Vaccine Based on the PRV Bartha-K61 Strain: Formulation Optimization, Stability Evaluation, and Preliminary Immunogenicity in Piglets"],"prefix":"10.3390","volume":"14","author":[{"given":"Yanhong","family":"Zhao","sequence":"first","affiliation":[{"name":"College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210014, China"},{"name":"Institute of Veterinary Immunology & Engineering, National Research Center of Engineering and Technology for Veterinary Biologicals, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China"},{"name":"Guotai (Taizhou) Center of Technology Innovation for Veterinary Biology, Taizhou 225300, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaoqing","family":"Pan","sequence":"additional","affiliation":[{"name":"Livestock Research Institute, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Fang","family":"Lv","sequence":"additional","affiliation":[{"name":"Institute of Veterinary Immunology & Engineering, National Research Center of Engineering and Technology for Veterinary Biologicals, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yalu","family":"Zhu","sequence":"additional","affiliation":[{"name":"Institute of Veterinary Immunology & Engineering, National Research Center of Engineering and Technology for Veterinary Biologicals, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China"},{"name":"Guotai (Taizhou) Center of Technology Innovation for Veterinary Biology, Taizhou 225300, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhen","family":"Wang","sequence":"additional","affiliation":[{"name":"Nanjing Jinsiyuan Biotechnology Co., Ltd., Xuanwu District, Nanjing 210014, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yu","family":"Lu","sequence":"additional","affiliation":[{"name":"College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210014, China"},{"name":"Institute of Veterinary Immunology & Engineering, National Research Center of Engineering and Technology for Veterinary Biologicals, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China"},{"name":"Guotai (Taizhou) Center of Technology Innovation for Veterinary Biology, Taizhou 225300, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Endong","family":"Bao","sequence":"additional","affiliation":[{"name":"College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210014, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,6,4]]},"reference":[{"key":"ref_1","first-page":"99","article-title":"Aujeszky\u2019s disease (pseudorabies) virus: The virus and molecular pathogenesis-state of the art, June 1999","volume":"31","author":"Mettenleiter","year":"2000","journal-title":"Vet. 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