{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,20]],"date-time":"2026-04-20T08:37:25Z","timestamp":1776674245116,"version":"3.51.2"},"reference-count":56,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2026,4,18]],"date-time":"2026-04-18T00:00:00Z","timestamp":1776470400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Cultivation and demonstration of new varieties of Culter alburnus with high quality and stress resistance","award":["2021C02069-3"],"award-info":[{"award-number":["2021C02069-3"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Animals"],"abstract":"<jats:p>To address cryodamage in Culter alburnus sperm, this study evaluated the effects of trehalose supplementation in a conventional cryomedium (D-15 + 10% ethylene glycol). Six experimental groups were established: fresh sperm (G1), a conventional cryomedium (G2), groups supplemented with 10, 100, or 200 mmol\/L trehalose (G3\u2013G5), and a control group with extender only (G6). The group with 100 mmol\/L trehalose (G4) was associated with improved post-thaw motility parameters (activation rate, movement time, and lifespan) and higher antioxidant (superoxide dismutase and catalase) and energy metabolism (ATPase, succinate dehydrogenase, lactate dehydrogenase) enzyme activities. Ultrastructural damage in G4 included partial plasma membrane rupture and mitochondrial swelling, while G6 exhibited additional damage features including membrane disintegration, mitochondrial disruption, and flagellar fracture. Proteomic analysis revealed that, compared to G1, G4 exhibited higher abundance of proteins (e.g., Histone H2A, cytochrome c oxidase, profilin) involved in structural integrity and energy homeostasis, whereas G6 showed signatures of oxidative stress and metabolic dysfunction (lower abundance of NADH dehydrogenase and higher abundance of calcium-transporting ATPase and glutathione S-transferase). In conclusion, 100 mmol\/L trehalose was associated with improved cryopreservation outcomes, and the proteins identified provide a basis for further investigation. This approach offers a framework for refining germplasm conservation strategies in aquaculture.<\/jats:p>","DOI":"10.3390\/ani16081245","type":"journal-article","created":{"date-parts":[[2026,4,20]],"date-time":"2026-04-20T07:15:08Z","timestamp":1776669308000},"page":"1245","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Cryogenic Damage and Trehalose Protection in Culter alburnus Sperm: An Integrated Assessment of Quality, Physiology, and Protein Expression"],"prefix":"10.3390","volume":"16","author":[{"given":"Shun","family":"Cheng","sequence":"first","affiliation":[{"name":"Key Laboratory of Healthy Freshwater Aquaculture, Zhejiang Institute of Freshwater Fisheries (Zhejiang Freshwater Fishery Environmental Monitoring Station), Ministry of Agriculture and Rural Affairs, Huzhou 313001, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3929-649X","authenticated-orcid":false,"given":"Shi-Li","family":"Liu","sequence":"additional","affiliation":[{"name":"Key Laboratory of Healthy Freshwater Aquaculture, Zhejiang Institute of Freshwater Fisheries (Zhejiang Freshwater Fishery Environmental Monitoring Station), Ministry of Agriculture and Rural Affairs, Huzhou 313001, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Mei-Li","family":"Chi","sequence":"additional","affiliation":[{"name":"Key Laboratory of Healthy Freshwater Aquaculture, Zhejiang Institute of Freshwater Fisheries (Zhejiang Freshwater Fishery Environmental Monitoring Station), Ministry of Agriculture and Rural Affairs, Huzhou 313001, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Wen-Ping","family":"Jiang","sequence":"additional","affiliation":[{"name":"Key Laboratory of Healthy Freshwater Aquaculture, Zhejiang Institute of Freshwater Fisheries (Zhejiang Freshwater Fishery Environmental Monitoring Station), Ministry of Agriculture and Rural Affairs, Huzhou 313001, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6795-0252","authenticated-orcid":false,"given":"Jian-Bo","family":"Zheng","sequence":"additional","affiliation":[{"name":"Key Laboratory of Healthy Freshwater Aquaculture, Zhejiang Institute of Freshwater Fisheries (Zhejiang Freshwater Fishery Environmental Monitoring Station), Ministry of Agriculture and Rural Affairs, Huzhou 313001, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Chao","family":"Zhu","sequence":"additional","affiliation":[{"name":"Key Laboratory of Healthy Freshwater Aquaculture, Zhejiang Institute of Freshwater Fisheries (Zhejiang Freshwater Fishery Environmental Monitoring Station), Ministry of Agriculture and Rural Affairs, Huzhou 313001, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jun-Zhi","family":"Luo","sequence":"additional","affiliation":[{"name":"Key Laboratory of Healthy Freshwater Aquaculture, Zhejiang Institute of Freshwater Fisheries (Zhejiang Freshwater Fishery Environmental Monitoring Station), Ministry of Agriculture and Rural Affairs, Huzhou 313001, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Fei","family":"Li","sequence":"additional","affiliation":[{"name":"Key Laboratory of Healthy Freshwater Aquaculture, Zhejiang Institute of Freshwater Fisheries (Zhejiang Freshwater Fishery Environmental Monitoring Station), Ministry of Agriculture and Rural Affairs, Huzhou 313001, China"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2026,4,18]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"444","DOI":"10.1111\/jai.13828","article-title":"Effects of 17\u03b1-methyltestosterone on the growth performance, development, and reproduction of gynogenetic topmouth culter (Culter alburnus Basilewsky)","volume":"35","author":"Chi","year":"2018","journal-title":"J. Appl. Ichthyol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"791","DOI":"10.1007\/s12562-019-01339-7","article-title":"Molecular characterization and expression analysis of cyp19a gene in Culter alburnus","volume":"85","author":"Zheng","year":"2019","journal-title":"Fish. Sci."},{"key":"ref_3","first-page":"483","article-title":"Cryopreservation of spermatozoa of silver carp, common carp, blunt snout bream and grass","volume":"38","author":"Chen","year":"1992","journal-title":"Acta Zool. Sin."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1672265","DOI":"10.1155\/2023\/1672265","article-title":"Preliminary analysis of quality and cryopreservation of sex-reversed female Culter alburnus sperm","volume":"2023","author":"Cheng","year":"2023","journal-title":"Aquac. Res."},{"key":"ref_5","first-page":"78","article-title":"Study on cyopreservation of sperm of Culter alburnus (\u2640) \u00d7 Magalobrame tarminalis (\u2642), Culter alburnus, and Magalobrame tarminalis","volume":"5","author":"Cheng","year":"2025","journal-title":"Sci. Fish Farming"},{"key":"ref_6","first-page":"49","article-title":"Proteomic analysis of extracellular medium of cryopreserved carp (Cyprinus carpio L.) semen","volume":"15","author":"Mariola","year":"2015","journal-title":"Comp. Biochem. Physiol.-Part D Genom. Proteom."},{"key":"ref_7","first-page":"174","article-title":"Short-term storage of sperm in common carp from laboratory research to commercial production\u2014A review","volume":"16","author":"Nururshopa","year":"2023","journal-title":"Rev. Aquac."},{"key":"ref_8","unstructured":"Jiang, X.R., Huo, X.Y., Huang, J.H., and Sun, Z.T. (2016). Biological Fermentation Industry Technology, China Light Industry Press."},{"key":"ref_9","first-page":"30","article-title":"The character and function of trehalose and its application","volume":"23","author":"Jin","year":"2015","journal-title":"Fine Spec."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"457","DOI":"10.1111\/j.1444-2906.2005.00985.x","article-title":"Cryopreservation of kelp grouper Epinephelus moara sperm using only a trehalose solution","volume":"71","author":"Miyaki","year":"2005","journal-title":"Fish. Sci."},{"key":"ref_11","first-page":"3","article-title":"Effect of trehalose on sperm cryopreservation of Acipenser schrenckii and preliminary study on the mechanism of its cryoinjury","volume":"45","author":"Chen","year":"2023","journal-title":"Mar. Fish."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"756","DOI":"10.1111\/j.1439-0426.2010.01545.x","article-title":"Quantitative semen parameters of Atlantic cod (Gadus morhua) and their physiological relationships with sperm activity and morphology","volume":"26","author":"Butts","year":"2010","journal-title":"J. Appl. Ichthyol."},{"key":"ref_13","first-page":"110","article-title":"The research review of prophenoloxidase activating system in Crustacean","volume":"30","author":"Meng","year":"1999","journal-title":"Oceanol. Limnol. Sin."},{"key":"ref_14","first-page":"158","article-title":"Effects of high level chromiumon on antioxidant enzyme system in gill and hepatopancreas of Procambarus","volume":"4","author":"Tan","year":"2007","journal-title":"J. Agro-Environ. Sci."},{"key":"ref_15","first-page":"143","article-title":"Research progress on function of catalase and its application in livestock and poultry production","volume":"45","author":"Zhu","year":"2022","journal-title":"Feed. Res."},{"key":"ref_16","first-page":"170","article-title":"Effects of cryopreservation on enzyme activity of Monopterus albus spermatozoa","volume":"35","author":"Yan","year":"2011","journal-title":"Acta Hydrobiol. Sin."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1046\/j.1439-0272.2001.00369.x","article-title":"Computerized analysis of cytochemical reactions for dehydrogenases and oxygraphic studies as methods to evaluate the function of the mitochondrial sheath in rat spermatozoa","volume":"33","author":"Piasecka","year":"2010","journal-title":"Andrologia"},{"key":"ref_18","first-page":"3","article-title":"Observation on freezing damage of sperm and embryos from several Chinese carps by scanning electron microscope","volume":"5","author":"Zhao","year":"1992","journal-title":"Freshw. Fish."},{"key":"ref_19","first-page":"286","article-title":"Ultrastructure of spermatozoa and effects of salinity temperature and pH on spermatozoa motility in Epinphelus coioides","volume":"10","author":"Zhao","year":"2003","journal-title":"J. Fish. Sci. China"},{"key":"ref_20","first-page":"20841","article-title":"Effects of trehalose, bovine serum albumin, and sucrose, on the integrity of the plasma membrane of Pseudosciaena crocea semen after cryopreservation","volume":"68","author":"Cheng","year":"2016","journal-title":"Isr. J. Aquac.-Bamidgeh"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"370","DOI":"10.1007\/BF02842852","article-title":"Evaluation of the damage in fish spermatozoa cryopreservation","volume":"24","author":"Li","year":"2006","journal-title":"Chin. J. Oceanol. Limnol."},{"key":"ref_22","first-page":"355","article-title":"Ultrastructure and ultralow temperature freezing effect of spermatozoon in black rockfish Sebastes schlegelii","volume":"35","author":"Zhang","year":"2020","journal-title":"J. Dalian Ocean. Univ."},{"key":"ref_23","first-page":"547","article-title":"Study of sperm cryopreservation in Silurus lanzhouensis and detection of cell damages after cryopreservation","volume":"45","author":"Xing","year":"2021","journal-title":"Acta Hydrobiol. Sin."},{"key":"ref_24","first-page":"291","article-title":"Physiological characteristics and ultrastructure of post-thaw sperm in turbot Scophthalmus maximus","volume":"44","author":"Zhang","year":"2013","journal-title":"Oceanol. Limnol. Sin."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"325","DOI":"10.3724\/SP.J.1035.2011.00325","article-title":"Effect of cryopreservation and short-term storage of Chinese giant salamander sperm","volume":"35","author":"Peng","year":"2011","journal-title":"Acta Hydrobiol. Sin."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"e12968","DOI":"10.1111\/raq.12968","article-title":"Influence of Dietary Fatty Acids on Fish Sperm Tolerance to Cryopreservation","volume":"17","author":"Villalobos","year":"2025","journal-title":"Rev. Aquac."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"124","DOI":"10.1016\/j.jprot.2016.02.029","article-title":"Motility of carp spermatozoa is associated with profound changes in the sperm proteome","volume":"138","author":"Dietrich","year":"2016","journal-title":"J. Proteom."},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Yang, Y., Wang, T., Yang, S., Wu, X., Huang, W.H., Fan, B., Xia, J.H., Meng, Z.N., and Liu, X.C. (2021). Corrigendum: The effects of storage in vitro on functions, transcriptome, proteome, and oxidation resistance of giant grouper sperm. Front. Mar. Sci., 8.","DOI":"10.3389\/fmars.2021.780244"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"359","DOI":"10.1038\/nmeth.1322","article-title":"Universal sample preparation method for proteome analysis","volume":"6","author":"Zougman","year":"2009","journal-title":"Nat. Methods"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1096","DOI":"10.2164\/jandrol.112.016535","article-title":"Reactive oxygen species and sperm function-in sickness and in health","volume":"33","author":"Aitken","year":"2012","journal-title":"J. Androl."},{"key":"ref_31","first-page":"438","article-title":"Effects of cryopreservation on enzyme activity of spermatozoa in Pseudosiaena crocea","volume":"34","author":"Huang","year":"2012","journal-title":"Mar. Fish."},{"key":"ref_32","first-page":"27","article-title":"Proteomic analysis between fresh and freeze-thawing semen in Acipenser schrenckii","volume":"49","author":"Chen","year":"2025","journal-title":"J. Fish. China"},{"key":"ref_33","first-page":"297","article-title":"Cryopreservational effects on the spermatic enzyme\u2019s activities of Japaneseell (Anguilla japonica)","volume":"30","author":"Huang","year":"2008","journal-title":"Mar. Fish."},{"key":"ref_34","first-page":"127","article-title":"Sperm Cryopreservation and Enzyme Activities Detection in Acrossocheilus fasciatus","volume":"25","author":"Wang","year":"2017","journal-title":"J. Agric. Biotechnol."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"347","DOI":"10.1016\/S0015-0282(16)59131-6","article-title":"Correlation between semen adenosine triphosphate and sperm fertilizing capacity","volume":"47","author":"Chan","year":"1987","journal-title":"Fertil. Steril."},{"key":"ref_36","first-page":"60","article-title":"Vitality and ultrastructure observation of fresh and cryopreservated sperm in Pseudosciaena crocea","volume":"44","author":"Cheng","year":"2013","journal-title":"Oceanol. Limnol. Sin."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"1013","DOI":"10.1071\/RD9961013","article-title":"Liquid storage of ram semen in the absence or presence of some antioxidants","volume":"8","author":"Maxwell","year":"1996","journal-title":"Reprod. Fertil. Dev."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"314","DOI":"10.1016\/j.anireprosci.2010.01.010","article-title":"Antioxidant systems of brown trout (Salmo trutta f. fario) semen","volume":"119","author":"Lahnsteiner","year":"2010","journal-title":"Anim. Reprod. Sci."},{"key":"ref_39","first-page":"50","article-title":"Effects of long-term cryopreservation on ultrastructure and enzyme activity of Epinephelus lanceolatus sperm","volume":"47","author":"Ding","year":"2023","journal-title":"J. Fish. China"},{"key":"ref_40","first-page":"22","article-title":"Ultrastructural changes of cryopreserved human spermatozoa","volume":"9","author":"Yu","year":"1995","journal-title":"Natl. J. Androl."},{"key":"ref_41","first-page":"526","article-title":"The morphological and ultrastructural variation of pacific oyster (Crassostrea gigas (Thunberg)) sperm after cryopreservation","volume":"32","author":"Li","year":"2002","journal-title":"J. Qingdao Ocean. Univ."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"59","DOI":"10.1016\/j.semcdb.2022.03.011","article-title":"Histone H2A variants: Diversifying chromatin to ensure genome integrity","volume":"135","author":"Oberdoerffer","year":"2022","journal-title":"Semin. Cell Dev. Biol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"310","DOI":"10.1016\/S0005-2728(99)00022-5","article-title":"The reactions of copper proteins with nitric oxide","volume":"1411","author":"Torres","year":"1999","journal-title":"BBA-Bioenergetics"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"155","DOI":"10.1016\/bs.ircmb.2020.05.005","article-title":"Profilin choreographs actin and microtubules in cells and cancer","volume":"355","author":"Pimm","year":"2020","journal-title":"Int. Rev. Cell Mol. Biol."},{"key":"ref_45","doi-asserted-by":"crossref","unstructured":"Maruf, A.A., Lee, O., and O\u2019Brien, P.J. (2014). Modifications of Mitochondrial Function by Toxicants. Reference Module in Biomedical Sciences, Elsevier.","DOI":"10.1016\/B978-0-12-801238-3.01900-0"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"366","DOI":"10.1016\/j.trsl.2014.07.001","article-title":"Protein phosphatase 1 catalytic isoforms: Specificity toward interacting proteins","volume":"164","author":"Esteves","year":"2014","journal-title":"Transl. Res."},{"key":"ref_47","first-page":"113139","article-title":"Small organic molecules targeting the energy metabolism of Mycobacterium tuberculosis","volume":"212","author":"Milan","year":"2020","journal-title":"Eur. J. Med. Chem."},{"key":"ref_48","doi-asserted-by":"crossref","unstructured":"Griffith, J., and Sansom, C. (1998). Calcium-transporting ATPase family. The Transporter FactsBook, Academic Press.","DOI":"10.1016\/B978-012303965-1\/50005-6"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"1392","DOI":"10.1016\/j.freeradbiomed.2009.02.025","article-title":"Glutathione S-transferase pi localizes in mitochondria and protects against oxidative stress","volume":"46","author":"Goto","year":"2009","journal-title":"Free. Radic. Biol. Med."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"618","DOI":"10.1016\/j.bbrc.2004.02.091","article-title":"RASL11A, member of a novel small monomeric GTPase gene family, is down-regulated in prostate tumors","volume":"316","author":"Louro","year":"2004","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"238","DOI":"10.1016\/0011-2240(88)90031-4","article-title":"Cryopreservation of sperm from the marine shrimp Sicyonia ingentis","volume":"25","author":"Anchordoguy","year":"1988","journal-title":"Cryobiology"},{"key":"ref_52","first-page":"114","article-title":"Trehalose and glycerol synergistically improve the cryopreservation of cashmere goat semen","volume":"57","author":"Song","year":"2021","journal-title":"Chin. J. Anim. Sci."},{"key":"ref_53","first-page":"256","article-title":"Effect of trehalose replacing partial glycerol on frozen-thawed sperm quality of Yanbian yellow cattle","volume":"49","author":"Chen","year":"2022","journal-title":"China Anim. Husb. Vet. Med."},{"key":"ref_54","first-page":"38","article-title":"Cryopreservation and short-term preservation of red spotted grouper (Epinephelus akaara) sperm","volume":"37","author":"Zheng","year":"2013","journal-title":"Mar. Sci."},{"key":"ref_55","first-page":"3744","article-title":"Establishment of cryopreservation methods for Fenneropenaeus chinensis sperm and investigation of ultrastructural damage","volume":"52","author":"Liu","year":"2025","journal-title":"China Anim. Husb. Vet. Med."},{"key":"ref_56","first-page":"1382","article-title":"Spermatozoa cryopreservation of Litopenaeus vannamei","volume":"44","author":"Yang","year":"2013","journal-title":"J. South. Agric."}],"container-title":["Animals"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-2615\/16\/8\/1245\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,4,20]],"date-time":"2026-04-20T07:39:07Z","timestamp":1776670747000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-2615\/16\/8\/1245"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,4,18]]},"references-count":56,"journal-issue":{"issue":"8","published-online":{"date-parts":[[2026,4]]}},"alternative-id":["ani16081245"],"URL":"https:\/\/doi.org\/10.3390\/ani16081245","relation":{},"ISSN":["2076-2615"],"issn-type":[{"value":"2076-2615","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,4,18]]}}}