{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,23]],"date-time":"2026-09-23T07:34:48Z","timestamp":1790148888126,"version":"4.0.1"},"reference-count":56,"publisher":"MDPI AG","issue":"19","license":[{"start":{"date-parts":[[2022,9,30]],"date-time":"2022-09-30T00:00:00Z","timestamp":1664496000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Technology System of Modern Agricultural Industry in Shandong Province","award":["SDAIT-10-08"],"award-info":[{"award-number":["SDAIT-10-08"]}]},{"name":"Technology System of Modern Agricultural Industry in Shandong Province","award":["1121010"],"award-info":[{"award-number":["1121010"]}]},{"name":"Start-up Fund for High-level Talents of Qingdao Agricultural University","award":["SDAIT-10-08"],"award-info":[{"award-number":["SDAIT-10-08"]}]},{"name":"Start-up Fund for High-level Talents of Qingdao Agricultural University","award":["1121010"],"award-info":[{"award-number":["1121010"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Animals"],"abstract":"<jats:p>Sperm cryopreservation contributes to the extensive utilization of artificial insemination (AI) in the daily livestock industry. However, due to the presence of few sperm with good biological function in post-thaw goat sperm, its use has been limited for AI purposes. Hence, its improvement has been the focus of many research studies. This study aimed to investigate the effects of proline supplementation of the freezing medium on goat sperm. The goat semen was cryopreserved with freezing medium supplementation of different concentrations of proline (0, 0.5, 1, 2 and 4 mM). The post-thaw sperm motility patterns, membrane integrity, acrosome integrity, lipid peroxidation (LPO) levels, malondialdehyde (MDA) levels, total antioxidant capacity (T-AOC), proline dehydrogenase (PRODH) activity, superoxide dis-mutase (SOD) activity, glutathione (GSH) levels and GSH\/GSSG were evaluated. Likewise, the expression and immunofluorescent localization of PRODH in post-thaw goat sperm was also detected. It was observed that addition of 2 mM proline to the freezing medium significantly enhanced post-thaw goat sperm total motility, progressive motility, straight-linear velocity (VSL), curvilinear velocity (VCL), average path velocity (VAP), straightness (STR), linearity (LIN), membrane integrity and acrosome integrity. Interestingly, PRODH was expressed in post-thaw goat sperm, especially in the post-acrosome and sperm tail. Addition of 2 mM proline also significantly increased the post-thaw sperm PRODH activity compared to the control. Moreover, post-thaw goat sperm LPO levels and MDA levels were reduced by supplementation of 2 mM proline. Furthermore, compared to the control, the values of post-thaw goat sperm T-AOC, SOD activity, GSH level and GSH\/GSSG were also significantly increased in 2 mM proline treatment. Reduction of post-thaw goat sperm apoptosis in 2 mM proline treatment was also observed as the levels of Caspase3 and Caspase9 were decreased by the supplementation with 2 mM proline. These observations suggest that the addition of 2 mM proline to the freezing medium increased post-thaw goat sperm quality by reducing oxidative stress during cryopreservation. These findings also provide novel insights into the use of proline as an efficient additive to enhance post-thaw goat sperm quality during cryopreservation.<\/jats:p>","DOI":"10.3390\/ani12192626","type":"journal-article","created":{"date-parts":[[2022,10,7]],"date-time":"2022-10-07T22:47:27Z","timestamp":1665182847000},"page":"2626","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":34,"title":["Beneficial Effect of Proline Supplementation on Goat Spermatozoa Quality during Cryopreservation"],"prefix":"10.3390","volume":"12","author":[{"given":"Weijing","family":"Zhang","sequence":"first","affiliation":[{"name":"College of Animal Science and Technology, Qingdao Agricultural University, Qingdao 266109, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lingjiang","family":"Min","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Qingdao Agricultural University, Qingdao 266109, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yajing","family":"Li","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Qingdao Agricultural University, Qingdao 266109, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yaning","family":"Lang","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Qingdao Agricultural University, Qingdao 266109, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3537-5892","authenticated-orcid":false,"given":"S. A. Masudul","family":"Hoque","sequence":"additional","affiliation":[{"name":"Department of Animal Breeding and Genetics, Faculty of Veterinary Medicine and Animal Science, Bangabandhu Sheikh Mujibur Rahman Agricultural University, Gazipur 1706, Bangladesh"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6611-126X","authenticated-orcid":false,"given":"Adedeji Olufemi","family":"Adetunji","sequence":"additional","affiliation":[{"name":"Department of Animal Sciences, North Carolina Agricultural and Technical State University, Greensboro, NC 27411, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhendong","family":"Zhu","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Qingdao Agricultural University, Qingdao 266109, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2022,9,30]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1002\/j.1939-4640.2000.tb03268.x","article-title":"Semen cryopreservation in domestic animals: A damaging and capacitating phenomenon","volume":"21","author":"Bailey","year":"2000","journal-title":"J. Androl."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"40","DOI":"10.1016\/j.cryobiol.2020.05.001","article-title":"Evaluation of used Purslane extracts in Tris extenders on cryopreserved goat sperm","volume":"94","author":"Azimi","year":"2020","journal-title":"Cryobiology"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"85","DOI":"10.1095\/biolreprod.115.128553","article-title":"Cholesterol-Loaded Cyclodextrin Increases the Cholesterol Content of Goat Sperm to Improve Cold and Osmotic Resistance and Maintain Sperm Function after Cryopreservation","volume":"94","author":"Salmon","year":"2016","journal-title":"Biol. Reprod."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"787","DOI":"10.1530\/REP-11-0436","article-title":"Osmotic stress and cryoinjury of koala sperm: An integrative study of the plasma membrane, chromatin stability and mitochondrial function","volume":"143","author":"Johnston","year":"2012","journal-title":"Reproduction"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"852","DOI":"10.1095\/biolreprod.110.085852","article-title":"Evaluating the Impacts of Osmotic and Oxidative Stress on Common Carp (Cyprinus carpio L.) Sperm Caused by Cryopreservation Techniques","volume":"83","author":"Li","year":"2010","journal-title":"Biol. Reprod."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"258","DOI":"10.1016\/j.smallrumres.2013.07.007","article-title":"The cryoprotective effects of amino acids supplementation on cooled and post-thaw Markhoz bucks semen quality","volume":"114","author":"Farshad","year":"2013","journal-title":"Small Rumin. Res."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"266","DOI":"10.1016\/j.theriogenology.2018.03.041","article-title":"Anti-apoptotic effects of minocycline on ram epididymal spermatozoa exposed to oxidative stress","volume":"114","author":"Merati","year":"2018","journal-title":"Theriogenology"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"401","DOI":"10.5424\/sjar\/2017151-9761","article-title":"Alpha-tocopherol improves frozen-thawed sperm quality by reducing hydrogen peroxide during cryopreservation of bull semen","volume":"15","author":"Motemani","year":"2017","journal-title":"Span. J. Agric. Res."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"403","DOI":"10.1007\/s10815-008-9232-3","article-title":"Effects of cryopreservation on sperm parameters and ultrastructural morphology of human spermatozoa","volume":"25","author":"Ozkavukcu","year":"2008","journal-title":"J. Assist. Reprod. Genet."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"327","DOI":"10.1016\/S0093-691X(01)00674-4","article-title":"Current status of sperm cryopreservation: Why isn\u2019t it better?","volume":"57","author":"Medeiros","year":"2002","journal-title":"Theriogenology"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"412","DOI":"10.1016\/j.theriogenology.2011.08.015","article-title":"Comparison of two commercial extenders for cryopreservation of goat semen without sperm washing","volume":"77","author":"Roof","year":"2012","journal-title":"Theriogenology"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"155","DOI":"10.1046\/j.1365-2605.1999.00162.x","article-title":"Effect of freeze\u2013thawing procedure on chromatin stability, morphological alteration and membrane integrity of human spermatozoa in fertile and subfertile men","volume":"22","author":"Hammadeh","year":"1999","journal-title":"Int. J. Androl."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"19","DOI":"10.1186\/s12014-018-9195-z","article-title":"Comparative analysis of human sperm glycocalyx from different freezability ejaculates by lectin microarray and identification of ABA as sperm freezability biomarker","volume":"15","author":"Xin","year":"2018","journal-title":"Clin. Proteom."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"59211503","DOI":"10.1155\/2019\/5921503","article-title":"Resveratrol Improves Boar Sperm Quality via 5\u2032AMP-Activated Protein Kinase Activation during Cryopreservation","volume":"2019","author":"Zhu","year":"2019","journal-title":"Oxidative Med. Cell. Longev."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"851","DOI":"10.2174\/0929867013373039","article-title":"Relative Impact of Oxidative Stress on Male Reproductive Function","volume":"8","author":"Sikka","year":"2001","journal-title":"Curr. Med. Chem."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1061","DOI":"10.1002\/j.1939-4640.2001.tb03446.x","article-title":"Osmotic Tolerance of Equine Spermatozoa and the Effects of Soluble Cryoprotectants on Equine Sperm Motility, Viability, and Mitochondrial Membrane Potential","volume":"22","author":"Ball","year":"2001","journal-title":"J. Androl."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"57","DOI":"10.1016\/j.jevs.2017.09.006","article-title":"Addition of Antioxidants Myoinositol, Ferulic Acid, and Melatonin and Their Effects on Sperm Motility, Membrane Integrity, and Reactive Oxygen Species Production in Cooled Equine Semen","volume":"59","author":"Affonso","year":"2017","journal-title":"J. Equine Veter. Sci."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1379","DOI":"10.1007\/s10815-019-01464-3","article-title":"Decreased very long chain polyunsaturated fatty acids in sperm correlates with sperm quantity and quality","volume":"36","author":"Craig","year":"2019","journal-title":"J. Assist. Reprod. Genet."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"771440","DOI":"10.3389\/fvets.2021.771440","article-title":"Effect of Glutathione on Sperm Quality in Guanzhong Dairy Goat Sperm During Cryopreservation","volume":"8","author":"Zou","year":"2021","journal-title":"Front. Veter. Sci."},{"key":"ref_20","first-page":"1","article-title":"The effects of green synthesized anionic cupric oxide nanoparticles on Zaraibi goat spermatozoa during cryopreservation with and without removal of seminal plasma","volume":"5","author":"Elshamy","year":"2022","journal-title":"Anim. Biotechnol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"29","DOI":"10.1016\/j.theriogenology.2019.02.011","article-title":"Lycium barbarum and Laminaria japonica polysaccharides improve Cashmere goat sperm quality and fertility rate after cryopreservation","volume":"129","author":"Ren","year":"2019","journal-title":"Theriogenology"},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Hassan, S.A., Khalil, W.A., Hassan, M.A.E., Yousif, A.I., Sabry, O.M., Wink, M., and Sobeh, M. (2021). Antioxidant and Antiapoptotic Effects of a Turraea fischeri Leaf Extract on Cryopreserved Goat Sperm. Animals, 11.","DOI":"10.3390\/ani11102840"},{"key":"ref_23","first-page":"146","article-title":"Effect of Idebenone, Resveratrol and Taurine on the Sperm Quality and Lipid Peroxidation of Cryopreserved Crossbred Ram Semen","volume":"42","author":"Naikoo","year":"2021","journal-title":"CryoLetters"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"151","DOI":"10.1016\/j.cryobiol.2021.02.004","article-title":"Supplementary role of vitamin E and amino acids added to diluent on goat sperm freezability","volume":"100","author":"Merati","year":"2021","journal-title":"Cryobiology"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"172","DOI":"10.1016\/j.cryobiol.2020.11.005","article-title":"Protective effects of Tribulus terrestris and Cinnamomum zeylanicum extracts and trehalose added to diluents on goat epididymal sperm freezability","volume":"98","author":"Ariyan","year":"2021","journal-title":"Cryobiology"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"876","DOI":"10.1111\/rda.14131","article-title":"Synergistic effects of myo-inositol and melatonin on cryopreservation of goat spermatozoa","volume":"57","author":"Vash","year":"2022","journal-title":"Reprod. Domest. Anim."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"89","DOI":"10.1016\/j.tplants.2009.11.009","article-title":"Proline: A multifunctional amino acid","volume":"15","author":"Szabados","year":"2010","journal-title":"Trends Plant Sci."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"1053","DOI":"10.1007\/s00726-010-0715-z","article-title":"Proline and hydroxyproline metabolism: Implications for animal and human nutrition","volume":"40","author":"Wu","year":"2011","journal-title":"Amino Acids"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"671","DOI":"10.1016\/j.freeradbiomed.2007.10.054","article-title":"Proline modulates the intracellular redox environment and protects mammalian cells against oxidative stress","volume":"44","author":"Krishnan","year":"2008","journal-title":"Free Radic. Biol. Med."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"3459","DOI":"10.1073\/pnas.0407960102","article-title":"From The Cover: Proline suppresses apoptosis in the fungal pathogen Colletotrichum trifolii","volume":"102","author":"Chen","year":"2005","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"1181","DOI":"10.1016\/j.freeradbiomed.2012.07.002","article-title":"Proline dehydrogenase is essential for proline protection against hydrogen peroxide-induced cell death","volume":"53","author":"Natarajan","year":"2012","journal-title":"Free Radic. Biol. Med."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"347","DOI":"10.1071\/R98053","article-title":"Effect of compatible solutes and diluent composition on the post-thaw motility of ram sperm","volume":"10","author":"Setchell","year":"1998","journal-title":"Reprod. Fertil. Dev."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"103393","DOI":"10.1016\/j.jevs.2021.103393","article-title":"L-Proline: An Effective Agent for Frozen and Post-thawed Donkey Semen Storage","volume":"101","author":"Li","year":"2021","journal-title":"J. Equine Veter. Sci."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"26326","DOI":"10.1038\/srep26326","article-title":"L-proline: A highly effective cryoprotectant for mouse oocyte vitrification","volume":"6","author":"Zhang","year":"2016","journal-title":"Sci. Rep."},{"key":"ref_35","first-page":"485","article-title":"Compatible solutes improve cryopreservation of human endothelial cells","volume":"33","author":"Sun","year":"2012","journal-title":"CryoLetters"},{"key":"ref_36","doi-asserted-by":"crossref","unstructured":"Feng, C., Zhu, Z., Bai, W., Li, R., Zheng, Y., Tian, X., Wu, D., Lu, H., Wang, Y., and Zeng, W. (2020). Proline Protects Boar Sperm Against Oxidative Stress through Proline Dehydrogenase-Mediated Metabolism and the Amine Structure of Pyrrolidine. Animals, 10.","DOI":"10.3390\/ani10091549"},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Zhu, Z., Ren, Z., Fan, X., Pan, Y., Lv, S., Pan, C., Lei, A., and Zeng, W. (2017). Cysteine protects rabbit spermatozoa against reactive oxygen species-induced damages. PLoS ONE, 12.","DOI":"10.1371\/journal.pone.0181110"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"166","DOI":"10.5713\/ab.21.0151","article-title":"Cysteine improves boar sperm quality via glutathione biosynthesis during the liquid storage","volume":"35","author":"Zhu","year":"2022","journal-title":"Anim. Biosci."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"856536","DOI":"10.3389\/fvets.2022.856536","article-title":"Reducing the Glucose Level in Pre-treatment Solution Improves Post-thaw Boar Sperm Quality","volume":"9","author":"Zhu","year":"2022","journal-title":"Front. Veter. Sci."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"266","DOI":"10.1111\/jog.13818","article-title":"Recent advances in fertility preservation","volume":"45","author":"Dolmans","year":"2019","journal-title":"J. Obstet. Gynaecol. Res."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"451","DOI":"10.1002\/mrd.1052","article-title":"Production of reactive oxygen species by spermatozoa undergoing cooling, freezing, and thawing","volume":"59","author":"Chatterjee","year":"2001","journal-title":"Mol. Reprod. Dev."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"87","DOI":"10.1080\/2090598X.2019.1599624","article-title":"Oxidative stress and sperm function: A systematic review on evaluation and management","volume":"17","author":"Dutta","year":"2019","journal-title":"Arab J. Urol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"1251","DOI":"10.1111\/rda.13508","article-title":"Effect of nicotinic acid on the plasma membrane function and polyunsaturated fatty acids composition during cryopreservation in boar sperm","volume":"54","author":"Lee","year":"2019","journal-title":"Reprod. Domest. Anim."},{"key":"ref_44","first-page":"523","article-title":"[The redox system in human semen and peroxidative damage of spermatozoa]","volume":"59","author":"Fraczek","year":"2005","journal-title":"Postepy. Hig. Med. Dosw."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"79","DOI":"10.1016\/j.anireprosci.2014.02.009","article-title":"Effect of extender and amino acid supplementation on sperm quality of cooled-preserved Andalusian donkey (Equus asinus) spermatozoa","volume":"146","author":"Dorado","year":"2014","journal-title":"Anim. Reprod. Sci."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"159","DOI":"10.1002\/ajp.10073","article-title":"Effect of amino acids on cryopreservation of cynomolgus monkey (Macaca fascicularis) sperm","volume":"59","author":"Li","year":"2003","journal-title":"Am. J. Primatol."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"423","DOI":"10.1006\/cryo.1993.1042","article-title":"Effect of Sucrose, Trehalose, Hypotaurine, Taurine, and Blood Serum on Survival of Frozen Bull Sperm","volume":"30","author":"Chen","year":"1993","journal-title":"Cryobiology"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"e14301","DOI":"10.1111\/and.14301","article-title":"L-proline as a novel additive to cryopreservation media improved post-thaw quality of human spermatozoon via reducing oxidative stress","volume":"54","author":"Moradi","year":"2022","journal-title":"Andrologia"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1006\/cryo.1998.2092","article-title":"Improvement of Ram Sperm Cryopreservation Protocols Assessed by Sperm Quality Parameters and Heterogeneity Analysis","volume":"37","author":"Ollero","year":"1998","journal-title":"Cryobiology"},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"122","DOI":"10.1016\/j.cryobiol.2020.11.012","article-title":"Effect of sulforaphane on apoptosis, reactive oxygen species and lipids peroxidation of human sperm during cryopreservation","volume":"99","author":"Valipour","year":"2021","journal-title":"Cryobiology"},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"257","DOI":"10.1016\/j.anireprosci.2008.04.014","article-title":"Oxidative stress, osmotic stress and apoptosis: Impacts on sperm function and preservation in the horse","volume":"107","author":"Ball","year":"2008","journal-title":"Anim. Reprod. Sci."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"913","DOI":"10.1007\/s10495-007-0756-2","article-title":"Mitochondria, oxidative stress and cell death","volume":"12","author":"Ott","year":"2007","journal-title":"Apoptosis"},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"451","DOI":"10.1016\/j.cmet.2012.02.013","article-title":"Impaired Insulin\/IGF1 Signaling Extends Life Span by Promoting Mitochondrial L-Proline Catabolism to Induce a Transient ROS Signal","volume":"15","author":"Zarse","year":"2012","journal-title":"Cell Metab."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"297","DOI":"10.1111\/ppl.12563","article-title":"Sterol glycosyltransferases required for adaptation of Withania somnifera at high temperature","volume":"160","author":"Singh","year":"2017","journal-title":"Physiol. Plant."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"689","DOI":"10.1080\/10715760802317663","article-title":"Glutathione and apoptosis","volume":"42","author":"Circu","year":"2008","journal-title":"Free Radic. Res."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"1767","DOI":"10.1016\/j.bbamcr.2012.06.019","article-title":"Glutathione and modulation of cell apoptosis","volume":"1823","author":"Circu","year":"2012","journal-title":"Biochim. Biophys. Acta"}],"container-title":["Animals"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-2615\/12\/19\/2626\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T00:44:22Z","timestamp":1760143462000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-2615\/12\/19\/2626"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,9,30]]},"references-count":56,"journal-issue":{"issue":"19","published-online":{"date-parts":[[2022,10]]}},"alternative-id":["ani12192626"],"URL":"https:\/\/doi.org\/10.3390\/ani12192626","relation":{},"ISSN":["2076-2615"],"issn-type":[{"value":"2076-2615","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,9,30]]}}}