{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T01:35:30Z","timestamp":1760146530467,"version":"build-2065373602"},"reference-count":16,"publisher":"MDPI AG","issue":"22","license":[{"start":{"date-parts":[[2024,11,11]],"date-time":"2024-11-11T00:00:00Z","timestamp":1731283200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100005880","name":"National Institute of Biological Resources","doi-asserted-by":"publisher","award":["NIBR202421101"],"award-info":[{"award-number":["NIBR202421101"]}],"id":[{"id":"10.13039\/501100005880","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Animals"],"abstract":"<jats:p>This study aims to develop an optimized method for cryopreserving the germ cells of Lissachatina fulica (L. fulica) using vitrification, as an alternative approach for conserving endangered snail species. First, we isolated several key reproductive organs, including the sperm oviduct, albumen gland, hermaphrodite gland (ovotestis), and hermaphrodite duct from L. fulica. When the ovotestis was finely chopped, numerous sperm with long tails and distinct heads were observed. The staining of sperm nuclei was confirmed using Hoechst 33342 dye. Since the hermaphrodite gland, referred to as the ovotestis, contains both male and female germ cells, we performed tissue staining on the ovotestis using hematoxylin and eosin (H&amp;E) dye. H&amp;E staining of the ovotestis revealed numerous oval-shaped acini containing sperm and early germ cells. Spermatocytes and spermatids were observed within distinct boundaries, with mature sperm appearing following spermatogenesis. To preserve the species of the L. fulica, we introduced vitrification technology to cryopreserve its reproductive organs. The non-vitrification group showed an average cell viability of 96.6%, while the vitrification group had 86.8% after thawing. This study presents a reliable cryopreservation protocol for L. fulica, with potential applications for other endangered snails, supporting conservation efforts to preserve genetic resources and biodiversity.<\/jats:p>","DOI":"10.3390\/ani14223229","type":"journal-article","created":{"date-parts":[[2024,11,11]],"date-time":"2024-11-11T08:46:54Z","timestamp":1731314814000},"page":"3229","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Development of Germ Cell Isolation and Optimal Cryopreservation Method for Lissachatina fulica (L. fulica)"],"prefix":"10.3390","volume":"14","author":[{"ORCID":"https:\/\/orcid.org\/0009-0004-7257-1061","authenticated-orcid":false,"given":"Jukyeong","family":"Jeong","sequence":"first","affiliation":[{"name":"Department of Biotechnology, College of Life and Applied Sciences, Yeungnam University, Gyeongsan 38541, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5035-4945","authenticated-orcid":false,"given":"Seungki","family":"Lee","sequence":"additional","affiliation":[{"name":"Biological and Genetic Resources Assessment Division, National Institute of Biological Resources, Incheon 22689, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4544-3668","authenticated-orcid":false,"given":"Jung Kyu","family":"Choi","sequence":"additional","affiliation":[{"name":"Department of Biotechnology, College of Life and Applied Sciences, Yeungnam University, Gyeongsan 38541, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2024,11,11]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Pellett, S., and O\u2019Brien, M. (2020). Giant African land snails. Handbook of Exotic Pet Medicine, John Wiley & Sons Ltd.","DOI":"10.1002\/9781119389934.ch24"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"693","DOI":"10.1007\/s10530-006-9069-6","article-title":"Rapid spread of an invasive snail in South America: The giant African snail, Achatina fulica, in Brasil","volume":"9","author":"Thiengo","year":"2007","journal-title":"Biol. Invasions"},{"key":"ref_3","unstructured":"Yap, C.K. (2017). Snails: Biodiversity, Biology and Behavioral Insights, Nova Science Publishers, Inc."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"205","DOI":"10.26832\/24566632.2020.0502019","article-title":"A review on persisting threats to snail\u2019s diversity and its Conservation approaches","volume":"5","author":"Dhiman","year":"2020","journal-title":"Arch. Agric. Environ. Sci."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"116","DOI":"10.1080\/13235818.2013.782793","article-title":"Australian land snails: A review of ecological research and conservation approaches","volume":"33","author":"Parkyn","year":"2013","journal-title":"Molluscan Res."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"126","DOI":"10.1093\/ilar.52.2.126","article-title":"Invertebrate models for biomedical research, testing, and education","volume":"52","year":"2011","journal-title":"ILAR J."},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Errajouani, F., Bakrim, H., Hourfane, S., Louajri, A., Rocha, J.M., El Aouad, N., and Laglaoui, A. (2023). Exploring the Potential Anti-Inflammatory and Wound-Healing Proprieties of Cepaea hortensis Snail Mucin. Cosmetics, 10.","DOI":"10.3390\/cosmetics10060170"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"e15827","DOI":"10.7717\/peerj.15827","article-title":"Biological properties of mucus from land snails (Lissachatina fulica) and freshwater snails (Pomacea canaliculata) and histochemical study of mucous cells in their foot","volume":"11","author":"Phrompanya","year":"2023","journal-title":"PeerJ"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"184","DOI":"10.1016\/j.jmau.2016.03.001","article-title":"Ultrastructure of ovotestis of young and adult pulmonate mollusk, Macrochlamys indica Benson","volume":"15","author":"Misra","year":"2016","journal-title":"J. Microsc. Ultrastruct."},{"key":"ref_10","first-page":"1066","article-title":"A brief review of parasitic castration in aquatic snails and its contribution in control of diverse vector snail populations and trematodiases in man and animals","volume":"9","author":"Choubisa","year":"2022","journal-title":"Austin J. Infect. Dis."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Jeong, J., Lee, S., and Choi, J.K. (2024). Ovotestis Isolation and Cryopreservation of Nesiohelix samarangae (Oriental Snail) as a Snail Model for Conserving Other Endangered Snail Species. Biology, 13.","DOI":"10.3390\/biology13040205"},{"key":"ref_12","first-page":"577","article-title":"Evaluation of euthanasia techniques for an invertebrate species, land snails (Succinea putris)","volume":"55","author":"Gilbertson","year":"2016","journal-title":"J. Am. Assoc. Lab. Anim. Sci."},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"L\u0151w, P., Moln\u00e1r, K., and Kriska, G. (2016). Atlas of Animal Anatomy and Histology, Springer.","DOI":"10.1007\/978-3-319-25172-1"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"6","DOI":"10.1007\/s10499-015-9900-0","article-title":"Sperm cryopreservation in marine mollusk: A review","volume":"23","author":"Liu","year":"2015","journal-title":"Aquac. Int."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"590","DOI":"10.1007\/s10126-021-10048-1","article-title":"Cryopreservation of Germ Cells of Banana Shrimp (Fenneropenaeus merguiensis) and Black Tiger Shrimp (Penaeus monodon)","volume":"23","author":"Rakbanjong","year":"2021","journal-title":"Mar. Biotechnol."},{"key":"ref_16","first-page":"493","article-title":"Sperm Cryopreservation in Fish and Shellfish","volume":"65","author":"Tiersch","year":"2007","journal-title":"Soc. Reprod. Fertil. Suppl."}],"container-title":["Animals"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-2615\/14\/22\/3229\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T16:30:21Z","timestamp":1760113821000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-2615\/14\/22\/3229"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,11,11]]},"references-count":16,"journal-issue":{"issue":"22","published-online":{"date-parts":[[2024,11]]}},"alternative-id":["ani14223229"],"URL":"https:\/\/doi.org\/10.3390\/ani14223229","relation":{},"ISSN":["2076-2615"],"issn-type":[{"type":"electronic","value":"2076-2615"}],"subject":[],"published":{"date-parts":[[2024,11,11]]}}}