Tissue storage manual for contaminant analysis in sea turtles
DOI:
https://doi.org/10.11606/issn.1678-4456.bjvras.2025.230332Keywords:
Sea turtles, Microplastics, Pollutants, Protocol, Sample TreatmentAbstract
Necropsies are essential tools for determining the cause of death in wildlife, including marine species like sea turtles. Through laboratory analysis of well-preserved tissues, wildlife disease specialists can investigate various potential causes of mortality in wildlife. The primary goal is to assist specialists in accurately identifying turtle organs and gathering the samples required for pathology and other vital tests. This protocol outlines methods for collecting ingested waste from sea turtles, as well as organizing and storing data related to these samples. The procedure starts with the meticulous gathering of deceased sea turtles, after which tissue samples are sorted according to their level of decomposition. Turtle necropsies should be conducted in certified facilities equipped for these procedures, ensuring compliance with established protocols and guidelines. Moreover, the protocol outlined here provides detailed guidance on the optimal methods for preserving tissue samples, tailored to the type of contaminant (microplastics, heavy metals, pesticides, or persistent organic pollutants) being analyzed. This contributes to a deeper understanding of the health and survival challenges faced by marine wildlife.
Downloads
References
Agusa T, Takagi K, Kubota R, Anan Y, Iwata H, Tanabe S. Specific accumulation of arsenic compounds in green turtles (Chelonia mydas) and hawksbill turtles (Eretmochelys imbricata) from Ishigaki Island, Japan. Environ Pollut. 2008;153(1):127-36. http://doi.org/10.1016/j.envpol.2007.07.013. PMid:17728031.
Alam SK, Brim MS. Organochlorine, PCB, PAH, and metal concentrations in eggs of loggerhead sea turtles (Caretta caretta) from northwest Florida, USA. J Environ Sci Health B. 2000;35(6):705-24. http://doi.org/10.1080/03601230009373303. PMid:11069014.
Aranda DA, Sindou P, Cauich Rodriguez JV, Saldaña GM, Coronado RFV, González WDN, Díaz ME, Escalante VC. A non-invasive method of microplastics pollution quantification in green sea turtle Chelonia mydas of the Mexican Caribbean. Mar Pollut Bull. 2024;200:116092. http://doi.org/10.1016/j.marpolbul.2024.116092. PMid:38359479.
Avery-Gomm S, Borrelle SB, Provencher JF. Linking plastic ingestion research with marine wildlife conservation. Sci Total Environ. 2018;637-638:1492-5. http://doi.org/10.1016/j.scitotenv.2018.04.409. PMid:29801242.
Barraza AD, Komoroske LM, Allen CD, Eguchi T, Gossett R, Holland E, Lawson DD, LeRoux RA, Lorenzi V, Seminoff JA, Lowe CG. Persistent organic pollutants in green sea turtles (Chelonia mydas) inhabiting two urbanized Southern California habitats. Mar Pollut Bull. 2020;153:110979. http://doi.org/10.1016/j.marpolbul.2020.110979. PMid:32275536.
Batista KA, Franciscone Mendes P. A virtópsia como complemento da necrópsia tradicional na medicina veterinária forense. Pubvet. 2022;16(7). http://doi.org/10.31533/pubvet.v16n07a1161.1-13.
Beckwith VK, Fuentes MMPB. Microplastic at nesting grounds used by the northern Gulf of Mexico loggerhead recovery unit. Mar Pollut Bull. 2018;131(Pt A):32-7. http://doi.org/10.1016/j.marpolbul.2018.04.001. PMid:29886953.
Bolten AB. Techniques for measuring sea turtles. In: Eckert KL, Bjorndal KA, Abreu-Grobois FA, Donnelly M, editors. Research and managment techniques for the conservation of sea turtles. Washington, DC: IUCN/SSC Marine Turtle Specialist Group; 1999. (IUCN/SSC Marine Turtle Specialist Group Publication, 4).
Bonanno G, Orlando-Bonaca M. Perspectives on using marine species as bioindicators of plastic pollution. Mar Pollut Bull. 2018;137:209-21. http://doi.org/10.1016/j.marpolbul.2018.10.018. PMid:30503427.
Browne MA, Crump P, Niven SJ, Teuten E, Tonkin A, Galloway T, Thompson R. Accumulation of microplastic on shorelines woldwide: sources and sinks. Environ Sci Technol. 2011;45(21):9175-9. http://doi.org/10.1021/es201811s. PMid:21894925.
Carr A. Impact of nondegradable marine debris on the ecology and survival outlook of sea turtles. Mar Pollut Bull. 1987;18(6):352-6. http://doi.org/10.1016/S0025-326X (87)80025-5.
Chaloupka M, Work TM, Balazs GH, Murakawa SKK, Morris R. Cause-specific temporal and spatial trends in green sea turtle strandings in the Hawaiian Archipelago (1982-2003). Mar Biol. 2008;154(5):887-98. http://doi.org/10.1007/s00227-008-0981-4.
Chen Y, Xia Z, Li H. Comparative analysis of the fecal bacterial communities of hawksbill sea turtles (Eretmochelys imbricata) and green sea turtles (Chelonia mydas). FEMS Microbiol Lett. 2022;369(1):fnac073. http://doi.org/10.1093/femsle/fnac073. PMid:35945331.
Clukey KE, Lepczyk CA, Balazs GH, Work TM, Li QX, Bachman MJ, Lynch JM. Persistent organic pollutants in fat of three species of Pacific pelagic longline caught sea turtles: accumulation in relation to ingested plastic marine debris. Sci Total Environ. 2018;610–611:402- 11. http://doi.org/10.1016/j.scitotenv.2017.07.242. PMid:28806556.
Colferai AS, Silva-Filho RP, Martins AM, Bugoni L. Distribution pattern of anthropogenic marine debris along the gastrointestinal tract of green turtles (Chelonia mydas) as implications for rehabilitation. Mar Pollut Bull. 2017;119(1):231-7. http://doi.org/10.1016/j.marpolbul.2017.03.053. PMid:28408048.
Eastman CB, Farrell JA, Whitmore L, Rollinson Ramia DR, Thomas RS, Prine J, Eastman SF, Osborne TZ, Martindale MQ, Duffy DJ. Plastic ingestion in post-hatchling sea turtles: assessing a major threat in Florida near shore waters. Front Mar Sci. 2020;7:693. http://doi.org/10.3389/fmars.2020.00693.
Eckert KL, Bjorndal KA, Abreu-Grobois FA, Donnelly M, editors. Research and managment techniques for the conservation of sea turtles. Washington, DC: IUCN/SSC Marine Turtle Specialist Group; 1999.
Escalona-Segura G, Borges-Ramírez MM, Estrella-Canul V, Rendón-von Osten J. A methodology for the sampling and identification of microplastics in bird nests. Green Anal Chem. 2022;3:100045. http://doi.org/10.1016/j.greeac.2022.100045.
European Union. Commission Decision (EU) 2017/848 of 17 May 2017 laying down criteria and methodological standards on good environmental status of marine waters and specifications and standardised methods for monitoring and assessment, and repealing Decision 2010/477/EU. Official Journal of the European Union; 2017 May 17.
Franzellitti S, Locatelli C, Gerosa G, Vallini C, Fabbri E. Heavy metals in tissues of loggerhead turtles (Caretta caretta) from the northwestern Adriatic Sea. Comp Biochem Physiol C Toxicol Pharmacol. 2004;138(2):187-94. http://doi.org/10.1016/j.cca.2004.07.008. PMid:15450866.
Geraci JR, Lounsbury VJ. Marine mammals ashore: a field guide for strandings. Galveston: Texas A&M Sea Grant; 2005.
Gómez-Ramírez P, Espín S, Navas I, Martínez-López E, Jiménez P, María-Mojica P, Peñalver J, García-Fernández AJ. Mercury and organochlorine pesticides in tissues of loggerhead sea turtles (Caretta caretta) stranded along the southwestern Mediterranean coastline (Andalusia, Spain). Bull Environ Contam Toxicol. 2020;104(5):559-67. http://doi.org/10.1007/s00128-020-02822-z. PMid:32185428.
Hermsen E, Mintenig SM, Besseling E, Koelmans AA. Quality criteria for the analysis of microplastic in biota samples: a critical review. Environ Sci Technol. 2018;52(18):10230-40. http://doi.org/10.1021/acs.est.8b01611. PMid:30137965.
Howard PH. Handbook of environmental fate and exposure data for organic chemicals. Vol. 2 Boca Raton: CRC Press; 2023.
Imhof HK, Sigl R, Brauer E, Feyl S, Giesemann P, Klink S, Leupolz K, Löder MG, Löschel LA, Missun J, Muszynski S, Ramsperger AF, Schrank I, Speck S, Steibl S, Trotter B, Winter I, Laforsch C. Spatial and temporal variation of macro-, meso- and microplastic abundance on a remote coral island of the Maldives, Indian Ocean. Mar Pollut Bull. 2017;116(1-2):340-7. http://doi.org/10.1016/j.marpolbul.2017.01.010. PMid:28109654.
Innis C, Nyaoke AC, Williams CR, Dunnigan B, Merigo C, Woodward DL, et al. Pathologic and parasitologic findings of cold-stunned kemp’s ridley sea turtles (lepidochelys kempii) stranded on Cape Cod, Massachusetts, 2001-2006. J Wildl Dis. 2009;45(3):594-610. http://doi.org/10.7589/0090-3558-45.3.594. PMid:19617470.
IUCN Red List of Threatened Species, version 2020-2 [Internet]. Gland (CH): IUCN; 2020 [cited 2020 Jul 9]. Available from: https://www.iucnredlist.org.
Jakimska A, Konieczka P, Skóra K, Namieśnik J. Bioaccumulation of metals in tissues of marine animals. Pt I, The role and impact of heavy metals on organisms. Pol J Environ Stud. 2011;20(5):1117-25.
Jerez S, Motas M, Cánovas RÁ, Talavera J, Almela RM, del Río AB. Accumulation and tissue distribution of heavy metals and essential elements in loggerhead turtles (Caretta caretta) from Spanish Mediterranean coastline of Murcia. Chemosphere. 2010;78(3):256-64. http://doi.org/10.1016/j.chemosphere.2009.10.062. PMid:19959203.
Keller JM, Kucklick JR, Stamper MA, Harms CA, McClellan- Green PD. Associations between organochlorine contaminant concentrations and clinical health parameters in loggerhead sea turtles from North Carolina, USA. Environ Health Perspect. 2004;112(10):1074-9. http://doi.org/10.1289/ehp.6923. PMid:15238280.
Lazar B, Gračan R. Ingestion of marine debris by loggerhead sea turtles, Caretta caretta, in the Adriatic Sea. Mar Pollut Bull. 2011;62(1):43-7. http://doi.org/10.1016/j.marpolbul.2010.09.013. PMid:21036372.
Lutcavage ME. Human impacts on sea turtle survival. In: Lutz PL, Musick JA, editors. The biology of sea turtles. Vol. 1. Boca Raton: CRC Press; 1997. p. 387-409.
Lutz PL, Musick JA, Wyneken J. The biology of sea turtles. Vol. 2. Boca Raton: CRC Press; 2003.
Matiddi M, DeLucia GA, Silvestri C, Darmon G, Tomás J, Pham CK, Camedda A, Vandeperre F, Claro F, Kaska Y, Kaberi H, Revuelta O, Piermarini R, Daffina R, Pisapia M, Genta D, Sözbilen D, Bradai MN, Rodríguez Y, Gambaiani D, Tsangaris C, Chaieb O, Moussier J, Loza AL, Miaud C. Data collection on marine litter ingestion in sea turtles and thresholds for good environmental status. J Vis Exp. 2019;2019(147). http://doi.org/10.3791/59466. PMid:31180365.
Nelms SE, Duncan EM, Broderick AC, Galloway TS, Godfrey MH, Hamann M, Lindeque PK, Godley BJ. Plastic and marine turtles: a review and call for research. ICES J Mar Sci. 2015;73(2):165-81. http://doi.org/10.1093/icesjms/fsv165.
Nuitja INS, Wijaya S. Studies on digestive organs of two sea turtles, Chelonia mydas L. and Eretmochelys imbricata L. Indones Fish Res J. 2017;11(1):47. http://doi.org/10.15578/ifrj.11.1.2005.47-52 .
Peixoto PV, Barros CSL. A importância da necropsia em medicina veterinária. Pesq Vet Bras. 1998;18(3-4):132-4. http://doi.org/10.1590/S0100-736X1998000300008.
Petry MV, Araújo LD, Brum AC, Benemann VRF, Finger JVG. Plastic ingestion by juvenile green turtles (Chelonia mydas) off the coast of Southern Brazil. Mar Pollut Bull. 2021;167:112337. http://doi.org/10.1016/j.marpolbul.2021.112337.
Rizzi M, Rodrigues FL, Medeiros L, Ortega I, Rodrigues L, Monteiro DS, Kessler F, Proietti MC. Ingestion of plastic marine litter by sea turtles in southern Brazil: abundance, characteristics and potential selectivity. Mar Pollut Bull. 2019;140:536-48. http://doi.org/10.1016/j.marpolbul.2019.01.054. PMid:30803675.
Rochman CM, Browne MA, Halpern BS, Hentschel BT, Hoh E, Karapanagioti HK, Rios-Mendoza LM, Takada H, Teh S, Thompson RC. Policy: classify plastic waste as hazardous. Nature. 2013;494(7436):169-71. http://doi.org/10.1038/494169a. PMid:23407523.
Roslan UAM, Harun FN. Modeling the impact of pollution on sea turtles. Mal J Fund Appl Sci. 2023;19(4):607-15. http://doi.org/10.11113/mjfas.v19n4.2913.
Salvarani P, Foster VC, Rendon J, Morgado F. Contaminants impact on marine turtle populations development. Ecotox Mar Organ. 2019a; 205:205-34. http://doi.org/10.1201/b22000-9.
Salvarani PI, Morgado F, Vieira LR, Rendón-von Osten J. Organochlorines contaminants in eggs of Hawksbill (Eretmochelys imbricata) and green sea turtles (Chelonia mydas) from Mexico coast. Arch Environ Contam Toxicol. 2019b;76(3):425-34. http://doi.org/10.1007/s00244-018-00589-3. PMid:30600338.
Short FS, Lôbo-Hajdu G, Guimarães SM, Laport MS, Silva R. Antimicrobial-resistant bacteria from free-living green turtles (Chelonia mydas). Antibiotics. 2023;12(8):1268. http://doi.org/10.3390/antibiotics12081268. PMid:37627688.
Thompson RC, Moore CJ, Saal FSV, Swan SH. Plastics, the environment and human health: current consensus and future trends. Philos Trans R Soc Lond B Biol Sci. 2009;364(1526):2153- 66. http://doi.org/10.1098/rstb.2009.0053. PMid:19528062.
Thomson JA, Burkholder D, Heithaus MR, Dill LM. Validation of a rapid visual-assessment technique for categorizing the body condition of green turtles (Chelonia Mydas) in the field. Copeia. 2009;2009(2):251-5. http://doi.org/10.1643/CE-07-227.
Tourinho PS, Ivar do Sul JA, Fillmann G. Is marine debris ingestion still a problem for the coastal marine biota of southern Brazil? Mar Pollut Bull. 2010;60(3):396-401. http://doi.org/10.1016/j.marpolbul.2009.10.013. PMid:19931101.
Underwood A. Research techniques in animal ecology: controversies and consequences. J Exp Mar Biol Ecol. 2002; 268(1):115-6. http://doi.org/10.1016/S0022-0981(01)00374-4.
van de Merwe JP, Hodge M, Olszowy HA, Whittier JM, Ibrahim K, Lee SY. Chemical contamination of green turtle (Chelonia mydas) eggs in peninsular Malaysia: implications for conservation and public health. Environ Health Perspect. 2009;117(9):1397-401. http://doi.org/10.1289/ehp.0900813. PMid:19750104.
Vanapalli KR, Samal B, Dubey BK, Bhattacharya J. Emissions and environmental burdens associated with plastic solid waste management. In: Al-Salem SM, editor. Plastics to energy: fuel, chemicals, and sustainability implications. Norwich: William Andrew; 2018. http://doi.org/10.1016/B978-0-12-813140-4.00012-1.
Vegter AC, Barletta M, Beck C, Borrero J, Burton H, Campbell ML, Costa MF, Eriksen M, Eriksson C, Estrades A, Gilardi KVK, Hardesty BD, Ivar do Sul JA, Lavers JL, Lazar B, Lebreton L, Nichols WJ, Ribic CA, Ryan PG, Schuyler QA, Smith SDA, Takada H, Townsend KA, Wabnitz CCC, Wilcox C, Young LC, Hamann M. Global research priorities to mitigate plastic pollution impacts on marine wildlife. Endanger Species Res. 2014;25(3):225-47. http://doi.org/10.3354/esr00623.
Wang Y, Qian H. Phthalates and their impacts on human health. Healthcare. 2021;9(5):603. http://doi.org/10.3390/healthcare9050603. PMid:34069956.
Work TM, Raskin RE, Balazs GH, Whittaker SD. Morphologic and cytochemical characteristics of blood cells from Hawaiian green turtles. Am J Vet Res. 1998;59(10):1252-7. http://doi.org/10.2460/ajvr.1998.59.10.1252. PMid:9781457.
Work TM. Sea turtle necropsy manual for biologists in remote refuges. Honolulu: National Wildlife Health Center, Hawaii Field Station; 2000.
Wyneken J. Respiratory anatomy-form and function in reptiles. Exotic DVM Magazine. 2001;3(2):17-22.
Yaghmour F, Samara F, Alam I. Analysis of polychlorinated biphenyls, polycyclic aromatic hydrocarbons and organochlorine pesticides in the tissues of green sea turtles, Chelonia mydas, (Linnaeus, 1758) from the eastern coast of the United Arab Emirates. Mar Pollut Bull. 2020;160:111574. http://doi.org/10.1016/j.marpolbul.2020.111574. PMid:33181917.
Yancheva V, Velcheva I, Georgieva E, Stoyanova S, Todorova B, Antal L, Nyeste K. Are Mytilus species suitable bioindicators for assessing aquatic pollution along the Black Sea Coast? A review. Ecol Balk. 2023;15(1).
Zhang T, Lin L, Li D, Wang J, Liu Y, Li R, Wu S, Shi H. Microplastic pollution at Qilianyu, the largest green sea turtle nesting grounds in the northern South China Sea. PeerJ. 2022;10:e13536. http://doi.org/10.7717/peerj.13536. PMid:35694388.
Downloads
Published
Issue
Section
License
Copyright (c) 2025 Brazilian Journal of Veterinary Research and Animal Science
This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License.
The journal content is authorized under the Creative Commons BY-NC-SA license (summary of the license: https://
