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Comet assay in Aegla platensis (Decapoda: Anomura) using a non-lethal hemolymph field sampling for in situ monitoring of freshwater genotoxicity

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Abstract

This study aimed to apply the comet assay on Aegla platensis crabs as a suitable non-destructive approach for in situ monitoring of freshwater genotoxicity. Animals were captured during four sampling periods in a stream under minor anthropogenic impacts in Southern Brazil. Crabs were captured with a hand net, then the hemolymph samples were collected, and the animals were released into the stream after a 20-min recovery time. Hemolymph samples were transported to the laboratory and used to perform the alkaline comet assay. Results showed an intermediate level in the DNA damage index (range 107.3–165.0 arbitrary unit). No significant differences were observed among the different sampling periods. Hemolymph was successfully used as a non-lethal source of biological samples, and the comet assay using A. platensis proved to be a feasible approach for genotoxicity studies.

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Fig. 1: Location of study area.
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Data availability

The datasets generated during and/or analyzed during the current study are available from the corresponding author upon reasonable request.

References

  • Andem AB, Agbor RB, Ekpo IA (2013) Review on Comet Assay: A reliable tool for assessing DNA damage in animal models. J Curr Res Sci 1:405–427

    Google Scholar 

  • Astley KN, Meigh HC, Glegg GA, Braven J, Depledge MH (1999) Multi-variate analysis of biomarker responses in Mytilus edulis and Carcinus maenas from the Tees Estuary (UK). Mar Pollut Bull 39:145–154

    Article  CAS  Google Scholar 

  • Azqueta A, Meier S, Priestley C, Gutzkow KB, Brunborg G, Sallette J, Soussaline F, Collins A (2011) The influence of scoring method on variability in results obtained with the comet assay. Mutagenesis 26:393–399. https://doi.org/10.1093/mutage/geq105

    Article  CAS  Google Scholar 

  • Bauchau AG (1981) Crustaceans. In: Ratcliffe NA, Rowley AF (ed) Invertebrate Blood Cells, vol 2. Academic Press, London, p 385–420

    Google Scholar 

  • Bond-Buckup G, Buckup L (1994) A família Aeglidae. Arq Zool 32:159–347

    Article  Google Scholar 

  • Bond-Buckup G, Jara CG, Perez-Losada M, Buckup L, Crandall KA (2008) Global diversity of crabs (Aeglidae: Anomura: Decapoda) in freshwater. Hydrobiologia 595:267–273. https://doi.org/10.1007/s10750-007-9022-4

    Article  Google Scholar 

  • Borges ACP, Piassão JFG, Paula MO, Sepp S, Bez CFS, Hepp LU, Valduga AT, Pereira AAM, Cansian RL (2018) Characterization of oxidative stress biomarkers in a freshwater anomuran crab. Braz J Biol 78:61–67. https://doi.org/10.1590/1519-6984.04816

    Article  CAS  Google Scholar 

  • Borges ACP, Piassão JFG, Albani SM, Albertoni EF, Martins MC, Cansian RL, Valduga AT, Hepp LU, Mielniczki-Pereira AA (2021) Multiple metals and agricultural use affects oxidative stress biomarkers in freshwater Aegla crabs. Braz J Biol 15:e230147. https://doi.org/10.1590/1519-6984.230147

    Article  Google Scholar 

  • Breitholtz M, Rudén C, Hansson SO, Bengtsson BE (2006) Ten challenges for improved ecotoxicological testing in environmental risk assessment. Ecotoxicol Environ Saf 63:324–335. https://doi.org/10.1016/j.ecoenv.2005.12.009

    Article  CAS  Google Scholar 

  • Brown RJ, Galloway TS, Lowe D, Browne MA, Dissanayake A, Jones MB, Depledge MH (2004) Differential sensitivity of three marine invertebrates to copper assessed using multiple biomarkers. Aquat Toxicol 66:267–287

    Article  CAS  Google Scholar 

  • Collins AR, Ma AG, Duthie SJ (1995) The kinetics of repair of oxidative DNA damage (strand breaks and oxidised pyrimidines) in human cells. Mutat Res 336:69–77. https://doi.org/10.1016/0921-8777(94)00043-6

    Article  CAS  Google Scholar 

  • Cumberlidge N, Ng PKL, Yeo DCJ, Magalhães C, Campos MR, Alvarez F, Naruse T, Daniels SR, Esser LJ, Attipoe FYK, Clotilde-Ba F-L, Darwall W, McIvor A, Baillie JEM, Collen B, Ram M (2009) Freshwater crabs and the biodiversity crisis: Importance, threats, status, and conservation challenges. Biol Conserv 142:1665–1673

    Article  Google Scholar 

  • da Silva Rocha AJ, Gomes V, Rocha Passos MJ, Hasue FM, Alves Santos TC, Bícego MC, Taniguchi S, Van Ngan P (2012) EROD activity and genotoxicity in the seabob shrimp Xiphopenaeus kroyeri exposed to benzo[a]pyrene (BaP) concentrations. Environ Toxicol Pharmacol 34:995–1003. https://doi.org/10.1016/j.etap.2012.07.006

    Article  CAS  Google Scholar 

  • Davanso MB, Moreira LB, Pimentel MF, Costa-Lotufo LV, Abessa LVS (2013) Biomarkers in mangrove root crab Goniopsis cruentata for evaluating quality of tropical estuaries. Mar Environ Res 92:80–88

    Article  Google Scholar 

  • Dhawan A, Bajpayee M, Parmar D (2009) Comet assay: A reliable tool for the assessment of DNA damage in different models. Cell Biol Toxicol 25:5–32

    Article  CAS  Google Scholar 

  • Di Donato G, De Matthaeis E, Ronci L, Setini A (2016) Genotoxicity biomarkers in the amphipod Gammarus elvirae exposed in vivo to mercury and lead, and basal levels of DNA damage in two cell types. Chem Ecol 32:843–857

    Article  Google Scholar 

  • Di Donato G, Davolos D, Chimenti C, De Matthaeis E (2020) A genotoxicity study on Talitrus saltator (Crustacea, Amphipoda) exposed to lead and benzo(a)pyrene. Eur Zool J 87:235–241. https://doi.org/10.1080/24750263.2020.1751887

    Article  CAS  Google Scholar 

  • Esteves F, Amaro R, Silva S, Sánchez-Flores M, Teixeira JP, Costa C (2020) The impact of comet assay data normalization in human biomonitoring studies outcomes. Toxicol Lett 332:56–64. https://doi.org/10.1016/j.toxlet.2020.06.024

    Article  CAS  Google Scholar 

  • Erraud A, Bonnard M, Geffard O, Chaumot A, Duflot A, Geffard A, Forget-Leray J, Xuereb B (2019a) Assessment of sperm DNA integrity within the Palaemon longirostris (H.) population of the Seine estuary. Environ Pollut 245:485–493. https://doi.org/10.1016/j.envpol.2018.11.006

    Article  CAS  Google Scholar 

  • Erraud A, Bonnard M, Geffard O, Coulaud R, Poret A, Duflot A, Forget-Leray J, Geffard A, Xuereb B (2019b) Signification of DNA integrity in sperm of Palaemon serratus (Pennant 1777): Kinetic responses and reproduction impairment. Mar Environ Res 144:130–140. https://doi.org/10.1016/j.marenvres.2019.01.005

    Article  CAS  Google Scholar 

  • Faria M, Huertas D, Soto DX, Grimalt JO, Catalan J, Riva MC, Barata C (2010) Contaminant accumulation and multi-biomarker responses in field collected zebra mussels (Dreissena polymorpha) and crayfish (Procambarus clarkii), to evaluate toxicological effects of industrial hazardous dumps in the Ebro river (NE Spain). Chemosphere 78:232–240

    Article  CAS  Google Scholar 

  • Gajski G, Žegura B, Ladeira C, Pourrut B, Del Bo’ C, Novak M, Sramkova M, Milić M, Gutzkow KB, Costa S, Dusinska M, Brunborg G, Collins A (2019) The comet assay in animal models: From bugs to whales – (Part 1 Invertebrates). Mutat Res 779:82–113. https://doi.org/10.1016/j.mrrev.2019.02.003

    Article  CAS  Google Scholar 

  • García O, Mandina T, Lamadrid AI, Diaz A, Reminigio A, Gonzalez Y, Piloto J, Gonzalez JE, Alvarez A (2004) Sensitivity and variability of visual scoring in the comet assay. Results of an inter-laboratory scoring exercise with the use of silver staining. Mutat Res 556:25–34. https://doi.org/10.1016/j.mrfmmm.2004.06.035

    Article  CAS  Google Scholar 

  • Glei M, Schneider T, Schlörmann W (2016) Comet assay: An essential tool in toxicological research. Arch Toxicol 90:2315–36. https://doi.org/10.1007/s00204-016-1767-y

    Article  CAS  Google Scholar 

  • Gruber C, Kortet R, Vainikka A, Hyvärinen P, Rantala MJ, Pikkarainen A, Jussila J, Makkonen J, Kokko H, Hirvonen H (2014) Variation in resistance to the invasive crayfish plague and immune defence in the native noble crayfish. Annal Zool Fenn 51:371–389

    Article  Google Scholar 

  • Halsband C, Dix MF, Sperre KH, Reinardy HC (2021) Reduced pH increases mortality and genotoxicity in an Arctic coastal copepod, Acartia longiremis. Aquat Toxicol 239:105961. https://doi.org/10.1016/j.aquatox.2021.105961

    Article  CAS  Google Scholar 

  • Jha AN (2008) Ecotoxicological applications and significance of the comet assay. Mutagenesis 23:207–221

    Article  CAS  Google Scholar 

  • Jussila J, Ruokonen TJ, JSyväranta J, Kokko H, Vainikka A, Makkonen J, Kortet R (2015) It takes time to see the menu from the body: An experiment on stable isotope composition in freshwater crayfishes. Knowl Manag Aquat Ec 416:25

    Article  Google Scholar 

  • Kankamol C, Salaenoi J (2018) Enzymes involved in immunity and characteristics of hemolymph in red sternum syndrome mud crabs (Scylla serrata). Agric Nat Resour 52:489–496

    Google Scholar 

  • Kumaravel TS, Vilhar B, Faux SP, Jha NA (2009) Comet Assay measurements: A perspective. Cell Biol Toxicol 25:53–64. https://doi.org/10.1007/s10565-007-9043-9

    Article  CAS  Google Scholar 

  • Lacaze E, Geffard O, Bony S, Devaux A (2010) Genotoxicity assessment in the amphipod Gammarus fossarum by use of the alkaline Comet assay. Mutat Res 700:32–38

    Article  CAS  Google Scholar 

  • Lee RF, Steinert S (2003) Use of the single cell gel electrophoresis/comet assay for detecting DNA damage in aquatic (marine and freshwater) animals. Mutat Res 544:43–64

    Article  CAS  Google Scholar 

  • Lewbart GA, Noga EJ, Hancock AL, Bullis RA (2006) Crustaceans. In: Lewbart GA (ed) Invertebrate medicine. Blackwell Publishing, IW, p 179–194

    Chapter  Google Scholar 

  • Martín-Díaz ML, Blasco J, Sales D, Delvalls TA (2007) Biomarkers study for sediment quality assessment in Spanish ports using the crab Carcinus maenas and the clam Ruditapes philippinarum. Arch Environ Contam Toxicol 53:66–77

    Article  Google Scholar 

  • Martins M, Costa PM (2015) The comet assay in Environmental Risk Assessment of marine pollutants: Applications, assets and handicaps of surveying genotoxicity in non-model organisms. Mutagenesis 30:89–106. https://doi.org/10.1093/mutage/geu037

    Article  CAS  Google Scholar 

  • Møller P, Azqueta A, Boutet-Robinet E et al. (2020) Minimum Information for Reporting on the Comet Assay (MIRCA): Recommendations for describing comet assay procedures and results. Nat Protoc 15:3817–3826. https://doi.org/10.1038/s41596-020-0398-1

    Article  CAS  Google Scholar 

  • Nadin SB, Vargas-Roig LM, Ciocca DR (2001) A silver staining method for single-cell gel assay. J Histochem Cytochem 49:1183–6

    Article  CAS  Google Scholar 

  • Noro CK, Buckup L (2002) Biologia reprodutiva e ecologia de Aegla leptodactyla Buckup & Rossi (Crustacea, Anomura, Aeglidae). Rev Bras de Zool 19:1063–1074

    Article  Google Scholar 

  • Rocha AJS, Botelho MT, Hasue FM, Arrud MJ (2015) Genotoxicity of shallow waters near the Brazilian Antarctic Station “Comandante Ferraz” (EACF), Admiralty Bay, King George Island, Antarctica. Braz J Oceanogr 63:63–70

    Article  Google Scholar 

  • Rodrigues ET, Pardal MÂ (2014) The crab Carcinus maenas as a suitable experimental model in ecotoxicology. Environ Int 70:158–182. https://doi.org/10.1016/j.envint.2014.05.018

    Article  CAS  Google Scholar 

  • Sahlmann A, Lode T, Heuschele J, Borgå K, Titelman J, Hylland K (2019) Genotoxic response and mortality in 3 marine copepods exposed to waterborne copper. Environ Toxicol Chem 38:2224–2232. https://doi.org/10.1002/etc.4541

    Article  CAS  Google Scholar 

  • Singh NP, McCoy MT, Tice RR, Schneider EL (1988) A simple technique for quantitation of low levels of DNA damage in individual cells. Exp Cell Res 175:184–91. https://doi.org/10.1016/0014-4827(88)90265-0

    Article  CAS  Google Scholar 

  • Strieder MN, Scherer RT, Viegas G (2006) Biomonitoramento da qualidade das águas em arroios na Bacia Hidrográfica do Rio dos Sinos, Rio Grande do Sul, Brasil. Unirevista 1:47–56

    Google Scholar 

  • Ternjej I, Mihaljevic Z, Iykovic M, Previsic A, Stankovic I, Maldini K, Zeljezic D, Kopjar N (2014) The impact of gypsum mine water: A case study on morphology and DNA integrity in the freshwater invertebrate, Gammarus balcanicus. Environ Pollut 189:229–238

    Article  CAS  Google Scholar 

  • Vazquez L, Alpuche J, Maldonado G (2009) Immunity mechanisms in crustaceans. Innate Immun 15:179–188

    Article  CAS  Google Scholar 

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Acknowledgements

AG received a student fellowship from the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES/PROSUP). LBdS received a fellowship of research productivity (PQ) granted by the national council for scientific and technological development (CNPq – 313546/2018-5). No funding was received for conducting this study.

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  1. Feevale University, ERS-239, 2755, Novo Hamburgo, Rio Grande do Sul, Brazil

    Angélica Goldoni, Marcelo Rech Pacheco & Luciano Basso da Silva

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  1. Angélica Goldoni
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  2. Marcelo Rech Pacheco
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  3. Luciano Basso da Silva
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LBS designed the research; MRP and AG performed data collection and analysis; AG wrote the first draft; all authors reviewed the manuscript.

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Correspondence to Luciano Basso da Silva.

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Goldoni, A., Pacheco, M.R. & da Silva, L.B. Comet assay in Aegla platensis (Decapoda: Anomura) using a non-lethal hemolymph field sampling for in situ monitoring of freshwater genotoxicity. Ecotoxicology 32, 160–165 (2023). https://doi.org/10.1007/s10646-023-02627-w

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