Revista de Gestão Costeira Integrada
Volume 22, Issue 2, September 2022, Pages 103-116

DOI: 10.5894/rgci-n441
* Submission: 3 APR 2021; Peer review: 9 JUL 2021; Revised: 26 SEP 2022; Accepted: 26 SEP 2022; Available on-line: 12 OCT 2022

Use of Brachionus Plicatilis (rotifera) to assess the quality of marine water in Callao bay, Peru

Marco Osorio¹, Seid Romero¹, Angélica Guabloche¹, Lorena Alvariño-Flores¹, Yuri Ayala-Sulca², Carlos Carrasco-Badajoz³, Luz Castañeda¹, Luis Carrasco⁴, Jose Alberto Iannacone^{@ 1, 5, 6}

---

@ Corresponding author: [email protected]
¹ Facultad de Ciencias Naturales y Matemática. Grupo de Investigación en Sostenibilidad Ambiental (GISA). Escuela Universitaria de Posgrado (EUP), Universidad Nacional Federico Villarreal. Lima, Perú.
² Laboratorio de Zoología. Facultad de Ciencias Biológicas. Universidad Nacional de San Cristóbal de Huamanga (Ayacucho-Perú). Miembro del grupo de investigación: Evolución Molecular de la Biota en el Perú. Universidad Nacional Mayor de San Marcos, Lima-Perú.
³ Laboratorio de Biodiversidad y Sistema de Información Geográfica BioSIG. Centro de Investigaciones en Ecosistemas y Biodiversidad Acuática CI-EBACUA. Facultad de Ciencias Biológicas. Universidad Nacional de San Cristóbal de Huamanga, Ayacucho-Perú.
⁴ Unidad de Investigación de la Facultad de Ingeniería Química. Universidad Nacional del Callao, Lima, Perú.
⁵ Laboratorio de Ingeniería Ambiental. Carrera de Ingeniería Ambiental. Coastal Ecosystems of Peru Research Group (COEPERU). Universidad Científica del Sur. Lima, Perú.
⁶ Grupo de Investigación "One Health". Facultad de Ciencias Biológicas. Universidad Ricardo Palma. Lima, Perú.

---

ASBTRACT
The present investigation used Brachionus plicatilis (Rotifera) to evaluate the water quality in Callao Bay, Peru. The water samples were taken in four areas: P1 in La Punta, P2 in Chucuito, P3 in front of the Callao Port Terminal and P4 in the San Lorenzo and El Frontón Islands, and in four seasonal periods: autumn-2015, winter- 2015, spring-2015, and summer-2016. Physical-chemical parameters, chlorophyll, phycocyanin, heavy metals, and bioassays with seawater at 24 h and 48 h of exposure to Brachionus plicatilis obtained from a standardized culture were evaluated in each area and seasonal period. High concentrations of Ag, Pb, Hg, Cu, Cr, Ni and Zn were found at all the sampling sites. The ​​ no observed effect concentration (NOEC) values (concentration at which a significant effect of mortality is not observed with respect to the control) and lowest observed effect concentration (LOEC) (minimum concentration at which a significant effect of mortality is observed with respect to the control) at 48 h of exposure to the B. plicatilis bioassay were lower for P1 (winter-2015) and P2 (winter-2015). Principal components analysis (PCA) showed that principal component 1 (PC1) contributed 38.70% and PC2 17.70%. PC1 was formed by LOEC of Cd, Hg, Ni, Ag, and Pb while PC2 was related to Cu, Cr and Zn. According to these results Brachionus plicatilis can be used as a bioindicator organism to assess water quality in the marine environment.

Keywords: acute bioassay, Brachionus, heavy metals, mortality, rotifer.

RESUMO
A presente investigação utilizou Brachionus plicatilis (Rotifera) para avaliar a qualidade da água na Bahía del Callao, Peru. As amostras de água foram coletadas em quatro locais: P1 em La Punta, P2 em Chucuito, P3 em frente ao Terminal Portuário de Callao e P4 nas Ilhas San Lorenzo e Frontón, e em quatro períodos sazonais: outono-2015, inverno-2015, primavera-2015 e verão-2016. Os parâmetros físico-químicos, clorofila, ficocianina, metais pesados ​​e bioensaios com água do mar às 24 he 48h de exposição com Brachionus plicatilis obtidos em cultura padronizada foram avaliados em cada área e período sazonal. Os metais Ag, Pb, Hg, Cu, Cr, Ni e Zn foram encontrados em maior concentração em todos os locais de amostragem. Os valores de NOEC (concentração onde um efeito significativo de mortalidade não é observado em relação ao controle) e LOEC (concentração mínima onde um efeito significativo de mortalidade é observado em relação ao controle) em 48 h de exposição com o bioensaio de B. plicatilis foram menores para P1 (inverno-2015) e P2 (inverno-2015). A técnica de Análise de Componentes Principais (ACP) mostrou que o componente principal 1 (CP1) contribuiu com 38,70% e o CP2 com 17,70%. CP1 foi formado por LOEC, Cd, Hg, Ni, Ag e Pb. CP2 relacionou as variáveis Cu, Cr e Zn. Propõe-se a utilização de Brachionus plicatilis como organismo bioindicador para avaliação da qualidade da água no meio marinho.

Palavras-chave: bioensaio agudo, Brachionus, metais pesados, mortalidade, rotífero.

 

Alvarado-Flores, J.; Rubio-Franchini, I.; Sánchez-Ávila, A.S.; Ramírez-Tlalolín, G.J.; Rico-Martínez, R. (2019) - Arsenic toxicity, bioaccumulation and risk assessment: A case study in Tolimique Dam, Aguascalientes, Mexico. Cogent Environmental Science, 5: 1650630. DOI:10.1080/23311843.2019.1650630.

AM - Acuerdo Ministerial (2015) - Norma de Calidad Ambiental y de Descarga de Efluentes: Recurso Agua. Norma de Calidad Ambiental y de descarga de efluentes al Recurso Agua. Reforma Texto Unificado Legislación Secundaria, Medio Ambiente, Libro VI, Decreto Ejecutivo 3516. Acuerdo Ministerial 97. Available on-line at https://www.ambiente.gob.ec/wp-content/uploads/downloads/2018/05/Acuerdo-097.pdf.

ANZECC - Australian and New Zealand Environment and Conservation Council & ARMCANZ - Agriculture and Resource Management Council of Australia and New Zealand (2000) - Australian and New Zealand guidelines for fresh and marine water quality. The guidelines / Australian and New Zealand Environment and Conservation Council, Agriculture and Resource Management Council of Australia and New Zealand. Australian Water Association, Available on-line at https://www.waterquality.gov.au/sites/default/files/documents/anzecc-armcanz-2000-guidelines-vol1.pdf.

Argüelles, J.; Quispe, J.; Ledesma, J.; León, V.; Sarmiento, M.; Robles, C. (2012). Estudio Línea base - ELBA Callao, primavera 2011 - Informe Interno IMARPE, 39: 3-4. Available on-line at http://biblioimarpe.imarpe.gob.pe/bitstream/123456789/2223/1/Informe%2039%283-4%291.pdf.

Beiras, R.; Vázquez, E.; Bellas, J.; Lorenzo, J.I.; Fernández, N.; Macho, G.; Marín, J.C: Casas, L. (2001) - Sea-urchin embryo bioassay for in situ evaluation of the biological quality of coastal seawater. Estuarine, Coastal and Shelf Science, 52: 29–32. DOI:10.1006/ecss.2000.0720.

Casanova-Rosero, R.; Suárez-Vargas, N.; Zambrano-Ortiz, M. (2015) - Valoración de algunas variables fisicoquímicas indicadoras de la calidad del agua en las principales bahías de la costa Pacífica colombiana–2009. Boletín Científico CIOH, 33: 195–214. Available on-line at https://www.oceandocs.org/handle/1834/14719.

CCMC - Canadian Environmental Quality Guidelines. (2007) - Canadian Water Quality Guidelines for the Protection of Aquatic Life. Chapter 4. Canadian Environmental Quality Guidelines. Canadian Council of Ministers of the Environment, Winnipeg. Available on-line at https://www.ccme.ca/en/resources/canadian_environmental_quality_guidelines.

Clark, M.S.; Denekamp, N.Y.; Thorne, M.A.S.; Reinhardt, R.; Drungowski, M.; Albrecht, M.W.; Klages, S.; Beck, A.; Kube, M.; Lubzens, E. (2012) - Long-term survival of hydrated resting eggs from Brachionus plicatilis. PLoS ONE, 7(1): e29365. DOI:10.1371/journal.pone.0029365.

Contreras-Sillero, M.E.; Pacheco-Vega, J.M.; Cadena-Roa, M.A.; Contreras-Chavarría, J.A.; Rangel-Dávalos, C.; Valdez-González, F.J.; González-Hermoso, J. P. (2019) - Evaluation of growth and bromatological composition of the rotifer Brachionus plicatilis fed different native microalgae from the Gulf of California. Revista Bio Ciencias, 6: e509. DOI:10.15741/revbio.06.e509.

Cousseau, M.; Perrotta, G. (2013) - Peces marinos de Argentina: biología, distribución, pesca. Publicaciones Especiales INIDEP. 4ta edición. Mar del Plata, Argentina. ISBN 978-987-1443-09-3.

Crane, M.; Babut, M. (2006) - Environmental quality standards for water framework directive priority substances: challenges and opportunities. Integrated Environmental Assessment and Management, 3: 290–296. DOI:10.1897/IEAM_2006-045.1.

Djinovic-Stojanovic, J. (2015) - Distribution of mercury in three marine fish species. Procedia Food Science, 5: 65-68. DOI:10.1016/j.profoo.2015.09.016.

ENFEN - Comité Multisectorial Encargado del Estudio Nacional del Fenómeno El Niño (2015) - Monitoreo y Pronóstico de El Niño y La Niña. Informe Técnico Enfen, 1 (9), 1-58. Available on-line at http://enfen.gob.pe/download/informe-tecnico-2015-12/?wpdmdl=890&ind=1524063682034.

Escobar-Chávez, C.; Alvariño, L.; Iannacone, J. (2019) - Evaluation of the aquatic environmental risk of the mixture of the pesticides imidacloprid (insecticide) and propineb (fungicide) in Daphnia magna straus, 1820. Paideia XXI, 9: 301–332. DOI:10.31381/paideia.v9i2.2759.

Fathallah, S.; Medhioub, M.N.; Medhioub, A.; Kraiem, M.M. (2011) - Ruditapes decussatus embryo-larval toxicity bioassay for assessment of Tunisian coastal water contamination. Journal of Environmental Chemistry and Ecotoxicology, 3: 277-285. Available on-line at https://academicjournals.org/article/article1380009080_Fathallah%20et%20al.pdf.

Ferrando, N.S.; Nandini, S.; Claps, M.C.; Sarma, S.S.S. (2019) - Effect of salinity and food concentration on competition between Brachionus plicatilis Müller, 1786 and Brachionus calyciflorus Pallas, 1776 (Rotifera). Marine and Freshwater Research, 71: 493-504. DOI:10.1071/MF18403.

Ferrari, L. (2015) - La ecotoxicología aplicada a la evaluación de la contaminación de los ríos: El caso del Río Reconquista. Ciencia e investigación, 65: 17-35. Available on-line at http://aargentinapciencias.org/wp-content/uploads/2018/01/RevistasCeI/tomo65-2/3-Ferrari-cei65-2-3.pdf.

Franco, A.L.; Nogueira, M.N.M.; Guimarães, K.S.; Frota, M.F.; Fernandes, C.M.S.; Sena, M.C. (2014) - Animal research: a bioethical reflection. Acta Bioethica, 20: 247-253. DOI:10.4067/S1726-569X2014000200012.

Fu, J.; Wang, H.; Billah, S.M.R.; Yu, H.; Zhang, X. (2014) - Heavy metals in seawater, sediments, and biota from the coastal area of Yancheng City, China. Environmental Toxicology and Chemistry, 33: 1697-1704. DOI:10.1002/etc.2575.

Gharaei, A.; Karimi, M.; Harijani, J.M.; Miri, M.; Faggio, C. (2020) - Population growth of Brachionus calyciflorus affected by deltamethrin and imidacloprid insecticides. Iranian Journal of Fisheries Sciences, 19: 588-601. Available on-line at http://jifro.ir/article-1-2987-en.html.

Guerrero-Jiménez, G.; Vannucchi, P.E.; Silva-Briano, M.; Adabache-Ortiz, A.; Rico-Martínez, R.; Roberts, D.; Neilson, R.; Elías-Gutiérrez, M. (2019) - Brachionus paranguensis sp. nov. (Rotifera, Monogononta), a member of the L group of the Brachionus plicatilis complex. ZooKeys, 880: 1–23. DOI: 10.3897/zookeys.880.28992.

Guillén, O.; Aquino, R.; Valdivia, B.; Calienes, R. (1978) - Contaminación en el Puerto del Callao. Instituto del Mar del Perú. Informe, 62: 1-55. Available on-line at http://biblioimarpe.imarpe.gob.pe/handle/123456789/287.

Herkovits, J.; Castañaga, L.A.; D’eramo, J.L.; Platonova-Jourani, V.P. (2015) - Living organisms influence on environmental conditions: pH modulation by amphibian embryos versus aluminum toxicity. Chemosphere, 139: 210–215. DOI:j.chemosphere.2015.05.025.

Hussain, M.B.; Laabir, M.; Yahia, M.N.D. (2020) - A novel index based on planktonic copepod reproductive traits as a tool for marine ecotoxicology studies. Science of the Total Environment, 727: 138621. DOI:10.1016/j.scitotenv.2020.138621.

Hwang, U.K.; Ryu, H.M.; Heo, S.; Chang, S.J.; Lee, K.W.; Lee, J.W. (2016) - Effect of heavy metals on the survival and population growth rates of marine rotifer, Brachionus plicatilis. korean journal of environmental biology, 34(4): 353~360. DOI: 10.11626/KJEB.2016.34.4.353.

INVEMAR (2003) - Informe del Estado de los Ambientes Marinos y Costeros en Colombia: 2003. Santa Marta, Colombia. Serie Documentos Generales, 294 pp. Available on-line at http://www.invemar.org.co/redcostera1/invemar/docs/IER_2003.pdf.

Jha, A. (2008) - Ecotoxicological applications and significance of the comet assay. Mutagenesis, 23:207–221. DOI:10.1093/mutage/gen014.

Jeong, Ch.B.; Kang, H.M.; Lee, M.Ch.; Byeon, E.; Park, H.G.; Lee, J.S. (2019a) - Effects of polluted seawater on oxidative stress, mortality, and reproductive parameters in the marine rotifer Brachionus koreanus and the marine copepod Tigriopus japonicas. Aquatic Toxicology, 208: 39–46. DOI: 10.1016/j.aquatox.2018.12.019.

Jeong, Ch.B.; Lee, Y.H.; Park, J.Ch.; Kang, H.M.; Hagiwara, A.; Lee, J.S. (2019b) – Effects of metal polluted seawater on life parameters and the induction of oxidative stress in the marine rotifer Brachionus koreanus. Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology, 225: 108576. DOI: 10.1016/j.cbpc.2019.108576.

Jolliffe, I.T.; Cadima, J. (2016) - Principal component analysis: a review and recent developments. Philosophical Transactions of the Royal Society A: Mathematical, Physical and Engineering Sciences, 374: 20150202. DOI:10.1098/rsta.2015.0202.

Kovalenko, V.F.; Goncharuk, V.V. and Golovkov, A.N. (2020) - Bioassay of seawater using the saltwater rotifer Brachionus plicatilis Müller. Journal of Water Chemistry and Technology, 42: 287–290. DOI: 10.3103/s1063455x20040062.

Larios-Meoño, J.F.; González-Taranco, C.; Olivares, Y.M. (2015). Las aguas residuales y sus consecuencias en el Perú. Revista de la Facultad de Ingeniería de la USIL, 2: 9-25. Available on-line at http://revistas.usil.edu.pe/index.php/syh/article/view/115#:~:text=Una%20preocupaci%C3%B3n%20es%20la%20contaminaci%C3%B3n,diabetes%20mellitus%2C%20y%20enfermedades%20cardiovasculares.

Li, X.D.; Wang, X.Y.; Xu M.E.; Jiang, Y.; Yan, T.; Wang X.C. (2020) - Progress on the usage of the rotifer Brachionus plicatilis in marine ecotoxicology: A review. Aquatic Toxicology, 229: 105678. DOI:10.1016/j.aquatox.2020.105678.

Li, Y.; Yu, J.; TSun, T.; Liu, C.; Sun, Y. and Wang, Y. (2018) - Using the marine rotifer Brachionus plicatilis as an endpoint to evaluate whether ROS-Dependent hemolytic toxicity is involved in the allelopathy induced by Karenia mikimotoi. Toxins, 10: 439. DOI:10.3390/toxins10110439.

López, C.; Steinitz-Kannan, M.; Stamou, G.; Michaloudi, E.; Papakostas, S.; Fontaneto, D. and Segers, H. (2019) - Brachionus rotundiformis Tschugunoff, 1921 from the Brachionus plicatilis species complex (Rotifera: Monogononta): A new record from Galápagos Archipelago, Ecuador. Papéis Avulsos de Zoologia, 59: e20195921. DOI:10.11606/1807-0205/2019.59.21.

MINAGRI – Ministerio de Agricultura y Riego (2016). Protocolo Nacional para el Monitoreo de la Calidad de los Recursos Hídricos Superficiales. Resolución Jefatural Nª 010-2016. MINAGRI, ANA (Autoridad Nacional del Agua). Available on-line at https://www.ana.gob.pe/sites/default/files/publication/files/protocolo_nacional_para_el_monitoreo_de_la_calidad_de_los_recursos_hidricos_superficiales.pdf.

MINAM - Ministerio del Ambiente (2017) - Aprueban Estándares de Calidad Ambiental (ECA) para Agua y establecen Disposiciones Complementarias. Decreto Supremo N° 004-2017-MINAM. Available on-line at http://www.minam.gob.pe/wp-content/uploads/2017/06/DS-004-2017-MINAM.pdf.

Oliveira, E.M.; Goldim, J.R. (2014) - Legislação de proteção animal para fins científicos e a não inclusão dos invertebrados: análise bioética. Revista Bioética, 22: 45-56. Available on-line at

https://www.scielo.br/pdf/bioet/v22n1/a06v22n1.pdf.

OECD - Organisation for Economic Cooperation and Development (2016) - Environmental Performance Reviews. Peru. Highlights and recommendations. United Nations Economic Commission for Latin America and the Caribbean (ECLAC). Available on-line at https://www.oecd.org/env/country-reviews/oecd-eclac-peru-highlights-recommendations.htm.

Orozco, R.; Castillo, S.; Enriquez, E.; Fernández, E.; Morón, O.; Flores, G. (1998) - Evaluación de los efectos de la contaminación sobre el ecosistema marino del Callao en agosto y diciembre de 1996. Informe Progresivo. Instituto del Mar del Perú, 81: 1-30. Available on-line at http://biblioimarpe.imarpe.gob.pe/handle/123456789/981.

Ortega-Salas, A.A.; Reyes-Bustamante, H. (2013) - Cultivation of the microalgae Chaetoceros gracilis to feed the rotifer Brachionus plicatilis. Cuadernos de Investigación UNED, 5: 189-193. DOI:10.22458/urj.v5i2.276.

Panduro, G.; Rengifo, G.C.; Barreto, J.L.; Arbaiza-Peña, Á.K.; Iannacone, J.; Alvariño, L.; Crnobrna, B. (2020) - Bioacumulación por mercurio en peces y riesgo por ingesta en una comunidad nativa en la amazonia peruana. Revista de Investigaciones Veterinarias del Perú, 31(3), e18177. Doi:10.15381/rivep.v31i3.18177.

Pascual, G.; Iannacone, J.; Alvariño, L. (2019) - Macroinvertebrados bentónicos y ensayos toxicológicos para evaluar la calidad del agua y del sedimento del río Rímac, Lima, Perú. Revista de Investigaciones Veterinarias del Perú, 30: 1421-1442. DOI: 10.15381/rivep.v30i4.17164.

Pérez-Legaspi, I. A.; Rico-Martínez, R. (1998) - Effect of temperature and food concentration in two species of littoral rotifers. Hydrobiologia, 387/388: 341- 348. DOI:10.1023/A:1017099906853.

Planes, E. and Fuchs, J. (2015) - Cuáles son los aportes de la ecotoxicología a las regulaciones ambientales. Ciencias e Investigación, 63: 45–62. Available on-line at http://aargentinapciencias.org/wp-content/uploads/2018/01/RevistasCeI/tomo65-2/5-Planes-cei65-2-5.pdf.

Price, M.H. (2019) - Switching to ArcGIS Pro from ArcMap. 172 p. Esri Press. Available on-line at https://esripress.esri.com/storage/esripress/images/370/switchingtoarcgispro_toc.pdf.

Rahman, A.R.A.; Cob, Z.C.; Jamari, Z.; Mohamed, A.M.; Toda, T.; Ross, O.H. (2018) - The effects of microalgae as live food for Brachionus plicatilis (rotifer) in intensive culture system. Tropical Life Sciences Research, 29: 127–138. DOI:10.21315/tlsr2018.29.1.9.

Rice, E.W.; Baird, R.B.; Eaton, A.D. (2017) - Standard Methods for the Examination of Water and Wastewater. 23 Ed. American Public Health Association, American Water Works Association, Water Environment Federation. Available on-line at https://www.wef.org/resources/publications/books/StandardMethods/.

Rico-Martinez, R.; Dingmann, B.; Snell, T.W. (1998) - Mating behavior of eight rotifer species. Hydrobiologia, 356: 165-173. DOI:10.1023/A:1003194216467.

Rico-Martínez, R.; Pérez-Legaspi, I.A.; Arias-Almeida, J.C.; Santos-Medrano, G.E. (2013) - Rotifers in Ecotoxicology. In: Encyclopedia of Aquatic Ecotoxicology. Springer-Verlag, Berlin-Heidelberg, Germany. DOI: 10.1007/SpringerReference_380004.

Roig, N.; Sierra, J.; Nadal, M.; Moreno-Garrido, I.; Nieto, E.; Hampel, M.; Perez-Gallego, E.; Schuhmacher, M.; Blasco, J. (2015) - Assessment of sediment ecotoxicological status as a complementary tool for the evaluation of surface water quality: the Ebro river basin case study. Science of The Total Environment, 503–504: 269-278. DOI:10.1016/j.scitotenv.2014.06.125.

Rojas-Jaimes, J.; Corcuera-Ciudad, R.; Iannacone, J.A. (2019) - Poisoning by inorganic mercury in minors in an urban area of illegal mining. Biotempo, 16: 113-116. DOI:10.31381/biotempo.v16i1.2182.

Sharma, J.G.; Masuda, R.; Tanaka, M.; Chakrabarti, R. (2018) - The Continuous Culture of rotifer Brachionus plicatilis with Sea water. Madridge Journal of Aquaculture Research and Development, 2: 40-42. DOI:10.18689/mjard-1000108.

Snell, W.; Moffat, B.; Janssen, C.; Persoone, G. (1991) - Acute toxicity tests using rotifers III effects of temperature strain and exposure time on the sensitivity of Brachionus plicatilis. Environmental Toxicology and Water Quality, 6: 63-75. DOI:10.1002/tox.2530060106.

Snell, T.W.; Johnston, R.K.; Matthews, A.B.; Park, N.; Berry, S.; Brashear, J. (2019) - Using Proales similis (Rotifera) for toxicity assessment in marine waters. Environmental Toxicology, 34: 634–644. DOI:10.1002/tox.22729.

Sotelo-Vásquez, D.; Iannacone, J. (2019) - Acute toxicity of three pesticides (butachlor, copper oxychloride and chlorpyrifos) on the marine benthic amphipod Apohyale grandicornis (Kroyer, 1945) (Crustacea: Hyalidae). Biotempo (Lima),16: 241–256. DOI:10.31381/biotempo.v16i2.2536.

Sun, Y.; Hou, X.; Xue, X.; Zhang, L.; Zhu, X.; Huang, Y.; Chen, Y. and Yang, Z. (2017) - Trade-off between reproduction and lifespan of the rotifer Brachionus plicatilis under different food conditions. Scientific Reports, 7: 15370. DOI:10.1038/​s41598-017-15863-y.

Toscano, E.; Severino, R. (2013) - Brachionidae (Rotifera: Monogononta) de la albufera El Paraíso y el reporte de Brachionus ibericus en el Perú. Revista peruana de biología, 20: 177–180. Available on-line at http://www.scielo.org.pe/pdf/rpb/v20n2/a11v20n2.pdf.

Tueros, I.; Borja, A.; Larreta, J.; Rodríguez, J.G.; Valencia, V.; Millán, E. (2009) - Integrating long-term water and sediment pollution data, in assessing chemical status within the European Water Framework Directive. Marine Pollution Bulletin, 58: 1389–1400. DOI:10.1016/j.marpolbul.2009.04.014.

USEPA - Environmental Protection Agency (1994) - Water topics. Available on-line at https://www.epa.gov/environmental-topics/water-topics.

USEPA - Environmental Protection Agency (2002) - Guidance for Quality Assurance Project Plans. EPA QA/G-5. EPA/240/R-02/009. EPA QA/G-5. https://www.epa.gov/sites/default/files/2015-06/documents/g5-final.pdf.

USEPA - Environmental Protection Agency (2009) - National Recommended Water Quality Criteria - Aquatic Life Criteria Table Available on-line at https://www.epa.gov/wqc/national-recommended-water-quality-criteria-aquatic-life-criteria-table.

Varea, R.; Piovano, S.; Ferreira, M. (2020) - Knowledge gaps in ecotoxicology studies of marine environments in Pacific Island Countries and Territories – A systematic review. Marine Pollution Bulletin, 156: 111264. DOI:10.1016/j.marpolbul.2020.111264.

Yona, G.K. (2018) - Effect of culture temperature on rotifers (Brachionus plicatilis sensu strictu) size and reproduction activities. Tanzania Journal of Agricultural Sciences, 17: 23-30. Available on-line at https://www.ajol.info/index.php/tjags/article/view/188252.

Zar, J. (1996) - Análisis bioestadístico. 718 p., Prentice-Hall. 3th ed. New Jersey, USA. ISBN: 0131008463 9780131008465.

 

em constru��o