J. Nucl. Phy. Mat. Sci. Rad. A.

In vitro Study of the Survival, Reproduction and Morphology of Daphnia pulicaria irradiated with a Low Energy Laser

F. Zaldivar, J. Montoya, S. Gonzalez, L.A. Mandujano, J. F. Mendez-Sanchez, L. Romero, J. Mulia, M. Paulin and D. Osorio-Gonzalez


Daphnia, laser irradiation, morphology

PUBLISHED DATE August 6, 2018
PUBLISHER The Author(s) 2018. This article is published with open access at www.chitkara.edu.in/publications.

Daphnia is a genus of crustaceans that is representative of freshwater communities. The species exhibit a high sensitivity to a wide range of toxic compounds so that they have been used internationally as biomonitors in toxicity tests to evaluate ecosystem conditions such as water quality. It is also a model genus in genetics, epigenetics and reproductive ecology. In this work, we used Daphnia pulicaria as a model to measure the effects of low-energy laser irradiation on survival, reproduction, and morphology variables of parental organisms and their offspring. We used (1) a single clone line of organisms to eliminate interindividual genetic variability; (2) individuals from more than 50 generations after the clone line was established, and offspring from the third brood onwards to dissipate maternal and epigenetic effects, and (3) neonates, those individuals of the species that have less than 48 hours of life, because they are the most sensitive stage to optical stimuli. We analyzed number of deaths, longevity, age at first reproduction, number of offspring per week, number of total offspring during all their life cycle, body size, size of the antennules, and length of the apical spine of the 4th and 5th brood of the irradiated individuals, who were exposed to a blue laser stimulus of 405 nm for 25 minutes with a power of 40 mW at a distance of 50 cm, compared to those of the control (non-irradiated) group.


Daphnia is worldwide considered a model organism in genetics, epigenetics and reproductive ecology experiments aside from being an excellent toxicity bioindicator in aquatic environments since they are the main representatives of the freshwater communities. By exhibiting high sensitivity to a wide range of toxic compounds, they have been internationally used in toxicity tests to measure and evaluate the conditions of an ecosystem. Besides, its adaptability and culture maintenance in the laboratory are relatively simple allowing the performance of quick and economic tests [1]. The reproductive cycle of Daphnia is one of its principal and greatest advantages as it is directly related to environmental stimuli. If the environmental conditions are ideal, Daphnia will carry out its reproduction through a parthenogenetic or asexual cycle, whereas when the environmental conditions are disturbed, its reproductive cycle will change to a sexual one.
Daphnia magna, Daphnia exilis, Daphnia pulex, Ceriodaphnia dubia and Moina macrocopa are used as toxicological models to measure the median lethal concentration (LC50), particularly of agricultural and industrial chemical products [2]. The toxins tend to accumulate in the flea body, and their subsequent effect is reflected as oxidative stress or damage. This property can be quantified through specific biomarkers, including antioxidant enzymatic defenses such as superoxide dismutase and catalase activity which are considered the first and second line of defense against reactive oxygen species. Oxidative damage can also be determined through the level of lipid peroxidation since lipids -especially unsaturated ones- are the molecules most susceptible to oxy-radicals. Moreover, in aquatic species, there is a close relationship between the increase in lipid peroxidation and oxidative deterioration [3].
Previous studies have exposed Daphnia to various toxins and evaluated their response to oxidative stressors as those generated from cyanobacteria blooming, artificial mixtures of cyanotoxins [3], exposition to wastewater effluents under ozonation treatments, and to advanced oxidation processes for removing drugs and their metabolites [4,5]. The results showed that exposure to toxic compounds might cause genetic or epigenetic changes in future generations [6,7]; such changes evaluated through differences in survival rate, longevity, growth, reproduction, and morphology. Another study conducted by Kushibiki et al. [8] showed that low-energy laser therapy induces a signaling cascade of second messengers which leads to the production of reactive oxygen species in irradiated cells, especially with a 405 nm blue laser. The increase in reactive oxygen species, adenosine triphosphate, and/or adenosine monophosphate concentrations seems to be to the fact that the components of the respiratory chain can absorb the energy of the laserinduced photon in the mitochondria. This phenomenon also promotes cell proliferation and cytoprotection. The benefits experimented by the cell and the production of reactive oxygen species depends on the exposure time, pulse frequency and dose delivered.

Page(s) 109-113
URL http://dspace.chitkara.edu.in/jspui/bitstream/123456789/749/1/19_JNP.pdf
ISSN Print : 2321-8649, Online : 2321-9289
DOI 10.15415/jnp.2018.61019

The irradiation with a solid-state blue laser (wavelength of 405 nm) for a time of 25 minutes and with a power of 40 mW produces significant changes in the survival, reproduction, and morphology of Daphnia pulicaria. This dose was lethal for 70% of the irradiated neonates and decreased the total number of offspring of the surviving individuals. The morphology was affected in neonates of the 4th and 5th brood of different irradiated mothers, where there was a decrease in the size of the body, a significant increase in the apical spine and a tendency to reduce the size of the antennule.

  • F. Martínez, R. Villase-or, F. Espinosa, G. Ríos, and R. Ramírez. Methodology for the production of neonates of Daphnia magna (Anomopoda: Daphnidae), as a test organism in aquatic toxicology. Zoology informs, 36–37, 59–81 (1997).
  • F. Martínez-Jerónimo, J. Rodríguez-Estrada, L. and Martínez-Jerónimo. Daphnia exilis Herrick, 1895 (Crustacea: Cladocera). A zooplankton species potentially useful as a test organism in acute toxicity bioassays in tropical and subtropical environments. International Journal of Environmental Pollution, 24 (4), 153–159 (2008).
  • Maranda Esterhuizen-Londt, Marie Von Schnehen, Sandra Kühn, Stephan Pflugmacher. Oxidative stress responses in the animal model, Daphnia pulex exposed to a natural bloom extract versus artificial cyanotoxin mixtures. Aquatic Toxicology, 179, 151–157 (2016). https://doi.org/10.1016/j.aquatox.2016.09.003
  • Ana Lourdes Oropesa, Sara C Novais, Marco F. L Lemos, Azahara Espejo, Carlos Gravato, Fernando Beltrán. Oxidative stress responses of Daphnia magna exposed to effluents spiked with emerging contaminants under ozonation and advanced oxidation processes. Environmental Science and Pollution Research, 24(2), 1735–1747 (2016). https://doi.org/10.1007/s11356-016-7881-9
  • A. L. Oropesa, A. M. Floro, P. Palma. Assessment of the effects of the carbamazepine on the endogenous endocrine system of Daphnia magna. Environmental Science and Pollution Research, 23(17), 17311–17321 (2016). https://doi.org/10.1007/s11356-016-6907-7
  • Warren W. Burggren. Dynamics of epigenetic phenomena: intergenerational and intragenerational phenotype ‘washout’. Journal of Experimental Biology, 218:80–87 (2015). https://doi.org/10.1242/jeb.107318
  • Kami D. M. Harris, Nicholas J. Bartlett, Vett K. Lloyd. Daphnia as an Emerging Epigenetic Model Organism. Genetics Research international, 2012, 8 pp (2012). http://dx.doi.org/10.1155/2012/147892
  • Toshihiro Kushibiki, Takeshi Hirasawa, Shinpei Okawa, Miya Ishihara. Blue Laser Irradiation Generates Intracellular Reactive Oxygen Species in Various Types of Cells. Photomedicine And Laser Surgery, 31(3), 95–104 (2013). https://doi.org/10.1089/pho.2012.3361
  • Secretariat of economy. Official standard NMX-AA- 087-scfi-2010. Water analysis–acute toxicity evaluation with daphnia magna, straus (crustacea – cladocera) test method 44 pp (2010).
  • Sarah J. Andrewartha and Warren W. Burggren. Transgenerational Variation in Metabolism and Life-History Traits Induced by Maternal Hypoxia in Daphnia magna. Physiological and Biochemical Zoology 85(6), 625–634 (2012). https://doi.org/10.1086/666657
  • Warren Burggren. Epigenetic Inheritance and Its Role in Evolutionary Biology: Re-Evaluation and New Perspectives. Biology 5(2), 24 (2016). https://doi.org/10.3390/biology5020024
  • Peter Stiling, Ecology: Global Insights & Investigations. Second edition. New York: McGraw-Hill Science Engineering (2014).