Opposite and synergistic physiological responses to water acidity and predator cues in spadefoot toad tadpoles

https://doi.org/10.1016/j.cbpa.2020.110654Get rights and content

Highlights

  • Predator cue recognition occurred regardless of water pH, as tadpoles developed anti-predator morphology even at pH = 4.

  • Predator presence induced lower levels of plasma corticosterone in tadpoles whereas acidic water increased them.

  • Predator presence reduced the activity of some antioxidant enzymes, congruent with a decrease in metabolism.

  • The combination of predator cues and low pH had negative synergistic effects on the redox status of tadpoles.

Abstract

Organisms are exposed to multiple environmental factors simultaneously to which they often respond behaviorally, morphologically and/or physiologically. Amphibian larvae are quite plastic and efficiently adjust their phenotype and physiology to the reigning local conditions. Here we tested whether the combination of predator presence and low water pH induces alterations in the morphology and physiology of spadefoot toad tadpoles. We raised Pelobates cultripes tadpoles in the laboratory in water at either pH 4 or 7, and in the presence or absence of caged dragonfly nymphs, and determined their changes in shape through geometric morphometrics to assess whether predator recognition was impaired or not at low pH. We also measured levels of plasma corticosterone, activity of four antioxidant enzymes, as well as markers of oxidative damage and redox status. We found that tadpoles altered their body shape in response to predator cues even at low pH, indicating that predator recognition was not interfered by water acidity and developmental responses were robust even under abiotic stress. Water acidity was associated with increased corticosterone levels in tadpoles, whereas predator presence consistently reduced corticosterone levels. Predator presence was linked to reduced antioxidant enzyme activity, whereas the combination of both factors resulted in negative synergistic effects on lipid peroxidation and the antioxidant capacity of tadpoles. Here we show that tadpoles detect predators even at low pH but that the development of adaptive anti-predatory morphology can magnify physiological imbalances when other stressors co-occur. These results emphasize the need to understand how multiple environmental perturbations can affect animal homeostasis.

Introduction

Antipredator responses are a widespread form of adaptive phenotypic plasticity that allows prey to alter their phenotype upon detection of predator cues (Agrawal, 2001; Tollrian and Harvell, 1998). These responses can be fine-tuned to the level of predation risk experienced (McCoy et al., 2012; Van Buskirk and Arioli, 2002) indicating the existence of accurate and reliable mechanisms of cue detection. Predator cue recognition, however, is critically dependent upon the joint evolutionary history between prey and predator (Ferrari et al., 2010b; Relyea, 2004). Because organisms live in rather complex environments, multiple factors can compromise the efficiency of inducible defenses even in the case of predator-prey systems with ample joint evolutionary history.

Stressful abiotic conditions (e.g. salinity, extreme pH or temperature) in particular have a two-fold potential for disruption of inducible defenses: they can interfere with cue quality by denaturalizing the cue itself or inhibiting its receptors (Troyer and Turner, 2015), or they can hamper antipredator responses by causing additional physiological stress (Hawlena and Schmitz, 2010). Exposure to multiple sources of stress could impair the organism's ability to develop adaptive antipredator phenotypes. For instance, altered water chemistry or pollution can hinder cue detection and activation of inducible defenses (Burraco et al., 2018; Ferrari et al., 2010a; Lürling and Scheffer, 2007; Polo-Cavia et al., 2016; Gabor et al., 2019), and it can also increase the costs of producing inducible defenses (Pestana et al., 2010; Teplitsky et al., 2005). Therefore the efficiency of inducible defenses depends on accurate and unobstructed predator cue recognition, but a sufficient body condition to overcome potential production costs of inducible defenses is also needed (Milot et al., 2014; Murren et al., 2015). From that perspective, additional stressors may prevent an appropriate expression of inducible defenses as a consequence of allocating resources to the maintenance of metabolic pathways linked to an induced stressful state, and away from the development of adaptive antipredator responses (Killen et al., 2013). Indeed, poorer body condition often entails increased vulnerability to predation (Hoey and McCormick, 2004; Murray, 2002). Conversely, antipredator responses can make prey more vulnerable to additional stressors, such as pesticides or viruses (Kerby et al., 2011; Relyea, 2003).

Here we study whether antipredator responses in larval amphibians are affected by simultaneous exposure to water acidity, using spadefoot toad tadpoles (Pelobates cultripes). Amphibians are sensitive to multiple stressors, often experiencing the activation of the hypothalamic-pituitary-interrenal (HPI) axis in response to them, which involves a physiological cascade that can induce oxidative stress and immunological imbalances (Burraco and Gomez-Mestre, 2016; Gervasi and Foufopoulos, 2008; Groner et al., 2013). However, the activation of the HPI-axis may be context dependent and result in unexpected dynamics (e.g. Gabor et al., 2018). The fact that multiple stressors may coincide, and the consequences of exposure to various concurrent stressors may be additive or even synergistic, combining to pose a much greater threat to amphibians than each factor individually (Blaustein et al., 2011; Boone et al., 2007; Davidson and Knapp, 2007). Water acidity and predator presence are common stressors to amphibians (Egea-Serrano et al., 2014), and they are the focus of this study, since their joint effect on amphibians has not been frequently studied. Low pH commonly poses a grave risk to embryonic and larval amphibians, causing an imbalance in their ionic regulation (Freda, 1986; Rasanen et al., 2002, Rasanen et al., 2003; Sadinski and Dunson, 1992). Water acidity is known to alter intraguild predator-prey interactions between amphibian species (Kiesecker et al., 1996), and to have very different impacts on swimming performance, growth and development across species (Freda and Dunson, 1985; Kutka, 1994; Rowe et al., 1992). Few studies have directly assessed the physiological stress responses of amphibian larvae to low pH, but they indicate that water acidity tends to increase their corticosterone levels (Burraco and Gomez-Mestre, 2016; Chambers et al., 2013). Likewise, the physiological stress responses to predator presence in amphibian larvae still require further research to confirm whether patterns found to date are dependent upon species identity, exposure duration, or a specific developmental stage. Wood frog (Rana sylvatica) tadpoles increase their corticosterone levels in the non-lethal presence of predators (Middlemis Maher et al., 2013), although this response may be dependent on the amount of time that tadpoles spend exposed to predator cues (Bennett et al., 2016). In contrast, green frog (Rana clamitans) and spadefoot toad tadpoles lower their corticosterone levels in response to predator cues (Burraco et al., 2013; Burraco and Gomez-Mestre, 2016; Fraker et al., 2009). In addition, the common frog (Rana temporaria) shows geographic and temporal variation in whether predators induce increased or decreased corticosterone levels in its tadpoles (Dahl et al., 2012). Our experiment was designed to test for additive or synergistic effects of low pH and predator presence on stress physiology of spadefoot toad tadpoles after chronic exposure to both sources of stress, quantifying the phenotypic responses of tadpoles to predators in both neutral and acidic water, their corticosterone levels and their oxidative status. We expected acidic water to interfere with the ability of tadpoles to detect predator cues. We also expected exposure to predator cues and acidic water to be associated with lower and higher corticosterone levels, respectively, and that both factors would involve redox imbalances in amphibian larvae. Finally, we predicted that the combination of acidic water and predator cues would magnify the individual effect of each factor, either in an additive or a synergistic way.

Section snippets

Methods and materials

This study was conducted at the Doñana Biological Reserve, located within the Doñana National Park, on the right bank of the Guadalquivir river mouth in southwestern Spain. The study area has a Mediterranean climate with an Atlantic influence, having hot and dry summers and rainfall occurring mostly in autumn or winter, from November to March (mean annual precipitation of 544.6 mm ± 211.3 mm; Diaz-Paniagua et al., 2010). The park comprises extensive marshes and a sandy area with shrubland,

Induced morphological responses despite water acidity

Using geometric morphometrics, we assessed the degree to which spadefoot toad tadpoles detected and responded to predator cues under either neutral water or acidic water conditions. Variation in the first relative warp (RW1) was strongly influenced by presence/absence of predators, regardless of water pH, whereas RW2 captured variation in tadpole shape was not associated with any of the experimental treatments (Fig. 1A). A linear model using RW1 as dependent variable indicated that tadpoles

Discussion

Contrary to our expectations, spadefoot toad tadpoles were capable of accurately detecting predator cues and developing an anti-predatory morphology in acidic water. This suggests that the cue recognition system is quite robust and that even at pH = 4 the cue remains recognizable and the tadpoles' receptors continue to be operative. Other studies have shown that chemoreception in aquatic systems can be obstructed by acidic substances such as humic acid, both in cases of conspecific recognition (

Acknowledgments

We are thankful to M. Merchand for his laboratory assistance during the experiment, as well as to C. Megía and F. Miranda for their assistance processing the samples for oxidative stress. Laboratory facilities were provided by ICTS-RBD. This research was funded by Plan Nacional I + D grant #CGL2017-83407-P.

References (84)

  • J. Menon et al.

    Oxidative stress, tissue remodeling and regression during amphibian metamorphosis

    Comp. Biochem. Physiol. Part C Toxicol. Pharmacol.

    (2007)
  • J.L.T. Pestana et al.

    Pesticide exposure and inducible antipredator responses in the zooplankton grazer, Daphnia magna Straus

    Chemosphere

    (2010)
  • N. Polo-Cavia et al.

    Low levels of chemical anthropogenic pollution may threaten amphibians by impairing predator recognition

    Aquat. Toxicol.

    (2016)
  • M.D. Prokić et al.

    Oxidative stress in Pelophylax esculentus complex frogs in the wild during transition from aquatic to terrestrial life

    Comp. Biochem. Physiol. Part A Mol. Integr. Physiol.

    (2019)
  • L. Serrano et al.

    Susceptibility to acidification of groundwater-dependent wetlands affected by water level declines, and potential risk to an early-breeding amphibian species

    Sci. Total Environ.

    (2016)
  • D.K. Skelly

    Activity level and the susceptibility of anuran larvae to predation

    Anim. Behav.

    (1994)
  • D.C. Adams et al.

    Geomorph: an R package for the collection and analysis of geometric morphometric shape data

    Methods Ecol. Evol.

    (2013)
  • A.A. Agrawal

    Phenotypic plasticity in the interactions and evolution of species

    Science

    (2001)
  • R. Arribas et al.

    Ecological consequences of amphibian larvae and their native and alien predators on the community structure of temporary ponds

    Freshw. Biol.

    (2014)
  • M. Barry et al.

    Metabolic responses of tadpoles to chemical predation cues

    Hydrobiologia

    (2013)
  • M.F. Benard

    Predator-induced phenotypic plasticity in organisms with complex life histories

    Annu. Rev. Ecol. Syst.

    (2004)
  • A.R. Blaustein et al.

    The complexity of amphibian population declines: understanding the role of cofactors in driving amphibian losses

    Ann. N. Y. Acad. Sci.

    (2011)
  • M.D. Boone et al.

    Multiple stressors in amphibian communities: effects of chemical contamination, bullfrogs, and fish

    Ecol. Appl.

    (2007)
  • P. Burraco et al.

    Physiological stress responses in amphibian larvae to multiple stressors reveal marked anthropogenic effects even below lethal levels

    Physiol. Biochem. Zool.

    (2016)
  • P. Burraco et al.

    Predator-induced physiological responses in tadpoles challenged with herbicide pollution

    Curr. Zool.

    (2013)
  • P. Burraco et al.

    Comparing techniques for measuring corticosterone in tadpoles

    Curr. Zool.

    (2015)
  • P. Burraco et al.

    Eucalypt leaf litter impairs growth and development of amphibian larvae, inhibits their antipredator responses and alters their physiology

    Conserv. Physiol.

    (2018)
  • D.L. Chambers et al.

    Pond acidification may explain differences in corticosterone among salamander populations

    Physiol. Biochem. Zool.

    (2013)
  • G. Cohen et al.

    Catalase-aminotriazole method for measuring secretion of hydrogen peroxide by microorganisms

    J. Bacteriol.

    (1969)
  • M.L. Collyer et al.

    A method for analysis of phenotypic change for phenotypes described by high-dimensional data

    Heredity (Edinb).

    (2015)
  • J.M. Cord et al.

    An enzymatic function for Erythrocuprein (Hemocuprein)

    J. Biol. Chem.

    (1969)
  • D. Costantini

    Understanding diversity in oxidative status and oxidative stress: the opportunities and challenges ahead

    J. Exp. Biol.

    (2019)
  • E. Dahl et al.

    Geographic variation in corticosterone response to chronic predator stress in tadpoles

    J. Evol. Biol.

    (2012)
  • C. Davidson et al.

    Multiple stressors and amphibian declines: dual impacts of pesticides and fish on yellow-legged frogs

    Ecol. Appl.

    (2007)
  • C. Diaz-Paniagua et al.

    Hatching success, delay of emergence and hatchling biometry of the spur-thighed tortoise, Testudo graeca, in South-Western Spain

    J.Zool.Lond.

    (1997)
  • C. Diaz-Paniagua et al.

    Los anfibios de Doñana, Naturaleza y Parques Nacionales

    (2005)
  • C. Diaz-Paniagua et al.

    Temporay ponds from Doñana National Park: a system of natural habitats for the preservation of aquatic flora and fauna

    Limnetica

    (2010)
  • A. Egea-Serrano et al.

    Multifarious selection through environmental change: acidity and predator-mediated adaptive divergence in the moor frog (Rana arvalis)

    Proc. R. Soc. B Biol. Sci.

    (2014)
  • M.C.O. Ferrari et al.

    Chemical ecology of predator-prey interactions in aquatic ecosystems: a review and prospectus

    Can. J. Zool.

    (2010)
  • H.S. Fisher et al.

    Alteration of the chemical environment disrupts communication in a freshwater fish

    Proc. R. Soc. B-Biological Sci.

    (2006)
  • J. Freda

    The influence of acidic pond water on amphibians: a review

    Water, air, soil Pollut. Historical Arch.

    (1986)
  • J. Freda et al.

    Field and laboratory studies of ion balance and growth rates of ranid tadpoles chronically exposed to low pH

    Copeia

    (1985)
  • Cited by (0)

    View full text