Vol. 7: 115–123, 2015 AQUACULTURE ENVIRONMENT INTERACTIONS Published online September 3 doi: 10.3354/aei00141 Aquacult Environ Interact OPENPEN ACCESSCCESS Effect of temperature on the metabolism, behaviour and oxygen requirements of Sparus aurata Mette Remen1,*, Marit A. J. Nederlof2, Ole Folkedal1, Grethe Thorsheim1, Ariadna Sitjà-Bobadilla3, Jaume Pérez-Sánchez3, Frode Oppedal1, Rolf Erik Olsen1,4 1Institute of Marine Research, 5984 Matredal, Norway 2Department of Aquaculture and Fisheries, Wageningen University, De Elst 1, 6708 WD Wageningen, The Netherlands 3Institute of Aquaculture Torre la Sal (IATS-CSIC), 12595 Ribera de Cabanes, Castellón, Spain 4Present address: Norwegian University of Science and Technology, Department of Biology, 7491 Trondheim, Norway ABSTRACT: We investigated the effect of temperature on the limiting oxygen saturation (LOS) of gilthead sea bream Sparus aurata. This threshold was defined as the % O2 saturation where fish no longer upheld their routine metabolic rate (RMR, the metabolic rate of fed and active fish) dur- ing a progressive decline in oxygen saturation. S. aurata (398 ± 10 g, mean ± SE) were kept in 3 replicate tanks and subjected to 3 changes in temperature: 16 to 20°C, 20 to 16°C and 16 to 12°C. At each temperature, fish were left to acclimatize for 8 to 10 d, before daily feed intake (DFI), the −1 −1 routine oxygen consumption rate (routine MO2, mg kg min ) and the LOS were measured. In addition, at 20°C the swimming speed was measured in fish subjected to a decline in O2 from full air saturation to levels below the LOS (minimum of 8−10% O2). For the temperature range tested (12−20°C), DFI, MO2 and LOS increased exponentially with temperature (7.5-, 3.6- and 2.2-fold, respectively) with mean (± SE) LOS being 17 ± 1, 21 ± 0 and 35 ± 5% O2 at 12, 16 and 20°C, respec- tively. A gradual decline in swimming activity was observed as O2 declined below the LOS, indi- cating increasing metabolic stress and/or a ‘sit-out’ coping strategy which may prolong survival time in severe hypoxia. The results show the importance of temperature as an influential variable over the environmental O2 requirements of S. aurata. KEY WORDS: Hypoxia · Aquaculture · Metabolism · Behaviour · Pcrit · Scrit · Oxygen threshold · Feeding rate · Temperature INTRODUCTION obic metabolism (Fry 1947, 1971). Studies in Atlantic salmon sea cages have revealed that O2 may drop to Gilthead sea bream Sparus aurata is an important alarmingly low levels, at times down to 30% O2. aquaculture species in countries surrounding the Factors contributing to these low O2 levels include Mediterranean Sea, with 154 000 t produced glob- water temperature, fish stocking density, algal den- ally in 2011 (FAO 2013). The growth phase is prima- sity and water exchange rate (Crampton et al. 2003, rily carried out in floating sea cages (Basurco et al. Johansson et al. 2006, 2007, Oppedal et al. 2011). 2011) where fish performance and welfare are There is currently little information on the sea cage closely linked to environmental conditions within O2 levels in S. aurata aquaculture, but variation in the sea cage (Fry 1971, Huntingford & Kadri 2008). the above mentioned factors is expected to cause The sea cage oxygen (O2) level is particularly variable levels of O2 in S. aurata production systems important as it is the main limiting factor of fish aer- as well. In order to develop production strategies © The authors 2015. Open Access under Creative Commons *Corresponding author: [email protected] by Attribution Licence. Use, distribution and reproduction are un restricted. Authors and original publication must be credited. Publisher: Inter-Research · www.int-res.com 116 Aquacult Environ Interact 7: 115–123, 2015 that ensure proper physiological function and sur- The LOS is expected to increase with any factor vival of the fish, it is important to define a limit for that increases the aerobic metabolic rate (Fry 1971, acceptable drops in O2. Neill et al. 1994). One of the main factors influencing When subjected to a gradual decline in O2, sparid metabolic rate is temperature (Fry 1947, 1971), which species are able to uphold their oxygen consumption is closely correlated with LOS and Scrit (e.g. Schur- rate (MO2) over a relatively wide range of O2 satura- mann & Steffensen 1997, Claireaux & Lagardère tions (Cerezo & García García 2004, Valverde et al. 1999, Barnes et al. 2011, Richards 2011, Mamun et al. 2006). They do this mainly by increasing gill ventila- 2013, Remen et al. 2013). In order to develop an tion and perfusion (Perry et al. 2009). At a certain applicable oxygen threshold for the S. aurata indus- level of O2, regulatory mechanisms are no longer suf- try, it is therefore necessary to investigate the rela- ficient to uphold MO2 and it starts to decrease with tionship between temperature and LOS for this spe- further reductions in O2 (Pörtner & Grieshaber 1993). cies. Further, it is important to do so under conditions This threshold is termed the critical oxygen satura- as similar to production conditions as possible (fed, tion (Scrit or Pcrit if O2 is presented in units of gas pres- undisturbed and swimming fish in groups), as RMR, sure, when determined in resting, fasted fish at a and thus the LOS, can be expected to vary consider- known, stable temperature, i.e. in fish with standard ably, e.g. with meal size (e.g. Guinea & Fernandez meta bolic rates (Schurmann & Steffensen 1997, 1997) and swimming speed (e.g. Steinhausen et al. Claire aux & Lagardère 1999, Behrens & Steffensen 2010) at a given temperature. 2007). However, when this threshold is determined The main aim of this study was to determine the in fed fish engaged in voluntary swimming activity, relationship between temperature (12−20°C) and i.e. fish with routine metabolic rates (RMRs), this LOS for S. aurata. To mimic an aquaculture setting, threshold has been termed the limiting oxygen satu- LOS was determined in fed and undisturbed fish. ration (LOS). The LOS departs from Scrit in being a Further, swimming speed and behaviour of S. aurata continuum which correlates with RMR at a given at O2 declining from full saturation to levels below temperature (e.g. Claireaux et al. 2000, Claireaux & the LOS were investigated in order to study the La gardère 1999, Remen et al. 2013). The LOS may behavioural response of this species to a progressive therefore change according to swimming speed drop in O2. (Stein hausen et al. 2010), specific dynamic action (SDA; the post-prandial increase in metabolism; reviewed by Secor 2009) or other factors that cause MATERIALS AND METHODS variation in RMR. In the present paper, the threshold O2 level below which MO2 decreases is presented as All experimental work was conducted in accor- LOS, since it is determined in fed S. aurata engaged dance with the laws and regulations for experiments in voluntary swimming activity, and because units of and procedures on live animals in Norway, following oxygen saturation (% of air saturation) are most com- the Norwegian Regulation on Animal Experimenta- monly used by aquaculturists. tion 1996. The experiment was approved by Forsøks- When O2 decreases below the LOS, the fish must dyrutvalget (FOTS ID 4580). increase their anaerobic metabolic rate to compen- sate for the decreasing ATP production (reviewed by Pörtner & Grieshaber 1993). Anaerobic ATP produc- Fish material and experimental facilities tion is far less efficient than aerobic ATP production, and the duration of survival becomes dependent on Sparus aurata juveniles (5 g) were purchased from the availability of substrates for anaerobiosis Ferme Marine de Douhet hatchery, Ile d’Oléron, (Richards 2009, 2011). Further, a drop in O2 to levels France, and transported to the Institute of Aquacul- below the LOS can be expected to induce stress ture Torre de la Sal in Spain in July 2011. Fish were (increased plasma levels of corticosteroids and cate- fed to satiety (EFICO YM 554, BioMar) and kept in an cholamines) and lactic acidosis (Van Raaij et al. 1996, open flow system (salinity 37.5 ppt) with natural pho- Vianen et al. 2001, Petersen & Gamperl 2011, Remen toperiod and water temperature until they reached et al. 2012, 2013). The LOS can therefore be imple- 150 g. They were then transported to Matre Research mented in aquaculture as the lower limit for accept- Station, Institute of Marine Research in Norway in able O2 levels with regard to the physiological func- January 2012. No fish died during transport. On tion and welfare of farmed fish (Huntingford & Kadri arrival, a total of 99 fish were distributed into three 2008). 500 l squared flow-through experimental tanks with Remen et al.: Temperature and O2 requirements of S. aurata 117 lids fitted with 18W fluorescent light tubes and auto- (12−20°C) was used because 12°C is a minimum tem- matic feeders (Arvo-Tec T drum 2000, www.arvotec.fi). perature for growth (Mingarro et al. 2002, Hernán- Feed was provided twice daily during the entire dez et al. 2003) and because 20°C was the maximum experiment (Amber Neptun 100, 5 mm, Skretting, temperature possible in the laboratory. The change Norway), aiming for >30% overfeeding. Spill feed in temperature was changed in a stepwise manner at was weighed, and feed intake was estimated accord- a rate of 2°C d−1. At each new temperature (20, 16, ing to the method of Helland et al. (1996). Calcula- 12°C) the fish were left to acclimatize for 8 to 10 d tions for estimation of the tank biomass over time are before measurements of MO2 and LOS (Days 12, 20 shown in ‘Calculations and statistics’ below. Fish and 28, respectively). After the final LOS measure- were kept on a 12:12 h light:dark h cycle at 34 ppt, ment on Day 28, all fish were anaesthetized (MS-222, −1 16°C and O2 saturation > 80% until fish reached a 150 mg l ) and weights and lengths measured.