Repeated exposure to noise increases tolerance in a coral reef fish.

Some anthropogenic noise is now considered pollution, with evidence building that noise 26 from human activities such as transportation, construction and exploration can impact 27 behaviour and physiology in a broad range of taxa. However, relatively little research has 28 considered the effects of repeated or chronic noise; extended exposures may result in 29 habituation or sensitisation, and thus changes in response. We conducted a field-based 30 experiment at Moorea Island to investigate how repeated exposure to playback of motorboat 31 noise affected a coral reef fish ( Dascyllus trimaculatus ). We found that juvenile D. 32 trimaculatus increased hiding behaviour during motorboat noise after two days of repeated 33 exposure, but no longer did so after one and two weeks of exposure. We also found that naïve 34 individuals responded to playback of motorboat noise with elevated ventilation rates, but that 35 this response was diminished after one and two weeks of repeated exposure. We found no 36 strong evidence that baseline blood cortisol levels, growth or body condition were affected by 37 three weeks of repeated motorboat-noise playback. Our study reveals the importance of 38 considering how tolerance levels may change over time, rather than simply extrapolating 39 from results of short-term studies, if we are to make decisions about regulation and 40 mitigation. 41


50
Some anthropogenic noise is now considered a global pollutant. As well as featuring in 51 national and international legislation (e.g. the European Commission Marine Strategy 52 Framework Directive and the United States National Environmental Policy Act), mounting 53 evidence shows that anthropogenic noise can impact behaviour (e.g. vocal communication, 54 anti-predator defence, foraging) and physiology (e.g. ventilation rate, metabolic rate, heart 55 rate) in at least some species from a broad range of taxa (Shannon et (Wale et al., 2013)). This is an important 62 consideration in the context of regulation, because human disturbance of natural habitats is 63 becoming more frequent and the pervasive nature of anthropogenic noise means that animals 64 are likely to be exposed multiple times during their lifetime. 65 Research in other fields reveals that animal responses to various stimuli can change one explanation for which could be habituation (Thorpe, 1963). For example, male white-  (Petrinovich and Patterson, 1979). If animals continue to respond to stimuli, they could 75 become chronically stressed (Cyr and Romero, 2009), with potential downstream effects on 76 growth and condition (Anderson et al., 2011). If an animal habituates fully to a stressor, 77 baseline cortisol concentration, behaviour and health will be the same as unstressed animals 78 (Cyr and Romero, 2009). Experimental data with repeat measures from the same individuals 79 over time are lacking in field studies of anthropogenic noise, so whether animals are able to 80 habituate to this stressor is unknown. 81 We used a field-based experiment on a coral reef fish to investigate the effects of variables such as extraneous noise, temperature and water quality, the acoustics of small 100 tanks mean that relevant sound exposure levels are very difficult to measure and control 101 (Parvelescu, 1967). However, field studies offer greater ecological relevance.

102
In this study, we exposed juvenile coral reef fish in their natural habitat to playbacks stress. Specifically, we tested whether hiding behaviour and ventilation rate responses to 112 motorboat-noise playback differed after repeat exposure. We predicted that these responses 113 would be heightened if fish tolerance to playbacks decreased, while these responses would 114 attenuate if tolerance increased. We also measured fish size, condition and baseline plasma 115 cortisol concentrations to test the longer term consequences of any change in tolerance to 116 repeated playback of motorboat noise.   were two different replicate playbacks for each treatment. Sound pressure levels (measured 154 using the hydrophone set-up described above) and particle acceleration levels (measured    week 2 were sacrificed using an overdose of MS222 before standard length (measured in the 236 same way) and wet mass (measured using a balance, to the nearest 0.001 g). were measured.

237
These were used to calculate condition factor using the following formula:  the entry of small amounts of blood into the capillary tube). After the sample was taken, a 253 further 18 µl of the EIA buffer was added to the capillary tube to achieve the desired dilution.

254
Samples were kept on ice until they were centrifuged for 10 min at 10 000 rpm, to separate 255 the plasma.   greater increase in OBR compared to ambient-noise playback, that effect was significantly 326 lessened by long-term exposure to playback of motorboat noise (Table 1; Fig. 5).

342
We found a behavioural and a physiological response to motorboat-noise playback in the 343 short term: after two days of exposure, juvenile Dascyllus trimaculatus were more likely to 344 hide during the period of a motorboat-pass playback than in the period immediately before, 345 and naïve fish (those that had not experienced motorboat-noise playback before) also showed 346 an increased ventilation rate (opercular beat rate, OBR) in response to noise in the short term

347
(1 min exposure). Our results concur with other studies that have found short-term The regime of sound exposure in our experiment was highly regular; one motorboat    Playback of ambient noise matched local ambient noise relatively well in sound pressure. As 591 the experiment was conducted on a sand flat, most ambient sound sources were not nearby, 592 particle acceleration ambient noise levels were thus lower as the particle motion component 593 of sound drops off with distance from the sound source faster than the pressure component.

594
Playbacks were affected by near-field effects and speaker performance meaning some 595 frequencies were louder and others quieter, but motorboats were louder than ambient noise 596 and motorboat-noise playbacks were louder than ambient-noise playbacks. Some recordings 597 contained harmonic noise at 50 Hz intervals; this was an artefact of recording via a laptop.