Final Report Summary - SCENT (Scaning chemical pollution as an ecological impact of non-native fish introductions: an experimental approach)
SCENT corresponds to an area of science where the full implication of pheromone communication has been overlooked whilst at the same time, it introduces the concept of pheromone pollution to test its effects and consequences for biological invasions and European fish biodiversity. Biological invasion is a major issue in the conservation of global biodiversity and represents a worldwide threat to ecosystem functioning and structure. The number of species introductions has dramatically increased in the last decades, however, the issue of pheromone pollution across species caused by invasive fishes had not been studied despite the high risk of ecological impact. Amongst the most impressive fish invasions in European inland waters in recent decades are those of the topmouth gudgeon Pseudorasbora parva. It is one of only three existing cyprinid species that have evolved nest guarding behaviours and each species is native to a different continent (i.e. P. parva Asia, L. delineatus Europe and P. promelas North America). Previous investigations demonstrated that reproductive chemical communications are in operation among these species. The overall aim of SCENT was to examine the concept of pheromone pollution as a facilitator in the establishment of P. parva in novel ecosystems through an experimental study of reproductive pheromone interactions with L. delineatus and P. promelas. The initial objectives of the project were: 1) That urinary pheromone releases by P. parva do not disrupt L. delinateus and P. promelas reproductive behaviour; 2) That reproductive pheromones released by P. parva do not disrupt the individual reproductive fitness of L. delinateus and P. promelas; and 3) Patterns of urinary response of reproductive P. parva females and males remain unchanged in the presence of L. delinateus and P. promelas. The final objective (4) was to identify the chemical structure of the sexual pheromones of P. parva. An extensive effort was made to achieve the stated objectives, although due to the first year experimental outcome some adjustments were made in the 2nd year experimental set up along with a new timetable. L. delinateus failed to reproduced in the first experimental set up despite having been successfully reproduced in the past in laboratory conditions. Thus, experimental design focused on the interactions between P. parva and P. promelas only. The intension to mark and monitor P. parva and P. promelas behaviour with a Passive Integrated Transponder (PIT) or colour tags could not be implemented for technical reasons. Consequently, it was decided to monitor fish behaviour using CCTV video cameras. An additional funding by a ‘Small Research Grant’ of the Fishery Society of the British Isles was achieved in support of the development of new experimental setup. During the first year of the fellowship Dr Kakareko focused his effort on designing and calibrating the new experimental setup, conducting the experiments to complete the objectives 1 and 2, and isolating active compounds from conditioned water derived from male P. parva for further analysis (objective 4). An advanced cooling system of stock tanks was engineered (this was crucial to control reproductive condition of P. parva). Indoor flow-through aquaria facilities with cameras attached from top view were established and calibrated. From May till July 2012 (i.e. spawning period of P. parva) series of experiments in tanks were conducted. The results of the experiments did not show a clear pattern of cross-species effect of pheromones on reproduction, and there were still issues that needed to be resolved before producing a comprehensive paper e.g. defining pre- and post-spawning patterns of fish behaviour, characterizing fish behaviour inside a nest. Therefore, during the second year of the fellowship a modified experimental set up was made with redesigned and improved protocols that covered all three initial objectives (objectives 1-3). Using indoor flow-through aquaria facilities with cameras mounted from side view, we tested cross-species and within-species effects of sex pheromones on fish reproduction. Our specific objective was to determine responses of mating males and females to chemical signals released by males of invasive P. parva. We conducted experiments in tanks containing pairs of females and males and artificial nests. During tests the fish were continuously exposed to conditioned water, i.e. water containing the active reproductive pheromone, obtained from reproductive conspecific males (treatment 1), heterospecific males (treatment 2) or to de-chlorinated water (control). Reproductive success recorded as the number and size of clutches, was recorded daily. Behavioural traits shown by males and females such as time spent on the nest and away by males, cleaning, tending, aggression have been identified and characterized. We found finally that male-specific sex pheromones, regardless of the species, have no cross-species impact on reproductive success of the fish studied. Preliminary results of behavioural data analyses indicated no substantial shift in behaviour that would have resulted in reduced reproductive outputs. Our results provide evidence that mating process of nest-guarding cyprinids is not dependent predominantly on olfactory cues as a singular mode of communication. As, so in waters with high visibility the invasive P. Parva, pheromones when introduced, seems to have a low potential to affect the reproduction of mating individuals of P. promelas through disruption of chemical communication. However, given that the species inhabits waters with high turbidity where the visibility is low, misinterpretation of heterospecific signals by naive receivers may have significant impact on native species, e.g. through arising fitness costs due to difficulties in finding a viable mate. That is why the phenomenon of pheromone pollution does require further investigation. The active components were successfully isolated from conditioned water derived from male P. parva using C-18 Solid Phase Extraction and High Pressure Liquid Chromatography. Work is currently undertaken in collaboration with Cardiff University to analyse the active HPLC fractions using both Nuclear Magnetic Resonance and Mass Spectrometry. After this the chemical structure of the fish pheromones will be characterized (currently analysed). In conclusion, due to the delay during the first stage of the fellowship the results will be published later than anticipated, nevertheless, SCENT has generated innovative knowledge on the issue of pheromone pollution across species caused by invasive fish, by using a multidisciplinary approach coupling an advanced experimental design and analytical techniques. As so, this proposal has made a highly valuable contribution to our understanding of ecological impact of alien species.
