The utility of eDNA as a tilapia surveillance tool

Background: Invasive fishes pose a major threat to aquatic ecosystems worldwide. Their impact can be severe in tropical regions, such as northern Australia, where over 20 invasive fish species are recorded from freshwater rivers and streams. The prevention of new incursions of invasive species is the goal, however when prevention fails, early detection of incursions is critical for successful control measures. To this end, environmental DNA (eDNA), the DNA that an organism leaves behind in the ecosystem, is proving to be a promising early detection tool for invasive aquatic species and has been used successfully to detect incursions of temperate invasive species. This research project aimed to modify conventional eDNA protocols developed by the University of Notre Dame (USA) for application in tropical environments to detect the invasive pest fish tilapia and to develop an understanding of the detection sensitivity of the method. Methods development: Broadly, the method involved collecting and filtering two litre water samples and testing the filtrate for the presence of tilapia eDNA with a species specific probe. Essential to implementation was the successful development of a species specific probe and selection of the appropriate filter types for turbid tropical environments. A tilapia specific probe was developed. Fine filters (from 3 to 10 μm) were inefficient in tropical environments, due to clogging from suspended particles and algae, but 20 μm filters were tested and proved successful for extraction of eDNA from water samples. The interpretation and application of eDNA surveillance methods required a knowledge of the rate of eDNA degradation and the sensitivity of the method. Thus, a set of controlled aquarium experiments were done that demonstrated eDNA persisted in the environment for between 15 and 29 days dependent on the temperature tested (23, 29 and 35°C). Further, to test sensitivity, low numbers of caged tilapia were introduced to large ponds (0.4 ML) to determine the probability of detection. The method proved effective and in experimental testing was demonstrated to successfully detect one fish/0.4 ML (after four days). Eureka Creek: Eureka Creek, in the Mitchell River catchment of the Gulf of Carpentaria, is an important site because it was the location of an incursion of tilapia identified in 2008. An eradication program was implemented to remove tilapia from the creek and prevent the spread into other Gulf of Carpentaria river catchments. Follow-up sampling using traditional methods suggested that the eradication program was successful but some concerns remained. Thus, as part of this project a new eDNA survey of Eureka Creek was conducted to follow-up the eradication program. Three locations surrounding the original infestation were sampled. At each location three 2 L water samples from three sites were collected and tested for the presence of tilapia eDNA. No tilapia eDNA was detected in any of these samples adding further weight to the evidence that tilapia were successfully eradicated from Eureka Creek. Comparison: To contrast the effectiveness of eDNA as a surveillance tool with the primary traditional sampling tool for tilapia, that of electrofishing, a survey was conducted using both methods at 14 sites, in the lower Fitzroy River catchment, where a recent invasion of tilapia was reported. Tilapia were detected by eDNA methods in eight of the 14 sites surveyed. Two sites had no detectable tilapia eDNA and the remaining four sites failed to meet quality control standards and results from these sites were not confirmed. Positive detections were obtained for the eDNA survey from all three sites where tilapia were detected using electrofishing and at a further five sites where electrofishing failed to detect tilapia. Environmental DNA surveillance has proven to be an effective early detection tool for tilapia incursions and likely to be more sensitive than traditional survey methods but requires considerable care and precision in its implementation due to the potential for sample contamination. Outputs: Environmental DNA technology was successfully adapted for the specific purpose of tilapia surveillance and this has resulted in a high quality service that will be beneficial to many organisations and associations to help early detection of tilapia incursions. Already, the uptake of eDNA as a method of tilapia surveillance and monitoring has occurred; the Fitzroy Basin Association and Catchment Solutions have employed the eDNA services developed from this project to survey the recent invasion of tilapia in central Queensland