Swimming Against the Flow: Environmental DNA Can Detect Bull Sharks (\u3ci\u3eCarcharhinus leucas\u3c/i\u3e) Across a Dynamic Deltaic Interface

© 2020 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd Human activities in coastal areas are accelerating ecosystem changes at an unprecedented pace, resulting in habitat loss, hydrological modifications, and predatory species declines. Understanding how these changes potentially cascade across marine and freshwater ecosystems requires knowing how mobile euryhaline species link these seemingly disparate systems. As upper trophic level predators, bull sharks (Carcharhinus leucas) play a crucial role in marine and freshwater ecosystem health. Telemetry studies in Mobile Bay, Alabama, suggest that bull sharks extensively use the northern portions of the bay, an estuarine–freshwater interface known as the Mobile-Tensaw Delta. To assess whether bull sharks use freshwater habitats in this region, environmental DNA surveys were conducted during the dry summer and wet winter seasons in 2018. In each season, 5 × 1 L water samples were collected at each of 21 sites: five sites in Mobile Bay, six sites in the Mobile-Tensaw Delta, and ten sites throughout the Mobile-Tombigbee and Tensaw-Alabama Rivers. Water samples were vacuum-filtered, DNA extractions were performed on the particulate, and DNA extracts were analyzed with Droplet Digital™ Polymerase Chain Reaction using species-specific primers and an internal probe to amplify a 237-base pair fragment of the mitochondrial NADH dehydrogenase subunit 2 gene in bull sharks. One water sample collected during the summer in the Alabama River met the criteria for a positive detection, thereby confirming the presence of bull shark DNA. While preliminary, this finding suggests that bull sharks use less-urbanized, riverine habitats up to 120 km upriver during Alabama\u27s dry summer season

Development of Highly Sensitive Environmental DNA Methods for the Detection of Bull Sharks, \u3ci\u3eCarcharhinus leucas\u3c/i\u3e (Müller and Henle, 1839), Using Droplet Digital\u3csup\u3eTM\u3c/sup\u3e PCR

Background: As apex and mesopredators, elasmobranchs play a crucial role in maintaining ecosystem function and balance in marine systems. Elasmobranch populations worldwide are in decline as a result of exploitation via direct and indirect fisheries mortalities and habitat degradation; however, a lack of information on distribution, abundance, and population biology for most species hinders their effective management. Environmental DNA analysis has emerged as a cost‐effective and non‐invasive technique to fill some of these data gaps, but often requires the development of species‐specific methodologies. Aims: Here, we established eDNA methodology appropriate for targeted species detections of Bull Sharks, Carcharhinus leucas, in estuarine waters in the northern Gulf of Mexico. Materials and Methods: We compared different QIAGEN®DNeasy® extraction kit protocols and developed a species‐specific Droplet Digital™ PCR (ddPCR) assay by designing primers and an internal probe to amplify a 237 base pair portion of the ND2 gene in the mitochondrial genome of C. leucas. To validate the developed methods, water samples were collected from known C. leucas habitat and from an ex situ closed environment containing a single C. leucas individual. The effectiveness of the assay in an open environment was then assessed by placing one C. leucas into a flow‐through mesocosm system and water samples were collected every 30 min for 3 hr. Results: The developed C. leucas ‐specific assay has the ability to detect target DNA concentrations in a reaction as low as 0.6 copies/μl. DdPCR reactions performed on water samples from known habitat and 30 min after a shark was added to the closed environment contained 1.62 copies/μl and 166.6 copies/μl of target C. leucas eDNA, respectively. Carcharhinus leucas eDNA was detected in the flow‐through system within 30 min, but concentrations remained low and variable throughout the duration of the experiment

An Environmental DNA Tool For Monitoring the Status of the Critically Endangered Smalltooth Sawfish, \u3ci\u3ePristis pectinata\u3c/i\u3e, In the Western Atlantic

The Critically Endangered Smalltooth Sawfish, Pristis pectinata, was once widespread in the tropical and subtropical waters of the Atlantic Ocean, but following substantial declines over the past century, the remaining population(s) are currently confined to Florida in the U.S., and the Bahamas. Recent research and verified public encounter reports suggest that the core population in south and southwest Florida may be stabilizing and potentially expanding into formerly occupied areas of their historic range in the western Atlantic; however, the status of this species outside of core waters is not well understood. Environmental DNA (eDNA) methods provide a relatively cost effective and rapid assessment tool for monitoring species occurrence in aquatic habitats. Here, we have developed an eDNA tool: a species-specific Droplet Digital™ PCR assay targeting a 100-base pair portion of the mitochondrial NADH dehydrogenase subunit 2 gene in P. pectinata, with the ability to reliably detect as little as 0.25 pg of target DNA. The assay was validated by analyzing a water sample from an occupied nursery in southwest Florida, which was found to contain an average of 11.54 copies of target DNA/µL (SE = 0.72) in the reaction. The assay was then further tested by placing a juvenile sawfish in an ex situ tank and analyzing water samples collected at time intervals. The implementation of this eDNA tool into field surveys will provide additional, reliable data to assess species recovery and aid in prioritizing localities in which to focus new research, conservation, and education initiatives