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Methodology

Have any sharks? Play GoFish and find out!

Journal source: Stoeckle MY, Das Mishu M, Charlop-Powers Z (2018) GoFish: A versatile nested PCR strategy for environmental DNA assays for marine vertebrates. PLoS ONE 13(12): e0198717. https://doi.org/10.1371/journal.pone.0198717

Introduction

Understanding and protecting marine ecosystems requires knowledge about the various species living in a habitat. It is also important to be able to understand how organism communities change over time, because changes in species composition, or the loss of one or more species, can be interpreted as a red flag that something is amiss. To accomplish this, scientists carry out long-term monitoring in various marine ecosystems. Historically, monitoring of species involved rather intrusive approaches, for example, collecting organisms with fish nets and bottom trawls, and counting how many types of species are present. However, such methods often result in the death of any collected creatures, which can thwart efforts to conserve those species. Isn’t there a better way to monitor marine communities without adding additional stress and without sacrificing accuracy of species identification?

Fish, mammals, and invertebrates that swim in the open oceans and coastal marine regions grow, reproduce, and slough off old cells into the surrounding water, leaving behind evidence of their presence in the form of environmental DNA (eDNA for short). eDNA is based upon the idea that all organisms in their environment slough off DNA, which stays in the water for a limited time, and scientists who want to survey what species are present in a habitat can simply collect a water sample, filter it, extract eDNA, and sequence that eDNA. Researchers then use Polymerase Chain Reaction (PCR), in which primers, short single-strands of DNA that target a certain region (gene) of the DNA, produce many copies of the desired region.

To address the limitations of current eDNA methods, Stoeckle and colleagues from The Rockefeller University in New York designed an eDNA protocol called “GoFish”, which uses nested PCR, in which a first round of PCR uses general primers to amplify a common gene region shared by many groups of animals. This first round PCR casts a “wide net” so to speak, to capture as many organism DNAs as possible from the initial sample. The amplified DNA from round one is amplified a second time with primers that target individual species. They tested their approach on weekly water samples taken from the lower Hudson River estuary and southern New York Bight during the summer of 2017.

Diagram of nested PCR workflow used in this study. Figure source: Stoeckle et al. 2018 modified Figure 1.

How well did their approach work?

The newly designed GoFish protocol was able to detect over a dozen fish species, including cunner, oyster toadfish, and seaboard goby, and a mammal, the bottlenose dolphin, which are common in the waters of the Hudson River estuary. They also detected three shark species, two skates, and four rays, and the time at which these were detected was consistent with migration patterns of these organisms. Additionally, GoFish correctly identified fish species that were expected to be found in certain habitat types within the study site. For example, GoFish detected more rocky bottom specialists (such as cunner) in the lower Hudson River, which is dominated by rocky habitat, than in outer New York harbor, which was mostly sandy bottom.

A Cownose Ray, a species that GoFish identified in the samples. Photo credit: cliff1066 via Creative Commons.

Cunner, another fish species identified with the GoFish protocol. Photo credit: Derek Keats via Creative Commons.

(Featured image) Map of study area and graphic showing the months in which organisms were identified by GoFish. Photo credit: Figure 3 from Stoeckle et al. 2018, via Creative Commons License.

The bigger picture

GoFish is a potential tool to be used alongside eDNA profiling of marine communities in which scientists are interested in the presence or absence of certain fish and mammal species. A major benefit of GoFish is the rapid turn-around time from sample collection to sequencing and analyzing those sequences. This would be useful for long-term monitoring studies that take samples repeatedly from one or multiple locations, allowing scientists to ID species in an area without catching or killing any marine creatures. In addition, because only water samples are taken, this approach could be useful for studies that are tracking populations of species of concern and endangered species.

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