Guestblog: Environmental DNA allows for the detection of cryptic seahorse species

I’m very proud to publish this guestblog by Georgia Nester. Georgia is a PhD-candidate at Curtin University, where she focuses on the use of environmental DNA on species that are otherwise hard to study. She has just published her first paper, which could be a game changer on how we detect and study seahorses and their relatives in the future.


Seahorses (members of the Syngnathidae family) have never been detected using environmental DNA (eDNA), despite the fact that globally there are 14 species classified as “Threatened” by the IUCN. We compared the ability to detect a wide range of fish including Syngnathidae of two existing fish metabarcoding assays (= methods to detect eDNA and two new fish metabarcoding assays which we developed. With our new assays, we detected three Syngnathidae species in eDNA survey of the Perth metropolitan area (Western Australia), while the existing assays did not detect any Syngnathidae. These detections include the seahorse species Hippocampus subelongatus and Hippocampus breviceps, which represents the first time a seahorse has been detected using eDNA.

 

H subelongatus

The West Australian Seahorse (Hippocampus subelongatus). Photo credit: Maarten De Brauwer

With increasing human pressures and climate change resulting in a continuous decline of global biodiversity, there is a growing demand for rapid and sensitive conservation and monitoring programs. Using traditional methods, accurate data on species presence/absence and distribution is often difficult to obtain in aquatic environments. Environmental DNA metabarcoding is an increasingly popular solution. eDNA metabarcoding is capable of revealing what species are present in an environment by detecting traces of DNA they leave behind in the environment (e.g. shed skin cells, scales, blood, faeces etc). While eDNA metabarcoding surveys have been applied to a wide range of aquatic environments, no one has reported the detection of a seahorse to the best of our knowledge.

Many Syngnathidae species are considered threatened, however many more species (over 30%) lack the data necessary to assess their extinction risk. With the risk of a ‘silent extinction’ for many Syngnathidae species, the design of a non-invasive method for monitoring and managing these cryptic species may be critical to their survival. False negatives (failure to detect a species when they are in fact present) are significant in conservation management. For this reason, we aimed to determine if the Syngnathidae family (seahorses, seadragons and pipefish) were being inadvertently missed in current eDNA surveys.

H breviceps (1)

Camouflaged species such as the shorthead seahorse (Hippocampus breviceps) can be hard to detect with the naked eye. Photo credit: Maarten De Brauwer

Australia is home to 128 species of Syngnathidae in 40 genera, 65 of which are found in Western Australian waters. The Perth metropolitan area in Western Australia was chosen as our study site as it encompasses several habitat types, including brackish and salt water. We sampled from five locations across the Perth metropolitan area and processed the samples back at the TrEnD Laboratory in Curtin University. The results of this study have recently been published in the scientific journal environmental DNA.

In total, we detected four species of Syngnathidae using our newly developed metabarcoding assays “16S_FishSyn_Short” and “16S_FishSyn_Long”. The Syngnathidae species we detected were the Western Australian seahorse (Hippocampus subelongatus), the shorthead seahorse (Hippocampus breviceps), the spotted pipefish (Stigmatopora argus) and the tiger pipefish (Filicampus tigris). With Syngnathidae populations declining due to exploitation for the aquarium trade and habitat degradation, we have shown that eDNA methodologies are capable of detecting Syngnathidae taxa in the environment. This will help inform conservation and management strategies by providing a much-needed non-invasive method for monitoring these populations. Importantly, our study represents the first time a seahorse species has been detected using eDNA methodologies.

H subelongatus_Dave

The Western Australia seahorse (Hippocampus subelongatus, one of the first seahorse species to be detected with eDNA. Photo credit: David Harasti

New publication: Critter diversity on the sand

It turns out that moving halfway across the world and diving into a new job is more time consuming than I expected, so I haven’t been keeping up with the blog recently. I’m slowly starting to get more organised and in the coming weeks I will try to catch up with summaries of papers that I’ve published recently.

The paper “High diversity, but low abundance of cryptobenthic fishes on soft sediment habitats in Southeast Asia” was published almost a year ago in the journal Estuarine, Coastal and Shelf Science. It was one of the key papers of my PhD and describes the diversity of critters on sandy habitats in Indonesia and the Philippines.


If you have ever been muck diving it won’t come as a surprise to you that there is some very exciting marine life to be found on sandy bottoms. When you mention places like Lembeh Strait, Anilao, or Dauin to keen divers – especially photographers – they either get lyrical about the amazing pictures they took there, or will tell you about their plans for visiting any of the above places to go see some crazy marine life.

Froggie yawning

Species like this painted frogfish (Antennarius pictus) are popular with muck divers

In fact, the popularity of these sandy critters is so great, that the divers visiting Southeast Asia for muck diving bring in more than $150 million of revenue each year, supporting thousands of sustainable jobs! With so much money and jobs involved, it would be normal to expect researchers and conservationists to be interested in knowing which animals live on tropical sandy slopes. Unfortunately, that assumption would be wrong, surprisingly little is known about soft sediment (=sandy) habitats in the tropics. Even basic knowledge such as which animals live where is often unknown.

Luckily things are changing! Scientific interest in “cryptobenthic species” – the small, camouflaged critters this site is all about – is definitely increasing, with excellent work being doing on coral reefs by colleagues from across the world. We are starting to understand just how important they are for coral reefs and how very diverse cryptobenthic species can be.

What I am interested in though, is what is going on with the critters that live away from reefs. Are the critters living on the sand as diverse as those one coral reefs? Which species are most common? What causes species to live in one area, but not another? To answer these questions I set of with my good friend Luke for a 3 month dive survey trip that took us to Lembeh Strait, the north coast of Bali, and the sandy slopes of Dauin.

Maarten Smile

Surveying soft sediment critters in Dauin. Photo: Luke Gordon

During our survey dives, we not only counted and identified the fish we saw, we also measured a bunch of other factors that could have an effect on the presence of critters. We wanted to know whether depth, benthic cover (growth of algae, coral, sponges etc), or the characteristics of the sediment played a role in which species we found.

So what did we find?

One of the most interesting results is that the diversity (number of species) of cryptobenthic species was very high, higher in fact than the cryptobenthic fish diversity on many coral reefs! In contrast, the abundance (number of individuals) was much, much lower than what is found on coral reefs. To put it in perspective, if a normal coral reef would be an aquarium with 300 cryptobenthic fish of 15 different species crammed inside, soft sediment habitats would be the same aquarium with 30 fish of 16 species.

When looking at environmental factors, one of the most important factors that influenced where species lived, was the characteristics of the sediment. For small critters it makes a big difference whether the sand is powdery fine, or coarse like gravel. There seems to be a middle ground where the size of the sediment seems ideal for many critters. The tricky part is that the characteristics of sediment are in a large part determined by other processes such as currents or wave action. For now it is too early to conclude whether critters are found in these places because of the type of sediment or because of other factors that shape the sediment!

The amount of growth on the bottom played a role as well, particularly when algae or sponges were present, which makes sense as it offers variation in the habitat and potential hiding places for some species. Depth differences played a minor role in some regions (Daiun, Bali), but did not make a real difference in Lembeh. The limited effect of depth could partially be due to the fact that we did not survey deeper than 16m (university diving regulations are quite restrictive). It would be a great follow-up study to compare with deeper depths, as I am sure they will give very different results.

What does it all mean?

This study was (as far as I know) the first one ever to investigate the cryptobenthic fish in soft sediment habitats. The unexpectedly high diversity and very low abundance means there is a lot more  species out there than what was assumed, but that we have to look much harder to find them. I mostly see our results as a starting point to guide further research. We have only uncovered a fraction of what is out there and are not even close to really understanding how tropical soft sediment systems function. While this provides an exciting opportunity for scientists like me to new research, it also means that we do not yet know how environmental threats such climate change or overfishing will impact  species living on soft sediment. We do not know yet if the species that muck dive tourism depends on need protection, or how to best protect them if they do.

Whiteface whaspfish

We might not know yet what the future holds for sand-specialists like this whiteface waspfish (Richardsonichthys leaucogaster), but I am hoping to find out!

What is a species?

As promised in a previous blogpost, it is time to get into another hotly debated topic in biology that most non-biologists wouldn’t even think was an issue at all. This is a big one, as it underpins pretty much all biology: “What is a species?”. I would argue that this question needs two important additional questions: “Why does it matter?” and “To whom does it matter?”. Since I am only human and like postponing difficult tasks at hand, let’s start with the follow-up questions.

speciesTo whom does it matter that we are capable of telling one species from another? Besides looking like a smart cookie when telling your fellow divers/birdwatchers/plant enthusiasts which species you’ve just seen, it doesn’t matter very much to be honest. We are no longer hunter-gatherers, so being able to tell which species you can eat, and which ones will will eat/kill you, doesn’t matter that much anymore. In other words, the discussion in the rest of this blog is mostly an issue for taxonomists, but it gives an interesting insight in how simple concepts can be quite complicated when you look closer.

Why does it matter then? For two reasons:

  1. People in general and scientists in particular like putting labels on objects around them, it helps us structure and understand the world we see.
  2. Being able to tell two species apart that look very similar can have big consequences for conservation action: Two birds/fish/plant might look similar, but they could be different species, one of which reasonably common, but the other one rare and on the brink of going extinct. If we don’t realise they are different, we might lose that species.

A logical follow-up question (in my mind) would be: “what does it matter if one species that looks a lot like the next goes extinct?”. This is a valid question, but would require a long (and interesting) scientific and philosophical discussion.

charlesdarwin1

Struggling to define what a species is? Don’t worry, so did Charles Darwin (photo source: www.brainpickings.org)

Back to the species concept, which seems obvious, but really is not. The question has been asked for centuries by many renowned scientists. Two of which were none other than Charles Darwin and Alfred Russel Wallace. It could even be claimed that this question is what eventually led to the theory of evolution, which essentially tries to explain how different species come into existence. It is obvious that you need to know what a species is before you can answer that question. Sometimes it is easy: the majority of people can tell the difference between a cat and a dog, but it becomes a lot more difficult when species look very similar. Try asking a marine biologist what the difference is between a Slingjaw wrasse (Epibulus insidiator) and a Latent slingjaw wrasse (Epibulus brevis), or between a Thin Ghostpipefish (Solenostomus leptosoma) and a Robust Ghostpipefish (Solenostomus cyanopterus). Go on, give it a go, it’ll be fun to see them struggle! (Smart-ass tip: a spot on the dorsal fin & smaller adult size / No difference, they are most likely the same species).

chi_dane

They might look different, but are the same species. Photo source: www.dogguide.net

The problem is how do you define a species? At what point is an individual that looks different than “the norm” a different species, rather than just natural variation? Dogs can look as different as Danish dogs and (rats) Chihuahuas but are all the same species. Compared to those two, lions and tigers look much more alike, yet they are very different species. The classic definition of a species (if they cannot have fertile offspring they are different species) works in most cases. Lion + Tiger = Liger, but ligers are infertile. I don’t know what Danish dog + Chihuahua would look like and I wouldn’t get ethics approval from my university to test it, but presumably the result would be a fertile dog.

So far the normal situation, but what happens when different species mate and have fertile offspring (hybrids)? This is surprisingly common in the ocean. Hybridising fish are not that rare if you know what to look for. I have personally seen it in Clownfish and Surgeonfish and almost certainly in Frogfish and Ghostpipefish. But is has also been recorded in groupers, manta rays, butterflyfish, angelfish and wrasses.

hybrid_surgeonfish

Hybridising surgeonfish: r: Acanthurus lineatus s: Acanthurus sohal t: A. lineatus x A. Sohal hybrid (Source)

In this case, can we say the parents are different species because they look different, behave differently, and typically live in different regions? Or are they a single, highly variable species, the way dogs are? Traditional taxonomy focused on what species looked like, would say they are different species, but not everyone agrees with this. A recent example from the pygmy seahorse world: Hippocampus severnsi and Hippocampus pontohi were described as two different species. However, new (yet unpublished) research shows that they are genetically identical, so the name H. severnsi was removed and they are now all called H. pontohi. I’d imagine much to the annoyance of Mike Severns, who no longer has a cool animal with his name on it.

Is genetics the solution to the problem? Depends on who you ask. Geneticists tend to say yes, old-school taxonomists tend to be a bit less convinced. Each side has very valid arguments, one of which is how genetically different do individuals have to be before they are considered different species (sound familiar?). Different cut-offs have been proposed, but as far as I know, there is no real consensus (please correct me if I am wrong geneticist-readers). Another serious issue is how to classify small life forms like microbes, this article has a great summary on that if you are interested.

If you think this is getting too complicated, it might be best not to become an evolutionary biologist or taxonomist, because things actually get a lot more complicated than what I described. Suffice to say, for biologists the term “species” is still not a clearly defined concept. But luckily for non-specialists, the essence of the debate is about such fine details that it really shouldn’t be keep you up at night.