Detecting Endangered Organisms

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Detecting Endangered Organisms

#1  Postby Calilasseia » Oct 12, 2017 2:53 pm

This is a big problem faced by many invertebrate zoologists - you know that an endangered species has been found at a given site in the past, but the only way to confirm that it's still there, is to go out to the site, capture specimens, and take them back to the laboratory for dissection.

Invertebrate zoologists run into this problem more frequently than they would like. Not least, because many of the groups of organisms they work with, are only reliably identifiable to species level via dissection.

A case in point is provided by an article in Phys.org. which centres upon an amphipod. The amphipod in question is Stygobromus hayi, the Hay's Spring Amphipod. Which is endemic to one location in Rock Creek Park in Washington D.C. This organism is on the endangered list, courtesy of its extremely restricted distribution and specialised niche. Matthew Niemiller, author of the latest paper on this species, explains thus:

The Hay's Spring amphipod lives in swampy areas called seepage springs where groundwater sometimes spills out onto the surface of Rock Creek Park, in Washington, D.C., its only known home in the world. Because it is so rare, and perhaps also because of its prestigious ZIP code and proximity to the National Zoo, this tiny crustacean is on the U.S. endangered species list.

"Yes, it's small, it's white, it's eyeless, it lives underground," said Matthew Niemiller, an ecologist with the Illinois Natural History Survey who led a recent study to search for the creature in its Rock Creek Park home. "It's not a cute, cuddly or charismatic species. But we're still learning more and more about groundwater ecosystems. And there is evidence that these crustaceans are important indicators of groundwater quality, and may play important roles in water purification and nutrient cycling over time."

The Hay's Spring amphipod's subterranean habitat makes it particularly difficult to study, Niemiller said.

"To find out where species are located and how many there are, we have to disturb the site," he said.

This normally involves sifting through waterlogged leaves in muddy seepage areas, where underground springs leak out to the surface.

"And because these critters are small - no more than a centimeter long - and a couple of other species look very similar to S. hayi, we would have to sacrifice any individuals we find and take them back to the lab to identify them," he said. "Since this is an endangered species, that's something we don't want to do."


I'll dispense quickly with the matter that the authors of the Phys.org article chose to unleash some humour when choosing the title thereof. Enjoy the dig at certain prominent individuals briefly, before moving on to the matter of the amphipod, and the tricky business of detecting whether it is there, without resorting to the usual methods.

Niemiller decided to try a different method, not least to determine if it actually worked.

That method is known as 'environmental DNA sampling'. Environmental DNA, or eDNA for short, consists of DNA strands that have been shed into the surroundings by various organisms. Every living organism jettisons at least some DNA during its life, by various means, and in the case of larger multicellular organisms, this fact has provided a useful tool for the detection of species in locations presenting various logistical difficulties. For example, the technique has already been tested - and found to work - with salamanders. But would the technique work with an amphipod just 1 cm long, and whose numbers in the site to be surveyed were uncertain to put it mildly?

There was, of course, only one way to find out - the classic empirical approach. Go forth and try it out.

To give an idea of the difficulties to be faced, there were several unanswered questions. Would a 1 cm amphipod, even if present in numbers, shed enough DNA into the environment to be detectable in an unequivocal manner? Would the special conditions of its environment - a seepage spring whose water contains high concentrations of humic acids - have a rapid degrading effect upon any DNA the amphipods did provide? Would it be possible to separate the DNA of the target species, from that of other, related species known to share the same habitat? Alternatively, does the DNA in question persist long beyond the death of its originator, adding to the difficulties of using it for auditing living organisms?

Other issues to be addressed, included the fact that the humic acids present in the groundwater being sampled, acted as inhibitors of the enzymes used for PCR amplification in the laboratory, so a means of dealing with these, in a manner not affecting any collected DNA, had to be devised as well.

After what sounds to the uninitiated, to be an inordinately huge effort on the part of a 1 cm amphipod, Niemiller had his results. Not only was he successful in detecting his target organism (and its relatives), but he found that it was present in a location where it had not been previously recorded, within the same park.

Though, Niemiller admits, it's early days for this technique. At the moment, it simply provides a binary yes-no answer, whether or not your target species is present or absent. It doesn't yet provide data on numbers of individuals present, which of course is a vital piece of data for conservation work. But, it's a step forward - a non-invasive technique has now been expanded to include aquatic invertebrates within its remit. Which could become even more valuable, if someone solves the problem of extending it to terrestrial invertebrates.

The paper in question is this one:

Evaluation Of eDNA For Groundwater Invertebrate Detection And Monitoring: A Case Study With Endangered Stygobromus (Amphipoda: Crangonyctidae) by Matthew L. Niemiller, Megan L. Porter, Jenna Keany, Heather Gilbert, Daniel W. Fong, David C. Culver, Christopher S. Hobson, K. Denise Kendall, Mark A. Davis & Steven J. Taylor, Conservation Genetics Resources, DOI: 10.1007/s12686-017-0785-2

Niemiller et al, 2017 wrote:Abstract

Effective conservation and management of biodiversity is limited by a lack of critical knowledge on species’ distributions and abundances. This problem is particularly exacerbated for species living in habitats that are exceptionally difficult to access or survey, such as groundwater habitats. Environmental DNA (eDNA) represents a rapid, noninvasive, and potentially cost-effective new tool for detection and monitoring of biodiversity that occur in such habitats. In this study, we investigated the utility of eDNA in detecting the federally endangered Hay’s Spring Amphipod Stygobromus hayi and a co-occurring common congener S. tenuis potomacus from unique groundwater-associated habitats—hypotelminorheic seepage springs—in the Washington, DC metro area. We developed taxon-specific primers and probes for each species to amplify Stygobromus DNA using qPCR. In silico and in vitro validation demonstrated specificity of each designed assay. Assays were then used to screen water samples collected from ten seepage springs. Stygobromus hayi was detected at four seepage springs, including one potential new locality, while S. t. potomacus was detected at four springs, two of which were new localities. This study is the first to our knowledge to successfully employ an eDNA approach to detect rare or threatened invertebrates from subterranean ecosystems. Our study highlights challenges of employing an eDNA approach for the detection and monitoring of invertebrates in groundwater habitats that are difficult to study, including accounting for PCR inhibition and the potential for cryptic genetic diversity.
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