Tom_marking-1
Photos provided by Dr. Thomas Hossie

When it comes to conservation biology, Trent University has the advantage of housing a few of the Ministry of Natural Resources (MNR)’s research labs. While that gives us a one-up on other universities, Dr. Craig Brunetti, Chair of the Biology Department, said that “private funding is much more variable.” A relatively new study led by Dr. Dennis Murray has just received funding from the Species At Risk Stewardship Fund (SARSF) to look into the population of small-mouthed salamanders on Pelee Island, a small island in Ontario on Lake Erie. Dr. Thomas Hossie, a post-doctoral  fellow at Trent and one of the researchers in the salamander project, talked to Arthur about what they hope to achieve over the next few years with their recent funding and what challenges they will face.

Tell me more about the funding from SARSF and what it means for Trent’s biology department?
The SARSF was created under the Endangered Species Act to encourage people to get involved in protecting and recovering [at-risk] species through stewardship activities. They are one of many grants and incentives available to help protect and recover plants and animals that are in danger of disappearing in Ontario.Trent has decades of research expertise in environmental research including in the areas of environmental modelling, genetics and molecular biology, and wildlife conservation. As such, this project is a natural fit for our institution.

This SARSF grant provides new opportunities for us to contribute to the protection of endangered wildlife, while simultaneously investigating ecological questions, and training our students.

For more information on SARSF, please visit:http://www.ontario.ca/document/species-risk-stewardship-fund-application-guidelines

Does Trent get funding like this often?
Other research groups at Trent have probably received funding through the SARSF program before, but this is the first time that our research group (Integrative Wildlife Conservation) has received funding of this kind.

What will you be doing with the money?
In Canada, small-mouthed salamanders occur only on Pelee Island, where they reach their northern range limit. The recent loss of some identified breeding ponds combined with the lack of data on this population has resulted in this species being listed as “Endangered” under the Species At Risk Act (SARA). Our multi-year project will fill critical knowledge gaps about the small-mouthed salamander.

Specifically, we will provide the first population size estimates for the small-mouthed salamander population on Pelee Island, study the genetic structure and composition of these salamanders, and characterize the aquatic and terrestrial habitat used by small-mouthed salamanders. We also seek to develop a genetic mark-recapture technique for “pure” small-mouthed salamanders.

How do the techniques and research methods for the salamander project compare to other population studies done either by Trent or population researchers in general?

Generating an estimate of population size for a natural population of mobile animals typically relies on a mark-recapture program, and this is the case with us as well. Briefly, mark-recapture involves collecting a subset of individuals from a population, marking these individuals, releasing them back into the wild, then returning and re-sampling the population. Your re-sample is made up of marked and unmarked individuals and this ratio can be used to estimate the population size.

Marking amphibians however poses a set of unique challenges. With turtles, you can notch their shells, paint on an alphanumeric code, or affix an external tag. With mammals, you can affix an ear-tag or a collar, and with birds you can add a ring or band around their leg. For frogs and salamanders, though, most of these techniques don’t really work. Some researchers have used toe-clipping (removing certain combinations of toes from the fore and hind legs) to individually mark frogs or toads, but even this fares poorly for salamanders because of their incredible ability to quickly regenerate and repair damaged tissue (i.e., the salamanders re-grow their toes before researchers are able to come back and re-sample the population!).

Pattern recognition has proven effective for species with distinctive dorsal or ventral [back or stomach] colour patterns (e.g., Fowler’s Toads), but many small-mouth salamanders lack conspicuous and highly distinctive patterns.
More recently, biologists have started to use visible implant elastomer (VIE), a flexible and brightly coloured material that is injected just below the skin. We give each individual four small injections of VIE and the various combinations of colour and location of these marks allows us to individually mark hundreds of salamanders.

These VIE tags fluoresce under UV light and remain visible to the researcher for the lifetime of the animal. Combining these unique marks with our genetic samples means that we can track their habitat use and never have to mark or take a tissue sample from those individuals ever again! This is great because these salamanders can live over 13 years!

What is meant in the press release when you say “state-of-the-art sampling and analytical methods?”
Our population estimate will be generated using the same general mark-recapture approach outlined above, but takes advantage of more sophisticated analytical techniques which can account for systematic differences in our ability to capture certain individuals (e.g., different genotypes, age-classes, sex), any environmental conditions that affect catch-ability, as well as any behavioural changes that result from having been captured (i.e., individuals may be easier or harder to find after having been captured once).

Are you collecting DNA from the population for analysis and how is that being done?
We collect DNA in two ways. First we collect a “swab sample” using a cytology brush to collect skin cells and second, we taking a small tissue sample from the tip of the salamander’s tail. The tail tip tissue provides us with a genetic sample that we can use to reliably distinguish unisexuals and small-mouthed salamanders.

We hope that we can cross-validate this with our swab samples and re-identify individual small-mouthed salamanders using their unique microsatellite [a genetic marker] signatures. If this is possible, then we could track the survival and habitat use (e.g. what ponds they breed in) of individual salamanders without having to mark them, or take tail tip tissue samples. Using the genetic information we can sort out which salamanders are unisexuals and which are small-mouthed salamanders.

Importantly, once the genetics have been run, we then also know the true identity of each recaptured salamander!

 

Salamander side-by-side

 

Could you explain the concept of the unisexual salamanders and why they are difficult to distinguish from pure small-mouthed salamanders?
Within the Great Lakes region, the distribution of blue-spotted salamander (Ambystoma laterale) and the small-mouthed salamanders (A. texanum) overlap, and in these areas of sympatry [two species living in the same geographic area], we also find “unisexual” Ambystoma salamanders that possess genetic material from multiple species. These all-female unisexuals differ from conventional hybrids in that they require sperm from a sympatric male to initiate egg development, but rarely incorporate the male’s DNA into that of her offspring (i.e., kleptogenesis).

Analysis of their mitochondrial DNA, however, has revealed that unisexuals are not a recent by-product of human-induced habitat change, but instead appear to have been evolving independently alongside other Ambystoma for more than three to five million years. Unisexual salamanders can have anywhere from two to five sets of chromosomes.

On Pelee Island, all unisexuals have at least one set of chromosomes from each the blue-spotted and small-mouthed salamanders. In fact, Pelee Island is the largest of chain of islands in southwestern Lake Erie and retains populations of A. laterale, A. texanum, and at least six unisexual genotypes with nuclear DNA from both species. In terms of their external appearance, it is not possible to distinguish small-mouthed salamanders from unisexuals. We are, however, taking photos and morphological data when we process the salamanders.

Another island in Lake Erie called Kelley’s Island has unisexuals that could have DNA from as many as three species! Some people may be familiar with the unisexuals in mainland Canada that have DNA from both Jefferson’s salamanders and Blue-spotted salamanders.

What will you be doing with the project going forward once you get your results on the increase or decrease of the population of salamanders?

Amphibian populations receive a surprisingly low amount of monitoring, especially given the fact that they are among the most threatened groups on the planet. Assessing whether a given amphibian population is in decline is made even more difficult by the typically high amount of year-to-year variation in population size.

My hope is that once we establish this mark-recapture program, we can implement some form of baseline population motioning to track further changes in the population over the longer-term.

Because Pelee Island is located both at the southern range limit for A. laterale and the northern range limit for A. texanum, monitoring these populations in particular could provide key insights into the effect of climate change on amphibian populations. Brunetti shared with Arthur that the Biology Department at Trent usually receives a tri-council funding from the Natural Science and Engineering Research Council (NSERC), Canadian Institutes of Health Research (CIHR), and the Social Science and Humanities Research Council (SSHRC).

There are also many other conservation research projects being conducted by Trent faculty on an ongoing basis. “Maggie Xenopoulos, Paul Frost, and Chris Metcalfe (ERS) have projects at the Experimental Lakes Area. Paul Wilson has a number of collaborations on caribou conservation,” said Brunetti.

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