This is a guest post by Cecilia di Bernardi.

Image Credit: Rick Heeres, CC BY 2.0, Image Cropped

Multiple species-specific molecular markers using nanofluidic array as a tool to detect prey DNA from carnivore scats (2021) Di Bernardi et al., Ecology & Evolution. https://doi.org/10.1002/ece3.7918

The Crux

Studying carnivore diet can be a crucial tool to inform both management and conservation of predators and their prey. If we’re going to ensure a carnivore’s survival, we need to know which species it relies on for food, and in what quantities.

Digging into an animal’s stomach isn’t the nicest way to get the crucial data we’re looking for, so non-invasive sampling of scats (that’s science for poop) has for been a more ideal approach to collecting valuable information on the occurrence, genetics, and diet of animals, especially when dealing with elusive and threatened species. Nowadays, DNA-based analyses of scats are allowing researchers to get more and more high-resolution data on predators’ food habits.

What We Did

We developed a DNA-based method to detect prey from wolf scats, taking advantage of the huge leaps the DNA analysis has been through in recent years. We also made use of nanotechnology (specifically Nanofluidic array technology fromFluidigm Inc.), which has been useful for detecting pathogen species in ticks, or traces of herbivores on browsed twigs, but has never applied to detect prey from predator scats!

Starting from the big bank of DNA sequences available online (GenBank, NCBI), we looked at specific areas of the genome, (the mitochondrial genome), in order to tell apart the different target prey species present in the wolf scat. We developed species-specific molecular markers (see Did You Know?) and tested them with reference tissue samples, kindly provided by the Swedish Museum of Natural History. After the protocol development and optimization, we ended up with a set of 80 markers for our 18 target species. We then applied the newly developed molecular method on a pilot sample of wolf scats collected in the field.

Did You Know: Molecular Markers

Since any species’ genome is an incredibly long sequence, scientists have developed more efficient ways of defining what DNA belongs to which species. The motivation is simple – if you’re trying to tell whether a genome belongs to a human or to a chimpanzee, you don’t want to be looking through the 99% of DNA we have in common, you want to go straight to that 1%. That’s why scientists develop ‘markers’. It helps them narrow down their search and identify species much more quickly.

What We Found

The molecular markers we developed did their job well, correctly detecting the 18 prey species, showing an overall good distinction between the tissue samples of the target and non-target species. In other words, this means that a tissue sample taken from a moose was detected by the moose markers but not by the reindeer markers, which is the sign of a successful marker!

When applied to the pilot of wolf scats collected in the wild, the method detected a total of 16 species, comprising wild ungulates (moose, roe deer, red deer, fallow deer, wild boar), domestic and semi-domestic animals (reindeer, cattle, sheep), small prey species (European badger, European hare, mountain hare, Western capercaillie, black grouse), and other carnivores (Eurasian lynx, wolverine, red fox).

Problems?

While the method detects the target species as we’d like, it cannot distinguish whether predation, scavenging, or territorial marking has occurred. Detection of fox DNA in wolf scats can mean a wolf predating on a fox, a wolf scavenging on a fox, but also a fox marking with its urine on a wolf scat! To partly disentangle this aspect, we are investigating the contribution of scavenging to wolves diet in Scandinavia, with data from GPS-collared wolves.

So What?

This molecular method, with its high-resolution prey detection, can help better understanding under what circumstances wolves eat certain prey and how that can affect ungulate populations, serving as a valuable complement to the current GPS technology used to investigate wolf predation. Wolf natural expansion is an ongoing and controversial phenomena in the Northern hemisphere, and any technique that tells us more about their impact is a welcome addition to our knowledge base.

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Cecilia Di Bernardi is an ecologist who is currently investigating wolf predation ecology within her PhD at the University of Rome La Sapienza in collaboration with SLU Swedish University of Agricultural Sciences. You can follow her on Twitter @c_dibernardi.