Experimental evidence of dispersal of invasive cyprinid eggs inside migratory waterfowl - Behind the scenes

Recently, our team published results of an experiment, where we demonstrated that fertilised eggs of two invasive cyprinid fish survive passing through the digestive tract of waterfowl. And yes, this is no joke guys, these fish eggs were consumed by mallards (with a little help), some of them were pooped out intact and some hatched and grew like nothing happened to them. Here we want to present the work in pictures and share some details that could not be shared in the published work.

The authors

One might ask, where this crazy idea was coming from. In 2013-14 Ádám was visiting Andy Green in Spain within the framework of a scholarship and they worked on exploring what organisms are transported by waterbirds in the Doñana National Park. During one of the field trips Adam collected shorebird pellets on the field. These pellets are material that the bird can't digest, usually indigestible plant material, bones, hairs, invertebrate exoskeleton, etc. But in these pellets Adam discovered a couple fish eggs and when he looked closely, he could see the embryo moving inside them. Although fish eggs are easily digestible, they probably got mixed up with indigestible material and the bird regurgitated them. This observation illustrated that fish eggs could survive at least for some time inside the digestive system of birds, but it was unclear whether they can get out alive if they go through the alimentary canal all the way. Nonetheless, this observation made it clear that endozoochory (i.e. the internal transportation of organisms) could play a role in the dispersal of fish.

Doñana National Park, where the story begins…

The picture bellow shows these eggs recovered from the inspected shorebird pellets. These eggs were placed to hatch in freshwater and the embryos died shortly, which made us think that the eggs originated from saltwater fish.

The fish eggs found in field collected faecal samples

Several members of the team had experience with experiments testing the endozoochorous potential of plant seeds and we believed that a similar experimental setup could be suitable to test if fish eggs survive passing the digestive tract of birds. We started planning the experiment using almost identical protocol to our previous experiments. Ducks were purchased from a local breeder, so they were habituated to humans and cages, but they were basically wild-type mallards, unaffected by domestication. Prior to the experiment, ducks were housed in their individual cages for a period of 24 hours, in order to habituate them to this new environment. Cages were large enough for the ducks to move around, while food and water was provided ad libitum throughout.

Setting up the experiment

All the eggs were produced in Szarvas by the Research Institute for Fisheries and Aquaculture. Carp has phytophilic eggs, which means they attach their eggs to submerged plants. Therefore, the eggs have a protective, gelatinous, adhesive overlay. This layer is responsible to attach the egg on the submerged leaf surface and protect them against physical impacts, it also has an antibacterial function. But, this adhesive layer has a disadvantage in artificial fish breeding: eggs tend to agglomerate to each other which annihilates most of them (isolate each other form oxygen).

In artificial breeding conditions, this protective layer usually cut by immersing the fertilized eggs into milk or a salt-carbamide solution. To maximize the number of living embryos we have to apply this treatment. Of course, in that way, we lost one protective layer of the egg that would somehow serve as a shield against the acidic media of the digestive system. On the other hand, this treatment also has a positive effect by hardening the remaining outer surface of fish eggs which also enhances their resistance.

Eggs were harvested, treated and fertilized according to the standard procedure used in artificial fish breeding programs at this institute. Eggs were transported to the experimental facility already fertilized and following a 24 hours incubation.

Fertilization rate was visually assessed using 3 x 100 randomly picked eggs in both species. We measured the weight of 3 x 500 random eggs of both Prussian and Common carp eggs. Both of these were ~3grams and we measured 3g batches to feed to the ducks.

Measuring the portion of eggs

During the experiment cages were placed apart to make individual sample collection possible. Plastic trays were placed under the cages to catch all the faeces they produce.

The cages placed separately to ensure individual fecal sample collection

Only after one hour following the feeding the treasure hunt began:

Fish egg retrievement from the fecal samples

Our enthusiasm was rather high in the beginning, and we didn’t have to wait long for the first eggs to appear in the faecal samples. They were present already in the samples collected in 1 hour following the feeding. This might sound surprising, but ducks have a very fast digestion. We were all heads over heels to see the first eggs, especially the embryos moving inside them:

Eggs were washed and placed in filtered river water. The work lasted very long, we had to stay up all night to follow the protocol, collect the samples in time and check every sample carefully for eggs.

The last check made at 3 am

Overall, we found 18 eggs in the feacal samples, but only 12 had viable embryo moving in these. The collected eggs were transported to our hatching facility in Debrecen. We used plastic tubes to hatch eggs recovered from the same sample separately. We additionally placed 50 eggs of both species to hatch as controls. The bottom of these tubes was made of fine mesh to allow proper aeration. Back then we thought this design will be appropriate, but later we realised that the aeration wasn’t perfect and several batches of eggs succumbed to fungal infection. This is a very typical problem of artificial hatcheries and similar fungal infections are rarely encountered in natural waters.

Retrieved eggs hatched in the lab

Eggs affected by fungal infection looked like this:

Unfortunately, our hatching infrastructure was not the best; many eggs died due to fungal infection

Eggs were continuously monitored. In the picture bellow you can see the carp egg inside the hatching tube. The developing embryo is also visible in the eggs.

Fortunately, some living embryos remained

After several hours of monitoring we saw the emergence of the first larvae from a passed common carp egg. Two more Prussian carp hatched later from passed eggs. All the control eggs succumbed to fungal infection, also illustrating that the infection was due to a faulty hatching design and not due to infection within the alimentary canal of the birds.

The hatched fishes

The whole team is very excited about future prospects of this study and we want to continue the work, exploring questions that this study raises. We are also very open to collaborations, so do not hesitate to contact us if you want to talk to us.

Details on the methodology, data, documentation, pictures and the preserved fishes that hatched are available from the authors. Contact us in case we missed to share something that you are interested in!

Ádám Lovas-Kiss (lovas-kiss.adam@okologia.mta.hu)
Orsolya Vincze (vincze.orsolya@okologia.mta.hu)
Balázs András Lukács (lukacs.balazs@okologia.mta.hu)


Read the full article here.