Here we go again – Many steps to an experiment on black worms

After a year as a teacher I came back to research in aquatic ecotoxicology. I’ll test a method to analyze fullerene nanoparticles in separated tissue fractions of black worms. Simply, I’ll expose the worms to fullerenes, collect organisms, fractionate their tissues, and then measure fullerene concentrations in each tissue fraction. But starting a new experiment requires a plenty of preparations in the lab before actual test can be started. Here I tell what is going on during the first two weeks.

I would need a test sediment treated with fullerenes. For the test sediment, I would need fullerenes suspended to water to be added to a natural sediment from Lake Höytiäinen. Luckily, we already had the sediment in our lab… if we didn’t have, I would have to wait for winter to go to the field and collect it through ice… I would also need my test species, black worms, synchronized to similar physiological condition.

Sediment sampling. Pictures by Kristiina Väänänen and Jarkko Akkanen
Sediment sampling during winter time.

As a very first job, I prepared artificial freshwater, which means a lab-made model of fresh water corresponding “average Finnish freshwater” with its hardness. Then, I used that water to suspend fullerenes. Making fullerene suspension takes time: fullerene powder must be vigorously mixed with water for two weeks before it can be used in the experiment. This mixing process must be done because fullerenes are not soluble in water, but they turn to water-stabile form via water flows and mixing. And when thinking about fullerenes’ fate in natural waters, they can enter to the environment e.g. in waste waters. Thus, water suspension is their first step to bottom sediments. Read more about fullerenes’ environmental fate here: http://onlinelibrary.wiley.com/doi/10.1002/etc.2175/full

Fullerene suspension, picture by Kukka Pakarinen
Fullerene suspension.

Black worms are sediment-dwelling benthic worms. They have important ecological roles in aquatic ecosystems as a food source for fish and as decomposers of sediment material. They can be exposed to fullerenes via wasted sediments. In this experiment I’ll need size-synchronized worms, as some other researchers in our group. That’s why we organized “a worm cutting day” to synchronize more than thousand worms. It means that four of us sat a day in the culture room picking worms from their aquariums to petri dishes, and then separating their head parts and tail parts by a surgeon knife: the head parts grow new tails and tail parts grow new heads. How to identify which part is which? Color of the head is a bit black and thicker whereas the tail is red and thinner. Then, we’ll wait for couple of weeks to let the worms create these new parts. Finally, we’ll get test worms with same size and condition. Dividing to heads and tails is also a normal way to reproduce for the black worms. Read more about fullerene-exposed black worms here: http://www.sciencedirect.com/science/article/pii/S0269749111003848

Worm cutting day
Worm cutting day
Head part, tail part and cutting
Head part, tail part and cutting

While fullerene suspension and the worms are underway, I can do some other preparations. Sediment dry weight must be known to adjust volume of fullerene suspension. Preparations for the dry weight could be favorite job for kids: wet sediment is homogenized with a perforated piston before samples are placed to weighing jars and dried.

Mixing and weighing the sediments
Mixing and weighing the sediments

Next week it’s time to measure fullerene concentration in the suspension, add fullerenes to sediment and let them stay to equilibrate before the experiment.

Text by Kukka Pakarinen

Pictures by Jarkko Akkanen, Kristiina Väänänen, Kukka Pakarinen, and Risto Pöhö