For months, veterinarians placed medication in the animals’ quarantine habitats at Chicago’s Shedd Aquarium to ensure that animals entering the building did not bring dangerous pests or pathogens with them. And for months the drug kept disappearing. Where did it go Who took it And what was your motive?
To solve this classic crime mystery, researchers from the Shedd Aquarium teamed up with microbiologists from Northwestern University to gather clues, follow up leads and ultimately track down the culprit.
After microbial and chemical analyzes of samples from the marine aquarium systems, the team determined that there was not just one culprit, but many: a family of microbes that are starving for nitrogen.
“Carbon, nitrogen, oxygen and phosphorus are basic needs that everything needs to be alive,” says Erica M. Hartmann from Northwestern, who led the study. “In this case, it looks like the microbes are using the drug as a source of nitrogen. When we examined how the drug was broken down, we found that the piece of the molecule that contained the nitrogen had disappeared. It would be like just eating the pickles from a cheeseburger and leaving the rest behind. “
An expert in indoor microbiology and chemistry, Hartmann is Assistant Professor of Civil and Environmental Engineering at the McCormick School of Engineering in Northwestern.
When a new animal enters the Shedd Aquarium, it must first go through a quarantine process before entering permanent residence. This allows the aquarium vets to monitor the animal for potentially contagious diseases or parasites without endangering other animals in the facility.
“The quarantine habitats of the Shedd Aquarium behind the scenes are a first point of contact for animals that enter the building – so we can safely welcome them so that no pathogens from outside are introduced into the animals that Shedd already call home” said Dr. Bill Van Bonn, Vice President Animal Health at Shedd Aquarium and co-author of the study. “We are grateful to have worked with Northwestern University to research scientifically what is happening microbially in our quarantine habitats, how we manage them and continue to ensure optimal welfare for the animals in our care.”
Antiparasitic drug mysteriously disappeared
During this quarantine process, all animals are given chloroquine phosphate, a common drug against parasites. Veterinarians proactively add it directly to the water as a pharmaceutical bath to treat a variety of diseases. After adding chloroquine to water, aquarists then measure the concentration of the drug. Then they realized that something was wrong.
“You have to maintain a certain concentration in the habitats in order to be able to treat the animals effectively,” says Hartmann. “But they noticed that the chloroquine was mysteriously disappearing. They added the right amount, then measured it and the concentration would be much lower than expected – to the point where it stopped working. “
Shedd Aquarium aquarists collected water samples and swab samples and sent them to Hartmann’s laboratory. Swab samples were taken from the sides of the habitats as well as from the incoming and outgoing pipes. In total, the team found around 754 different microbes.
“Of course there are microbes in the water, but there are also microbes that stick to the sides of surfaces,” said Hartmann. “If you’ve had an aquarium at home before, you’ve probably noticed dirt growing on the sides. Sometimes people add snails or algae-eating fish to clean the sides. So we wanted to investigate what is in the water and what is stuck to the sides of the surfaces. “
Study ‘leftovers’ from food
By examining these samples, the Northwestern and Shedd Aquarium teams first determined that microbes were causing the drug to go away, and then located the microbes responsible. Hartmann’s team cultivated the collected microbes and then supplied chloroquine as the sole source of carbon. When the results of this experiment were inconclusive, the team performed delicate analytical chemistry to examine the degraded chloroquine.
“When the chloroquine was eaten, we essentially looked at the leftovers,” said Hartmann. “That’s when we noticed that nitrogen is the most important driver.”
The unusual suspects
Of the 754 microbes collected, the researchers narrowed it down to at least 21 different guilty suspects – belonging to the Phyla Actinobacteria, Bacteroidetes, Chloroflexi, and Proteobacteria – that live in the habitat’s outlet pipes. In fact, some of the microbes appear to be brand new and never studied before.
“We couldn’t find a single perpetrator, but we were able to isolate the specific location,” says Hartmann. “Our results indicated that simply flushing the quarantine habitats with fresh water would not be enough to fix the problem as the responsible microbes were clinging to the sides of the pipes.”
Hartmann said the pipes may need to be scrubbed or replaced to prevent chloroquine from disappearing in the future. Another possible solution might be to switch between fresh and salt water regularly, as microbes are usually sensitive to one or the other.
“Everyone in the Shedd Aquarium is obviously very committed to the health and welfare of the animals and is very enthusiastic about research,” said Hartmann. “It was really cool to work with them because we could help the animals and we might have discovered new organisms.”