News: Sexually Confused Fish Are the Canary in the Coal Mine of Water Pollution—But Bacteria Could Come to the Rescue

Exposed to hormones, pharmaceuticals, and other chemicals, the beautiful wild fish in Canada's Grand River have taken on some pretty odd characteristics—they're turning into females. A long-term study suggests using bacteria to manage polluted water could turn the tide for feminized fish.

A native species, Etheostoma caeruleum, known as the rainbow darter, comes by its name honestly. Colorfully banded in shades of blue and red-orange, rainbow darters grow to about three inches and reach old age by five. Fond of clean headwater streams, darters are a sentinel species given their high sensitivity—and vulnerability—to pollution.

In a study from the University of Waterloo, published in the journal of Environmental Science & Technology, scientists had the opportunity to evaluate darters before and after improvements were made to their habitat, located downstream from municipal wastewater treatment facilities.

In southern Ontario, pollution in the water is linked to the intersex changes in rainbow darters that have been observed for years. In the research, intersex refers to a condition when male fish possess mixed gender physical attributes, including ova (female reproductive cells) located within the testes of male fish. As could be expected, intersex changes align with lower reproductive success, among other impacts.

What Causes Male Fish to Be Feminized?

The darters in this research were studied in their natural environment—downstream from effluent discharged from wastewater treatment plants in Kitchener and Waterloo, Ontario. The habitat, known as the Grand River watershed, is the largest to drain into Lake Erie, supporting a human population approaching one million.

Because of their rapid growth, large numbers, and relatively short lifespan, darters make good study subjects. In addition to the development of ovum, male darters in these waters also grow smaller testicles, have reduced steroid levels, and show alterations in how their genetic material expresses itself. Overall, not a good development. So why is it happening?

For fish in a wild environment, it is difficult to pin down exactly why off-kilter genetic and physical changes take place. That said, compounds identified in the darter habitat known to cause feminizing characteristics in fish include:

• natural hormones
• synthetic hormones from industrial chemicals
• pharmaceuticals including ibuprofen (non-steroidal anti-inflammatory), naproxen (NSAID), and carbamazepine (anticonvulsant)

These are all examples of endocrine-disrupting compounds (EDCs)—natural or synthetic chemicals that interfere with developmental, immune, neurological, reproductive, and other processes in humans and wildlife. Endocrine disruption (changes in hormone levels caused by chemicals in the environment) in fish or mammals is a serious concern and may underpin abnormal intersex expressions in fish and other forms of life. In humans, endocrine disruptors may cause other negative symptoms like lowered fertility, an increased incidence of endometriosis and some cancers, and negative effects in children and developing embryos during pregnancy.

Years ago, compounds like DDT were used for household and industrial purposes. Only later, DDT was discovered to have serious and lasting harmful impacts on human health because of its hormone-disrupting capabilities.

More recently, research about plasticizers that mimic hormones, (the most well known is bisphenol A, also called BPA), brought awareness that endocrine disrupting chemicals are accumulating in our bodies even before birth. While some animal studies suggest that infants and children may harmed by BPA, that link hasn't been confirmed in human subjects.

To figure out how endocrine-disrupting chemicals impact fish feminization, and if intervention methods can stop these effects, the researchers closely evaluated changes in darter populations for more than 10 years. These fish were living in streams in the outflow of municipal water treatment plants, and during the study, they started using beneficial bacteria to filter their water output.

Why Nitrification?

To improve water quality and efficiency, significant changes to the Kitchener Municipal Wastewater Treatment Plant were conducted and completed in 2013. Part of the changes involved implementing nitrification. Well—what is nitrification and how can it help?

Nitrification is a matter of microbes. If you know someone with a healthy aquarium of fish, they can tell you about the need for beneficial bacteria that form compounds called nitrites and nitrates. When you set up a fish tank, or say, a wastewater treatment plant, using microorganisms to create a landscape of beneficial bacteria helps break down waste like feces, urine, and even pharmaceutical products, into harmless compounds.

The nitrogen cycle starts with ammonia. Ammonia concentrations are typically high in wastewater treatment facilities, due to human waste, breakdown of proteins, chemicals, and other materials. Nitrifying bacteria are naturally present in the environment. When ammonia builds up, nitrifying bacteria quickly colonize, consume, and convert the ammonia to nitrite. The nitrite is then consumed and converted by another type of nitrifying bacteria (Nitrosomonas and Nitrobacter, respectively) to more harmless nitrate.

In your aquarium, this community of nitrifying bacteria is located in the gravel, or the filter, and is called a bacteria field. In a wastewater treatment facility, a similar process results in nitrifying activated sludge.

In the Kitchener Wastewater Treatment facility, the installation of the sludge bed and increased aeration resulted in significant drops in the level of ammonia (which is toxic to fish). As effluent percolated through the nitrifying bacteria, levels of ammonia and endocrine disrupting compounds went down in the outflow water.

Mark Servos, Canada Research Chair in Water Quality Protection in the Biology department at the University of Waterloo, and one of the study's researchers, noted to Invisiverse that "nitrifying conditions promote different bacterial communities and this results in better treatment. [Endocrine disruptors] are expected to be removed more effectively."

So What About the Fish?

Prior to introduction of the nitrification process at the Kitchener Water Treatment Plant, between 70% and 100% of male darters living in effluent waters downstream of the plant displayed intersex characteristics, one of the highest rates of any location in the world.

The implementation of nitrification at the Kitchener plant dramatically improved the plant's overall effluent quality in terms of observed concentrations of nutrients, concentrations of pharmaceuticals, and total estrogenicity.

Within three years after upgrades were complete at the facility, less than 10% of the fish carried markers of intersex confusion. In a press release, Servos states:

Rainbow darters are the Grand River's canary in the coal mine. They're extremely sensitive to the concentration of estrogens and other hormone disrupters in the water. Still, we didn't expect them to recover so quickly.

The study also suggests native fish can recover from earlier toxic exposure, with individuals showing less physical feature confusion over time, as the water quality of their habitat improves. Servos told Invisiverse that, although they did not sample other fish, "we have data that other species were impacted (by the effluent) and we think the improvements will be across many species."

Small but powerful, the nitrifying bacteria are digesting the pollution and reducing the risk of EDCs to those living downstream, whether it is fish or families.

"Having long-term data of the fish population, before and after the wastewater treatment upgrades makes this a truly unique study," Servos said. "The changes to Kitchener's wastewater treatment system have had a much larger positive impact then we had anticipated."

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