Fecal forensics track sources of water contamination

Escherichia coli, or E. coli, as it is widely known, is the go-to microbial marker to test whether water is safe to drink or to swim in. A common strain of bacteria found in the gut of humans and all warm-blooded animals, E. coli is a practical tool for detecting fecal contamination of our waters. The testing is easy to do, is relatively quick, and does not require a lot of training or expensive equipment. It entails taking a water sample, filtering it and culturing the E. coli to find out how many cells are present. Any detection of E. coli in drinking water triggers a boil-water advisory and a reading of 100 or more E. coli per 100 milliliters of water closes a beach.

It's not a perfect tool, though. As Dr. Tom Edge, an Environment and Climate Change Canada researcher specializing in waterborne pathogens, explains, "There are situations where E. coli testing will have limited or no value in indicating a waterborne health risk."

View of Cryptosporidium parvum oocysts and Giardia lamblia under a microscope

Cryptosporidium parvum oocysts and Giardia lamblia (intestinalis). Bar = 10 microns. Credit: H.D.A. Lindquist, U.S. EPA" http://www.epa.gov/microbes/images.html

As an example, he points to Milwaukee. In April 1993, some 400,000 people – half the city's population – were stricken by gastrointestinal problems caused by the waterborne parasite Cryptosporidium. More than 50 people died as a result, and health care costs spiraled over $50 million. The water supply had been carefully monitored for chlorine-sensitive bacteria like E. coli, which, unfortunately, may not indicate the presence of a chlorine-resistant pathogen like Cryptosporidium.

E. coli testing also does not identify the source of the fecal contamination, which needs to be determined to identify solutions to the contamination. Dr. Edge thought the emerging field of microbial source tracking could provide new forensic tools that would track the pollution back to its precise source.

His initial research applied DNA fingerprinting and antibiotic resistance profiling to E. coli from fecal pollution samples. The work was not pretty. It involved collecting thousands of E. coli isolates from sewage treatment plants, along with dog, cat and bird droppings, in Hamilton, Ottawa and Toronto. What Dr. Edge found was that the E. coli in water samples from beaches often better matched the DNA fingerprint and antibiotic resistance profile of E. coli from seagulls and Canada geese than that from humans, dogs or cats.

That's when, Dr. Edge says, "We started realizing that birds were an important part of beach closures around the Great Lakes." And if that was true, it was going to be crucial information for municipal officials, who needed to track the source of E. coli contamination in order to fix the problem. If it was a human source, then the sewage system was the problem, usually requiring a costly engineering solution. If it was an animal source, then less costly mitigation measures could be implemented.

Advances in genomics offered a way to pursue this line of investigation without the time-consuming and expensive need to collect thousands of E. coli from fecal samples. Rather, all the microorganisms in a water sample could be trapped by a filter and recovered, and then all of their DNA extracted and examined for DNA sequences of microorganisms that could only have come from a specific animal's gut.

The problem was that not much work of this type was being done in Canada on birds. With funding from the Genomics Research and Development Initiative (GRDI), Dr. Edge was able to team up with scientists at the U.S. Environmental Protection Agency, who were also looking at microorganism DNA sequences from fecal samples to see if they could find a microorganism marker for seagulls that was unique from other birds and mammals.

Seagulls on a beach in Toronto, Canada

Large seagull colonies are a fact of urban life in Canada. Given that each seagull produces up to 64 droppings a day, it's little wonder that they are a major cause of beach closures. Credit: Tom Edge

Together they pursued cloning and sequencing of the DNA of common strains of bacteria found in the gut of the seagulls from around Toronto, and hit what seemed to be pay dirt. After testing their discovery against Dr. Edge's many fecal samples and those from across the United States, they were able to confirm it. They had, indeed, found a new and unique bacterial marker that could distinguish seagull fecal contamination from that of all other birds and animals.

Dr. Edge says the big surprise was just how widespread the seagull marker was in several Ontario cities. In urban centers with large colonies of seagulls, it was everywhere in the water samples – from the beaches to runoff from parking lots. Consider that over 100,000 seagulls nest at Toronto's Leslie Spit alone and that they range all over the city, and you have a serious source of potential contamination.

The results from this genomics research are changing how municipalities manage their beaches. City beach managers no longer automatically default to looking for a problem with the sewage system. Rather, they are now considering contamination from seagulls and taking measures to control the birds' access to beaches. In Ottawa, overhead wires have been installed at the Mooney's Bay and Britannia beaches to deter seagulls and Canada geese. Dogs are also used at Britannia Beach to chase the birds away, and an all-natural fertilizer made of fish compost and saw dust is being used successfully at the Petrie Island beach to keep Canada geese from living there.

The seagull marker is now being used around the world, based on the published results. This is an important phase in the research cycle, as it will validate whether the seagull marker has universal application as a tracking tool for contamination caused by seagulls. And so far, it is looking like it does. Only a small number of seagull populations have been found to have lower levels of the bacteria in their gut, and it very rarely appears in other bird species. Considering the diversity of seagulls and bacteria in the world, this new and unique DNA marker is indeed novel.

Award-winning innovation

The U.S. Environmental Protection Agency (EPA) presented its Scientific and Technical Achievement Award in 2010 to Jorge Santo Domingo (EPA), and Tom Edge and Stephen Hill, both with Environment and Climate Change Canada, for their breakthrough work on the seagull marker. The U.S. EPA presented its Scientific and Technical Achievement Award to this team again in 2014 for additional novel molecular assays to detect waterfowl fecal contamination in environmental waters.

Their research used genomics techniques to discover new DNA markers for bacteria found only in the gut of seagulls and other waterfowl. The result is new tools to detect and measure bird fecal contamination in water samples, allowing a more targeted and cost-effective solution for the clean up of pollution sources, and ultimately reducing the contaminants entering the environment.

Dr. Edge's research is continuing under the new Government of Canada GRDI's interdepartmental EcoBiomics Project, where it will use next generation DNA sequencing techniques to characterize the enormous microbial diversity found in water samples. For the first time, these techniques are enabling scientists to see far beyond the DNA from E. coli and seagull gut bacteria, to the DNA from many millions of microorganisms. Better characterization of aquatic microbiomes (all the microorganisms and their gene diversity) will provide more comprehensive information on the composition of water samples to help identify pollution threats and guide decisions to protect water quality.

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