Australasian Science: Australia's authority on science since 1938

Probiotics for the Planet’s Polluted Plumbing

Credit: Eraxion/iStock

Credit: Eraxion/iStock

By Matthew Lee & Mike Manefield

Imagine a world where billions of tiny creatures were deployed in the environment to degrade industrial pollutants that contaminate the world’s crucial groundwater reserves.

Fresh water is one of the most precious resources on Earth. The vast majority of fresh water sits below ground in aquifers, from which it is sourced for human consumption and agriculture in many parts of the world. Keeping this precious resource clean from contamination is a crucial concern.

We have been developing ways to use microbes to clean up contaminated water, and recently cracked one of the world’s most difficult chemical pollution problems: breaking down the toxic organochlorine chloroform. Not only have we discovered bacteria that can do the job, but we’ve also found how to get them to work eight times more efficiently than normal.

Organochlorines are a group of chemicals that have been mass-produced by humankind since the 1960s because their volatility, low solubility, high density and stability make them very useful for manufacturing plastics, rubber and refrigerants, degreasing engines and dry cleaning clothes.

Unfortunately, owing to poor handling and improper disposal, organochlorines routinely find their way into aquifers and contaminate groundwater. Here the properties that make them so useful are the same that make them a nightmare in the environment. Because of their low solubility and stability they take centuries to break down. Because of their density they sink to the bottom of aquifers and are hard to access. Because of their volatility, human and environmental health are at risk when the groundwater meets surface water bodies such as rivers, bays, harbours and oceans.

In acute doses organochlorines are toxic. Extended exposure to low doses will give you cancer.

Fortunately, bacteria in groundwater can degrade organochlorines. They are not very abundant and they are fastidious, fragile and slow-growing, but scientists have developed ways and means of exploiting these microscopic critters to clean up our mess.

They are called organochlorine-respiring bacteria (ORB), which means they don’t break these compounds down by eating them; rather, they breathe them. Ironically, they find oxygen toxic in the same way we find what they breathe toxic, which makes them that bit more challenging to handle.

Over the past decade our laboratory at the University of NSW has been ORB-hunting. We’ve taken groundwater and sediment samples from some of the most polluted sites in Australia – Botany Industrial Park in Sydney, and the Altona Chemical Complex in Melbourne – and incubated them under very specific conditions in the laboratory to select for the bacteria we want. Through painstaking nurturing over years, during which we needed to protect them from oxygen, we have discovered bacteria that can break down the most common organochlorine groundwater pollutants: perchloroethene for dry cleaning, chloroform for refrigeration, and dichloroethane for PVC production.

Having these special bacteria in isolation in the laboratory gives us a chance to sequence their genomes and find out what makes them tick. By studying the expression of the genes encoded in their DNA, looking at both gene transcription (the production of RNA from the DNA code) and gene translation (the production of protein from the RNA code), we can learn how these bacteria have evolved to breathe toxic groundwater pollutants.

Furthermore, by isolating these bacteria we can then grow them in large numbers and add them back into contaminated groundwater to accelerate the natural clean-up process. We grow the bacteria up in beer kegs in a similar way to which brewers grow yeast to make beer. Then we transport these kegs of pollution-degrading bacteria to contaminated sites and inject them below ground through narrow wells drilled into the aquifer. If conditions are friendly the bacteria will use the pollutants to grow. It’s a win for them and a win for us.

We are proud to have been responsible for the first deployments of chlorinated ethene-, ethane- and methane-degrading bacteria for the bioremediation of organochlorine-contaminated groundwater on the Australian continent.

Three years ago we established an Australian business, Micronovo Pty Ltd, to ensure that the expertise and cultures we have developed are accessible to custodians of contaminated sites and environmental consulting companies. These people send contaminated groundwater samples to Micronovo and we use molecular tools to check for the presence of microbes that can break down the particular pollutant at the site from which the samples came. If the right microbes are there, we offer advice about how to make conditions just right for them to flourish along with nutrient formulations to support this. If the right microbes are missing, we offer them the matching bacterial culture.

On one hand our bacterial cultures are like very clever micromachines that unpack themselves in the aquifer and mop up the toxins. On the other hand our cultures are like probiotics for the polluted plumbing of the planet. Once the pollutant is gone, the bacteria we injected rapidly die off because they have nothing to breathe. Like us, they can last for a while without food, but not so long if they are suffocating.

In 2012 we discovered the first bacterial culture in the world that can completely degrade the infamous organochlorine chloroform. Once used as an anaesthetic (until it was recognised as a human carcinogen and a cause of cardiac failure), chloroform has been mass-produced as an industrial solvent since the 1960s. At the height of production in the 1980s, the US produced 250,000 tonnes of chloroform every year. Chloroform is ranked eleventh in the US EPA’s Priority List of Hazardous Substances.

Currently, 474 of the 1287 polluted sites on the US priorities list are contaminated with chloroform. In Australia, the numbers are smaller but the proportions are the same. Chloroform contamination in the environment is a major problem globally.

The tricky thing about chloroform is that it inhibits the growth and activity of bacteria that can break down other pollutants. At most polluted sites you have more than one contaminant, so the presence of chloroform effectively puts on hold the natural attenuation of the contamination. For this reason, the discovery of a chloroform-degrading bacterial culture is a major international scientific breakthrough enabling the removal of chloroform so that bacteria that are indigenous to the site can degrade the remaining pollutants.

But it’s not all about the microbiology. As microbiologists we play our part alongside environmental engineers, hydrologists, geologists and chemists to understand what is happening in the water bodies below our feet. Groundwater ecosystems are much like those on land or in the oceans – they consist of a high diversity of organisms interacting with each other and with the surrounding abiotic environment. It’s a little like a subterranean rainforest – complex, catalytic, beautiful and valuable.

In recent work we have been exploring the interplay between different remediation technologies. One of the alternatives to bioremediation is the use of iron. Elemental iron (not iron oxide) can do much the same trick chemically as the bacteria do biologically, but it has some important limitations: it is more difficult to get into the ground, and there are some organochlorines that bacteria can break down but iron cannot.

In the industry these are considered competing technologies, but this “one versus the other” mentality has obscured an important point revealed by our latest research: they work best in combination. In an award-winning study published this year in Environmental Science and Technology, we have shown that chloroform is broken down eight times faster when a culture of our chloroform-degrading bacteria is combined with iron, compared with just iron alone.

Using bacteria to degrade pollutants is the most cost-effective and sustainable solution to groundwater contamination owing to the low demand for energy. The trade-off is that it is usually slower and requires microbiology expertise to administer successfully.

In comparison with other industrialised countries, Australia has been slow to adopt biological remediation technologies, possibly owing to its mining and agriculture heritage – if it doesn’t involve a big machine it isn’t going to work. With the development of world-class expertise, the commercial availability of services and cultures and countless addresses to government regulators, community groups and clients, the Australian remediation industry is warming to the little guys that turn the biogeochemical cycles of our planet.

Minima maxima sunt. The small things are the great things.

Dr Matthew Lee and Associate Professor Mike Manefield work in the School of Biotechnology and Biomolecular Sciences at The University of NSW.