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Explainer: Wastewater

COVID-19 detections in wastewater have been making headlines for the past 12 months – including this week in Melbourne. 

Prior to the pandemic, our wastewater was sampled and tested for a variety of reasons – including to monitor the environment, look for other viruses, and track illicit drugs. Those activities are all still happening, alongside COVID wastewater monitoring programs running across Australia, with detailed information available for NSW, VictoriaQueensland,South AustraliaWestern Australiaand the ACT.

So, with wastewater unlikely to leave the news anytime soon, it’s worth taking a look at how the testing is actually done.

How is the water collected?

“The wastewater is collected using an automated sampler, which is housed at the wastewater treatment plant,” say Richard Bade and Cobus Gerber, both researchers in analytical chemistry at the University of South Australia.

“In Australia the water utilities or local councils manage and operate the plants.”

The specific type of sampler varies depending on the state and council. Since sewage flow changes throughout the day – with residential areas peaking in mornings and evenings, for instance – samples are usually combined together from a 24-hour period. A sampler might collect the same volume of water every 15 minutes, or it might collect larger and smaller amounts across the day. In total, a few litres per day are collected for sampling.

 The wastewater treatment plant refrigerates or freezes the liquid until all samples have been collected, and then sends some of this 24 hour-composite for analysis by Bade and Gerber.

“For our work, we collect between 100 and 600 mL of the 24h composite sample, but the operators generally use additional volumes for their own tests,” says Bade.

How is it tested?

Once in the lab, analyses vary depending on what you’re looking for. For instance, COVID-19 is most easily detected in faecal matter – solids. Drugs can be different.

“For our drug analysis, the wastewater samples are first filtered to remove any solid matter. As the drugs that we monitor are predominantly excreted in urine, they should not be in the solid fraction,” says Bade. 

The sample is then chemically treated depending on the drugs they’re looking for – the aim is to have a concentrated amount of the compounds they’re after, without too many other things to confuse their readings. To identify the molecules, they usually use liquid chromatography and then mass spectrometry. Chromatography separates the mixture into individual compounds, and mass spectrometry identifies the atoms and molecules, as well as their abundance.

“We look both at specific drugs (targeted) or screen generally for compounds of interest,” says Bade. 

The molecules in the COVID-19 virus are too large and complicated to show up in this type of analysis.

Testing for COVID – and other viruses – requires a PCR test. This is the same test that’s done on nose and throat swabs: it amplifies any COVID RNA that might be present. To do this, a small section of RNA that appears in the SARS-CoV-2 genome is used as a probe, to find suspicious RNA. This probe then looks for matching pieces of RNA in the sample, and – if they’re present – the viral RNA is copied enough times that it can be easily detected. The process is done at a high enough temperature to denature the RNA without destroying it completely, so it doesn’t pose a risk to the researchers.

As most state authorities will tell you, testing for SARS-CoV-2 in wastewater can be a helpful indicator of presence of COVID-19 in the area, but it’s not a guarantee that there’s an infectious person in the community. Viral shedding can happen for several months after infection in some people.

Similarly, the virus denatures and dissolves fast in wastewater, so a clean sewage reading doesn’t mean there’s no COVID-19 around.

What else can we look for?

Gerber says looking for SARS-CoV-2 is similar to looking for other viruses, but you need to know what you’re looking for beforehand. “The only difference would be that in order to check for the sensitivity and the specificity of the method, actual viral RNA is required to develop and validate the method.” For instance, to look for hepatitis, we’d need a section of hepatitis RNA.

With smaller molecules like drugs, there’s more flexibility. “A general screening method allows, in theory, an infinite number of compounds to be analysed. The only drawback of the screening approach is that you can only say whether or not a drug is present rather than how much of a drug is present,” says Bade.

Alongside ingested drugs, which were the subject of a recent paper whose authors included Bade and Gerber, authorities can also look for drugs that weren’t used – or by-products from illicit drug manufacture.

“In terms of the general procedures and methods, there is no difference between ingested drugs and their precursors,” says Bade.

“The main difference is in which compound to look for. For example, a metabolite of a drug will only show ingestion, while the parent drug (or precursor/byproducts from manufacture) can show manufacture or direct disposal.”

We need to examine how illicit drugs are metabolised by the body – and therefore how they might show up in urine – to tell whether or not they’ve been used.

And other testing techniques can find yet more information.  “Pesticides, antibiotics, personal care products, pharmaceuticals, alcohol and tobacco have all also been measured in wastewater,” says Bade.

Can testers catch COVID?

Sewage is something we’re taught to avoid, for very good reasons. There are a lot of bugs in our effluent, so professionals need to be careful when working with it.

The risk from COVID infection in particular is pretty low, though.

“Wastewater contains microbes which secrete enzymes which in turn degrade viral RNA quite rapidly. Thus, the infection risk is relatively low and by the time the water is treated at a plant, very little viable virus would remain,” says Gerber.

“Bear in mind, this risk is not really any different for the usual operations of those in charge of the sewer network, as various pathogens exist in wastewater and precautions would (should) already be in place.”