What we learn about the Corona virus through waste water research

When more people are infected in a city, more viruses enter the sewers through the stools. What we want to know now is whether the virus is present in sewage water. Not because we think that the virus is likely to spread via water, but mainly because we could get an indication of the number of virus infections in the population of a city by monitoring the levels of the virus in sewage water.

Since only COVID-19 patients with severe symptoms are tested in the hospital laboratories, the reported figures underestimate the real number of COVID-19 patients. With regular sewage measurements we could provide additional information on the circulation of viruses in the population. Using this method we might also be able to measure whether the number of virus infections in a city will increase again next winter.

How do we do this research?

Together with the water boards and assisted by the managers of wastewater treatment plants (wwtp), we take samples of sewage water entering the wwtp over 24 hours. From these samples we extract the virus using methods that have been developed for other viruses (which are transmitted via water, such as norovirus). We purify the genetic material of the virus (RNA) from the concentrate and very specifically test whether SARS-CoV-2 virus genes are present. We do this using the RT-PCR method. To have more certainty that the virus is really present, we test against different gene fragments and multiple genes of the virus.

The investigation started on 6 February 2020, three weeks before the first official coronavirus infection in the Netherlands. This first measurement was needed to test the method and see if we did not see signals while the virus was not present. We took the second series of samples on 5 March, a week after the first COVID-19 patient in the Netherlands was reported. The third series of samples were taken on March 15.

What are the results?

The method we use in KWR’s microbiological laboratory is developed by ‘on-the-fly’. In this crisis situation, there is no time to first develop, test and validate a method as we would normally do. This means that we have preliminary results of our measurements. We currently examine all samples several times and look at the reproducibility of the results. Furthermore, we double check and focus on fragments of multiple genes.

In the samples taken on 6 February, none of the tests showed a reaction, so there was no presence of the virus in sewage water. Which is what we expected, since the virus was not present in the Netherlands yet.

For the 5 March samples, one of the gene fragments did show a reaction at four waste water treatment plants (wwtp). Because the other gene fragments showed no reaction, we were less confident about the results and reported them as ‘inconclusive’. For us, however, it was an indication that there was a change compared to February 6. We did additional tests to try to prevent inhibition of the test. That worked and that made the test result clearer.

In the 15 March samples, the gene fragments, which showed a reaction on March 5, showed a reaction again, which was also stronger than on March 5. Two other gene fragments also showed a reaction in some of the samples. Therefore, we reported that we found the new coronavirus in the sewage water (again: as a preliminary result).

We are fairly sure that what we detect is actually SARS-CoV-2 for two reasons.

1. Multiple gene fragments and genes of the virus show a reaction.
2. Since we saw no signal of the virus in the Netherlands before the circulation, and in the measurement series of March 5 we see a reaction at one of the gene fragments and in the measurement series of this week we see a reaction at one (Coevorden), two (Harnaschpolder, Amersfoort and Apeldoorn), three (Amsterdam West, Schiphol, Tilburg) or four (Utrecht (incl. Overvecht)) of the gene fragments. The water after sewage treatment at wwt Tilburg was negative on both days tested.

How do we interpret the results of our research so far?

• The results indicate that SARS-CoV-2 is present in sewage water
• SARS-CoV-2 was not found in effluent (that is sewage water treated by the wwtp). So far we have only investigated this in Tilburg.
• The method is not yet quantitative, but based on the strength of the signal, the concentration of virus in sewage seems low.
• Our estimate is that the risk that employees at the wwtp run of getting COVID-19 through contact with sewage (aerosols) is very low. See for this subject the website of STOWA and RIONED (both in Dutch).
• We think that the SARS-CoV-2 screening of sewage water can be used as a tool to measure the virus circulation in a population (e.g. a city or a smaller municipality). If we can further substantiate and validate our method, the water sector will have a tool that provides valuable additional information about the spread of the virus in the population.

The current total picture of virus circulation in the population of the Netherlands is incomplete, because the test kits are limited. Most people who (fortunately) have mild symptoms are not tested, since  testing is mainly (and rightly so) reserved for use in hospitals for patients with serious medical conditions.

Our sewage screening can help to get a better picture of the virus circulation. When the current peak (hopefully it will stay as flat as possible!) is over, sewage screening also helps to detect early – possibly coming winter – if the virus circulation is increasing.

Wat het riool vertelt over het coronavirus from KWR Water Research Institute on Vimeo.