Detecting Spread Of Virus Over Large Populations: Analysing Sewage Water Is Far Easier And Accurate Than Individual Testing

In regular life, wastewater may be something to walk away from. But when it comes to monitoring a disease outbreak, it is a resource to embrace.

Wastewater, or sewage, can give researchers a front-row seat to the real-time movement of a pandemic and allow public health officials to prepare a response — before things escalate.

Data from wastewater analysis can supplement and enrich the information gathered through existing Covid-19 surveillance systems. Best of all, it's a safe method because there is no known case to date of an individual falling sick from direct exposure to sewage.

Last month, the Karnataka government had launched sewage surveillance across Bengaluru as Covid-19 cases shot up in the garden city. They may aim to identify infection clusters, as well as the direction in which the infection is moving — essentially if the cases are rising or falling — in various areas of the city by evaluating the sewage samples.

While this process is underway in Bengaluru, sewage surveillance has been in use in India since last year, courtesy, a team of researchers from Dr Rakesh Mishra's group at Council Of Scientific And Industrial Research-Centre for Cellular & Molecular Biology (CSIR-CCMB), Hyderabad, and from two other CSIR laboratories — CSIR–Indian Institute of Chemical Technology (CSIR-IICT) and CSIR–National Environmental Engineering Research Institute (CSIR-NEERI).

Dr Mishra is currently the Director of Tata Institute of Genetics and Society, Bengaluru.

Rounding up wastewater samples from sewage treatment plants (STPs) in Hyderabad for detecting SARS-CoV-2 RNA, a CSIR-CCMB–CSIR-IICT collaboration estimated that in August last year, about 1-7 per cent of the city's population was infected. This was when the number of Covid-19 cases by means of individual testing for the entire state of Telangana read just over 91,000 (as of 15 August 2020).

The Covid-19 testing facility at CSIR-CCMB was the stage for the wastewater analysis.

"It is known that SARS-CoV-2, the causative agent of Covid-19, is shed in the stools of patients. This shedding happens regardless of whether the patient has any gastrointestinal upset or not, or whether the patient is symptomatic or not, and the virus can be detected in the stool for several days even after the patient tests negative for Covid-19 in conventional nasopharyngeal tests," says Dr Shivranjani Moharir, post-doctoral researcher and part of the sewage surveillance team at CSIR-CCMB.

The process is rather simple. Samples are collected from STPs and brought to the laboratory, where they are filtered and concentrated. The refined samples are then used for RNA isolation. With the help of the real-time polymerase chain reaction (PCR) technique, the RNA of SARS-CoV-2 is detected and quantified. With some mathematical modelling, researchers are then able to get an estimate of the number of people infected.

"If we know the volume or capacity of the STP, or the number of people who are contributing to that STP, then we can get the number of people who are positive for the disease... because the amount of virus that is shed by a positive person is known in literature. Based on that, we can get an idea of the number of people infected in that particular community," Shivranjani says.

It doesn't take long to arrive at this number either. There is variation in the time taken for the sample to arrive at the lab, but thereafter, the processing time at the lab, along with the mathematical modelling and calculations, generally take only a day or two — or even just a few hours if things proceed smoothly.

According to Shivranjani, this output is "quite accurate". Compared to regular, individual testing, sewage surveillance throws out a more robust figure. In individual testing, asymptomatic individuals may choose not to get tested and therefore won't feature in the infection spread figure — despite carrying the virus.

But results from wastewater analysis roughly include all individuals — symptomatic, asymptomatic, and those who have recently recovered from Covid-19.

Similarly, other factors, such as a lack of access to healthcare among some communities, don't influence results in wastewater testing.

"The sewage surveillance technique doesn't have any bias. The sample has input from the whole community or area," Shivranjani says.

Wastewater-based epidemiology, which is the proper term for this kind of study, was used initially to determine poliovirus prevalence. It came to be used prominently during the outbreak of paralytic poliomyelitis in Finland in 1984 and 1985.

Although only 10 patients developed the polio disease in the country at that time, wastewater surveillance estimated that at least 1,00,000 persons were carrying the causative wild-type serotype 3 poliovirus. The outbreak quickly led to an extensive vaccination campaign.

The sewage water analysis continued during and after vaccination, as they went after the poliovirus trail till there wasn't any left behind in June 1985. It also spoke to the efficacy of the vaccination campaign underway in Finland at the time.

Sewage surveillance for poliovirus came to be used similarly through the late 1980s, 1990s, and 2000s in various countries, including in India.

After the turn of the century, however, a new context emerged for wastewater studies — to determine illicit drug use in communities.

In 2005, cocaine and its main urinary metabolite, benzoylecgonine, were found in the surface waters of populated areas in Italy using this method. The study found that River Po, the largest Italian river, carried the equivalent of about 4 kg cocaine per day. It helped shine a light on the real extent to which cocaine was being consumed in the area.

Since 2011, a multi-city study in Europe has analysed wastewater to estimate drug consumption in various communities.

Over the years, the scope of wastewater analysis has expanded to include such things as understanding general public health and lifestyle and identifying chemical pollutants in the environment.

With the outbreak of SARS-CoV-2 in 2019, the benefit of using wastewater-based epidemiology to monitor the pandemic was clear. However, its use has been limited to a few labs and small pockets of the population only.

This is unfortunate because not only can sewage surveillance provide an estimate of the infection spread at the community level, it can be a useful tool in identifying the variants of a virus, in addition to providing an estimate of the rise and fall of an outbreak — "the waves".

In a study published mid-June by mSystems, a journal of the American Society for Microbiology, researchers said they detected variant B.1.1.7 in the sewage samples collected from London, United Kingdom.

Impressively, they detected the variant in a sample from weeks earlier than was spotted by clinical surveillance.

"For variants, you need to sequence the viral genome," says Shivranjani, adding that "essentially, we can know if any particular variant is enriched".

Researchers from another CSIR laboratory, National Chemical Laboratory (CSIR-NCL), Pune, have successfully sequenced the SARS-CoV-2 genome from sewage samples.

The burning question of whether there will be future waves of Covid-19 can be investigated using wastewater-based epidemiology. Regular measurements can detect a rise in the viral load as it happens, preventing delays in observing and responding to disease trends.

"In villages and small towns, where proper drainage systems and sewage treatment plants may not be available, the samples can be collected from drains/nala. One can analyse the viral load over time at a particular sampling point. The peaks and troughs in the trend of viral load over time will give an idea of the prevalence of infection in the surrounding area," Shivranjani says.

Wastewater-based epidemiological studies on a larger scale can give public health officials a bird's eye view of population-level infection dynamics and immunity.

Perhaps, most important, it can act as an early warning system for potential future outbreaks.