In this exclusive interview, we speak with Dr. Kendra Maas to find out how she is developing a monitoring tool to analyze SARS-CoV-2 levels in wastewater. As the facility director for the Microbial Analysis Resources and Services (MARS) department at UConn, Maas is spearheading a program that aims to pinpoint where and when SARS-CoV-2 enters a wastewater system. By using Nanotrap® particles to rapidly isolate the virus from wastewater, this tool could prove crucial in limiting the potential scope of the outbreak.
KM: As a scientist at a fee-for-service core facility, I am a technical expert for other people. We analyze samples from all sorts of environments and work on multiple projects at once, making sure a project is functional before handing the research back to the scientist who commissioned it.
By March 2020, it was apparent that COVID-19 testing in the U.S. was lacking, so I started petitioning for MARS to bring some of that testing in-house. Since most microbiome analysis is PCR-based, it was easy to switch to looking for viruses with PCR as well.
I have prior experience working in wastewater research, so it was a good starting point for my first self-owned project at UConn. Once people get over their initial disgust with wastewater, understanding the implications of research based on ‘sewage’ is fascinating to them. Wastewater research enables broad population-level epidemiology approaches, which UConn is very interested in. Moreover, UConn has its own wastewater treatment plant that helped me skip the bureaucratic hurdles and approvals to get those initial samples, and the plant management was excited to collaborate.
KM: The goal of this work is to get wastewater monitoring for SARS-CoV-2 up and running as quickly as possible – which is the need of the hour. We want the public to get engaged in thinking – “How do we want to use this huge wastewater surveillance tool in a way that benefits the town?” Then, once we have this infrastructure up and running, the immediate goal is to circumvent broad shutdowns to keep schools and towns open. This tool will help us locate and catch outbreaks early so we can make targeted shutdowns.
KM: A huge proportion of the population - about 40% of the people who are infected and infectious - are asymptomatic or pre-symptomatic. Young adults are even more likely to be asymptomatic. So, we need a way to monitor an entire population frequently, without being completely reliant on self-reporting and contact tracing.
Wastewater sampling enables this. It needs very little privacy infrastructure to be built because we cannot trace it back to any one individual and it helps detect people who are asymptomatic.
There is one caveat with monitoring SARS-CoV-2 in wastewater: it can be difficult to distinguish if the virus reported is from asymptomatic patients or from those who have recovered but are still shedding the virus. Studies are finding that on infection, the viral load increases first in the upper respiratory tract. Then, the day before or the day that someone develops symptoms, there is a higher viral load in the feces. This effect persists for up to six weeks after they recover. From my very first sample, I was detecting SARS-CoV-2 on campus in June when there were no known cases on campus because Connecticut was one of the early hotspots in the U.S. The important thing to consider when monitoring an outbreak in wastewater is looking at the trendline and not the absolute levels.
KM: The field has been evolving rapidly. The method that nearly everybody used in April to concentrate viruses out of wastewater was a polyethylene glycol (PEG) precipitation. They took between 250 mL and a half-liter of wastewater, added PEG, and then spun it. I only have access to one centrifuge that can take up to 250 mL and can only hold six samples at a time. That meant I was severely limited for time as I needed to spin my sample for hours.
The other classic and less troublesome method is to filter wastewater through negatively charged membranes. It would take about 30-45 minutes to filter 100 mL of wastewater treatment plant samples, but we were aiming to put upstream samplers across the campus to help isolate and test more cross-sections. Once you go upstream from the plant, the filtration time with this method increases massively: to filter 100 mL it would take three to four hours. Moreover, I can only put four filtration setups in my biosafety cabinet, which would again limit my sample processing speed.
KM: In early June, a leading Australian group published a great methods comparison paper on wastewater, where they mentioned filtration or PEG precipitation were not the only methods available. When I started looking into that, I came across Ceres. The idea of being able to just add a solution, let it incubate for 20 minutes, throw it on a magnet, and being done was amazing. It's all about saving time!
There is a small downside; it takes a couple more PCR cycles to detect SARS-CoV-2 in the samples that I processed with the Nanotrap particles versus the filter samples. This could be because either the Nanotrap particle method is not as efficient or because it concentrates only complete viruses (as opposed to viral bits).
So, I can either process dozens of samples with the Nanotrap particles, or I can process four with filtration. I’m happy to lose some detection power if that means I can process more samples and test more places. So, that's what I've adopted.
KM: It’s super easy. They recommend taking 50 mL, letting everything sit for 10 minutes, removing the top 40 mL, and then adding in the Nanotrap particles for incubation with a little bit of inversion. The biggest hurdle in using them is trying to find a magnet that works. I went online and found an industrial magnet that is only $16 or $17 and that clears the sample in 20 minutes.
KM: Befriend your facilities people; they have a huge amount of expertise and practical knowledge in your system. Seek other people out, ask lots of questions, and look for protocols. So many scientists are willing to help, at least in ecology. I could not have been up and running, producing data in a month, without the advice of all these people who have worked through these problems for years.
KM: The biggest thing is to make this accessible to as many researchers as possible. We are exploring partnerships, for example, with special needs schools where students have a hard time with mask compliance. Wastewater surveillance could help inform such schools in advance of an outbreak to help them implement alternative preventative measures.
We can perhaps start monitoring for flu or even STD outbreaks in the wastewater if we can add the corresponding primers and probes to the detection PCR. More awareness may be needed before such widespread monitoring is commonly adopted. However, once there are more universities with tools to monitor for outbreaks, there are so many applications for this!
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