There’s Something in the Water: Accurately Identifying Freshwater Microbial Communities

By Caroline Reisiger

8 December 2022


A biologist collects samples from a freshwater stream.


Across the globe, species are declining at an alarming rate. One way that biologists can help protect species is through ecological assessments, which help identify and inventory the species present in a particular ecosystem. Such inventories are no small undertaking, however, and biologists usually end up leaving microbes out of the picture, due to the difficulty involved in identifying them.

But including microbes in ecological assessments could dramatically improve how we evaluate ecosystem health, says Christopher Hempel, a PhD student in the Department of Integrative Biology who studies the diversity of freshwater systems.

Microbial diversity gives you a really good idea of the state of a specific environment; it can let you know whether an ecosystem is healthy or if it needs restoration or environmental protection.”

Currently, the most common way to survey microbial diversity in an ecosystem is through DNA-based identification. However, once DNA samples have been collected, there are many different processing and analysis strategies. To date, no one has figured out a perfect strategy for using and analyzing this DNA to determine microbial diversity as part of a freshwater ecological assessment. To add to the challenge, new strategies are developed continuously, including some that even go beyond DNA and also incorporate RNA analysis.

To improve current strategies, Hempel — working with Dr. Dirk Steinke, an adjunct professor of molecular ecology and evolution — compared two uncommon but promising methods of microbial identification: metagenomics and total RNA sequencing.

“These two techniques are cutting edge and it is important to figure out which one works best in freshwater environments to identify microbes”, Hempel says.

Metagenomics is a method that looks at all the DNA in a sample. This approach gives a broad picture of the biodiversity present but can miss species that are low in abundance. In addition, 40-90% of the DNA present in a water system at any time is from non-living organisms, so it is difficult to get a real-time picture of the species that currently live in an ecosystem.

Alternatively, total RNA sequencing mostly looks at ribosomal RNA, a type of RNA that is most abundant when microbes are alive and growing and works particularly well to identify microbes. This method can thus give a more accurate picture of the current microbial diversity of a system.

Over three years, Hempel compared these DNA and RNA-based methods using a known “mock” microbial community, so that their level of accuracy could be properly compared.  He also looked at over 600 ways of processing the resulting data by computer for analysis, with the goal of determining which combination of methods worked best.

Overall, total RNA sequencing emerged as the preferable choice. Not only does it more accurately detect microbial species, it also requires less sequencing data, making this method much more cost-effective.

The study is an exciting glimpse of the potential of microbes to help shape how we assess and monitor ecosystem health.

“Interest in microbial diversity and the importance of microbial communities for environmental research is just starting to get broader recognition,” says Hempel.

While more research is needed to confirm the findings using field-collected samples, the study is a first step towards providing researchers across the globe with an accurate and efficient method to include microbial diversity in ecological assessments.

This study was funded by the Canada First Research Excellence Fund-Food from Thought project at the University of Guelph.


Read the full study in the journal Nucleic Acids Research.

Read about other CBS Research Highlights.