Collect What You Need, Preserve What Matters

You Can't Study What You Can't Save Intro

There's an old joke about a heart surgeon and a car mechanic. The mechanic asks, "Why do you get paid so much more than I do? We both fix complex machines." The surgeon replies, "Yes—but I have to work on mine while it's still running."

The punchline serves as a reminder of how much complexity is added when working on a dynamic, living system. And this principle is just as relevant in microbiome research as it is in medicine. When you collect a sample—be it soil, feces, tissue, or wastewater—you're capturing a dynamic, living community. The microbes are active, adapting, growing, dying, and interacting, and unlike a heart, these microbial systems don't stop ticking just because they've been scooped into a tube.

Even after the microbes themselves are no longer alive, the enzymes they produced—DNases, RNases, proteases—are often still very much active. These enzymes continue breaking down nucleic acids and other biomolecules in the sample. Worse yet, this degradation doesn't happen uniformly. Depending upon storage conditions, some organisms' DNA or RNA degrades faster than others, skewing the apparent makeup of the community. Additionally, if the microbes are not frozen or killed immediately, the change in conditions will start favoring certain microbes to "bloom" and others to stop growing entirely or start dying. Those that bloom often do so by consuming those that died when conditions changed. A sample that isn't properly preserved doesn't just lose data—it creates false data.

This is why microbiome sample preservation isn't just a technical detail—it's a foundational requirement and why every good microbiome pipeline must begin with the basics of sample collection.

The Fragility of Microbial Signatures Effects of Degradation

Microbial communities are incredibly dynamic. Even minor environmental changes can start shifting the structure of a sample within minutes. Some microbes bloom rapidly, others degrade, and many begin releasing enzymes that break down genetic material.

This degradation introduces a unique kind of error—*bias*. DNA and RNA from more fragile organisms may vanish quickly, leaving a skewed microbial fingerprint that looks trustworthy but is biologically misleading. If the likely patterns created by collection and preservation bias sound like the exact opposite of lysis bias, it is because they generally are. Combine collection/preservation bias selecting for tougher organisms with lysis bias selecting against these tougher organisms and the consistency and accuracy of your results will be anybody's guess.

Preservation isn't about stopping decay eventually—it's about freezing that community in time, right at the moment of collection.

How Samples Fall Apart: Freeze/Thaw and Degradation in Action Causes of Degradation

Freezing may seem like a good way to preserve samples. While frozen, the sample will maintain its integrity over the long term, but the inevitable thaw will quickly destroy that integrity unless appropriate measures are taken. The freezing process will cause cells to rupture, with physically weaker cells (often gram-negative) lysing at a higher rate. All of the spilled cell contents containing degradative enzymes and nucleic acids will be protected by the cold, so no sample integrity is lost at this point. When frozen samples thaw—even briefly—enzymes reactivate, and nucleic acids degrade. This sets off a cascade of degradation that disproportionately affects more fragile microbes. Repeat this process several times and major taxa such as Bacteroidetes can disappear.

While using extreme cold as a preservative suffers from its deficiencies, chemical preservatives inactivate enzymes and halt biological activity at the source.

Zymo Research's DNA/RNA Shield™, for example, stabilizes nucleic acids immediately—even in challenging matrices like feces or wastewater—and works at room temperature, making it ideal for field or clinical settings. Additionally, unlike other chemical preservatives, DNA/RNA Shield™ can be put through the lysis and extraction process, rather needing to be discarded before lysis and extraction due to chemical incompatibilities with the downstream efforts. Chemical preservatives are all lytic to some extent, and microbes that lyse in a preservative that must later be discarded are at high risk of being discarded with the preservative waste.

Case Study: When Preservation Fails, E. coli Takes Over Case Study

In a high-profile publication, researchers found Escherichia coli and other gammaproteobacteria being over-represented in many human stool samples. The culprit? Samples were shipped without stabilization. During transit, E. coli bloomed, outcompeting the native community.

The microbial distortion was so severe, the researchers had to develop a technique to bioinformatically exclude E. coli from the analysis (Song et al., mSystems 2016).

This example underscores that cold shipping alone can't prevent bias. Without immediate stabilization, fast-growing organisms can quickly overwhelm the sample's original makeup, often consuming other organisms to fuel this growth.

Best Practices for Preserving Microbial Truth at the Point of Collection Best Practices

Two people in immersion suits walking in Antarctic ice.

 

Preserve immediately: Don't wait to stabilize your sample.
Avoid freeze-thaw cycles: Damage is cumulative and selective.
Use preservatives validated for both DNA and RNA: Compatibility matters.
Match your kit to your matrix: Feces, soil, and wastewater all pose unique challenges.
Plan for field conditions: Room-temperature preservatives simplify logistics.

Zymo's DNA/RNA Shield™ and collection kits are built to address these challenges—stopping degradation, inactivating pathogens, and preserving nucleic acids without a cold chain.

Making Sample Collection Easy and Error-free Workflow Optimization

While the ideas of "easy" and "sample collection" do not always go together, considering the ease of use and simplicity of collection instruments can be easily missed when developing a pipeline. Zymo Research offers a variety of sample collection systems utilizing our DNA/RNA Shield™ preservative to facilitate sample collection for both scientists and at-home collection for research subjects. Examples of this include the Bunny Wipe™ to simplify at-home fecal sample collection with no training required, SafeCollect™ collection devices to prevent spills by keeping the DNA/RNA Shield™ sequestered until after sealing of the sample collection device and facilitate self-collection, and systems optimized for room-temperature shipping that can facilitate a cost-effective world-wide sample chain.

Additionally, some products such as our DNA/RNA Shield Fecal Collection Tubes with built in scoops and the Bunny Wipe™ have systems in place to prevent a problem that is common with fecal sample collection: overwhelming the preservative volume with excess sample material. Both of these systems not only facilitate collection of fecal samples in a manner that is more appealing to both scientists and research subjects, but also make it easy to ensure that an appropriate mass of sample is added to the preservative and that it does not get overloaded, thus risking a compromise of sample integrity.

Preserving the Moment That Matters Conclusion

Every microbial sample is a moment frozen in time. But without preservation, that moment degrades—quietly, invisibly, and irreversibly.

Preserving the truth of that sample at the time of collection is critical. Whether you're sampling from a patient, a river, or a wastewater plant, Zymo Research offers tools to help you protect what matters most: the biological truth of your sample.

Because good data starts with good collection.

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