Hannah Nowers — Cracking the case of the wasting sea stars 🌟

Sea star wasting disease has wiped out 20+ species from Alaska to Baja, melting animals into goo. The obvious hypothesis: maybe the susceptible species just don’t have enough immune genes. Hannah put that idea to the test, combing through annotated sea star genomes and sorting immune genes into functional categories. The plot twist? Gene count isn’t the answer. Susceptible and resistant species carry similar immune gene arsenals — so the real story is about how those genes are switched on and off, not how many there are. One clean result, and suddenly the whole field is asking a better question.

Download Hannah’s capstone presentation (PDF) · Read Hannah’s capstone paper (PDF) · GitHub repo

Noah Ozguner — Do oysters get stressed when you shake them? 🦪

Oyster farmers “tumble” their oysters to sculpt prettier shells — but is it secretly stressing the animals out? Noah grabbed two sizes of oyster seed, gave them a 20-minute shake, and tracked their metabolism for a week with a color-changing assay. Tiny oysters? Totally unbothered. But the big ones threw a delayed metabolic curveball — no reaction at first, then a dramatic spike a full week later (p = 0.0006). Turns out bigger oysters carry a hidden energetic hangover, and it shows up on a timer.

Download Noah’s poster (PDF)

Hazel Abrahamson-Amerine — What a heatwave does to an oyster’s fuel tank 🔥

Oysters bank energy as glycogen — their cellular savings account. Hazel asked what happens to that account during a marine heatwave. Using a glow-in-the-dark glycogen assay, they cranked oysters to 32 °C for 48 hours and watched the reserves crash. The hopeful part: give them a break from the heat, and they slowly rebuild the stash. Recovery is real — it just takes time. Next up: where in the body it happens, and whether oysters toughen up for round two.

Download Hazel’s presentation (PDF)

Christina Zhang — Can oysters be trained to take the heat? 🌡️

If a heatwave hits twice, does surviving the first one help with the second? Christina tested whether oysters can be “primed” — toughened up by an earlier non-lethal heat exposure. She gave Pacific oyster seed different thermal histories (cool or hot, then cool or hot again) and read out their metabolism across a temperature gradient. The verdict: priming changes the game. Below 35 °C, primed oysters ran a calmer, lower metabolic response — a sign of tolerance — while unprimed oysters spiked. But push past 37 °C and the stress response overrides any priming benefit, and mortality climbs. Oysters can learn to take the heat, but only up to a point.

Download Christina’s poster (PDF)

Four students, four big swings, four home runs. Congrats Hannah, Noah, Hazel, and Christina — the lab is lucky to have you. 🎉

Defense Recording

Why Standard Tools Fall Short

If you work in genomics, you are probably familiar with tools like DESeq2 and edgeR for differential gene expression analysis. These are workhorses of the field, but they are built for pairwise comparisons: treatment vs. control, timepoint A vs. timepoint B. Our coral data has a richer structure — expression measured across multiple genes, multiple species, and multiple samples over time — that these tools cannot natively capture.

We needed methods that could discover coordinated expression patterns spanning all three of those dimensions simultaneously.

Barnacle: Sparse Tensor Decomposition for Multi-Species Expression

Enter Barnacle, a sparse tensor decomposition tool originally developed for marine metatranscriptomics. The idea is elegant: instead of flattening your data into a two-dimensional matrix, you represent it as a 3D tensor (Gene x Taxon x Sample) and decompose it into a small number of sparse components. Each component captures a distinct co-expression program — a group of genes that behave similarly across species and conditions.

The critical tuning decisions are:

• How many components (rank)? Too few and you miss real signals; too many and you start fitting noise. We use a cross-validation strategy — fit on half the data, test reconstruction on the held-back half — to find the sweet spot.
• How sparse? Sparsity controls which genes are assigned to each component. We use split-half reproducibility: if both halves of a random split identify the same core genes for a component, the sparsity threshold is keeping real members.

Applied to our E5 coral timeseries data, Barnacle revealed biologically meaningful components. One component, for example, aligned closely with known calcification genes — a result that would be difficult to extract from traditional pairwise analyses.

With these components in hand, we now have groups of co-regulated genes that we can interrogate for epigenetic regulatory patterns and use to test hypotheses about how environmental stress propagates through molecular networks.

The Epigenetic Layer: More Than One Regulator

Epigenetics — heritable changes in gene activity that do not alter the DNA sequence itself — is central to our project. In marine invertebrates, the epigenetic landscape includes:

• DNA methylation — In contrast to mammals, marine invertebrate methylation is mosaic and concentrated in gene bodies rather than promoters.
• Long non-coding RNA (lncRNA) — Regulatory RNA molecules numbering in the tens of thousands.
• MicroRNA (miRNA) — Short regulatory RNAs that target messenger RNA for degradation or translational repression.

Each of these layers can influence gene expression, and they likely interact with each other. Simple pairwise correlations between a single epigenetic mark and expression only tell part of the story. We wanted a model that could integrate all three layers and identify which features — and which combinations — actually drive expression.

Elastic Net Regression: Predicting Expression from Epigenetics

This is where Elastic Net regression comes in. Elastic Net is a regularized regression method that blends two complementary penalties:

• Lasso (the “harsh editor”) drives coefficients to exactly zero, performing feature selection by removing irrelevant predictors.
• Ridge (the “compromiser”) shrinks correlated predictors together rather than arbitrarily choosing one, handling multicollinearity gracefully.

Our predictor space is formidable: roughly 50 miRNAs, 10,000 lncRNAs, and 20,000 gene-body methylation features, all predicting expression across approximately 40 samples per species. This is a classic p >> n problem, and Elastic Net is well suited for it.

We trained models using 80/20 train/test splits repeated across 50 replicates, validated predictions using noise injection (if the model is real, adding noise to predictors should degrade performance), and applied stability selection (Faletto and Bien 2022) to focus on features that are consistently selected across resampled datasets rather than artifacts of any single split.

Video

What We Found

The results are striking and represent, to our knowledge, the first integrated picture of multi-layer epigenetic regulation of gene expression in a marine invertebrate system:

• Per-gene regulatory profiles reveal that individual genes differ substantially in which epigenetic features predict their expression. Some are primarily methylation-driven, others lncRNA-driven, and many are regulated by a combination.
• High-level type compositions show systematic patterns in how regulatory strategies are distributed across the genome.
• Regulatory network maps capture how epigenetic features interact, suggesting coordinated regulatory programs rather than independent, isolated effects.

Looking Ahead

This work is a starting point. The tensor decomposition framework can be extended to incorporate additional data types (e.g., lncRNA expression, genes found in fewer than all species), and the elastic net models can be refined as more samples and species are added. The broader goal — understanding how environmental stress rewires epigenetic regulation and whether those changes can be inherited — remains one of the most important questions in coral biology and climate adaptation.

We are grateful to the eScience Incubator Program and particularly to Vaughn Iverson for pushing us to think about our data in new ways. The combination of domain expertise in marine biology with data science methods has opened analytical doors that neither community would have found alone.

For more information, visit the E5 project and Barnacle documentation.

Training oyster parents’ immune systems

In this study, adult Pacific oysters were briefly exposed to Poly(I:C), a harmless synthetic molecule that mimics viral infection and activates the oyster immune system. This kind of immune priming has been shown to increase disease resistance in oysters—but here we asked a different question:

Does immune priming in parents change how their offspring handle heat and metabolic stress?

After the exposure, oysters were spawned, and their larvae were reared in commercial hatchery conditions and then grown into seed oysters over several months. These offspring were later tested for growth, survival under heat stress, and metabolic performance.

Faster growth in primed families

The first surprise was growth.

Although offspring from primed parents were slightly smaller early on, they grew faster over time and ended up larger than controls by 236 days post-fertilization. In other words, immune priming in parents altered how energy was allocated during development—suggesting a shift in developmental strategy.

This matters for hatcheries, where faster-growing seed often perform better during nursery and grow-out phases.

Heat survival depends on how hot it gets

Next, the team exposed oysters to temperatures ranging from normal conditions to extreme heat (up to 45°C) and measured survival.

The results were strikingly temperature-specific:

• At 40°C, offspring from primed parents were 35% more likely to survive than controls 

• At 42°C, however, those same primed offspring were more likely to die than controls 

This shows that immune priming doesn’t simply make oysters “tougher”—it reshapes where their thermal limits lie. In a warming ocean, that nuance matters.

Metabolism tells the story

Why did primed offspring survive better at 40°C but worse at 42°C? The answer appears to lie in their metabolism.

Using a high-throughput resazurin assay, the team measured how quickly oysters processed energy under different temperatures  .

They found:

• At 36°C, primed offspring had 43% higher metabolic activity than controls

• At 40°C, primed offspring showed lower metabolic activity than controls 

This pattern suggests that immune priming increases metabolic flexibility—the ability to ramp metabolism up under moderate stress and suppress it when conditions become extreme. That flexibility is likely what allows primed oysters to survive better at 40°C by entering a kind of protective metabolic slowdown, a known survival strategy in stressed marine invertebrates.

A form of environmental memory

These effects persisted for more than seven months after fertilization, long after early maternal provisioning should have faded. This points to deeper mechanisms of environmental memory, potentially involving:

• altered maternal lipid and energy provisioning

• immune-related transcripts passed into eggs

• changes in microbiomes

• epigenetic regulation (DNA methylation, non-coding RNAs) 

Together, these mechanisms may allow oysters to transmit information about parental stress environments directly to the next generation.

What does this mean for aquaculture?

This research opens the door to a new idea:

Hatcheries could condition broodstock before spawning to improve offspring performance.

Because Poly(I:C) can be delivered by simple immersion, it could be integrated into existing hatchery workflows as a scalable, non-invasive treatment. When paired with selective breeding, immune priming might help produce oyster seed that is more resilient to heatwaves and disease—two of the biggest threats to modern shellfish farming.

At the same time, the study makes clear that priming is not a magic bullet. Benefits depended strongly on temperature, and extreme heat erased them. Field trials and longer-term studies are needed before these tools can be used widely.

Why this matters

As oceans warm and pathogens spread, oysters are being pushed toward their physiological limits. This study shows that parents matter—and that immune experiences in one generation can shape how the next generation grows, survives, and uses energy.

That’s environmental memory in action.

And it may become one of our most powerful tools for building climate-resilient aquaculture in the decades ahead.

🌊 A New Colorful Window into Oyster Metabolism

Resazurin, a redox-sensitive dye used for decades in biomedical and microbial assays, shifts from blue to pink as it’s reduced by metabolic activity. Huffmyer et al. adapted this dye-based reaction — familiar to many cell biologists — to whole oysters, showing that fluorescence intensity closely tracks oxygen consumption and reflects metabolic performance at the organismal level .

The team developed a standardized, scalable protocol using multi-well plates and fluorometric readings to quantify metabolic rate across hundreds of individuals at once. This makes it one of the first high-throughput metabolic assays for live shellfish, an enormous advance for aquaculture and climate resilience research.

You can explore their detailed methods and ongoing assay development at the Roberts Lab Resazurin Assay Development site.

🔥 Metabolism Under Pressure: What the Color Shift Reveals

By exposing oysters (Crassostrea gigas and C. virginica) to temperature gradients and acute heat shocks, the researchers observed predictable “thermal performance curves” — metabolism peaked near 36 °C before declining sharply at higher temperatures. Notably, oysters that suppressed metabolism under extreme stress were more likely to survive, suggesting that metabolic depression is a hallmark of thermal resilience.

The assay also captured family-level differences in metabolic response, indicating that some oyster lineages are genetically predisposed to handle stress better than others. In a large-scale breeding experiment at the Virginia Institute of Marine Science, metabolic rates measured by resazurin correlated with predicted survival performance in selectively bred oyster families .

🧬 Why This Matters

Metabolic rate is one of the most fundamental indicators of animal health and stress, but it’s historically been difficult and time-consuming to measure in shellfish. The resazurin assay overcomes that bottleneck, offering:

• High-throughput capability: hundreds of oysters per run

• Low cost and minimal equipment (a simple plate reader suffices)

• Compatibility with live animals — no lethal sampling required

• Potential for integration with breeding and monitoring programs

These advantages make it a game-changer for aquaculture, ecology, and conservation, enabling rapid screening of stress resilience across populations and environments.

🌍 Toward Smarter, More Resilient Aquaculture

As marine ecosystems face intensifying heatwaves and acidification, tools like the resazurin assay can help hatcheries, farmers, and researchers identify the oysters most likely to thrive under stress. The study’s message is clear: color can tell a powerful physiological story.

Blue means rest. Pink means life. And somewhere between those hues lies the key to building a more resilient future for oysters — and the people who depend on them.

📘 Citation

Huffmyer, A. S., Ozguner, N., Baird, M., Elvrum, C., Kounellas, C., Dicksion, D., White, S. J., Plough, L., Gavery, M. R., Krebs, N., Walton, W., Small, J., Pitsenbarger, M., Ealy-Whitfield, H., & Roberts, S. (2025). From Blue to Pink: Resazurin as a High-Throughput Proxy for Metabolic Rate in Oysters. bioRxiv. https://doi.org/10.1101/2025.11.06.686367