The research, led by Marino Korlević of the Ruđer Bošković Institute in Rovinj, Croatia, together with colleagues from several Croatian marine research institutions, set out to answer a deceptively simple question: which microbes genuinely live on mussel gills, and which are merely passing through? This distinction matters because resident taxa may perform functions that benefit the host, such as contributing to nutrition or defense, while transient taxa simply reflect what the animal has recently filtered from the water column. Separating the two requires sampling that spans both space and time, which is precisely what the team undertook.

The researchers sampled mussels at two locations on the eastern coast of the Adriatic Sea, Lim Bay on the Istrian peninsula and Mali Ston Bay in Dalmatia, two of Croatia’s most important shellfish farming areas. Crucially, they did not stop at the mussels themselves. At the same time and at the same locations, they also characterized the microbial communities in the surrounding seawater and in the sediment. This three-way comparison is what gives the study its analytical power: if a microbe found on the gills is also abundant in seawater, it may simply be a hitchhiker; if it appears on the gills regardless of its abundance in the environment, it is more likely a true associate of the host.

To characterize the communities, the team used Illumina MiSeq sequencing of the V4 region of the 16S rRNA gene, the standard workhorse of modern microbiome research. This approach amplifies and reads a short, highly conserved stretch of bacterial DNA that allows researchers to identify which taxa are present and in what relative proportions. Sequences were grouped into operational taxonomic units, or OTUs, providing a fine-grained picture of community structure that could be compared across individuals, sites, and months.

The first major finding was that the gill microbiome is genuinely distinct. When the researchers compared the structure of the gill communities, measured at the OTU level, with the microbial communities of the surrounding seawater and sediment, the gills stood apart from both. This is not a trivial result. Mussels pump enormous volumes of water across their gills every day, and if the gill community were simply a snapshot of the planktonic bacteria in that water, the two communities would look statistically indistinguishable. The fact that they do not suggests that the gill environment actively selects for particular microbes, whether through the gill’s epithelial surfaces, its mucus chemistry, the immune defenses of the host, or the microconditions of oxygen and nutrients that prevail there.

Equally striking was what the comparison between the two bays showed, or rather did not show. The structure of the gill microbiome did not differ significantly between Lim Bay and Mali Ston Bay, despite the geographic separation and the environmental differences between the sites. In other words, whatever forces shape the mussel gill community, they appear to operate consistently across the Adriatic farming grounds studied. The gill microbiome, in this sense, behaves like a host-associated community rather than a local environmental artifact. What did differ was time: the gill communities showed clear differences between months, indicating temporal dynamics in their composition over the course of the year-long sampling program.

The taxonomic analysis then delivered the study’s central insight. Within the family Endozoicomonadaceae, a group of Gammaproteobacteria well known from associations with marine invertebrates, the researchers found members without known relatives at the genus level, so-called unclassified Endozoicomonadaceae, present in the gills of mussels from both locations and throughout the entire study period. This persistence across space and time marks them as resident members of the gill microbiome, microbes that maintain a stable association with the host rather than drifting in and out with the seasons. Their consistent presence hints at a functional role, though the study, based on 16S rRNA gene profiles, does not directly demonstrate what that role might be.

In contrast, other members of the same family behaved very differently. Endozoicomonas and the taxon provisionally known as Candidatus Endoecteinascidia appeared in the gill microbiome of mussels from both locations only in certain months, marking them as transient taxa whose presence ebbs and flows with the seasons. The picture became even more site-specific with the Vibrionaceae. Vibrio and unclassified members of the Vibrionaceae were also transient, appearing only in certain months, but with a telling distinction: they were specific to Lim Bay. That a well-known group of bacteria, which includes both harmless marine organisms and significant pathogens, showed up as a seasonal, location-limited component of the gill community is likely to attract attention from aquaculture researchers, since Vibrio dynamics in farmed bivalves are a persistent concern for the industry.

The conceptual framework the study reinforces is one that microbiome science has been converging on for years: host-associated communities are mixtures of core and casual members, and only long-term, repeated sampling can tell them apart. A single snapshot of a gill microbiome might have recorded the unclassified Endozoicomonadaceae and concluded they were the whole story, or it might have caught a Vibrio bloom and drawn alarming conclusions. By sampling monthly across a year and pairing the mussel data with seawater and sediment references, the Croatian team was able to show that the gill community combines a stable backbone with a shifting overlay of seasonal and local arrivals. The temporal dynamics they observed, with community composition differing between months even as the overall gill signature remained distinct from the environment, suggest that both the host and the changing conditions of the Adriatic play a role in shaping who lives on the gills.

For the aquaculture sector, the implications are practical as well as conceptual. Mytilus galloprovincialis is farmed intensively in the Mediterranean, and the health of farmed mussels is increasingly understood in terms of their microbial associations. Knowing that a stable, resident core of Endozoicomonadaceae persists on the gills across sites and seasons provides a baseline against which future perturbations, whether from warming waters, disease outbreaks, or changes in farming practice, can be measured. Knowing that Vibrio appears only transiently and only at one of the two studied bays provides a template for monitoring programs that distinguish routine seasonal variation from genuine warning signs. And methodologically, the study demonstrates the value of the three-compartment design, comparing host, water, and sediment simultaneously, as a way to separate true host association from environmental background. As microbiome research moves from description toward prediction, studies of this kind, grounded in a full year of observation at real farming sites, provide the empirical foundation on which that transition will depend. The gills of the Mediterranean mussel, it turns out, are not just a filter but a curated habitat, one whose permanent residents and temporary guests can now be told apart.

Subject of Research: Resident and transient bacterial taxa in the gill microbiome of the Mediterranean mussel Mytilus galloprovincialis in the Adriatic Sea

Article Title: Resident and Transient Microbial Taxa in the Gill Microbiome of the Mussel Mytilus galloprovincialis

Article References: Korlević, M., Markovski, M., Kapetanović, D., Vardić Smrzlić, I., Orlić, K., Bolotin, J., Kožul, V., Bobanović-Ćolić, S., Nerlović, V., & Perić, L. (2026). Resident and Transient Microbial Taxa in the Gill Microbiome of the Mussel Mytilus galloprovincialis. Microbial Ecology. https://doi.org/10.1007/s00248-026-02888-y

Image Credits: AI Generated

DOI: 10.1007/s00248-026-02888-y

Keywords: gill microbiome, Mytilus galloprovincialis, Mediterranean mussel, Endozoicomonadaceae, Vibrionaceae, 16S rRNA gene sequencing, Adriatic Sea, aquaculture, marine microbiology, temporal dynamics, Gammaproteobacteria, microbial ecology

The research, led by Teresa Bednarek of the University of Western Australia and the Australian Institute of Marine Science, focused on two closely related corymbose corals, Acropora millepora and Acropora digitifera. Corals are not solitary organisms but holobionts, integrated ecological units comprising the animal host, its photosynthetic dinoflagellate symbionts of the family Symbiodiniaceae, and a diverse community of bacteria, archaea and fungi. The algal partners supply the bulk of the coral’s energy through photosynthesis, while the bacterial associates contribute to nutrient cycling, immune function and metabolite exchange. Understanding how external partners such as resident fish perturb this internal ecosystem is a frontier in coral biology, particularly as reefs face intensifying thermal stress.

In May 2023, the team sampled 155 healthy adult colonies across two sites near Coral Bay, at depths of two to five metres. Before collecting tissue, divers conducted visual censuses of every damselfish swimming between the branches or hovering immediately above each colony, recording species, abundance and estimated body length. Five damselfish species were documented, including Dascyllus reticulatus, Dascyllus aruanus, Dascyllus trimaculatus, Pomacentrus moluccensis and Pomacentrus coelestis. Branch tips were snapped from the centre of each colony, flash-frozen in liquid nitrogen and later processed for symbiont cell counts, chlorophyll quantification and DNA extraction from the intact coral fragment, skeleton included.

The molecular work relied on amplicon sequencing of two genetic markers. The internal transcribed spacer 2 region, or ITS2, was used to characterise the Symbiodiniaceae communities, while the V5-V6 hypervariable regions of the 16S rRNA gene profiled the bacterial and archaeal assemblages. Sequences were denoised into amplicon sequence variants using the DADA2 pipeline, taxonomy was assigned against the SILVA and GeoSymbio reference databases, and the LULU algorithm was applied to curate the highly variable ITS2 variants. Statistical models then tested whether damselfish presence, abundance or biomass explained variation in symbiont density, pigmentation and community composition, while controlling for colony size and sampling site.

The first major finding was that host identity, not fish residency, is the dominant force shaping the holobiont. A. millepora carried roughly twice the symbiont cell density of A. digitifera, at 2.36 million cells per square centimetre versus 1.25 million. Chlorophyll a per symbiont cell was similar between species, but A. digitifera packed significantly more of the accessory pigment chlorophyll c2 into each algal cell, suggesting a distinct light-harvesting strategy that extends absorption into spectral ranges where chlorophyll a performs poorly. Because chlorophyll c2 is among the first pigments lost during thermal stress, this baseline difference could influence how each species approaches bleaching thresholds.

Both corals were dominated by symbionts of the genus Cladocopium, yet at finer resolution their assemblages diverged sharply. A. millepora hosted predominantly the C21 type, which averaged 91.67 percent of its symbiont community, while A. digitifera was dominated by C3k at 85.23 percent. Bacterial communities told a parallel story. Both species were overwhelmingly dominated by members of the family Endozoicomonadaceae, bacteria often regarded as markers of coral health, but the specific strains differed. A. millepora harboured a richer and more even bacterial community, with Endozoicomonadaceae comprising 83.48 percent of its microbiome, whereas A. digitifera’s community was more narrowly concentrated, with the family accounting for 95.27 percent. Coral species alone explained nearly a quarter of the variation in bacterial community composition across all colonies.

The damselfish themselves were not evenly distributed. A. millepora colonies hosted significantly more fish than A. digitifera, averaging 11 individuals per occupied colony compared with 6, although total fish biomass was similar between the two coral species. The assemblage composition also differed, with D. reticulatus dominating on A. millepora at 73 percent of recorded individuals, while D. aruanus and P. moluccensis were proportionally more common on A. digitifera. In total, 573 damselfish were counted on A. millepora colonies and 168 on A. digitifera, underscoring how strongly fish settlement preferences track coral architecture and species identity.

When the researchers tested whether fish occupancy altered the algal symbionts, the answer was largely no. Symbiont density, chlorophyll a content and overall symbiont community composition remained essentially unaffected by damselfish presence, abundance or biomass in both coral species. The one exception was chlorophyll c2 per cell, which responded in opposite directions: increasing fish numbers were associated with higher chlorophyll c2 in A. digitifera but lower chlorophyll c2 in A. millepora. The authors suggest this divergence may reflect photoacclimation to improved light availability, as damselfish remove sediment and algae from their host colonies, or nutrient-stimulated pigment production, with the direction of the response shaped by the different symbiont types each coral harbours.

The bacterial communities, by contrast, told a more nuanced story. In A. millepora, colonies hosting damselfish showed significantly lower Shannon diversity, and both diversity and evenness declined steadily as the number of resident fish increased. Remarkably, the interspecies gap in bacterial diversity that distinguished A. millepora from A. digitifera in fish-free colonies disappeared entirely in fish-occupied ones, as though the damselfish pulled A. millepora’s microbiome down to levels comparable with its neighbour. Crucially, bacterial richness was unaffected, meaning the fish did not eliminate taxa but rather shifted their relative abundances, allowing certain bacteria to dominate. Damselfish-occupied colonies also showed significantly lower community heterogeneity, a pattern that contrasts with the erratic, high-variability microbiomes typically associated with dysbiosis under stress.

The authors propose that fish-derived nutrients, primarily ammonium and phosphate excreted directly onto the colony, act as a consistent selective pressure favouring particular bacterial taxa, while the physical agitation of water within the branches and direct microbial transfer via contact or waste deposition may further reshape the local microbial environment. Notably, Terasakiellaceae, a bacterial family associated with nitrogen fixation in coral tissue, was weakly enriched in fish-occupied A. millepora colonies, consistent with nutrient-enriched conditions. In A. digitifera, where fish were fewer and the assemblage composition differed, bacterial communities remained essentially untouched, and the authors caution that host morphology or the species’ inherently lower bacterial diversity may also contribute to its apparent resistance.

The study offers an initial snapshot rather than a definitive causal account, and the authors are explicit about its limits. The observations were made under non-stressful autumn conditions, and whether the damselfish-associated microbial shifts confer resilience benefits under thermal stress remains a critical open question. Long-term monitoring would clarify whether these community changes persist as fish populations turn over through recruitment and mortality, and experimental manipulations using clonal coral genotypes would help isolate mechanisms from the variability inherent in observational work. Still, the message is clear: the coral holobiont is not an isolated system, and the animals that live in and around a colony are active participants in its microbial architecture. For reef conservation and restoration programmes, the implication is that transplant success may depend not only on choosing the right coral species but on understanding the fish tenants that come with it.

Subject of Research: Species-specific effects of resident damselfish on the coral holobiont microbiome of Acropora corals

Article Title: Resident damselfish influence the coral holobiont: species-specific responses in Acropora millepora and Acropora digitifera

Article References: Bednarek, T., Brooker, R. M., Martin, B. C., Gleeson, D. B., & Thomas, L. (2026). Resident damselfish influence the coral holobiont: species-specific responses in Acropora millepora and Acropora digitifera. Coral Reefs. https://doi.org/10.1007/s00338-026-02938-4

Image Credits: AI Generated

DOI: 10.1007/s00338-026-02938-4

Keywords: coral holobiont, damselfish, Acropora millepora, Acropora digitifera, Symbiodiniaceae, Cladocopium, Endozoicomonadaceae, microbiome, Ningaloo Reef, coral reefs, symbiosis, 16S rRNA sequencing