# Copepod-associated microbial biogeography in the epipelagic ocean

Ximena Velasquez, Tal Ozer, Maria Grazia Mazzocchi, François Carlotti, Dalit Meron, Mireille Pujo-Pay, Pascal Conan, Xavier Durrieu de Madron, Tamar Guy-haim

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Ximena Velasquez, Tal Ozer, Maria Grazia Mazzocchi, François Carlotti, Dalit Meron, et al. Copepod-associated microbial biogeography in the epipelagic ocean. Limnology and Oceanography Letters, 2025, 10.1002/lol2.70054. hal-05312289

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*Limnology and Oceanography Letters* 2025

© 2025 The Author(s). *Limnology and Oceanography Letters* published by Wiley Periodicals LLC on behalf of Association for the Sciences of Limnology and Oceanography. doi: 10.1002/loI2.70054

LETTER

Copepod-associated microbial biogeography in the epipelagic ocean

Ximena Velasquez ID, ¹,²* Tal Ozer,¹ Maria Grazia Mazzocchi,³ Francois Carlotti,⁴ Dalit Meron,²,⁵

Mireille Pujo-Pay,⁶ Pascal Conan,⁶,⁷ Xavier Durrieu de Madron,⁸ Tamar Guy-Haim ID ¹,⁹*

¹National Institute of Oceanography, Israel Oceanographic and Limnological Research (IOLR), Haifa, Israel; ²Department of Marine Biology, Leon H. Charney School of Marine Sciences, University of Haifa, Haifa, Israel; ³Integrative Marine Ecology Department, Stazione Zoologica Anton Dohrn, Naples, Italy; ⁴Université Aix-Marseille, Université de Toulon, CNRS, IRD, Mediterranean Institute of Oceanography (MIO), Marseille, France; ⁵Morris Kahn Marine Research Station, University of Haifa, Sdot-Yam, Israel; ⁶LOMIC, CNRS, Sorbonne Université, Banyuls/Mer, France; ⁷OSU STAMAR, CNRS-Sorbonne Université, Paris, France; ⁸CEFREM, CNRS-Université de Perpignan Via Domitia, Perpignan, France; ⁹Department of Life Sciences, Ben-Gurion University of the Negev, Beer Sheva, Israel

**Scientific Significance Statement**

Copepods support diverse microbial life that differs from that found free-living in seawater. Over broad spatial scales, coexisting copepods may share their microbes, forming a microbial metacommunity. However, the mechanisms shaping the spatial distribution of these host-associated microbial communities remain poorly understood. Identifying the ecological processes that govern these associations is essential for understanding their biogeography and how it relates to oceanographic patterns. Our study shows that copepod-associated microbial metacommunities more closely reflect environmental gradients and current circulation in the Mediterranean Sea than seawater microbial metacommunities, suggesting their potential as indicators of microbial biogeographic patterns and ecosystem change.

**Abstract**

Zooplankton-microbial interactions play crucial roles in epipelagic ecosystem functions. The distinct west-to-east gradients and complex circulation patterns in the Mediterranean Sea, combined with the ubiquity of pelagic copepods, provide an ideal model to study the ecological processes driving host-associated microbial spatial distribution. Here, we characterized and compared the copepod-associated microbial metacommunities (CAMC) with those from seawater microbial metacommunities (SMC). Copepod-associated microbial metacommunities displayed spatial dissimilarity between the western and eastern basins, while SMC exhibited similar microbial compositions. The within-basin similarity observed in CAMC was associated with connectivity by the surface currents. Ecological drift explained most of CAMC variability, likely as a response to the restricted

This is an open access article under the terms of the [Creative Commons Attribution License](https://creativecommons.org/licenses/by/4.0/), which permits use, distribution and reproduction in any medium, provided the original work is properly cited.

**Associate editor:** Mar Benavides

**Data Availability Statement:** Demultiplexed V4-16S rRNA amplicon reads and associated metadata are deposited in the SRA database under the BioProject accession number PRJNA1240050, https://www.ncbi.nlm.nih.gov/bioproject/?term=PRJNA1240050. Copepod barcoding data are available in NCBI Genbank under accession numbers PV274503.1-PV274530.1, PV282463.1-PV282506.1. Environmental data and metadata are available in the Zenodo repository at https://doi.org/10.5281/zenodo.15981975. The full reproducible code for the statistical analyses and bioinformatic pipeline is available at https://github.com/ximvel/Copepod-associated_microbial_metacommunities_CAMC.

<footnote>*Correspondence: ximvel@ocean.org.il; tamar.guy-haim@ocean.org.il</footnote>
co-dispersal of the hosts with their microbes, which presented low prevalence and abundance. Seawater micro-
bial metacommunities displayed higher homogenizing dispersal, with widely distributed generalist taxa. We
conclude that CAMC better reflect cross-basin gradients and connectivity patterns than SMC, suggesting that
CAMC may serve as a useful proxy for studying microbial biogeography.

Copepods are the most numerous metazoans in the ocean (Verity and Smetacek 1996), playing a central role in the pelagic trophic webs and the biological pump (Buitenhuis et al. 2006; Steinberg and Landry 2017). They harbor diverse microbial communities acquired horizontally from the surrounding waters and the food, or vertically—from the mother to the offspring (Tang 2005; Tang et al. 2010). At local scales, copepod microbial communities are host-specific and strongly influenced by host biological traits such as diet and feeding behavior (Shoemaker and Moisander 2015, 2017; Velasquez et al. 2023). However, over large spatial scales, coexisting copepods may share their microbes through direct host–host interactions or indirectly via the external environment, forming a microbial metacommunity (Adair and Douglas 2017; Araujo et al. 2024).

The composition and distribution of microbial meta-
communities depend not only on microbial dispersal
through ocean currents or host distribution, but also on eco-
logical drift and environmental filtering (i.e., selection)
(Nemergut et al. 2013). Drift refers to random changes
(e.g., mortality) in the relative abundances of microbial taxa
within a metacommunity, while selection involves abiotic
factors that shape metacommunity structure (Zhou and
Ning 2017). These ecological processes strongly interact, often
leading to a decline in microbial similarity with increasing geo-
graphical distance (Villarino et al. 2022). Copepods have rela-
tively limited dispersal due to restricted connectivity by currents
or environmental barriers (Sommeria-Klein et al. 2021), which
may favor the proliferation of specialist (rare) and opportunist
microbial taxa, leading to stronger distance-decay relationships
(Clark et al. 2021; Dickey et al. 2021; Troussellier et al. 2017). In
contrast, free-living microbial communities tend to more widely
disperse and are dominated by abundant, generalist taxa (Ruiz-
González et al. 2019).

The Mediterranean Sea provides an excellent environment to study copepod microbial biogeography. It has unique oceanographic features, with a pronounced west-to-east gradient of decreasing nutrient concentration and plankton biomass and increasing sea surface salinity and temperature (Pujo-Pay et al. 2011; Siokou-Frangou et al. 2010; Tanhua et al. 2013). Furthermore, the Mediterranean Sea exhibits complex surface circulation patterns and mesoscale features (Berline et al. 2021; Bouzaiene et al. 2020), which can act as a corridor or barrier for plankton dispersal (McManus and Woodson 2012; Sommeria-Klein et al. 2021; Villarino et al. 2018), with their associated microbes (Mony et al. 2022). Copepods benefit opportunistic microbes that use them as a source of nutrients in less favorable environments, as well as a

dispersal vector, allowing the maintenance of ecosystem pro-
cesses at a large spatial scale (Troussellier et al. 2017).

Copepods were previously used as a proxy for
mesozooplankton biogeographical patterns in the Mediterra-
nean Sea (Acha et al. 2020; Nowaczyk et al. 2011). Thus, we
hypothesized that the distinctive environmental factors and
circulation patterns of the western vs. eastern basin in the
Mediterranean Sea would result in greater horizontal spatial
variability in copepod-associated microbial metacommunities
(CAMC) compared to seawater microbial metacommunities
(SMC). To test this hypothesis, we collected physicochemical
data from 46 stations at the epipelagic zone (0–200 m) along
the Mediterranean Sea. We sampled epipelagic water and
mesozooplankton from the Tyrrhenian to the Levantine
regions. We used 16S rRNA gene amplicon sequence variants
(ASVs) obtained from dominant copepod taxa (*Oncaea*,
*Farranula*, and *Clausocalanus*) and seawater to characterize
CAMC and SMC metacommunities, as well as to determine
the mechanisms shaping their spatial distribution.

Methods

Physicochemical data acquisition and sampling collection

Sampling was conducted in the framework of the French program PERLE (Pelagic Ecosystem Response in the Levant Experiment PERLE-4, https://doi.org/10.17600/18001980). A rosette sampler equipped with a CTD SeaBirdTM 911plus was used to collect seawater samples (for DNA extraction and inorganic nutrient measurements) and record physical variables in the epipelagic zone (0–200 m) at 46 stations distributed in the Mediterranean Sea, onboard the R/V *L'Atalante* between March 28 and April 26, 2021 (Fig. 1A). *See* Supporting Information Methods for more details in sampling collection.

Copepod sampling and identification

Vertical hauls with WP2 nets (200-µm mesh size) were deployed at 16 stations to collect mesozooplankton. Individual adults of calanoid *Clausocalanus* and cyclopoid *Farranula* and *Oncaea* copepods were manually picked from the net samples and identified onboard (see Supporting Information Methods). We selected these copepod genera because they are widely distributed throughout the Mediterranean Sea, were dominant in our epipelagic samples, and play distinct functional roles within mesozooplankton communities (Benedetti et al. 2023). Five replicates of five individuals per taxon (whole organism) were collected into a sterile 1.5-mL tube with molecular-grade absolute ethanol and stored at -20°C until DNA extraction. Additionally, single individuals of *Farranula* (n = 2) and *Oncaea* (n = 3) per station were collected
<figure><img src="image_3.png" /><figcaption>**Fig. 1.** Map of sampling locations, vertical profiles of physical and chemical variables, and circulation patterns. (A) Geographical distribution of the 46 stations sampled during the PERLE-4 cruise across the Mediterranean Sea from March 29 to April 24, 2021. (B) Average depth profiles of water properties (epipelagic zone, 0–200 m) at the Tyrrhenian, Ionian-west (western), Ionian-east, Hellenic arc, Cretan, and Levantine (eastern) regions. Nutrient measurements (average, 0–200 m) are fitted to GAMs. (C) Surface water circulation from Copernicus Marine Service (CMEMS) physical reanalysis model at selected dates (western region: 2 April, 2021; eastern region: 10 April, 2021), matching the period of sampling.</figcaption></figure>
and preserved in molecular ethanol for molecular identification (barcoding, *see* Supporting Information Methods).

## Seawater vertical profiles and circulation models

Profile data of temperature (°C), salinity, density (kg m⁻³), dissolved oxygen (µmol kg⁻¹) and fluorescence-Chl-*a* (µg L⁻¹) were recorded and used to calculate the average depth profiles of the environmental variables at each station in the Tyrrhenian, Ionian-west, Ionian-east, Hellenic arc, Cretan, and Levantine regions, and plotted using Ocean data view (ODV 5.7.2) (Schlitzer 2018). Physical reanalysis model data was used to estimate the surface currents at the time of sampling (Mediterranean Sea Physical Reanalysis, CMEMS MED-Currents; https://doi.org/10.25423/CMCC/MEDSEA_MULTIYEAR PHY_006_004_E3R1) (*see* Supporting Information Methods).

## Next-Generation Sequencing, host barcoding, and bioinformatic analyses

Prior to DNA extraction, copepod specimens were rinsed with sterilized molecular water. Total genomic DNA for NGS (Next-Generation Sequencing) of microbial communities and copepod barcoding (using Sanger sequencing) was extracted using the DNeasy Blood & Tissue Kit (Qiagen, Germany). The 16S rRNA gene (V4 hypervariable region) was PCR amplified by the modified primer pair 515F/806R (Apprill et al. 2015; Parada et al. 2016) and sequenced using Illumina MiniSeq. The obtained 16S sequences (available in NCBI under Project PRJNA1240050) were processed by the nf-core/ampliseq pipeline (Straub et al. 2020), using DADA2 with default parameters (https://nf-co.re/ampliseq/2.11.0/) to determine the amplicon sequence variants (ASVs) and assign taxonomy against the Silva 16S database v. 138 (Quast et al. 2012). Full methods, including PCR conditions and sequence processing and bioinformatic pipelines, are described in Velasquez et al. (2023) and in the Supporting Information Methods.

## Statistical and phylogenetic null model analyses

Microbial data analyses and visualization were performed using R v. 4.1.0 (R Development Core Team 2021) the “phyl-oseq” “vegan” and “ggplot2” packages (McMurdie and Holmes 2013; Oksanen et al. 2013; Wickham et al. 2016) Principal coordinate analysis (PCoA) was used to visualize the β-diversity (species composition per station) dissimilarities among regions. The Mantel test was used to determine the effects of the environmental factors and geographical distances in driving CAMC and SMC β-diversity. The relative contribution of ecological processes (selection, dispersal, and ecological drift) to structuring CAMC and SMC was quantified based on the phylogenetic null model analysis (Stegen et al. 2013; Stegen et al. 2015). The model estimation involves two main steps. First, the contribution of selection is assessed by calculating the beta Nearest Taxon Index (βNTI), which measures phylogenetic distances between each ASV and its closest relative in each pair of samples (Junger et al. 2023). Second, the influence of dispersal and ecological drift is

evaluated by computing the Raup–Crick metric based on Bray–Curtis dissimilarities (RCbray), using ASV compositional turnover (Gazulla et al. 2022). Detailed methods of the null model estimation are available in the Supporting Information Methods.

The frequency in which each microbial ASV taxa (CAMC and SMC) occurs across stations and their mean relative abundance across all samples was calculated to distinguish between rare and abundant taxa with restricted or wide distributions (for more details on the analysis, *see* the Supporting Information Methods). The abundance-occupancy relationship (the correlation between the occurrence of each ASV taxa and their mean relative abundance) (Logares et al. 2020) was evaluated and plotted using non-linear least-squares fitting analysis by a sigmoid formula (“minpack.lm” package in R, Elzhov et al. 2013). Furthermore, we checked for differences in the relative abundances across regions of microbial taxa that were mostly associated with the copepods using the Kruskal–Wallis test, followed by multiple pairwise comparisons with multiple testing. The full description of the methods is detailed in the Supporting Information Methods.

## Results

### Environmental gradients and surface water circulation

The Mediterranean Sea transect showed contrasting physical and chemical features in the western compared to the eastern basin. The western regions displayed more stratified waters with lower temperatures and salinities and higher static (water column) stability than the eastern regions, which were characterized by mixed waters with higher temperatures and salinities (Fig. 1B). Fluorescence-Chl-*a* values were higher in the western regions than in the eastern regions. Oxygen depletion with depth displayed strong changes in the western Mediterranean Sea, where high concentrations were recorded in the upper 80 m (Fig. 1B). Water density was low in the upper 50 m and strongly increased with depth in the Tyrrhenian and Ionian-west, while the eastern regions displayed much smaller vertical variability. PO₄ and NOₓ concentrations decreased eastward (Fig. 1B).

The circulation models showed a strong signal of surface water outflowing from the Tyrrhenian to the Ionian-west via two pathways, the Sicily Channel and the Strait of Messina (Fig. 1C), contributing to the connectivity between western regions. In the eastern basin, the current analysis showed a signal of surface seawater intrusion from the Cretan region to the Hellenic arc and eastern Ionian. Similarly, there was a weak outflow of seawater from the Levantine to the other eastern regions (Fig. 1C).

### Microbial β-diversity patterns

The PCoA showed spatial dissimilarity in CAMC. Two main clusters were observed; the first one including the western regions, Tyrrhenian and Ionian-west, and the second
encompassing the eastern regions, Ionian-east, Cretan, Hellenic arc, and Levantine (Fig. 2A). The first PCoA axis indicated significant differences in CAMC composition between the two western regions (Wilcox test, *p* = 0.001). However, the Tyrrhenian and Ionian-west regions were markedly different from the CAMC of the eastern regions (*p* ≤ 0.00005; Table S1). Non-significant differences (*p* > 0.05) were observed in CAMC among some of the eastern regions (Table S1). In contrast, SMC showed low spatial variability, displaying similar microbial composition among regions (Fig. 2B). Seawater microbial metacommunities clustered by epipelagic depth layer, showing significant differences between samples from the lower epipelagic (100–200 m) vs. upper (5–75 m) layer and the DCM (39–80 m) (*p* ≤ 0.0000014; Table S2).

## Biotic and abiotic factors influencing the microbial composition

The genetic distances of *Farranula* and *Oncaea* (p-distances), indicating host genetic variation, are detailed in Tables S3 and S4, respectively. A non-significant correlation was found between the genetic distances and the microbial dissimilarity (Bray–Curtis distances within stations; Fig. S1) of *Farranula*

(*R*² = −0.03; *p* = 0.5) and *Oncaea* (*R*² = 0.01; *p* = 0.3). See Supporting Information Results—Host genetic diversity for further information.

The correlation between geographic distances and Bray–Curtis dissimilarity, as a proxy for β-diversity in CAMC, was positive (Mantel *r* = 0.35, *p* = 0.001; Fig. 3A), showing a significant distance–decay relationship. The factors that contribute mostly to the variability in CAMC were salinity (Mantel *r* = 0.40), temperature (*r* = 0.34), and NOₓ (*r* = 0.32), presenting a positive and significant correlation (Fig. 3B). Seawater density (*r* = 0.27), PO₄ (*r* = 0.26) and dissolved oxygen (*r* = 0.21) had a smaller and significant influence on CAMC, while Chl-*a* did not show a significant effect on CAMC (*r* = −0.08, *p* = 0.98). In contrast, SMC displayed a weaker distance-decay relationship (*r* = 0.18, *p* = 0.01; Fig. 3A). All environmental factors had a significant positive effect on the SMC, with NOₓ, dissolved oxygen, and PO₄ having a higher influence than temperature, salinity, Chl-*a*, and water density (Fig. 3B).

## Ecological processes structuring microbial β-diversity

The null model analysis showed that the relative contributions of the mechanisms shaping the microbial β-diversity in

<figure><img src="image_4.png" /><figcaption>**Fig. 2.** Differences in the microbial β-diversity among regions across the Mediterranean Sea. (A) Principal coordinates analysis (PCoA) of the CAMC composition (Bray–Curtis distances). Boxplots of the first PCoA axis showing the differences between regions. (B) Principal coordinates analysis (PCoA) of the SMC composition (Bray–Curtis distances). Boxplots of the first PCoA axis showing differences by depth layer in the epipelagic zone.</figcaption></figure>
<figure><img src="image_5.png" /><figcaption>**Fig. 3.** Relationships between microbial β-diversity, geographical distances, and environmental factors. (A) Distance (km) decay of dissimilarities for CAMC and SMC along the Mediterranean Sea. (B) Heatmap showing the Mantel correlations of CAMC and SMC dissimilarity (Bray–Curtis distances) with each environmental factor (Euclidian distances). Red and blue colors indicate positive or negative correlation, respectively. * indicates significant effect (*p* < 0.05) of the environmental factor on the microbial composition.</figcaption></figure>

the Mediterranean Sea highly differed between CAMC and SMC (Fig. 4A). Ecological drift had the largest impact on CAMC composition (57%), while homogenizing dispersal was the most important process structuring SMC (46%). The role of selection was very limited in CAMC (9%) compared to SMC (22%) (Fig. 4A). Heterogeneous and homogeneous selection explained 8% and 1% of CAMC turnover, while 9% and 13% in SMC, respectively. Dispersal limitation contributed 10% and 17% to CAMC and SMC, respectively, whereas homogenizing dispersal explained half of the proportion (24%) in CAMC compared to SMC (46%). Drift had a low impact on SMC, contributing 15% to the changes in the composition. The dominant role of ecological drift in the CAMC turnover was supported by the abundance-occupancy relationship analysis where CAMC showed taxa with highly restricted distribution and low mean relative abundances (91%; Fig. 4B) and only 0.3% widely distributed and highly abundant microbial taxa. In contrast, SMC displayed a lower proportion of rare taxa with restricted distribution (55%) but a higher proportion of widely distributed and highly abundant taxa (4%; Fig. 4B).

## Geographical patterns of dominant copepod-associated microbial taxa

The relative abundances of the dominant CAMC taxa (Fig. S2) varied according to the region, showing similar

patterns in regions with similar environmental factors and spatial connectivity (Fig. 5A). Dominant taxa in CAMC and their correlation with environmental factors are detailed in Fig. 5B and the Supporting Information Results.

## Discussion

Our study represents the first large-scale investigation of epipelagic copepod-associated microbial metacommunities (CAMC), revealing that the pronounced spatial gradients and surface circulation patterns in the Mediterranean Sea drive CAMC β-diversity. Here, we show higher CAMC similarity between regions with lower environmental variability. These results support the assumption that host species inhabiting similar abiotic conditions tend to harbor similar microbial metacommunities (Härer and Rennison 2023; Schellenberg and Clarke 2020), as they share microbes from the same external environment and through direct host–host interactions (Adair and Douglas 2017; Araujo et al. 2024). The variability of salinity and temperature had the strongest influence on CAMC, increasing the microbial dissimilarity between the western and eastern basins. This finding is consistent with studies in corals (van Oppen and Blackall 2019), sponges (Yang et al. 2019), and macroalgae (Pearman et al. 2024),
<figure><img src="image_6.png" /><figcaption>**Fig. 4.** Relative contribution of the ecological processes to CAMC and SMC and abundance-occupancy relationship. (A) Bar plots indicating the effects of ecological drift, dispersal, and selection (environmental filtering) on CAMC and SMC. The circled illustrations represent the CAMC (left) and SMC (right). (B) Mean relative abundance vs. occurrence (i.e., number of samples in which each taxon is present) of CAMC and SMC taxa in all samples and stations. The yellow lines indicate the restricted (< 10% occurrence) and rare taxa (< 0.0001 rel. abundance) and the widely distributed (> 70% occurrence) and abundant taxa (> 0.001 rel. abundance).</figcaption></figure>

demonstrating that variation in these environmental factors changes host microbial composition.

Water currents play an important role in creating either dispersion or mixing barriers that lead to connectivity or lack of it, affecting planktonic communities' distribution (Sommeria-Klein et al. 2021; Srivastava and Kratina 2013). Here, we found that physical connectivity results in high similarity in the CAMC within each of the two main basins. The intrusion of surface waters from the Tyrrhenian to the western Ionian via the Sicily Channel and the Straits of Messina increased the connectivity between the Tyrrhenian and the western Ionian, possibly facilitating the co-dispersal of the hosts with their associated microbes. Similarly, the water circulation from the Cretan region to the Hellenic arc and eastern Ionian can explain the

CAMC resemblance in the eastern basin. These results support the hypothesis that associated microbial distribution often depends on the dispersal dynamics of the hosts (Dickey et al. 2021; Mony et al. 2022). Mesoscale oceanographic features (currents) can last several days to many months (Bonaduce et al. 2021), driving the distribution dynamics of both the host and associated microbes. We found a stronger distance-decay relationship in CAMC than SMC, indicating a greater similarity in copepod microbial composition with decreased geographic distance. This pattern aligns with a meta-analysis by Clark et al. (2021) comparing distance-decay relationships in different free-living and host-associated microbial communities.

Our results indicated that the compositional variability in the CAMC was not related to interspecific genetic differences
<figure><img src="image_7.png" /><figcaption>**Fig. 5.** Dominant taxa in the CAMC and their correlations with environmental factors. (A) Relative abundance of microbial taxa that significantly changed between the western and eastern regions (*p* < 0.05, pairwise comparisons). (B) Heatmap showing Spearman rank correlation between the relative abundance of the associated microbial taxa and each environmental factor.</figcaption></figure>

within each copepod host, as congeneric copepods can have convergent microbial composition. Different copepods that share ecological traits (e.g., diet, feeding behavior) tend to harbor similar microbial communities (Velasquez et al. 2023). Previous studies have indicated that, at large spatial scales, host distribution and environment are stronger drivers of microbial assembly than host identity (Boscaro et al. 2022; Freeman et al. 2020), suggesting that higher variability in the microbial composition may mask the influence of host identity. While the host regulates some microbial taxa, ecological processes (dispersal, selection, and ecological drift), operating at the metacommunity level, shape the dynamics of other microbial taxa (Adair and Douglas 2017).

Our analysis revealed that ecological drift explains a high proportion of the microbial compositional variability in CAMC, likely as a response to the restricted CAMC dispersal between the western and eastern basins. When microbial dispersal is limited, stochastic processes within the local metacommunity can lead to compositional drift (Adair and Douglas 2017). The abundance and distribution of species are typically positively correlated—species that are more abundant also tend to occupy more sites. This correlation is known as the abundance-occupancy relationship (Gaston et al. 2000). In our study, however, we found a negative abundance-occupancy relationship in CAMC, where copepods harbor a high proportion of rare microbial taxa with narrow distribution. This suggests that CAMC are more susceptible to the effect of ecological drift since rare taxa are less dispersed and more likely to become locally extinct (Nemergut et al. 2013; Zhou and Ning 2017). Ecological drift has a stronger influence on the microbial metacommunity when selection (environmental filtering) is weaker (Chase and Myers 2011). Indeed,

we found a very limited effect of environmental filtering and a high influence of ecological drift on the CAMC. The constraint influence of environmental conditions on the host microbes is the result of the selective pressures imposed by the hosts (Delleuze et al. 2024; Härer and Rennison 2023). The observed homogeneous dispersal reflects the physical connectivity within each basin, reinforcing the assumption that surface circulation patterns are a key driver in structuring the CAMC at the epipelagic Mediterranean Sea. The dominant CAMC taxa recorded during our study were characterized by copiotrophic bacteria, which can benefit from the host by increasing their abundance and activity (Troussellier et al. 2017). The majority of these opportunistic taxa were more abundant in eastern than western regions, suggesting that in warm and nutrient-depleted environments, copepods may provide a rich habitat for microbe colonization, enhancing their ability to survive (Velasquez et al. 2023).

Surface seawater microbes (SMC) can exhibit high dispersal rates since they can cope with adverse environmental conditions and circulation barriers by aerial dispersal and dormancy (Echenique-Subiabre et al. 2025; Mestre and Höfer 2021), reducing compositional differences at a large spatial scale (Villarino et al. 2022). Contrary to CAMC, SMC had a higher spatial similarity between regions, resulting in a weaker distance-decay relationship that suggests a homogenized distribution in the epipelagic zone of the Mediterranean Sea. The observed SMC shift by depth is consistent with previous studies that showed vertical stratification of the seawater microbial taxa (Mapelli et al. 2013; Sebastián et al. 2021; Sunagawa et al. 2015). Changes in SMC were shown to be strongly influenced by abiotic factors, such as water temperature, dissolved oxygen, and nutrient concentrations (Junger et al. 2023;
Sunagawa et al. 2015). Similarly, we found that dissolved oxygen and nutrient availability significantly affect the SMC. High environmental variability can favor dominant microbes with broad environmental tolerance (i.e., cosmopolitan taxa), leading to wider distribution (Hanson et al. 2012; Ruiz-González et al. 2019), and explaining the enhanced effect of homogeneous dispersal on the SMC. Generalist taxa are less susceptible to local extinction due to their more stable relative abundances (Junger et al. 2023; Villarino et al. 2022), featuring limited ecological drift, which aligns with our findings. The most abundant free-living microbial taxa include cosmopolitan oligotrophs such as *Prochlorococcus* and *SAR11* (Ruiz-González et al. 2019; Troussellier et al. 2017), as was also observed in our study. These microbes are susceptible to environmental variability, making selection an important ecological process in shaping the microbial composition in surface waters (Langenheder and Lindström 2019). We found that homogeneous selection had a larger effect on SMC than heterogeneous selection. This is likely because certain microbial taxa are adapted to specific environmental conditions, resulting in similar microbial composition (Gazulla et al. 2022; Martiny et al. 2006).

Here, we showed that CAMC reflect the cross-basin spatial gradients and circulation patterns of the epipelagic Mediterranean Sea better than SMC. These findings suggest that CAMC can serve as proxies of biogeographical patterns across the global surface ocean. The link we found between oligotrophic environments and copepod-associated copiotrophic taxa indicates that CAMC can be potentially used as bioindicators of the ongoing and projected global change, with the increasing stratification and consequent oligotrophy of the epipelagic layer. Future studies on microbial metacommunities associated with copepods and other plankters should investigate the change in the role of selection, dispersal, and drift along latitudinal and depth gradients to better understand the link between microbial metacommunities and pelagic hosts.

## Author Contributions

This study was conceived by Tamar Guy-Haim and Ximena Velasquez. Ximena Velasquez and Tamar Guy-Haim co-led the study, designed and conducted the field sampling, wrote the manuscript, and contributed equally to the study. Ximena Velasquez performed the molecular and statistical analyses. Tal Ozer performed the oceanographic modeling. Maria Grazia Mazzocchi confirmed the morphological identification of the copepods used in this study. Francois Carlotti assisted with field sampling. Mireille Pujo-Pay, Pascal Conan, and Xavier Durrieu de Madron have supplied the physical and chemical data. Tamar Guy-Haim and Dalit Meron supervised the student. All authors provided comments and edits on the manuscript.

## Acknowledgments

This study is a contribution of the PERLE project, a joint initiative of the “Chantier Méditerranée” MERMEX supported by CNRS-INSU, IFREMER, CEA, and Météo-France as part of the program MISTRALS coordinated by INSU, and the Israeli Oceanographic and Limnological Research (IOLR). We would like to thank the R/V *L'Atalante* crew for helping at sea. The study was funded by the Southern Marine Science and Engineering Guangdong Laboratory (Guangzhou) grant SMSEGL20SC02 to Tamar Guy-Haim and Dalit Meron.

## Conflicts of Interest

None declared.

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**Supporting Information**

Additional Supporting Information may be found in the
online version of this article.

Submitted 30 March 2025

Revised 18 July 2025

Accepted 01 August 2025