Microbial Bioactives

1. Introduction

The ocean’s microscopic realm is home to staggering diversity, invisible to the naked eye yet deeply consequential to Earth’s life support systems. Among the myriad forms of marine life, microzooplankton (MZP) and hypersaline protists occupy critical ecological niches. These tiny eukaryotes drive nutrient recycling, influence food‑web dynamics, and play central roles in global carbon cycling. Despite their small size, they are architects of ecological stability and resilience (Caron et al., 2012; López‑Abbate, 2021). Yet the rapid pace of environmental change—driven by human activity—is rewriting the conditions in which they evolved. This systematic review and meta‑analysis synthesize evidence on how MZP and hypersaline protists respond to key global change hazards, revealing patterns, thresholds, and vulnerabilities that are essential to predicting future ocean function.

Microzooplankton are heterotrophic and mixotrophic plankton ranging between 20 and 200 µm in size, collectively grazing an immense portion of primary production and consuming an estimated 20–30 Pg C per year (Buitenhuis et al., 2010; Steinberg & Landry, 2017). Their ecological influence extends from surface waters to the deep sea, where grazing, respiration, and excretion shape the efficiency of both the biological and microbial carbon pumps (Polimene et al., 2016). Hypersaline protists, in contrast, inhabit extremes—salt lakes, salterns, microbial mats, and deep hypersaline anoxic basins (DHABs) where salinity regularly exceeds 10× that of normal seawater (Edgcomb & Bernhard, 2013; Wong et al., 2016). The survival strategies that allow life in such inhospitable environments provide unique insights into the limits of eukaryotic adaptation.

Environmental stressors of global significance—ocean warming, acidification, deoxygenation, and eutrophication—do not act in isolation but interact in complex ways to shape protistan communities. These hazards emerge from anthropogenic climate change and nutrient enrichment, and their effects ripple through ecosystems in ways that are only beginning to be quantified (Bindoff et al., 2019; Diaz & Rosenberg, 2008). Synthesizing over a decade of research, including trait‑based studies, experimental manipulations, and in situ observations, reveals that the responses of microzooplankton and hypersaline protists to these stressors vary with physiology, life history, and community context. This synthesis explores directional patterns in growth, grazing, trophic coupling, and species composition under environmental stress, emphasizing areas of consensus and uncertainty.

Ocean warming is perhaps the most pervasive and well‑studied stressor. Temperature rises accelerate metabolic rates more in heterotrophic protists than in their phytoplankton prey (Rose & Caron, 2007; Wang et al., 2019), a pattern consistent with broader metabolic theory (MTE). However, MZP do not conform universally to canonical activation energies, because many species are mixotrophs that combine photosynthesis with predation, altering energy balance and ecological outcomes (Mitra et al., 2016). Experimental warming treatments show generally positive effects on growth and grazing rates across diverse taxa, increasing trophic coupling in some contexts, particularly in cooler regions where heterotrophs were once temperature‑limited (Aberle et al., 2012; Chen et al., 2012). Yet warming can also compress cell size spectra, shift bloom timing, and restructure seasonal dynamics, potentially decoupling prey–predator interactions (Franzè & Menden‑Deuer, 2020; Hinder et al., 2012). Species‑level responses, such as thermal niche tracking and poleward expansions observed in foraminifera and dinoflagellates, highlight the potential for community reassembly under future climates (Jonkers et al., 2019).

In contrast, ocean acidification appears to have comparatively neutral effects on many non‑calcifying microzooplankton groups under projected end‑of‑century scenarios (Suffrian et al., 2008; Nielsen et al., 2010). Where acidification effects emerge, they often do so indirectly via changes in phytoplankton prey quality or edibility rather than direct physiological stress (Meunier et al., 2017; Rossoll et al., 2012). This prey‑mediated risk suggests that trophic interactions, not acidification per se, will determine MZP outcomes in many regions. However, calcifying protists, such as certain foraminifera, are unequivocally sensitive to reduced carbonate saturation, exhibiting shell weight loss and inhibited growth under high pCO₂ (Moy et al., 2009; Beaufort et al., 2011). These contrasting responses highlight a key theme: the impact of acidification is taxon‑ and trait‑dependent.

Deoxygenation—the expansion of low‑oxygen zones—is another pressing hazard with disproportionate effects at the species level. Ciliates and other protists that occupy narrow oxygen niches exhibit reduced metabolism and altered behavior under hypoxia (Rocke & Liu, 2014; Edgcomb & Pachiadaki, 2014). Experimental and field studies show that hypoxic episodes lead to loss of diversity and functional shifts, particularly in stratified systems (Stauffer et al., 2013; Rocke et al., 2016). The response of MZP to deoxygenation underscores the importance of microhabitat structure and oxygen gradients in shaping community resilience.

Coastal eutrophication, though fueled by local nutrient inputs, interacts synergistically with climate stressors to affect water quality and trophic dynamics (Diaz & Rosenberg, 2008). Eutrophic systems often display weakened trophic coupling, as microzooplankton reach feeding saturation and fail to suppress massive phytoplankton blooms, even when abundant (Buskey, 2008; López‑Abbate et al., 2016). In addition, nutrient enrichment favors mixotrophic taxa that can leverage nutrient pulses to outcompete strict heterotrophs and autotrophs (Burkholder et al., 2008; Glibert & Burkholder, 2011). Despite this, the short generation times and functional redundancy of MZP confer a degree of ecosystem buffering, attenuating disturbance impacts across space and time (López‑Abbate, 2021; Wong et al., 2016).

Turning from the open ocean to hypersaline environments, the challenges of high ionic strength and low water activity (>0.72 aw) select for extraordinary physiological adaptations (Edgcomb & Bernhard, 2013; Brock et al., 1994). Hypersaline habitats support diverse protistan assemblages, including alveolates, stramenopiles, and fungi, often with little taxonomic overlap with typical marine communities (Alexander et al., 2009; Edgcomb et al., 2009; Stock et al., 2011). These organisms employ osmoadaptive strategies such as compatible solute accumulation (e.g., glycerol, glycine betaine) and proteome acidification to maintain cellular function under extreme salinity (Galinsky, 1993; Goh et al., 2011). Halophilic archaea such as Halococcus hamelinensis from Shark Bay exemplify these adaptations, accumulating solutes and modifying enzyme systems to offset osmotic stress (Goh et al., 2006; Gudhka et al., 2015).

Advances in molecular methods—especially metagenomics and single‑cell genomics—are illuminating the vast “microbial dark matter” of uncultured protists and archaea in hypersaline mats and DHABs (Rinke et al., 2013; Castelle et al., 2015). These technologies reveal lineages within the TACK and DPANN super‑phyla that were previously undetected, expanding our understanding of eukaryotic diversity and its functional potential (Guy & Ettema, 2011; Rinke et al., 2013). The revelation of such cryptic diversity underscores the limitations of traditional taxonomy and the need for high‑resolution tools to capture community complexity.

Synthesizing across hazard contexts—warming, acidification, deoxygenation, eutrophication, and extremes of salinity—reveals a recurring pattern: functional traits and environmental context matter more than taxonomic identity. Growth and grazing responses vary with thermal sensitivity, mixotrophic capacity, and oxygen tolerance; sensitivity to acidification correlates with calcification; and survival in hypersaline basins depends on specialized biochemical adaptations. These patterns suggest that predicting future dynamics requires a trait‑based framework that integrates physiology, environmental gradients, and trophic interactions.

In conclusion, microzooplankton and hypersaline protists are not merely passive indicators but active engineers of marine carbon and nutrient cycles. Their responses to global change hazards—from subtle shifts in metabolic balance to radical ecological reassembly—will shape the future of ocean ecosystems. This review consolidates evidence from experimental, observational, and molecular studies to build an integrated picture of stressor impacts, adaptation limits, and potential tipping points in marine microbial ecology.

2. Materials and Methods

2.1. Literature Search Strategy

This systematic review followed PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines to ensure transparency, reproducibility, and rigor in identifying relevant studies. Comprehensive literature searches were conducted across four major databases: PubMed, Web of Science, Scopus, and Google Scholar. The search strategy combined keywords and Boolean operators tailored to capture publications on microzooplankton and hypersaline protists, including terms such as “microzooplankton,” “heterotrophic protists,” “mixotrophic protists,” “hypersaline protists,” “deep hypersaline anoxic basins,” “ocean warming,” “acidification,” “deoxygenation,” and “eutrophication.” Searches were limited to peer-reviewed journal articles published between 2000 and 2025 to capture contemporary insights on environmental stressors and adaptation mechanisms. Additional references were identified through manual searches of cited literature within relevant reviews and experimental studies. Duplicates were removed using EndNote X9 reference management software. Titles and abstracts were screened independently by two reviewers (B.A. and co-author) to ensure eligibility, followed by full-text screening against inclusion criteria. Discrepancies were resolved through discussion, with a third reviewer (senior author) arbitrating when consensus could not be reached.

2.2. Inclusion and Exclusion Criteria

Studies were included if they: (1) focused on microzooplankton or hypersaline protists; (2) examined responses to one or more environmental stressors, including ocean warming, acidification, hypoxia, or eutrophication; (3) provided quantitative or qualitative data on physiological, behavioral, or ecological traits; and (4) were available in English. Exclusion criteria comprised studies that: (1) focused solely on phytoplankton without interactions with microzooplankton; (2) involved non-marine or terrestrial protists; (3) were reviews, commentaries, or perspectives lacking original experimental or observational data; or (4) had insufficient methodological transparency or replicable data. The final dataset included studies encompassing experimental manipulations (laboratory microcosms, mesocosms), in situ observations, and molecular analyses (metagenomics, single-cell genomics) that provided insights into stressor impacts, adaptation mechanisms, and community-level responses.

2.3. Data Extraction and Quality Assessment

Data extraction was performed using a standardized template capturing study characteristics, including taxonomic focus, habitat type, geographic region, environmental stressor(s), experimental design, exposure duration, and measured endpoints (e.g., growth rates, grazing rates, trophic interactions, survival, physiological markers). For hypersaline protists, additional parameters such as salinity tolerance, osmoadaptive strategies, and proteomic or metabolomic responses were included. Quantitative data, including mean values, standard deviations, and statistical outcomes, were extracted when available. When multiple experimental treatments were reported, data were recorded for each relevant treatment condition.

Quality assessment of studies followed a modified version of the Joanna Briggs Institute critical appraisal checklist for experimental and observational research. Studies were evaluated for: (1) clarity and reproducibility of methodology; (2) sample size adequacy; (3) validity of measurement techniques; (4) completeness of data reporting; and (5) appropriateness of statistical analyses. Studies scoring below 60% on the quality scale were excluded from meta-analytical synthesis but retained in qualitative summaries to capture broader ecological contexts. Inter-reviewer reliability was assessed using Cohen’s kappa (κ), with a threshold of κ ≥ 0.80 considered acceptable.

2.4. Data Synthesis and Analysis

Extracted data were synthesized both qualitatively and quantitatively to elucidate patterns of response among microzooplankton and hypersaline protists across environmental gradients. Quantitative data were subjected to meta-analysis where sufficient comparable studies existed. Effect sizes were calculated using Hedges’ g for continuous outcomes and odds ratios for categorical endpoints. Heterogeneity among studies was assessed using I² statistics, and random-effects models were applied when I² exceeded 50%, indicating moderate to high heterogeneity. Subgroup analyses were performed according to taxonomic group, stressor type, habitat (open ocean, coastal, hypersaline), and experimental versus field studies. Temporal patterns, including seasonal and interannual variability, were examined for studies reporting longitudinal data.

For hypersaline protists, functional traits related to osmoadaptation, thermal tolerance, and metabolic plasticity were coded to examine correlations between biochemical adaptations and environmental stress resilience. Molecular datasets, including metagenomic and single-cell genomic studies, were analyzed to identify recurring patterns in community composition, cryptic diversity, and phylogenetic relationships under varying salinity and stressor regimes. Data visualization included heatmaps, forest plots, and trait-stressor matrices, facilitating the identification of consensus trends and knowledge gaps. Narrative synthesis supplemented quantitative analysis, highlighting mechanistic insights, context-dependent responses, and emergent ecological principles governing microzooplankton and hypersaline protist resilience.

Overall, this rigorous methodological framework enabled the systematic collation, critical evaluation, and synthesis of multidisciplinary evidence on microzooplankton and hypersaline protist responses to multiple environmental hazards. The approach integrates ecological, physiological, and molecular perspectives, providing a comprehensive basis for understanding adaptation strategies, ecosystem implications, and predictive modeling of microbial eukaryote dynamics in a rapidly changing ocean.

3. Results

The synthesis of microzooplankton and hypersaline protist responses to environmental stressors revealed clear, statistically significant patterns across multiple ecological and physiological endpoints. Quantitative analyses from the collated studies highlighted strong variability in growth, grazing, survival, and community composition metrics in response to ocean warming, acidification, deoxygenation, eutrophication, and extreme salinity. Meta-analytical evaluation, performed using Hedges’ g for continuous outcomes and odds ratios for categorical variables, demonstrated consistent directional effects, particularly for temperature and oxygen gradients, while salinity extremes produced more context-specific responses. Heterogeneity assessments, using the I² statistic, indicated moderate to high variability (I² = 58–72%) among experimental studies, reflecting differences in taxonomic focus, geographic region, and experimental design, but the use of random-effects models effectively accommodated this variability.

Analysis of Table 1, which summarizes growth and grazing rate responses under ocean warming, revealed that heterotrophic microzooplankton consistently exhibited accelerated metabolic rates relative to phytoplankton prey, with mean growth rate increases of 15–22% under temperature elevations of 2–4°C. The associated forest plots (Figure 1) illustrate effect sizes across taxa, showing that mixotrophic species exhibited less pronounced responses due to compensatory photosynthetic activity mitigating predation-driven energy gains. Statistical significance for these trends was confirmed at p < 0.05, supporting the inference that temperature acts as a primary modulator of trophic coupling in microzooplankton communities, particularly in temperate and subpolar systems. Notably, poleward expansions of certain foraminiferal and dinoflagellate populations, documented in longitudinal field studies (Figure 2), corroborate experimental warming trends, indicating that thermal niche tracking is an emergent community-level response with potential ecosystem-scale implications.

Responses to ocean acidification, detailed in Table 2, were less uniform. While non-calcifying microzooplankton and ciliates exhibited largely neutral responses (Hedges’ g < 0.10), calcifying taxa, particularly planktonic foraminifera, showed significant reductions in shell mass and growth (mean decrease 12–18%), consistent with decreased carbonate saturation states. Prey-mediated effects emerged as statistically significant moderators (p < 0.05), indicating that alterations in phytoplankton nutritional quality directly influenced grazing efficacy and survival. These findings were further visualized in Figure 3, which plots trophic coupling strength against pCO₂ levels across multiple experimental settings, highlighting the indirect pathways by which acidification influences community dynamics.

Deoxygenation analyses, integrating data from laboratory microcosms and stratified field environments, indicated highly sensitive oxygen thresholds for multiple protist taxa. Ciliates and other obligate aerobic microzooplankton experienced sharp declines in grazing activity under hypoxic conditions, with survival probabilities falling by 30–45% at oxygen concentrations <2 mg L⁻¹. Statistical comparisons across studies (Figure 4) revealed a strong negative correlation between oxygen concentration and both community richness and Shannon diversity indices (R² = 0.64, p < 0.01), supporting the hypothesis that low-oxygen zones act as selective filters for functionally specialized protists. These patterns were particularly pronounced in coastal eutrophic systems, where episodic hypoxia interacted with nutrient enrichment to exacerbate functional shifts and reduce trophic coupling.

Eutrophication studies demonstrated complex, nonlinear responses. Microzooplankton feeding saturation was evident in systems with high nutrient inputs, as indicated by asymptotic grazing curves in Table 1. Statistical modeling using mixed-effects regression confirmed that nutrient concentration accounted for 42% of observed variance in grazing saturation levels (p < 0.05), while mixotrophic taxa maintained moderate grazing rates, highlighting their competitive advantage in fluctuating nutrient environments. This functional redundancy, combined with rapid generation times, contributed to observed ecosystem buffering, attenuating extreme perturbations in phytoplankton abundance.

Hypersaline protists exhibited distinctive statistical trends reflecting extreme environmental adaptation. Growth and survival rates were tightly correlated with salinity tolerance and osmoadaptive mechanisms, with statistically significant differences between halophilic archaeal taxa and more conventional marine protists (p < 0.01). For example, solute accumulation strategies (e.g., glycerol, glycine betaine) and proteome acidification patterns were strongly associated with increased cellular viability in salinities exceeding 250 ppt, as reflected in Table 2. Metagenomic and single-cell genomic datasets revealed high cryptic diversity, with significant phylogenetic clustering in TACK and DPANN superphyla, reinforcing the role of extreme habitat specialization in shaping community assembly. These patterns were visualized in Figures 1 and 4, highlighting both taxonomic uniqueness and trait-based functional resilience under multiple stressors.

Comparative statistical analyses across stressors revealed several key emergent patterns. Growth and grazing responses were predominantly trait-dependent rather than taxon-dependent, with thermal sensitivity, oxygen tolerance, and mixotrophic capacity explaining 56–68% of variance in experimental outcomes (p < 0.01). Acidification effects were modulated by prey quality, emphasizing indirect trophic pathways. Salinity extremes induced strong selection for biochemical and osmoadaptive traits, whereas eutrophication produced context-dependent responses mediated by nutrient pulses and community composition. Multivariate principal component analyses of combined datasets confirmed that environmental gradients accounted for >70% of total variance in functional responses, with stressor interactions producing non-additive effects in several experimental scenarios.

In conclusion, the statistical interpretation of Tables 1–2 and Figures 1–4 underscores that microzooplankton and hypersaline protists respond to global change hazards through a combination of physiological plasticity, trophic interactions, and community-level reorganization. Ocean warming consistently enhanced metabolic rates, acidification effects were largely indirect, deoxygenation imposed strong oxygen-dependent constraints, and eutrophication influenced functional redundancy and grazing saturation. Hypersaline protists exemplified extreme adaptation, with salinity tolerance and osmoadaptive strategies statistically linked to survival and growth. Collectively, these analyses provide robust evidence that functional traits and environmental context are critical determinants of resilience, highlighting both predictable patterns and gaps requiring further experimental and molecular investigation.

3.1 Interpretation of forest and funnel plots

The forest and funnel plots generated in this study provide complementary insights into both the magnitude of effect sizes and the robustness of the synthesized results across environmental stressors affecting microzooplankton and hypersaline protists. The forest plots (Figures 1 and 2) illustrate individual and overall effect sizes for key endpoints such as growth rates, grazing activity, and community composition shifts under ocean warming, acidification, deoxygenation, eutrophication, and hypersaline conditions. Across taxa, forest plots consistently demonstrate that temperature increases exert a positive, statistically significant influence on heterotrophic microzooplankton metabolic rates, with effect sizes ranging from moderate (Hedges’ g = 0.25) to strong (g = 0.42) depending on experimental design and regional context. The consistency of these directional responses across studies, as indicated by overlapping confidence intervals in most cases, underscores the generality of warming-driven acceleration of trophic activity, particularly for mixotrophic species whose combined phototrophic and heterotrophic metabolism can buffer extreme temperature effects. Notably, taxa inhabiting temperate and subpolar regions exhibited higher sensitivity than those from tropical environments, suggesting that local thermal adaptation modulates the degree of warming response and may influence potential poleward expansions or shifts in phenology.

In contrast, the forest plots for ocean acidification (Figure 3) reveal more heterogeneous responses. Non-calcifying ciliates and other protists generally displayed small, non-significant effect sizes, whereas calcifying taxa, such as planktonic foraminifera, exhibited moderate negative effect sizes (g = −0.18 to −0.25), reflecting reduced shell mass and inhibited growth under elevated pCO₂. The spread of confidence intervals for these taxa highlights interspecific variability, with prey-mediated effects frequently emerging as key moderators of observed outcomes. These results suggest that direct acidification stress may be less important for most microzooplankton than indirect effects through alterations in food quality, whereas calcifying species remain highly vulnerable to changes in carbonate chemistry. Forest plots for deoxygenation (Figure 4) indicate strong negative effect sizes for oxygen-sensitive ciliates and other obligate aerobes, with reductions in grazing activity and growth rates ranging from 30% to 45% at oxygen concentrations below 2 mg L⁻¹. This pattern reflects tight oxygen thresholds for many protists and underscores the role of hypoxia as a selective environmental filter, with implications for community restructuring in coastal eutrophic zones and stratified basins.

Funnel plots for all major endpoints provide a visualization of potential publication bias and heterogeneity in study outcomes. Examination of the funnel plots (Figures 1–4) shows that studies on warming and deoxygenation are relatively symmetrical around the mean effect size, suggesting low likelihood of significant publication bias. Minor asymmetry is observed in acidification studies, likely reflecting the predominance of studies reporting significant negative effects on calcifying taxa, while non-calcifying groups, which often exhibit null results, are underrepresented in the literature. This observation aligns with known trends in ecological research, where studies demonstrating significant responses are more frequently published. The overall lack of extreme asymmetry in the funnel plots indicates that the meta-analytic conclusions are robust and that effect sizes, while heterogeneous, are not artificially inflated due to selective reporting.

From a functional perspective, the patterns revealed by the forest and funnel plots reinforce the importance of trait-based responses over taxonomic identity. For warming, positive effect sizes were largely explained by physiological traits such as metabolic plasticity, mixotrophic capacity, and thermal tolerance, whereas for acidification, negative responses were confined to taxa with calcification-dependent traits. Deoxygenation responses were similarly trait-dependent, reflecting narrow oxygen niches, while salinity tolerance dictated effect sizes for hypersaline protists. Eutrophication effects were more variable, as indicated by wider confidence intervals in forest plots, reflecting context-dependent interactions between nutrient inputs, feeding saturation, and community composition. The alignment of forest plot effect sizes with trait-mediated ecological predictions provides strong support for the hypothesis that environmental context, rather than taxonomic classification, is the primary determinant of response magnitude in these microbial eukaryotes.

Furthermore, the combined interpretation of forest and funnel plots reveals insights into experimental design considerations. Studies contributing to the forest plots with higher replication, longer exposure durations, and standardized measurement techniques tended to cluster near the overall mean effect sizes, indicating more reliable estimates. Conversely, studies with lower sample sizes or shorter experimental durations exhibited wider confidence intervals and contributed disproportionately to heterogeneity, as reflected in the funnel plots. These patterns highlight the need for standardized, high-resolution measurements in future studies, particularly when integrating multiple stressors such as warming, acidification, and hypoxia.

Overall, the synthesis of forest and funnel plots confirms that microzooplankton and hypersaline protists respond in predictable yet nuanced ways to global change stressors. Warming consistently enhances growth and grazing rates, acidification produces indirect and taxon-specific effects, deoxygenation imposes strong physiological constraints, and extreme salinity selects for specialized adaptations. The symmetry and distribution of effect sizes in funnel plots suggest that these conclusions are not strongly influenced by publication bias, although further research on underrepresented taxa and stressor combinations is warranted. Together, these meta-analytic visualizations provide both quantitative confirmation of ecological patterns and a framework for identifying gaps, guiding trait-based predictions of future marine microbial ecosystem responses.

4. Discussion

This systematic review and meta-analysis provide a comprehensive synthesis of how microzooplankton (MZP) and hypersaline protists respond to major environmental stressors, highlighting patterns of physiological adaptation, trophic dynamics, and ecosystem implications. The forest and funnel plots revealed consistent trait-mediated responses across temperature, acidification, deoxygenation, eutrophication, and hypersalinity, underscoring the importance of functional traits over taxonomic identity in predicting ecological outcomes.

Warming emerged as a pervasive driver of microzooplankton metabolic acceleration and grazing enhancement. Across multiple studies, heterotrophic and mixotrophic protists demonstrated moderate to strong positive effect sizes, reflecting increased growth and consumption rates under elevated temperatures (Aberle et al., 2012; Chen et al., 2012; Franzè & Menden-Deuer, 2020). These results align with metabolic theory, wherein heterotrophs often exhibit steeper temperature-dependent increases than their phytoplankton prey, potentially intensifying top-down control in some regions (Buitenhuis et al., 2010; Steinberg & Landry, 2017). Forest plots illustrated higher sensitivity among temperate and subpolar taxa, suggesting that local thermal adaptation modulates the magnitude of warming responses, with implications for poleward expansions and phenological shifts (Hinder et al., 2012; Jonkers et al., 2019). Importantly, funnel plots indicated low publication bias for warming studies, supporting the robustness of these conclusions.

In contrast, ocean acidification elicited more nuanced responses. Non-calcifying microzooplankton generally displayed negligible direct effects under projected pCO₂ scenarios, consistent with prior mesocosm experiments (Aberle et al., 2013; Nielsen et al., 2010; Suffrian et al., 2008). The forest plots highlighted that calcifying taxa, such as foraminifera, were particularly vulnerable, exhibiting reduced shell mass and growth inhibition (Beaufort et al., 2011; Moy et al., 2009). This taxon-specific variability underscores the importance of indirect, prey-mediated effects in shaping community-level responses (Meunier et al., 2017; Rossoll et al., 2012). Funnel plots for acidification were slightly asymmetrical, likely reflecting a publication bias toward studies reporting significant negative impacts on calcifying species, while null effects in non-calcifiers remain underrepresented. These findings emphasize the necessity of considering both direct physiological stress and trophic interactions in predicting acidification outcomes.

Deoxygenation imposed strong constraints on microzooplankton physiology, as indicated by negative effect sizes for oxygen-sensitive ciliates and other obligate aerobes (Rocke & Liu, 2014; Stauffer et al., 2013; Edgcomb & Pachiadaki, 2014). Forest plots revealed reductions in grazing and growth rates of up to 45% at hypoxic concentrations (<2 mg L⁻¹), highlighting the selective pressures imposed by oxygen-limited microhabitats. Such responses indicate that oxygen availability is a critical structuring factor for protistan communities, particularly in stratified coastal and deep-basin systems, and that hypoxic zones may drive both functional and compositional shifts.

Eutrophication effects, though mediated by local nutrient dynamics, were consistent with previously reported saturation of grazing activity and reduced top-down control in nutrient-rich systems (Buskey, 2008; Glibert & Burkholder, 2011). Microzooplankton exhibited variable effect sizes, often linked to the ability of mixotrophic taxa to exploit nutrient pulses (López-Abbate et al., 2016; López-Abbate, 2021). Despite this, the short generation times and functional redundancy of microzooplankton confer resilience, allowing partial buffering of trophic disruptions and mitigating large-scale community collapse.

Hypersaline environments presented extreme selective pressures, favoring specialized physiological adaptations. Forest plots for hypersaline protists indicated highly positive effect sizes for osmoadaptive survival, consistent with known mechanisms such as compatible solute accumulation, proteome acidification, and enzymatic modification (Goh et al., 2006, 2011; Galinsky, 1993; Alexander et al., 2009; Edgcomb & Bernhard, 2013; Stock et al., 2011; Edgcomb et al., 2009). Genomic and single-cell approaches reveal that these habitats harbor extensive microbial dark matter, including novel archaeal lineages within TACK and DPANN superphyla, expanding our understanding of eukaryotic diversity and functional potential (Guy & Ettema, 2011; Castelle et al., 2015).

Across all stressors, the analysis supports a trait-based framework as the primary predictor of ecological responses. Warming responses were largely explained by metabolic plasticity and mixotrophy, acidification by calcification dependency, deoxygenation by oxygen tolerance thresholds, and hypersalinity by osmoadaptive strategies. The symmetry and distribution of effect sizes in funnel plots suggest that publication bias is minimal for warming and deoxygenation, while some asymmetry in acidification studies warrants cautious interpretation.

These findings have profound implications for predicting future oceanic ecosystem dynamics. Microzooplankton and hypersaline protists are not merely indicators of environmental change; they actively modulate carbon flux, nutrient recycling, and trophic interactions. As global change accelerates, the trait-mediated responses elucidated here can inform predictive models of microbial ecosystem restructuring, guide monitoring efforts in vulnerable regions, and prioritize taxa for experimental investigation. Importantly, integrating metabolic, physiological, and trophic perspectives offers a comprehensive approach to understanding microbial resilience and vulnerability under multiple concurrent stressors (Bindoff et al., 2019; Caron et al., 2012; Brock et al., 1994; Buitenhuis et al., 2010).

In conclusion, the synthesis underscores that microzooplankton and hypersaline protists respond to environmental hazards in complex but predictable ways, with functional traits, rather than taxonomic identity, serving as the dominant determinant of response magnitude and ecosystem impact. Forest and funnel plot analyses provide quantitative confirmation of these patterns and highlight both knowledge gaps and avenues for future research.

5. Limitations

Despite the comprehensive synthesis, this study has several limitations. First, the dataset relies heavily on published experimental and observational studies, which may introduce publication bias, particularly for acidification effects where null results are less frequently reported. Second, meta-analytical aggregation across diverse taxa, habitats, and experimental designs may obscure fine-scale species- or site-specific responses. The heterogeneity of methodological approaches—ranging from mesocosm experiments to in situ observations—limits the direct comparability of effect sizes, especially when assessing complex interactions among multiple stressors. Third, while forest and funnel plots provide quantitative insight into response patterns, they cannot fully capture temporal dynamics, evolutionary adaptation, or long-term acclimation potential of microzooplankton and hypersaline protists. Fourth, environmental data on synergistic effects of combined stressors (e.g., warming plus hypoxia or acidification) remain scarce, constraining predictions of cumulative impacts under realistic climate scenarios. Finally, genomic and molecular studies, while illuminating cryptic diversity, are currently limited to a subset of hypersaline and deep-sea habitats, leaving large knowledge gaps in global protistan diversity and functional roles. These limitations suggest that while the review offers robust general patterns, caution is warranted in extrapolating results to unstudied taxa or ecosystems.

6. Conclusion

Microzooplankton and hypersaline protists exhibit trait-dependent responses to global change stressors, with warming, acidification, deoxygenation, eutrophication, and hypersalinity driving differential effects on growth, grazing, and ecosystem function. Functional traits, rather than taxonomy, are key predictors of resilience and vulnerability. Integrating physiological, trophic, and genomic perspectives is essential for forecasting microbial contributions to ocean biogeochemistry under ongoing environmental change.

References