Periodic Reporting for period 2 - MitoRescue (Role of mitochondria in mediating evolutionary rescue by anthropogenic pollutants in an invasive crab)
Reporting period: 2025-07-01 to 2026-06-30
Summary of the context and overall objectives of the project
Invasive alien species (IAS) are a major driver of biodiversity loss and generate substantial ecological and socio-economic costs worldwide. Their impacts are expected to become increasingly difficult to manage under global environmental change, as climate warming and pollution alter both ecosystem conditions and the capacity of species to establish, persist and spread. Some invasive populations display a remarkable capacity to rapidly adjust to novel environmental conditions, yet the physiological and cellular mechanisms underlying this adaptive potential remain poorly understood. Identifying these mechanisms is essential to improve our capacity to anticipate biological invasions and develop more effective monitoring, risk-assessment and management strategies.
MitoRescue addressed this knowledge gap by investigating the role of mitochondria in the rapid adaptation and “evolutionary rescue” of invasive species. Mitochondria are central to cellular energy production and redox regulation and are highly responsive to environmental stress, making mitochondrial plasticity a potentially important mechanism allowing organisms to maintain performance under rapidly changing environmental conditions. Using the European green crab Carcinus maenas, one of the world's most successful marine invaders, MitoRescue investigated how ancestral and recently established invasive populations respond to the combined effects of warming and anthropogenic pollution. The project integrated field and laboratory approaches across multiple levels of biological organisation, combining whole-organism physiology, biochemical and redox responses, high-resolution mitochondrial respirometry, mitochondrial imaging and transcriptomics.
The project has generated new evidence supporting mitochondrial plasticity as an important mechanistic component of adaptive responses to environmental change, extending the concept of evolutionary rescue beyond classical population-genetic approaches. It has also revealed marked sex-specific differences in mitochondrial physiology, redox homeostasis and biochemical responses, highlighting the importance of considering biological sex when evaluating the adaptive capacity of natural populations. In parallel, MitoRescue identified candidate mitochondrial and bioenergetic biomarkers with potential for assessing stress tolerance and adaptive potential and developed methodological advances for mitochondrial phenotyping, including optimised permeabilised-tissue approaches for high-resolution respirometry in marine crustaceans.
These results establish a pathway from fundamental mechanistic knowledge towards improved environmental assessment and management. By identifying physiological traits associated with the capacity to tolerate multiple environmental stressors, MitoRescue provides a scientific basis for the future development of biomarkers that could help identify invasive populations with enhanced adaptive potential and support more mechanistically informed ecological risk assessment. Such approaches could ultimately contribute to earlier and more targeted management interventions and a more efficient allocation of monitoring resources, helping reduce the ecological and economic impacts of biological invasions on fisheries, aquaculture and coastal ecosystems.
The project's impact extends beyond its immediate scientific findings. Advanced expertise in mitochondrial physiology, high-resolution respirometry, tissue permeabilisation, imaging and transcriptomic approaches acquired through the international fellowship has been transferred to European research institutions and is now being applied in ongoing and future research. The international collaborations established through MitoRescue are continuing beyond the fellowship and have already generated joint publications, research initiatives and opportunities for future funding.
In the longer term, MitoRescue contributes to the European objective of protecting and restoring biodiversity by improving understanding of the mechanisms that determine the establishment and persistence of invasive species under climate change and pollution. Its results are relevant to the EU Biodiversity Strategy for 2030, as well as to the Water Framework Directive, Marine Strategy Framework Directive and the broader objectives of the European Green Deal concerning biodiversity conservation, pollution reduction and climate resilience. By linking mitochondrial mechanisms to organismal performance and adaptive capacity, MitoRescue provided new scientific knowledge and methodological capacity for anticipating biological responses to environmental change and supporting evidence-based management of aquatic ecosystems.
MitoRescue addressed this knowledge gap by investigating the role of mitochondria in the rapid adaptation and “evolutionary rescue” of invasive species. Mitochondria are central to cellular energy production and redox regulation and are highly responsive to environmental stress, making mitochondrial plasticity a potentially important mechanism allowing organisms to maintain performance under rapidly changing environmental conditions. Using the European green crab Carcinus maenas, one of the world's most successful marine invaders, MitoRescue investigated how ancestral and recently established invasive populations respond to the combined effects of warming and anthropogenic pollution. The project integrated field and laboratory approaches across multiple levels of biological organisation, combining whole-organism physiology, biochemical and redox responses, high-resolution mitochondrial respirometry, mitochondrial imaging and transcriptomics.
The project has generated new evidence supporting mitochondrial plasticity as an important mechanistic component of adaptive responses to environmental change, extending the concept of evolutionary rescue beyond classical population-genetic approaches. It has also revealed marked sex-specific differences in mitochondrial physiology, redox homeostasis and biochemical responses, highlighting the importance of considering biological sex when evaluating the adaptive capacity of natural populations. In parallel, MitoRescue identified candidate mitochondrial and bioenergetic biomarkers with potential for assessing stress tolerance and adaptive potential and developed methodological advances for mitochondrial phenotyping, including optimised permeabilised-tissue approaches for high-resolution respirometry in marine crustaceans.
These results establish a pathway from fundamental mechanistic knowledge towards improved environmental assessment and management. By identifying physiological traits associated with the capacity to tolerate multiple environmental stressors, MitoRescue provides a scientific basis for the future development of biomarkers that could help identify invasive populations with enhanced adaptive potential and support more mechanistically informed ecological risk assessment. Such approaches could ultimately contribute to earlier and more targeted management interventions and a more efficient allocation of monitoring resources, helping reduce the ecological and economic impacts of biological invasions on fisheries, aquaculture and coastal ecosystems.
The project's impact extends beyond its immediate scientific findings. Advanced expertise in mitochondrial physiology, high-resolution respirometry, tissue permeabilisation, imaging and transcriptomic approaches acquired through the international fellowship has been transferred to European research institutions and is now being applied in ongoing and future research. The international collaborations established through MitoRescue are continuing beyond the fellowship and have already generated joint publications, research initiatives and opportunities for future funding.
In the longer term, MitoRescue contributes to the European objective of protecting and restoring biodiversity by improving understanding of the mechanisms that determine the establishment and persistence of invasive species under climate change and pollution. Its results are relevant to the EU Biodiversity Strategy for 2030, as well as to the Water Framework Directive, Marine Strategy Framework Directive and the broader objectives of the European Green Deal concerning biodiversity conservation, pollution reduction and climate resilience. By linking mitochondrial mechanisms to organismal performance and adaptive capacity, MitoRescue provided new scientific knowledge and methodological capacity for anticipating biological responses to environmental change and supporting evidence-based management of aquatic ecosystems.
Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far
MitoRescue investigated the role of mitochondrial plasticity in the capacity of invasive species to cope with environmental change, using the European green crab Carcinus maenas as a model. The scientific programme combined field sampling and controlled laboratory experiments with analyses spanning multiple levels of biological organisation, from whole-organism performance and biochemical responses to mitochondrial function, morphology and gene expression. Particular attention was given to the combined effects of temperature and anthropogenic pollution and to differences between ancestral and recently established invasive populations.
During the first part of the project, ancestral and invasive populations of C. maenas originating from contrasting environmental conditions were characterised and experimentally exposed to different temperatures and levels of pollution. Whole-organism physiological performance was integrated with biochemical and redox biomarkers to determine whether invasive populations display physiological characteristics that may contribute to enhanced tolerance of environmental stress. These experiments generated a comprehensive comparative dataset on population-specific responses to warming and pollution and fulfilled the first scientific objective of the project.
The project subsequently investigated mitochondrial function as a potential mechanism underlying these differences in environmental tolerance. High-resolution respirometry was used to characterise mitochondrial bioenergetics and determine how environmental exposure to pollutants modifies mitochondrial performance. A major technical component of this work was the optimisation of tissue-permeabilisation approaches for crustacean gills, allowing mitochondrial respiratory function to be measured directly in intact permeabilised tissues. This approach preserves tissue organisation and provides an alternative to conventional mitochondrial isolation. The methodological work also revealed functionally distinct mitochondrial phenotypes between different crab gill types, highlighting previously underappreciated mitochondrial specialisation within this organ.
During the return phase, the mitochondrial phenotype was further characterised through complementary imaging and transcriptomic approaches. All planned biological material was collected and processed, and quantitative imaging analyses provided a comprehensive characterisation of mitochondrial morphology across experimental groups. RNA extraction, library preparation and high-throughput sequencing were also completed for all selected samples, generating the transcriptomic dataset required to investigate differential gene expression and population-level genetic variation. Together, these datasets enable mitochondrial functional, structural and molecular responses to be examined within a common experimental framework.
A major scientific outcome of MitoRescue is the establishment of an integrated, multi-scale framework linking organismal performance to mitochondrial bioenergetics, redox regulation, morphology and gene expression. The results provide evidence that mitochondrial plasticity represents an important mechanistic component of responses to environmental change and support its consideration in the context of evolutionary rescue and invasion biology. The project thereby extends the study of evolutionary rescue beyond predominantly population-genetic approaches by identifying physiological and cellular mechanisms that may contribute to the persistence of populations exposed to rapidly changing environmental conditions.
Another important outcome was the identification of marked sex-specific differences in mitochondrial physiology, redox homeostasis and biochemical responses. These findings demonstrate that biological sex can substantially influence physiological responses to environmental stress and should therefore be explicitly considered when evaluating the adaptive capacity of natural populations.
Finally, MitoRescue identified candidate mitochondrial and bioenergetic biomarkers associated with environmental stress tolerance and adaptive potential. These results provide a foundation for future work assessing whether mitochondrial phenotypes can contribute to predicting the vulnerability or adaptive capacity of populations exposed to multiple environmental stressors. Although developed using C. maenas, the methodological and conceptual framework has potential application to other aquatic organisms and to broader questions in environmental physiology and ecotoxicology.
Overall, the project successfully generated the experimental and methodological framework required to address its central hypothesis and produced complementary physiological, biochemical, mitochondrial, imaging and transcriptomic datasets. All planned experimental activities were completed. Technical difficulties experienced by the external sequencing provider delayed completion of the transcriptomic component, as several samples required resequencing to achieve the contracted sequencing depth; however, this did not compromise the quality or completeness of the final dataset. The resulting integrated dataset provides a mechanistic basis for understanding how mitochondrial plasticity may contribute to environmental tolerance and evolutionary rescue in invasive populations.
During the first part of the project, ancestral and invasive populations of C. maenas originating from contrasting environmental conditions were characterised and experimentally exposed to different temperatures and levels of pollution. Whole-organism physiological performance was integrated with biochemical and redox biomarkers to determine whether invasive populations display physiological characteristics that may contribute to enhanced tolerance of environmental stress. These experiments generated a comprehensive comparative dataset on population-specific responses to warming and pollution and fulfilled the first scientific objective of the project.
The project subsequently investigated mitochondrial function as a potential mechanism underlying these differences in environmental tolerance. High-resolution respirometry was used to characterise mitochondrial bioenergetics and determine how environmental exposure to pollutants modifies mitochondrial performance. A major technical component of this work was the optimisation of tissue-permeabilisation approaches for crustacean gills, allowing mitochondrial respiratory function to be measured directly in intact permeabilised tissues. This approach preserves tissue organisation and provides an alternative to conventional mitochondrial isolation. The methodological work also revealed functionally distinct mitochondrial phenotypes between different crab gill types, highlighting previously underappreciated mitochondrial specialisation within this organ.
During the return phase, the mitochondrial phenotype was further characterised through complementary imaging and transcriptomic approaches. All planned biological material was collected and processed, and quantitative imaging analyses provided a comprehensive characterisation of mitochondrial morphology across experimental groups. RNA extraction, library preparation and high-throughput sequencing were also completed for all selected samples, generating the transcriptomic dataset required to investigate differential gene expression and population-level genetic variation. Together, these datasets enable mitochondrial functional, structural and molecular responses to be examined within a common experimental framework.
A major scientific outcome of MitoRescue is the establishment of an integrated, multi-scale framework linking organismal performance to mitochondrial bioenergetics, redox regulation, morphology and gene expression. The results provide evidence that mitochondrial plasticity represents an important mechanistic component of responses to environmental change and support its consideration in the context of evolutionary rescue and invasion biology. The project thereby extends the study of evolutionary rescue beyond predominantly population-genetic approaches by identifying physiological and cellular mechanisms that may contribute to the persistence of populations exposed to rapidly changing environmental conditions.
Another important outcome was the identification of marked sex-specific differences in mitochondrial physiology, redox homeostasis and biochemical responses. These findings demonstrate that biological sex can substantially influence physiological responses to environmental stress and should therefore be explicitly considered when evaluating the adaptive capacity of natural populations.
Finally, MitoRescue identified candidate mitochondrial and bioenergetic biomarkers associated with environmental stress tolerance and adaptive potential. These results provide a foundation for future work assessing whether mitochondrial phenotypes can contribute to predicting the vulnerability or adaptive capacity of populations exposed to multiple environmental stressors. Although developed using C. maenas, the methodological and conceptual framework has potential application to other aquatic organisms and to broader questions in environmental physiology and ecotoxicology.
Overall, the project successfully generated the experimental and methodological framework required to address its central hypothesis and produced complementary physiological, biochemical, mitochondrial, imaging and transcriptomic datasets. All planned experimental activities were completed. Technical difficulties experienced by the external sequencing provider delayed completion of the transcriptomic component, as several samples required resequencing to achieve the contracted sequencing depth; however, this did not compromise the quality or completeness of the final dataset. The resulting integrated dataset provides a mechanistic basis for understanding how mitochondrial plasticity may contribute to environmental tolerance and evolutionary rescue in invasive populations.
Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)
MitoRescue advances the state of the art by introducing mitochondrial plasticity as a mechanistic dimension in the study of evolutionary rescue and biological invasions. While evolutionary rescue has traditionally been considered predominantly from a population-genetic perspective, MitoRescue integrated organismal physiology, mitochondrial bioenergetics, redox biology, mitochondrial morphology and transcriptomics to investigate how physiological and cellular plasticity may contribute to population persistence under rapid environmental change. Using the invasive European green crab Carcinus maenas, the project established a multi-scale framework linking environmental stress, mitochondrial phenotype and organismal performance, representing one of the first comprehensive applications of this approach to an invasive marine invertebrate.
Among its main results, MitoRescue identified candidate mitochondrial and bioenergetic biomarkers associated with environmental stress tolerance and adaptive potential. It also revealed marked sex-specific differences in mitochondrial physiology, redox homeostasis and biochemical responses, highlighting biological sex as an important factor when assessing the adaptive capacity of natural populations. At the methodological level, the project optimised tissue-permeabilisation protocols for marine crustacean gills, enabling high-resolution measurements of mitochondrial respiration while preserving tissue organisation. Together with the imaging and transcriptomic approaches implemented during the project, these developments provide a methodological platform for investigating mitochondrial responses to environmental stress across complementary levels of biological organisation.
These results have potential applications extending beyond C. maenas. In the longer term, mitochondrial biomarkers could contribute to more mechanistically informed environmental monitoring and ecological risk assessment and help identify invasive populations with enhanced capacity to tolerate environmental change. Such information could support the prioritisation of monitoring and management efforts and a more efficient allocation of resources devoted to biological invasions, with potential benefits for fisheries, aquaculture and coastal ecosystem management.
Further research is required before these candidate biomarkers can be translated into operational monitoring or management tools. Key priorities are to validate relationships between mitochondrial phenotype, stress tolerance and adaptive potential across additional populations, species and environmental conditions; establish their predictive value under realistic field conditions; and determine the generality of the MitoRescue conceptual framework. The methodological developments will similarly benefit from further validation and standardisation across species and tissue types and from inter-laboratory comparisons to facilitate broader uptake.
Follow-up competitive funding will provide the main pathway for this validation and scaling-up. The scientific results, methodologies and international collaborations established through MitoRescue provide the foundation for future funding applications, including a planned ERC proposal. Such follow-up projects will enable the MitoRescue approach to be expanded beyond the original model system, test the generality and predictive capacity of the identified mitochondrial biomarkers, and ultimately bridge the gap between fundamental mitochondrial ecophysiology and applications in environmental assessment and management. The collaborations established during the fellowship are already continuing through joint research initiatives and publications, providing a strong international network through which this next research stage can be implemented.
At present, the principal exploitation pathway for MitoRescue is therefore scientific and methodological rather than commercial. Successful uptake will depend primarily on publication and open dissemination of results and datasets, further biological validation, methodological standardisation, continued international collaboration and follow-up research funding. The project has already strengthened European capacity in high-resolution mitochondrial phenotyping through the transfer and implementation of its methodologies at European partner institutions. In accordance with the project's Open Science strategy, definitive datasets will also be made publicly available alongside the corresponding publications, facilitating their independent validation and reuse.
In the longer term, these advances have the potential to improve understanding and assessment of the capacity of aquatic organisms to respond to interacting environmental stressors. By linking mitochondrial mechanisms with adaptive capacity under warming and pollution, MitoRescue provides knowledge relevant to European priorities for biodiversity conservation, invasive-species management, pollution assessment and climate resilience, including the EU Biodiversity Strategy for 2030, the Water Framework Directive, the Marine Strategy Framework Directive and the European Green Deal.
Among its main results, MitoRescue identified candidate mitochondrial and bioenergetic biomarkers associated with environmental stress tolerance and adaptive potential. It also revealed marked sex-specific differences in mitochondrial physiology, redox homeostasis and biochemical responses, highlighting biological sex as an important factor when assessing the adaptive capacity of natural populations. At the methodological level, the project optimised tissue-permeabilisation protocols for marine crustacean gills, enabling high-resolution measurements of mitochondrial respiration while preserving tissue organisation. Together with the imaging and transcriptomic approaches implemented during the project, these developments provide a methodological platform for investigating mitochondrial responses to environmental stress across complementary levels of biological organisation.
These results have potential applications extending beyond C. maenas. In the longer term, mitochondrial biomarkers could contribute to more mechanistically informed environmental monitoring and ecological risk assessment and help identify invasive populations with enhanced capacity to tolerate environmental change. Such information could support the prioritisation of monitoring and management efforts and a more efficient allocation of resources devoted to biological invasions, with potential benefits for fisheries, aquaculture and coastal ecosystem management.
Further research is required before these candidate biomarkers can be translated into operational monitoring or management tools. Key priorities are to validate relationships between mitochondrial phenotype, stress tolerance and adaptive potential across additional populations, species and environmental conditions; establish their predictive value under realistic field conditions; and determine the generality of the MitoRescue conceptual framework. The methodological developments will similarly benefit from further validation and standardisation across species and tissue types and from inter-laboratory comparisons to facilitate broader uptake.
Follow-up competitive funding will provide the main pathway for this validation and scaling-up. The scientific results, methodologies and international collaborations established through MitoRescue provide the foundation for future funding applications, including a planned ERC proposal. Such follow-up projects will enable the MitoRescue approach to be expanded beyond the original model system, test the generality and predictive capacity of the identified mitochondrial biomarkers, and ultimately bridge the gap between fundamental mitochondrial ecophysiology and applications in environmental assessment and management. The collaborations established during the fellowship are already continuing through joint research initiatives and publications, providing a strong international network through which this next research stage can be implemented.
At present, the principal exploitation pathway for MitoRescue is therefore scientific and methodological rather than commercial. Successful uptake will depend primarily on publication and open dissemination of results and datasets, further biological validation, methodological standardisation, continued international collaboration and follow-up research funding. The project has already strengthened European capacity in high-resolution mitochondrial phenotyping through the transfer and implementation of its methodologies at European partner institutions. In accordance with the project's Open Science strategy, definitive datasets will also be made publicly available alongside the corresponding publications, facilitating their independent validation and reuse.
In the longer term, these advances have the potential to improve understanding and assessment of the capacity of aquatic organisms to respond to interacting environmental stressors. By linking mitochondrial mechanisms with adaptive capacity under warming and pollution, MitoRescue provides knowledge relevant to European priorities for biodiversity conservation, invasive-species management, pollution assessment and climate resilience, including the EU Biodiversity Strategy for 2030, the Water Framework Directive, the Marine Strategy Framework Directive and the European Green Deal.