Reproductive toxicity and transgenerational effects of petroleum mixtures in fish

Crude oil is an extremely complex mixture, containing thousands of compounds, many of which remain unexplored. It is also highly toxic to marine life, posing a significant threat to fish species of both commercial and ecological importance. One of the main challenges for environmental scientists and policymakers is predicting the full impact of oil spills on fish populations. While modeling approaches are valuable for risk and impact assessments, they often focus on a single life stage or a single generation, missing the bigger picture of long-term, multigenerational effects. Emerging understanding of the complexity of oil compounds, their toxicity to not only early life stages, and the potential transfer of toxicity from parent to offspring suggests that current models may lack the necessary robustness and accuracy. In response to this need, the ToxiGen project aims to answer key questions: Does exposure to oil disrupt reproduction in adult fish? What specific compounds are most toxic, and how do they cause harm? Most importantly, do the effects of crude oil persist across multiple generations? Our research links molecular changes in parent fish with the compounds they absorb and examines how this affects the health and reproductive success of their offspring over three generations. In large-scale in vivo studies conducted on arctic keystone species cod and polar cod, we have discovered significant disruptions in key reproductive organs, both at molecular and physiological levels, leading to effects in early life stages of the first generation. Our team has also successfully developed a new analytical method to identify the chemical structures bioaccumulated in the brain, liver, and gonads of these fish. Results show a similar proportion of compounds across tissues, with naphthalenes being the most prevalent, followed by alkylated benzenes, cyclic monoaromatics, and, finally, multicyclic aromatics and PAHs. For each of these classes, the most abundant compounds have been identified and will undergo toxicity testing using tissue slice cultures. Ongoing analyses reveal significant effects on the brain, liver, and gonads, with multi-omics data integration expected to provide a comprehensive view of the pathways involved in the observed toxicity and contribute to understanding the risks for future generations. Preliminary data from ex vivo exposures of cod tissue to a crude oil water-accommodated fractions (WAF) and various WAF extracts suggest the potential importance of the resin fraction, above that of the well-known and toxic PAH fraction. These results are further supported by behavioral changes observed in zebrafish larvae exposed to the same oil extracts. Upcoming studies, using the zebrafish model, will explore if and how these toxic effects might be passed on to future generations.

A fundamental and enduring challenge for ecosystem managers developing risk and impact assessment tools is to predict the extent of impact that spilled oil can have on the immediate fish populations and on future generations. Modelling approaches are important tools for these assessments, but they currently suffer from two major shortcomings: 1) the failure to acknowledge that effects can be transferred through several generations and 2) the assumption that a small group of toxic compounds, polycyclic aromatic hydrocarbons, can be used as chemical proxy for exposure to complex petroleum mixtures. Emerging understanding within petroleum ecotoxicology now suggests that model outputs lack robustness. Up until now studies have not been able to determine the effects over multiple generations, and to properly identify the causative agents within petroleum. ToxiGen is an interdisciplinary study that will determine the effects and toxic mechanisms of petroleum on the reproductive success of fishes and subsequent generations. Our ambition is to identify the compounds responsible for alterations in reproductive success in adults and survival and fitness of the progeny. We will work with Atlantic cod and polar cod that are of high ecological and commercial importance, respectively. Zebrafish will be used for high-risk and generational experiments. We will work at the frontier of analytical chemistry to characterize the toxic fractions of petroleum. To disentangle mechanisms of toxicity, novel methods such as micro-injecting compounds into eggs will be used. We will combine different approaches to identify new biomarkers of exposure and effects. The new knowledge will then be integrated into reproductive adverse outcome pathways to be available through an internationally harmonized knowledgebase. Ultimately, ToxiGen will provide environmental managers with knowledge and data for models, and methods to detect and monitor the presence and impact of these complex mixtures in situ.