Transgenerational effects of parental crude oil exposure on the morphology of adult Fundulus grandis

Highlights

• Swimming fitness and two-dimensional landmark-based geometric morphometrics with multivariate generalized linear modeling were used to identify the transgenerational effects of crude oil exposure in adult gulf killifish.

• Statistical models demonstrate that paternal (F0) oiling rather than maternal (F0) oiling explains more of the variance in fish shape, including effects such as reduced body length and altered body depth and aberrant spinal curvature and altered angles of head projection relative to the body axis.

• Anatomical effects persisted in progeny for at least two generations despite progeny being raised continuously in clean water.

• The aerobic swimming performance of progeny derived from oiled fathers was also impaired for at least two generations.

Abstract

The current study involved exposing adult F0 Gulf killifish (Fundulus grandis) to Macondo-252 oil for 36 to 44 days and assessing the effects of this oiling on the swimming performance and morphology in two generations of progeny reared in clean water. Following exposure to oil, the F0 fish were used as broodstock to generate four lineages of F1 fish using a full-matrix mating design derived from the gametes of clean and oil-exposed parents. Later, the four lineages of F1 fish were used as broodstock to create an F2 generation of the same four lineages. We found few differences in embryonic outcome (% dead,% hatched, and% unhatched) in any of the four lineages of F1 and F2 fish. However, as adults, F1 and F2 fish derived from oil-exposed males from the F0 generation had significantly lower critical swimming speeds (U_crit) than both the control and maternally oil-exposed lineages. Additionally, progeny of oil-exposed fish had altered body shape based on the statistical analysis of two-dimensional landmark-based geometric morphometrics. Fish from oil-exposed lineages showed increased body depth, altered spinal curvature, and changes in the upward angle of projection of the head. Both generations had a significant main effect of maternal and paternal oil exposure on shape; however, F0 paternal oil exposure explained more of the variance in shape across both generations relative to F0 maternal exposure. Our findings demonstrate that parental exposure to oil can impact the shape and aerobic swimming capacity of offspring for at least two generations after the original paternal oiling.

Introduction

The 2010 Deepwater Horizon oil spill resulted in the release of an estimated 507 million liters of crude oil into the northern Gulf of Mexico (nGoM) and oiled 2100 km of coastline (Deepwater Horizon Natural Resource Damage Assessment Trustees, 2016). Like with other large marine oil spills, this 2010 spill prompted considerable research to characterize the biological effects of acute crude oil exposure in aquatic animals (Incardona, 2017). In fish, crude oil toxicity has been attributed to the induction of impaired cardiovascular development in embryos (Incardona et al., 2006, 2004; Jung et al., 2013; Khursigara et al., 2017; Perrichon et al., 2021; Sørhus et al., 2016; Xu et al., 2017). Other acute studies have also shown that oil exposure can produce teratogenic effects such as abnormal craniofacial, spinal, and muscular development in both freshwater and marine teleosts (de Soysa et al., 2012b; Incardona et al., 2014; Incardona and Scholz, 2016; Philibert et al., 2019). Despite the importance of these studies, little is known about the long-term consequences of abnormal embryonic or larval development to the morphology and fitness of animals late in life.

Although oil is most acutely toxic in fish during the embryonic and larval life stages, the tissues of adult fish exposed to oil show altered gene expression patterns suggestive of impaired reproduction (Pilcher et al., 2014; Whitehead et al., 2012). These studies suggest the potential for persistent impacts of transient oil exposure, including effects that may span generations. Given our growing understanding of epigenetic modes of inheritance in fish (Labbé et al., 2017), there has been increasing interest in the transgenerational effects of crude oil exposure. Recent studies on Macondo oil from the Deepwater Horizon oil spill have begun to characterize the multigenerational impacts of parental crude oil exposure (Bautista and Burggren, 2019; Jasperse et al., 2019a, 2019b; Vignet et al., 2015). Traditionally, most studies have investigated the effects of both parental lineages on transgenerational inheritance or have preferentially focused solely on the maternal input. Collectively, these studies have shown somatic, developmental, behavioral, and reproductive effects when both parents are exposed to oil (Bautista and Burggren, 2019; Jasperse et al., 2019a, 2019b). However, there is growing interest in the role of paternal modes of epigenetic inheritance (Bautista et al., 2020). In zebrafish (Danio rerio), it was found that the methylome is responsible for epigenetic gene regulation, and that inheritance occurs via the sperm and not the oocyte  (Jiang et al., 2013). Overall, these studies demonstrate the persistence of altered pheno- and epigenotypes across multiple generations, although additional work is needed to assess the maternal versus paternal modes of transgenerational inheritance. Also, assessments of biological effects in progeny should be extended to include adult phenotypes and other fitness level endpoints.

Traditional assessments of commonly observed adult phenotypes such as altered spinal and craniofacial development have been conducted in a presence-absence manner in oil toxicity studies (de Soysa et al., 2012a; Hansen et al., 2019; Raimondo et al., 2014). However, these studies do not account for the variance in total body shape for a treatment group. An alternate approach that would allow for the identification of fine-scale changes in body shape across treatments would be through the usage of landmark-based geometric morphometrics (GM). In past studies, landmark-based GM has been used to identify patterns of altered morphology in response to marine pollution in wild populations of fish (Gharred et al., 2020), bivalves (Reyna et al., 2019; Scalici et al., 2017), and gastropods (Primost et al., 2016). Only a few laboratory studies have utilized GM techniques to assess the effects of aquatic toxicants in fish; although morphology has been shown altered by PCB or TCDD exposure in fathead minnows (Coulter et al., 2019) and zebrafish (Chollett et al., 2014), respectively. Although two-dimensional (2D) GM application to environmental toxicity is less common in the context of transgenerational studies, this technique could provide a powerful approach to identify subtle changes in animal structure due to toxicant exposure.

In the current study, we investigated the effects of parental crude oil exposure on adult offspring for two generations using an assessment of swimming performance and GM techniques to quantify variation in body shape changes. The ubiquitous nGoM marsh resident, the Gulf killifish (Fundulus grandis), was used for this study as it is an ecologically relevant model. The Gulf killifish was one of the many nGOM marsh species that experienced oiling during its peak spawning period following the Deepwater Horizon oil spill. Male and female Gulf killifish were exposed for 36–44 days to sublethal concentrations of Macondo-252 light sweet crude oil. These fish were then used to generate one of four lineages of F1 fish using a full-matrix mating design with gametes from control and oil-exposed F0 adults. The F1 generation was not exposed to oil directly at any time following fertilization. This F1 generation was then used to create an F2 generation of fish that also was never exposed directly to oil. We noticed upon maturation that morphological abnormalities were present. For this reason, we assessed the aerobic swimming capacity for both F1 and F2 fish. We additionally conducted ad hoc landmark-based GM and subsequent multivariate generalized linear models (GLM) to determine the effects of parental crude oil exposure on (1) fish morphology and (2) patterns of maternal or paternal transmission of altered phenotypes in two generations.