This collaborative research project examined the physiological and molecular effects of Louisiana Sweet Crude Oil in the killifish model system.  Killifish is a well-described, out-bred vertebrate model that exhibits population differences in hydrocarbon sensitivity.  Following the Deepwater Horizon oil spill (DHOS), large expanses of the northern Gulf of Mexico were heavily oiled, particularly during the reproductive season of the Gulf killifish.  There were two major interconnected aims of this research project.  The first aim was to screen populations of Gulf killifish embryos for variability in sensitivity to Louisiana Sweet Crude Oil.  This aim exploited the fact that differences in oil/hydrocarbon sensitivity exist in this and other closely-related fish species, and that differences in physiological/genomic responses to oil between populations could provide insights into the mechanisms of oil toxicity in developing vertebrates.  The second aim was to test for impacts, and identify molecular mechanisms of early-life and adult oil exposures on: early life stage development; early life stage performance; and adult performance and reproduction.  This final aim was extended to include oil spill impacts that span generations.

In early work in my laboratory, we demonstrated that coastal fish along the northern Gulf of Mexico had been exposed for an extended period to bioactive concentrations of oil.  Furthermore, oiling in the environment occurred during the reproductive season of the Gulf killifish.  Given the possibility that Gulf killifish were exposed chronically in the field to crude oil, studies were then conducted to evaluate the long-term impacts of oil in this species.  During 2014 and 2015, Gulf killifish were collected from five sites ranging from Texas to Florida.  Three of these populations were from sites in Florida, Louisiana, and Texas with no history of hydrocarbon exposure, and two of the populations were from polluted sites, including an U.S. EPA Superfund Site within the Houston Ship Channel and another from a site in Louisiana very heavily oiled following the DHOS.  Embryos were obtained from the adult fish collected from each site and these embryos were exposed acutely to oil at 6 or more concentrations from fertilization to hatch to assess their relative sensitivity to oil during embryo development.  In brief, we observed a significant interaction between population and oil concentration.  Whereas embryos from the clean sites and from the oiled site in Louisiana were highly sensitive to oiling, the embryos from the Superfund site in Texas were refractory to most oil effects.   Furthermore, oiling of these adults in the laboratory reduced gamete quality and reduced the hatching success of progeny for all these populations except for the Texas population from the Superfund site.  These data suggest that the progeny of adult fish adapted to survival in the Superfund site in Texas is better able to tolerate subsequent oiling.  Furthermore, there is evidence that embryos derived from the previously oiled site in Louisiana may in fact be more sensitive to acute oiling.  

In the fall of 2015/spring 2016, we undertook three large experiments, which became the focus for much of the remaining research.  One of these experiments involved exposing adult killifish to clean or oiled water for 40-60 days, and then using these adults as a brood stock for the generation of an F1 generation of killifish.  We created four specific lineages of F1 progeny derived by crossing the gametes of control and oiled parents (i.e., F0 generation). The resulting F1 generation consisted of the four following lineages: control females x control males (CxC), exposed females x exposed females (ExE), control females x exposed males (CxE), and exposed females x control males (ExC).  The F1 progeny were grown for at least two years in clean water before they were used to generate an F2 generation of CxC; ExE; CxE; and ExC.  In 2017, once fish were reproductively active, we produced the F2 generation by crossing F1 females and F1 males of the lineage to create four lineages of F2 fish.  Following assessment of fertilization success, embryos from each lineage were randomly distributed into clean or oiled water.  Embryos were monitored over a 21-day period for heart rate, time to hatch, and hatching success, and embryos collected at several stages of development for transcriptomics and hatchlings sampled for transcriptomics and morphometric analysis.  First, there is clear evidence that the F0 oiling has significant effects in the F1 progeny, particularly in fish derived from oiled fathers.  F1 adults of paternally-oiled fish showed severe scoliosis, altered fin development, and discernible craniofacial deformities, including altered jaw development.  We have also demonstrated that F1 fish of paternally-oiled fish show impaired feeding efficiency and swimming capacity.  Oiling of F0 adults produces effects in the physiology and morphology of the F2 generation, despite the fact the F1 fish were held their entire lives in clean water.  These data support the potential of intergenerational impacts of crude oil in killifish.  

Last Modified: 10/19/2018
Modified by: Fernando Galvez