Project Overview

Overview: Dr. Ed Chesney at the Louisiana Universities Marine Consortium (LUMCON) was awarded an RFP-II grant at $657,945 to conduct the RFP-II project titled “Defining Ecologically Relevant Sublethal Effects: How do Low Levels of Exposure to Oil and Dispersants Affect Performance and Survival of Larvae of Gulf Nekton?”. The project consisted of 1 institution (LUMCON), 1 principal investigator (Chesney), 1 co-PI (Portier), 2 post docs (Duffy, Webb), 1 LSU graduate student (Saal), and participation by several research staff and undergraduate students.   The goal of the proposed project was to address Theme III.  We defined ecologically relevant outcomes for the response of the early life history stages of GOM fish and invertebrate species to exposures of lethal and sub-lethal levels of oil under conditions that mimic natural scenarios in play during DWH.  The objective was to provide a basis for better understanding impacts related to DWH for ecologically and economically important coastal species of nekton.  It is likely that the most significant direct impacts of DWH oil releases for fisheries of the nGOM were impacts to the early life stages of spring and summer spawning fishes and invertebrates that reside in the offshore to inshore regions of Louisiana and Mississippi where the most abundant and valuable Gulf of Mexico fisheries exist.  Many important species were affected because spring and summer is the principal spawning period for most species.  In spite of the challenge of studying the ELH stages of fishes, it is critically important to understand environmental impacts to this life stage because embryonic and larval fish often exhibit greater sensitivity to xenobiotic compounds than do adult fishes.  

The area most impacted by oil are also the most productive waters of the GOM and characterized by the very high primary productivity (i.e. the fertile-crescent) with coastal condition that range from oligotrophic (offshore bluewater) to eutrophic (inshore) plus high turbidity zones from river-borne sediments that are easily re-suspended by tides and wind in shallow coastal waters.  Although larvae and juveniles were the most heavily impacted life stages of nekton they are also the most difficult to assess at the population level because of their naturally high mortality rates from predation and other factors.  The backbone of modern toxicological methods is to focus on the repeatability, traceability of exposure levels and the ability for intra- and interspecies comparisons of the lethal toxicity of various toxic compounds including crude oil.  While this provides a standard for comparisons of toxic compounds, it does not provide a means to easily account for specific environmental conditions or the behavioral effects that can alter larval survival under the real world conditions that occur during a spill.

 

During an oil spill exposure levels vary over a large part of the environment, many biota are exposed for short durations and many exposures are below lethal concentrations as toxicity is dispersed and oil degrades.  Especially important ecologically relevant kinematics that can be affected by sub-lethal exposures include prey search behavior (swimming), prey capture and predator avoidance behaviors.  Typical laboratory exposures also do not account for the effects of the presence of phytoplankton and suspended sediments on the toxicity of oil and dispersants.  In order to get around some of these limitations, we conducted site relevant studies with native species under conditions that mimic natural exposure scenarios to provide a better understanding of the effects of the oil spill on early life stages of marine organisms.  We tested 5 prominent species, evaluating both lethal and sub-lethal effects of oil on marine larvae:

 

We established 24 hour LC50s with marine larvae (bay anchovy,  blue crab, stone crab, red drum and, red snapper) exposing them to CEWAFs (chemically enhanced water accommodated fractions) HEWAFs (high energy water accommodated fractions) and/or WAFs (water accommodated fractions) of crude oil as a baseline for sub-lethal tests

1. We conducted sublethal exposures of larvae to HEWAFs with different exposure durations at both lethal and sub-lethal concentrations
2. We conducted experiments with aged aged/dispersed oil (CEWAFs) and evaluate the degree that toxicity is moderated by aging at different salinities
3. We conducted separate toxicity experiments with and without phytoplankton added to the test waters to evaluate the effects of the presence of plankton on the toxicity of HEWAFs
4. We evaluated post exposure bay anchovy larval performance (feeding, growth, swimming) and used those performance responses to quantify sub-lethal effects to different crude oil concentrations, exposure durations and exposure conditions.

 

The array of experiments outline above were designed to provide a better basis for predicting impacts to larvae exposed to crude oil from DWH in the nGOM.  Some of the results of these studies will be used to make projections of sub-lethal effects on the ELH stages of the studied biota based upon their performance in response to contact with crude oil.  These results will allow managers to better assess impacts from DWH as well as develop strategies to reduce or minimize impacts to ELH stages of fish from future oil releases.

 

Research Highlights

Dr. Chesney’s research, which included 2 outreach products and activities, resulted in 1 peer-reviewed publications and 6 datasets being submitting to the GoMRI Information and Data Cooperative (GRIIDC), which are available to the public. Significant outcomes of their research (all related to GoMRI Research Theme III) are highlighted below.

 

Year 1 (2013):  In year 1, we ran 41 exposure experiments with 4 different species of larvae (focused primarily on two species, bay anchovy and blue crab) in an effort to compare responses among different species of larvae to oil exposure.  We exposed fish larvae and zoea to oil in different forms that are relevant to the DWH oil spill.  Multiple early life-stages of larvae were exposed to low energy water- accommodated fractions (WAF) of Macondo surrogate oil and chemically-enhanced WAFs, (CEWAF), and a dispersant (Corexit 9500).  No mortality was observed in 24 or 48 hour WAF exposures made up in 20 psu seawater, but CEWAF exposures induced different rate of mortality between species and among different life stages of a single species beyond the typical dose-dependent responses.  Comparable-age bay anchovy larvae were nearly seven times more sensitive to CEWAF than blue crab zoea.  Bay anchovy larvae demonstrated similar patterns of sensitivity to CEWAF between 5 dph and 20 dph, with sensitivity to oil less for older better developed larvae.  Further, CEWAF produced with 0 psu water was considerably more toxic to larvae than CEWAFmade up in 25 psu seawater for all species.  Blue crab zoea proved to be more sensitive to dispersants than fish larvae.

 

The establishment of baseline exposure mortality data allowed us to determine WAF and CEWAF concentrations that could potentially induce changes in behavior and development in early life stages of larvae without acute effects on survival.  We later used those results to study sub-lethal effects of relatively brief (24 hr) oil exposure over a longer time period during early development.

 

Twenty four hour sub-lethal exposures of bay anchovy and subsequent growout after transfer to oil free water showed that growth and survival can be compromised over a periods of several days.  These results show that sub-lethal exposures that occurred much more widely than lethal exposures during DWH can have significant effects on cohorts of early life history stages of fishes and invertebrates through effects on growth and delayed mortality effects.  We published those results in 2016.

 

Year 2 (2014):  In 2014, we attended GOMRI conference in Mobile where we presented 2 posters and 1 oral presentation.

 

The project postdoc departed in September 2014 and was not replace until January 2015, so progress in 2014 was affected by Dr. Duffy’s departure.  The new postdoc (Dr. Sarah Webb) began to pick up where Dr. Duffy left the project.   In an effort to standardize our exposure methods, we began to conduct the bulk of our exposures with HEWAFs.  This had several advantages including providing a means to compare our data to the vast amount of exposure data that was being generated by the NRDA process related to the Deepwater Horizon oil spill.  It also provided much more consistent and predictable exposure conditions so it reduced the need to repeat experiment plagued by a high degree of variability in the exposure fluids.

 

In the fall of 2014 we conducted a series of exposures with red drum embryos, plus 6 DPH and 21 DPH larvae.  These experiments showed that red drum are a good candidate for comparative exposures with fish larvae and provided baseline exposure results.

 

At the end of 2015, we ran a series of experiments with phytoplankton-rotifer mixes to look at the effects of plankton on partitioning of oil.  GCMS analyses were completed with these preliminary tests (although final oil concentrations were later recalculated). We found that oil was retained at higher rates in exposures with plankton present than without.

 

Year 3 (2015):  Dr. Chesney attended the annual GOMRI conference in Houston where he gave an oral presentation on toxicity of oil to fish larvae and co-authored 1 poster on bay anchovy transcriptome with Dr. Neigels group.

 

In 2015, we conducted numerous plankton-no plankton oil exposures with bay anchovy embryos and yolksac larvae in an effort to understand how the presence of plankton affects the loss of oil components during an exposure because early exposures with plankton showed that non feeding stages of bay anchovy larvae are just as sensitive to oil exposure with plankton present as feeding stages.  Our initial hypothesis was that intake of oil for larvae might be through feeding along with contact exposure.

 

A graduate student from Louisiana State University (Erin Saal, M.S.) completed the analytical chemistry from experiments designed to determine where oil compounds are sequestered during HEWAF exposures (i.e. sticking to beaker walls, plankton uptake or biotransformation, retained in the water, volitalized).  This was an unfunded collaboration.  Erin was able to partition the oil’s fate during beaker exposures with plankton present, although it did not fully resolve our understanding of the “plankton” effect.   Erin defended her master’s thesis in December of 2015.

 

In 2015, we established an LC50 for red snapper embryos and larvae.  This established a HEWAF LC50 for another important Gulf species to the mix that we can directly compare to our other species.  We also opportunistically conducted additional exposure trials with stone crab zoea in order to complete range finding experiments and established an LC50 for 2 dph stone crab zoea.  This was not one of our original target species but provided data that could be compared to blue crab and other larvae.  We also established red drum broodstock during summer 2015 for generating eggs/larvae of a sciaenid for exposure experiments planned for the fall of 2015.  However, we lost those fish because of a pump failure so we were unable to complete further exposures of red drum larvae.

 

Late in 2015, we attempted to focus on the mechanism of the plankton effect by investigating the relative conditions of the plankton (phytoplankton and rotifers) at different exposure concentrations.  As part of that we established an LC50 for our rotifers stock to help us to better understand the plankton-no plankton responses to oil exposure. We also conducted toxicity testing with the phytoplankton we used in exposures, Nannochloropsis oculatus.  Dr. Webb attempted to optimize her methods for detecting live and dead phytoplankton during oil exposures.  Dr. Webb attended and presented the results of her work at the national SETAC meeting in Utah, November 2015.

 

Year 4 (2016):   Dr. Webb departed for a teaching professorship on January 12th 2016.   Dr. Chesney presented the results of Dr. Webb’s work at the annual GOMRI conference in Tampa Florida.  We also lost the research assistant that had worked on the project since the beginning.  Mr. Childress departed for graduate school at the end of 2015.  Those departures forced us to rethink the final year of the project.  We hired two research assistants in March and May 2016 to complete the research with increased participation in experiments by Dr. Chesney.

 

In 2015 we detected a problem with the early GCMS data from the project.  This created some a problem with submitting data to complete datasets to GRDCII.  Once those issues were resolved, data from the early experiments (2013-2014) were revised and submitted to the GOMRI GRDIIC database.  We published a paper based upon those data (after they were revised) on lethal and sublethal scenarios of exposure of bay anchovy larvae to crude oil.  A PDF of that publication was submitted to GoMRI as part of a quarterly report.

 

Dr. Webb returned for two weeks in June 2016 and continued her experiments to look at the effect of plankton on oil dynamics during exposures.

 

Progress ramped up in the 2nd quarter of 2016 as we completed personnel training and experiments were underway again.  The highlights of the 2016 research were a series of extremely short (2 and 6 hr) exposures of bay anchovy embryos and 1 dph larvae.  These studies were to evaluate how short duration exposures affect both lethal and non-lethal outcomes of oil exposure.  Those results showed that extremely short duration exposure, although not highly lethal during a short exposure, could prove to be lethal (post exposure) or damaging to newly hatched larvae.

 

The other highlight of the 2016 research was the sublethal exposures of 3 DPH bay anchovy larvae and the effects of those low dose, short duration (2 h) exposures on their swimming performance.  These studies showed that the swimming ability of first feeding bay anchovy larvae are significantly affected by low dose, short duration exposure.

 

Late in the project, we developed a collaborative relationship with another group of researchers that wanted us to conduct single compound exposures with our principal test species, the bay anchovy.  Once we had worked out the methodology, we began a series of single compound exposures with bay anchovy embryos and larvae.  We had some execution challenges with this new type of experiment but results of toluene exposures were good and those data are available on the GRCII database.

 

Dr. Chesney submitted an abstract to the early Life history chapter meeting of the American Fisheries Society entitled: Toxicological testing with an estuarine icon, the bay anchovy, Anchoa mitchilli.  The talk was accepted for an oral presentation that he gave on June 23rd, 2016 in Solomons, Md.  He gave a similar talk at the 2017 GoMRI meeting in NOLA.

 

Summary results to date and scientific highlights

 

1. Our 24 hr sub-lethal exposures of bay anchovy and subsequent growout after transfer to oil free water showed that growth and survival can be compromised over a periods of several days after exposure.  The significance of those results is that exposure conditions believed to be sub-lethal can in fact be lethal just a few days post exposure.  If lethal exposure levels from laboratory LC50s are used to predict outcomes in the field, then delayed mortality from supposedly sublethal exposures would underestimate environmental impacts to marine larvae during oil spills.  That suggests that low dose exposures should be better evaluated in the future, especially for sensitive fish and invertebrate larvae. We published those results in 2016.
   
   
2. A highlight of the 2015 research was that the presence of marine plankton (rotifers and phytoplankton) in exposure waters can affect the outcomes of exposures with marine larvae.  The main effect seems to be related to higher concentrations of oil being retained in the exposure fluids when plankton are present but only in exposures at lower concentrations.  The significance of this research is that exposures of marine larvae are normally done without plankton present although plankton is present in coastal environments.  The significance of these results may be primarily for oil exposure methods with larvae in the lab, especially at low exposure doses.
   
   
3. The highlights of the 2016 research was that a series of extremely short (2 and 6 hr) exposures of bay anchovy embryos and 1 dph larvae can affect both lethal and non-lethal outcomes of oil exposure.  The other highlight of the 2016 research was the sublethal exposures of 3 DPH bay anchovy larvae and the effects of those low dose, short duration (2 h) exposures on their swimming performance.  All levels of exposure reduced swimming speeds of first feeding bay anchovy larvae. These results showed that extremely short duration exposures, although not always lethal within the exposure time, can prove to be lethal (post exposure) or damaging to first feeding larvae.  They also showed that even at sublethal doses of short exposure the swimming performance of larvae can be compromised. These results should have consequences for how we conduct exposures with highly sensitive life stages or species in the future.
   
   
4. A highlight of the entire projects results is that when we exposed a number of species of marine larvae (a total of 5 species of vertebrate and invertebrate larvae) under similar conditions and at similar stages of development our results showed that there was a lot of variability in their sensitivity to crude oil and dispersants. The significance of those results is that there is a lot of variability in sensitivity to these substances that is difficult to predict without toxicological testing.