Genomic and physiological footprint of the Deepwater Horizon oil spill on resident marsh fishes

The biological consequences of the Deepwater Horizon oil spill are unknown, especially for resident organisms. Here, we report results from a field study tracking the effects of contaminating oil across space and time in resident killifish during the first 4 mo of the spill event. Remote sensing and analytical chemistry identified exposures, which were linked to effects in fish characterized by genome expression and associated gill immunohistochemistry, despite very low concentrations of hydrocarbons remaining in water and tissues. Divergence in genome expression coincides with contaminating oil and is consistent with genome responses that are predictive of exposure to hydrocarbon-like chemicals and indicative of physiological and reproductive impairment. Oil-contaminated waters are also associated with aberrant protein expression in gill tissues of larval and adult fish. These data suggest that heavily weathered crude oil from the spill imparts significant biological impacts in sensitive Louisiana marshes, some of which remain for over 2 mo following initial exposures

Novel Method for Analysis of Allele Specific Expression in Triploid Oryzias latipes Reveals Consistent Pattern of Allele Exclusion

Assessing allele-specific gene expression (ASE) on a large scale continues to be a technically challenging problem. Certain biological phenomena, such as X chromosome inactivation and parental imprinting, affect ASE most drastically by completely shutting down the expression of a whole set of alleles. Other more subtle effects on ASE are likely to be much more complex and dependent on the genetic environment and are perhaps more important to understand since they may be responsible for a significant amount of biological diversity. Tools to assess ASE in a diploid biological system are becoming more reliable. Non-diploid systems are, however, not uncommon. In humans full or partial polyploid states are regularly found in both healthy (meiotic cells, polynucleated cell types) and diseased tissues (trisomies, non-disjunction events, cancerous tissues). In this work we have studied ASE in the medaka fish model system. We have developed a method for determining ASE in polyploid organisms from RNAseq data and we have implemented this method in a software tool set. As a biological model system we have used nuclear transplantation to experimentally produce artificial triploid medaka composed of three different haplomes. We measured ASE in RNA isolated from the livers of two adult, triploid medaka fish that showed a high degree of similarity. The majority of genes examined (82%) shared expression more or less evenly among the three alleles in both triploids. The rest of the genes (18%) displayed a wide range of ASE levels. Interestingly the majority of genes (78%) displayed generally consistent ASE levels in both triploid individuals. A large contingent of these genes had the same allele entirely suppressed in both triploids. When viewed in a chromosomal context, it is revealed that these genes are from large sections of 4 chromosomes and may be indicative of some broad scale suppression of gene expression

RNA-Seq reveals complex genetic response to deepwater horizon oil release in <it>Fundulus grandis</it>

<p>Abstract</p> <p>Background</p> <p>The release of oil resulting from the blowout of the Deepwater Horizon (DH) drilling platform was one of the largest in history discharging more than 189 million gallons of oil and subject to widespread application of oil dispersants. This event impacted a wide range of ecological habitats with a complex mix of pollutants whose biological impact is still not yet fully understood. To better understand the effects on a vertebrate genome, we studied gene expression in the salt marsh minnow <it>Fundulus grandis</it>, which is local to the northern coast of the Gulf of Mexico and is a sister species of the ecotoxicological model <it>Fundulus heteroclitus</it>. To assess genomic changes, we quantified mRNA expression using high throughput sequencing technologies (RNA-Seq) in <it>F. grandis</it> populations in the marshes and estuaries impacted by DH oil release. This application of RNA-Seq to a non-model, wild, and ecologically significant organism is an important evaluation of the technology to quickly assess similar events in the future.</p> <p>Results</p> <p>Our <it>de novo</it> assembly of RNA-Seq data produced a large set of sequences which included many duplicates and fragments. In many cases several of these could be associated with a common reference sequence using blast to query a reference database. This reduced the set of significant genes to 1,070 down-regulated and 1,251 up-regulated genes. These genes indicate a broad and complex genomic response to DH oil exposure including the expected <it>AHR</it>-mediated response and CYP genes. In addition a response to hypoxic conditions and an immune response are also indicated. Several genes in the choriogenin family were down-regulated in the exposed group; a response that is consistent with AH exposure. These analyses are in agreement with oligonucleotide-based microarray analyses, and describe only a subset of significant genes with aberrant regulation in the exposed set.</p> <p>Conclusion</p> <p>RNA-Seq may be successfully applied to feral and extremely polymorphic organisms that do not have an underlying genome sequence assembly to address timely environmental problems. Additionally, the observed changes in a large set of transcript expression levels are indicative of a complex response to the varied petroleum components to which the fish were exposed.</p

Comparison of calculated ASE values to whole gene expression.

<p>(A) Correlation of sum of ASE values per transcript (calculated only from dSNP sites) to geometric mean of each gene (calculated from coverage over entire transcript). (B) Plot of geometric mean of read coverage against raw fragment count showing a strong correlation between the two.</p

Parent strains and variant calling.

<p>(A) Parent strains and gender of donor genomes (images provided by MS). HNI-II males were mated with SOK females to produce F₁ embryos. At the blastula stage, cells were separated and diploid nuclei from them were injected into OR ova where they fused with the haploid nucleus of the oocyte. (B) Examples of variants called by VarScan. Only variant positions in which at least one strain was completely and unambiguously different from the others could be used (variant at position 343). Variant positions were unsuitable where one strain was only partially different (i.e. from heterozygosity) or where a strain had insufficient coverage to confidently call a set of observed nucleotides (variants at positions 47 and 998). The total number of dSNPs identified for each strain is indicated in the bottom line of red text.</p

dSNP-complete transcripts which can be placed in chromosomes.

<p>Each chromosome is represented by large vertical white bar outlined in gray. Transcripts are represented by horizontal bars of uniform size and are placed in the rank order in which they occur in each chromosome. The bars are colored to indicate the allelic imbalance category to which the transcript belongs based on exceptional high or low expressing alleles. Blank spaces represent transcripts that could not be said to deviate from equal allele expression. Each chromosome is divided into a left and right half by a black line. The left half of each chromosome gives the plot for trpA, while the right half gives the plot for trpB. The tallest bar (that for chromosome 8) is comprised of 329 transcripts in the order they occur on the chromosome with ties being assigned sequential ranks. The total numbers of transcripts that fall into the same category in both trpA and trpB are indicated in parenthesis after each category name in the legend.</p