The cytotoxicity and genotoxicity of hexavalent chromium in medaka (Oryzias latipes) cells.

Chromium is an increasing health concern for aquatic environments, however, the mechanism of chromium toxicity in aquatic species is yet unknown. We used a medaka (Oryzias latipes) fin cell line to investigate the cytotoxicity and genotoxicity of sodium chromate, a soluble form of hexavalent chromium. We used a clonogenic cytotoxicity assay to measure sodium chromate cytotoxicity, gamma-H2A.X immunofluoresence to measure DNA double-strand breaks, and chromosome damage to measure clastogenicity. We found that sodium chromate is cytotoxic to medaka fin cells, with toxicity increasing in a concentration-dependent manner. Treatments of 0.5, 1, 5, 10, 25, 50 and 100 microM sodium chromate caused 100, 103.5, 87.8, 77.5, 40.9, 15 and 2.7% survival, respectively, relative to the control. We visualized DNA double-strand breaks in medaka cells through the formation of gamma-H2A.X foci. Breaks could be detected at concentrations as low as 1 microM. We also found that sodium chromate induces chromosomal aberrations, causing chromatid lesions and exchanges that increase with concentration. Treatments of 0, 1, 5, 10 and 25 microM sodium chromate damaged 10.3, 17, 32.3, 43 and 51.6% of metaphases and induced 13, 23, 44, 69 and 118 total aberrations in 100 metaphases, respectively. These data show that hexavalent chromium is both cytotoxic and genotoxic to fish cells. Our results set the context for future work in the medaka cell culture model and provide important tools for investigating mechanisms of toxicity in aquatic organisms.

A three year study of metal levels in skin biopsies of whales in the Gulf of Mexico after the Deepwater Horizon oil crisis.

In response to the explosion of the Deepwater Horizon and the massive release of oil that followed, we conducted three annual research voyages to investigate how the oil spill would impact the marine offshore environment. Most investigations into the ecological and toxicological impacts of the Deepwater Horizon Oil crisis have mainly focused on the fate of the oil and dispersants, but few have considered the release of metals into the environment. From studies of previous oil spills, other marine oil industries, and analyses of oil compositions, it is evident that metals are frequently encountered. Several metals have been reported in the MC252 oil from the Deepwater Horizon oil spill, including the nonessential metals aluminum, arsenic, chromium, nickel, and lead; genotoxic metals, such as these are able to damage DNA and can bioaccumulate in organisms resulting in persistent exposure. In the Gulf of Mexico, whales are the apex species; hence we collected skin biopsies from sperm whales (Physeter macrocephalus), short-finned pilot whales (Globicephala macrorhynchus), and Bryde's whales (Balaenoptera edeni). The results from our three-year study of monitoring metal levels in whale skin show (1) genotoxic metals at concentrations higher than global averages previously reported and (2) patterns for MC252-relevant metal concentrations decreasing with time from the oil spill.

Whole Body Melanoma Transcriptome Response in Medaka.

The incidence of malignant melanoma continues to increase each year with poor prognosis for survival in many relapse cases. To reverse this trend, whole body response measures are needed to discover collaborative paths to primary and secondary malignancy. Several species of fish provide excellent melanoma models because fish and human melanocytes both appear in the epidermis, and fish and human pigment cell tumors share conserved gene expression signatures. For the first time, we have examined the whole body transcriptome response to invasive melanoma as a prelude to using transcriptome profiling to screen for drugs in a medaka (Oryzias latipes) model. We generated RNA-seq data from whole body RNA isolates for controls and melanoma fish. After testing for differential expression, 396 genes had significantly different expression (adjusted p-value <0.02) in the whole body transcriptome between melanoma and control fish; 379 of these genes were matched to human orthologs with 233 having annotated human gene symbols and 14 matched genes that contain putative deleterious variants in human melanoma at varying levels of recurrence. A detailed canonical pathway evaluation for significant enrichment showed the top scoring pathway to be antigen presentation but also included the expected melanocyte development and pigmentation signaling pathway. Results revealed a profound down-regulation of genes involved in the immune response, especially the innate immune system. We hypothesize that the developing melanoma actively suppresses the immune system responses of the body in reacting to the invasive malignancy, and that this mal-adaptive response contributes to disease progression, a result that suggests our whole-body transcriptomic approach merits further use. In these findings, we also observed novel genes not yet identified in human melanoma expression studies and uncovered known and new candidate drug targets for further testing in this malignant melanoma medaka model.

A RAD-tag genetic map for the platyfish (Xiphophorus maculatus) reveals mechanisms of karyotype evolution among teleost fish.

Mammalian genomes can vary substantially in haploid chromosome number even within a small taxon (e.g., 3-40 among deer alone); in contrast, teleost fish genomes are stable (24-25 in 58% of teleosts), but we do not yet understand the mechanisms that account for differences in karyotype stability. Among perciform teleosts, platyfish (Xiphophorus maculatus) and medaka (Oryzias latipes) both have 24 chromosome pairs, but threespine stickleback (Gasterosteus aculeatus) and green pufferfish (Tetraodon nigroviridis) have just 21 pairs. To understand the evolution of teleost genomes, we made a platyfish meiotic map containing 16,114 mapped markers scored on 267 backcross fish. We tiled genomic contigs along the map to create chromosome-length genome assemblies. Genome-wide comparisons of conserved synteny showed that platyfish and medaka karyotypes remained remarkably similar with few interchromosomal translocations but with numerous intrachromosomal rearrangements (transpositions and inversions) since their lineages diverged ∼120 million years ago. Comparative genomics with platyfish shows how reduced chromosome numbers in stickleback and green pufferfish arose by fusion of pairs of ancestral chromosomes after their lineages diverged from platyfish ∼195 million years ago. Zebrafish and human genomes provide outgroups to root observed changes. These studies identify likely genome assembly errors, characterize chromosome fusion events, distinguish lineage-independent chromosome fusions, show that the teleost genome duplication does not appear to have accelerated the rate of translocations, and reveal the stability of syntenies and gene orders in teleost chromosomes over hundreds of millions of years.