Production of a reference transcriptome and transcriptomic database (PocilloporaBase) for the cauliflower coral, Pocillopora damicornis

<p>Abstract</p> <p>Background</p> <p>Motivated by the precarious state of the world's coral reefs, there is currently a keen interest in coral transcriptomics. By identifying changes in coral gene expression that are triggered by particular environmental stressors, we can begin to characterize coral stress responses at the molecular level, which should lead to the development of more powerful diagnostic tools for evaluating the health of corals in the field. Furthermore, the identification of genetic variants that are more or less resilient in the face of particular stressors will help us to develop more reliable prognoses for particular coral populations. Toward this end, we performed deep mRNA sequencing of the cauliflower coral, <it>Pocillopora damicornis</it>, a geographically widespread Indo-Pacific species that exhibits a great diversity of colony forms and is able to thrive in habitats subject to a wide range of human impacts. Importantly, <it>P. damicornis </it>is particularly amenable to laboratory culture. We collected specimens from three geographically isolated Hawaiian populations subjected to qualitatively different levels of human impact. We isolated RNA from colony fragments ("nubbins") exposed to four environmental stressors (heat, desiccation, peroxide, and hypo-saline conditions) or control conditions. The RNA was pooled and sequenced using the 454 platform.</p> <p>Description</p> <p>Both the raw reads (n = 1, 116, 551) and the assembled contigs (n = 70, 786; mean length = 836 nucleotides) were deposited in a new publicly available relational database called PocilloporaBase <url>http://www.PocilloporaBase.org</url>. Using BLASTX, 47.2% of the contigs were found to match a sequence in the NCBI database at an E-value threshold of ≤.001; 93.6% of those contigs with matches in the NCBI database appear to be of metazoan origin and 2.3% bacterial origin, while most of the remaining 4.1% match to other eukaryotes, including algae and amoebae.</p> <p>Conclusions</p> <p><it>P. damicornis </it>now joins the handful of coral species for which extensive transcriptomic data are publicly available. Through PocilloporaBase <url>http://www.PocilloporaBase.org</url>, one can obtain assembled contigs and raw reads and query the data according to a wide assortment of attributes including taxonomic origin, PFAM motif, KEGG pathway, and GO annotation.</p

Rapid Evolution of Coral Proteins Responsible for Interaction with the Environment

Christian R. Voolstra is with King Abdullah University of Science and Technology, Shinichi Sunagawa is with the European Molecular Biology Laboratory, Mikhail V. Matz is with UT Austin, Till Bayer is with King Abdullah University of Science and Technology, Manuel Aranda is with King Abdullah University of Science and Technology, Emmanuel Buschiazzo is with University of California Merced, Michael K. DeSalvo is with University of California San Francisco, Erika Lindquist is with the Department of Energy Joint Genome Institute, Alina M. Szmant is with University of North Carolina Wilmington, Mary Alice Coffroth is with State University of New York at Buffalo, Mónica Medina is with University of California Merced.Background -- Corals worldwide are in decline due to climate change effects (e.g., rising seawater temperatures), pollution, and exploitation. The ability of corals to cope with these stressors in the long run depends on the evolvability of the underlying genetic networks and proteins, which remain largely unknown. A genome-wide scan for positively selected genes between related coral species can help to narrow down the search space considerably. Methodology/Principal Findings -- We screened a set of 2,604 putative orthologs from EST-based sequence datasets of the coral species Acropora millepora and Acropora palmata to determine the fraction and identity of proteins that may experience adaptive evolution. 7% of the orthologs show elevated rates of evolution. Taxonomically-restricted (i.e. lineage-specific) genes show a positive selection signature more frequently than genes that are found across many animal phyla. The class of proteins that displayed elevated evolutionary rates was significantly enriched for proteins involved in immunity and defense, reproduction, and sensory perception. We also found elevated rates of evolution in several other functional groups such as management of membrane vesicles, transmembrane transport of ions and organic molecules, cell adhesion, and oxidative stress response. Proteins in these processes might be related to the endosymbiotic relationship corals maintain with dinoflagellates in the genus Symbiodinium. Conclusion/Relevance -- This study provides a birds-eye view of the processes potentially underlying coral adaptation, which will serve as a foundation for future work to elucidate the rates, patterns, and mechanisms of corals' evolutionary response to global climate change.This work was supported by DEB-1054766 to M.V.M. and National Science Foundation grants IOS-0644438 and OCE-0313708 to M.M., and by a Collaborative Travel Fund to C.R.V. made by King Abdullah University of Science and Technology (KAUST). The work conducted by the U.S. Department of Energy Joint Genome Institute is supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC02-05CH11231. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Biological Sciences, School o

Symbiodinium Transcriptomes: Genome Insights into the Dinoflagellate Symbionts of Reef-Building Corals

Dinoflagellates are unicellular algae that are ubiquitously abundant in aquatic environments. Species of the genus Symbiodinium form symbiotic relationships with reef-building corals and other marine invertebrates. Despite their ecologic importance, little is known about the genetics of dinoflagellates in general and Symbiodinium in particular. Here, we used 454 sequencing to generate transcriptome data from two Symbiodinium species from different clades (clade A and clade B). With more than 56,000 assembled sequences per species, these data represent the largest transcriptomic resource for dinoflagellates to date. Our results corroborate previous observations that dinoflagellates possess the complete nucleosome machinery. We found a complete set of core histones as well as several H3 variants and H2A.Z in one species. Furthermore, transcriptome analysis points toward a low number of transcription factors in Symbiodinium spp. that also differ in the distribution of DNA-binding domains relative to other eukaryotes. In particular the cold shock domain was predominant among transcription factors. Additionally, we found a high number of antioxidative genes in comparison to non-symbiotic but evolutionary related organisms. These findings might be of relevance in the context of the role that Symbiodinium spp. play as coral symbionts

Evaluating vertical migration behavior of harmful raphidophytes in the Delaware Inland Bays utilizing quantitative real-time PCR

Mixed blooms of 4 species of harmful raphidophytes (Chattonella cf. verruculosa, Chattonella subsalsa, Heterosigma akashiwo, and Fibrocapsa japonica) occur in the shallow (1 to 2 m) Delaware Inland Bays (DIB), USA. Raphidophytes vertically migrate in other deeper water ecosystems to utilize deep nutrient stocks at night, and thus obtain an advantage over non-migrating algae. Anoxic DIB sediments release high levels of bioavailable phosphate, which could potentially be used by vertically migrating flagellates. This study aimed to characterize and understand the migration patterns of DIB raphidophytes, and determine whether benthic phosphate fluxes could provide the cells with P. We demonstrated vertical migration of isolated DIB raphidophyte cultures in the laboratory, where differences in the response of C. subsalsa and H. akashiwo to light:dark period manipulations suggested possible differences in external versus endogenous regulation of migration behavior in the 2 species. Natural blooms in the field (enclosed in a mesocosm system) also exhibited patterns of diel vertical migration, as determined by quantitative real-time PCR (QPCR) used to enumerate the diel vertical distributions of each species. Our data suggested that these 2 photoautotrophic species spend daylight hours near the surface and are found directly on the sediment surface at night. However, diel changes in particulate C:P ratios did not support the hypothesis that there is preferential uptake of sedimentary phosphate at night. Our results also suggested that the migration behavior may have important implications for designing sampling strategies for monitoring programs. QPCR has a number of decisive advantages over traditional microscopic counting methods, making this a powerful tool for fine spatial and temporal scale detection and enumeration of vertically migrating harmful algal species. © Inter-Research 2005

Improved quantitative real-time PCR assays for enumeration of harmful algal species in field samples using an exogenous DNA reference standard

© 2005, by the American Society of Limnology and Oceanography, Inc. Quantitative real-time PCR (QPCR) is a powerful and sensitive method for quantitative detection of microorganisms. Application of this methodology for enumeration of harmful algal bloom (HAB) species has the potential to revolutionize our approach to HAB research, making it possible to identify correlations between cell abundances and factors that regulate bloom dynamics. Its application to ecological studies, however, has produced mixed results. QPCR assays typically rely on the generation of standard curves from plasmids or laboratory cultures that may be unrealistic when compared to amplification of DNA extracted from field samples. In addition, existing methods often fail to incorporate controls to assess variability in extraction and amplification efficiencies, or include controls that are sequence-specific and preclude the investigation of multiple species. Here, we describe the development and rigorous analysis of QPCR assays for two HAB species, Chattonella subsalsa and Heterosigma akashiwo, in which we introduce a known concentration of exogenous DNA plasmid into the extraction buffer as a reference standard. Since the target DNA is extracted in the presence of the reference standard, inherent variability in extraction and amplification efficiencies affect both target and standard equally. Furthermore, the reference standard is applicable to QPCR analysis of any microbial species. Using environmental bloom samples as calibrators, we evaluated the accuracy of the comparative Ct method for enumeration of target species in several field samples. Our investigation demonstrates that the comparative Ct method with an exogenous DNA reference standard provides both accurate and reproducible quantification of HAB species in environmental samples