Biology
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Item Astyanax surface and cave fish morphs(Springer Nature, 2020-07-11) Jeffery, William R.The small teleost fish Astyanax mexicanus has emerged as an outstanding model for studying many biological topics in the context of evolution. A major attribute is conspecific surface dwelling (surface fish) and blind cave dwelling (cavefish) morphs that can be raised in the laboratory and spawn large numbers of transparent and synchronously developing embryos. More than 30 cavefish populations have been discovered, mostly in northeastern Mexico, and some are thought to have evolved independently from surface fish ancestors, providing excellent models of parallel and convergent evolution. Cavefish have evolved eye and pigmentation regression, as well as modifications in brain morphology, behaviors, heart regenerative capacity, metabolic processes, and craniofacial organization. Thus, the Astyanax model provides researchers with natural “mutants” to study life in the challenging cave environment. The application of powerful genetic approaches based on hybridization between the two morphs and between the different cavefish populations are key advantages for deciphering the developmental and genetic mechanisms regulating trait evolution. QTL analysis has revealed the genetic architectures of gained and lost traits. In addition, some cavefish traits resemble human diseases, offering novel models for biomedical research. Astyanax research is supported by genome assemblies, transcriptomes, tissue and organ transplantation, gene manipulation and editing, and stable transgenesis, and benefits from a welcoming and interactive research community that conducts integrated community projects and sponsors the International Astyanax Meeting (AIM).Item Long-term time-lapse live imaging reveals extensive cell migration during annelid regeneration(BioMed Central, 2016) Zattara, Eduardo E.; Turlington, Kate W.; Bely, Alexandra E.Background: Time-lapse imaging has proven highly valuable for studying development, yielding data of much finer resolution than traditional “still-shot” studies and allowing direct examination of tissue and cell dynamics. A major challenge for time-lapse imaging of animals is keeping specimens immobile yet healthy for extended periods of time. Although this is often feasible for embryos, the difficulty of immobilizing typically motile juvenile and adult stages remains a persistent obstacle to time-lapse imaging of post-embryonic development. Results: Here we describe a new method for long-duration time-lapse imaging of adults of the small freshwater annelid Pristina leidyi and use this method to investigate its regenerative processes. Specimens are immobilized with tetrodotoxin, resulting in irreversible paralysis yet apparently normal regeneration, and mounted in agarose surrounded by culture water or halocarbon oil, to prevent dehydration but allowing gas exchange. Using this method, worms can be imaged continuously and at high spatial-temporal resolution for up to 5 days, spanning the entire regeneration process. We performed a fine-scale analysis of regeneration growth rate and characterized cell migration dynamics during early regeneration. Our studies reveal the migration of several putative cell types, including one strongly resembling published descriptions of annelid neoblasts, a cell type suggested to be migratory based on “still-shot” studies and long hypothesized to be linked to regenerative success in annelids. Conclusions: Combining neurotoxin-based paralysis, live mounting techniques and a starvation-tolerant study system has allowed us to obtain the most extensive high-resolution longitudinal recordings of full anterior and posterior regeneration in an invertebrate, and to detect and characterize several cell types undergoing extensive migration during this process. We expect the tetrodotoxin paralysis and time-lapse imaging methods presented here to be broadly useful in studying other animals and of particular value for studying post-embryonic development.Item Transcriptome characterization via 454 pyrosequencing of the annelid Pristina leidyi, an emerging model for studying the evolution of regeneration(2012-06-29) Nyberg, Kevin G.; Conte, Matthew A.; Kostyun, Jamie L.; Forde, Alison; Bely, Alexandra E.Background: The naid annelids contain a number of species that vary in their ability to regenerate lost body parts, making them excellent candidates for evolution of regeneration studies. However, scant sequence data exists to facilitate such studies. We constructed a cDNA library from the naid Pristina leidyi, a species that is highly regenerative and also reproduces asexually by fission, using material from a range of regeneration and fission stages for our library. We then sequenced the transcriptome of P. leidyi using 454 technology. Results: 454 sequencing produced 1,550,174 reads with an average read length of 376 nucleotides. Assembly of 454 sequence reads resulted in 64,522 isogroups and 46,679 singletons for a total of 111,201 unigenes in this transcriptome. We estimate that over 95% of the transcripts in our library are present in our transcriptome. 17.7% of isogroups had significant BLAST hits to the UniProt database and these include putative homologs of a number of genes relevant to regeneration research. Although many sequences are incomplete, the mean sequence length of transcripts (isotigs) is 707 nucleotides. Thus, many sequences are large enough to be immediately useful for downstream applications such as gene expression analyses. Using in situ hybridization, we show that two Wnt/β-catenin pathway genes (homologs of frizzled and β-catenin) present in our transcriptome are expressed in the regeneration blastema of P. leidyi, demonstrating the usefulness of this resource for regeneration research. Conclusions: 454 sequencing is a rapid and efficient approach for identifying large numbers of genes in an organism that lacks a sequenced genome. This transcriptome dataset will be a valuable resource for molecular analyses of regeneration in P. leidyi and will serve as a starting point for comparisons to non-regenerating naids. It also contributes significantly to the still limited genomic resources available for annelids and lophotrochozoans more generally.Item Breaking the A-P axis: Evolution of diverse asexual reproduction strategies in Convolutriloba acoels(2009) Sikes, James M.; Bely, Alexandra E.; Behavior, Ecology, Evolution and Systematics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The defining characteristic of the Bilateria is the presence of a distinct head end and tail end, which defines the anterior-posterior (A-P) axis, a feature that is established during embryogenesis and generally remains unaltered during the lifetime of an organism. While a few bilaterians have evolved asexual reproduction strategies that allow them to subdivide the A-P axis, acoels in the genus Convolutriloba have an unparalleled ability to alter the A-P axis during modes of transverse fission, longitudinal fission, and reversed polarity budding. Convolutriloba acoels thus offer an exceptional opportunity to investigate the mechanisms that allow for the radical modification of an already established A-P body axis and to explore the evolution and development of diverse asexual reproduction strategies among related species. In this study, I reconstruct the evolutionary history of asexual reproduction in the Convolutriloba and compare the diverse modes of asexual reproduction at the level of body-wall musculature, nervous system development, and cell proliferation while also exploring the regenerative potentials of tissues across species with different modes of asexual reproduction. In addition, I further explore the unusual process of A-P axis reversal that occurs during reversed polarity budding in C. retrogemma through studies of body patterning and regeneration. The results of these analyses suggest that a rich developmental toolkit of regenerative abilities, including the ability to utilize both epimorphosis and morphallaxis, to regenerate all parts of its body even from a small fragment, and to produce bifurcated A-P axes were present in the ancestor of the Convolutriloba allowing for the evolution of A-P axis modifications unlike any other bilaterian group. This toolkit along with the evolution of a seemingly unpatterned zone of tissue within the body of C. retrogemma capable of generating new anterior axes appear to have allowed this species to evolve the ability to form reversed A-P axes during budding.