Population-level transcriptome sequencing of nonmodel organisms Erynnis propertius and Papilio zelicaon

<p>Abstract</p> <p>Background</p> <p>Several recent studies have demonstrated the use of Roche 454 sequencing technology for <it>de novo </it>transcriptome analysis. Low error rates and high coverage also allow for effective SNP discovery and genetic diversity estimates. However, genetically diverse datasets, such as those sourced from natural populations, pose challenges for assembly programs and subsequent analysis. Further, estimating the effectiveness of transcript discovery using Roche 454 transcriptome data is still a difficult task.</p> <p>Results</p> <p>Using the Roche 454 FLX Titanium platform, we sequenced and assembled larval transcriptomes for two butterfly species: the Propertius duskywing, <it>Erynnis propertius </it>(Lepidoptera: Hesperiidae) and the Anise swallowtail, <it>Papilio zelicaon </it>(Lepidoptera: Papilionidae). The Expressed Sequence Tags (ESTs) generated represent a diverse sample drawn from multiple populations, developmental stages, and stress treatments.</p> <p>Despite this diversity, > 95% of the ESTs assembled into long (> 714 bp on average) and highly covered (> 9.6× on average) contigs. To estimate the effectiveness of transcript discovery, we compared the number of bases in the hit region of unigenes (contigs and singletons) to the length of the best match silkworm (<it>Bombyx mori</it>) protein--this "ortholog hit ratio" gives a close estimate on the amount of the transcript discovered relative to a model lepidopteran genome. For each species, we tested two assembly programs and two parameter sets; although CAP3 is commonly used for such data, the assemblies produced by Celera Assembler with modified parameters were chosen over those produced by CAP3 based on contig and singleton counts as well as ortholog hit ratio analysis. In the final assemblies, 1,413 <it>E. propertius </it>and 1,940 <it>P. zelicaon </it>unigenes had a ratio > 0.8; 2,866 <it>E. propertius </it>and 4,015 <it>P. zelicaon </it>unigenes had a ratio > 0.5.</p> <p>Conclusions</p> <p>Ultimately, these assemblies and SNP data will be used to generate microarrays for ecoinformatics examining climate change tolerance of different natural populations. These studies will benefit from high quality assemblies with few singletons (less than 26% of bases for each assembled transcriptome are present in unassembled singleton ESTs) and effective transcript discovery (over 6,500 of our putative orthologs cover at least 50% of the corresponding model silkworm gene).</p

Thermal Variability Increases the Impact of Autumnal Warming and Drives Metabolic Depression in an Overwintering Butterfly

Increases in thermal variability elevate metabolic rate due to Jensen's inequality, and increased metabolic rate decreases the fitness of dormant ectotherms by increasing consumption of stored energy reserves. Theory predicts that ectotherms should respond to increased thermal variability by lowering the thermal sensitivity of metabolism, which will reduce the impact of the warm portion of thermal variability. We examined the thermal sensitivity of metabolic rate of overwintering Erynnis propertius (Lepidoptera: Hesperiidae) larvae from a stable or variable environment reared in the laboratory in a reciprocal common garden design, and used these data to model energy use during the winters of 1973–2010 using meteorological data to predict the energetic outcomes of metabolic compensation and phenological shifts. Larvae that experienced variable temperatures had decreased thermal sensitivity of metabolic rate, and were larger than those reared at stable temperatures, which could partially compensate for the increased energetic demands. Even with depressed thermal sensitivity, the variable environment was more energy-demanding than the stable, with the majority of this demand occurring in autumn. Autumn phenology changes thus had disproportionate influence on energy consumption in variable environments, and variable-reared larvae were most susceptible to overwinter energy drain. Therefore the energetic impacts of the timing of entry into winter dormancy will strongly influence ectotherm fitness in northern temperate environments. We conclude that thermal variability drives the expression of metabolic suppression in this species; that phenological shifts will have a greater impact on ectotherms in variable thermal environments; and that E. propertius will be more sensitive to shifts in phenology in autumn than in spring. This suggests that increases in overwinter thermal variability and/or extended, warm autumns, will negatively impact all non-feeding dormant ectotherms which lack the ability to suppress their overwinter metabolic thermal sensitivity

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

Castor canadensis: an analysis of foraging patterns in two different habitats of northern Michigan.

This study compares two different foraging habitats for the American beaver, Castor canadensis, in Michigan. This study was designed to test optimal foraging theory's application to a field population. This study analyzes evidence of beaver activity in order to investigate efficiency in foraging. We expected to observe beaver preference for species and stem size, as well as decreased removal rates with respect to increased distance from the water's edge. Changing stem size selection as a function of increased distance from the water was also expected. All such observations would illustrate that beaver make economic decisions about maximizing energetic gains. Some debate has centered on foraging in beaver, and in review of the literature, one finds several conflicting reports. This research was modeled after past studies on beaver foraging in order to contribute commentary to the on-going debate in optimal foraging theory's application to central foragers, such as the beaver. In our study, beaver were found to show a species preference and a preference for stems located close to the water's edge. However, no significant relationship was found for stem size choice and distance. No overall stem preference was found either. Due to within and between site variation in species composition, these results show how foraging behavior can differ under different environmental circumstances. It is also discussed that beaver may not be optimal foragers, but rather are constrained by factors other than needs for energy maximization, such as predator avoidance. This study is unable to make a decisive conclusion that beavers forage optimally.http://deepblue.lib.umich.edu/bitstream/2027.42/54530/1/2969.pdfDescription of 2969.pdf : Access restricted to on-site users at the U-M Biological Station

Considering Local Adaptation in Issues of Lepidopteran Conservation a Review and Recommendations

Butterflies are among the most charismatic and alluring of invertebrate organisms. Their complex life histories and diverse habitat affiliations often produce regional variation and local adaptation among different populations. Unfortunately, numerous lepidopteran species are facing extinction. In this paper we address the importance of considering local adaptation in butterfly conservation and encourage investigations of regional specialization that may enhance the effectiveness of conservation strategies such as captive rearing, habitat restoration and the introduction of populations into new localities. We also give suggestions for dealing with problems arising from local adaptation in particular instances

Select but diverse countries are reducing both climate vulnerability and CO2 emissions

Mitigation of greenhouse gas (GHG) emissions and adaptation to climate risk are two essential ingredients of climate change policy. Both are needed and co-benefits may exist. Yet, mitigation and adaptation are not usually pursued together. Part of remedying this shortcoming is understanding the relationship between GHG emissions and climate vulnerability reduction and recognizing when and where they trend together. Here, we compare changes in fossil fuel CO2 emissions per capita and in climate vulnerability scores over the past two decades in 179 countries. We use climate vulnerability scores from the well-established ND-GAIN Country Index, a composite metric constructed from thirty-six indicators covering three components of vulnerability (exposure, sensitivity and adaptive capacity). We find that 69% of the countries decreased climate vulnerability, while increasing their per capita fossil fuel CO2 emissions. These countries are successfully reducing climate vulnerability but are increasing their GHG emissions and thus failing in mitigation efforts. In contrast, 23% of the countries have been successful in simultaneously reducing per capita CO2 emissions and climate vulnerability. Furthermore, in highly vulnerable countries, increasing CO2 emissions are not correlated with decreasing climate vulnerability. These findings underscore that climate vulnerability reduction may be due only partly to economic development. This finding also changes our prevailing view that increases in CO2 emissions are associated with vulnerability reduction. Finally, examining mitigation and climate-vulnerability reduction by sector, we show that a majority of countries are able to reduce vulnerability in ecosystem services. Those countries and sectors with positive trends provide examples for others to follow, as solutions at the mitigation-climate vulnerability reduction interface are essential for sustainable economic development

Appendix A. Range map of Quercus garryana and map of study sites on Vancouver Island, British Columbia.

Range map of Quercus garryana and map of study sites on Vancouver Island, British Columbia

Appendix B. Detailed methods of observational surveys, exclusion experiment, and statistical analyses.

Detailed methods of observational surveys, exclusion experiment, and statistical analyses

Appendix D. Measuring the performance of Erynnis propertius: size, survivorship, and butterfly production.

Measuring the performance of Erynnis propertius: size, survivorship, and butterfly production

Appendix B. Phenology of Neuroterus saltatorius and Erynnis propertius on Vancouver Island, British Columbia.

Phenology of Neuroterus saltatorius and Erynnis propertius on Vancouver Island, British Columbia