The planktonic stages of the salmon louse (Lepeophtheirus salmonis) are tolerant of end-of-century pCO2 concentrations.

The copepod Lepeophtheirus salmonis is an obligate ectoparasite of salmonids. Salmon lice are major pests in salmon aquaculture and due to its economic impact Lepeophtheirus salmonis is one of the most well studied species of marine parasite. However, there is limited understanding of how increased concentration of pCO2 associated with ocean acidification will impact host-parasite relationships. We investigated the effects of increased pCO2 on growth and metabolic rates in the planktonic stages, rearing L. salmonis from eggs to 12 days post hatch copepodids under three treatment levels: Control (416 µatm), Mid (747 µatm), and High (942 µatm). The pCO2 treatment had a significant effect on oxygen consumption rate with the High treatment animals exhibiting the greatest respiration. The treatments did not have a significant effect on the other biological endpoints measured (carbon, nitrogen, lipid volume, and fatty acid content). The results indicate that L. salmonis have mechanisms to compensate for increased concentration of pCO2and that populations will be tolerant of projected future ocean acidification scenarios. The work reported here also describes catabolism during the lecithotrophic development of L. salmonis, information that is not currently available to parameterize models of dispersal and viability of the planktonic free-living stages.

Cancer cell redirection biomarker discovery using a mutual information approach.

Introducing tumor-derived cells into normal mammary stem cell niches at a sufficiently high ratio of normal to tumorous cells causes those tumor cells to undergo a change to normal mammary phenotype and yield normal mammary progeny. This phenomenon has been termed cancer cell redirection. We have developed an in vitro model that mimics in vivo redirection of cancer cells by the normal mammary microenvironment. Using the RNA profiling data from this cellular model, we examined high-level characteristics of the normal, redirected, and tumor transcriptomes and found the global expression profiles clearly distinguish the three expression states. To identify potential redirection biomarkers that cause the redirected state to shift toward the normal expression pattern, we used mutual information relationships between normal, redirected, and tumor cell groups. Mutual information relationship analysis reduced a dataset of over 35,000 gene expression measurements spread over 13,000 curated gene sets to a set of 20 significant molecular signatures totaling 906 unique loci. Several of these molecular signatures are hallmark drivers of the tumor state. Using differential expression as a guide, we further refined the gene set to 120 core redirection biomarker genes. The expression levels of these core biomarkers are sufficient to make the normal and redirected gene expression states indistinguishable from each other but radically different from the tumor state.

Characterization of Calcification Events Using Live Optical and Electron Microscopy Techniques in a Marine Tubeworm.

Characterizing the first event of biological production of calcium carbonate requires a combination of microscopy approaches. First, intracellular pH distribution and calcium ions can be observed using live microscopy over time. This allows identification of the life stage and the tissue with the feature of interest for further electron microscopy studies. Life stage and tissues of interest are typically higher in pH and Ca signals. Here, using H. elegans, we present a protocol to characterize the presence of calcium carbonate structures in a biological specimen on the scanning electron microscope (SEM), using energy-dispersive X-ray spectroscopy (EDS) to visualize elemental composition, using electron backscatter diffraction (EBSD) to determine the presence of crystalline structures, and using transmission electron microscopy (TEM) to analyze the composition and structure of the material. In this protocol, a focused ion beam (FIB) is used to isolate samples with dimension suitable for TEM analysis. As FIB is a site specific technique, we demonstrate how information from the previous techniques can be used to identify the region of interest, where Ca signals are highest.

Mechanical robustness of the calcareous tubeworm Hydroides elegans: warming mitigates the adverse effects of ocean acidification.

Development of antifouling strategies requires knowledge of how fouling organisms would respond to climate change associated environmental stressors. Here, a calcareous tube built by the tubeworm, Hydroides elegans, was used as an example to evaluate the individual and interactive effects of ocean acidification (OA), warming and reduced salinity on the mechanical properties of a tube. Tubeworms produce a mechanically weaker tube with less resistance to simulated predator attack under OA (pH 7.8). Warming (29°C) increased tube volume, tube mineral density and the tube's resistance to a simulated predatory attack. A weakening effect by OA did not make the removal of tubeworms easier except for the earliest stage, in which warming had the least effect. Reduced salinity (27 psu) did not affect tubes. This study showed that both mechanical analysis and computational modeling can be integrated with biofouling research to provide insights into how fouling communities might develop in future ocean conditions.

Weakening mechanisms of the serpulid tube in a high-CO2 world.

Many benthic marine organisms produce calcium carbonate (CaCO3) structures for mechanical protection through a biologically controlled calcification process. However, the oceans are becoming unfavorable for calcification because of the stress associated with ocean acidification (OA) and associated chemical changes such as declining saturation state of CaCO3 and decreasing seawater pH. This work studies the impacts of OA-driven decreased pH on the calcareous tubes produced by the serpulid tubeworm Hydroides elegans. Tubes grown under control and OA experimental conditions were measured for structural and mechanical properties, and their mechanical properties were further interpreted using finite element analysis (FEA). The near-future predicted pH value of 7.8 altered tube ultrastructure, volume, and density and decreased the mean tube hardness and elasticity by ∼ 80 and ∼ 70%, respectively. The crushing force required for breaking the tube was reduced by 64%. The FEA results demonstrated how a simulated predator attack may affect the structure with different structural and mechanical properties and consequently shift the stress development and distribution in the tubes, causing a more concentrated stress distribution and therefore leading to a lower ability to withstand attacks.

Interactive effects of ocean acidification, elevated temperature, and reduced salinity on early-life stages of the pacific oyster.

Ocean acidification (OA) effects on larvae are partially attributed for the rapidly declining oyster production in the Pacific Northwest region of the United States. This OA effect is a serious concern in SE Asia, which produces >80% of the world's oysters. Because climate-related stressors rarely act alone, we need to consider OA effects on oysters in combination with warming and reduced salinity. Here, the interactive effects of these three climate-related stressors on the larval growth of the Pacific oyster, Crassostrea gigas, were examined. Larvae were cultured in combinations of temperature (24 and 30 °C), pH (8.1 and 7.4), and salinity (15 psu and 25 psu) for 58 days to the early juvenile stage. Decreased pH (pH 7.4), elevated temperature (30 °C), and reduced salinity (15 psu) significantly delayed pre- and post-settlement growth. Elevated temperature lowered the larval lipid index, a proxy for physiological quality, and negated the negative effects of decreased pH on attachment and metamorphosis only in a salinity of 25 psu. The negative effects of multiple stressors on larval metamorphosis were not due to reduced size or depleted lipid reserves at the time of metamorphosis. Our results supported the hypothesis that the C. gigas larvae are vulnerable to the interactions of OA with reduced salinity and warming in Yellow Sea coastal waters now and in the future.

Temperature dependent effects of elevated CO2 on shell composition and mechanical properties of Hydroides elegans: insights from a multiple stressor experiment.

The majority of marine benthic invertebrates protect themselves from predators by producing calcareous tubes or shells that have remarkable mechanical strength. An elevation of CO2 or a decrease in pH in the environment can reduce intracellular pH at the site of calcification and thus interfere with animal's ability to accrete CaCO3. In nature, decreased pH in combination with stressors associated with climate change may result in the animal producing severely damaged and mechanically weak tubes. This study investigated how the interaction of environmental drivers affects production of calcareous tubes by the serpulid tubeworm, Hydroides elegans. In a factorial manipulative experiment, we analyzed the effects of pH (8.1 and 7.8), salinity (34 and 27‰), and temperature (23°C and 29°C) on the biomineral composition, ultrastructure and mechanical properties of the tubes. At an elevated temperature of 29°C, the tube calcite/aragonite ratio and Mg/Ca ratio were both increased, the Sr/Ca ratio was decreased, and the amorphous CaCO3 content was reduced. Notably, at elevated temperature with decreased pH and reduced salinity, the constructed tubes had a more compact ultrastructure with enhanced hardness and elasticity compared to decreased pH at ambient temperature. Thus, elevated temperature rescued the decreased pH-induced tube impairments. This indicates that tubeworms are likely to thrive in early subtropical summer climate. In the context of climate change, tubeworms could be resilient to the projected near-future decreased pH or salinity as long as surface seawater temperature rise at least by 4°C.

Decreased pH does not alter metamorphosis but compromises juvenile calcification of the tube worm Hydroides elegans.

Using CO2 perturbation experiments, we examined the pre- and post-settlement growth responses of a dominant biofouling tubeworm (Hydroides elegans) to a range of pH. In three different experiments, embryos were reared to, or past, metamorphosis in seawater equilibrated to CO2 values of about 480 (control), 980, 1,480, and 2,300 µatm resulting in pH values of around 8.1 (control), 7.9, 7.7, and 7.5, respectively. These three decreased pH conditions did not affect either embryo or larval development, but both larval calcification at the time of metamorphosis and early juvenile growth were adversely affected. During the 24-h settlement assay experiment, half of the metamorphosed larvae were unable to calcify tubes at pH 7.9 while almost no tubes were calcified at pH 7.7. Decreased ability to calcify at decreased pH may indicate that these calcifying tubeworms may be one of the highly threatened species in the future ocean.