Proteomic analysis of oyster larvae reveals molecular mechanism of ocean acidification and multiple stressor effects

The increase in carbon dioxide emissions due to human activities has led to drastic variations in global climate. In addition to global warming and extreme weather patterns, the high CO2 levels have been leading to progressive ocean acidification. Compounded with other climate change related stressors, ocean acidification will hinder the ability of marine organisms to adapt to the ensuing changes and might affect human dependence on oceans as a source of food. Most marine organisms have complex life cycles, involving metamorphosis from larval to adult forms. In the early stages of life, oysters have calcium carbonate shells that are particularly sensitive to low pH, and the rapid climatic changes can compromise their metamorphosis. High temperature, low salinity and low pH resulting from ocean acidification are detrimental to both native and cultivated oyster populations. Although mechanistic studies to understand the tolerance responses of closely related species would be significant in this context, none have been reported to date. Therefore, this thesis aims to reveal the mechanisms that distinguish the “winners” from the “losers” among the selected aquatic species of commercial importance, in withstanding the stress induced by climate change. The present study employed molecular approaches to evaluate the interactive and cumulative effects of multiple stressors on large-scale cultures of pediveliger larvae from two oyster populations, Crassostrea hongkongensis and Crassostrea gigas. The study undertook transcriptomic and proteomic profiling of changes induced by ocean acidification in the larvae. The results revealed that oyster larvae could adopt an energy ‘trade-off’ strategy through metabolic suppression and adjust cell signalling pathways to overcome the stress induced by ocean acidification. Information from the oyster genome database facilitated the shotgun proteomics investigations on oyster larvae remarkably revealed over 1350 proteins in both the species. The study identified species- and stressor-specific tolerance responses, and survival mechanisms that preserved calcification, in oyster larvae. The larvae showed depletion of energy reserves due to enhanced metabolism, oxidative damage-induced immune response and metabolic suppression. The study reveals the existence of tolerance mechanisms in oysters that help them adapt to stresses resulting from climate change. It pioneered the use of a proteomics approach to understand the impact of multiple stressors on oyster larvae and the molecular mechanisms underlying their successful adaptation to them. Highlighted several potential possible biomarkers in this study will likely to play an important role in identifying oyster species showing heritable tolerance for future aquaculture.published_or_final_versionBiological SciencesDoctoralDoctor of Philosoph

Proteomic profiles of Platynereis spp. collected from inside and outside the CO2 vent (Ischia, Italy) and used in a reciprocal transplant experiment in September 2013

Platynereis spp. were collected via snorkelling or scuba from either inside (40°43′53″N, 13°57′47″E) (stations A1-A3) or outside (40°43'33.33"N, 13°57'36.38"E and 40°44′48″N, 13°56′39″E) (stations C1-C3) the carbon dioxide (CO2) vent on the island of Ischia (Italy) and used in a reciprocal transplant experiment. The effect of exposure to high or low partial pressures of CO2 (pCO2) conditions on the proteome of worms from different pCO2 regimes was investigated to understand the effect of exposure to different pCO2 conditions on the cellular physiological response. This experiment was conducted between 04/09/2013 and 16/09/2013. The experiment was staggered during this time so all worms could be processed. After five days exposure to either low or high CO2 conditions worms were snap frozen in liquid nitrogen and shipped to the University of Hong Kong for proteomic analysis which was finalised on 02/08/2015. Proteomic profiles of worms were characterised using a liquid chromatography-mass spectrometry/mass spectrometry triple time of flight (LC-MS/ MS Triple TOF) approach

Synthesis of Bimetallic BiPO₄/ZnO Nanocomposite: Enhanced Photocatalytic Dye Degradation and Antibacterial Applications

Multidrug-resistant strains (MDRs) are becoming a major concern in a variety of settings, including water treatment and the medical industry. Well-dispersed catalysts such as BiPO4, ZnO nanoparticles (NPs), and different ratios of BiPO4/ZnO nanocomposites (NCs) were synthesized through hydrothermal treatments. The morphological behavior of the prepared catalysts was characterized using XRD, Raman spectra, PL, UV–Vis diffuse reflectance spectroscopy (UV-DRS), SEM, EDX, and Fe-SEM. MDRs were isolated and identified by the 16s rDNA technique as belonging to B. flexus, B. filamentosus, P. stutzeri, and A. baumannii. The antibacterial activity against MDRs and the photocatalytic methylene blue (MB) dye degradation activity of the synthesized NPs and NCs were studied. The results demonstrate that the prepared BiPO4/ZnO-NCs (B1Z4-75:300; NCs-4) caused a maximum growth inhibition of 20 mm against A. baumannii and a minimum growth inhibition of 12 mm against B. filamentosus at 80 μg mL−1 concentrations of the NPs and NCs. Thus, NCs-4 might be a suitable alternative to further explore and develop as an antibacterial agent. The obtained results statistically justified the data (p ≤ 0.05) via one-way analysis of variance (ANOVA). According to the results of the antibacterial and photocatalytic study, we selected the best bimetallic NCs-4 for the photoexcited antibacterial effect of MDRs, including Gram ve+ and Gram ve− strains, via UV light irradiation. The flower-like NCs-4 composites showed more effectiveness than those of BiPO4, ZnO, and other ratios of NCs. The results encourage the development of flower-like NCs-4 to enhance the photocatalytic antibacterial technique for water purification

Synthesis of Bimetallic BiPO4/ZnO Nanocomposite: Enhanced Photocatalytic Dye Degradation and Antibacterial Applications

Multidrug-resistant strains (MDRs) are becoming a major concern in a variety of settings, including water treatment and the medical industry. Well-dispersed catalysts such as BiPO4, ZnO nanoparticles (NPs), and different ratios of BiPO4/ZnO nanocomposites (NCs) were synthesized through hydrothermal treatments. The morphological behavior of the prepared catalysts was characterized using XRD, Raman spectra, PL, UV–Vis diffuse reflectance spectroscopy (UV-DRS), SEM, EDX, and Fe-SEM. MDRs were isolated and identified by the 16s rDNA technique as belonging to B. flexus, B. filamentosus, P. stutzeri, and A. baumannii. The antibacterial activity against MDRs and the photocatalytic methylene blue (MB) dye degradation activity of the synthesized NPs and NCs were studied. The results demonstrate that the prepared BiPO4/ZnO-NCs (B1Z4-75:300; NCs-4) caused a maximum growth inhibition of 20 mm against A. baumannii and a minimum growth inhibition of 12 mm against B. filamentosus at 80 μg mL−1 concentrations of the NPs and NCs. Thus, NCs-4 might be a suitable alternative to further explore and develop as an antibacterial agent. The obtained results statistically justified the data (p ≤ 0.05) via one-way analysis of variance (ANOVA). According to the results of the antibacterial and photocatalytic study, we selected the best bimetallic NCs-4 for the photoexcited antibacterial effect of MDRs, including Gram ve+ and Gram ve− strains, via UV light irradiation. The flower-like NCs-4 composites showed more effectiveness than those of BiPO4, ZnO, and other ratios of NCs. The results encourage the development of flower-like NCs-4 to enhance the photocatalytic antibacterial technique for water purification

Environmental conditions inside and outside the CO2 vent (Ischia, Italy) before and during a reciprocal transplant experiment with Platynereis spp. in September 2013

Platynereis spp. were collected via snorkelling or scuba from either inside (40°43′53″N, 13°57′47″E) (stations A1-A3) or outside (40°43'33.33"N, 13°57'36.38"E and 40°44′48″N, 13°56′39″E) (stations C1-C3) the carbon dioxide (CO2) vent on the island of Ischia (Italy) and used in a reciprocal transplant experiment between 04/09/2013 and 16/09/2013. The effect of exposure to high or low partial pressures of CO2 (pCO2) conditions on the proteome and metabolome (metabolome, and lipidome) of worms from different pCO2 regimes was investigated to understand the effect of exposure to different pCO2 conditions on the cellular physiological response. Before and during the transplant experiment water samples were taken to characterise the physio-chemical parameters of sea water used when exposing the worms to: (i) low pCO2 conditions (‘control’ stations C1, C2 and C3) and (ii) high pCO2 conditions (‘acidified’ stations A1, A2, A3). Salinity, temperature, pH NBS (Mettler-Toledo pH meter, Beaumont Leys, UK), total alkalinity (TA) (AS-ALK2, Apollo SciTech, Bogart, USA), dissolved inorganic carbon (DIC), carbon dioxide partial pressure (pCO2), bicarbonate and carbonate ion concentration ([HCO3–] and [CO32–), calcite and aragonite saturation state (Ωcal and Ωara) are provided

Metabolomic profiles of Platynereis spp. collected from inside and outside the CO2 vent (Ischia, Italy) and used in a reciprocal transplant experiment in September 2013

Platynereis spp. were collected via snorkelling or scuba from either inside (40°43′53″N, 13°57′47″E) or outside (40°43'33.33N, 13°57'36.38E and 40°44′48″N, 13°56′39″E) the carbon dioxide (CO2) vent on the island of Ischia (Italy) and used in a reciprocal transplant experiment. The effect of exposure to high or low partial pressures of CO2 (pCO2) conditions on the metabolome (metabolome, and lipidome) of worms from different pCO2 regimes was investigated to understand the effect of exposure to different pCO2 conditions on the cellular physiological response. This experiment was conducted between 04/09/2013 and 16/09/2013. The experiment was staggered during this time so all worms could be processed. After five days exposure to either low or high CO2 conditions worms were snap frozen in liquid nitrogen and shipped to the University of Birmingham for metabolomic analysis which was finalised on 21/01/2016. Metabolomic profiles of worms were characterised using a mass spectrometry approach. A standard mass spectrometry based metabolomics workflow was used to analyse both the polar and lipid extracts from the samples (Kirwan et al. 2014). Raw mass spectral data were processed using the SIM-stitching algorithm, using an in-house Matlab script. The data matrices were normalized using the PQN algorithm. Missing values were imputed using the KNN algorithm. The resulting data matrix was analysed using univariate statistics, described below. The same matrix was transformed using the generalised logarithm to stabilise the technical variance across the measured peaks prior to analysis using multivariate statistics. Signals were putatively annotated with empirical formulae calculated by the MIPack software (Weber et al. 2010), searching the KEGG (Kanehisa et al. 2012) and LipidMaps (Fahy et al. 2007) databases, and confirmed by performing calculations based on the original spectra in Xcalibur 2.0.7 (Thermo Fisher Scientific)