Reversal of ocean acidification enhances net coral reef calcification.

Approximately one-quarter of the anthropogenic carbon dioxide released into the atmosphere each year is absorbed by the global oceans, causing measurable declines in surface ocean pH, carbonate ion concentration ([CO3(2-)]), and saturation state of carbonate minerals (Ω). This process, referred to as ocean acidification, represents a major threat to marine ecosystems, in particular marine calcifiers such as oysters, crabs, and corals. Laboratory and field studies have shown that calcification rates of many organisms decrease with declining pH, [CO3(2-)], and Ω. Coral reefs are widely regarded as one of the most vulnerable marine ecosystems to ocean acidification, in part because the very architecture of the ecosystem is reliant on carbonate-secreting organisms. Acidification-induced reductions in calcification are projected to shift coral reefs from a state of net accretion to one of net dissolution this century. While retrospective studies show large-scale declines in coral, and community, calcification over recent decades, determining the contribution of ocean acidification to these changes is difficult, if not impossible, owing to the confounding effects of other environmental factors such as temperature. Here we quantify the net calcification response of a coral reef flat to alkalinity enrichment, and show that, when ocean chemistry is restored closer to pre-industrial conditions, net community calcification increases. In providing results from the first seawater chemistry manipulation experiment of a natural coral reef community, we provide evidence that net community calcification is depressed compared with values expected for pre-industrial conditions, indicating that ocean acidification may already be impairing coral reef growth.

Insights into the autotransport process of a trimeric autotransporter, Yersinia Adhesin A (YadA).

Trimeric autotransporter adhesins (TAAs) are a subset of a larger protein family called the type V secretion systems. They are localized on the cell surface of Gram-negative bacteria, function as mediators of attachment to inorganic surfaces and host cells, and thus include important virulence factors. Yersinia adhesin A (YadA) from Yersinia enterocolitica is a prototypical TAA that is used extensively to study the structure and function of the type Vc secretion system. A solid-state NMR study of the membrane anchor domain of YadA previously revealed a flexible stretch of small residues, termed the ASSA region, that links the membrane anchor to the stalk domain. In this study, we present evidence that single amino acid proline substitutions produce two different conformers of the membrane anchor domain of YadA; one with the N-termini facing the extracellular surface, and a second with the N-termini located in the periplasm. We propose that TAAs adopt a hairpin intermediate during secretion, as has been shown before for other subtypes of the type V secretion system. As the YadA transition state intermediate can be isolated from the outer membrane, future structural studies should be possible to further unravel details of the autotransport process.

The inverse autotransporters of Yersinia ruckeri, YrInv and YrIlm, contribute to biofilm formation and virulence.

Yersinia ruckeri causes enteric redmouth disease (ERM) that mainly affects salmonid fishes and leads to significant economic losses in the aquaculture industry. An increasing number of outbreaks and the lack of effective vaccines against some serotypes necessitates novel measures to control ERM. Importantly, Y. ruckeri survives in the environment for long periods, presumably by forming biofilms. How the pathogen forms biofilms and which molecular factors are involved in this process, remains unclear. Yersinia ruckeri produces two surface-exposed adhesins, belonging to the inverse autotransporters (IATs), called Y. ruckeri invasin (YrInv) and Y. ruckeri invasin-like molecule (YrIlm). Here, we investigated whether YrInv and YrIlm play a role in biofilm formation and virulence. Functional assays revealed that YrInv and YrIlm promote biofilm formation on different abiotic substrates. Confocal microscopy revealed that they are involved in microcolony interaction and formation, respectively. The effect of both IATs on biofilm formation correlated with the presence of different biopolymers in the biofilm matrix, including extracellular DNA, RNA and proteins. Moreover, YrInv and YrIlm contributed to virulence in the Galleria mellonella infection model. Taken together, we propose that both IATs are possible targets for the development of novel diagnostic and preventative strategies to control ERM.

Trapping of redox-mediators at the surface of Chlorella vulgaris leads to error in measurements of cell reducing power.

The reduction of the redox mediator ferricyanide, [Fe(CN)6](3-), by a range of algal and bacterial species, is frequently measured to probe plasma membrane ferrireductase activity or to quantify the reducing power of algal/bacterial biofilms and suspensions. In this study we have used rotating disk electrochemistry (RDE) to investigate the reduction of ferricyanide by the model organism Chlorella vulgaris. Importantly, we have seen that the diffusion limited current due to the oxidation of ferrocyanide, [Fe(CN)6](4-), at the electrode decreased linearly as C. vulgaris was added to the solution, even though in a pure ferrocyanide solution the algae are not able to reduce the mediator further and are simply spectator 'particles'. We attribute this effect to trapping of ferrocyanide at the cell surface, with up to 14% of the ferrocyanide missing from the solution at the highest cell concentration. The result has important implications for all techniques that use electrochemistry and other concentration dependent assays (e.g. fluorescence and colourimetry) to monitor ferrocyanide concentrations in the presence of both biofilms and cell suspensions. Analyte trapping could lead to a substantial underestimation of the concentration of reduced product.

EC-SERS detection of thiabendazole in apple juice using activated screen-printed electrodes.

Thiabendazole (TBZ), a benzimidazole fungicide used for post-harvest treatment, may be a trace contaminant of food matrices. In this work, we report the first EC-SERS (electrochemical-surface enhanced Raman spectroscopy) detection of TBZ in spiked apple juice using electrochemically (EC) roughened, gold-based screen-printed electrodes (AuSPEs) and portable instrumentation. Polarizing the substrate (-0.8 V vs Ag/AgCl) improves the recorded SERS signal of TBZ, allowing to reach a limit of detection (LOD) in juice of 0.061 ppm with a relatively wide linear range (0.5-10 µM) and good intermediate precision (%RSD < 10). The recovery of TBZ from unprocessed juice was found to be more than 82 %. Furthermore, a proof-of-concept integration of AuSPEs with a miniaturized flow cell for the preconcentration of TBZ and the controlled delivery of sample and reagents has been demonstrated. This approach paves the way for integrated, portable analytical systems applicable for on-site sample collection, processing, and analysis.

Surface morphology and surface energy of anode materials influence power outputs in a multi-channel mediatorless bio-photovoltaic (BPV) system.

Bio-photovoltaic cells (BPVs) are a new photo-bio-electrochemical technology for harnessing solar energy using the photosynthetic activity of autotrophic organisms. Currently power outputs from BPVs are generally low and suffer from low efficiencies. However, a better understanding of the electrochemical interactions between the microbes and conductive materials will be likely to lead to increased power yields. In the current study, the fresh-water, filamentous cyanobacterium Pseudanabaena limnetica (also known as Oscillatoria limnetica) was investigated for exoelectrogenic activity. Biofilms of P. limnetica showed a significant photo response during light-dark cycling in BPVs under mediatorless conditions. A multi-channel BPV device was developed to compare quantitatively the performance of photosynthetic biofilms of this species using a variety of different anodic conductive materials: indium tin oxide-coated polyethylene terephthalate (ITO), stainless steel (SS), glass coated with a conductive polymer (PANI), and carbon paper (CP). Although biofilm growth rates were generally comparable on all materials tested, the amplitude of the photo response and achievable maximum power outputs were significantly different. ITO and SS demonstrated the largest photo responses, whereas CP showed the lowest power outputs under both light and dark conditions. Furthermore, differences in the ratios of light : dark power outputs indicated that the electrochemical interactions between photosynthetic microbes and the anode may differ under light and dark conditions depending on the anodic material used. Comparisons between BPV performances and material characteristics revealed that surface roughness and surface energy, particularly the ratio of non-polar to polar interactions (the CQ ratio), may be more important than available surface area in determining biocompatibility and maximum power outputs in microbial electrochemical systems. Notably, CP was readily outperformed by all other conductive materials tested, indicating that carbon may not be an optimal substrate for microbial fuel cell operation.

Breast cancer risk in relation to occupations with exposure to carcinogens and endocrine disruptors: a Canadian case-control study.

BACKGROUND: Endocrine disrupting chemicals and carcinogens, some of which may not yet have been classified as such, are present in many occupational environments and could increase breast cancer risk. Prior research has identified associations with breast cancer and work in agricultural and industrial settings. The purpose of this study was to further characterize possible links between breast cancer risk and occupation, particularly in farming and manufacturing, as well as to examine the impacts of early agricultural exposures, and exposure effects that are specific to the endocrine receptor status of tumours. METHODS: 1005 breast cancer cases referred by a regional cancer center and 1146 randomly-selected community controls provided detailed data including occupational and reproductive histories. All reported jobs were industry- and occupation-coded for the construction of cumulative exposure metrics representing likely exposure to carcinogens and endocrine disruptors. In a frequency-matched case-control design, exposure effects were estimated using conditional logistic regression. RESULTS: Across all sectors, women in jobs with potentially high exposures to carcinogens and endocrine disruptors had elevated breast cancer risk (OR = 1.42; 95% CI, 1.18-1.73, for 10 years exposure duration). Specific sectors with elevated risk included: agriculture (OR = 1.36; 95% CI, 1.01-1.82); bars-gambling (OR = 2.28; 95% CI, 0.94-5.53); automotive plastics manufacturing (OR = 2.68; 95% CI, 1.47-4.88), food canning (OR = 2.35; 95% CI, 1.00-5.53), and metalworking (OR = 1.73; 95% CI, 1.02-2.92). Estrogen receptor status of tumors with elevated risk differed by occupational grouping. Premenopausal breast cancer risk was highest for automotive plastics (OR = 4.76; 95% CI, 1.58-14.4) and food canning (OR = 5.70; 95% CI, 1.03-31.5). CONCLUSIONS: These observations support hypotheses linking breast cancer risk and exposures likely to include carcinogens and endocrine disruptors, and demonstrate the value of detailed work histories in environmental and occupational epidemiology.

Living coral tissue slows skeletal dissolution related to ocean acidification.

Climate change is causing major changes to marine ecosystems globally, with ocean acidification of particular concern for coral reefs. Using a 200 d in situ carbon dioxide enrichment study on Heron Island, Australia, we simulated future ocean acidification conditions, and found reduced pH led to a drastic decline in net calcification of living corals to no net growth, and accelerated disintegration of dead corals. Net calcification declined more severely than in previous studies due to exposure to the natural community of bioeroding organisms in this in situ study and to a longer experimental duration. Our data suggest that reef flat corals reach net dissolution at an aragonite saturation state (ΩAR) of 2.3 (95% confidence interval: 1.8-2.8) with 100% living coral cover and at ΩAR > 3.5 with 30% living coral cover. This model suggests that areas of the reef with relatively low coral mortality, where living coral cover is high, are likely to be resistant to carbon dioxide-induced reef dissolution.

Sulfide-induced nitrate reduction in the sludge of an anaerobic digester of a zero-discharge recirculating mariculture system.

The anaerobic digester is a vital component in a zero-discharge mariculture system as therein most of the organic matter is mineralized and nitrogen-containing compounds are converted to gaseous N(2). Although denitrification is a major respiratory process in this nitrate-rich treatment stage, also sulfate respiration takes place and may cause undesirable high sulfide concentrations in the effluent water. To examine the effect of sulfide on nitrate reduction, in situ depth profiles of inorganic nitrogen and sulfur compounds were determined. Additionally, nitrate reduction was examined as a function of ambient sulfide concentrations in sludge collected from different locations in the anaerobic reactor. Depth profiles showed high concentrations of nitrate and low concentrations of sulfide and ammonia in the aqueous layer of the reactor. A sharp decrease of nitrate and an increase in sulfide and ammonia concentrations was measured at the water-sludge interface. Nitrate reduction was highest in this interface zone with rates of up to 8.05+/-0.57 micromol NO(3)(-)h(-1)g((sludge))(-1). Addition of sulfide increased the nitrate reduction rate at all sludge depths, pointing to the important role of autotrophic denitrification in the anaerobic reactor. Dissimilatory nitrate reduction to ammonia (DNRA) was found to be low in all sludge layers but was enhanced when sludge was incubated at high sulfide concentrations. Although nitrate reduction rates increased as a result of sulfide addition to sludge samples, no differences in nitrate reduction rates were observed between the samples incubated with different initial sulfide concentrations. This as opposed to sulfide oxidation rates, which followed Michaelis-Menten enzymatic kinetics. Partial oxidation of sulfide to elemental sulfur instead of a complete oxidation to sulfate, could explain the observed patterns of nitrate reduction and sulfide oxidation in sludge incubated with different initial sulfide concentrations.

Linoleic acid isomerase from Propionibacterium acnes: purification, characterization, molecular cloning, and heterologous expression.

Propionibacterium acnes strain ATCC 6919 catalyzes the isomerization of the double bond at the C9 position in linoleic acid (c9,c12, 18:2) to form t10,c12 conjugated linoleic acid (CLA, 18:2). CLA has significant health benefits in animal and human. The linoleic acid C9 isomerase was purified to an apparent homogeneity by successive chromatography on diethylaminoethyl (DEAE) anion exchange, hydrophobic interaction, and chromatofocusing columns. Two degenerated oligonucleotide primers were synthesized according to the N-terminal peptide sequence to clone, by polymerase chain reaction (PCR), a short nucleotide sequence (62 bp) of the isomerase gene. The linoleic acid isomerase gene (lai) was subsequently cloned by inverse PCR. The amino acid sequence deduced from the lai coding sequence predicts a protein of 424 amino acid residues (48 kDa), excluding the N-terminal methionine, which was absent in the polypeptide purified from the native host. The isomerase shares no significant sequence homology to other enzymes except a flavin-binding domain in the N-terminal region. The recombinant isomerase purified from Escherichia coli showed a typical ultraviolet spectrum for FAD-bound proteins. The recombinant enzyme produced a single isomer of t10,c12-CLA from linoleic acid, as demonstrated by gas chromatography and gas chromatography-mass spectrum analysis. The recombinant isomerase protein was expressed at high levels in E. coli, but it was almost totally sequestered in inclusion bodies. The level of active isomerase was increased 376-fold by medium and process optimization in bench-scale fermentors.

Occupation and breast cancer: a Canadian case-control study.

A local collaborative process was launched in Windsor, Ontario, Canada to explore the role of occupation as a risk factor for cancer. An initial hypothesis-generating study found an increased risk for breast cancer among women aged 55 years or younger who had ever worked in farming. On the basis of this result, a 2-year case-control study was undertaken to evaluate the lifetime occupational histories of women with breast cancer. The results indicate that women with breast cancer were nearly three times more likely to have worked in agriculture when compared to the controls (OR = 2.80 [95% CI, 1.6-4.8]). The risk for those who worked in agriculture and subsequently worked in automotive-related manufacturing was further elevated (OR = 4.0 [95% CI, 1.7-9.9]). The risk for those employed in agriculture and subsequently employed in health care was also elevated (OR = 2.3 [95% CI, 1.1-4.6]). Farming tended to be among the earlier jobs worked, often during adolescence. While this article has limitations including the small sample size and the lack of information regarding specific exposures, it does provide evidence of a possible association between farming and breast cancer. The findings indicate the need for further study to determine which aspects of farming may be of biological importance and to better understand the significance of timing of exposure in terms of cancer risk.

An investigation of anode and cathode materials in photomicrobial fuel cells.

Photomicrobial fuel cells (p-MFCs) are devices that use photosynthetic organisms (such as cyanobacteria or algae) to turn light energy into electrical energy. In a p-MFC, the anode accepts electrons from microorganisms that are either growing directly on the anode surface (biofilm) or are free floating in solution (planktonic). The nature of both the anode and cathode material is critical for device efficiency. An ideal anode is biocompatible and facilitates direct electron transfer from the microorganisms, with no need for an electron mediator. For a p-MFC, there is the additional requirement that the anode should not prevent light from perfusing through the photosynthetic cells. The cathode should facilitate the rapid reaction of protons and oxygen to form water so as not to rate limit the device. In this paper, we first review the range of anode and cathode materials currently used in p-MFCs. We then present our own data comparing cathode materials in a p-MFC and our first results using porous ceramic anodes in a mediator-free p-MFC.

Estimation of Tumor Interstitial Fluid Pressure (TIFP) Noninvasively.

Tumor interstitial fluid pressure (TIFP), is a physiological parameter with demonstrated predictive value for a tumor's aggressiveness, drug delivery, as well as response to treatments such as radiotherapy and chemotherapy. Despite its utility, measurement of TIFP has been limited by the need for invasive procedures. In this work, the theoretical basis for approaching the absolute value of TIFP and the experimental method for noninvasively measuring TIFP are presented. Given specific boundary and continuity conditions, we convert theoretical variables into measurable variables by applying MRI technology. The work shows that TIFP in the central region of the tumor can be estimated by an analysis of the variation of tissue fluid motion in the tumor rim and surrounding tissue. It is determined from three noninvasive measurable parameters: i) an estimate of the velocity of the tumor interstitial fluid at the tumor surface, which is maximal, ii) a measurement of the distance from the tumor surface to where the tumor exudates are absorbed (or normalized), and iii) an estimate of the hydraulic conductivity of the interstitium through which the tumor exudate travels. We experimentally show that the fluid flow within the tumor rim is not uniform, even for a round shaped tumor, and demonstrate the procedures for the noninvasive measurement of TIFP.

Evidence for Rhythmicity Pacemaker in the Calcification Process of Scleractinian Coral.

Reef-building scleractinian (stony) corals are among the most efficient bio-mineralizing organisms in nature. The calcification rate of scleractinian corals oscillates under ambient light conditions, with a cyclic, diurnal pattern. A fundamental question is whether this cyclic pattern is controlled by exogenous signals or by an endogenous 'biological-clock' mechanism, or both. To address this problem, we have studied calcification patterns of the Red Sea scleractinian coral Acropora eurystoma with frequent measurements of total alkalinity (AT) under different light conditions. Additionally, skeletal extension and ultra-structure of newly deposited calcium carbonate were elucidated with (86)Sr isotope labeling analysis, combined with NanoSIMS ion microprobe and scanning electron microscope imaging. Our results show that the calcification process persists with its cyclic pattern under constant light conditions while dissolution takes place within one day of constant dark conditions, indicating that an intrinsic, light-entrained mechanism may be involved in controlling the calcification process in photosymbiotic corals.

Iron reduction by the cyanobacterium Synechocystis sp. PCC 6803.

Synechocystis sp. PCC 6803 uptakes iron using a reductive mechanism, similar to that exhibited by many other microalgae. Various bio-electrochemical technologies have made use of this reductive cellular capacity, but there is still a lack of fundamental understanding of cellular reduction rates under different conditions. This study used electrochemical techniques to further investigate the reductive interactions of Synechocystis cells with Fe(III) from the iron species potassium ferricyanide, with varying cell and ferricyanide concentrations present. At the lowest cell concentrations tested, cell reduction machinery appeared to kinetically limit the reduction reaction, but ferricyanide reduction rates were mass transport controlled at the higher cell and ferricyanide concentrations studied. Improving the understanding of the reduction of Fe(III) by whole cyanobacterial cells is important for improving the efficiencies of technologies that rely on this interaction.

Six Month In Situ High-Resolution Carbonate Chemistry and Temperature Study on a Coral Reef Flat Reveals Asynchronous pH and Temperature Anomalies.

Understanding the temporal dynamics of present thermal and pH exposure on coral reefs is crucial for elucidating reef response to future global change. Diel ranges in temperature and carbonate chemistry parameters coupled with seasonal changes in the mean conditions define periods during the year when a reef habitat is exposed to anomalous thermal and/or pH exposure. Anomalous conditions are defined as values that exceed an empirically estimated threshold for each variable. We present a 200-day time series from June through December 2010 of carbonate chemistry and environmental parameters measured on the Heron Island reef flat. These data reveal that aragonite saturation state, pH, and pCO2 were primarily modulated by biologically-driven changes in dissolved organic carbon (DIC) and total alkalinity (TA), rather than salinity and temperature. The largest diel temperature ranges occurred in austral spring, in October (1.5 - 6.6°C) and lowest diel ranges (0.9 - 3.2°C) were observed in July, at the peak of winter. We observed large diel total pH variability, with a maximum range of 7.7 - 8.5 total pH units, with minimum diel average pH values occurring during spring and maximum during fall. As with many other reefs, the nighttime pH minima on the reef flat were far lower than pH values predicted for the open ocean by 2100. DIC and TA both increased from June (end of Fall) to December (end of Spring). Using this high-resolution dataset, we developed exposure metrics of pH and temperature individually for intensity, duration, and severity of low pH and high temperature events, as well as a combined metric. Periods of anomalous temperature and pH exposure were asynchronous on the Heron Island reef flat, which underlines the importance of understanding the dynamics of co-occurrence of multiple stressors on coastal ecosystems.

A small-scale air-cathode microbial fuel cell for on-line monitoring of water quality.

The heavy use of chemicals for agricultural, industrial and domestic purposes has increased the risk of freshwater contamination worldwide. Consequently, the demand for efficient new analytical tools for on-line and on-site water quality monitoring has become particularly urgent. In this study, a small-scale single chamber air-cathode microbial fuel cell (SCMFC), fabricated by rapid prototyping layer-by-layer 3D printing, was tested as a biosensor for continuous water quality monitoring. When acetate was fed as the rate-limiting substrate, the SCMFC acted as a sensor for chemical oxygen demand (COD) in water. The linear detection range was 3-164 ppm, with a sensitivity of 0.05 µA mM(-1) cm(-2) with respect to the anode total surface area. The response time was as fast as 2.8 min. At saturating acetate concentrations (COD>164 ppm), the miniature SCMFC could rapidly detect the presence of cadmium in water with high sensitivity (0.2 µg l(-1) cm(-2)) and a lower detection limit of only 1 µg l(-1). The biosensor dynamic range was 1-25 µg l(-1). Within this range of concentrations, cadmium affected only temporarily the electroactive biofilm at the anode. When the SCMFCs were again fed with fresh wastewater and no pollutant, the initial steady-state current was recovered within 12 min.

A short-term in situ CO2 enrichment experiment on Heron Island (GBR).

Ocean acidification poses multiple challenges for coral reefs on molecular to ecological scales, yet previous experimental studies of the impact of projected CO2 concentrations have mostly been done in aquarium systems with corals removed from their natural ecosystem and placed under artificial light and seawater conditions. The Coral-Proto Free Ocean Carbon Enrichment System (CP-FOCE) uses a network of sensors to monitor conditions within each flume and maintain experimental pH as an offset from environmental pH using feedback control on the injection of low pH seawater. Carbonate chemistry conditions maintained in the -0.06 and -0.22 pH offset treatments were significantly different than environmental conditions. The results from this short-term experiment suggest that the CP-FOCE is an important new experimental system to study in situ impacts of ocean acidification on coral reef ecosystems.

Carbon cycling in a zero-discharge mariculture system.

Interest in mariculture systems will rise in the near future due to the decreased ability of the ocean to supply the increasing demand for seafood. We present a trace study using stable carbon and nitrogen isotopes and chemical profiles of a zero-discharge mariculture system stocked with the gilthead seabream (Sparus aurata). Water quality maintenance in the system is based on two biofiltration steps. Firstly, an aerobic treatment step comprising a trickling filter in which ammonia is oxidized to nitrate. Secondly, an anaerobic step comprised of a digestion basin and a fluidized bed reactor where excess organic matter and nitrate are removed. Dissolved inorganic carbon and alkalinity values were higher in the anaerobic loop than in the aerobic loop, in agreement with the main biological processes taking place in the two treatment steps. The δ13C of the dissolved inorganic carbon (δ13C(DIC)) was depleted in 13C in the anaerobic loop as compared to the aerobic loop by 2.5-3‰. This is in agreement with the higher dissolved inorganic carbon concentrations in the anaerobic loop and the low water retention time and the chemolithotrophic activity of the aerobic loop. The δ13C and δ15N of organic matter in the mariculture system indicated that fish fed solely on feed pellets. Compared to feed pellets and particulate organic matter, the sludge in the digestion basin was enriched in 15N while δ13C was not significantly different. This latter finding points to an intensive microbial degradation of the organic matter taking place in the anaerobic treatment step of the system.