Evaluation of individual protein errors in silver-stained two-dimensional gels

The relationship between spot volume and variation for all protein spots observed on large format 2D gels when utilising silver stain technology and a model system based on mammalian NSO cell extracts is reported. By running multiple gels we have shown that the reproducibility of data generated in this way is dependent on individual protein spot volumes, which in turn are directly correlated with the coefficient of variation. The coefficients of variation across all observed protein spots were highest for low abundant proteins which are the primary contributors to process error, and lowest for more abundant proteins. Using the relationship between spot volume and coefficient of variation we show it is necessary to calculate variation for individual protein spot volumes. The inherent limitations of silver staining therefore mean that errors in individual protein spot volumes must be considered when assessing significant changes in protein spot volume and not global error. (C) 2003 Elsevier Science (USA). All rights reserved.

Cell death and degeneration in the symbiotic dinoflagellates of the coral Stylophora pistillata during bleaching

Rising sea temperatures are increasing the incidences of mass coral bleaching (the dissociation of the coral-algal symbiosis) and coral mortality. In this study, the effects of bleaching (induced by elevated light and temperature) on the condition of symbiotic dinoflagellates (Symbiodinium sp.) within the tissue of the hard coral Stylophora pistillata (Esper) were assessed using a suite of techniques. Bleaching of S. pistillata was accompanied by declines in the maximum potential quantum yield of photosynthesis (F-v/F-m, measured using pulse amplitude modulated [PAM] fluorometry), an increase in the number of Sytox-green-stained algae (indicating compromised algal membrane integrity and cell death), an increase in 2',7'-dichlorodihydrofluroscein diacetate (H(2)DCFDA)stained algae (indicating increased oxidative stress), as well as ultrastructural changes (vacuolisation, losses of chlorophyll, and an increase in accumulation bodies). Algae expelled from S. pistillata exhibited a complete disorganisation of cellular contents; expelled cells contained only amorphous material. In situ samples taken during a natural mass coral bleaching event on the Great Barrier Reef in February 2002 also revealed a high number of Sytox-labelled algae cells in symbio. Dinoflagellate degeneration during bleaching seems to be similar to the changes resulting from senescence-phase cell death in cultured algae. These data support a role for oxidative stress in the mechanism of coral bleaching and highlight the importance of algal degeneration during the bleaching of a reef coral.

Scleractinian corals with photoprotective host pigments are hypersensitive to thermal bleaching

Recent episodes of mass coral bleaching, the loss of symbiotic dinoflagellates or photosynthetic pigment from hermatypic corals, have been triggered by elevated sea temperatures. Photosynthetic irradiance is an important secondary factor. Host based pigments (pocilloporins or Green Fluorescent Protein homologues) have been proposed to reduce the impact of elevated temperature by shading the dinoflagellate symbionts of corals, thereby reducing light stress. This study investigates this phenomenon in the reef-building coral Acropora aspera from Heron Island Research Station (Great Barrier Reef, Australia), which occurs as 3 distinct colour morphs. Experimental data showed that the host pigments are photoprotective at normal temperatures or

Testing the 'photoinhibition' model of coral bleaching using chemical inhibitors

Explants of the hard coral Seriatopora hystrix were exposed to sublethal concentrations of the herbicide diuron DCMU (N'-(3,4-dichlorophenyl,-N,N-dimethylurea)) and the heavy metal copper. Pulse amplitude modulated (PAM) chlorophyll fluorescence techniques were used to assess the effects on the photosynthetic efficiency of the algal symbionts in the tissue (in Symbio), and chlorophyll fluorescence and counts of symbiotic algae (normalised to surface area) were used to assess the extent of coral bleaching. At 30 mug DCMU l(-1), there was a reduction in both the maximum effective quantum yield (DeltaF/F-m') and maximum potential quantum yield (F-v/F-m) of the algal symbionts in symbio. Corals subsequently lost their algal symbionts and discoloured (bleached), especially on their upper sunlight-exposed surfaces. At the same DCMU concentration but under low light (5% of growth irradiance), there was a marked reduction in DeltaF/F-m' but only a slight reduction in F-v/F-m and slight loss of algae. Loss of algal symbionts was also noted after a 7 d exposure to concentrations as low as 10 mug DCMU l(-1) under normal growth irradiance, and after 14 d exposure to 10 mug DCMU l(-1) under reduced irradiance. Collectively the results indicate that DCMU-induced bleaching is caused by a light-dependent photoinactivation of algal symbionts, and that bleaching occurs when F-v/F-n, (measured 2 h after sunset) is reduced to a value of less than or equal to 0.6. Elevated copper concentrations (60 mug Cu l(-1) for 10 h) also induced a rapid bleaching in S. hystrix but without affecting the quantum yield of the algae in symbio. Tests with isolated algae indicated that substantially higher concentrations (300 mug Cu l(-1) for 8 h) were needed to significantly reduce the quantum yield. Thus, copper-induced bleaching occurs without affecting the algal photosynthesis and may be related to effects on the host (animal). It is argued that warm-water bleaching of corals resembles both types of chemically induced bleaching, suggesting the need for an integrated model of coral bleaching involving the effect of temperature on both host (coral) and algal symbionts.

Endolithic algae photoacclimate to increased irradiance during coral bleaching

The photoacclimation of endolithic algae ( of the genus Ostreobium) inhabiting the skeleton of the Mediterranean coral Oculina patagonica during a bleaching event was examined. Pulse amplitude modulated (PAM) chlorophyll fluorescence techniques in situ were used to assess the photosynthetic efficiency of endolithic algae in the coral skeleton and the symbiotic dinoflagellates (zooxanthellae) in the coral tissue. Relative photosynthetic electron transport rates (ETRs) of the endolithic algae under bleached areas of the colony were significantly higher than those of endolithic algae from a healthy section of the colony and those of zooxanthellae isolated from the same section. Endolithic algae under healthy parts of the colony demonstrated an ETRmax of 16.5% that of zooxanthellae from tissue in the same section whereas endolithic algae under bleached sections showed ETRmax values that were 39% of those found for healthy zooxanthellae. The study demonstrates that endolithic algae undergo photoacclimation with increased irradiance reaching the skeleton. As PAM fluorometry has become a major tool for assessing levels of stress and bleaching in corals, the importance of considering the contribution of the endolithic algae to the overall chlorophyll fluorescence measured is highlighted.

Global assessment of coral bleaching and required rates of adaptation under climate change

Elevated ocean temperatures can cause coral bleaching, the loss of colour from reef-building corals because of a breakdown of the symbiosis with the dinoflagellate Symbiodinium. Recent studies have warned that global climate change could increase the frequency of coral bleaching and threaten the long-term viability of coral reefs. These assertions are based on projecting the coarse output from atmosphere-ocean general circulation models (GCMs) to the local conditions around representative coral reefs. Here, we conduct the first comprehensive global assessment of coral bleaching under climate change by adapting the NOAA Coral Reef Watch bleaching prediction method to the output of a low- and high-climate sensitivity GCM. First, we develop and test algorithms for predicting mass coral bleaching with GCM-resolution sea surface temperatures for thousands of coral reefs, using a global coral reef map and 1985-2002 bleaching prediction data. We then use the algorithms to determine the frequency of coral bleaching and required thermal adaptation by corals and their endosymbionts under two different emissions scenarios. The results indicate that bleaching could become an annual or biannual event for the vast majority of the world's coral reefs in the next 30-50 years without an increase in thermal tolerance of 0.2-1.0 degrees C per decade. The geographic variability in required thermal adaptation found in each model and emissions scenario suggests that coral reefs in some regions, like Micronesia and western Polynesia, may be particularly vulnerable to climate change. Advances in modelling and monitoring will refine the forecast for individual reefs, but this assessment concludes that the global prognosis is unlikely to change without an accelerated effort to stabilize atmospheric greenhouse gas concentrations.

Coral bleaching following wintry weather

Extensive coral bleaching Occurred intertidally in early August 2003 in the Capricorn Bunker group (Wistari Reef, Heron and One Tree Islands; Southern Great Barrier Reef). The affected intertidal coral had been exposed to unusually cold (minimum = 13.3degreesC; wet bulb temperature = 9degreesC) and dry winds (44% relative humidity) for 2 d during predawn low tides. Coral bleached in the upper 10 cm of their branches and had less than 0.2 x 10(6) cell cm(-2) as compared with over 2.5 x 10(6), Cell cm(-2) in nonbleached areas. Dark-adapted quantum yields did not differ between symbionts in bleached and nonbleached areas. Exposing symbionts to light, however, led to greater quenching of Photosystem 11 in symbionts in the bleached coral. Bleached areas of the affected colonies had died by September 2003, with areas that were essentially covered by more than 80% living coral decreasing to less than 10% visible living coral cover. By January 2004, coral began to recover, principally from areas of colonies that were not exposed during low tide (i.e., from below dead, upper regions). These data highlight the importance of understanding local weather patterns as well as the effects of longer term trends in global climate.

Increased mortality and photoinhibition in the symbiotic dinoflagellates of the Indo-Pacific coral Stylophora pistillata (Esper) after summer bleaching

Coral bleaching (the loss of symbiotic dinoflagellates from reef-building corals) is most frequently caused by high-light and temperature conditions. We exposed the explants of the hermatypic coral Stylophora pistillata to four combinations of light and temperature in late spring and also in late summer. During mid-summer, two NOAA bleaching warnings were issued for Heron Island reef (Southern Great Barrier Reef, Australia) when sea temperature exceeded the NOAA bleaching threshold, and a 'mild' (in terms of the whole coral community) bleaching event occurred, resulting in widespread S. pistillata bleaching and mortality. Symbiotic dinoflagellate biomass decreased by more than half from late spring to late summer (from 2.5x10(6) to 0.8x10(6) dinoflagellates cm(2) coral tissue), and those dinoflagellates that remained after summer became photoinhibited more readily (dark-adapted F (V) : F (M) decreased to (0.3 compared with 0.4 in spring), and died in greater numbers (up to 17% dinoflagellate mortality compared with 5% in the spring) when exposed to artificially elevated light and temperature. Adding exogenous antioxidants (D-mannitol and L-ascorbic acid) to the water surrounding the coral had no clear effect on either photoinhibition or symbiont mortality. These data show that light and temperature stress cause mortality of the dinoflagellate symbionts within the coral, and that susceptibility to light and temperature stress is strongly related to coral condition. Photoinhibitory mechanisms are clearly involved, and will increase through a positive feedback mechanism: symbiont loss promotes further symbiont loss as the light microenvironment becomes progressively harsher.

Monitoring coral bleaching using a colour reference card

Assessment of the extent of coral bleaching has become an important part of studies that aim to understand the condition of coral reefs. In this study a reference card that uses differences in coral colour was developed as an inexpensive, rapid and non-invasive method for the assessment of bleaching. The card uses a 6 point brightness/saturation scale within four colour hues to record changes in bleaching state. Changes on the scale of 2 units or more reflect a change in symbiont density and chlorophyll a content, and therefore the bleaching state of the coral. When used by non-specialist observers in the field (here on an intertidal reef flat), there was an inter-observer error of I colour score. This technique improves on existing subjective assessment of bleaching state by visual observation and offers the potential for rapid, wide-area assessment of changing coral condition.