Sharks are one of the most threatened groups of marine animals worldwide, mostly owing to overfishing and habitat degradation/loss. Although these cartilaginous fish have evolved to fill many ecological niches across a wide range of habitats, they have limited capability to rapidly adapt to human-induced changes in their environments. Contrary to global warming, ocean acidification was not considered as a direct climate-related threat to sharks. Here we show, for the first time, that an early ontogenetic acclimation process of a tropical shark (Chiloscyllium punctatum) to the projected scenarios of ocean acidification (?pH = 0.5) and warming (+4°C; 30°C) for 2100 elicited significant impairments on juvenile shark condition and survival. The mortality of shark embryos at the present-day thermal scenarios was 0% both at normocapnic and hypercapnic conditions. Yet routine metabolic rates (RMRs) were significantly affected by temperature, pH and embryonic stage. Immediately after hatching, the Fulton condition of juvenile bamboo sharks was significantly different in individuals that experienced future warming and hypercapnia; 30 days after hatching, survival rapidly declined in individuals experiencing both ocean warming and acidification (up to 44%). The RMR of juvenile sharks was also significantly affected by temperature and pH. The impact of low pH on ventilation rates was significant only under the higher thermal scenario. This study highlights the need of experimental-based risk assessments of sharks to climate change. In other words, it is critical to directly assess risk and vulnerability of sharks to ocean acidification and warming, and such effort can ultimately help managers and policy-makers to take proactive measures targeting most endangered species.
The morphological plasticity of scleractinian corals can be influenced by numerous factors in their natural environment. However, it is difficult to identify in situ the relative influence of a single biotic or abiotic factor, due to potential interactions between them. Light is considered as a major factor affecting coral skeleton morphology, due to their symbiotic relation with photosynthetic zooxanthellae. Nonetheless, most studies addressing the importance of light on coral morphological plasticity have focused on photosynthetically active radiation (PAR) intensity, with the effect of light spectra remaining largely unknown. The present study evaluated how different light spectra affect the skeleton macro- and microstructures in two coral species (Acropora formosa sensu Veron (2000) and Stylophora pistillata) maintained under controlled laboratory conditions. We tested the effect of three light treatments with the same PAR but with a distinct spectral emission: 1) T5 fluorescent lamps with blue emission; 2) Light Emitting Diodes (LED) with predominantly blue emission; and 3) Light Emitting Plasma (LEP) with full spectra emission. To exclude potential bias generated by genetic variability, the experiment was performed with clonal fragments for both species. After 6 months of experiment, it was possible to detect in coral fragments of both species exposed to different light spectra significant differences in morphometry (e.g., distance among corallites, corallite diameter, and theca thickness), as well as in the organization of their skeleton microstructure. The variability found in the skeleton macro- and microstructures of clonal organisms points to the potential pitfalls associated with the exclusive use of morphometry on coral taxonomy. Moreover, the identification of a single factor influencing the morphology of coral skeletons is relevant for coral aquaculture and can allow the optimization of reef restoration efforts.
There is growing concern that modifications to the global environment such as ocean acidification and increased ultraviolet radiation may interact with anthropogenic pollutants to adversely affect the future marine environment. Despite this, little is known about the nature of the potential risks posed by such interactions. Here, we performed a multifactorial microcosm experiment to assess the impact of ocean acidification, ultraviolet radiation B (UV-B) and oil hydrocarbon contamination on sediment chemistry, the microbial community (composition and function) and biochemical marker response of selected indicator species. We found that increased ocean acidification and oil contamination in the absence of UV-B will significantly alter bacterial composition by, among other changes, greatly reducing the relative abundance of Desulfobacterales, known to be important oil hydrocarbon degraders. Along with changes in bacterial composition, we identified concomitant shifts in the composition of aromatic hydrocarbons in the sediment and an increase in oxidative stress effects on our indicator species. Interestingly, our study identifies UV-B as a critical component in the interaction between these factors, since its presence alleviates harmful effects caused by the combination of reduced pH and oil pollution. The model system used here shows that the interactive effect of reduced pH and oil contamination can adversely affect the structure and functioning of sediment benthic communities, with the potential to exacerbate the toxicity of oil hydrocarbons in marine ecosystems. This article is protected by copyright. All rights reserved.
Aquaculture facilities worldwide continue to experience significant economic losses because of disease caused by pathogenic bacteria, including multidrug-resistant strains. This scenario drives the search for alternative methods to inactivate pathogenic bacteria. Phage therapy is currently considered as a viable alternative to antibiotics for inactivation of bacterial pathogens in aquaculture systems. While phage therapy appears to represent a useful and flexible tool for microbiological decontamination of aquaculture effluents, the effect of physical and chemical properties of culture waters on the efficiency of this technology has never been reported. The present study aimed to evaluate the effect of physical and chemical properties of aquaculture waters (e.g. pH, temperature, salinity and organic matter content) on the efficiency of phage therapy under controlled experimental conditions in order to provide a basis for the selection of the most suitable protocol for subsequent experiments. A bioluminescent genetically transformed Escherichia coli was selected as a model microorganism to monitor real-time phage therapy kinetics through the measurement of bioluminescence, thus avoiding the laborious and time-consuming conventional method of counting colony-forming units (CFU). For all experiments, a bacterial concentration of ? 10(5) CFU ml(-1) and a phage concentration of ? 10(6-8) plaque forming unit ml(-1) were used. Phage survival was not significantly affected by the natural variability of pH (6.5-7.4), temperature (10-25 °C), salinity (0-30 g NaCl l(-1) ) and organic matter concentration of aquaculture waters in a temperate climate. Nonetheless, the efficiency of phage therapy was mostly affected by the variation of salinity and organic matter content. As the effectiveness of phage therapy increases with water salt content, this approach appears to be a suitable choice for marine aquaculture systems. The success of phage therapy may also be enhanced in non-marine systems through the addition of salt, whenever this option is feasible and does not affect the survival of aquatic species being cultured.
Cleaning symbioses play an important role in the health of certain coastal marine communities. These interspecific associations often occur at specific sites (cleaning stations) where a cleaner organism (commonly a fish or shrimp) removes ectoparasites/damaged tissue from a 'client' (a larger cooperating fish). At present, the potential impact of climate change on the fitness of cleaner organisms remains unknown. This study investigated the physiological and biochemical responses of tropical (Lysmata amboinensis) and temperate (L. seticaudata) cleaner shrimp to global warming. Specifically, thermal limits (CTMax), metabolic rates, thermal sensitivity, heat shock response (HSR), lipid peroxidation [malondialdehyde (MDA) concentration], lactate levels, antioxidant (GST, SOD and catalase) and digestive enzyme activities (trypsin and alkaline phosphatase) at current and warming (+3 °C) temperature conditions. In contrast to the temperate species, CTMax values decreased significantly from current (24-27 °C) to warming temperature conditions (30 °C) for the tropical shrimp, where metabolic thermal sensitivity was affected and the HSR was significantly reduced. MDA levels in tropical shrimp increased dramatically, indicating extreme cellular lipid peroxidation, which was not observed in the temperate shrimp. Lactate levels, GST and SOD activities were significantly enhanced within the muscle tissue of the tropical species. Digestive enzyme activities in the hepatopancreas of both species were significantly decreased by warmer temperatures. Our data suggest that the tropical cleaner shrimp will be more vulnerable to global warming than the temperate Lysmata seticaudata; the latter evolved in a relatively unstable environment with seasonal thermal variations that may have conferred greater adaptive plasticity. Thus, tropical cleaning symbioses may be challenged at a greater degree by warming-related anthropogenic forcing, with potential cascading effects on the health and structuring of tropical coastal communities (e.g. coral reefs).
For several years, knowledge on the microbiome associated with marine invertebrates was impaired by the challenges associated with the characterization of bacterial communities. With the advent of culture independent molecular tools it is possible to gain new insights on the diversity and richness of microorganisms associated with marine invertebrates. In the present study, we evaluated if different preservation and processing methodologies (prior to DNA extraction) can affect the bacterial diversity retrieved from snakelocks anemone Anemonia viridis. Denaturing gradient gel electrophoresis (DGGE) community fingerprints were used as proxy to determine the bacterial diversity retrieved (H'). Statistical analyses indicated that preservation significantly affects H'. The best approach to preserve and process A. viridis biomass for bacterial community fingerprint analysis was flash freezing in liquid nitrogen (preservation) followed by the use of a mechanical homogenizer (process), as it consistently yielded higher H'. Alternatively, biomass samples can be processed fresh followed by cell lyses using a mechanical homogenizer or mortar &pestle. The suitability of employing these two alternative procedures was further reinforced by the quantification of the 16S rRNA gene; no significant differences were recorded when comparing these two approaches and the use of liquid nitrogen followed by processing with a mechanical homogenizer.
Little is known about the capacity of early life stages to undergo hypercapnic and thermal acclimation under the future scenarios of ocean acidification and warming. Here, we investigated a comprehensive set of biological responses to these climate change-related variables (2°C above winter and summer average spawning temperatures and ?pH=0.5 units) during the early ontogeny of the squid Loligo vulgaris. Embryo survival rates ranged from 92% to 96% under present-day temperature (13-17°C) and pH (8.0) scenarios. Yet, ocean acidification (pH 7.5) and summer warming (19°C) led to a significant drop in the survival rates of summer embryos (47%, P<0.05). The embryonic period was shortened by increasing temperature in both pH treatments (P<0.05). Embryo growth rates increased significantly with temperature under present-day scenarios, but there was a significant trend reversal under future summer warming conditions (P<0.05). Besides pronounced premature hatching, a higher percentage of abnormalities was found in summer embryos exposed to future warming and lower pH (P<0.05). Under the hypercapnic scenario, oxygen consumption rates decreased significantly in late embryos and newly hatched paralarvae, especially in the summer period (P<0.05). Concomitantly, there was a significant enhancement of the heat shock response (HSP70/HSC70) with warming in both pH treatments and developmental stages. Upper thermal tolerance limits were positively influenced by acclimation temperature, and such thresholds were significantly higher in late embryos than in hatchlings under present-day conditions (P<0.05). In contrast, the upper thermal tolerance limits under hypercapnia were higher in hatchlings than in embryos. Thus, we show that the stressful abiotic conditions inside the embryo's capsules will be exacerbated under near-future ocean acidification and summer warming scenarios. The occurrence of prolonged embryogenesis along with lowered thermal tolerance limits under such conditions is expected to negatively affect the survival success of squid early life stages during the summer spawning period, but not winter spawning.
The chemical diversity associated with marine natural products (MNP) is unanimously acknowledged as the "blue gold" in the urgent quest for new drugs. Consequently, a significant increase in the discovery of MNP published in the literature has been observed in the past decades, particularly from marine invertebrates. However, it remains unclear whether target metabolites originate from the marine invertebrates themselves or from their microbial symbionts. This issue underlines critical challenges associated with the lack of biomass required to supply the early stages of the drug discovery pipeline. The present review discusses potential solutions for such challenges, with particular emphasis on innovative approaches to culture invertebrate holobionts (microorganism-invertebrate assemblages) through in toto aquaculture, together with methods for the discovery and initial production of bioactive compounds from these microbial symbionts.
Kleptoplasty is a remarkable type of photosynthetic association, resulting from the maintenance of functional chloroplasts--the 'kleptoplasts'--in the tissues of a non-photosynthetic host. It represents a biologically unique condition for chloroplast and photosynthesis functioning, occurring in different phylogenetic lineages, namely dinoflagellates, ciliates, foraminiferans and, most interestingly, a single taxon of metazoans, the sacoglossan sea slugs. In the case of sea slugs, chloroplasts from macroalgae are often maintained as intracellular organelles in cells of these marine gastropods, structurally intact and photosynthetically competent for extended periods of time. Kleptoplasty has long attracted interest owing to the longevity of functional kleptoplasts in the absence of the original algal nucleus and the limited number of proteins encoded by the chloroplast genome. This review updates the state-of-the-art on kleptoplast photophysiology, focusing on the comparative analysis of the responses to light of the chloroplasts when in their original, macroalgal cells, and when sequestered in animal cells and functioning as kleptoplasts. It covers fundamental but ecologically relevant aspects of kleptoplast light responses, such as the occurrence of photoacclimation in hospite, operation of photoprotective processes and susceptibility to photoinhibition. Emphasis is given to host-mediated processes unique to kleptoplastic associations, reviewing current hypotheses on behavioural photoprotection and host-mediated enhancement of photosynthetic performance, and identifying current gaps in sacoglossan kleptoplast photophysiology research.
The advent of industrial activities in the deep sea will inevitably expose deep-sea organisms to potentially toxic compounds. Although international regulations require environmental risk assessment prior to exploitation activities, toxicity tests remain focused on shallow-water model species. Moreover, current tests overlook potential synergies that may arise from the interaction of chemicals with natural stressors, such as the high pressures prevailing in the deep sea. As pressure affects chemical reactions and the physiology of marine organisms, it will certainly affect the toxicity of pollutants arising from the exploitation of deep-sea resources. We emphasize the need for environmental risk assessments based on information generated from ecotoxicological trials that mimic, as close as possible, the deep-sea environment, with emphasis to a key environmental factor - high hydrostatic pressure.
This review covers the literature published for marine natural products isolated from macroalgae and addresses the taxonomic details of source organisms, the chemical types of isolated compounds and the location of sampling sites. The emphasis of this review is on the identification of the most bioprospected taxa and regions, as well as on how these trends have shifted over time.
The ability to understand and predict the effects of ocean warming (under realistic scenarios) on marine biota is of paramount importance, especially at the most vulnerable early life stages. Here we investigated the impact of predicted environmental warming (+3 °C) on the development, metabolism, heat shock response and antioxidant defense mechanisms of the early stages of the common octopus, Octopus vulgaris. As expected, warming shortened embryonic developmental time by 13 days, from 38 days at 18 °C to 25 days at 21 °C. Concomitantly, survival decreased significantly (~29.9 %). Size at hatching varied inversely with temperature, and the percentage of smaller premature paralarvae increased drastically, from 0 % at 18 °C to 17.8 % at 21 °C. The metabolic costs of the transition from an encapsulated embryo to a free planktonic form increased significantly with warming, and HSP70 concentrations and glutathione S-transferase activity levels were significantly magnified from late embryonic to paralarval stages. Yet, despite the presence of effective antioxidant defense mechanisms, ocean warming led to an augmentation of malondialdehyde levels (an indicative of enhanced ROS action), a process considered to be one of the most frequent cellular injury mechanisms. Thus, the present study provides clues about how the magnitude and rate of ocean warming will challenge the buffering capacities of octopus embryos and hatchlings physiology. The prediction and understanding of the biochemical and physiological responses to warmer temperatures (under realistic scenarios) is crucial for the management of highly commercial and ecologically important species, such as O. vulgaris.
Some species of sacoglossan sea slugs can maintain functional chloroplasts from specific algal food sources in the cells of their digestive diverticula. These stolen chloroplasts (kleptoplasts) can survive in the absence of the plant cell and continue to photosynthesize, in some cases for as long as one year. Within the Metazoa, this phenomenon (kleptoplasty) seems to have only evolved among sacoglossan sea slugs. Known for over a century, the mechanisms of interaction between the foreign organelle and its host animal cell are just now starting to be unravelled. In the study of sacoglossan sea slugs as photosynthetic systems, it is important to understand their relationship with light. This work reviews the state of knowledge on autotrophy as a nutritional source for sacoglossans and the strategies they have developed to avoid excessive light, with emphasis to the behavioural and physiological mechanisms suggested to be involved in the photoprotection of kleptoplasts. A special focus is given to the advantages and drawbacks of using pulse amplitude modulated fluorometry in photobiological studies addressing sacoglossan sea slugs. Finally, the classification of photosynthetic sacoglossan sea slugs according to their ability to retain functional kleptoplasts and the importance of laboratory culturing of these organisms are briefly discussed.
Herbivory in corals, especially for symbiotic species, remains controversial. To investigate the capacity of scleractinian and soft corals to capture microalgae, we conducted controlled laboratory experiments offering five algal species: the cryptophyte Rhodomonas marina, the haptophytes Isochrysis galbana and Phaeocystis globosa, and the diatoms Conticribra weissflogii and Thalassiosira pseudonana. Coral species included the symbiotic soft corals Heteroxenia fuscescens and Sinularia flexibilis, the asymbiotic scleractinian coral Tubastrea coccinea, and the symbiotic scleractinian corals Stylophora pistillata, Pavona cactus and Oculina arbuscula. Herbivory was assessed by end-point PCR amplification of algae-specific 18S rRNA gene fragments purified from coral tissue genomic DNA extracts. The ability to capture microalgae varied with coral and algal species and could not be explained by prey size or taxonomy. Herbivory was not detected in S. flexibilis and S. pistillata. P. globosa was the only algal prey that was never captured by any coral. Although predation defence mechanisms have been shown for Phaeocystis spp. against many potential predators, this study is the first to suggest this for corals. This study provides new insights into herbivory in symbiotic corals and suggests that corals may be selective herbivorous feeders.
Marine natural products (NP) are unanimously acknowledged as the blue gold in the urgent quest for new pharmaceuticals. Although corals are among the marine organisms with the greatest diversity of secondary metabolites, growing evidence suggest that their symbiotic bacteria produce most of these bioactive metabolites. The ex hospite culture of coral symbiotic microbiota is extremely challenging and only limited examples of successful culture exist today. By contrast, in toto aquaculture of corals is a commonly applied technology to produce corals for aquaria. Here, we suggest that coral aquaculture could as well be a viable and economically feasible option to produce the biomass required to execute the first steps of the NP-based drug discovery pipeline.
An experimental life support system (ELSS) was constructed to study the interactive effects of multiple stressors on coastal and estuarine benthic communities, specifically perturbations driven by global climate change and anthropogenic environmental contamination. The ELSS allows researchers to control salinity, pH, temperature, ultraviolet radiation (UVR), tidal rhythms and exposure to selected contaminants. Unlike most microcosms previously described, our system enables true independent replication (including randomization). In addition to this, it can be assembled using commercially available materials and equipment, thereby facilitating the replication of identical experimental setups in different geographical locations. Here, we validate the reproducibility and environmental quality of the system by comparing chemical and biological parameters recorded in our ELSS with those prevalent in the natural environment. Water, sediment microbial community and ragworm (the polychaete Hediste diversicolor) samples were obtained from four microcosms after 57 days of operation. In general, average concentrations of dissolved inorganic nutrients (NO3 (-) ; NH4 (+) and PO4 (-3) ) in the water column of the ELSS experimental control units were within the range of concentrations recorded in the natural environment. While some shifts in bacterial community composition were observed between in situ and ELSS sediment samples, the relative abundance of most metabolically active bacterial taxa appeared to be stable. In addition, ELSS operation did not significantly affect survival, oxidative stress and neurological biomarkers of the model organism Hediste diversicolor. The validation data indicate that this system can be used to assess independent or interactive effects of climate change and environmental contamination on benthic communities. Researchers will be able to simulate the effects of these stressors on processes driven by microbial communities, sediment and seawater chemistry and to evaluate potential consequences to sediment toxicity using model organisms such as Hediste diversicolor.
The "PortoNovo" project was developed to standardize the methodologies for water quality management in the port areas of coastal Atlantic regions to improve the Water Frame Directive (WFD) for these specific water bodies. Under this scope, water and sediment samples were collected from five sites within the Port of Aveiro, Portugal. According to the physical and chemical parameters that were analyzed (i.e., metals, total organic carbon, polychlorinated biphenyls and polycyclic aromatic hydrocarbons), the sediments were not considered at risk based on European sediment quality laws. However, the bioassays that were performed on the sediment samples (Microtox®) and the standardized acute toxicity test using the marine rotifer, Brachionus plicatilis, on sediment elutriates revealed higher toxicity levels. The use of bioassays to assess sediment quality clearly complements more conservative approaches and highlights current gaps within the WFD. The approach presented here can be easily transferred to other port areas for more reliable water quality management.
In the present study, we assessed the bacterial richness and composition of sediment samples collected in and around the port of Aveiro, on the Atlantic coast of mainland Portugal. Sediment samples were collected in five locations: two within the port harbor, two in port areas along a channel adjacent to the harbor and one in a relatively undisturbed reference location. These areas were characterized as under high, medium and no port activity, respectively. In-depth, barcoded-pyrosequencing analysis indicated that port activity affects the composition and abundance of bacterial communities colonizing surface sediments. Sampling sites under the influence of port activities (channel and harbor) were associated with higher relative abundances of Desulfobacterales and a marked decline in the abundance of Flavobacteriia. In addition, there was a pronounced prevalence of operational taxonomic units (OTUs) in the port area that were closely related to hydrocarbon-degrading bacteria (Desulfococcus spp.), antifouling paint (bacterium strain WH6-7) and copper rich sediments (bacterium strain CanalPD16A). Here we provide evidence that specific phylotypes detected have the potential to be used as biomarkers and should be evaluated in future studies as proxies for sediment disturbance associated with port activity.
Global climate change has the potential to seriously and adversely affect marine ecosystem functioning. Numerous experimental and modeling studies have demonstrated how predicted ocean acidification and increased ultraviolet radiation (UVR) can affect marine microbes. However, researchers have largely ignored interactions between ocean acidification, increased UVR and anthropogenic pollutants in marine environments. Such interactions can alter chemical speciation and the bioavailability of several organic and inorganic pollutants with potentially deleterious effects, such as modifying microbial-mediated detoxification processes. Microbes mediate major biogeochemical cycles, providing fundamental ecosystems services such as environmental detoxification and recovery. It is, therefore, important that we understand how predicted changes to oceanic pH, UVR, and temperature will affect microbial pollutant detoxification processes in marine ecosystems. The intrinsic characteristics of microbes, such as their short generation time, small size, and functional role in biogeochemical cycles combined with recent advances in molecular techniques (e.g., metagenomics and metatranscriptomics) make microbes excellent models to evaluate the consequences of various climate change scenarios on detoxification processes in marine ecosystems. In this review, we highlight the importance of microbial microcosm experiments, coupled with high-resolution molecular biology techniques, to provide a critical experimental framework to start understanding how climate change, anthropogenic pollution, and microbiological interactions may affect marine ecosystems in the future.
The present study combined a DGGE and barcoded 16S rRNA pyrosequencing approach to assess bacterial composition in the water of a recirculating aquaculture system (RAS) with a shallow raceway system (SRS) for turbot (Scophthalmus maximus) and sole (Solea senegalensis). Barcoded pyrosequencing results were also used to determine the potential pathogen load in the RAS studied. Samples were collected from the water supply pipeline (Sup), fish production tanks (Pro), sedimentation filter (Sed), biofilter tank (Bio), and protein skimmer (Ozo; also used as an ozone reaction chamber) of twin RAS operating in parallel (one for each fish species). Our results revealed pronounced differences in bacterial community composition between turbot and sole RAS, suggesting that in the systems studied there is a strong species-specific effect on water bacterial communities. Proteobacteria was the most abundant phylum in the water supply and all RAS compartments. Other important taxonomic groups included the phylum Bacteriodetes. The saltwater supplied displayed a markedly lower richness and appeared to have very little influence on bacterial composition. The following potentially pathogenic species were detected: Photobacterium damselae in turbot (all compartments), Tenacibaculum discolor in turbot and sole (all compartments), Tenacibaculum soleae in turbot (all compartments) and sole (Pro, Sed and Bio), and Serratia marcescens in turbot (Sup, Sed, Bio and Ozo) and sole (only Sed) RAS. Despite the presence of these pathogens, no symptomatic fish were observed. Although we were able to identify potential pathogens, this approach should be employed with caution when monitoring aquaculture systems, as the required phylogenetic resolution for reliable identification of pathogens may not always be possible to achieve when employing 16S rRNA gene fragments.
The combined effects of future ocean acidification and global warming on the hypoxia thresholds of marine biota are, to date, poorly known. Here, we show that the future warming and acidification scenario led to shorter embryonic periods, lower survival rates and the enhancement of premature hatching in the cuttlefish Sepia officinalis. Routine metabolic rates increased during the embryonic period, but environmental hypercapnia significantly depressed pre-hatchlings energy expenditures rates (independently of temperature). During embryogenesis, there was also a significant rise in the carbon dioxide partial pressure in the perivitelline fluid (PVF), bicarbonate levels, as well as a drop in pH and oxygen partial pressure (pO?). The critical partial pressure (i.e. hypoxic threshold) of the pre-hatchlings was significantly higher than the PVF oxygen partial pressure at the warmer and hypercapnic condition. Thus, the record of oxygen tensions below critical pO? in such climate scenario indicates that the already harsh conditions inside the egg capsules are expected to be magnified in the years to come, especially in populations at the border of their thermal envelope. Such a scenario promotes untimely hatching and smaller post-hatching body sizes, thus challenging the survival and fitness of early life stages.
Marine invertebrates are rich sources of bioactive compounds and their biotechnological potential attracts scientific and economic interest worldwide. Although sponges are the foremost providers of marine bioactive compounds, cnidarians are also being studied with promising results. This diverse group of marine invertebrates includes over 11,000 species, 7500 of them belonging to the class Anthozoa. We present an overview of some of the most promising marine bioactive compounds from a therapeutic point of view isolated from cnidarians in the first decade of the 21st century. Anthozoan orders Alcyonacea and Gorgonacea exhibit by far the highest number of species yielding promising compounds. Antitumor activity has been the major area of interest in the screening of cnidarian compounds, the most promising ones being terpenoids (monoterpenoids, diterpenoids, sesquiterpenoids). We also discuss the future of bioprospecting for new marine bioactive compounds produced by cnidarians.
Aquaculture activities are increasing worldwide, stimulated by the progressive reduction of natural fish stocks in the oceans. However, these activities also suffer heavy production and financial losses resulting from fish infections caused by microbial pathogens, including multidrug resistant bacteria. Therefore, strategies to control fish infections are urgently needed, in order to make aquaculture industry more sustainable. Antimicrobial photodynamic therapy (aPDT) has emerged as an alternative to treat diseases and prevent the development of antibiotic resistance by pathogenic bacteria. The aim of this work was to evaluate the applicability of aPDT to inactivate pathogenic fish bacteria. To reach this objective a cationic porphyrin Tri-Py(+)-Me-PF was tested against nine pathogenic bacteria isolated from a semi-intensive aquaculture system and against the cultivable bacteria of the aquaculture system. The ecological impact of aPDT in the aquatic environment was also tested on the natural bacterial community, using the overall bacterial community structure and the cultivable bacteria as indicators. Photodynamic inactivation of bacterial isolates and of cultivable bacteria was assessed counting the number of colonies. The impact of aPDT in the overall bacterial community structure of the aquaculture water was evaluated by denaturing gel gradient electrophoresis (DGGE). The results showed that, in the presence of Tri-Py(+)-Me-PF, the growth of bacterial isolates was inhibited, resulting in a decrease of ?7-8 log after 60-270 min of irradiation. Cultivable bacteria were also considerably affected, showing decreases up to the detection limit (?2 log decrease on cell survival), but the inactivation rate varied significantly with the sampling period. The DGGE fingerprint analyses revealed changes in the bacterial community structure caused by the combination of aPDT and light. The results indicate that aPDT can be regarded as a new approach to control fish infections in aquaculture systems, but it is clearly more difficult to inactivate the complex natural bacterial communities of aquaculture waters than pure cultures of bacteria isolated from aquaculture systems. Considering the use of aPDT to inactivate pathogenic microbial community of aquaculture systems the monitoring of microorganisms is needed in order to select the most effective conditions.
A new methodology for the assessment of thiocyanate (SCN(-)) is proposed based on optical fiber (OF) detection coupled to a liquid chromatography system (LC). The developed methodology showed an adequate performance for the analysis of SCN(-) comparable to a high performance liquid chromatography with UV detector (HPLC-UV) methodology: a detection limit of 3 µg L(-1), a linear range from 4 to 400 µg L(-1), and an analytical time of less than 6 min. The OF based methodology was of compact design and easy operation. This simple system has the potential to be used as a sensing approach for SCN(-) in seawater.
Mangroves are complex and dynamic ecosystems varying in salinity, water level and nutrient availability; they also contain diverse and distinct microbial communities. Studies of microbes and their interactions with other ecosystem components (e.g., tree roots) are critical for our understanding of mangrove ecosystem functioning and remediation. Using a barcoding pyrosequencing approach, we previously noted the persistence of terrestrial bacterial populations on mangrove roots when nursery raised saplings were transplanted back to their natural environment. Here we go into further detail about the potential functional associations of bacterial guilds with distinct mangrove microhabitats including the rhizosphere. We also use a nonparametric richness estimator to show that estimated operational taxonomic unit (OTU) richness is more than twice that observed. In the transplant microhabitat, our estimate suggests that there are almost 7,000 OTUs for a sample size of 10,400 individual sequences with no sign of an asymptote, indicating that "true" richness for this microhabitat is substantially larger. Results on the number of bacterial OTUs should, however, be viewed with caution given that the barcoding pyrosequencing technique used can yield sequencing artifacts that may inflate richness estimates if not properly removed.
Kleptoplasty is a particularly remarkable type of symbiosis, consisting of the presence of functional chloroplasts in the tissues of a host of another species. One of the most well-studied types of kleptoplasty is the association between sacoglossan molluscs (sea slugs) and algal chloroplasts. After ingestion, the chloroplasts remain photosynthetically functional and provide photosynthates to the host, therefore named as "solar-powered" sea slugs. This study evaluated the use of two optical methods, spectral reflectance analysis and in vivo Chl fluorescence, as measured by pulse amplitude modulated (PAM) fluorometry, for the in vivo quantification of kleptoplastic chlorophyll (Chl) a content in the sacoglossan Elysia viridis (Montagu, 1804) bearing chloroplasts of the macroalgae Codium tomentosum var. mucronatum (G. Hamel) Ardré. The Chl a content of E. viridis specimens was compared to a number of reflectance-based indices and to the dark-level fluorescence, F(o). Most reflectance-based indices varied linearly with the symbiosis Chl a content over the whole range of pigment content variation. Most significant correlations (P < 0.001) were found between indices using as reference the reflectance at 750 nm, with the proportion of pigment content explained by the indices varying between 63.5% and 85.9%. F(o) varied linearly with the Chl a content only for low pigment levels (below 4-6 microg Chl a per individual), above which it followed a saturation-like pattern. The use of optical methods was illustrated by monitoring the changes in Chl a content of specimens during periods of starvation and subsequent recovery. The results of this study suggest that, if basic requirements of signal detection and reproducible measuring geometry are verified, these optical methods may be readily applied to other photosynthetic symbioses.
The wide geographical distribution of the Norway lobster, Nephrops norvegicus, results in a delay, with latitudinal decrease, in the larval season from spring to winter. Newly hatched larvae of the species may therefore be exposed to suboptimal levels or types of prey and face intermittent periods of starvation at low latitudes. This work investigated the feeding response of the first two zoeal stages of N. norvegicus under variable prey densities, prey types, feeding histories, and photoperiods. Both zoeae (Z) I and II increased the number of consumed prey with increasing food levels. ZI preferred Artemia sp. nauplii over larger metanauplii, while in ZII, higher ingestion was observed only for metanauplii at higher food concentrations. The number of prey ingested by larvae previously starved or under low food conditions was always higher than that of larvae exposed to high food levels. These findings seem to indicate that larvae may maximize prey ingestion in the presence of plankton patches with higher food abundance and minimize the deleterious effects induced by previous periods of intermittent starvation or unsuitable prey densities or types. Extreme photoperiods (24 and 0 h of light) did not improve larval feeding ability and are not a suitable option for larviculture.
Photosynthetic sea slugs have the ability to "steal" chloroplasts (kleptoplasts) from marine macroalgae and keep them structurally intact and physiologically functional. The photosynthetic activity of these symbioses has been assessed using pulse amplitude modulated (PAM) fluorometry. However, the movement of these sacoglossan slugs can impair specific photobiological studies on kleptoplasts. Thus, immobilizing sacoglossan slugs while not interfering with the photosynthetic activity would be a methodological advance for research in this field. We evaluated the effect of two anesthetics, eugenol and MS-222, on the photosynthetic activity of kleptoplasts and on the behavior of the kleptoplasts-bearing slug Elysia viridis. Anesthetics promoted relaxation of sea slug muscle with no touch reaction in about 6 min. Sea slugs immobilized for 120 min completely recovered after anesthetic removal. No significant differences were found on photosynthetic parameters measured immediately (0-1 min) after immobilization. The effective quantum yield of photosystem II of E. viridis after 120 min of immobilization was significantly decreased by 12% in the MS-222 treatment, while eugenol promoted no significant effect. Photosynthetic activity assessed by rapid light-response curves (RLC) of relative electron transport rates (rETR) revealed a significant decrease in both initial response to light (-34%) and maximum rETR (rETR(m)) (-60%), after 120 min of immobilization using MS-222. After 120 min of immobilization with eugenol, the initial response to light significantly decreased 15% and rETR(m) decreased 27%. We conclude that, whenever photobiological studies employing PAM fluorometry require immobilization of photosynthetic sea slugs, eugenol can be used as a powerful anesthetic with little impact on the photosynthetic activity of kleptoplasts.
Sea-Nine 211™ is a new biocide specifically formulated for antifouling paints and being considered to have a low environmental impact. Even with a short environmental half-life, this compound can cause toxic effects on marine organisms. This study used PAM fluorometry and biomarkers of oxidative stress (GST, CAT and LPO) to monitor potential toxic effects of Sea-Nine 211™ on fragments of the soft coral Sarcophyton cf. glaucum. After exposure to concentrations of 1-100 ?g l(-1) for 72 h, CAT activity was inhibited under the two highest concentrations, being in accordance with the activity of GST. LPO activity (as TBARS) and photosynthetic efficiency of endosymbiotic zooxanthellae were not significantly affected. These results show that PAM fluorometry alone cannot detect the full effects of Sea-Nine 211™ on Sarcophyton cf. glaucum and should be used together with other biomarkers. This holobiont driven approach to evaluate chemical toxicity in photosynthetic corals is therefore recommended for biocides which are not photosystem II inhibitors.
The present study aimed to assess if ragworm fatty acids (FA) profiles could be used to discriminate their spatial distribution in an historically mercury-contaminated estuarine environment, i.e., if it was possible to differentiate ragworms present in salt marsh sediments surrounding plant roots and rhizomes (rhizosediment) from adjacent unvegetated sediment. Additionally, we also tried to determine if ragworms differed in mercury content and if these values could also be used to identify the habitat they occur in. Results show that, within the same area, ragworms can be distinguished using FA profiles and that in halophyte rhizosediment ragworms display more than twice the levels of alpha-linolenic acid (18:3n-3). The ratio cis-vaccenic/oleic acids (18:1n-7/18:ln-9) in ragworms suggests higher carnivory in unvegetated sediments. Our study indicates that ragworm FA profiles can be used to identify their habitat, their trophic interaction with halophytes and reveal a spatially contrasting feeding behaviour, which also reflects mercury accumulation.
Bioprospecting for new marine natural products (NPs) has increased significantly over the last decades, leading to an unprecedented discovery of new molecules. Marine invertebrates have been the most important source of these NPs, with researchers commonly targeting particular taxonomic groups, marine regions and/or molecules from specific chemical groups. The present review focuses on new NPs identified from marine invertebrates between 2000 and 2009, and performs a detailed analysis on: (1) the chemical groups of these NPs; (2) the association of particular chemical groups to specific marine invertebrate taxa; and (3) the yielding of molecules from the same chemical group from organisms occurring in a particular geographic region. Our survey revealed an increasing number of new terpenoids being discovered between 2000 and 2009, contrasting with the decreasing trend in the discovery of new alkaloids and aliphatic molecules. Overall, no particular association was identified between marine invertebrate taxa and chemical groups of new NPs. Nonetheless, it is worth noting that most NPs recorded from cnidarians and mollusks were terpenoids, while most NPs identified in echinoderms were aliphatic compounds or carbohydrates. The geographical trends observed in our study do not support the idea of particular chemical groups of new NPs being associated with marine invertebrates from any specific geographical region, as NPs from different chemical groups were commonly distributed worldwide.
Cyanide fishing is a method employed to capture marine fish alive on coral reefs. They are shipped to markets for human consumption in Southeast Asia, as well as to supply the marine aquarium trade worldwide. Although several techniques can be used to detect cyanide in reef fish, there is still no testing method that can be used to survey the whole supply chain. Most methods for cyanide detection are time-consuming and require the sacrifice of the sampled fish. Thiocyanate anion (SCN(-)) is a metabolite produced by the main metabolic pathway for cyanide anion (CN(-)) detoxification. Our study employed an optical fiber (OF) methodology (analytical time <6 min) to detect SCN(-) in a non-invasive and non-destructive manner. Our OF methodology is able to detect trace levels (>3.16 µg L(-1)) of SCN(-) in seawater. Given that marine fish exposed to cyanide excrete SCN(-) in the urine, elevated levels of SCN(-) present in the seawater holding live reef fish indicate that the surveyed specimens were likely exposed to cyanide. In our study, captive-bred clownfish (Amphiprion clarkii) pulse exposed for 60 s to either 12.5 or 25 mg L(-1) of CN(-) excreted up to 6.96±0.03 and 9.84±0.03 µg L(-1) of SCN(-), respectively, during the 28 days following exposure. No detectable levels of SCN(-) were recorded in the water holding control organisms not exposed to CN(-), or in synthetic seawater lacking fish. While further research is necessary, our methodology can allow a rapid detection of SCN(-) in the holding water and can be used as a screening tool to indicate if live reef fish were collected with cyanide.
It is acknowledged that marine invertebrates produce bioactive natural products that may be useful for developing new drugs. By exploring untapped geographical sources and/or novel groups of organisms one can maximize the search for new marine drugs to treat human diseases. The goal of this paper is to analyse the trends associated with the discovery of new marine natural products from invertebrates (NMNPI) over the last two decades. The analysis considers different taxonomical levels and geographical approaches of bioprospected species. Additionally, this research is also directed to provide new insights into less bioprospected taxa and world regions. In order to gather the information available on NMNPI, the yearly-published reviews of Marine Natural Products covering 1990-2009 were surveyed. Information on source organisms, specifically taxonomical information and collection sites, was assembled together with additional geographical information collected from the articles originally describing the new natural product. Almost 10000 NMNPI were discovered since 1990, with a pronounced increase between decades. Porifera and Cnidaria were the two dominant sources of NMNPI worldwide. The exception was polar regions where Echinodermata dominated. The majority of species that yielded the new natural products belong to only one class of each Porifera and Cnidaria phyla (Demospongiae and Anthozoa, respectively). Increased bioprospecting efforts were observed in the Pacific Ocean, particularly in Asian countries that are associated with the Japan Biodiversity Hotspot and the Kuroshio Current. Although results show comparably less NMNPI from polar regions, the number of new natural products per species is similar to that recorded for other regions. The present study provides information to future bioprospecting efforts addressing previously unexplored taxonomic groups and/or regions. We also highlight how marine invertebrates, which in some cases have no commercial value, may become highly valuable in the ongoing search for new drugs from the sea.
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