Do fish gut microbiotas vary across spatial scales? A case study of Diplodus vulgaris in the Mediterranean Sea.

BACKGROUND: Biogeography has been linked to differences in gut microbiota in several animals. However, the existence of such a relationship in fish is not clear yet. So far, it seems to depend on the fish species studied. However, most studies of fish gut microbiotas are based on single populations. In this study, we investigated the gut microbiota of fish from three wild populations of the two-banded sea bream Diplodus vulgaris (Geoffroy Saint-Hilaire, 1817) to determine whether its diversity, structure and potential functionality reflect the geographic origin of the fish, at large and small geographical scale. Additionally, we explored the host- and environmental-related factors explaining this relationship. RESULTS: We showed that the taxonomy and potential functionality of the mucosa-associated gut microbiota of Diplodus vulgaris differ to varying degrees depending on the spatial scale considered. At large scale, we observed that both the taxonomical structure and the potential functionality of the fish microbiota differed significantly between populations. In contrast, the taxonomical diversity of the microbial community displayed a significant relationship with factors other than the geographic origin of the fish (i.e. sampling date). On the other hand, at small scale, the different composition and diversity of the microbiota differ according to the characteristics of the habitat occupied by the fish. Specifically, we identified the presence of Posidonia oceanica in the benthic habitat as predictor of both the microbiota composition and diversity. Lastly, we reported the enrichment of functions related to the metabolism of xenobiotics (i.e. drugs and 4-aminobenzoate) in a population and we indicated it as a potential target of future monitoring. CONCLUSIONS: With this study, we confirmed the importance of investigating the gut microbiota of wild fish species using multiple populations, taking into account the different habitats occupied by the individuals. Furthermore, we underscored the use of the biodegradation potential of the gut microbiota as an alternative means of monitoring emerging contaminants in Mediterranean fish.

Development and validation of a DNA-based multi-species biomonitoring toolkit using a high-throughput qPCR platform: A case study of Irish shellfish species.

Biomonitoring of marine life has been enhanced in recent years by the integration of innovative DNA-based approaches, which offer advantages over more laborious techniques (e.g. microscopy). However, trade-offs between throughput, sensitivity and quantitative measurements must be made when choosing between the prevailing molecular methodologies (i.e. metabarcoding or qPCR/dPCR). Thus, the aim of the present study was to demonstrate the utility of a microfluidic-enabled high-throughput quantitative PCR platform (HTqPCR) for the rapid and cost-effective development and validation of a DNA-based multi-species biomonitoring toolkit, using larvae of 23 commercially targeted bivalve and crustacean species as a case study. The workflow was divided into three main phases: definition of (off-) target taxa and establishment of reference databases (PHASE 1); selection/development and assessment of molecular assays (PHASE 2); and protocol optimization and field validation (PHASE 3). 42 assays were eventually chosen and validated. Genetic signal not only showed good correlation with direct visual counts by microscopy but also showed the ability to provide quantitative data at the highest taxonomic resolution (species level) in a time- and cost-effective fashion. This study developed a biomonitoring toolkit, demonstrating the considerable advantages of this state-of-the-art technology in boosting the developmental testing and application of panels of molecular assays for the monitoring and management of natural resources. Once developed, this approach provides a cost and time-effective alternative compared to other multi-species approaches (e.g. metabarcoding). In addition, it is transferable to a wide range of species and will aid future monitoring programmes.

Importance of suspended particulate organic matter in the diet of Nephrops norvegicus (Linnaeus, 1758).

The extent to which commercially important Nephrops norvegicus lobsters feed on particulates in the wild is unknown, even though this could be an important way for burrow-dwelling females to avoid starvation during the long breeding season. This was investigated using δ13C and δ15N isotopic signatures in tissues with long and short turnover rates to provide diet discrimination and compare this between males and females. Secondary objectives examined size-related differences and calculated the trophic position based on the new results. Almost half the diet (47%) was made up of suspended particulate organic matter (POMsusp) alone. Fish was another important item in the diet, with plankton and invertebrate sources coming much lower down in dietary importance. Significantly more suspension feeding was observed in small or medium sized individuals than large ones in both sexes. However, there were no sex-related patterns, despite females being restricted to burrows for part of the analysis period. Female diet was almost identical to males and POMsusp comprised a large component of the diet in both sexes. The trophic position was estimated at 2.94 ± 0.16 (mean ± SD), which was at the lower end of the range reported in previous studies (2.60 to 4.32).

Acid treatment of Atlantic salmon (Salmo salar) scales prior to analysis has negligible effects on δ13C and δ15N isotope ratios.

There is debate in the literature as to whether scales of fishes require acidification to remove inorganic carbonates prior to stable isotope analysis. Acid-treated and untreated scales from 208 Atlantic salmon from nine locations on both sides of the Atlantic were analysed for δ13C and δ15N. Linear mixed-effect models determined the effect of acid treatment to be statistically significant. However, the mean difference was small (δ13C 0.1 ± 0.2‰, δ15N -0.1 ± 0.2‰) and not of biological relevance. This study concludes that Atlantic salmon scales do not need to be acidified prior to stable isotope analysis.

Trace element fingerprinting of blue mussel (Mytilus edulis) shells and soft tissues successfully reveals harvesting locations.

Globally, aquaculture products are expected to account for >60% of total seafood produce by 2030. In the European Union, the seafood sector is of considerable economic importance to member states with household spending on seafood produce totalling €54.8 billion in 2016. Within the EU, shellfish aquaculture supports livelihoods and employment in many rural communities throughout the region. Harmful algal blooms pose considerable risk to consumer safety and in turn, stability of the shellfish market. If contaminated produce was to make it to the market the health risk to the public could be considerable, but the damage to the sector through loss of trust in producers would also be significant. Mytilus edulis account for a considerable portion of the aquaculture sector in the Northeast Atlantic. At present, no scientific tool is available to industry regulators, to allow them to trace mussel produce to its source, uphold food safety standards and ensure consumer confidence. The present study uses chemical analysis of shells and soft tissues to classify individual M. edulis to their site of harvest. The use of random forest classification of trace element composition has revealed location specific elemental signatures for all examined sites. This led to the correct classification of 100% of individuals sampled to their respective harvesting locations, including two sites located just 6 km apart within the same bay. The protocol demonstrated here provides the basis for a scientifically driven traceability framework for shellfish produce.

Spatio-temporal trace element fingerprinting of king scallops (Pecten maximus) reveals harvesting period and location.

A rapidly growing human population is increasingly relying on seafood as a source of protein and other essential nutrients. Bivalve shellfish, both from wild populations and aquaculture, will undoubtedly continue to account for a significant portion of overall seafood production, but consumption of such shellfish carries potential health risks. Biotoxins, disease causing organisms and pollution contribute to this risk, as shellfish are indiscriminate, passive filter feeders. While government bodies, industry regulators and producers are capable of managing this risk, counterfeit produce can risk public safety, in turn damaging the reputation of the entire industry. Traceability tools provide a means to uphold food safety standards and mitigate remaining risk to consumers. Here, we show how the use of trace element (TE) signatures in shells and soft tissues of king scallops combined, can predict geographic origin with 100% accuracy. Importantly, we explore the temporal stability of this method, successfully classifying 100% of individuals correctly between two dates just 42 days apart from the same harvesting location. The most important elements in the trace element signatures of the scallops, discriminating between harvesting sites and dates were barium, boron, chromium, lead, manganese, molybdenum and selenium. The traceability tool described here offers a viable method to trace produce to its source, empowering industry regulators, government authorities, aquaculture practitioners and retailers in terms of tracking shellfish throughout the supply chain, which would comply with legislation and boost consumer confidence.

Stable-isotope analysis reveals the importance of soft-bodied prey in the diet of lesser spotted dogfish Scyliorhinus canicula.

This study examines the diet of lesser spotted dogfish Scyliorhinus canicula, one of the most ubiquitous predators of European coastal waters. This species is of increasing ecological significance as other large predatory fish decline as it has known interactions with fisheries. Scyliorhinus canicula diet was investigated in Irish coastal waters during June and July 2014 using both stomach-content analysis and δ13 C and δ15 N stable-isotope ratios. Prey contribution to the diet from dual stable-isotope data was estimated using Bayesian mixing models. It was found that only stable-isotope analysis provided a time-integrated picture of the diet of S. canicula and allowed for a new estimation of their trophic position. Trophic positions from stomach-content analysis within the present study and previous studies were found to be higher than revealed by isotopes. Stomach-content analysis could not reveal the importance of soft-bodied animals in the diet of these fish, however this approach remains a valuable tool to understand the potential prey spectrum in advance of isotope analysis and allows for a better resolution down to species level. The results highlight a greater variety in the diet of this important predator and the benefit of taking a multidisciplinary approach in dietary studies.

Growth in Nephrops norvegicus from a tag-recapture experiment.

Nephrops norvegicus is a commercially valuable fishery in the EU but management of stocks is challenging due to difficulties in aging individuals and calculating growth and biomass production. Growth of N. norvegicus was estimated by releasing 1177 tagged individuals in western Ireland in Summer 2013 and recapturing these in 2014 (n = 207, an average of 344 days later) and 2015 (n = 38, 654-665 days later). Moulting occurred twice per year in approximately half of the males and only once in females. Mean growth increments after approximately one year were 5.1 mm Carapace Length (CL) in males and 1.4 mm CL in females. After two years, males had grown by 12.0 mm CL and females by 4.6 mm CL, on average, across size classes. Low variation in growth increments was seen across female size classes, but significantly lower growth was observed in larger males, meeting an important assumption of the Von Bertalanffy Growth Function. Asymptotic carapace lengths were 70.8 mm (males) and 55.2 mm (females) with respective growth constants (k) of 0.161 yr-1 and 0.077 yr-1. The results suggest that this is a very productive fishery and that survivability of returns from creel fishing is high.

Development of non-lethal sampling of carbon and nitrogen stable isotope ratios in salmonids: effects of lipid and inorganic components of fins.

The preferred tissue for analyses of fish stable isotope ratios for most researchers is muscle, the sampling of which typically requires the specimen to be sacrificed. The use of non-destructive methods in fish isotopic research has been increasing recently, but as yet is not a standard procedure. Previous studies have reported varying levels of success regarding the utility of non-lethally obtained stable isotope materials, e.g. fins, but none have accounted for the potential compounding effects of inorganic components of fin rays or lipids. Comparisons of carbon (δ(13)C) and nitrogen (δ(15)N) stable isotope ratios of muscle with adipose and caudal fin of two salmonids, Atlantic salmon (Salmo salar L.) and brown trout (Salmo trutta L.), revealed that caudal fin can be used as a non-destructive surrogate for muscle in stable isotope analysis, but that adipose fin, where available, is a better proxy. The use of a published model to inexpensively counteract the confounding effect of lipids, which are depleted in (13)C, greatly improved the relationship between fish muscle and fins. However, efforts to account for the inorganic components of fin rays were counterproductive and required twice the biomass of fins clipped from each fish. As this experiment was conducted on wild fish, controlled laboratory studies are required to confirm these field observations.