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The microbiota of recirculating aquaculture systems (RAS) is of major importance for optimal fish health. However, the microbial communities in commercial RAS are highly complex and more knowledge is needed to potentially control and maintain beneficial microbial communities for good fish production. In this study we monitored microbial communities in a commercial RAS producing Atlantic salmon fry (Salmo salar) during seven consecutive production batches. The water of rearing tanks and the water sump downstream of the biofilter/upstream of the UV, as well as biofilm of the wall of the rearing tanks and the fixed bed biofilter were analysed using 16S rRNA gene amplicon sequencing to elucidate the spatial-temporal microbial dynamics. The results showed that the microbiota composition of water and biofilm varied within and between the production batches, and that the fallowing periods had a substantial effect on the microbial communities. The correlation of the water and biofilm microbiota to fish presence in the system was confirmed by supervised machine learning. Shifts in the composition of the microbiota were identified in conjunction with variations in organic matter loading both during production and fallowing. In addition, variables like oxygen saturation, biomass, and feed type, showed good correlation with variations in the water microbiota composition. Although microbiota changed at fallowing, the microbiota returned to similar compositions during the production phases and was especially evident for the water microbiota. This indicates that the development of microbiota composition is strongly dictated by the similar selection pressure in the system. Nitrifying communities were dominated by Nitrospira, and the third most abundant Nitrospira OTUs were related to the comammox Nitrospira nitrificans. The microbial communities in the biofilter biofilm and water were significantly different but shared abundant taxa and followed the same temporal microbial dynamics and indicates an interaction between the biofilter biofilm and the suspended bacteria. CFU analysis showed that the fraction of rapid-growing bacteria was significantly higher in the rearing water than in the water sump upstream the UV disinfection, indicating that disinfection upstream the rearing tanks allowed for growth of opportunistic bacteria. A community with considerable potential for opportunistic regrowth can have consequences for the microbial water quality and the resistance against pathogen invasion The absence of an in-line disinfection step or placing the disinfection unit upstream the biofilter might provide better microbial water quality and a more resilient system against pathogen proliferation.

Recirculating aquaculture systems (RAS) harbour complex microbial communities which can have an impact on the growth and development of the reared fish. This study aimed to improve our understanding of microbial community dynamics in a RAS involving three production batches of Atlantic salmon fry and parr during a period of 20 months. Water for analysis of microbiota was sampled at different positions in the RAS, and we also examined the effect of UV treatment on the water microbiota. Microbial communities were characterized by 16S rDNA amplicon sequencing of water samples taken directly upstream and downstream of the UV treatment unit and from three of the rearing tanks. In total 6 sampling events were made during a 20-month period. The study showed that: 1) Two of the production batches had a highly similar water microbiota despite disinfection of the system between the batches and rearing fish of different stages. In contrast, the first production batch showed a different water microbiota with variable composition through the system and over time. A more immature biofilter in the first batch may explain these differences. 2) The full-flow UV treatment directly upstream the rearing tanks had no observable effect on the community composition of the water microbiota in the different sampling positions in the RAS. This was likely a consequence of the low hydraulic retention time (HRT) (23 min) in rearing tanks, low bacterial regrowth in the fish tanks and community changes throughout the RAS loop. 3) The disinfection effect on viable bacterial densities in the water directly downstream of the UV treatment was around 89%, when the water was clear. Regrowth of bacteria following disinfection was low compared to those reported for marine RAS with UV disinfection and long HRT in fish tanks. The study shows that UV disinfection can be used to efficiently reduce bacterial density without compromising the microbial water quality in the fish tanks in RAS with low HRT.

Decline in fish oil and fish meal availability has forced the aquaculture sector to investigate alternative and sustainable aquafeed ingredients. Despite that several studies have evaluated the effect of fish oil replacement in aquaculture fish species, there is a knowledge gap on the effects of alternative dietary lipid sources on the gut microbiota in early life stages of Salmo salar. The present study evaluated the influence of dietary administration of two different lipid sources (fish oil and vegetable oil) on the intestinal microbiota of first feeding Atlantic salmon (S. salar) up to 93 days post first feeding (dpff). The two diets used in this study, FD (fish oil diet) and VD (blend of rapeseed, linseed and palm oils diet), were formulated to cover the fish nutritional requirements. Apart from the lipid source, the rest of the feed components were identical in the two diets. Hindgut samples were collected at 0, 35, 65, and 93 dpff. Moreover, fertilized eggs, yolk sac larvae, rearing water and feed were also collected in order to assess a possible contribution of their microbiota to the colonization and bacterial succession of the fish intestines. To analyze the bacterial communities, amplicon sequencing was used targeting the V3–V4 region of the 16S rRNA gene. The findings indicate that feeding on either fish oil or vegetable oil-based diet, fish growth variables (mean wet weight and total length) did not differ significantly during the experiment (p > 0.05). No significant differences were also found between the two dietary groups, regarding their gut bacteria composition, after the analysis of the 16S rRNA sequencing data. Instead, gut microbiota changed with age, and each stage was characterized by different dominant bacteria. These operational taxonomic units (OTUs) were related to species that provide different functions and have been isolated from a variety of environments. The results also show little OTUs overlap between the host and rearing environment microbiota. Overall, this study revealed the occurrence of a core microbiota in early life of Atlantic salmon independent of the feed-contained oil origin. Copyright © 2021 Nikouli, Kormas, Jin, Olsen, Bakke and Vadstein. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

Oil spilled in the Arctic may drift into ice-covered areas and become trapped until the ice melts. To determine if exposure to oil during freezing may have a priming effect on degradation of the oil, weathered dispersed oil (2-3 mg/L) was frozen into solid ice for 200 days at -10 °C, then melted and incubated for 64 days at 4 °C. No degradation was measured in oil frozen into ice prior to melting. Both total amount of oil and target compounds were biotransformed by the microbial community from the melted ice. However, oil released from melted ice was degraded at a slower rate than oil incubated in fresh seawater at the same temperature (4 °C), and by a different microbial community. These data suggest negligible biodegradation of oil frozen in sea ice, while oil-degrading bacteria surviving in the ice may contribute to biodegradation when the ice melts.

An important step in water treatment in recirculating aquaculture systems (RAS) is the removal of particulate organic matter. While commonly applied treatment technologies remove most of the particles, fine particulate organic matter typically accumulate in RAS, and serve as substrate for heterotrophic bacteria in the system. Membrane ultrafiltration has been shown to reduce the concentrations of total organic carbon and to counteract blooms of opportunistic bacteria in periods with high organic loading. The potential influence of such opportunistic bacteria on the fish microbiomes and fish welfare is poorly understood. In this study, we investigated the effects of high organic loading on the water and hindgut microbiomes in RAS with Atlantic salmon parr. Two pilot-scale RAS, one with membrane ultrafiltration of a 10–15% side-stream (mRAS), and one without (cRAS), were operated in periods with low and high organic loadings, and the water and hindgut microbiomes were characterized by Illumina sequencing of 16S rDNA amplicons. At the end of the experiment, after a period of high organic loading, two OTUs representing Mycobacterium and Spartobacteria dominated the water microbiomes in cRAS, with relative abundances as high as 24 and 17%, respectively. In the mRAS water they were rare (relative abundances of 1.9 and 0.1%). The membrane ultrafiltration stabilized the water microbiota and efficiently prevented the growth of these bacterial populations in RAS. The parr hindgut microbiomes were generally highly distinct from the RAS water microbiomes and were dominated by the bacterial phyla Bacilli and Firmicutes. Furthermore, an OTU probably representing Carnobacterium inhibens, originally isolated from the gut of adult salmon, was highly abundant in the gut microbiomes, with relative abundances reaching over 30%. However, for the cRAS fish after a period with high organic loading, the relative abundance of this Carnobacterium OTU was reduced, and an increased similarity between the gut and the water microbiomes was observed. The Mycobacterium and Spartobacteria OTUs that were abundant in the cRAS water were also abundant in the cRAS hindgut samples. This indicates a direct influence of the water microbiomes on the gut microbiomes, and that high organic loading in the cRAS resulted in growth of opportunistic bacteria that were able to colonize the fish gut. The present study shows that membrane filtration is an applicable strategy to counteract growth of opportunistic bacteria in RAS and to promote positive fish – microbe interactions.

Lumpfish (Cyclopterus lumpus L.) in Norway is currently produced in traditional flow-through systems (FTS). Hatcheries frequently show signs of bacterial infections, unstable microbial communities in the rearing water and varying mortality. Recirculating aquaculture systems (RAS) is proposed to create stable and healthy microbial environments, with less probabilities for blooming of opportunistic microbes. Studies have also shown that RAS increases the survival of marine fish. The aim with this study was to investigate the effect of various RAS water treatment designs on water and biofilm microbiota, survival, growth and gill health of lumpfish. An experiment with lumpfish was conducted, from 2 months post hatch to the transfer into sea cages. Five different water treatment regimens were compared: 1. RAS with no additional water treatment, 2. RAS with a filtration unit for removal of small particles, 3. RAS with filtration and disinfection with UV-irradiation, 4. RAS with filtration and disinfection with UV-irradiation and ozone and 5. FTS as a reference. The microbiota of the rearing water and tank wall biofilm were sampled and characterized by Illumina sequencing of 16S rDNA amplicons. Lumpfish juveniles reared in the RAS treatments were exposed to a more stable and diverse rearing water microbiota, with a lower share of opportunistic bacteria, a probable reason for the higher survival and better gill health of the fish compared to siblings reared in the FTS. Lumpfish reared in RAS without disinfection were exposed to a more diverse and stable water microbiota, with a lower share of opportunistic and potential harmful bacteria, compared to the lumpfish reared in RAS with disinfection and FTS. This resulted in better gill health. Fish in RAS with filtration, but no disinfection, had a better gill health than the fish in the RAS without filtration, possibly due to the reduction of small particles. The lumpfish were exposed to different microbial communities of both water and biofilm, due to the different treatments of the incoming tank water. In conclusion, our results indicate that implementation of RAS in the production of lumpfish has a potential to increase both survival, growth and gill health of the fish and that RAS with filtration of small particles, but without disinfection, result in the best fish health and performance among the investigated treatments. © 2020 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

As ice extent in the Arctic is declining, oil and gas activities will increase, with higher risk of oil spills to the marine environment. To determine biotransformation of dispersed weathered oil in newly formed ice, oil dispersions (2–3 ppm) were incubated in a mixture of natural seawater and frazil ice for 125 days at −2 °C. Dispersed oil in seawater without frazil ice were included in the experimental setup. Presence or absence of frazil ice was a strong driver for microbial community structures and affected the rate of oil degradation. n-alkanes were degraded faster in the presence of frazil ice, the opposite was the case for naphthalenes and 2–3 ring PAHs. No degradation of 4–6 ring PAHs was observed in any of the treatments. The total petroleum oil was not degraded to any significant degree, suggesting that oil will freeze into the ice matrix and persist throughout the icy season. © 2020 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

A key challenge in recirculating aquaculture systems (RAS) is the accumulation of particulate organic matter, especially the fine and colloidal fraction due to low removal efficiency of today's technology. The supply of organic matter is typically the limiting resource determining the carrying capacity (CC) of heterotrophic bacteria in the system. An appropriate and stable CC is proposed as a strategy for an optimal microbial environment in RAS with less blooms of opportunistic bacteria and more stable community dynamics. In this study, we investigated the effects of including a membrane for ultrafiltration in the RAS water treatment loop (treating 10–15% of the total water flow) to reduce the amount of fine and colloidal organic matter. Atlantic salmon parr (Salmo salar) were reared in two pilot-scale RAS (mRAS: membrane, cRAS: conventional). To evaluate the bacterial dynamics with and without membrane filtration at different organic loadings, the water exchange rates of the systems were manipulated equally to create periods with high and low loading of organic matter. The results showed that in the mRAS water, the level of organic matter was more stable throughout the experiment for the changing organic matter loadings. As a consequence, water in mRAS had higher microbial diversity, lower and shorter bacterial blooms and generally lower bacterial densities than in cRAS. All variables indicate a better microbial environment in the water of the system with membrane filtration. Also, the physicochemical water quality was better in mRAS in terms of lower turbidity and particulate organic matter (POC), and slightly lower concentrations of total ammonia nitrogen (TAN). The composition of the microbial communities was significantly different between the two systems, and temporal variations in the community dynamics were observed in both systems during the periods with different organic loadings. At high organic loading, the genus Mycobacterium had high relative abundance in the cRAS water (up to 0.25) compared to mRAS (0.01–0.03). The fish in mRAS were significantly bigger (14%) than fish in cRAS at the end of the experiment, however it is hard to conclude whether the better growth in mRAS was due to higher temperatures (caused by membrane operation) or better water quality, as it was probably a combination of both. We can conclude that membrane filtration gave more stable and better physicochemical and microbial water quality, which will reduce the probability for microbially related accidents in RAS. © 2020 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

The rearing environment is important for a stable production of good quality lobster juveniles. By providing an environment excluding pathogens and dominated by mutualistic bacteria, the probability of developing healthy host-microbe relationships and produce healthy juveniles is increased. Disinfection of water and sudden increase in the supply of organic matter in culture tanks are processes that open for uncontrolled microbial regrowth in the rearing water. This increase the variability in the development of the microbiota between replicate rearing tanks and promotes selection for potentially harmful opportunistic bacteria. In two start feeding experiments with European lobster (Homarus gammarus) we compared the bacterial environment in three types of rearing systems: a recirculating aquaculture system (RAS) with UV treatment directly in front of the rearing raceways, a RAS without disinfection, and a conventional flow through system (FTS). The RAS with no disinfection was hypothesised to stabilise the microbiota of the rearing water, select against opportunistic bacteria, and reduce variability in production outcome between replicate tanks compared to the other systems. As predicted, the three different systems developed significantly different compositions of the microbiota in the rearing water and the larvae. On average, the survival of larvae in RAS without disinfection increased with 43 and 275 %, in the first experiment, and 64 and 18 % in the second experiment, compared to RAS with UV and FTS, respectively. Also, the RAS without disinfection showed less variability in the survival of larvae between replicate tanks and batches compared to the other treatments. The results are promising for controlling the microbiota of the rearing water to improve, increase and stabilise the production of marine larvae by competent use of water treatment and selection regimes. Based on the presented and previous work, RAS is recommended over FTS, and in RAS it is recommended to avoid point-disinfection of the recirculating water, to provide a stable and beneficial microbial environment in the cultivation of marine larvae. https://doi.org/10.1016/j.aquaeng.2021.102176 © 2021 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

Fish live in close contact with microbes in their surrounding water. For cultivated marine fish larvae, stable microbial environments, characterized by slow-growing, K-selected bacteria, have been found to improve growth and survival. Compared to flow-through systems, recirculating aquaculture systems (RAS) offer a better possibility for obtaining microbial stability in the rearing water. However, the microbial ecology in RAS is complex and little studied, and to optimize the microbial water quality more knowledge is needed on microbial community dynamics and interactions between microbes and fish. In the present study we investigated microbial community dynamics in three large-scale RAS, operated with different salinities (12, 22 and 32 ppt), for production of Atlantic salmon post-smolts. We focused on the stability of microbial communities over time in the bioreactors and the fish tanks, and compared the microbial communities within and between the three RAS. The study showed that: 1) The microbial community of the water was similar between the compartments of each RAS at the same time point, 2) The microbial communities of the water (and to a lesser extent the biofilm) within each system underwent large changes over time, 3) The microbiota of biofilm and water differed significantly within each system, 4) Biofilm community profiles were more similar between RAS than the water community profiles, and 5) Salinity structured the microbial community composition. The present study provides new knowledge on how the microbiota in RAS varies with time and how it is influenced by environmental factors.

The selection pressure can be used to reduce the opportunities for proliferation of potentially harmful bacteria. Hypothetically the chance of proliferation of opportunistic bacteria in the fish tank can be minimized by microbial maturation of the incoming water at a microbial carrying capacity similar to that in the rearing tanks. In a start feeding experiment with Ballan wrasse (Labrus bergylta) we compared the bacterial environment in two flow through systems: a microbially matured system (MMS), where the water presented to the fish was matured in a biofilter, and a fed microbially matured system (F-MMS), where the biofilter was fed organic matter to increase the microbial carrying capacity. As predicted, the MMS showed a more variable and often high microbial growth potential in the tank water. The microbial community composition of the tank water was more stable, diverse and species rich in the F-MMS than in the MMS. The results are promising for controlling the microbiota of the rearing water by competent use of water treatment and selection regimes. Statement of relevance The experiment shows that small changes in management (organic load and maturation of water) of water treatment give significant different microbiota in fish tanks. The experiment also shows that by increasing the microbial carrying capacity in a maturation unit to the level of the tank microbial carrying capacity, the microbial community in the fish tank becomes more stable and less open for opportunistic proliferation. This work reveals promising possibilities for controlling the microbiota of the rearing water in land based aquaculture by competent use of water treatment and selection regimes. Improved control and understanding of microbial control is very relevant for the aquaculture industry.

An Atlantic cod larva (Gadus morhua) feeding experiment was carried out to compare three different live feed enrichment products for Brachionus ibericus and Artemia salina: 1) Nannochloropsis sp. microalgal concentrate (frozen) (N), 2) Pavlova viridis microalgal concentrate (frozen) (P), and 3) Larviva Multigain (R) (BioMar, Denmark)-formulated commercial product (CP). Over the experimental period of 42 days larvae of groups P and CP showed significantly higher instantaneous growth rates (G: 0.074 +/- 0.001 d(-1) and 0.079 +/- 0.009 d(-1)) than larvae of group N (G: 0.04 +/- 0.0 d(-1)). Survival at the end of the trial of group CP was highest (20 +/- 8%), whereas significantly lower survival was observed for groups P (7 +/- 2%) and N (

The use of probiotic bacteria is a strategy suggested to overcome microbial problems during early stages of farmed animals, including fish. However, few studies have investigated the efficiency in the establishment of probiotics in the fish and how lasting such treatments are. The objective of this study was to evaluate whether the microbiota of Atlantic cod larvae Gadus morhua L can be steered by introduction of selected probiotic candidates, and if there are particular developmental stages where this is more easily obtained. Cod larvae were given a mixture of 4 candidate probiotic strains (Microbacterium (ID3-10), Ruegeria (RA4-1), Pseudoalteromonas (RA7-14) and Vibrio (RD5-30)) through the live feed and the water during a treatment period of 24 h. The strains originated from reared cod larvae and are proven to have probiotic properties. The larvae were treated at seven different ages, namely days 0, 2, 4, 8, 16, 30 and 45 post-hatching, with one new experimental group started each time. A real-time PCR strategy was developed to quantify the relative amounts of the four added bacterial strains in live feed, and in water and cod larvae during 10 days after exposure. Only ID3-10 was measured to constitute considerable fractions of the larval microbiota for most of the treatments, despite the fact that all the probiotic candidates originated from cod larvae intestines. In most cases, the amounts of the added strains decreased to approximately background levels after a maximum of 4 days, indicating only a transient presence in the larvae. The results indicate that probiotic treatment aiming for colonization with new strains in the cod larvae is difficult, and challenges the probiotic concept for the larval stage unless continuous or repeated addition to the fish larvae is used.

A high share of opportunistic and fast-growing microbes is generally negative for the performance of marine fish larvae. It is hypothesised that larvae that are reared in water dominated by K-strategists (mature microbial communities) will perform better, because they are less likely to encounter opportunistic (r-selected) microbes and develop detrimental host–microbe interactions. K-selection can be set up by securing competition for resources by maintaining a low nutrient supply per microbe in a maturing biofilter in a flow through system or in a recirculation system without disinfection. In an experiment with Atlantic cod larvae (Gadus morhua L.) the development of the microbial community in the inflowing water to tanks as well as in the tank water was studied in a conventional flow through system (FTS) and two hypothesised K-selective systems: a microbially matured flow through system (MMS) and a recirculation aquaculture system (RAS). As predicted, the K-selective treatments resulted in a significantly more mature and stable microbial community composition in the water going in to the fish tanks compared to the FTS. The larvae in the K-selective treatments showed 72% higher survival than the larvae in the FTS. In K-selective systems where the inflowing water to the fish tanks is matured at a microbial carrying capacity that is significantly lower than the carrying capacity in the rearing tank (i.e. MMS) there is hypothetically an opening for proliferation of opportunists to match the higher carrying capacity in the tank water. As predicted, the RAS showed K-selection of the incoming water at a carrying capacity similar to that of the tank water, which resulted in a significantly more stable microbial community composition in the tank water of the RAS than in the MMS and FTS. The results of this experiment showed a high potential for increasing fish survival by using K-selection of bacteria, which is a cheap and easy method that can be used in all kinds of new or existing aquaculture systems.

Particle accumulation in recirculating aquaculture systems (RAS) are generated from uneaten food and feces, and bacteria tends to proliferate. Small organic particles act as substrate for bacterial growth and may by direct interference with gills cause stress and reduce disease resistance for the cultivated species. Most diseases in aquaculture of marine fish larvae are caused by opportunistic microorganisms that become pathogenic when the fish is under stressed conditions. The present study investigated the potential of using a membrane bioreactor (MBR) as a part of the water treatment in a RAS for production of marine fish larvae, since the small pore size (50 nm) efficiently removes organic particles including the colloidal fraction in the system. The experiment was a 50 day start-feeding experiment with Atlantic cod ( Gadus morhua L.), comparing water quality and larval performance in a conventional RAS (cRAS) with a membrane filtrated RAS (mRAS). In mRAS, 8.5% of the water flow was at any time membrane filtrated. The present study showed that the MBR improved water quality significantly by reducing turbidity and by lowering the total number of bacteria in the rearing water (up to 80% reduction). Microbial communities in water samples were significantly different between cRAS and mRAS already on 5 days post-hatching (dph) and throughout the experiment. The microbial community composition of the rearing water was significantly different from the composition of the larval microbiota during the experiment, although differences between larval samples were different only on 50 dph. The present study showed the potential of using MBR to lower the bacterial carrying capacity (CC) by efficiently removing organic particles and bacteria. In average a 13% higher cod larval growth (weight, %) at 40 dph and a 3.0% higher survival rate at 50 dph were measured in the mRAS scheme.

The minke whale (Balaenoptera acutorostrata) is subject to commercial whaling, but stock identification and assessment are still uncertain. Mitochondrial DNA (mtDNA) sequences were determined to examine the population structure of minke whales from the central and northeastern parts of the North Atlantic, as well as the Antarctic regions IV and V. The analyses include 345 nucleotide positions of the control region of 110 individuals, and 250 nucleotide positions of the NADH dehydrogenase subunit 2 gene for a representative selection of North Atlantic minke whales. Maximum parsimony analyses and sequence divergence calculations did not reveal any genetic differentiation between individuals from the central and northeastern parts of the North Atlantic. These results do not support the International Whaling Commission's separation of minke whales in this area into different management units, and they are in conflict with previously reported results from allozyme analyses. Comparison of minke whale control region sequences showed that the sequence diversity of North Atlantic minke whales is substantially lower (0.0065) than that of Antarctic minke whales (0.0166), and clearly demonstrated that individuals from these two areas represent genetically distinct populations.