Phylogeographic mitogenomics of Atlantic cod Gadus morhua: Variation in and among trans‐Atlantic,trans‐Laurentian,Northern cod,and landlocked fjord populations

The historical phylogeography, biogeography, and ecology of Atlantic cod (Gadus morhua) have been impacted by cyclic Pleistocene glaciations, where drops in sea temperatures led to sequestering of water in ice sheets, emergence of continental shelves, and changes to ocean currents. High‐resolution, whole‐genome mitogenomic phylogeography can help to elucidate this history. We identified eight major haplogroups among 153 fish from 14 populations by Bayesian, parsimony, and distance methods, including one that extends the species coalescent back to ca. 330 kya. Fish from the Barents and Baltic Seas tend to occur in basal haplogroups versus more recent distribution of fish in the Northwest Atlantic. There was significant differentiation in the majority of trans‐Atlantic comparisons (Φ_ST = .029–.180), but little or none in pairwise comparisons within the Northwest Atlantic of individual populations (Φ_ST = .000–.060) or defined management stocks (Φ_ST = .000–.023). Monte Carlo randomization tests of population phylogeography showed significantly nonrandom trans‐Atlantic phylogeography versus absence of such structure within various partitions of trans‐Laurentian, Northern cod (NAFO 2J3KL) and other management stocks, and Flemish Cap populations. A landlocked meromictic fjord on Baffin Island comprised multiple identical or near‐identical mitogenomes in two major polyphyletic clades, and was significantly differentiated from all other populations (Φ_ST = .153–.340). The phylogeography supports a hypothesis of an eastern origin of genetic diversity ca. 200–250 kya, rapid expansion of a western superhaplogroup comprising four haplogroups ca. 150 kya, and recent postglacial founder populations.     

Antimicrobial activity in serum of Atlantic cod Gadus morhua L.: comparison between cultured and wild fish

The present investigation determined the antibacterial activities in the sera of both cultured and wild Atlantic cod Gadus morhua . Serum samples from both groups of fish were quantified for total protein, and their effects against fish pathogens Listonella anguillarum , Aeromonas salmonicida and Yersinia ruckeri were determined using co-incubation assay. Total serum protein concentrations were not significantly different between farmed and wild Atlantic cod. Sera of cultured Atlantic cod significantly decreased growth of L. anguillarum by at least two-log₁₀ reductions in bacterial count, while those of the wild Atlantic cod were able to inhibit the growth of all three fish bacterial pathogens. The present study showed that sera from wild fish possess broader antibacterial activities than cultured Atlantic cod and that these could provide an explanation for the differences regarding their immunity to bacterial infections as well as their health status.     

Distribution of viral haemorrhagic septicaemia virus in wild fish species of the North Sea, north east Atlantic Ocean and Irish Sea.

A surveillance programme was initiated on the occurrence and distribution of viral haemorrhagic septicaemia virus (VHSV) in wild marine fish. Six research cruises were undertaken in an 18 mo period during 1997 and 1998, covering the North Sea, the Atlantic waters off the north and west coasts of Scotland and the Irish Sea. A total of 19,293 fish were sampled from 23 different species including cod, haddock, Norway pout, herring and sprat. Individual fish lengths were recorded and the fish were checked for lesions, haemorrhaging and other signs of disease. Pools of organ samples were taken for virus assay. The majority of fish sampled did not display clinical signs indicative of viral haemorrhagic septicaemia. A small number of cod were found with skin lesions and haddock with skin haemorrhaging. Of the 2081 organ and skin sample pools collected, 21 tested positive for VHSV by tissue culture and enzyme-linked immunosorbent assay. Seventeen of the isolates originated from Norway pout Trisopterus esmarkii, one from cod Gadus morhua (skin lesion), one from herring Clupea harengus, one from whiting Merlangius merlangus, and one from a previously unreported host species, poor cod Trisopterus minutus.     

Gene flow and lack of population differentiation in Atlantic cod, Gadus morhua L., from Iceland, and comparison of cod from Norway and Newfoundland

Restriction analysis of mitochondrial DNA was used to study genetic variation and geographic population structure of Atlantic cod from localities around Iceland. Gene phylogenies were constructed and geographic locations superimposed on these. The variation was not localized. Estimated gene flow was large. Thus, Atlantic cod in Iceland belong to a single genetic population. Analyses of published sequence variation of cod from Norway and Newfoundland showed that the extensive continuity of intraspecific phylogeny of cod in Iceland, which is very similar to other marine organisms with a similar life history, extends from Norway to Newfoundland and possibly to larger areas of the Atlantic.     

Strong genetic differentiation and postglacial origin of populations in the marine midge Clunio marinus (Chironomidae,Diptera)

The marine midge Clunio marinus (Chironomidae, Diptera) is characterized by a one‐dimensional distribution along the European Atlantic coast, where its lunar and circadian emergence rhythms are genetically adapted to the local tidal regimes, resulting in a series of ‘temporal races’. Clunio marinus is restricted to rocky coasts and thus the temporal races occur in different rocky patches. We studied 10 populations of Clunio marinus from five different regions, spanning the major rocky mainland coasts from Spain to Norway, using amplified fragment length polymorphisms (AFLP), microsatellites and mitochondrial cytochrome oxidase I (COI) sequences. Star‐like patterns of COI haplotypes within regions indicate postglacial colonization. A high degree of shared polymorphisms in AFLP markers suggests colonization from a single source, implying postglacial evolution of timing adaptations in relation to the local tidal regime. In contrast, no COI haplotypes are shared among regions. We hypothesize that different levels of differentiation of nuclear vs. mitochondrial markers in the source region were carried forward during postglacial expansion. Despite the recent origin of populations, all markers reveal distinct genetic differentiation between rocky coasts on a scale of 650–1150 km. Differentiation between rocky coasts is not correlated to timing adaptations, suggesting that geographic isolation is prevalent between rocky coasts and that this facilitated the evolution of local timing adaptations. At the same time there is little genetic differentiation within rocky coasts on a scale of 2–6 km; leaving open the possibility that within rocky coasts with large variation in tidal regimes, temporal adaptations evolved in the face of gene flow.     

Karyotyping and cytogenetic mapping of Atlantic cod (Gadus morhua Linnaeus, 1758)

The Atlantic cod (Gadus morhua) is an important natural resource for northern societies and is now also considered to be a promising candidate for aquaculture. In recent years, much effort has been directed towards the development of genomic tools, and genome initiatives for Atlantic cod have been established. Despite the growing attention devoted to the Atlantic cod genomics, basic aspects of its genome structure and organization remain unknown. Thus, the present work aims to study cytogenetic features of the Atlantic cod as a contribution to the knowledge of this species' genome. The Atlantic cod displays a diploid number of 46 chromosomes, with a karyotypic formula 16 m/sm + 30 st/t. Conventional karyotyping was improved by chromosomal mapping of two classes of repetitive sequences. 18S rDNA clusters were assigned to pairs 2 and 4; small amounts of 18S rDNA clusters were occasionally detected on pair 5. These findings could not be related to the geographical origin of the specimens, but were consistent with the variability of these repeated genes in fish in general. 5S ribosomal gene clusters, apparently corresponding to a single 5S rDNA class, were detected on twelve chromosomes (pairs 11, 12, 14, 17, 20 and 21). The present update of the existing but meagre information on the karyotype of Atlantic cod, plus the first physical mapping of repetitive genes in this species herein, opens the way for an integrated approach that combines genetic and physical mapping with the assembly of the genome of this commercially important species.     

Molecular Genetic Analysis of Stomach Contents Reveals Wild Atlantic Cod Feeding on Piscine Reovirus (PRV) Infected Atlantic Salmon Originating from a Commercial Fish Farm

In March 2012, fishermen operating in a fjord in Northern Norway reported catching Atlantic cod, a native fish forming an economically important marine fishery in this region, with unusual prey in their stomachs. It was speculated that these could be Atlantic salmon, which is not typical prey for cod at this time of the year in the coastal zone. These observations were therefore reported to the Norwegian Directorate of Fisheries as a suspected interaction between a local fish farm and this commercial fishery. Statistical analyses of genetic data from 17 microsatellite markers genotyped on 36 partially-degraded prey, samples of salmon from a local fish farm, and samples from the nearest wild population permitted the following conclusions: 1. The prey were Atlantic salmon, 2. These salmon did not originate from the local wild population, and 3. The local farm was the most probable source of these prey. Additional tests demonstrated that 21 of the 36 prey were infected with piscine reovirus. While the potential link between piscine reovirus and the disease heart and skeletal muscle inflammation is still under scientific debate, this disease had caused mortality of large numbers of salmon in the farm in the month prior to the fishermen''s observations. These analyses provide new insights into interactions between domesticated and wild fish.     

Harvest Pressure on Coastal Atlantic Cod (Gadus morhua) from Recreational Fishing Relative to Commercial Fishing Assessed from Tag-Recovery Data

Marine recreational fishing is a popular outdoor activity. However, knowledge about the magnitude of recreational catches relative to commercial catches in coastal fisheries is generally sparse. Coastal Atlantic cod (Gadus morhua) is a target species for recreational fishers in the North Atlantic. In Norway, recreational fishers are allowed to use a variety of traps and nets as well as long-line and rod and line when fishing for cod. From 2005 to 2013, 9729 cod (mean size: 40 cm, range: 15–93 cm) were tagged and released in coastal Skagerrak, southeast Norway. Both high-reward (NOK 500) and low-reward tags (NOK 50) were used in this study. Because some harvested fish (even those posting high-reward tags) may go unreported by fishers, reporting rates were estimated from mark-recovery models that incorporate detection parameters in their structure, in addition to survival and mortality estimates. During 2005 to 2013, a total of 1707 tagged cod were recovered and reported by fishers. We estimate the overall annual survival to be 33% (SE 1.5). Recreational rod and line fishing were responsible for 33.7% (SE 2.4) of total mortality, followed by commercial fisheries (15.1% SE 0.8) and recreational fixed gear (6.8% SE 0.4). Natural mortality was 44.4% (SE 2.5) of total mortality. Our findings suggest that recreational fishing—rod and line fishing in particular—is responsible for a substantial part of fishing mortality exerted on coastal cod in southern Norway.     

Disentangling structural genomic and behavioural barriers in a sea of connectivity

Genetic divergence among populations arises through natural selection or drift and is counteracted by connectivity and gene flow. In sympatric populations, isolating mechanisms are thus needed to limit the homogenizing effects of gene flow to allow for adaptation and speciation. Chromosomal inversions act as an important mechanism maintaining isolating barriers, yet their role in sympatric populations and divergence with gene flow is not entirely understood. Here, we revisit the question of whether inversions play a role in the divergence of connected populations of the marine fish Atlantic cod (Gadus morhua), by exploring a unique data set combining whole‐genome sequencing data and behavioural data obtained with acoustic telemetry. Within a confined fjord environment, we find three genetically differentiated Atlantic cod types belonging to the oceanic North Sea population, the western Baltic population and a local fjord‐type cod. Continuous behavioural tracking over 4 year revealed temporally stable sympatry of these types within the fjord. Despite overall weak genetic differentiation consistent with high levels of gene flow, we detected significant frequency shifts of three previously identified inversions, indicating an adaptive barrier to gene flow. In addition, behavioural data indicated that North Sea cod and individuals homozygous for the LG12 inversion had lower fitness in the fjord environment. However, North Sea and fjord‐type cod also occupy different depths, possibly contributing to prezygotic reproductive isolation and representing a behavioural barrier to gene flow. Our results provide the first insights into a complex interplay of genomic and behavioural isolating barriers in Atlantic cod and establish a new model system towards an understanding of the role of genomic structural variants in adaptation and diversification.     

Plasmid profiling of Vibrio salmonicida for epidemiological studies of cold-water vibriosis in Atlantic salmon (Salmo salar) and cod (Gadus morhua).

In 1988, a new plasmid profile was observed for Vibrio salmonicida isolated from cod (Gadus morhua) and Atlantic salmon (Salmo salar) in fish farms in northern Norway. This new plasmid profile, which consisted of plasmids of 61, 21, 3.4, and 2.8 megadaltons, is 1 of 11 plasmid profiles which have so far been observed for V. salmonicida. Plasmid profiling and plasmid DNA hybridization were used in epidemiological studies of cold-water vibriosis. Our results indicate that V. salmonicida was transmitted from Atlantic salmon to cod and vice versa. The 61-megadalton plasmid was found exclusively in V. salmonicida strains originating from northern Norway, which is the only area in which this plasmid has ever been observed. Plasmid DNA hybridization and restriction endonuclease analysis show that the plasmid DNA of V. salmonicida remained stable throughout a 7-year survey.     

Plasmid profiling of Vibrio salmonicida for epidemiological studies of cold-water vibriosis in Atlantic salmon (Salmo salar) and cod (Gadus morhua)

In 1988, a new plasmid profile was observed for Vibrio salmonicida isolated from cod (Gadus morhua) and Atlantic salmon (Salmo salar) in fish farms in northern Norway. This new plasmid profile, which consisted of plasmids of 61, 21, 3.4, and 2.8 megadaltons, is 1 of 11 plasmid profiles which have so far been observed for V. salmonicida. Plasmid profiling and plasmid DNA hybridization were used in epidemiological studies of cold-water vibriosis. Our results indicate that V. salmonicida was transmitted from Atlantic salmon to cod and vice versa. The 61-megadalton plasmid was found exclusively in V. salmonicida strains originating from northern Norway, which is the only area in which this plasmid has ever been observed. Plasmid DNA hybridization and restriction endonuclease analysis show that the plasmid DNA of V. salmonicida remained stable throughout a 7-year survey.     

Previously unrecognised division within Moritella viscosa isolated from fish farmed in the North Atlantic

Previously undocumented phenotypical and genetic variation was identified amongst isolates of Moritella viscosa collected from various geographical locations and from different fish species. The studied isolates could be split into 2 major phenotypically and genetically different clusters, one of which was consistent with the species type strain (NCIMB 13548). Isolates consistent with the type strain originated exclusively from Atlantic salmon farmed in Norway, Scotland and the Faroe Isles, although a single isolate from farmed Norwegian cod clustered closely with this group. The 'variant' cluster comprised isolates originating from Norwegian farmed rainbow trout, Icelandic farmed rainbow trout and salmon, Canadian farmed (Atlantic) salmon, Icelandic lumpsucker and only exceptionally from Norwegian salmon. With the exception of the single aforementioned cod isolate, all isolates from Norwegian farmed cod belonged to the variant cluster. Phenotypically, the clusters could be absolutely separated only by elevated haemolytic activity in the variant strain, although approximately half of these isolates also produced acid from mannose, in contrast to the typical (type) strain. While 16S rRNA gene sequencing was unable to separate the 2 clusters, Western blot analyses, plasmid profile analysis, pulsed field gel electrophoresis and gyrB gene sequence analysis produced clusters consistent with the phenotypic data. Macroscopically and histologically the disease in rainbow trout caused by the variant strain was consistent with that previously described in Atlantic salmon. The results of the present study may indicate a degree of host specificity of the typical strain for Atlantic salmon.     

Changes in the Intestinal Microbiota of Wild Atlantic cod Gadus morhua L. Upon Captive Rearing

The commensal microbiota plays an important role in the well-being of the host organism, and it would be worthwhile to know the tenacious communities among them. Therefore, a study was undertaken to examine the changes in constitution of the intestinal microbiota of wild fish consequential to captivity. At first, the composition of intestinal microorganisms of Atlantic cod caught from the coastal area off Bod?, Norway, was examined. Thereafter, the changes in the bacterial community of the captive fish after offering them artificial feed or subjecting them to starvation were studied. The microbiota from the intestinal contents and wall segments were analyzed quantitatively by spread plate technique and DAPI staining and qualitatively by denaturing gradient gel electrophoresis. The study revealed that the counts of intestinal microbes in wild-caught Atlantic cod were not affected by captive rearing for 6?weeks, either when fed or when starved. However, the diversity of intestinal bacterial community was reduced in response to artificial feeding, whereas the change was restricted upon starvation.     

Expression of vasa and nanos3 during primordial germ cell formation and migration in Atlantic cod (Gadus morhua L.)

Primordial germ cells (PGCs), progenitors of gametes, are specified very early in embryonic development and undergo an active migration to the site where the future gonads will form. While the developmental pattern of PGCs during embryogenesis has been documented in few model teleost fishes, there is currently no information available for any representative of Superorder Paracanthopterygii. This includes Atlantic cod (Gadus morhua), which is a historically important food fish in both fisheries and aquaculture industries. In the present study, we cloned and characterized vasa and nanos3 and used them as germ cell markers in Atlantic cod. Sequencing results showed prospective vasa and nanos3 mRNA contained the domains used to describe their respective protein family. Furthermore, phylogenetic analysis using the amino acid sequence placed Atlantic cod Vasa distinct from representatives of three other taxonomic Superorders. Atlantic cod Nanos3 was placed with other homologues from the Nanos3 subfamily. Expression of both genes was detected from the first cleavage division; both were specifically expressed in Atlantic cod PGCs from the 32-cell stage. While nanos3 expression ceased during early somitogenesis, vasa was strongly expressed throughout embryonic development. Using vasa as a marker, we described the Atlantic cod PGC migration pattern. We demonstrated that Atlantic cod PGCs migrate ventral to the trunk mesoderm. With the exception of Pacific herring (Clupea pallasii), PGCs in other described teleost fishes migrate lateral to the trunk. The results from this study are the first step toward understanding germ line formation in Atlantic cod.     

A reappraisal of activity metabolism in Atlantic cod (Gadus morhua)

Atlantic cod ( Gadus morhua ) were forced to swim in a swim tunnel respirometer until fatigued; oxygen consumption rate (O₂) was measured during swimming at incremental speeds until the fish was exhausted and during recovery from exhaustion. Maximal oxygen consumption (O_2max) occurred during maximal activity as has been found for other fish species, but at odds with the current paradigm for Atlantic cod. Earlier experiments had drawn the conclusion that O_2max in Atlantic cod occurs during recovery from exhaustive exercise. We found no support for this paradigm in our experiments and we propose that the respiratory physiology of Atlantic cod is not unlike that of other fishes.     

Ancient DNA reveals a southern presence of the Northeast Arctic cod during the Holocene

Climate change has been implicated in an increased number of distributional shifts of marine species during the last century. Nonetheless, it is unclear whether earlier climatic fluctuations had similar impacts. We use ancient DNA to investigate the long-term spawning distribution of the Northeast Arctic cod (skrei) which performs yearly migrations from the Barents Sea towards spawning grounds along the Norwegian coast. The distribution of these spawning grounds has shifted northwards during the last century, which is thought to be associated with food availability and warming temperatures. We genetically identify skrei specimens from Ruskeneset in west Norway, an archaeological site located south of their current spawning range. Remarkably, ¹⁴C analyses date these specimens to the late Holocene, when temperatures were warmer than present-day conditions. Our results either suggest that temperature is not the only driver influencing the spawning distribution of Atlantic cod, or could be indicative of uncertainty in palaeoclimate reconstructions in this region. Regardless, our findings highlight the utility of aDNA to reconstruct the historical distribution of economically important fish populations and reveal the complexity of long-term ecological interactions in the marine environment.     

Differences in lens optical plasticity in two gadoid fishes meeting in the Arctic

Arctic and boreal/temperate species are likely to be evolutionary adapted to different light regimes. Currently, the boreal/temperate Atlantic cod (Gadus morhua) is coexisting with the native polar cod (Boreogadus saida) in the Arctic waters around Svalbard, Norway. Here, we studied light/dark adaptative optical plasticity of their eye lenses by exposing fish to bright light during the polar night. Schlieren photography, high-definition laser scanning and ray tracing were used to determine the optical properties of excised crystalline lenses. Both species have multifocal lenses, an optical adaptation for improved color vision. In polar cod, the optical properties of the lens were independent of light exposure. In the more southern Atlantic cod, the optical properties of the lens changed within hours upon exposure to light, even after months of darkness. Such fast optical adjustment has previously only been shown in a tropical cichlid. During the polar night the Atlantic cod lens seems to be unregulated and dysfunctional since it had an unsuitable focal length and severe spherical aberration. We present a system, to our knowledge unique, for studying visual plasticity on different timescales in relation to evolutionary history and present the first study on the polar cod visual system.     

The genome size and the structure and content of ribosomal RNA genes in Atlantic cod (Gadus morhua L.).

1. The haploid genome size of the Atlantic cod was estimated to 3.4 x 10(8)kb by reassociation kinetics analysis of cod sperm DNA. 2. The size of the small and large subunit ribosomal RNAs is 1.85 and 4.1 kb, respectively. 3. Restriction enzyme mapping of the rRNA coding unit revealed conservation of an Eco RI site in the coding regions of 18 S and 28 S rRNA and a Bam HI site in the 28 S rRNA coding region compared to other fish species. 4. The length of the repeat unit of the cod rDNA was found to be 30 kb. 5. The rRNA genes are repeated approximately 50 times in the cod genome and constitutes 0.08% of the cod genetic material.     

Ancient chromosomal rearrangement associated with local adaptation of a postglacially colonized population of Atlantic Cod in the northwest Atlantic

Intraspecific diversity is central to the management and conservation of exploited species, yet knowledge of how this diversity is distributed and maintained in the genome of many marine species is lacking. Recent advances in genomic analyses allow for genome‐wide surveys of intraspecific diversity and offer new opportunities for exploring genomic patterns of divergence. Here, we analysed genome‐wide polymorphisms to measure genetic differentiation between an offshore migratory and a nonmigratory population and to define conservation units of Atlantic Cod (Gadus morhua) in coastal Labrador. A total of 141 individuals, collected from offshore sites and from a coastal site within Gilbert Bay, Labrador, were genotyped using an ~11k single nucleotide polymorphism array. Analyses of population structure revealed strong genetic differentiation between migratory offshore cod and nonmigratory Gilbert Bay cod. Genetic differentiation was elevated for loci within a chromosomal rearrangement found on linkage group 1 (LG1) that coincides with a previously found double inversion associated with migratory and nonmigratory ecotype divergence of cod in the northeast Atlantic. This inverted region includes several genes potentially associated with adaptation to differences in salinity and temperature, as well as influencing migratory behaviour. Our work provides evidence that a chromosomal rearrangement on LG1 is associated with parallel patterns of divergence between migratory and nonmigratory ecotypes on both sides of the Atlantic Ocean.