Nasal responses to elevated temperature and Francisella noatunensis infection in Atlantic cod (Gadus morhua).

We report the histological and transcriptomic changes in the olfactory organ of Atlantic cod exposed to Francisella noatunensis. Experimental infection was performed at either 12 °C or 17 °C. Infected fish presented the classic gross pathologies of francisellosis. Nasal morpho-phenotypic parameters were not significantly affected by elevated temperature and infection, except for the number of mucus cells in the 12 °C group seven weeks after the challenge. A higher number of genes were altered through time in the group reared at 17 °C. At termination, the nasal transcriptome of infected fish in both groups was similar to the control. When both infected groups were compared, 754 DEGs were identified, many of which were involved in signalling, defence, transmembrane and enzymatic processes. In conclusion, the study reveals that elevated temperature could trigger responses in the olfactory organ of Atlantic cod and shape the nasal response to F. noatunensis infection.

Comparative basal transcriptome profiles of the olfactory rosette and gills of Atlantic salmon (Salmo salar) unveil shared and distinct immunological features.

The molecular repertoire of the mucosa-associated lymphoid tissue (MALT) in the olfactory rosette in most teleost fish is unknown. Here we present the basal transcriptome of the olfactory rosette of Atlantic salmon (Salmo salar). To investigate its mucosal immune features, we performed a comparative transcriptomic analysis with the gills, one of the most studied organs possessing MALT. Pathway enrichment revealed that cytokine-cytokine interaction and the neuroactive ligand-receptor interaction pathways were at the core of the shared similarity between the two organs. The immunological features of the two organs were further characterised by the overrepresentation of several immune-related pathways, particularly important for pathogen recognition. The immunological differences between the two organs were underlined with the differential regulation of markers for interleukins, extracellular matrix, antimicrobial peptides, and complement. The basal transcriptome of Atlantic salmon olfactory rosette is a valuable molecular toolbox that will advance our understanding of nasal immunity in teleost fish.

Immunologic Profiling of the Atlantic Salmon Gill by Single Nuclei Transcriptomics.

Anadromous salmonids begin life adapted to the freshwater environments of their natal streams before a developmental transition, known as smoltification, transforms them into marine-adapted fish. In the wild, smoltification is a photoperiod-regulated process, involving radical remodeling of gill function to cope with the profound osmotic and immunological challenges of seawater (SW) migration. While prior work has highlighted the role of specialized "mitochondrion-rich" cells (MRCs) and accessory cells (ACs) in delivering this phenotype, recent RNA profiling experiments suggest that remodeling is far more extensive than previously appreciated. Here, we use single-nuclei RNAseq to characterize the extent of cytological changes in the gill of Atlantic salmon during smoltification and SW transfer. We identify 20 distinct cell clusters, including known, but also novel gill cell types. These data allow us to isolate cluster-specific, smoltification-associated changes in gene expression and to describe how the cellular make-up of the gill changes through smoltification. As expected, we noted an increase in the proportion of seawater mitochondrion-rich cells, however, we also identify previously unknown reduction of several immune-related cell types. Overall, our results provide fresh detail of the cellular complexity in the gill and suggest that smoltification triggers unexpected immune reprogramming.

Photoperiod-dependent developmental reprogramming of the transcriptional response to seawater entry in Atlantic salmon (Salmo salar).

The developmental transition of juvenile salmon from a freshwater resident morph (parr) to a seawater (SW) migratory morph (smolt), known as smoltification, entails a reorganization of gill function to cope with the altered water environment. Recently, we used RNAseq to characterize the breadth of transcriptional change which takes place in the gill in the FW phase of smoltification. This highlighted the importance of extended exposure to short, winter-like photoperiods (SP) followed by a subsequent increase in photoperiod for completion of transcriptional reprogramming in FW and efficient growth following transfer to SW. Here, we extend this analysis to examine the consequences of this photoperiodic history-dependent reprogramming for subsequent gill responses upon exposure to SW. We use RNAseq to analyze gill samples taken from fish raised on the photoperiod regimes we used previously and then challenged by SW exposure for 24 hours. While fish held on constant light (LL) throughout were able to hypo-osmoregulate during a 24 hours SW challenge, the associated gill transcriptional response was highly distinctive from that in fish which had experienced a 7-week period of exposure to SP followed by a return to LL (SPLL) and had consequently acquired the characteristics of fully developed smolts. Fish transferred from LL to SP, and then held on SP for the remainder of the study was unable to hypo-osmoregulate, and the associated gill transcriptional response to SW exposure featured many transcripts apparently regulated by the glucocorticoid stress axis and by the osmo-sensing transcription factor NFAT5. The importance of these pathways for the gill transcriptional response to SW exposure appears to diminish as a consequence of photoperiod mediated induction of the smolt phenotype, presumably reflecting preparatory developmental changes taking place during this process.

Diversified regulation of circadian clock gene expression following whole genome duplication.

Across taxa, circadian control of physiology and behavior arises from cell-autonomous oscillations in gene expression, governed by a networks of so-called 'clock genes', collectively forming transcription-translation feedback loops. In modern vertebrates, these networks contain multiple copies of clock gene family members, which arose through whole genome duplication (WGD) events during evolutionary history. It remains unclear to what extent multiple copies of clock gene family members are functionally redundant or have allowed for functional diversification. We addressed this problem through an analysis of clock gene expression in the Atlantic salmon, a representative of the salmonids, a group which has undergone at least 4 rounds of WGD since the base of the vertebrate lineage, giving an unusually large complement of clock genes. By comparing expression patterns across multiple tissues, and during development, we present evidence for gene- and tissue-specific divergence in expression patterns, consistent with functional diversification of clock gene duplicates. In contrast to mammals, we found no evidence for coupling between cortisol and circadian gene expression, but cortisol mediated non-circadian regulated expression of a subset of clock genes in the salmon gill was evident. This regulation is linked to changes in gill function necessary for the transition from fresh- to sea-water in anadromous fish. Overall, this analysis emphasises the potential for a richly diversified clock gene network to serve a mixture of circadian and non-circadian functions in vertebrate groups with complex genomes.

Correction: RNA profiling identifies novel, photoperiod-history dependent markers associated with enhanced saltwater performance in juvenile Atlantic salmon.

[This corrects the article DOI: 10.1371/journal.pone.0227496.].

RNA profiling identifies novel, photoperiod-history dependent markers associated with enhanced saltwater performance in juvenile Atlantic salmon.

Atlantic salmon migrate to sea following completion of a developmental process known as smolting, which establishes a seawater (SW) tolerant phenotype. Smolting is stimulated by exposure to long photoperiod or continuous light (LL) following a period of exposure to short photoperiod (SP), and this leads to major changes in gill ion exchange and osmoregulatory function. Here, we performed an RNAseq experiment to discover novel genes involved in photoperiod-dependent remodeling of the gill. This revealed a novel cohort of genes whose expression rises dramatically in fish transferred to LL following SP exposure, but not in control fish maintained continuously on LL or on SP. A follow-up experiment revealed that the SP-history dependence of LL induction of gene expression varies considerably between genes. Some genes were inducible by LL exposure after only 2 weeks exposure to SP, while others required 8 weeks prior SP exposure for maximum responsiveness to LL. Since subsequent SW growth performance is also markedly improved following 8 weeks SP exposure, these photoperiodic history-dependent genes may be useful predictive markers for full smolt development.

Phenology and cover of plant growth forms predict herbivore habitat selection in a high latitude ecosystem.

The spatial and temporal distribution of forage quality is among the most central factors affecting herbivore habitat selection. Yet, for high latitude areas, forage quantity has been found to be more important than quality. Studies on large ungulate foraging patterns are faced with methodological challenges in both assessing animal movements at the scale of forage distribution, and in assessing forage quality with relevant metrics. Here we use first-passage time analyses to assess how reindeer movements relate to forage quality and quantity measured as the phenology and cover of growth forms along reindeer tracks. The study was conducted in a high latitude ecosystem dominated by low-palatable growth forms. We found that the scale of reindeer movement was season dependent, with more extensive area use as the summer season advanced. Small-scale movement in the early season was related to selection for younger stages of phenology and for higher abundances of generally phenologically advanced palatable growth forms (grasses and deciduous shrubs). Also there was a clear selection for later phenological stages of the most dominant, yet generally phenologically slow and low-palatable growth form (evergreen shrubs). As the summer season advanced only quantity was important, with selection for higher quantities of one palatable growth form and avoidance of a low palatable growth form. We conclude that both forage quality and quantity are significant predictors to habitat selection by a large herbivore at high latitude. The early season selectivity reflected that among dominating low palatability growth forms there were palatable phenological stages and palatable growth forms available, causing herbivores to be selective in their habitat use. The diminishing selectivity and the increasing scale of movement as the season developed suggest a response by reindeer to homogenized forage availability of low quality.

Detection of transgenes in local maize varieties of small-scale farmers in eastern cape, South Africa.

Small-scale subsistence farmers in South Africa have been introduced to genetically modified (GM) crops for more than a decade. Little is known about i) the extent of transgene introgression into locally recycled seed, ii) what short and long-term ecological and socioeconomic impacts such mixing of seeds might have, iii) how the farmers perceive GM crops, and iv) to what degree approval conditions are followed and controlled. This study conducted in the Eastern Cape, South Africa, aims primarily at addressing the first of these issues. We analysed for transgenes in 796 individual maize plants (leaves) and 20 seed batches collected in a village where GM insect resistant maize was previously promoted and grown as part of an governmental agricultural development program over a seven year period (2001-2008). Additionally, we surveyed the varieties of maize grown and the farmers' practices of recycling and sharing of seed in the same community (26 farmers were interviewed). Recycling and sharing of seeds were common in the community and may contribute to spread and persistence of transgenes in maize on a local or regional level. By analysing DNA we found that the commonly used transgene promoter p35s occurred in one of the 796 leaf samples (0.0013%) and in five of the 20 seed samples (25%). Three of the 20 seed samples (15%) included herbicide tolerant maize (NK603) intentionally grown by the farmers from seed bought from local seed retailers or acquired through a currently running agricultural development program. The two remaining positive seed samples (10%) included genes for insect resistance (from MON810). In both cases the farmers were unaware of the transgenes present. In conclusion, we demonstrate that transgenes are mixed into seed storages of small-scale farming communities where recycling and sharing of seeds are common, i.e. spread beyond the control of the formal seed system.