A temperature shift during embryogenesis impacts prevalence of deformity in diploid and triploid Atlantic salmon (Salmo salar L.)

The study investigated the effects of a temperature shift during embryogenesis on diploid and triploid Atlantic salmon (Salmo salar L.) embryo development and juvenile skeletal deformities. From fertilization, sibling populations were incubated under one of three temperatures (6, 8 or 11°C) until 400 °days when all fish were then reared under a common temperature until smolt. Survival was negatively impacted by increasing temperatures irrespective of ploidy. There was no effect of incubation temperatures on growth in diploids, but triploids incubated at 6°C had improved growth rates (thermal growth coefficient, TGC: 6°C: 1.05, 8°C: 0.94, 11°C: 0.48). Fish from 11°C in both ploidies showed increased jaw and vertebral deformity prevalence. In response to the temperature change at 400 °days post fertilization, upregulation of bmp2, bmp4, col2a1, mmp13, opn and sparc, and downregulation of ocn further suggest that bone and cartilage formation is compromised after experiencing a thermal shift. The data show that temperature profile during embryogenesis strongly influences future growth and deformity prevalence. Triploids appear to require a lower incubation temperature than the current industry standard of 8°C to promote better overall performance; however, a thermal shift during embryogenesis was shown to impact expression of important developmental genes

Characterization of External Mucosal Microbiomes of Nile Tilapia and Grey Mullet Co-cultured in Semi-Intensive Pond Systems

The external mucosal surfaces of the fish harbor complex microbial communities, which may play pivotal roles in the physiological, metabolic, and immunological status of the host. Currently, little is known about the composition and role of these communities, whether they are species and/or tissue specific and whether they reflect their surrounding environment. Co-culture of fish, a common practice in semi-intensive aquaculture, where different fish species cohabit in the same contained environment, is an easily accessible and informative model toward understanding such interactions. This study provides the first in-depth characterization of gill and skin microbiomes in co-cultured Nile tilapia (Oreochromis niloticus) and grey mullet (Mugil capito) in semi-intensive pond systems in Egypt using 16S rRNA gene-based amplicon sequencing. Results showed that the microbiome composition of the external surfaces of both species and pond water was dominated by the following bacterial phyla: Proteobacteria, Fusobacteriota, Firmicutes, Planctomycetota, Verrucomicrobiota, Bacteroidota, and Actinobacteriota. However, water microbial communities had the highest abundance and richness and significantly diverged from the external microbiome of both species; thus, the external autochthonous communities are not a passive reflection of their allochthonous communities. The autochthonous bacterial communities of the skin were distinct from those of the gill in both species, indicating that the external microbiome is likely organ specific. However, gill autochthonous communities were clearly species specific, whereas skin communities showed higher commonalities between both species. Core microbiome analysis identified the presence of shared core taxa between both species and pond water in addition to organ-specific taxa within and between the core community of each species. These core taxa included possibly beneficial genera such as Uncultured Pirellulaceae, Exiguobacterium, and Cetobacterium and opportunistic potential pathogens such as Aeromonas, Plesiomonas, and Vibrio. This study provides the first in-depth mapping of bacterial communities in this semi-intensive system that in turn provides a foundation for further studies toward enhancing the health and welfare of these cultured fish and ensuring sustainability

Optimization of Low-Biomass Sample Collection and Quantitative PCR-Based Titration Impact 16S rRNA Microbiome Resolution

The major aquatic interface between host and environment in teleost finfish species is the gill. The diversity of this infraclass, high complexity of the organ, and its direct exposure to the surrounding environment make it an ideal candidate for furthering our understanding of the intertwined relationships between host and microbiome. Capturing the structure and diversity of bacterial communities from this low-biomass, inhibitor-rich tissue can, however, prove challenging. Lessons learned in doing so are directly applicable to similar sample types in other areas of microbiology. Through the development of a quantitative PCR assay for both host material and 16S rRNA genes, we tested and developed a robust method for low-biomass sample collection which minimized host DNA contamination. Quantification of 16S rRNA facilitated not only the screening of samples prior to costly library construction and sequencing but also the production of equicopy libraries based on 16S rRNA gene copies. A significant increase in diversity of bacteria captured was achieved, providing greater information on the true structure of the microbial community. Such findings offer important information for determining functional processes. Results were confirmed across fresh, brackish, and marine environs with four different fish species, with results showing broad homology between samples, demonstrating the robustness of the approach. Evidence presented is widely applicable to samples similar in composition, such as sputum or mucus, or those that are challenging due to the inherent inclusion of inhibitors. IMPORTANCE The interaction between the fish gill and surrounding bacteria-rich water provides an intriguing model for examining the interaction between the fish, free-floating bacteria, and the bacterial microbiome on the gill surface. Samples that are inherently low in bacteria, or that have components that inhibit the ability to produce libraries that identify the components of microbial communities, present significant challenges. Gill samples present both of these types of challenges. We developed methods for quantifying both the bacterial and host DNA material and established a sampling method which both reduced inhibitor content and maximized bacterial diversity. By quantifying and normalizing bacteria prior to library construction, we showed significant improvements with regards to the fidelity of the final data. Our results support wide-ranging applications for analyzing samples of similar composition, such as mucus and sputum, in other microbiological spheres

Exploring the impact of thermal delousing on gill health and microbiome dynamics in farmed Atlantic Salmon

The economic cost of salmon louse (Lepeophtheirus salmonis and Caligus rogercresseyi) infestations in the Atlantic salmon (Salmo salar) industry has been estimated to be around $900 million annually. This high cost has driven a concerted effort to develop, examine, understand, and implement various methods for louse control. Husbandry interventions utilising warm water exposure have been highly successful in complementing traditional chemotherapeutants, especially as the efficiency of the latter has reduced in recent years. In this study, we sought to examine the impact of thermal delousing on gill health in two commercial sites with different historical husbandry and treatment interventions prior to and post-thermal treatment. Methods to characterise gill health and the response to thermal treatment included a detailed examination of the gill for microparasites using both histology and qPCR and targeted immune gene expression analysis, most notably antigen-presenting cells (mhc ii), proinflammatory cytokines (il-1β and tnf-α) and inhibitory cytokines (tgf-β and il-10). Furthermore, we examined the bacterial communities present on the gill surface using 16S rRNA amplicon sequencing. Data obtained from these trials indicated a minimal impact on gill microparasite prevalence in response to the thermal treatment. The expression of immune markers exhibited a significant decrease across both sites after treatment. Intriguingly, marked differences in the gill bacterial communities in response to treatment between the two sites were clearly observed. This divergence could be attributed to the notable differences in husbandry history and health status of the fish at the two sites prior to the thermal treatment. Our data suggest that microbiome diversity is an informative indicator of fish gill health and could be used to define appropriate interventions when treating sea lice

Net cleaning impacts Atlantic salmon gill health through microbiome dysbiosis

Introduction: Net biofouling has a significant impact for the global salmon industry in the seawater grow-out stage in terms of its management. Current mitigation strategies occur primarily through the regular removal of biofouling using in situ cleaning. While in situ net cleaning is effective there is uncertainty as to whether the equipment or dispersed material has an impact upon the fish in the cages. Through direct contact with the environment, the significant surface area of the gill including its microbiome is directly exposed to the acute environmental changes generated by net cleaning. This study aimed to provide a detailed understanding of the impact of in situ net cleaning on Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss) gill health. Methods: Three field trials were conducted on commercial fish farms in western Norway. Fouling organisms on net pens and flushed particles during in situ cleaning were identified and screened for major fish pathogens. Hydrographic profile measurements were performed to measure the impact on water quality. Gill samples were examined for histopathological changes, immune gene expression, and the prevalence of major pathogens. 16S rRNA amplicon sequencing was employed to explore the impact of net cleaning on gill microbiome. Results and discussion: Data obtained from these trials identified a diversity of fouling species including hydroids, algae, skeleton shrimps, and filter feeders on net pens, a direct impact on measured water quality indicators, a moderate change in gill inflammatory and antigen presentation activity at the level of mRNA, and a large significant change in gill microbiome. Observed changes in gill microbial community involved a decrease in bacterial richness coupled to an increase in identified bacterial genera related to negative health consequences. Parallel analyses for pathogens load in biofouling organisms and flushed particles highlighted the presence of several fish bacteria and parasites. However, minor changes were detected in salmon gill pathogen diversity and loading. Our results suggest that biofouling organisms may act as transient reservoirs for some fish pathogens but not viruses and that gill microbial dysbiosis could be related to the host stress response during and post net cleaning

Impact of lighting conditions on the developmental physiology of Atlantic salmon (Salmo salar)

The Atlantic salmon (Salmo salar) lifecycle is punctuated by distinct ontogenic stages which are routinely manipulated commercially by photoperiod regimes to enable year-round production. As such, light plays a critical role throughout the production cycle, however, it remains poorly characterised and light spectrum and intensity have not been defined optimally yet. This thesis was therefore set out to test the effects of narrow bandwidth light (Blue-λ(max) 444 nm, Green-λ(max) 523 nm, Red-λ(max) 632 nm and White) and intensity in freshwater (FW). Fry-parr development, out-of-season smoltification and ocular and vertebral health were examined as was the long-term effects of FW light regimes on seawater (SW) growth and muscle structure. In addition, the impact of photoperiod regimes on out-of-season smolts following transfer to SW was investigated. Major findings from the trials conducted show that light spectrum and intensity influence parr development with lower intensities performing better than higher intensities. Both the initiation and duration of smoltification was impacted by spectrum. Importantly, this doctoral work showed that daily changes in light intensity, from low during the scotophase to high during the photophase applied for the duration of a standard out-of-season smoltification regime was capable of providing a sufficient cue for the induction of smoltification. Historic FW light exposure impacted SW performance and post-transfer SW photoperiod had significant impact upon growth and maturation development. Results based on changes to the gonadosomatic index provide important guidance for suitable post-transfer photoperiods for smolt transferred to SW around the winter solstice. Importantly, from the parameters tested, exposure to different spectrum or light intensities did not adversely affect vertebral or ocular health. This thesis did not only focus on the physiological effects of light but also aimed to characterise better the pathways involved in light perception and integration. To do so, the neural response to both broad spectrum white light, darkness and Red and Blue light was investigated through deep brain insitu-hybridisation and high throughput sequencing (NGS) of the pituitary gland. Results showed substantial spectral and light/ dark changes in the both the deep brain and pituitary transcriptome. Overall, this research provides both scientifically interesting and commercially relevant guidance for the optimisation of lighting systems for use in captive salmon aquaculture.Major findings from the trials conducted show that light spectrum and intensity influence parr development with lower intensities performing better than higher intensities. Both the initiation and duration of smoltification was impacted by spectrum. Importantly, this doctoral work showed that daily changes in light intensity, from low during the scotophase to high during the photophase applied for the duration of a standard out-of-season smoltification regime was capable of providing a sufficient cue for the induction of smoltification. Historic FW light exposure impacted SW performance and post-transfer SW photoperiod had significant impact upon growth and maturation development. Results based on changes to the gonadosomatic index provide important guidance for suitable post-transfer photoperiods for smolt transferred to SW around the winter solstice. Importantly, from the parameters tested, exposure to different spectrum or light intensities did not adversely affect vertebral or ocular health. This thesis did not only focus on the physiological effects of light but also aimed to characterise better the pathways involved in light perception and integration. To do so, the neural response to both broad spectrum white light, darkness and Red and Blue light was investigated through deep brain insitu-hybridisation and high throughput sequencing (NGS) of the pituitary gland. Results showed substantial spectral and light/ dark changes in the both the deep brain and pituitary transcriptome. Overall, this research provides both scientifically interesting and commercially relevant guidance for the optimisation of lighting systems for use in captive salmon aquaculture