Transcriptome analyses reveal key roles of alternative splicing regulation in atlantic salmon during the infectious process of Piscirickettsiosis disease.

In the Chilean salmon farming industry, infection by Piscirickettsia salmonis is the primary cause of the main bacterial disease known as Piscirickettsiosis, which has an overwhelming economic impact. Although it has been demonstrated that Piscirickettsiosis modifies the expression of numerous salmonids genes, it is yet unknown how alternative splicing (AS) contributes to salmonids bacterial infection. AS, has the potential to create heterogeneity at the protein and RNA levels and has been associated as a relevant molecular mechanism in the immune response of eukaryotes to several diseases. In this study, we used RNA data to survey P. salmonis-induced modifications in the AS of Atlantic salmon and found that P. salmonis infection promoted a substantial number (158,668) of AS events. Differentially spliced genes (DSG) sensitive to Piscirickettsiosis were predominantly enriched in genes involved in RNA processing, splicing and spliceosome processes (e.g., hnRNPm, hnRPc, SRSF7, SRSF45), whereas among the DSG of resistant and susceptible to Piscirickettsiosis, several metabolic and immune processes were found, most notably associated to the regulation of GTPase, lysosome and telomere organization-maintenance. Furthermore, we found that DSG were mostly not differentially expressed (5-7 %) and were implicated in distinct biological pathways. Therefore, our results underpin AS achieving a significant regulatory performance in the response of salmonids to Piscirickettsiosis.

Attention Deficit-Hyperactivity Disorder (ADHD): From Abnormal Behavior to Impairment in Synaptic Plasticity.

Attention deficit-hyperactivity disorder (ADHD) is a neurodevelopmental disorder with high incidence in children and adolescents characterized by motor hyperactivity, impulsivity, and inattention. Magnetic resonance imaging (MRI) has revealed that neuroanatomical abnormalities such as the volume reduction in the neocortex and hippocampus are shared by several neuropsychiatric diseases such as schizophrenia, autism spectrum disorder and ADHD. Furthermore, the abnormal development and postnatal pruning of dendritic spines of neocortical neurons in schizophrenia, autism spectrum disorder and intellectual disability are well documented. Dendritic spines are dynamic structures exhibiting Hebbian and homeostatic plasticity that triggers intracellular cascades involving glutamate receptors, calcium influx and remodeling of the F-actin network. The long-term potentiation (LTP)-induced insertion of postsynaptic glutamate receptors is associated with the enlargement of spine heads and long-term depression (LTD) with spine shrinkage. Using a murine model of ADHD, a delay in dendritic spines' maturation in CA1 hippocampal neurons correlated with impaired working memory and hippocampal LTP has recently reported. The aim of this review is to summarize recent evidence that has emerged from studies focused on the neuroanatomical and genetic features found in ADHD patients as well as reports from animal models describing the molecular structure and remodeling of dendritic spines.

Resultados de una evaluación y seguimiento online de problemas de salud mental en universitarios chilenos / Results of an online mental health problems screening and follow-up with Chilean university students

Resumen Este estudio examina la detección y posterior seguimiento online de problemas de salud mental en estudiantes universitarios y expone los resultados de ambos procesos. Para ello, se aplicó a través de la plataforma de la universidad a 1 707 estudiantes una batería de cuestionarios destinados a detectar desesperanza y riesgo de suicidio, síntomas depresivos y ansiosos, estrés, trastornos de la conducta alimentaria y apoyo social percibido. Una vez aplicados los instrumentos, los estudiantes recibían una retroalimentación acerca de sus niveles de síntomas e información acerca de redes y establecimientos para buscar ayuda y recibir atención en los casos requeridos. A mediados del año siguiente, se realizó una encuesta de seguimiento a 577 de los estudiantes que previamente habían respondido todos los cuestionarios. Los resultados de la batería de cuestionarios aplicados indican altos índices de problemas de salud mental, con diferencias estadísticamente significativas según sexo, siendo las mujeres quienes presentan mayor prevalencia. Estos resultados concuerdan con diversos estudios que muestran altos porcentajes de síntomas de problemas de salud mental en los universitarios. A su vez, el posterior seguimiento permitió evidenciar que el feedback entregado activó la búsqueda de ayuda, y se obtuvieron correlaciones positivas entre sentirse mejor y haber buscado un tratamiento de salud mental o haber pedido ayuda a los amigos o a la familia. El uso de tecnologías de la información en la detección y prevención de salud mental tendría un alcance positivo y resultó ser una herramienta útil, accesible y cercana para los jóvenes.

Abstract The exponential increase in higher education students and the new university profile that incorporates more vulnerable social groups are associated with an increase in mental health problems in this population. Likewise, the complex stage of the life cycle in which these young people find themselves and the stress of the transition between adolescence and adulthood are other risk factors for the development of mental health problems. The emergence of mental health problems in this group could then be due to a confluence of factors, also facilitating risk behaviors. National and international researches have reported the worrying rates of mental health disorders in university students, mainly in relation to the presence of depression, anxiety, stress, alcohol or drug use, eating disorders, and suicidal ideation. These disorders tend to be aggravated, because the university population is prone not to seek help. The stigma of presenting mental health problems, skepticism about treatments, not wanting to worry the closest family and the few economic resources or not having a health plan, lead to not seeking help or do this late. Studies have found that more than half of students who need mental health and support services are not receiving them. On the other hand, a high percentage seeks help late, which implies that the disorders are accentuated or become chronic, which makes subsequent treatment and improvement difficult, even becoming a public health problem due to the high economic and professional demand that these treatments require.

Unveiling the Role of Dynamic Alternative Splicing Modulation After Infestation with Sea Lice (Caligus rogercresseyi) in Atlantic Salmon.

Sea lice are pathogenic marine ectoparasite copepods that represent a severe risk to the worldwide salmon industry. Several transcriptomic investigations have characterized the regulation of gene expression response of Atlantic salmon to sea lice infestation. These studies have focused on the levels of transcript, overlooking the potentially relevant role of alternative splicing (AS), which corresponds to an essential control mechanism of gene expression through RNA processing. In the present study, we performed a genome-wide bioinformatics characterization of differential AS event dynamics in control and infested C. rogercresseyi Atlantic salmon and in resistant and susceptible phenotypes. We identified a significant rise of alternative splicing events and AS genes after infestation and 176 differential alternative splicing events (DASE) from 133 genes. In addition, a higher number of DASE and AS genes were observed among resistant and susceptible phenotypes. Functional annotation of AS genes shows several terms and pathways associated with behavior, RNA splicing, immune response, and RNA binding. Furthermore, three protein-coding genes were identified undergoing differential transcript usage events, among resistant and susceptible phenotypes. Our findings support AS performing a relevant regulatory role in the response of salmonids to sea lice infestation.

Phytochemical Screening and Antioxidant Activity of Seven Native Species Growing in the Forests of Southern Chilean Patagonia.

The genus Nothofagus is one of the most abundant in the subantarctic Patagonian forests. Five species inhabit these ecosystems, three evergreen (Nothofagus betuloides, Nothofagus dombeyi, and Nothofagus nitida) and two deciduous (Nothofagus pumilio and Nothofagus antarctica). This is the first report on the levels of secondary metabolites and the antioxidant capacity of Patagonian tree species growing in natural environments. The aim of this work was to carry out a phytochemical screening, to determine the antioxidant capacity, the sun protection factor, and the α-glucosidase and tyrosinase inhibitory activity of foliar extracts of the five previous species. Besides, Aristotelia chilensis and Berberis microphylla, two species of Patagonian shrubs growing in the same forests, were used as reference. N. dombeyi was the Nothofagus with the best antioxidant capacity. B. microphylla differed from all studied species. Moreover, the Nothofagus was split into two groups. N. betuloides and N. dombeyi are the most similar species to A. chilensis. The α-glucosidase was completely inhibited by all studied extracts. Furthermore, N. antarctica, N.pumilio, and N. nitida inhibited about 70% of the tyrosinase activity. All the results found in this study for the species of the genus Nothofagus support further research on their potential beneficial properties for human health.

Temporal Gene Expression Signature of Plasma Extracellular Vesicles-MicroRNAs from Post-Smolt Coho Salmon Challenged with Piscirickettsia salmonis.

Piscirickettsiosis is the most important bacterial disease in the Chilean salmon industry, which has borne major economic losses due to failure to control it. Cells use extracellular vesicles (EVs) as an inter-cellular communicators to deliver several factors (e.g., microRNAs) that may regulate the responses of other cells. However, there is limited knowledge about the identification and characterization of EV-miRNAs in salmonids or the effect of infections on these. In this study, Illumina sequencing technology was used to identify Coho salmon plasma EV-miRNAs upon Piscirickettsia salmonis infection at four different time points. A total of 118 novels and 188 known EV-miRNAs, including key immune teleost miRNAs families (e.g., miR-146, miR-122), were identified. A total of 245 EV-miRNAs were detected as differentially expressed (FDR < 5%) in terms of control, with a clear down-regulation pattern throughout the disease. KEGG enrichment results of EV-miRNAs target genes showed that they were grouped mainly in cellular, stress, inflammation and immune responses. Therefore, it is hypothesized that P. salmonis could potentially benefit from unbalanced modulation response of Coho salmon EV-miRNAs in order to promote a hyper-inflammatory and compromised immune response through the suppression of different key immune host miRNAs during the course of the infection, as indicated by the results of this study.

Vitellogenesis in the Patagonian toothfish (Dissostichus eleginoides) conditioned to a recirculating aquaculture system.

The Patagonian toothfish (Dissostichus eleginoides) is a new promising fish species for diversifying the aquaculture industry in Chile because of its high economic value and high international demand. However, when attempting to start aquaculture of a new species, one of the major challenges is successfully achieving conditions to reproduce them. This is particularly difficult when the information on the biology and physiology of the reproduction process of the species in question is scarce, as is the case with D. eleginoides. Additionally, female reproductive dysfunction is more prevalent under culture conditions and it is very important to have tools to evaluate the progress of oocyte maturation. Therefore, evaluation of the vitellogenesis process in addition to measuring gonadosomatic index (GSI) and oocyte diameter is an important parameter for allowing the monitoring of females from a broodstock that will spawn in the reproductive season. This study aimed to develop an enzyme-linked immunosorbent assay (ELISA) specific for the Patagonian toothfish (D. eleginoides) vitellogenine (Vtg) and quantify the plasma level in the fishes, maintained in a recirculation aquaculture system (RAS), throughout their reproductive cycle. A polyclonal antibody was prepared using the isolated major egg protein as antigen. This antibody was specific to the major plasma phosphoprotein identified as Vtg and was used to develop and standardize an indirect ELISA assay. The assay standard curve was linear from 0.1 to 1 µg/ml purified egg yolk protein and the average r2 was 0.995. We corroborated our ELISA assay by demonstrating a strong correlation between high levels of plasma Vtg obtained by the assay and the intensity of the corresponding bands in both SDS-PAGE coomassie stained gels and Western Blot. During the two reproductive seasons analyzed, the highest Vtg plasma level was obtained in the majority of the females in the last three months before spawning (December-January). This differs from the wild population in which the spawning occurs during the austral winter (June-September). Therefore, the RAS condition established to maintain in captivity the D. eleginoides allows females to develop mature oocytes normally, as was evidenced by picks of Vtg plasma levels.

Temporal genome-wide DNA methylation signature of post-smolt Pacific salmon challenged with Piscirickettsia salmonis.

Piscirickettsiosis is the most important bacterial disease in the Chilean salmon industry, which has sorted several efforts to its control, generating enormous economic losses. Epigenetic alterations, such as DNA methylation, can play a relevant role in the modulation of the metazoans response to pathogens. Bacterial disease may activate global and local immune responses generating intricate responses with significant biological impact in the host. However, it is scarcely understood how bacterial infections influence fish epigenetic alterations. In the present study, we utilized Pacific salmon and Piscirickettsiosis as model, to gain understanding into the dynamics of DNA methylation among fish-bacterial infection interactions. A genome-wide analysis of DNA methylation patterns in female spleen tissue of Pacific salmon was achieved by reduced representation bisulphite sequencing from a time course design. We determined 2,251, 1,918, and 2,516 differentially methylated regions DMRs among infected and control Pacific salmon in 1 dpi, 5 dpi, and 15 dpi, respectively. The mean methylation difference per DMR among control and infected groups was of ~35%, with an oscillatory pattern of hypo, hyper, and hypomethylation across the disease. DMCs, among the control and infected group, showed that they were statistically enriched in intergenic regions and depleted in exons. Functional annotation of the DMR genes demonstrated three KEGG principal categories, associated directly with the host response to pathogens infections. Our results provide the first evidence of epigenetic variation in fish provoked by bacterial infection and demonstrate that this variation can be modulated across the disease.

Identification and characterization of miRNAs and lncRNAs of coho salmon (Oncorhynchus kisutch) in normal immune organs.

MicroRNAs (miRNAs) and long noncoding RNAs (lncRNAs) are two relevant non-coding RNAs (ncRNAs) class. Oncorhynchus kisutch (coho salmon) is an important aquaculture pacific salmon species without report of miRNAs and a very limited register of lncRNAs. To gain knowledge about the interaction and discovery of miRNAs and lncRNAs in coho salmon we used high-throughput sequencing technology to sequence small and transcriptome libraries from three immune organs. A total of 163 mature miRNAs and 4,975 lncRNAs were discovered. The profiles of expression of both ncRNAs indicated that liver and head-kidney share relatively similar expression patterns. We identified 814 and 181 putative target sequences for 1048 lncRNAs and 47 miRNAs, respectively. The results obtained provide new information and enlarge our understanding of the diversities of ncRNAs in coho salmon.

Novel microsatellite markers discovery in Patagonian toothfish (Dissostichus eleginoides) using high-throughput sequencing.

Patagonian toohfish (Dissostichus eleginoides), is a sub Antartic notothenioid fish key in the marine ecosystem that sustains fishery of higher commercial value in the world. However, there are a scarce knowledge or information about its population genetic background, product of the almost null information of molecular markers available for this species. Here, we use high-throughput sequencing technology (Illumina platform) to develop 1071 microsatellite loci, of which 22 loci were selected to evaluation. Polymorphism and genetic diversity of each locus was assessed in two locations distant by 2370 km. Considering both locations, a mean PIC value of 0.748 was estimated. Selected microsatellite loci showed among two to seventeen alleles by locus in the first location and two to twelve in the second. The observed heterozygosity varied from 0.18 to 0.91 and from 0.12 to 0.87 for the first and second location, respectively. While, the expected heterozygosity ranged from 0.15 to 0.92 and from 0.11 to 0.90. Three loci were monomorphic in only one location. Microsatellite markers developed here will be useful in future studies on conservation, fishery and population genetics of this species.

Metal concentrations and source identification in Chilean public children's playgrounds.

This study is focused on four Biobio region cities, Concepcion, Talcahuano, Los Ángeles, and Tomé; these cities flourished very close to different industrial activities. We determined a pseudo total concentration of seven heavy metals (Cr, Ni, Cu, Zn, As, Cd, and Pb) in playground soils through inductively coupled plasma-mass spectrometry (ICP-MS). Principal components analysis (PCA) revealed that contamination in soils comes from three principal sources. Firstly, industrial and burning activities; secondly, the use of phytosanitary and chemical products; and thirdly, vehicular traffic emissions. Zn and Cu are the most abundant analyzed elements in all the playground's soils. Concepción reflected the lowest values of pollutants and Talcahuano the highest, reflecting the industrial effects. The average values of the analyzed elements were Cr = 32.90 mg kg-1; Ni = 23.76 mg kg-1; Cu = 31.51 mg kg-1; Zn = 63.69 mg kg-1; As = 19.51 mg kg-1; Cd = 0.50 mg kg-1; and Pb = 17.59 mg kg-1. Anomalously high values of some elements were found Cu = 462.73 mg kg-1, Zn = 364.39 mg kg-1, As = 34.7 mg kg-1 in Talcahuano, Cd = 1.6 mg kg-1 in Tome, and Pb = 55.59 mg kg-1 in Los Ángeles. Nevertheless, according to international guideline values of pollutants (VROM 2000 and ADEC 2010) there is no risk for children in any playground studied but all playgrounds are a potential risk for the environment. It points out the necessity to continue studying and monitoring Chilean urban playground to prevent health problems in the population. Graphical abstract.

Author Correction: Selection of reference genes for microRNA analysis associated to early stress response to handling and confinement in Salmo salar.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

Functional Annotation of All Salmonid Genomes (FAASG): an international initiative supporting future salmonid research, conservation and aquaculture.

We describe an emerging initiative - the 'Functional Annotation of All Salmonid Genomes' (FAASG), which will leverage the extensive trait diversity that has evolved since a whole genome duplication event in the salmonid ancestor, to develop an integrative understanding of the functional genomic basis of phenotypic variation. The outcomes of FAASG will have diverse applications, ranging from improved understanding of genome evolution, to improving the efficiency and sustainability of aquaculture production, supporting the future of fundamental and applied research in an iconic fish lineage of major societal importance.

Selection of reference genes for microRNA analysis associated to early stress response to handling and confinement in Salmo salar.

MicroRNAs are key non-coding RNA molecules that play a relevant role in the regulation of gene expression through translational repression and/or transcript cleavage during normal development and physiological adaptation processes like stress. Quantitative reverse transcription polymerase chain reaction (RT-qPCR) has become the approach normally used to determine the levels of microRNAs. However, this approach needs the use of endogenous reference. An improper selection of endogenous references can result in confusing interpretation of data. The aim of this study was to identify and validate appropriate endogenous reference miRNA genes for normalizing RT-qPCR survey of miRNAs expression in four different tissues of Atlantic salmon, under handling and confinement stress conditions associated to early or primary stress response. Nine candidate reference normalizers, including microRNAs and nuclear genes, normally used in vertebrate microRNA expression studies were selected from literature, validated by RT-qPCR and analyzed by the algorithms geNorm and NormFinder. The results revealed that the ssa-miR-99-5p gene was the most stable overall and that ssa-miR-99-5p and ssa-miR-23a-5p genes were the best combination. Moreover, the suitability of ssa-miR-99-5p and ssa-miR-23a-5p as endogeneuos reference genes was demostrated by the expression analysis of ssa-miR-193-5p gene.

The Atlantic salmon genome provides insights into rediploidization.

The whole-genome duplication 80 million years ago of the common ancestor of salmonids (salmonid-specific fourth vertebrate whole-genome duplication, Ss4R) provides unique opportunities to learn about the evolutionary fate of a duplicated vertebrate genome in 70 extant lineages. Here we present a high-quality genome assembly for Atlantic salmon (Salmo salar), and show that large genomic reorganizations, coinciding with bursts of transposon-mediated repeat expansions, were crucial for the post-Ss4R rediploidization process. Comparisons of duplicate gene expression patterns across a wide range of tissues with orthologous genes from a pre-Ss4R outgroup unexpectedly demonstrate far more instances of neofunctionalization than subfunctionalization. Surprisingly, we find that genes that were retained as duplicates after the teleost-specific whole-genome duplication 320 million years ago were not more likely to be retained after the Ss4R, and that the duplicate retention was not influenced to a great extent by the nature of the predicted protein interactions of the gene products. Finally, we demonstrate that the Atlantic salmon assembly can serve as a reference sequence for the study of other salmonids for a range of purposes.

Ssa miRNAs DB: Online repository of in silico predicted miRNAs in Salmo salar.

UNLABELLED: The Atlantic salmon (Salmo salar) is a very valuable commercial salmonid species. As with other aquaculture species, intensive aquaculture of Atlantic salmon often faces disease problems especially in early life stages which can limit stable production of the species. 'Ssa miRNAs DB', a bioinformatics and manually curated database, aims at providing a comprehensive resource of microRNA in Altantic salmon, with a user friendly interface for a convenient retrieval of each entry by microRNA ID or target gene. The current version of Ssa miRNAs DB involved the prediction of 41 and 266 homologous and novel microRNAs, respectively. AVAILABILITY: The database is available for free at http://www.molgenv.com/ssa_mirnas_db_home.php.

Sequencing the genome of the Atlantic salmon (Salmo salar).

The International Collaboration to Sequence the Atlantic Salmon Genome (ICSASG) will produce a genome sequence that identifies and physically maps all genes in the Atlantic salmon genome and acts as a reference sequence for other salmonids.

Characterization of new microsatellite markers derived from sequence databases for the emu (Dromaius novaehollandiae).

The emu (Dromaius novaehollandiae), a member of ratite family, is native to Australia and has been introduced to other countries worldwide. In this work, 10 polymorphic microsatellite loci were isolated and characterized for emu from public sequences. Polymorphism was surveyed in 22 individuals from two different populations kept in captivity. Between two and 11 alleles were found per locus, and the observed heterozygosity ranged from 0.05 to 0.85, in accordance with expectations. These markers will be useful as tools for detecting levels of genetic variation, reconstructing pedigrees (for quantitative genetic analysis) and identifying markers associated to fitness traits in emu populations.

Phylogeny, biogeography, and rates of diversification of New World Astragalus (Leguminosae) with an emphasis on South American radiations.

This study uses phylogenetic relationships of New World representatives of the species-rich genus Astragalus (Leguminosae; Papilionoideae) to follow up on recent evidence pointing to rapid and recent plant diversification patterns in the Andes. Bayesian and maximum likelihood phylogenetic analyses were done using nuclear rDNA ITS and chloroplast spacers trnD-trnT and trnfM-trnS1, either separately or in combination. The effect of using partitioned vs. nonpartitioned analyses in a Bayesian approach was evaluated. Highest resolution was obtained when the data were combined in partitioned or nonpartitioned Bayesian analyses. All phylogenies support two clades of South American species nested within the North American species, implying two separate invasions from North to South America. These two clades correspond to the original morphological classification of Johnston (1947 Journal of the Arnold Arboretum 28: 336-409). The mean ages of the South American clades were very recent but still significantly different (1.89 and 0.98 Ma). Upper and lower bounds on rates of diversification varied between 2.01 and 0.65 species/Ma for the older clade and 2.06 and 1.24 species/Ma for the younger clade. Even the lower bounds are still very high, reasserting Neo-Astragalus in the growing list of recent rapid radiations of plants, especially in areas with a high physiographic diversity, such as the Andes.

PHYLOGEOGRAPHY OF THE GENUS SPONGITES (CORALLINALES, RHODOPHYTA) FROM CHILE(1).

Both the records and the descriptions of the crustose species of coralline algae on the southeastern coast of South America are from the early 1900s. Unlike other algae species on the coast of Chile, the biogeography and distribution of crustose corallines have not been studied despite their abundance. Through recent studies, it has been determined that the genus Spongites is the most conspicuous genus along the rocky intertidal of the Chilean coasts. It is also common to the entire coast of the Southern Hemisphere; however, the relationship between species and the possible reasons for their distribution is unknown. We used nuclear and mitochondrial genetic markers and SEM observations of morphological characters to examine Spongites samples from the Southern Hemisphere and to establish the phylogeographic relationships of Chilean Spongites with specimens from other southern coasts. The combination of these analyses revealed the following: (i) a monophyletic clade that represents the Chilean Spongites and (ii) a paraphyletic clade for South African, New Zealand, and Argentine samples. Consequently, we postulate two nonexclusive hypotheses regarding the relationship of Spongites species in the Southern Hemisphere: (i) a complex history of extinction, speciation, and recolonization that might have erased original Gondwanan split patterns, and (ii) an Antarctic Peninsula origin for the Chilean Spongites species.