Shining light on the transcriptome: Molecular regulatory networks leading to a fast-growth phenotype by continuous light in an environmentally sensitive teleost (Atherinopsidae).

Photoperiod can profoundly affect the physiology of teleost fish, including accelerated growth here defined as "fast growth phenotypes". However, molecular regulatory networks (MRNs) and biological processes being affected by continuous illumination and which allow some teleost species evident plasticity to thrive under this condition are not yet clear. Therefore, to provide a broad perspective of such mechanisms, Chirostoma estor fish were raised and sampled for growth under a simulated control (LD) 12 h Light: 12 h Dark or a continuous illumination (LL) 24 h Light: 0 h Dark since fertilization. The experiment lasted 12 weeks after hatching (wah), the time at which fish were sampled for growth, length, and whole-body cortisol levels. Additionally, 3 heads of fish from each treatment were used to perform a de novo transcriptome analysis using Next-Generation Sequencing. Fish in LL developed the fast growth phenotype with significant differences visible at 4 wah and gained 66% more mass by 12 wah than LD fish. Cortisol levels under LL were below basal levels at all times compared to fish in LD, suggesting circadian dysregulation effects. A strong effect of LL was observed in samples with a generalized down-regulation of genes except for Reactive Oxygen Species responses, genome stability, and growth biological processes. To our knowledge, this work is the first study using a transcriptomic approach to understand environmentally sensitive MRNs that mediate phenotypic plasticity in fish submitted to continuous illumination. This study gives new insights into the plasticity mechanisms of teleost fish under constant illumination.

Loss of gut microbial diversity in the cultured, agastric fish, Mexican pike silverside (Chirostoma estor: Atherinopsidae).

Teleost fish are the most diverse group of extant vertebrates and have varied digestive anatomical structures and strategies, suggesting they also possess an array of different host-microbiota interactions. Differences in fish gut microbiota have been shown to affect host development, the process of gut colonization, and the outcomes of gene-environment or immune system-microbiota interactions. There is generally a lack of studies on the digestive mechanisms and microbiota of agastric short-intestine fish however, meaning that we do not understand how changes in gut microbial diversity might influence the health of these types of fish. To help fill these gaps in knowledge, we decided to study the Mexican pike silverside (Chirostoma estor) which has a simplified alimentary canal (agastric, short-intestine, 0.7 gut relative length) to observe the diversity and metabolic potential of its intestinal microbiota. We characterized gut microbial populations using high-throughput sequencing of the V3 region in bacterial 16S rRNA genes while searching for population shifts resulting associated with fish development in different environments and cultivation methods. Microbiota samples were taken from the digesta, anterior and posterior intestine (the three different intestinal components) of fish that grew wild in a lake, that were cultivated in indoor tanks, or that were raised in outdoor ponds. Gut microbial diversity was significantly higher in wild fish than in cultivated fish, suggesting a loss of diversity when fish are raised in controlled environments. The most abundant phyla observed in these experiments were Firmicutes and Proteobacteria, particularly of the genera Mycoplasma, Staphylococcus, Spiroplasma, and Aeromonas. Of the 14,161 OTUs observed in this experiment, 133 were found in all groups, and 17 of these, belonging to Acinetobacter, Aeromonas, Pseudomonas, and Spiroplasma genera, were found in all samples suggesting the existence of a core C. estor microbiome. Functional metagenomic prediction of bacterial ecological functions using PICRUSt2 suggested that different intestinal components select for functionally distinct microbial populations with variation in pathways related to the metabolism of amino acids, vitamins, cofactors, and energy. Our results provide, for the first time, information on the bacterial populations present in an agastric, short-gut teleost with commercial potential and show that controlled cultivation of this fish reduces the diversity of its intestinal microbiota.

High levels of docosahexaenoic acid are present in eight New World silversides (Pisces: Atherinopsidae)

Docosahexaenoic acid (DHA) is the most critical and least available omega-3 fatty acid in the Western human diet. Currently, the source of omega-3 long chain polyunsaturated fatty acids (LC-PUFA) is mainly dependent on wild fisheries, making this resource unsustainable in the foreseeable future. In recent years, a high rate of biosynthesis and accumulation of DHA has been discovered in a freshwater species (Chirostoma estor) belonging to the Atherinopsidae family. Interest in evaluating fatty acid composition in other members of the family has emerged, so this study compiles original data of flesh composition of eight atherinopsid species from freshwater and brackish environments, either wild or cultured. High levels of DHA (16 to 31%) were found in all analyzed members of the family, except in C. grandocule, independently of their habitat or origin. The analyzed species of the Jordani group (C. estor, C. promelas and C. humboldtianum) showed high DHA and low EPA levels (<0.5%) as previously reported for cultured C. estor. The low trophic niche of these atherinopsids and their fatty acid accumulation capabilities are factors that make these species noteworthy candidates for sustainable aquaculture.(AU)

O ácido docosahexaenóico (DHA) é o ácido graxo ômega-3 mais importante e menos disponível na dieta humana ocidental. Atualmente, a fonte de ácidos graxos poliinsaturados de cadeia longa ômega-3 (LC-PUFA) depende principalmente da pesca extrativista, tornando esse recurso insustentável em um futuro próximo. Nos últimos anos, uma alta taxa de biossíntese e acúmulo de DHA foi descoberta em uma espécie de água doce (Chirostoma estor) pertencente à família Atherinopsidae. Deste modo, surgiu o interesse em avaliar a composição de ácidos graxos em outros membros da família. Portanto, este estudo compila dados originais da composição de carne de oito espécies de aterinopsídeos de ambientes de água doce e salobra, selvagens ou cultivadas. Altos níveis de DHA (16 a 31%) foram encontrados em todos os membros da família analisados, exceto em C. grandocule, independentemente de seu habitat ou origem. As espécies analisadas do grupo Jordani (C. estor, C. promelas e C. humboldtianum) apresentaram altos níveis de DHA e EPA baixos (<0,5%), como relatado anteriormente para C. estor cultivado. O baixo nicho trófico desses aterinopsídeos e sua capacidade de acumulação de ácidos graxos são fatores que tornam essas espécies notáveis candidatas à aquicultura sustentável.(AU)

Experimental evidence of masculinization by continuous illumination in a temperature sex determination teleost (Atherinopsidae) model: is oxidative stress involved?

The present study evaluates the influence of continuous light on phenotypic sex ratios in Chirostoma estor, a temperature sex determination animal model. Relative gene expression levels of 5 day old larvae were performed on two early gonad differentiation genes (sox9 and foxl2), two stress axis activation genes (gcr1 and crf) and four reactive oxygen species (ROS) antagonist effector genes (sod2, ucp2, gsr and cat). Two light treatments were applied from fertilization; control (12L:12D) simulated natural photoperiod and a continuous illumination photoperiod. By the end of the trial (12 weeks after hatching), differentiated and normal gonads were clearly identifiable in both treatments by histological observations. Regarding sex ratio, 73% of phenotypic males were found in continuous illumination compared with 40% in controls. Consistently, the sox9 gene (involved in early testis differentiation) showed an over expression in 64% of the individual larvae analysed compared with foxl2 (ovarian differentiation) suggesting a masculinization tendency in continuous illumination. On the other hand, only 36% of individuals showed the same tendency in the control treatment consistent with phenotypic sex ratios found under normal culture conditions. Relative gene expression results did not show significant difference in sod2, ucp2 and gcr1 levels, but cat, gsr and crf showed significantly higher expression levels in the continuous illumination treatment suggesting that both, the stress axis and ROS response mechanisms were activated at this time. This study suggests, a link between continuous light, oxidative stress and environmental sex determination in vertebrates. However, further research is necessary to describe this possible upstream mechanism that may drive some aspects of sexual plasticity in vertebrates.

3,5-Diiodothyronine-mediated transrepression of the thyroid hormone receptor beta gene in tilapia. Insights on cross-talk between the thyroid hormone and cortisol signaling pathways.

T3 and cortisol activate or repress gene expression in virtually every vertebrate cell mainly by interacting with their nuclear hormone receptors. In contrast to the mechanisms for hormone gene activation, the mechanisms involved in gene repression remain elusive. In teleosts, the thyroid hormone receptor beta gene or thrb produces two isoforms of TRß1 that differ by nine amino acids in the ligand-binding domain of the long-TRß1, whereas the short-TRß1 lacks the insert. Previous reports have shown that the genomic effects exerted by 3,5-T2, a product of T3 outer-ring deiodination, are mediated by the long-TRß1. Furthermore, 3,5-T2 and T3 down-regulate the expression of long-TRß1 and short-TRß1, respectively. In contrast, cortisol has been shown to up-regulate the expression of thrb. To understand the molecular mechanisms for thrb modulation by thyroid hormones and cortisol, we used an in silico approach to identify thyroid- and cortisol-response elements within the proximal promoter of thrb from tilapia. We then characterized the identified response elements by EMSA and correlated our observations with the effects of THs and cortisol upon expression of thrb in tilapia. Our data show that 3,5-T2 represses thrb expression and impairs its up-regulation by cortisol possibly through a transrepression mechanism. We propose that for thrb down-regulation, ligands other than T3 are required to orchestrate the pleiotropic effects of thyroid hormones in vertebrates.

3,5-Diiodothyronine (T2) is on a role. A new hormone in search of recognition.

Thyroid hormone (TH) actions are mediated by triiodothyronine (T3), which acts by binding to the TH receptors (TRs). Since TH exert pleiotropic effects, interest has grown in identifying other possible bioactive thyronines that could explain their diversity of functions. Accordingly, 3,5-diiodothyronine (T2) has been shown to be bioactive. In mammals, T2 regulates mRNA expression of several T3-regulated genes, but doses up to 100-fold greater than those of T3 were required to generate comparable effects. In teleosts, T2 and T3 regulate gene expression in vivo with equivalent potency. Furthermore, in vivo and in vitro studies support the notion that T2 binds to and activates a specific, long TRß1 isoform that contains a nine amino acid insert at the beginning of the ligand binding domain, whereas T3 can interact also with a different TRß1 isoform that lacks this insert. Similarly, T2 and T3 differentially regulate long- and short-TRß1 expression, respectively, strongly suggesting a different signaling pathway for each hormone, at least in the species that express both receptors. In vivo, T2 effectively triggers a burst of body growth in tilapia by interacting with the long TRß1 isoform, supporting the notion that T2 is physiologically relevant in this species. Current knowledge of T2 effects and action mechanisms lead us to propose that there is an extra level in the thyroid hormone signaling cascade, and that T2 is produced and regulated specifically for this purpose.

3,5-di-iodothyronine stimulates tilapia growth through an alternate isoform of thyroid hormone receptor ß1.

Recent studies in our laboratory have shown that in some teleosts, 3,5-di-iodothyronine (T2 or 3,5-T2) is as bioactive as 3,5,3'-tri-iodothyronine (T3) and that its effects are in part mediated by a TRß1 (THRB) isoform that contains a 9-amino acid insert in its ligand-binding domain (long TRß1 (L-TRß1)), whereas T3 binds preferentially to a short TRß1 (S-TRß1) isoform that lacks this insert. To further understand the functional relevance of T2 bioactivity and its mechanism of action, we used in vivo and ex vivo (organotypic liver cultures) approaches and analyzed whether T3 and T2 differentially regulate the S-TRß1 and L-TRß1s during a physiological demand such as growth. In vivo, T3 and T2 treatment induced body weight gain in tilapia. The expression of L-TRß1 and S-TRß1 was specifically regulated by T2 and T3 respectively both in vivo and ex vivo. The TR antagonist 1-850 effectively blocked thyroid hormone-dependent gene expression; however, T3 or T2 reversed 1-850 effects only on S-TRß1 or L-TRß1 expression, respectively. Together, our results support the notion that both T3 and T2 participate in the growth process; however, their effects are mediated by different, specific TRß1 isoforms.