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The heat waves on the South Pacific coast could lead to thermal stress in native fish. The red cusk-eel (Genypterus chilensis) is relevant for Chilean artisanal fisheries and aquaculture diversification. This study examined the effect of high-temperature stress in the gills of G. chilensis in control (14 °C) and high-temperature stress (19 °C) conditions. High-temperature stress induces a significant increase in gills cortisol levels. Additionally, oxidative damage was observed in gills (protein carbonylation and lipoperoxidation). RNA-seq data was used to build the first transcriptome assembly of gills in this species (23,656 annotated transcripts). A total of 1138 down-regulated and 1531 up-regulated transcripts were observed in response to high-temperature stress in gills. The enrichment analysis showed immune response and replication enriched processes (on down-regulated transcripts), and processes related to the folding of proteins, endoplasmic reticulum, and transporter activity (on up-regulated transcripts). The present study showed how gills could be affected by high-temperature stress.
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Gadiformes , Branquias , Animales , Peces , Transcriptoma , Estrés Oxidativo , Anguilas/genética , InmunidadRESUMEN
The genes encoding for estrogen receptor (ESR2) and follicle-stimulating hormone receptor (FSHR) play crucial roles in ovarian follicular development. This study aimed to determine the expression levels of miRNAs predicted against FSHR and ESR2 mRNAs in follicular cells related to their target genes during the estrous cycle in canines. Antral follicles were dissected from 72 ovaries following ovariohysterectomies. MiRNAs regulating FSHR and ESR2 genes were selected from miRNA databases, and mature miRNA and mRNA expression profiling was performed using real-time polymerase chain reaction (PCR). The best miRNA for each target gene was selected considering the quantitative PCR (qPCR) performance and target prediction probability, selecting only miRNAs with a binding p-value of 1.0, and choosing cfa-miR-34a and cfa-let-7c for FSHR and ESR2, respectively. The expression levels comparing the different phases of the estrous cycle were evaluated using ANOVA. Pearson correlations between the expression pattern of each miRNA and their target genes were performed. Each miRNA and its target genes were expressed in the granulosa cells in all estrous phases. FSHR remained low in anestrus and proestrus, increased (p < 0.05) to the highest level in estrus, and decreased (p < 0.05) in diestrus. ESR2 showed the same trend as FSHR, with the highest (p < 0.05) expression in estrus and the lowest (p < 0.05) in anestrus and proestrus. A tendency for an inverse relationship was observed between the expression of miR-34a and FSHR only in the anestrus phase, while an inverse correlation (r = -0.8) was found between miRNA-7c and ESR2 (p < 0.01). The expression profile of miR-34a and miR-let-7c and their predicted target genes of dog ovarian follicles throughout the estrous cycle observed in this study suggest a role in the transcriptional regulation of FSHR and ESR2, which is the first evidence of the involvement of these miRNAs in the canine follicular function.
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The salmonid rickettsial syndrome (SRS) is a systemic bacterial infection caused by Piscirickettsia salmonis that generates significant economic losses in Atlantic salmon (Salmo salar) aquaculture. Despite this disease's relevance, the mechanisms involved in resistance against P. salmonis infection are not entirely understood. Thus, we aimed at studying the pathways explaining SRS resistance using different approaches. First, we determined the heritability using pedigree data from a challenge test. Secondly, a genome-wide association analysis was performed following a complete transcriptomic profile of fish from genetically susceptible and resistant families within the challenge infection with P. salmonis. We found differentially expressed transcripts related to immune response, pathogen recognition, and several new pathways related to extracellular matrix remodelling and intracellular invasion. The resistant background showed a constrained inflammatory response, mediated by the Arp2/3 complex actin cytoskeleton remodelling polymerization pathway, probably leading to bacterial clearance. A series of biomarkers of SRS resistance, such as the beta-enolase (ENO-ß), Tubulin G1 (TUBG1), Plasmin (PLG) and ARP2/3 Complex Subunit 4 (ARPC4) genes showed consistent overexpression in resistant individuals, showing promise as biomarkers for SRS resistance. All these results together with the differential expression of several long non-coding RNAs show the complexity of the host-pathogen interaction of S. salar and P. salmonis. These results provide valuable information on new models describing host-pathogen interaction and its role in SRS resistance.
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Enfermedades de los Peces , Piscirickettsia , Infecciones por Piscirickettsiaceae , Salmo salar , Animales , Salmo salar/genética , Estudio de Asociación del Genoma Completo , Piscirickettsia/fisiología , Transcriptoma , Interacciones Huésped-Patógeno , CitoesqueletoRESUMEN
Environmental stressors, such as temperature, are relevant factors that could generate a negative effect on several tissues in fish. A key fish species for Chilean aquaculture diversification is the red cusk-eel (Genypterus chilensis), a native fish for which knowledge on environmental stressors effects is limited. This study evaluated the effects of high-temperature stress on the liver of red cusk-eel in control (14 °C) and high-temperature (19 °C) groups using multiple approaches: determination of plasmatic hepatic enzymes (ALT, AST, and AP), oxidative damage evaluation (AP sites, lipid peroxidation, and carbonylated proteins), and RNA-seq analysis. High-temperature stress generated a significant increase in hepatic enzyme activity in plasma. In the liver, a transcriptional regulation was observed, with 1239 down-regulated and 1339 up-regulated transcripts. Additionally, high-temperature stress generated oxidative stress in the liver, with oxidative damage and transcriptional modulation of the antioxidant response. Furthermore, an unfolded protein response was observed, with several pathways enriched, as well as a heat shock response, with several heat shock proteins up regulated, suggesting candidate biomarkers (i.e., serpinh1) for thermal stress evaluation in this species. The present study shows that high-temperature stress generated a major effect on the liver of red cusk-eel, knowledge to consider for the aquaculture and fisheries of this species.
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The buoyancy of eggs and embryos is associated with successful development in pelagic fish. Buoyancy is the result of oocyte hydration, which depends on the osmotic force exerted by free amino acids (FAA) generated by yolk proteolysis, and cathepsins are the main enzymes involved in this process. Seriola lalandi is a pelagic fish whose farming has been hampered by development failure that have been partially attributed to decreased buoyancy of embryos. Therefore, the aim of this study was to compare the mRNA expression and activity of cathepsins B, D, and L, as well as the FAA content in floating and low-floating embryos at different developmental stages. The chosen stages were eggs, morula, blastula, gastrula and 24 h embryos. Complementary assessments showed that there were no differences attributed to buoyancy status in embryo and oil droplet diameters, as well as the transcriptional status at any developmental stage. Cathepsin B did not show differences in mRNA expression or activity related to buoyancy at any stage. Cathepsin D displayed higher transcript and activity levels only in low-floating eggs compared with those floating. Cathepsin L showed higher expression in floating eggs and 24 h embryos compared with that of low-floating, but the activity of this enzyme was higher in floating eggs and morula. Total FAA content constantly decreased throughout development in floating embryos, but it was always higher than low-floating embryos until gastrula stage. In 24 h embryos floating and low-floating embryos share similar quantities of FAA. In summary, differences in the expression and activity of cathepsins between floating and low-floating embryos could be revealed at specific embryonic stages, suggesting different functions of these enzymes throughout development. Besides 24 h embryos, FAA content seems to be a decisive factor for buoyancy of embryos during early development of S. lalandi. Overall, considering the main role of cathepsins and FAA in buoyancy acquisition process and therefore in both embryo quality and viability, our study identifies good marker candidates to evaluate embryo quality in the farming of this species.
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Understanding the genetic status of aquaculture strains is essential for developing management guidelines aimed at sustaining the rates of genetic gain for economically important traits, as well as securing populations that will be robust to climate change. Coho salmon was the first salmonid introduced to Chile for commercial purposes and now comprises an essential component of the country's aquaculture industry. Several events, such as admixture, genetic bottlenecks, and rapid domestication, appear to be determinants in shaping the genome of commercial strains representing this species. To determine the impact of such events on the genetic diversity of these strains, we sought to estimate the effective population size (Ne) of several of these strains using genome-wide approaches. We compared these estimates to commercial strains from North America and Japan, as well as a hatchery strain used for supportive breeding of wild populations. The estimates of Ne were based on a method robust to assumptions about changes in population history, and ranged from low (Ne = 34) to relatively high (Ne = 80) in the Chilean strains. These estimates were higher than those obtained from the commercial North American strain but lower than those observed in the hatchery population and the Japanese strain (with Ne over 150). Our results suggest that some populations require measures to control the rates of inbreeding, possibly by using genomic information and incorporating new genetic material to ensure the long-term sustainability of these populations.
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Salmonid rickettsial septicemia (SRS) is the major infectious disease of the Chilean salmonid aquaculture industry caused by Piscirickettsia salmonis. Intensive farming conditions generate stress and increased susceptibility to diseases, being skeletal muscle mainly affected. However, the interplay between pathogen infection and stress in muscle is poorly understood. In this study, we perform an RNA-seq analysis on rainbow trout myotubes that are pretreated for 3 h with cortisol (100 ng/mL) and then infected with P. salmonis strain LF-89 for 8 h (MOI 50). Twelve libraries are constructed from RNA samples (n = 3 per group) and sequenced on Illumina HiSeq 4000. A total of 704,979,454 high-quality reads are obtained, with 70.25% mapped against the reference genome. In silico DETs include 175 total genes-124 are upregulated and 51 are downregulated. GO enrichment analysis reveals highly impacted biological processes related to apoptosis, negative regulation of cell proliferation, and innate immune response. These results are validated by RT-qPCR of nine candidate transcripts. Furthermore, cortisol pretreatment significantly stimulated bacterial gene expression of ahpC and 23s compared to infection. In conclusion, for the first time, we describe a transcriptomic response of trout myotubes infected with P. salmonis by inducing apoptosis, downregulating cell proliferation, and intrinsic immune-like response that is differentially regulated by cortisol.
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Brucella abortus, B. canis, and pathogenic Leptospira are zoonotic pathogens that infect humans, as well as domestic and wild animals. In wild canids, they may affect their fertility and reproduction, threatening their conservation. Wild canids play a crucial role in the environment as meso- and top-predators and environmental sentinels for zoonotic pathogens. In Chile, three species of wild canids are present, and due to changes in land use and environmental dynamics, it is of utmost relevance to determine the role of these species in the epidemiology of brucellosis and leptospirosis. This study aimed to detect the exposure to B. abortus, B. canis, and pathogenic Leptospira by serologic, bacteriologic, and molecular techniques in native foxes from rehabilitation and exhibition centers in Central Chile. Forty-six blood samples were obtained from Lycalopex culpaeus and L. griseus, detecting 10.9% of seropositivity to B. canis and 7.7% to L. Javanica. No seropositivity was seen for B. abortus. Exposure was not registered by culture and qPCR in any of the sampled animals. Our findings are the first register of exposure to any Brucella species in wild canids in Chile and highlight the need to establish surveillance programs of these emerging pathogens.
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Skeletal muscle is the most abundant tissue in teleosts and is essential for movement and metabolism. Recently, it has been described that skeletal muscle can express and secrete immune-related molecules during pathogen infection. However, the role of this tissue during infection is poorly understood. To determine the immunocompetence of fish skeletal muscle, juvenile rainbow trout (Oncorhynchus mykiss) were challenged with Piscirickettsia salmonis strain LF-89. P. salmonis is the etiological agent of piscirickettsiosis, a severe disease that has caused major economic losses in the aquaculture industry. This gram-negative bacterium produces a chronic systemic infection that involves several organs and tissues in salmonids. Using high-throughput RNA-seq, we found that 60 transcripts were upregulated in skeletal muscle, mostly associated with inflammatory response and positive regulation of interleukin-8 production. Conversely, 141 transcripts were downregulated in association with muscle filament sliding and actin filament-based movement. To validate these results, we performed in vitro experiments using rainbow trout myotubes. In myotubes coincubated with P. salmonis strain LF-89 at an MOI of 50, we found increased expression of the proinflammatory cytokine il1b and the pattern recognition receptor tlr5s 8 and 12 h after infection. These results demonstrated that fish skeletal muscle is an immunologically active organ that can implement an early immunological response against P. salmonis.
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Enfermedades de los Peces/inmunología , Inflamación/inmunología , Músculo Esquelético/inmunología , Oncorhynchus mykiss/inmunología , Piscirickettsia/fisiología , Infecciones por Piscirickettsiaceae/inmunología , Transcriptoma , Animales , Acuicultura , Enfermedades de los Peces/genética , Enfermedades de los Peces/microbiología , Perfilación de la Expresión Génica , Inflamación/genética , Inflamación/microbiología , Músculo Esquelético/metabolismo , Músculo Esquelético/microbiología , Oncorhynchus mykiss/genética , Oncorhynchus mykiss/microbiología , Infecciones por Piscirickettsiaceae/microbiologíaRESUMEN
In pelagic fish, embryo buoyancy is a noteworthy aspect of the reproductive strategy, and is associated with overall quality, survival, and further developmental success. In captivity, the loss of buoyancy of early embryos correlates with high mortality that might be related to massive cell death. Therefore, the aim of this study was to evaluate under captivity conditions the expression of genes related to the apoptosis process during the early embryonic development of the pelagic fish Seriola lalandi, and its relationship to the buoyancy of embryos. The relative expression of bcl2, bax-like, casp9, casp8, and casp3 was evaluated by RT-qPCR and FasL/Fas protein levels by western blot in five development stages of embryos sorted as floating or low-floating. All genes examined were expressed in both floating and low-floating embryos up to 24 h of development. Expression of the pro-apoptotic factors bax, casp9, casp8, and casp3 was higher in low-floating as compared with floating embryos in a developmental stage-specific manner. In contrast, there was no difference in expression of bcl2 between floating and low-floating embryos. Fas protein was detected as a single band in floating embryos without changes in expression throughout development; however, in low-floating embryos, three higher intensity reactive bands were detected in the 24-h embryos. Interestingly, FasL was only detected at 24-h in floating embryos, whereas in low-floating samples this ligand was present at all stages, with a sharp increase as development progressed. Cell death, as evaluated by the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay, was highly increased in low-floating embryos as compared to floating embryos throughout all developmental stages, with the highest levels observed during the gastrula stage and at 24 h. The results of this study suggest that an increase in cell death, probably associated with the intrinsic and extrinsic apoptosis pathways, is present in low-floating embryos that might explain their lower developmental potential under captivity conditions.
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Shiga toxin-producing Escherichia coli (STEC) is a zoonotic pathogen that causes severe illness in humans, often associated with foodborne outbreaks. Antimicrobial resistance among foodborne E. coli has increased over the last decades becoming a public health issue. In this study, the presence and features of STEC were investigated in samples of meat, seafood, vegetables and ready-to-eat street-vended food collected in Chile, using a genomic and microbiological approach. Phenotypic and genotypic antimicrobial resistance profiles were determined, and serotype, phylogroup, sequence type (ST) and phylogenomics were predicted using bioinformatic tools. Three thousand three hundred samples collected in 2019 were screened, of which 18 were positive for STEC strains (0.5%), with stx2a (61.1%) being the predominant stx subtype. The presence of the virulence genes lpfA (100%), iha and ehaA (94.4%), and ehxA, hlyA and saa (83.3%) was confirmed among the STEC strains; the Locus of adhesion and autoaggregation (LAA) was predicted in 14 (77.8%) strains. Strains displayed resistance to colistin (100%), and intermediate resistance to enrofloxacin (11.1%) and chloramphenicol (5.6%). In this regard, mutations in the two-component regulatory system genes pmrA (S29G), pmrB (D283G) and phoP (I44L), and the presence of the qnrB19 gene were confirmed. STEC strains belonged to ST11231 (38.9%), ST297 and ST58 (16.7% each), and ST1635, ST11232, ST446, ST442 and ST54 (5.6% each), and the most frequently detected serotypes were O113:H21 (44.4%), O130:H11 and O116:H21 (16.7% each), and O174:H21 (11.1%). Strains belonging to the international ST58 showed genomic relatedness with worldwide strains from human and non-human sources. Our study reports for the first time the genomic profile of STEC strains isolated from food in Chile, highlighting the presence of international clones and sequence types commonly associated with human infections in different geographical regions, as well as the convergence of virulence and resistance in STEC lineages circulating in this country.
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Antibacterianos/farmacología , Microbiología de Alimentos , Escherichia coli Shiga-Toxigénica/genética , Chile , Farmacorresistencia Bacteriana , Genoma Bacteriano , Genómica , Humanos , Filogenia , Secuenciación Completa del GenomaRESUMEN
Red cusk-eel (Genypterus chilensis) is a native species with potential for Chilean aquaculture diversification. However, no information exists on the effects of temperature on oxidative stress and eggs quality markers in post-ovulatory eggs and ovary of this species. We determine that high and low temperature generate oxidative damage on post-ovulatory eggs, with no effect on ovary. Temperature induces thermal stress markers expression on post-ovulatory eggs, and modulates antioxidant and eggs quality markers on post-ovulatory eggs and ovary, information to consider for quality evaluation in the red cusk-eel management.
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Peces/fisiología , Ovario/fisiología , Óvulo/fisiología , Estrés Oxidativo/fisiología , Temperatura , Animales , Acuicultura , Chile , Femenino , Proteínas de Peces/genética , Peces/genética , Regulación de la Expresión Génica/fisiología , Estrés Oxidativo/genéticaRESUMEN
BACKGROUND: Brucella canis is the etiological agent of canine brucellosis, a worldwide neglected zoonosis that constitutes one of the major infectious causes of infertility and reproductive failure in dogs. Although genomic information available for this pathogen has increased in recent years, here we report the first genome sequencing of a B. canis strain in Chile, and the differences in virulence genes with other B. canis strains. RESULTS: Genome assembly produced a total length of 3,289,216 bp, N50 of 95,163 and GC% of 57.27, organized in 54 contigs in chromosome I, and 21 contigs in chromosome II. The genome annotation identified a total of 1981 CDS, 3 rRNA and 36 tRNA in chromosome I, and 1113 CDS and 10 tRNA in chromosome II. There is little variation between the different strains and the SCL isolate. Phylogenetic analysis showed that the Chilean SCL strain is closely related to B. canis and B. suis strains. Small differences were found when compared to the Serbian isolate, but all strains shared the same recent common ancestor. Finally, changes in the sequence of some virulence factors showed that the SCL strain is similar to other South American B. canis strains. CONCLUSIONS: This work sequenced and characterized the complete genome of B. canis strain SCL, evidencing the complete presence of all the genes of the virB operon, and minor changes in outer membrane proteins and in the urease operon. Our data suggest that B. canis was introduced from North America and then spread throughout the South American continent.
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Animales , Perros , Brucelosis/epidemiología , Brucella canis/genética , Brucella canis/patogenicidad , Ureasa/genética , Brucelosis/transmisión , Zoonosis , Chile , GenomaRESUMEN
The red cusk-eel (Genypterus chilensis) is a native species with strong potential to support Chilean aquaculture diversification. Environmental stressors, such as temperature, may generate important effects in fish physiology with negative impact. However, no information exists on the effects of thermal stress in Genypterus species or how this stressor affects the skeletal muscle. The present study evaluated for the first time the effect of high temperature stress in red cusk-eel juveniles to determine changes in plasmatic markers of stress (cortisol, glucose and lactate dehydrogenase (LDH)), the transcriptional effect in skeletal muscle genes related to (i) heat shock protein response (hsp60 and hsp70), (ii) muscle atrophy and growth (foxo1, foxo3, fbxo32, murf-1, myod1 and ddit4), and (iii) oxidative stress (cat, sod1 and gpx1), and evaluate the DNA damage (AP sites) and peroxidative damage (lipid peroxidation (HNE proteins)) in this tissue. Thermal stress generates a significant increase in plasmatic levels of cortisol, glucose and LDH activity and induced heat shock protein transcripts in muscle. We also observed an upregulation of atrophy-related genes (foxo1, foxo3 and fbxo32) and a significant modulation of growth-related genes (myod1 and ddit4). Thermal stress induced oxidative stress in skeletal muscle, as represented by the upregulation of antioxidant genes (cat and sod1) and a significant increase in DNA damage and lipid peroxidation. The present study provides the first physiological and molecular information of the effects of thermal stress on skeletal muscle in a Genypterus species, which should be considered in a climate change scenario.
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Anguilas , Enfermedades de los Peces , Trastornos de Estrés por Calor , Animales , Glucemia/análisis , Daño del ADN , Anguilas/sangre , Anguilas/genética , Anguilas/fisiología , Enfermedades de los Peces/sangre , Enfermedades de los Peces/genética , Enfermedades de los Peces/metabolismo , Enfermedades de los Peces/patología , Proteínas de Peces/genética , Trastornos de Estrés por Calor/sangre , Trastornos de Estrés por Calor/genética , Trastornos de Estrés por Calor/patología , Trastornos de Estrés por Calor/veterinaria , Hidrocortisona/sangre , Músculo Esquelético/metabolismo , Músculo Esquelético/patología , Atrofia Muscular , Estrés Oxidativo , TranscriptomaRESUMEN
Canine brucellosis caused by Brucella canis is a zoonotic disease that causes reproductive alterations in dogs, such as infertility, abortion, and epididymitis. This pathogen is especially prevalent in South America, and due to the lack of official control programs and the growing trend of adopting dogs it constitutes a public health risk that must be addressed. The aim of this study was to determine the prevalence of B. canis infection in kennel, shelter, and household dogs and to characterize the genomic properties of circulating strains, including ure and virB operons and omp25/31 genes. Samples from 771 dogs were obtained, and the infection was detected by blood culture and/or serology in 7.0% of the animals. The complete ure and virB operons and the omp25/31 genes were detected. Interestingly, we found different single-nucleotide polymorphisms (SNPs) in some of the analyzed genes, which could mean a change in the fitness or virulence of these strains. This study provides further evidence about dogs as a source of B. canis strains that can infect people. This also highlights the need to implement official control programs, including the mandatory testing of dogs, especially stray dogs, before adoption.
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In echinoderms, the immune system plays a relevant role in defense against infection by pathogens. Particularly, in sea urchins, the immune system has been shown to be complex, especially in terms of the variety of immune genes and molecules described. A key component of the response to external pathogens are the Toll-like receptors (TLRs), which are a well-characterized class of pattern recognition receptors (PRRs) that participate in the recognition of pathogen-associated molecular patterns (PAMPs). Despite the fact that TLRs have been described in several sea urchin species, for the red sea urchin (Loxechinus albus), which is one of the most important sea urchins across the world in terms of fisheries, limited information on the TLR-mediated immune response exists. In the present study, for the first time, we evaluated the effect of thermal stress, LPS and poly I:C treatment on the coelomocyte immune response of Loxechinus albus to determine how these factors modulate TLR and strongylocin (antimicrobial peptides of echinoderms) responses. We show that the tlr3-like, tlr4-like, tlr6-like and tlr8-like transcripts are modulated by poly I:C, while LPS only modulates the tlr4-like response; there was no effect of temperature on TLR expression, as evaluated by RT-qPCR. Additionally, we showed that strongylocin-1 and strongylocin-2 are modulated in response to simulated viral infection with poly I:C, providing the first evidence of strongylocin expression in L. albus. Finally, we determined that temperature and LPS modify the viability of coelomocytes, while poly I:C treatment did not affect the viability of these cells. This study contributes to the knowledge of immune responses in sea urchins to improve the understanding of the role of TLRs and strongylocins in echinoderms.
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Inmunidad , Lipopolisacáridos/farmacología , Poli I-C/farmacología , Erizos de Mar/inmunología , Temperatura , AnimalesRESUMEN
The red cusk-eel (Genypterus chilensis) is a native species with strong potential to support Chilean aquaculture diversification. Under commercial conditions, fish are exposed to several stressors. To date, little is known about the mechanism involved in the stress response of red cusk-eel, and there is no information related to the regulation mediated by long noncoding RNAs (lncRNAs). The objective of this work was to identify for the first time the lncRNAs in the transcriptome of G. chilensis and to evaluate the differential expression levels of lncRNAs in the liver, head kidney, and skeletal muscle in response to handling stress. We used previously published transcriptome data to identify the lncRNAs by applying a series of filters based on annotation information in several databases to discard coding sequences. We identified a total of 14,614 putative lncRNAs in the transcriptome of red cusk-eel, providing a useful lncRNA reference resource to be used in future studies. We evaluated their differential expression in response to handling stress in the liver, head kidney, and skeletal muscle, identifying 112, 323, and 108 differentially expressed lncRNAs, respectively. The results suggest that handling stress in red cusk-eel generate an altered metabolic status in liver, altered immune response in head kidney, and skeletal muscle atrophy through an important coding and noncoding gene network. This is the first study that identifies lncRNAs in Genypterus genus and that evaluates the relation between handling stress and lncRNAs in teleost fish, thereby providing valuable information regarding noncoding responses to stress in Genypterus species.
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Anguilas/genética , ARN Largo no Codificante , Estrés Fisiológico , Animales , Acuicultura , Anguilas/metabolismo , Riñón Cefálico/metabolismo , Hígado/metabolismo , Músculo Esquelético/metabolismo , RNA-SeqRESUMEN
The red cusk-eel (Genypterus chilensis) is a native Chilean species with a high-value market, with the potential to diversify Chilean aquaculture. The objective of this study was to develop a set of microsatellite markers, estimate genetic parameters, determine population differentiation, and identify the population structure of wild and commercial populations of G. chilensis. We discovered 6427 microsatellites markers from RNA-seq data, of which 54.9%, 20.2% and 16.8% were di-, tri-, and tetranucleotides, respectively. We used 12 of these markers to genotype two sets of broodstock, one group from commercial fish, and one group from wild fish from the Coquimbo Region of G. chilensis. We estimate the genetic parameters of the markers, selecting ten polymorphic markers (PIC > 0.5). We observed differences in the inbreeding coefficient among populations, with high values of inbreeding in one broodstock set and lower values in the other groups. The evaluation of population differentiation using Fst showed small (0.0195) to large (0.1888) genetic differentiation between the groups. The structure analysis showed that commercial and wild groups were formed by three clusters, without relevant evidence of admixture process, suggesting that groups evaluated in this study are formed of at least three subpopulations of G. chilensis, which could be explained by the low or lack of migration suggested for this species. This is the first study that identifies a high number of molecular markers in G. chilensis, providing relevant information of the genetic structure of commercial and wild population of this species.
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Peces/genética , Repeticiones de Microsatélite/genética , Transcriptoma/genética , Animales , ADN/análisis , ADN/genética , Explotaciones Pesqueras , Variación GenéticaRESUMEN
Growth differentiation factor 9 (GDF-9) and bone morphogenetic protein 15 (BMP-15) have pivotal roles in oocyte development in many species, therefore the aim was to investigate these factors during in vitro maturation (IVM) of canine oocytes. Canine cumulus oocytes complexes (COCs) were cultured in six groups for 72 hr in a supplemented TCM199-Hepes medium as (a) Control group; (b) GDF-9 antibody (Ab); (c) BMP-15 Ab; (d) recombinant human (rh) GDF-9; (e) rh BMP-15 or (f) rh BMP-15 and GDF-9. Data were evaluated by ANOVA. The Abs against GDF-9 or BMP-15 had a negative impact on meiotic development. Higher (p < 0.05) number of oocytes was arrested at GVBD stage when they were incubated with either GDF-9 Ab (64.4 ± 2.1%) or BMP-15 Ab (67.2%± 4.9%) in comparison to those in control group (32.4 ± 7.8%). In contrast, more (p < 0.05) oocytes in control group reached MI (37.4 ± 1.3%) and MII stages (10.2 ± 2.1%) comparing to those groups with GDF-9 Ab (23.1 ± 4.7% MI; 0.0% MII) or BMP-15 Ab (16.4 ± 2.4%MI; 5.9% ± 2.1 MII). Higher rates (p < 0.05) of oocytes in control group stayed still arrested at GV (19.9 ± 8.6%) in comparison to those cultured with either rhGDF-9 (3.7 ± 0.4%) or rhBMP-15 (10.9 ± 0.7%). However, there were no differences in MII rates between oocytes cultured with GDF-9 (14.7 ± 3.1) and BMP-15 (7.8 ± 2.5) separately. But, more oocytes (p < 0.05) reached the MII stage (20.5 ± 3.8%) compared to those exposed to each protein separately and to the control group. These results suggest that these proteins likely contribute to the meiotic development in dogs.
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Proteína Morfogenética Ósea 15/farmacología , Factor 9 de Diferenciación de Crecimiento/farmacología , Técnicas de Maduración In Vitro de los Oocitos/veterinaria , Oocitos/fisiología , Animales , Anticuerpos/farmacología , Células Cultivadas , Perros , Femenino , Humanos , Oocitos/efectos de los fármacos , Oogénesis , Proteínas Recombinantes/farmacologíaRESUMEN
In pelagic species such as Seriola lalandi, survival of both the eggs and embryos depends on yolk processing during oocyte maturation and embryo development. The main enzymes involved in these processes are the cathepsins, which are essential for the hydration process, acquiring buoyancy and nutrition of the embryo before hatching. This study aimed to investigate the mRNA expression profiles of cathepsins B, D and L (catb, catd and catl) and the activity of these enzymes during early development in S. lalandi. We included previtellogenic oocytes (PO). All three enzymes were highly expressed in PO, but the expression was reduced throughout development. Between PO and recently spawned eggs (E1) the transcript to catb and catd decreased, unlike catl. Cathepsin B activity, showed stable levels between PO until blastula stage (E4). High activities levels of cathepsins D and L were observed in E1 in comparison with later developmental stages. Cathepsin L activity remained constant until E1, consistent with observations in other pelagic spawners, where its participation in a second protolithic cleavage of the yolk proteins, has been proposed for this enzyme. Their profiles of both mRNA expression and enzymatic activity indicate the importance of these enzymes during early development and suggest different roles in egg yolk processing for the hydration process and nutrition in early embryos in this species.