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The obligate air-breathing Amazonian fish, Arapaima gigas, hatch as water-breathing larvae but with development, they modify their swim bladder to an air-breathing organ (ABO) while reducing their gill filaments to avoid oxygen loss. Here, we show that significant changes already take place between 4 weeks (1.6 g) and 11 weeks (5 g) post hatch, with a reduction in gill lamellar surface area, increase in gill diffusion distance, and proliferation of the parenchyma in the ABO. By using a variety of methods, we quantified the surface area and diffusion distances of the gills and skin, and the swim bladder volume and anatomical complexity from hatch to 11-week-old juveniles. In addition, we identified the presence of two ionocyte types in the gills and show how these change with development. Until 1.6 g, A. gigas possess only the H+ -excreting/Na+ -absorbing type, while 5-g fish and adults have an additional ionocyte which likely absorbs H+ and Cl- and excretes HCO3- . The ionocyte density on the gill filaments increased with age and is likely a compensatory mechanism for maintaining ion transport while reducing gill surface area. In the transition from water- to air-breathing, A. gigas likely employs a trimodal respiration utilizing gills, skin, and ABO and thus avoid a respiratory-ion regulatory compromise at the gills.

Made available in DSpace on 2022-04-28T19:46:56Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-12-01 Julius-Maximilians-Universität Würzburg Arapaima gigas is one of the largest freshwater fish species of high ecological and economic importance. Overfishing and habitat destruction are severe threats to the remaining wild populations. By incorporating a chromosomal Hi-C contact map, we improved the arapaima genome assembly to chromosome-level, revealing an unexpected high degree of chromosome rearrangements during evolution of the bonytongues (Osteoglossiformes). Combining this new assembly with pool-sequencing of male and female genomes, we identified id2bbY, a duplicated copy of the inhibitor of DNA binding 2b (id2b) gene on the Y chromosome as candidate male sex-determining gene. A PCR-test for id2bbY was developed, demonstrating that this gene is a reliable male-specific marker for genotyping. Expression analyses showed that this gene is expressed in juvenile male gonads. Its paralog, id2ba, exhibits a male-biased expression in immature gonads. Transcriptome analyses and protein structure predictions confirm id2bbY as a prime candidate for the master sex-determiner. Acting through the TGFβ signaling pathway, id2bbY from arapaima would provide the first evidence for a link of this family of transcriptional regulators to sex determination. Our study broadens our current understanding about the evolution of sex determination genetic networks and provide a tool for improving arapaima aquaculture for commercial and conservation purposes. Developmental Biochemistry Biocenter University of Wuerzburg, Am Hubland The Xiphophorus Genetic Stock Center Department of Chemistry and Biochemistry Texas State University, San Marcos Biochemistry and Cell Biology Biocenter University of Wuerzburg, Am Hubland Sigenae GenPhySE INRAE ENVT Université de Toulouse MIAT INRA Université de Toulouse, Chemin de Borde Rouge INRAE LPGP Department of Ecology and Evolution University of Lausanne and Swiss Institute of Bioinformatics Embrapa Amazônia Ocidental Embrapa Pesca E Aquicultura Reproductive and Molecular Biology Group Department of Morphology Institute of Biosciences UNESP GeT-PlaGe INRAE Genotoul Leibniz-Institute of Freshwater Ecology and Inland Fisheries IGB, Müggelseedamm 301 & 310 Amphibian Research Center Hiroshima University Comprehensive Cancer Center Mainfranken University Hospital Reproductive and Molecular Biology Group Department of Morphology Institute of Biosciences UNESP

The use of biopesticides based on Bacillus thuringiensis (Berliner 1915) in agriculture has been considered harmless for non-target organisms such as fish. The present study aimed to investigate the effects of a biopesticide based on B. thuringiensis on the physiology and histology of the liver, kidney and intestine of Arapaima gigas (Schinz, 1822), via exposure to water (0.13 g/L) and in the diet (0.13 g), after 24 and 48 h. Fish subjected to B. thuringiensis in the water of their breeding and feeding tanks exhibited mortality due to changes in erythrogram (hematocrit, hemoglobin, erythrocytes), thrombogram and leucogram blood parameters, and plasma (sodium, chloride, potassium, cholesterol, glucose, triglycerides, cortisol and total proteins) and enzymatic (Aspartate Amino Transferase and Alanine Amino Transferase) biochemistry. Histopathological alterations in the liver and kidney ranged from mild to severe and were characterized by the presence of cytoplasmic vacuolization, nuclear hypertrophy and atrophy, melanomacrophage centers and necrosis, and in the intestine by changes to the number of villi and goblet cells. Therefore, these physiological and histopathological alterations indicate that care should be taken with the dispersion of biopesticides based on agricultural B. thuringiensis in fish farming.

Background: Arapaima gigas (Schinz, 1822) is the largest freshwater scaled fish in the world, and an emerging species for tropical aquaculture development. Conservation of the species, and the expansion of aquaculture requires the development of genetic tools to study polymorphism, differentiation, and stock structure. This study aimed to investigate genomic polymorphism through ddRAD sequencing, in order to identify a panel of single nucleotide polymorphisms (SNPs) and to simultaneously assess genetic diversity and structure in wild (from rivers Amazon, Solimões, Tocantins and Araguaia) and captive populations. Results: Compared to many other teleosts, the degree of polymorphism in A. gigas was low with only 2.3% of identified RAD-tags (135 bases long) containing SNPs. A panel of 393 informative SNPs was identified and screened across the five populations. Higher genetic diversity indices (number of polymorphic loci and private alleles, Shannon?s Index and HO) were found in populations from the Amazon and Solimões, intermediate levels in Tocantins and Captive, and very low levels in the Araguaia population. These results likely reflect larger population sizes from less urbanized environments in the Amazon basin compared to Araguaia. Populations were significantly differentiated with pairwise FST values ranging from 0.086 (Amazon × Solimões) to 0.556 (Amazon × Araguaia). Mean pairwise relatedness among individuals was significant in all populations (P < 0.01), reflecting a degree of inbreeding possibly due to severe depletion of natural stocks, the species sedentary behaviour and possible sampling biases. Although Mantel test was not significant (P = 0.104; R2 = 0.65), Bayesian analysis in STRUCTURE and discriminant analysis of principal components (DAPC) showed populations of Amazon and Solimões to be genetically differentiated from Araguaia, with Tocantins comprising individuals from both identified stocks. Conclusions: This relatively rapid genotyping by sequencing approach proved to be successful in delineating arapaima stocks. The approach and / or SNP panels identified should prove valuable for more detailed genetic studies of arapaima populations, including the elucidation of the genetic status of described discrete morphotypes and aid in delivery of conservation programs to maintain genetic diversity in reservoirs across the Amazon region. Made available in DSpace on 2019-02-02T23:30:37Z (GMT). No. of bitstreams: 1 CNPASA2019bmcgenetics.pdf: 1798542 bytes, checksum: 978ae6a34bd5f90bccc338c1b3ad1d10 (MD5) Previous issue date: 2019-01-31

The fish heart ventricle has varied morphology and may have a specific morpho-functional design in species adapted to extreme environmental conditions. In general, the Amazonian ichthyofauna undergoes constant variations in water temperature, pH and oxygen saturation, which makes these species useful for investigations of cardiac morphology. Arapaima gigas, a member of the ancient teleost group Osteoglossomorpha, is one of the largest freshwater fish in the world. This species has a specific heart metabolism that uses fat as the main fuel when O2 supplies are abundant but also can change to glycogen fermentation when O2 content is limiting. However, no information is available regarding its heart morphology. Here, we describe the heart of A. gigas, with emphasis on the ventricular anatomy and myoarchitecture. Specimens of A. gigas weighing between 0.3 and 4040 g were grouped into three developmental stages. The hearts were collected and the anatomy analyzed with a stereomicroscope, ultrastructure with a scanning electron microscope, and histology using toluidine blue, Masson's trichrome and Sirius red stains. The ventricle undergoes morphological changes throughout its development, from the initial saccular shape with a fully trabeculated myocardium and coronary vessel restricted to the subepicardium (Type I) (group 1) to a pyramidal shape with mixed myocardium and coronary vessels that penetrate only to the level of the compact layer (Type II) (groups 2 and 3). The trabeculated myocardium has a distinct net-like organization in all the specimens, differing from that described for other teleosts. This arrangement delimits lacunae with a similar shape and distribution, which seems to allow a more uniform blood distribution through this myocardial layer. Made available in DSpace on 2019-05-01T00:45:39Z (GMT). No. of bitstreams: 1 CPAFAP2019HeartstructureintheAmazonian.pdf: 3145035 bytes, checksum: 8816d442538e9dac41c9bb58fad5e867 (MD5) Previous issue date: 2019

This work was supported by Rede de Pesquisa em Gen?mica Populacional Humana (RPGPH)?3381/2013 CAPES-BioComputacional; FADESP/PROPESP/UFPA (Universidade Federal do Par?); and CNPq (Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico). It was also supported by CNPq/Produtividade grant 304413/2015-1 to A.R.S. and CNPq/Produtividade grant 305258/2013-3 to S.S. Universidade Federal do Par?. Instituto de Ci?ncias Biol?gicas. Laborat?rio de Gen?tica Humana e M?dica. Bel?m, PA, Brazil. Universidade Federal do Rio Grande do Norte. Bioinformatics Multidisciplinary Environment. Natal, RN, Brazil. Universidade Federal do Par?. Instituto de Ci?ncias Biol?gicas. Laborat?rio de Gen?tica Humana e M?dica. Bel?m, PA, Brazil. Universidade Federal do Rio Grande do Norte. Bioinformatics Multidisciplinary Environment. Natal, RN, Brazil. Universidade Federal do Rio Grande do Norte. Bioinformatics Multidisciplinary Environment. Natal, RN, Brazil. Universidade Federal do Par?. Instituto de Ci?ncias Biol?gicas. Laborat?rio de Gen?tica Humana e M?dica. Bel?m, PA, Brazil / Universidade Federal do Rio Grande do Norte. Departmento de Gen?tica. Natal, RN, Brazil. Instituto Nacional de C?ncer. Programa de Gen?tica. Rio de Janeiro, RJ, Brazil. Universidade Federal de Minas Gerais. Instituto de Ci?ncias Biol?gicas. Departamento de Bioqu?mica e Imunologia. Belo Horizonte, MG, Brazil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Laborat?rio de Cultura de Tecidos e Citogen?tica. Ananindeua, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Laborat?rio de Cultura de Tecidos e Citogen?tica. Ananindeua, PA, Brasil. Universidade Federal Rural da Amaz?nia. Laborat?rio de Gen?tica Aplicada. Bel?m, PA, Brazil. Universidade Federal do Par?. N?cleo de Pesquisas em Oncologia. Bel?m, PA, Brazil. Universidade Federal do Par?. Instituto de Ci?ncias Biol?gicas. Laborat?rio de Gen?tica Humana e M?dica. Bel?m, PA, Brazil / Universidade Federal do Par?. N?cleo de Pesquisas em Oncologia. Bel?m, PA, Brazil. Universidade Federal do Rio Grande do Norte. Bioinformatics Multidisciplinary Environment. Natal, RN, Brazil / Universidade Federal do Rio Grande do Norte. Departamento de Bioqu?mica. Natal, RN, Brazil. Instituto Nacional de C?ncer. Programa de Gen?tica. Rio de Janeiro, RJ, Brazil / Universidade Federal do Rio de Janeiro. Departamento de Gen?tica. Rio de Janeiro, RJ, Brazil. Universidade Federal do Rio Grande do Norte. Bioinformatics Multidisciplinary Environment. Natal, RN, Brazil / Universidade Federal do Rio Grande do Norte. Instituto do C?rebro. Natal, RN, Brazil. Universidade Federal do Par?. Instituto de Ci?ncias Biol?gicas. Laborat?rio de Gen?tica Humana e M?dica. Bel?m, PA, Brazil / Universidade Federal do Par?. N?cleo de Pesquisas em Oncologia. Bel?m, PA, Brazil. The Pirarucu (Arapaima gigas) is one of the world?s largest freshwater fishes and member of the superorder Osteoglossomorpha (bonytongues), one of the oldest lineages of ray-finned fishes. This species is an obligate air-breather found in the basin of the Amazon River with an attractive potential for aquaculture. Its phylogenetic position among bony fishes makes the Pirarucu a relevant subject for evolutionary studies of early teleost diversification. Here, we present, for the first time, a draft genome version of the A. gigas genome, providing useful information for further functional and evolutionary studies. The A. gigas genome was assembled with 103-Gb raw reads sequenced in an Illumina platform. The final draft genome assembly was ?661?Mb, with a contig N50 equal to 51.23?kb and scaffold N50 of 668?kb. Repeat sequences accounted for 21.69% of the whole genome, and a total of 24,655 protein-coding genes were predicted from the genome assembly, with an average of nine exons per gene. Phylogenomic analysis based on 24 fish species supported the postulation that Osteoglossomorpha and Elopomorpha (eels, tarpons, and bonefishes) are sister groups, both forming a sister lineage with respect to Clupeocephala (remaining teleosts). Divergence time estimations suggested that Osteoglossomorpha and Elopomorpha lineages emerged independently in a period of ?30 Myr in the Jurassic. The draft genome of A. gigas provides a valuable genetic resource for further investigations of evolutionary studies and may also offer a valuable data for economic applications.

The study was funded by the Center of Technological Innovation (CIT), Evandro Chagas Institute (IEC) and National Council of Scientific and Technological Development (CNPq) (Bionorte Project: 550939/2010-5) Universidade Federal do Par? - Campus de Bragan?a. Instituto de Estudos Costeiros. Laborat?rio de Gen?tica e Biologia Molecular. Bragan?a, PA, Brazil. Universidade Federal do Par? - Campus de Bragan?a. Instituto de Estudos Costeiros. Laborat?rio de Gen?tica e Biologia Molecular. Bragan?a, PA, Brazil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil. Universidade Federal do Par? - Campus de Bragan?a. Instituto de Estudos Costeiros. Laborat?rio de Gen?tica e Biologia Molecular. Bragan?a, PA, Brazil. Universidade Federal do Par? - Campus de Bragan?a. Instituto de Estudos Costeiros. Laborat?rio de Gen?tica e Biologia Molecular. Bragan?a, PA, Brazil. BACKGROUND: The Arapaima (Arapaima gigas) is one of the world's largest freshwater bony fish, and is found in the rivers of the Amazon basin. This species is a potential aquaculture resource, although reproductive management in captivity is limited in particular due to the lack of external sexual dimorphism. In this study, using the 454 Roche platform (pyrosequencing) techniques, we evaluated a major portion of the transcriptome of this important Amazonian species. RESULTS: Four libraries obtained from the liver and skin tissue of juvenile specimens (representing males and females separately) were sequenced, yielding 5,453,919 high-quality reads. The de novo transcriptome assembly resulted in 175,792 contigs, with 51,057 significant blast hits. A total of 38,586 transcripts were mapped by Gene Ontology using Blast2GO. We identified 20,219 genes in the total transcriptome (9,551 in the liver and 16,818 in the skin). The gene expression analyses indicated 105 genes in the liver and 204 in the skin with differentiated expression profiles, with 95 being over-expressed in the females and 214 in the males. The log2 Fold Change and heatmap based on Reads Per Kilobase per Million mapped reads (RPKM) revealed that the gene expression in the skin is highly differentiated between male and female arapaima, while the levels of expression in the liver are similar between the sexes. CONCLUSION: Transcriptome analysis based on pyrosequencing proved to be a reliable tool for the identification of genes with differentiated expression profiles between male and female arapaima. These results provide useful insights into the molecular pathways of sexual dimorphism in this important Amazonian species, and for comparative analyses with other teleosts

Parental investment in Arapaima gigas includes nest building and guarding, followed by a care provision when a cephalic fluid is released from the parents' head to the offspring. This fluid has presumably important functions for the offspring but so far its composition has not been characterised. In this study the proteome and peptidome of the cephalic secretion was studied in parental and non-parental fish using capillary electrophoresis coupled to mass spectrometry (CE-MS) and GeLC-MS/MS analyses. Multiple comparisons revealed 28 peptides were significantly different between males and parental males (PC-males), 126 between females and parental females (PC-females), 51 between males and females and 9 between PC-males and PC-females. Identification revealed peptides were produced in the inner ear (pcdh15b), eyes (tetraspanin and ppp2r3a), central nervous system (otud4, ribeye a, tjp1b and syn1) among others. A total of 422 proteins were also identified and gene ontology analysis revealed 28 secreted extracellular proteins. From these, 2 hormones (prolactin and stanniocalcin) and 12 proteins associated to immunological processes (serotransferrin, α-1-antitrypsin homolog, apolipoprotein A-I, and others) were identified. This study provides novel biochemical data on the lateral line fluid which will enable future hypotheses-driven experiments to better understand the physiological roles of the lateral line in chemical communication. Made available in DSpace on 2017-10-28T23:13:12Z (GMT). No. of bitstreams: 1 CNPASA2017plos.pdf: 6615081 bytes, checksum: 5b4346a70e2c175296d1aac3fed12435 (MD5) Previous issue date: 2017-10-27

Arapaima gigas (Osteoglossidae) is one of the largest fish species in the Amazon Basin, attaining lengths of over 2.5 m and weights of over 100 kg. Its flesh is prized, and it has great potential for production in aquaculture systems. However, live pirarucu cannot be reliably sexed visually, even after sexual development, since this species does not have clear external sexual dimorphism. Simple and inexpensive methods for sexing immature pirarucu based on DNA markers would facilitate production of this species in commercial operations. We analyzed A. gigas male and female DNA pools with 566 RAPD primers, generating 2609 fragments, with an estimated 1341 segregating polymorphic markers, and an estimated average spacing of 714 kb, which corresponds to less than 0.1% of the species' genome. Two putative sex-specific fragments were initially identified in bulked samples; but they were not confirmed in a study of individual male and female samples. We suggest that A. gigas has developed a non-chromosomal system of sex determination or, alternatively, that the species has undergone a recent loss of the chromosome carrying the sex-determining locus. Made available in DSpace on 2018-01-26T23:41:24Z (GMT). No. of bitstreams: 1 gmr3157.pdf: 658920 bytes, checksum: 8a9b0b3d54721ca4478e2e902e26a21b (MD5) Previous issue date: 2015-02-05

The common gonadotrophic hormone α-subunit (GTHα) has been previously isolated by our research group from A. gigas pituitaries; in the present work the cDNA sequences encoding FSHβ and LHβ subunits have also been isolated from the same species of fish. The FSH β-subunit consists of 126 amino acids with a putative 18 amino acid signal peptide and a 108 amino acid mature peptide, while the LH β-subunit consists of 141 amino acids with a putative 24 amino acid amino acid signal peptide and a 117 amino acid mature peptide. The highest identity, based on the amino acid sequences, was found with the order of Anguilliformes (61%) for FSHβ and of Cypriniformes (76%) for LHβ, followed by Siluriformes, 53% for FSHβ and 75% for LHβ. Interestingly, the identity with the corresponding human amino acid sequences was still remarkable: 45.1% for FSHβ and 51.4% for LHβ. Three dimensional models of ag-FSH and ag-LH, generated by using the crystal structures of h-FSH and h-LH as the respective templates and carried out via comparative modeling and molecular dynamics simulations, suggested the presence of the so-called "seat-belt", favored by a disulfide bond formed between the 3rd and 12th cysteine in both β-subunits. The sequences found will be used for the biotechnological synthesis of A. gigas gonadotrophic hormones (ag-FSH and ag-LH). In a first approach, to ascertain that the cloned transcripts allow the expression of the heterodimeric hormones, ag-FSH has been synthesized in human embryonic kidney 293 (HEK293) cells, preliminarily purified and characterized.

In the present study we report a spatial autocorrelation analysis of molecular data obtained for Arapaima gigas, and the implication of this study for conservation and management. Arapaima is an important, but critically over-exploited giant food fish of the Amazonian várzea. Analysis of 14 variable microsatellite loci and 2,347 bp of mtDNA from 126 individuals sampled in seven localities within the Amazon basin suggests that Arapaima forms a continuous population with extensive genetic exchange among localities. Weak effect of isolation-by-distance is observed in microsatellite data, but not in mtDNA data. Spatial autocorrelation analysis of genetic and geographic data suggests that genetic exchange is significantly restricted at distances greater than 2,500 km. We recommend implementing a source-sink metapopulation management and conservation model by proposing replicate high quality várzea reserves in the upper, central, and lower Amazon basin. This conservation strategy would: 1) preserve all of the current genetic diversity of Arapaima; 2) create a set of reserves to supply immigrants for locally depleted populations; 3) preserve core várzea areas in the Amazon basin on which many other species depend. We stress that conservation strategies should not only preserve current genetic diversity, but also the evolutionary processes which have generated the observed patterns.

Third-stage larvae (L3) of Hysterothylacium sp. were collected by the first time in juveniles of pirarucu Arapaima gigas farmed in the Rio Preto da Eva, Amazonas state. Ninety-eight (98) out of 100 examined fish showed to be parasitized. Five hundred and ninety larvae of Hysterothylacium sp. were collected from the intestines, stomach and pyloric caeca. The mean intensity of parasite indexes was 6.02 (±5.75) ranging from 1 to 40 larvae per host and the mean abundance was 5.9 (±5.76). The A. gigas is the new host record for larvae of Hysterothylacium sp. in Brazil, and this is the first record of larvae of Hysterothylacium (Nematoda: Anisakidae) with zoonotic potential in the pirarucu from South America.