Haplotypes traceability and genetic variability of the breeding population of pacu (Piaractus mesopotamicus) revealed by mitochondrial DNA.

The main objective of this study was to estimate the genetic diversity levels and haplotype traceability in pacu Piaractus mesopotamicus from the breeding program located in Brazil by analyses of the mitochondrial DNA control region (mtDNA). Moreover, broodstocks from eight commercial fish farms were used for comparative evaluation, four from Brazil (Br1-Br4) and four from Argentina (Ar1-Ar4). The descriptive results revealed 47 polymorphic sites and 51 mutations, which evidenced 34 haplotypes. Ten haplotypes were shared among fish farms and 24 were exclusive. The nucleotide diversity (π) ranged from 0.00031 to 0.01462 and haplotype diversity (Hd) from 0.125 to 0.868. The analysis of molecular variance (AMOVA) indicated high structure present in the analyzed stocks (FST = 0.13356 and ФST = 0.52707). The genetic diversity was high in most of the commercial broodstocks, especially those from Brazil. We observed seven haplotypes in the genetic breeding population, of which four were exclusive and three shared among the commercial fish farms. The genetic diversity was moderate (π = 0.00265 and Hd = 0.424) and considered appropriated for this breeding population of pacu. Our results provide support for the genetic diversity maintenance and mtDNA traceability of pacu commercial broodstocks.

Development of microsatellite markers using next-generation sequencing for the fish Colossoma macropomum.

Tambaqui (Colossoma macropomum) is a fish species from the Amazon and Orinoco Rivers, with favorable characteristics to the cultivation system and great market acceptance in South America. However, the construction of a genetic map for the genetic improvement of this species is limited by the low number of molecular markers currently described. Thus, this study aimed to validate gene-associated and anonymous (non-genic) microsatellites obtained by next generation sequencing (RNA-seq and whole genome shotgun-WGS, respectively), for future construction of a genetic map and search for quantitative trait loci (QTL) in this species. In the RNA-seq data, the observed and expected heterozygosity (Ho and He) ranged from 0.09 to 0.73, and 0.09 to 0.85, respectively. In the WGS data, Ho and He ranged from 0.33 to 0.95, and 0.28 to 0.92, respectively. In general, the evaluation of 200 markers resulted in 45 polymorphic loci, of which 14 were gene-associated (RNA-Seq) and 31 were anonymous (WGS). Moreover, some markers were related to genes of the immune system, biological regulation/control and biogenesis. This study contributes to increase the number of molecular markers available for genetic studies in C. macropomum, which will allow the development of breeding programs assisted by molecular markers.

Genetic Characterization of the Fish Piaractus brachypomus by Microsatellites Derived from Transcriptome Sequencing.

The pirapitinga, Piaractus brachypomus (Characiformes, Serrasalmidae), is a fish from the Amazon basin and is considered to be one of the main native species used in aquaculture production in South America. The objectives of this study were: (1) to perform liver transcriptome sequencing of pirapitinga through NGS and then validate a set of microsatellite markers for this species; and (2) to use polymorphic microsatellites for analysis of genetic variability in farmed stocks. The transcriptome sequencing was carried out through the Roche/454 technology, which resulted in 3,696 non-redundant contigs. Of this total, 2,568 contigs had similarity in the non-redundant (nr) protein database (Genbank) and 2,075 sequences were characterized in the categories of Gene Ontology (GO). After the validation process of 30 microsatellite loci, eight markers showed polymorphism. The analysis of these polymorphic markers in farmed stocks revealed that fish farms from North Brazil had a higher genetic diversity than fish farms from Southeast Brazil. AMOVA demonstrated that the highest proportion of variation was presented within the populations. However, when comparing different groups (1: Wild; 2: North fish farms; 3: Southeast fish farms), a considerable variation between the groups was observed. The FST values showed the occurrence of genetic structure among the broodstocks from different regions of Brazil. The transcriptome sequencing in pirapitinga provided important genetic resources for biological studies in this non-model species, and microsatellite data can be used as the framework for the genetic management of breeding stocks in Brazil, which might provide a basis for a genetic pre-breeding programme.

Microsatellites Associated with Growth Performance and Analysis of Resistance to Aeromonas hydrophila in Tambaqui Colossoma macropomum.

Tambaqui, Colossoma macropomum, is the main native fish species produced in Brazil, and is an important species for genetic improvement in aquaculture. In addition, breeding studies on this species can be optimized with the use of molecular markers associated with productive phenotypes. The objective of the present study was to test the performance of growth traits and resistance to the bacteria, Aeromonas hydrophila, in association with microsatellite markers in C. macropomum. In this study, three full-sib families were subjected to bacterial challenge and morphometric growth assessments. Tambaqui families subjected to the bacterial challenge differed significantly in death time and mortality rate. There was, however, no association between resistance to bacteria and microsatellite markers. In relation to growth traits, we observed a marker/phenotype association in two microsatellites. The marker in the 6b isoform x5 gene (TNCRC6b) was associated with length, whereas an anonymous marker was associated with height. The present study highlighted the evaluation of molecular markers associated with growth traits, and can serve as the basis for future marker-assisted selection (MAS) of tambaqui.