Taís da Silva Lopes

Cooling of pacu (Piaractus mesopotamicus) embryos at various stages of development for 6 or 10 hours

The objective of this research was to verify the effects of cooling embryos of pacu, Piaractus mesopotamicus, in four stages of development during two stocking periods. The stages of embryo development were at: blastoderm, ∼ 64 cells-1.4 h after fertilization (haf); 25% of the epiboly movement--5.2 haf; blastoporous closing--8.0 haf; and optical vesicle appearing--13.3 haf. Embryos were exposed to a cryoprotectant solution containing methanol (10%) and sucrose (0.5 M). Thereafter, embryos were submitted to a cooling curve until they reached -8 °C, and then kept cooled for 6 or 10 h. In addition, for each stage of embryonic development, a control group with uncooled embryos was used to compare hatching rates. The total number of larvae from the first two stages of ontogenetic development (1.4 and 5.2 haf) was lower compared to the other stages (0.0 and 8.0 haf). There was no significant difference between stages 8.0 and 13.3 haf for the total number of larvae (49.9 ± 6.7% and 55.2 ± 6.7%, respectively). Embryo diameter varied according to embryonic stage, providing evidence of differences in membrane permeability. There was a negative correlation between embryo diameter and the total number of larvae (r = -0.372). In conclusion, use of embryonic stages 8.0 and 13.3 haf were recommended for maintaining cooled pacu embryos at -8 °C for 6 or 10 h.

Diversidade genética de estoques de reprodutores de Colossoma macropomum

The genetic diversity of tambaqui (Colossoma macropomum) broodstocks from two hatchery station in Rondonia State was studied by the RAPD marker. Ten primers were used to analyze 30 broodstocks samples from the hatchery stations of Boa Esperanca and Vale Verde. The polymorphic fragments percentage and Shannon genetic diversity index were high in the two broodstocks. The Boa Esperanca broodstock presented an exclusive fragment. The genetic differentiation was low and the number of migrants per generation was high among the broodstocks. The dendrogram did not separate the broodstocks individuals in different groups. The results indicate a high genetic variability in the broodstocks, being a little bit lower in the Vale Verde broodstock. Besides, there is a genetic proximity among the broodstocks.

Chilling curves for Piaractus mesopotamicus (Holmberg, 1887) embryos stored at -8 ◦ C

The present study investigates the effect of different slow chilling curves on the storage of pacu (Piaractus mesopotamicus) embryos submitted to chilling at -8°C. Embryos at the blastopore closure stage were divided into two groups: G1 - embryos exposed to cryoprotectant solution containing methanol (10%) and sucrose (0.5 M), treated as follows: (T1) taken directly from room temperature to the refrigerator without being submitted to the curve; (T2) chilling curve of 0.5°C/min; and (T3) chilling curve of 1°C/min; and G2 - the cryoprotectant solution alone was submitted to these same temperatures, receiving the embryos only after temperature had decreased, corresponding to treatments T4, T5 and T6, respectively. Treatments were kept at -8°C for a period of 6 h. Embryo development was evaluated for each treatment, with six replicates in an entirely randomized design. Survival among embryos not submitted to refrigeration was 94.3 ± 8.05%. Percentage of total larvae (TL) and addled eggs (AE) did not differ statistically between the groups, although percentage of swimming larvae (SL) exhibited higher values in G1 for the 1°C/min curve. Furthermore, when comparing the three chilling curves, a decrease of 1°C/min resulted in the highest TL percentage (90.85%), followed by the 0.5°C/min curve (78.52%). Thus, the use of 1°C/min chilling curves is recommended for P. mesopotamicus embryos stored for 6 h at -8°C.

Monitoramento da variabilidade genética de pacu, Piaractus mesopotamicus, do programa de aumento de estoque do rio Paranapanema

Avaliou-se a variabilidade genetica dos estoques de reprodutores e dos peixes jovens de Piaractus mesopotamicus de tres pisciculturas do estado do Parana, utilizadas no programa de aumento de estoque de peixes no rio Paranapanema. Foi utilizado o marcador RAPD para avaliar as amostras do estoque de reprodutores e dos peixes jovens das pisciculturas de Palotina, Cambara e Andira. A porcentagem de fragmentos polimorficos e o indice de diversidade genetica de Shannon dos estoques de reprodutores variaram de 75,0% a 71,4% e de 0,434 a 0,376, respectivamente. Os peixes jovens das pisciculturas apresentaram valores mais elevados para ambos os parâmetros, com excecao da piscicultura de Palotina, na qual o indice de diversidade genetica de Shannon foi semelhante. Os estoques de reprodutores apresentaram alta variabilidade genetica, e esta foi mantida nos peixes jovens.

Microsatellite analysis of pacu broodstocks used in the stocking program of Paranapanema River, Brazil

Monitoring the genetic diversity has fundamental importance for fish stocking programs. This experiment aims to evaluate the genetic diversity in two hatchery stations (A and B) with pacu Piaractus mesopotamicus (Holmberg, 1887) in Andira, state of Parana, Brazil used in stocking programs of Paranapanema River. Six microsatellite loci were amplified using DNA extracted from 60 fin-clipping samples. The broodstock B had the average number of alleles and the mean heterozygosity (alleles: 3.7 and HO: 0.628) higher than the broodstock A (alleles: 3.5 and HO: 0.600). Alleles with low frequency levels were observed in the both broodstocks. The positive coefficients of endogamy in the locus Pme2 (broodstock A: FIS = 0.30 and broodstock B: FIS = 0.20), Pme5 (broodstock B: FIS = 0.15), Pme14 (broodstock A: FIS = 0.07) and Pme28 (broodstock A: FIS = 0.24 and broodstock B: FIS = 0.20) indicated deficiency of heterozygotes. Presence of null allele in the locus Pme2 was detected. The negative estimates in loci Pme4 (broodstock A: FIS = - 0.43 and broodstock B: FIS = - 0.37), Pme5 (broodstock A: FIS= - 0.11), Pme14 (broodstock B: FIS= - 0.15) and Pme32 (broodstock A: FIS = - 0.93 and broodstock B: FIS = - 0.60) were indicating the excess of heterozygotes. Evidence of linkage disequilibrium and lower allelic richness was found only in the broodstock A. Neis gene diversity was high in both broodstocks. The genetic distance (0.085) and identity (0.918) showed similarity between broodstocks, which reflects a possible common origin. 6.05% of the total genetic variance was due to differences among broodstocks. Recent bottleneck effect in two broodstocks was observed. The results indicated a higher genetic diversity in the two broodstocks and they presented low genetic difference. This was caused by the reproductive management in both hatchery stations, reduction of population size and genetic exchange between the hatchery stations.

Diversidad genética de Piaractus mesopotamicus utilizado en programas de repoblación

Nowadays it has been verified the decrease and the disappearance of many fish species in several aquatic environments in the Brazil. As form of minimizing that impact, stocking programs come being used with more intensity, meantime, little importance has been given to the genetic variability, fundamental parameter for any conservation practices. The objective of this study was to analyze the genetic diversity of Piaractus mesopotamicus stocks used in stocking programs, through the RAPD molecular marker. Sixty broodstocks of two fish farms stations located in the Sapopema (S) and Cambara (C) cities in the Parana State (Brazil) and 30 larvae of the Sapopema offspring (PS) were analyzed. The 10 selected primers yielded 69 fragments of which 62 (89.85%) were polymorphic. Differences (p<0.05) in the frequency of 42 fragments were observed, with four exclusive for the Sapopema stock. The genetic variability results estimated by the percentage of polymorphic fragments (S: 70.15%; PS: 73.13% and C: 77.81%) and for the genetic diversity of Shannon index (S: 0.365; PS: 0.379 and C: 0.468) showed that the genetic variability was maintained in the offspring, due possibly to the good reproductive management. On the other hand, the opposing genetic variability among the two broodstocks indicated a genetic differentiation among them, possibly due to the founder effect. According with the AMOVA, most of the variation is within each stock (62.43%). This result was corroborated with the FST (0.375) values and the effective number of migrants (2.20) that showed a high genetic differentiation and a low gene flow. It was also verified that S and SP were those but similar genetically (genetic identity, IG= 0.851) and that S and C presented less genes in common (IG= 0.786).