Applying a heat transfer mathematical model for the cryopreservation of rainbow trout (Oncorhynchus mykiss) sperm: How straw location over liquid nitrogen level affects freezing rate and fertilization yield.

Cryopreservation of rainbow trout semen under field conditions was analyzed. Straw location over liquid nitrogen level is a crucial variable that affects freezing rate and fertilization yield due to changes in nitrogen vapor external temperature. The objectives were: to analyze cryopreservation protocols by experimentally measuring the cooling rates and fertilization yield of 0.5 ml plastic straws located in nitrogen vapor at different heights corresponding to different external temperatures; to numerically simulate the freezing process, by solving the heat transfer partial differential equations with the corresponding thermo-physical properties of the biological system and the plastic straw; to evaluate and analyze the surface heat transfer coefficient (h) during the freezing process of the straws; to introduce a new variable, the characteristic freezing time (tc), that enables comparison between protocols; this variable was defined as the elapsed period between the initial freezing temperature and a final reference temperature of -40 °C (temperature in which more than 80 % of the water is in a frozen state). The mathematical model predicted the temperature distribution inside the straw, showing a low effect of straw plastic materials (polyethylene-terephthalate glycol, polyvinyl-chloride, and polypropylene) on freezing rates. The average h value obtained from numerical simulations was 25.5 W/m2 K, close to that obtained from the analytical Nusselt correlation for natural convection. An improvement on fertilization trials was observed when the average external nitrogen temperature was -129.6 °C (temperature range: -94 to -171 °C) with an average tc of 56.8 s (ranging between 47 and 72 s). These results corresponded to a height above the level of liquid nitrogen of 2 cm. Comparison with literature reported data showed satisfactory results. Applying mathematical models in the cryobiology field achieved results that are relevant for cryopreservation activities.

Morphologic and genetic variation within a relict Andean catfish, Hatcheria macraei , and its relationship with Trichomycterus areolatus and Bullockia maldonadoi (Siluriformes: Trichomycteridae).

The South American siluriform fishes are found primarily in the Neotropical region, north and east of the Colorado River of Argentina, with a few relict species distributed southward and westward on both sides of the Andes Mountains. Three of these, the closely related trichomycterids Hatcheria macraei, Trichomycterus areolatus and Bullockia maldonadoi, have been subject to historical taxonomic and nomenclatural arrangements. Here, we amplify a 652-bp fragment of COI mtDNA from 55 H. macraei individuals and use publicly available Cytb mtDNA sequences of the three taxa to assess their relationship, genetic variation and haplotype distribution in relation to hydrographic basins. In addition, we extend a recent morphometric study on H. macraei by analyzing body shape in 447 individuals collected from 24 populations across their entire cis-Andean distribution. We identified some lineages previously assigned to T. areolatus that show a closer relationship to either B. maldonadoi or H. macraei, revealing new boundaries to their currently known trans-Andean distribution. We found a great morphologic variation among H. macraei populations and a high genetic variation in H. macraei, T. areolatus and B. maldonadoi associated with river basins. We highlight further integrative studies are needed to enhance our knowledge of the southern Andean trichomycterid diversity.

Thermal effects in rainbow trout (Oncorhynchus mykiss) F1 embryos (farmed female × wild thermal-resistant male).

The aim of this work was to investigate the response of rainbow trout embryos (Oncorhynchus mykiss) (i.e., survival, size at hatching, time to hatching, malformations) to four incubation temperatures (5.8, 8.9, 14.0 and 16.8°C), taking into account the origin of the male parental genome and comparing pure farmed and F1 embryos (farmed female × wild thermal-resistant male). Several consequences of thermal stress were observed: lower accumulated thermal units (ATU) at hatching at high temperatures, and lower survival, shorter hatched free embryos and less-consumed yolk sac at extreme temperatures. The effect of the thermal-adapted male parental genome was shown only in the lower percentage of incompletely hatched free embryos in the F1 families. It appears that to obtain greater modification of thermal performance during early development, the adapted genome of the wild thermal-resistant population has to be included through maternal inheritance, thus producing a stabilized strain selected for domesticity, growth and thermal adaptation.

Early morphological variation and induction of phenotypic plasticity in Patagonian pejerrey

The aim of this work was to study two aspects of phenotypic plasticity in the Patagonian pejerrey Odontesthes hatcheri (Teleostei: Atherinopsidae) the dependence of the early morphology on developmental time and temperature, and the induction of morphological changes by controlled feeding in juveniles. Newly hatched free embryos, incubated at two different temperatures (13 and 18oC), and juveniles were used for the study and induction of phenotypic plasticity. Body and head shapes were analyzed with geometric morphometrics and linear measurements. Our results showed that shape variation at hatching was related to the bending of the embryo head on the yolk sac, increasing the head-trunk angle due to progressive straightening of the embryo. The head-trunk angle was related with temperature at incubation, with embryos incubated at higher temperature being more bent. Embryos that hatched earlier had bigger yolk sacs than those that hatched later. In juveniles, controlled feeding experiments added new morphological variation to that of wild juveniles. In all comparisons, the slenderness of the head, the size of premaxilla and jaw, and the position of the eye showed an enlarged variation due to controlled feeding. These results will contribute to comprehending the complexity of the morphological variation of O. hatcheri.

O objetivo deste trabalho foi estudar a variação morfológica e plasticidade fenotípica do peixe-rei da Patagônia Odontesthes hatcheri (Teleostei: Atherinopsidae), a dependência da morfologia inicial no tempo de desenvolvimento e temperatura, e a indução de alterações morfológicas pela alimentação controlada em juvenis. Embriões recém-nascidos, incubados a duas temperaturas diferentes (13 e 18oC) e juvenis foram utilizados para o estudo de indução de plasticidade fenotípica. Formas do corpo e cabeça foram analisadas com técnicas de morfometria geométrica e medições lineares. Os nossos resultados mostraram que a variação da forma no nascimento foi relacionada com a curvatura da cabeça do embrião no saco vitelino, aumentando o ângulo de cabeça-tronco devido ao endireitamento progressivo do embrião. O ângulo da cabeça-tronco relacionou-se com a temperatura de incubação, com os embriões incubados na temperatura elevada sendo mais curvados. Os embriões que eclodiram mais cedo tinham sacos vitelinos maiores do que aqueles que eclodiram tardiamente. Em juvenis, os experimentos de alimentação controlada adicionaram nova variação morfológica àquela dos juvenis selvagens. Em todas as comparações, a espessura da cabeça, o tamanho da pré-maxila e mandíbula, e a posição do olho mostraram uma maior variação devido à alimentação controlada. Estes resultados irão contribuir para a compreensão da complexidade da variação morfológica de O. hatcheri.

The effect of rearing temperature in larval development of pejerrey, Odontesthes bonariensis: morphological indicators of development

It is well known that in pejerrey water temperature not only affects growth rates but also directs the sexual differentiation process. This fact rise the question of how different the development of pejerrey larvae of the same age is when reared at different temperatures. A description of developmental stages for the embryonic and larval periods of the pejerrey, Odontesthes bonariensis, and the influence of rearing temperature on larval development are presented. Then, larval development was studied at three rearing temperatures, and changes in general morphology, fin morphology, and caudal fin structure have been taken into consideration within the thermal range involved in the temperature sex determination of this species. Fin fold reabsorption, caudal fin formation, and body shape were selected to follow the events leading to the acquisition of the juvenile morphology. The juvenile phenotype was defined when the fin fold was reabsorpted and the caudal fin acquired its definitive homocercal structure. The moment at which the juvenile phenotype was achieved, was evaluated in relation to larval age, size and, shape. The size resulted as the best indicator of development in pejerrey.

A temperatura da água não afeta apenas as taxas de crescimento no peixe-rei, mas também direciona o processo de diferenciação sexual. Este fato levanta o questionamento de quão diferente é o desenvolvimento de larvas do peixe-rei da mesma idade quando criadas em temperaturas diferentes. Este trabalho teve como objetivo apresentar uma descrição do de desenvolvimento de embriões e larvas do peixe-rei, Odontesthes bonariensis, e a influência da temperatura de criação no desenvolvimento das larvas. Neste trabalho, o desenvolvimento das larvas foi estudado em três temperaturas diferentes de cultivo. Foram consideradas as alterações ocorridas na morfologia geral, assim como na morfologia e na estrutura da nadadeira caudal dentro da variação termal da temperatura de determinação sexual desta espécie. A taxa de reabsorção da membrana embrionária, a formação da cauda e o formato do corpo foram selecionados para acompanhar os eventos que levam à aquisição da morfologia juvenil. O fenótipo juvenil foi definido quando a nadadeira caudal foi reabsorvida e a cauda adquiriu a estrutura homocerca. O momento no qual o fenótipo juvenil foi atingido, foi avaliado quanto à idade, tamanho e formato da larva, sendo que o tamanho resultou no melhor indicador do desenvolvimento do peixe-rei.

Ten novel microsatellite loci characterized for a remarkably widespread fish: Galaxias maculatus (Galaxiidae).

Ten polymorphic microsatellite markers (five tetra-, one compound tetra-, one octa- and three dinucleotides) were isolated and characterized for Galaxias maculatus, a fish species widely distributed in the Southern Hemisphere. Markers were tested in 89 individual samples from a single location and the number of alleles ranged between 2 and 28. Observed and expected heterozygosities ranged from 0.103 to 0.910 and 0.098 to 0.935 respectively. No evidence was detected for either linkage disequilibrium (P-values > 0.05 for each locus pair) or deviations from HWE (P-values > 0.05 for every loci).

Climate control on ancestral population dynamics: insight from Patagonian fish phylogeography.

Changes in lake and stream habitats during the growth and retreat of Pleistocene glaciers repeatedly altered the spatial distributions and population sizes of the aquatic fauna of the southern Andes. Here, we use variation in mtDNA control region sequences to infer the temporal dynamics of two species of southern Andean fish during the past few million years. At least five important climate events were associated with major demographic changes: (i) the widespread glaciations of the mid-Pliocene (c. 3.5 Ma); (ii) the largest Patagonian glaciation (1.1 Ma); (iii) the coldest Pleistocene glaciation as indicated by stacked marine delta(18)O (c. 0.7 Ma); (iv) the last southern Patagonian glaciation to reach the Atlantic coast (180 ka); and (v) the last glacial maximum (LGM, 23-25,000 years ago). The colder-water inhabitant, Galaxias platei, underwent a strong bottleneck during the LGM and its haplotype diversity coalesces c. 0.7 Ma. In contrast, the more warm-adapted and widely distributed Percichthys trucha showed continuous growth through the last two glacial cycles but went through an important bottleneck c. 180,000 years ago, at which time populations east of the Andes may have been eliminated. Haplotype diversity of the most divergent P. trucha populations, found west of the Andes, coalesces c. 3.2 Ma. The demographic timelines obtained for the two species thus illustrate the continent-wide response of aquatic life in Patagonia to climate change during the Pleistocene, but also show how differing ecological traits and distributions led to distinctive responses.

Isolation and characterization of 13 microsatellite loci for Percichthys trucha (Percichthyidae).

Thirteen polymorphic microsatellite loci are described for the South American freshwater fish Percichthys trucha. Number of alleles per locus ranged from two to 21 and observed heterozygosities ranged from 0.304 to 0.915 in a sample of 47 individuals from four different sampling locations.

Phylogeography of the Percichthyidae (Pisces) in Patagonia: roles of orogeny, glaciation, and volcanism.

We used molecular evidence to examine the roles that vicariance mechanisms (mountain-building and drainage changes during the Pleistocene) have played in producing phylogeographical structure within and among South American fish species of the temperate perch family Percichthyidae. The percichthyids include two South American genera, Percichthys and Percilia, each containing several species, all of which are endemic to southern Argentina and Chile (Patagonia). Maximum-likelihood phylogenies constructed using mitochondrial DNA (mtDNA) control region haplotypes and nuclear GnRH3-2 intron allele sequences support the current taxonomy at the genus level (both Percichthys and Percilia form strongly supported, monophyletic clades) but indicate that species-level designations need revision. Phylogeographical patterns at the mtDNA support the hypothesis that the Andes have been a major barrier to gene flow. Most species diversity occurs in watersheds to the west of the Andes, together with some ancient divergences among conspecific populations. In contrast, only one species (Percichthys trucha) is found east of the Andes, and little to no phylogeographical structure occurs among populations in this region. Mismatch analyses of mtDNA sequences suggest that eastern populations last went through a major bottleneck c. 188 000 bp, a date consistent with the onset of the penultimate and largest Pleistocene glaciation in Patagonia. We suggest that eastern populations have undergone repeated founder-flush events as a consequence of glacial cycles, and that the shallow phylogeny is due to mixing during recolonization periods. The area of greater diversity west of the Andes lies outside the northern limit of the glaciers. mtDNA mismatch analysis of the genus Percilia which is restricted to this area suggests a long-established population at equilibrium. We conclude that patterns of genetic diversity in these South American genera have been primarily influenced by barriers to gene flow (Andean orogeny, and to a lesser extent, isolation in river drainages), and by glacial cycles, which have resulted in population contraction, re-arrangement of some watersheds, and the temporary breakdown of dispersal barriers among eastern river systems.