Assessing paternity in japanese quails (Coturnix japonica) using microsatellite markers - inferences for its mating system and reproductive success

Microsatellite markers were analyzed in Japanese quails, Coturnix japonica, using different methodologies (PAGE and automated genotyping), in order to evaluate their use in paternity testing. Ten animal triplets composed by a female and two males were used to mate and generate an offspring. Paternity was determined in five-day-old embryos, and the data generated by fluorescent labeled and tailored primers in PCR and further automated genotyping were robust. Three microsatellite markers were polymorphic (Na = 5-8, H E = 0.75) and no loci were found to deviate significantly from Hardy-Weinberg equilibrium or showed any evidence of linkage disequilibrium (p > 0.05). A slight heterozygote deficiency and some incompatibilities between the female known parent and its offspring that involved homozygous genotypes were observed at GUJ0001 locus and may indicate the presence of null alleles. Although a reduced set of microsatellite primers were applied, it was possible to determine the paternity of 96.87% of the embryos, using combined data of three loci. The approach was useful for parentage inferring in a captive population of C. japonica and the results evidenced a potential polyandric mating system in the species, in which no advantage mechanism of last-male sperm precedence seems to occur.

Cytogenetic analyses of two Curimatidae species (Pisces; Characiformes) from the Paranapanema and Tietê Rivers

Cytogenetic analyses were performed in two Curimatidae species (Steindachnerina insculpta and Cyphocharax modesta) from the Paranapanema and Tietê Rivers (São Paulo State, Brazil), showing a karyotype composed of 54 meta-submetacentric chromosomes in both species. Silver- and chromomycyn-staining and fluorescent in situ hybridization (FISH) using a 18S rDNA probe indicated that the nucleolar organizer regions (NORs) of both species are localized in the terminal region of the long arm of two metacentric chromosomes. Although a single NOR system was evidenced in both analyzed species, S. insculpta and C. modesta presented the nucleolar organizer regions in distinct chromosome pairs, indicating that these cistrons can be considered cytogenetic markers. Variation on the amount and distribution of the constitutive heterochromatin (C-bands) could also be detected between the two species - while S. insculpta presented few heterochromatic blocks, intensely stained C-bands were evidenced in C. modesta specially in the terminal region of the long arm of the NOR-bearing chromosomes. Although most Curimatidae species have been characterized by homogeneous karyotypes, isolated populations could be established under different environmental conditions leading to karyotype micro-structure variations specially related to the NORs localization and C-banding distribution. The obtained data were useful for the cytogenetic characterization and differentiation of S. insculpta and C. modesta and could be used in evolutionary inferences in the Curimatidae group.

Análises citogenéticas foram realizadas em duas espécies de Curimatidae (Steindachnerina insculpta e Cyphocharax modestus) provenientes dos rios Paranapanema e Tietê (Estado de São Paulo, Brasil), evidenciando um cariótipo composto por 54 cromossomos meta-submetacêntricos em ambas as espécies. Coloração com nitrato de prata e cromomicina e hibridação in situ fluorescente (FISH), utilizando uma sonda de DNAr 18S, mostraram que as regiões organizadoras de nucléolos (RONs) de ambas as espécies estão localizadas na região terminal do braço longo de dois cromossomos metacêntricos. Embora as espécies analisadas tenham apresentado um sistema de RONs simples, S. insculpta e C. modesta apresentaram as regiões organizadoras de nucléolos em distintos pares de cromossomos, indicando que estes cístrons podem ser considerados marcadores citogenéticos. Variação na quantidade e distribuição de heterocromatina constitutiva (bandas C) também pôde ser detectada entre as duas espécies - enquanto S. insculpta apresentou poucos blocos heterocromáticos, bandas C intensamente coradas foram evidenciadas em C. modesta especialmente na região terminal do braço longo dos cromossomos portadores de RONs. Embora a maioria das espécies de Curimatidae seja caracterizada por cariótipos homogêneos, populações isoladas podem ter se estabelecido sob condições ambientais distintas, levando à ocorrência de variações na micro-estrutura cariotípica especialmente relacionadas à localização das RONs e à distribuição das bandas C. Os dados obtidos mostraram-se úteis para caracterização e diferenciação citogenética de S. insculpta e C. modesta e podem ser utilizados em inferências evolutivas no grupo Curimatidae.