Paraphyly and evolutionary independent lineages in Gymnotus pantherinus (Gymnotiformes: Gymnotidae) in the Brazilian Atlantic Forest Coastal Streams.

In the Brazilian Atlantic Forest (AF), many terrestrial species with broad geographical distributions show high diversity and endemism of intraspecific lineages, as revealed by molecular genetic data. This pattern, however, is less explored in freshwater fishes. Gymnotus pantherinus is an electric fish endemic to the Brazilian coastal drainages that shows a wide distribution, ranging from the states of Bahia to Santa Catarina, an unusual pattern for AF fishes. It has been hypothesized that G. pantherinus is a species complex because distinct morphotypes were described for the species based on morphometric and meristic features. We used mitochondrial and nuclear data to test this hypothesis. Based on phylogenetic inference and multi-locus, multispecies coalescent methods, we identified six independent lineages, flagging them as candidate species. One such lineage is the recently described species G. refugio that is nested within G. pantherinus and renders it paraphyletic, showing it is a species complex. We named G. pantherinus stricto sensu the lineage that includes samples from the type locality (Santos, SP). Our results show that genetic lineages correspond only partially and far exceed the number of previously reported morphotypes. Genetic breaks in the group correspond to landscape features associated with the Serra do Mar mountain range and with riverine dynamics caused by sea level changes during the last glacial maximum. Moreover, we found evidence of river capture events affecting phylogeographic structure in the group. We uncovered an important dimension of diversity in the group and encourage further integration of genetic and phenotypic data. Such integration is a fruitful approach not only to reduce the gap between taxonomy and evolutionary history in Gymnotidae, but also to uncover the real AF biodiversity.

Mixed-stock analysis in green turtles Chelonia mydas: mtDNA decipher current connections among west Atlantic populations.

The green turtle Chelonia mydas undertakes wide-ranging migrations between feeding and nesting sites, resulting in mixing and isolation of genetic stocks. We used mtDNA control region to characterize the genetic composition, population structure, and natal origins of C. mydas in the West Atlantic Ocean, at one feeding ground (State of Rio de Janeiro, Brazil), and three Caribbean nesting grounds (French Guiana, Guadeloupe, and Suriname). The feeding ground presented considerable frequency of common haplotypes from the South Atlantic, whereas the nesting sites presented a major contribution of the most common haplotype from the Caribbean. MSA revealed multiple origins of individuals at the feeding ground, notably from Ascension Island, Guinea Bissau, and French Guiana. This study enables a better understanding of the dispersion patterns and highlights the importance of connecting both nesting and feeding areas. Effective conservation initiatives need to encompass these ecologically and geographically distinct sites as well as those corridors connecting them.

Cytogenetic and molecular analyses in troglobitic and epigean species of Pimelodella (Siluriformes: Heptapteridae) from Brazil

Samples from seven different locations of the genus Pimelodella were genetically examined, two caves (exclusively subterranean, upper Tocantins River and São Francisco River) and five epigean (from upper Paraná River basin). Cytogenetic analyses revealed the same diploid number (2n=46) for all species besides similarities in both number and location of nucleolar organizer regions and C bands. FISH with 5S rDNA probes and CMA3 staining indicated significant differences among the studied species. Application of PCR-RFLP in ATPase 6 and 8 mitochondrial genes allowed building a minimum evolution phenogram identifying the close evolutionary relationship among groups. Both chromosomal and molecular data were useful to infer the relationships among studied Pimelodella species.

Amostras de sete diferentes localidades do gênero Pimelodella foram geneticamente analisadas, duas cavernícolas (exclusivamente subterrâneas, alto rio Tocantins e rio São Francisco) e cinco epígeas (provenientes da bacia do alto Paraná). Análises citogenéticas revelaram o mesmo número diploide (2n=46) para todas as espécies, além de similaridades no número e localização das regiões organizadoras de nucléolo e bandas C. FISH com sondas de rDNA 5S e marcação com CMA3 indicaram diferenças significativas entre as espécies estudadas. A aplicação da técnica de PCR-RFLP nos genes mitocondriais ATPase 6 e 8 permitiu a construção de um fenograma de evolução mínima identificando uma estreita relação evolutiva entre as espécies estudadas.

Astyanax hastatus Myers, 1928 (Teleostei, Characidae): a new species complex within the genus Astyanax?

Four populations of Astyanax hastatus Myers 1928 from the Guapimirim River basin (Rio de Janeiro State) were analyzed and three distinct cytotypes identified. These cytotypes presented 2n = 50 chromosomes, with 4M+8SM+10ST+28A (Cytotype A), 8M+10SM+14ST+18A (Cytotype B), 6M+8SM+4ST+32A (Cytotype C) and scanty heterochromatin, mainly located throughout pericentromeric regions of several chromosomal pairs. No homologies with the As-51 satellite DNA were observed in the three cytotypes, although all of them presented multiple 18S rDNA sites, as detected by both silver nitrate staining and FISH (fluorescent in situ hybridization). The application of the term "species complex" in Astyanax is discussed from a cytotaxonomic viewpoint.

Cytogenetic characterization of the strongly electric Amazonian eel, Electrophorus electricus (Teleostei, Gymnotiformes), from the Brazilian rivers Amazon and Araguaia

A karyotype analysis of the electric eel, Electrophorus electricus (Teleostei, Gymnotiformes), a strongly electric fish from northern South America, is presented. Two female specimens were analyzed, one from the Amazon River and one from the Araguaia River. The specimens had a chromosomal number of 2n = 52 (42M-SM + 10A). C-bands were present in a centromeric and pericentromeric position on part of the chromosomes; some interstitial C-bands were also present. Heteromorphic nucleolus organizer regions (NORs) were detected in two chromosome pairs of the specimen from the Amazon River. The chromosome number and karyotype characteristics are similar to those of other Gymnotidae species. The genera Electrophorus and Gymnotus are positioned as the basal lineages in the Gymnotiformes phylogeny.

Use of chromosome microdissection in fish molecular cytogenetics

Chromosome microdissection is a technique in which whole chromosomes or chromosomal segments are dissected under an inverted microscope yielding chromosome-specific sequences. Several protocol modifications introduced during the past 15 years reduced the number of chromosomes required for most applications. This is of particular interest to fish molecular cytogenetics, since most species present highly uniform karyotypes which make impossible the collection of multiple copies of the same chromosome. Probes developed in this manner can be used to investigate chromosome homologies in closely related species. Here we describe a protocol recently used in the gymnotiform species group Eigenmannia and review the major steps involved in the generation of these markers focusing on protocol modifications aiming to reduce the number of required chromosomes.