Karyotype diversity and evolutionary trends in the Asian swamp eel Monopterus albus (Synbranchiformes, Synbranchidae): a case of chromosomal speciation?

BACKGROUND: Synbranchidae or swamp eels are fishes belonging to the order Synbranchiformes that occur in both freshwater and occasionally in brackish. They are worldwide distributed in tropical and subtropical rivers of four different continents. A large degree of chromosomal variation has been found in this family, mainly through the use of conventional cytogenetic investigations. Inside this group, a still almost unexplored species under the cytogenetic point of view is the Asian swamp eel Monopterus albus, a widely distributed species throughout Asia. Here, we tested the hypothesis of chromosomal speciation, where a case of sympatric speciation may occur as the primary consequence of chromosomal rearrangements. We performed a comparative chromosomal analysis of M. albus from 22 different localities in Thailand, using distinct staining methods (C-banding, Ag-NO3, and Chromomycin A3), and FISH with repetitive DNA probes (5S rDNA, 18S rDNA, Rex1 element and microsatellite repeats). RESULTS: This approach evidenced two contrasting karyotypes (named karyomorphs A and B) that varied concerning their 2n and repetitive DNAs distribution, where chromosomal fusions and pericentric inversions were involved in such differentiation. While the karyomorph A has 2n = 24 chromosomes, the karyomorph B has only 2n = 18, both with NF = 24. In addition, karyomorph A contains only acrocentric chromosomes, while karyomorph B contains three unique metacentric pairs. These features highlight that M. albus has already gone through a significant genomic divergence, and may include at least two cryptic species. CONCLUSIONS: This marked chromosomal differentiation, likely linked to the lifestyle of these fishes, point to the occurrence of a chromosomal speciation scenario, in which fusions and inversions had a prominent role. This highlights the biodiversity of M. albus and justifies its taxonomic revision, since this nominal species may constitute a species complex.

Sex Chromosome Evolution and Genomic Divergence in the Fish Hoplias malabaricus (Characiformes, Erythrinidae).

The Erythrinidae family (Teleostei: Characiformes) is a small Neotropical fish group with a wide distribution throughout South America, where Hoplias malabaricus corresponds to the most widespread and cytogenetically studied taxon. This species possesses significant genetic variation, as well as huge karyotype diversity among populations, as reflected by its seven major karyotype forms (i.e., karyomorphs A-G) identified up to now. Although morphological differences in their bodies are not outstanding, H. malabaricus karyomorphs are easily identified by differences in 2n, morphology and size of chromosomes, as well as by distinct evolutionary steps of sex chromosomes development. Here, we performed comparative genomic hybridization (CGH) to analyse both the intra- and inter-genomic status in terms of repetitive DNA divergence among all but one (E) H. malabaricus karyomorphs. Our results indicated that they have close relationships, but with evolutionary divergences among their genomes, yielding a range of non-overlapping karyomorph-specific signals. Besides, male-specific regions were uncovered on the sex chromosomes, confirming their differential evolutionary trajectories. In conclusion, the hypothesis that H. malabaricus karyomorphs are result of speciation events was strengthened.

First Chromosomal Analysis in Hepsetidae (Actinopterygii, Characiformes): Insights into Relationship between African and Neotropical Fish Groups.

Hepsetidae is a small fish family with only the genus Hepsetus, with six described species distributed throughout the South, Central and Western regions of Africa, showing a close relationship with the Alestidae and some Neotropical fish families. However, no cytogenetic information is available for both Hepsetidae and Alestidae species, thus preventing any evolutionary comparative studies at the chromosomal level. In the present study, we are providing new cytogenetic data for Hepsetus odoe, including the standard karyotype, C-banding, repetitive DNAs mapping, comparative genomic hybridization (CGH) and whole chromosome painting (WCP), providing chromosomal patterns and subsidies for comparative cytogenetics with other characiform families. Both males and females H. odoe have 2n = 58 chromosomes (10m + 28sm + 20st/a), with most of the C-band positive heterochromatin localized in the centromeric and subtelomeric regions. Only one pair of chromosomes bears proximal 5S rDNA sites in the short arms, contrasting with the 18S rDNA sequences which are located in the terminal regions of four chromosome pairs. Clear interstitial hybridization signals are evidenced for the U1 and U2 snDNA probes, but in only one and two chromosome pairs, respectively. Microsatellite motifs are widely distributed in the karyotype, with exception for the (CGG)10, (GAA)10 and (GAG)10 probes, which highlight conspicuous interstitial signals on an unique pair of chromosomes. Comparative data from conventional and molecular cytogenetics, including CGH and WCP experiments, indicate that H. odoe and some Erythrinidae species, particularly Erythrinus erythrinus, share similar chromosomal sequences suggesting some relatedness among them, although bearing genomic specificities in view of their divergent evolutionary histories.

Karyotype and Mapping of Repetitive DNAs in the African Butterfly Fish Pantodon buchholzi, the Sole Species of the Family Pantodontidae.

The monophyletic order Osteoglossiformes represents one of the most ancestral groups of teleosts and has at least 1 representative in all continents of the southern hemisphere, with the exception of Antarctica. However, despite its phylogenetic and biogeographical importance, cytogenetic data in Osteoglossiformes are scarce. Here, karyotype and chromosomal characteristics of the lower Niger River population of the African butterfly fish Pantodon buchholzi, the sole species of the family Pantodontidae (Osteoglossiformes), were examined using conventional and molecular cytogenetic approaches. All specimens examined had 2n = 46 chromosomes, with a karyotype composed of 5 pairs of metacentric, 5 pairs of submetacentric, and 13 pairs of acrocentric chromosomes in both sexes. No morphologically differentiated sex chromosomes were identified. C-bands were located in the centromeric/pericentromeric region of all chromosomes and were associated with the single AgNOR site. FISH with ribosomal DNA probes revealed that both 5S and 18S rDNA were present in only 1 pair of chromosomes each, but did not colocalize. CMA3+ bands were observed near the telomeres in several chromosome pairs and also at the 18S rDNA sites. The mapping of di- and trinucleotide repeat motifs, Rex6 transposable element, and U2 snRNA showed a scattered distribution over most of the chromosomes, but for some microsatellites and the U2 snRNA also a preferential accumulation at telomeric regions. This study presents the first detailed cytogenetic analysis in the African butterfly fish by both conventional and molecular cytogenetic protocols. This is the first of a series of further cytogenetic and cytogenomic studies on osteoglossiforms, aiming to comprehensively examine the chromosomal evolution in this phylogenetically important fish order.

Hypermethylated Chromosome Regions in Nine Fish Species with Heteromorphic Sex Chromosomes.

Sites and amounts of 5-methylcytosine (5-MeC)-rich chromosome regions were detected in the karyotypes of 9 Brazilian species of Characiformes fishes by indirect immunofluorescence using a monoclonal anti-5-MeC antibody. These species, belonging to the genera Leporinus, Triportheus and Hoplias, are characterized by highly differentiated and heteromorphic ZW and XY sex chromosomes. In all species, the hypermethylated regions are confined to constitutive heterochromatin. The number and chromosome locations of hypermethylated heterochromatic regions in the karyotypes are constant and species-specific. Generally, heterochromatic regions that are darkly stained by the C-banding technique are distinctly hypermethylated, but several of the brightly fluorescing hypermethylated regions merely exhibit moderate or faint C-banding. The ZW and XY sex chromosomes of all 9 analyzed species also show species-specific heterochromatin hypermethylation patterns. The analysis of 5-MeC-rich chromosome regions contributes valuable data for comparative cytogenetics of closely related species and highlights the dynamic process of differentiation operating in the repetitive DNA fraction of sex chromosomes.