Research Article |
Corresponding author: Karine Frehner Kavalco ( kavalco@ufv.br ) Academic editor: Inna Kuznetsova
© 2018 Karina de Oliveira Brandão, Dinaíza Abadia Rocha-Reis, Caroline Garcia, Rubens Pazza, Lurdes Foresti de Almeida-Toledo, Karine Frehner Kavalco.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Brandão KO, Rocha-Reis DA, Garcia C, Pazza R, Almeida-Toledo LF, Kavalco KF (2018) Studies in two allopatric populations of Hypostomus affinis (Steindachner, 1877): the role of mapping the ribosomal genes to understand the chromosome evolution of the group. Comparative Cytogenetics 12(1): 1-12. https://doi.org/10.3897/CompCytogen.v12i1.22052
|
Several cytogenetic markers show chromosomal diversity in the fish such as “armoured catfish”. Although studies have characterized many species in the major genera representing these Siluridae, particularly in the genus Hypostomus Lacépède, 1803, trends in chromosome evolution of this group remain unclear. The Paraíba do Sul river basin contains the armoured catfish Hypostomus affinis Steindachner, 1877, which is unique because of its distribution of repetitive DNAs, the 5S and 18S rDNA. Identified samples and registered collections in Brazilian museums were identified as the same typological species, while we observed wide variations in the physical location of this gene in the karyotype based on fluorescent in situ hybridization results. In this study, we propose that these species can represent evolutionarily independent units, as these fish frequently undergo processes such as dispersion and vicariance and that the rDNA is associated with DNA that spreads in the genome, such as transposons. Additionally, the absence of gene flow due to the distance of the sample location could intensify evolutionary processes. The phenotypes found for the 18S rDNA showed minor changes in relation to the number of sites between the lower and upper drainage regions of Paraíba do Sul. The large difference in the number of sites found for the 5S rDNA entered the same region (upper drainage of the basin) and the literature data could represent a population dynamics where an expansion of the 5S rDNA sites provides an extinct or non-sampled cytotype in this work.
Biodiversity, Catfish, Cytogenetics, Hypostominae , Teleostei
With a wide geographic distribution in nearly all of the Neotropical region from Costa Rica to Argentina, Loricariidae is considered one of the largest Neotropical fish families and the largest Family of catfishes (Siluriformes) (
The great diversity of armored catfish is also reflected in the available cytogenetic data of the group. Loricariidae exhibits large variations in diploid number, ranging from 2n = 36 chromosomes in Loricaria latirostris Boulenger, 1900 (
Most of this great diversity is related to the genus Hypostomus Lacépède, 1803, which contains approximately 200 valid species (
Fluorescent in situ hybridization (FISH) for localization of the 18S ribosomal RNA (18S rRNA) gene was one of the first cytogenetic-molecular markers applied in Neotropical fish (
In contrast, data for 5S ribosomal DNA (5S rDNA) are limited. This marker has been defined in only approximately a dozen species of the genus for some Neotropical populations (
Although Hypostomus affinis Steindachner, 1877 was found in the Mucuri and Doce river basin, most of the records are related to the Paraíba do Sul river, indicating a wide distribution of this species in this river basin (
Two populations of H. affinis were collected from Jacuí creek, Cunha/SP (-23.04052/-44.93408, Fig.
Hydrographic map of the southeast coast of Brazil with the collection points of Hypostomus affinis. Point “a” corresponds to Cunha/SP and point “b” corresponds to Itaocara/RJ. Hydrographic basins: Paraíba do Sul (in purple), São Francisco (in green), Upper Paraná (in red) and Rios Costeiros (in blue).
The chromosomal preparations were obtained from kidney cells of the animals as described by
The physical location of the ribosomal genes was detected via FISH (
Hybridization was detected with avidin and fluorescein isothiocyanate for 18S rDNA probes and Cy3 for 5S rDNA probes. Blade assembly was performed with antifade and propidium iodide, and antifade and DAPI for 18S rDNA and rDNA 5S probes, respectively. High-stringency washes with >75% (20% formamide/0.1× SSC) were performed for 15 min, and the signals were amplified using biotin-conjugated anti-avidin solution and incubated in non-fat dry milk buffer. Images were acquired with a camera coupled to an OLYMPUS BX41 microscope (Olympus Inc., Tokyo, Japan) using QCapture 6.0 (QImaging Surrey, BC, Canada) software.
To assemble the karyotypes, chromosomes were classified as metacentric (m), submetacentric (sm), subtelocentric (st), or acrocentric (a) according to the arm ratio proposed by
Male and female fish in both populations showed a diploid number of 2n = 66 chromosomes, karyotype composed of 12m+12m+14st+28a, and fundamental number FN = 104 (Fig.
C-banding staining of both populations revealed subtle pericentromeric markers on several chromosomes, as well as conspicuous terminal blocks on two pairs of acrocentric chromosomes (a) (25, 29), although a size heteromorphism was found in one of the pairs in the population of Itaocara (25) (Fig.
Silver nitrate staining of both populations revealed the existence of multiple systems of NORs. In Cunha/SP specimens, two pairs of chromosome a (21, 24) exhibited large markers on their long arms (Fig.
Hybridization of the 5S rDNA probe revealed two sites marked in the lowest metacentric pair of the complement in both populations (pair 6) (Fig.
From a cytogenetic perspective, only one sample of H. affinis from the Jacuí creek Cunha/SP has been previously studied (
Although the chromosome number is the same and the karyotypic formula observed in the populations studied slightly differs from the previously sampled population, other cytogenetic features revealed differentiated evolutionary units. The difference in karyotype symmetry observed between the chromosomes of both samplings from Cunha/SP were clear; this was also clear when the relative size of the chromosomes was organized based on type, even when the same measurement and classification criterion proposed by
The difference among the observed chromosomal sites in the populations cannot be attributed to the use of 18S and 5S ribosomal DNA probes isolated from different species. The rRNA in eukaryotes presents as two subunits (one formed by 28S, 18S and 5.8S and another one formed by 5S) and their DNA sequences vary very slowly due to selective pressure, being considered highly conserved (
For the location of 18S rDNA, we observed conservation of the number and position of sites in samples of the upper drainage region (
The existence of different chromosomal formulas in close groups of different organisms, or nominally similar species, is attributed to chromosomal rearrangements. In armored catfish, two major types of chromosomal rearrangements appear to be involved in karyotype differences, depending on fixation of the diploid number (non-Robertsonian) or their variation (Robertsonian) (
The divergent phenotype observed by
The speciation by allopatry can be an important source of diversity in Neotropics and could be responsible for the biodiversity of fishes from Brazilian rivers and it is possible that very short time periods can produce new phenotypes on Hypostomus chromosomes. At the same time, the extensive chromosomal variation observed in the sample of H. affinis analyzed previously by
Minor chromosome changes were found between the two sampled populations, especially regarded to an extra chromosome pair bearing 18S rDNA in population from Cunha. In addition, 18S rDNA distribution in Cunha was the same as previously sample. However, the remarkable difference in the 5S rDNA distribution between two sampling at Cunha, separated by four years between the collections, could represent a population dynamic where an expansion of the 5S rDNA sites provide a phenotype furtherly extinct or not sampled in this work.
This work was supported by the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq, 484626/2013-2) and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES - Fellowship for M.Sc. degree).