Molecular cytogenetic analysis and the establishment of a cell culture in the fish species Hollandichthys multifasciatus (Eigenmann &amp; Norris, 1900) (Characiformes, Characidae)

Letícia Batista Soares,Claudio Oliveira,Lucas Peres Ramos,Fausto Foresti,Fabilene Gomes Paim

doi:10.1590/1678-4685-gmb-2020-0260

Molecular cytogenetic analysis and the establishment of a cell culture in the fish species Hollandichthys multifasciatus (Eigenmann & Norris, 1900) (Characiformes, Characidae)

Letícia Batista Soares, Claudio Oliveira + Show 3 more

Open Access

https://doi.org/10.1590/1678-4685-gmb-2020-0260

Copy DOI

Abstract

Hollandichthys is a fish genus of the family Characidae that was until recently considered to be monotypic, with cytogenetic, morphological, and molecular data being restricted to a few local populations. In the present study, the karyotype of a population of Hollandichthys multifasciatus was analyzed using classical and molecular cytogenetic approaches for the investigation of potential markers that could provide new perspectives on the cytotaxonomy. H. multifasciatus presented a diploid number of 2n=50 chromosomes and a karyotype formula of 8m+10sm+32st. A single pair of chromosomes presented Ag-NORs signals, which coincided with the 18S rDNA sites visualized by FISH, whilst the 5S rDNA sequences were mapped in two chromosome pairs. The distribution of the U snRNA genes was mapped on the Hollandichthys chromosomes for the first time, with the probes revealing the presence of the U1 snDNA on the chromosomes of pair 20, U2 on pairs 6 and 19, U4 on pair 16, and U6 on the chromosomes of pair 11. The results of the present study indicated karyotypic differences in comparison with the other populations of H. multifasciatus studied previously, reinforcing the need for further research to identify isolated populations or the potential existence of cryptic Hollandichthys species.

Highlights

Characidae (Teleostei, Characiformes) is the most diverse family of Neotropical fishes, with approximately 1,200 smallbodied species (Fricke et al, 2020; Malabarba and Malabarba, 2020)
We performed a detailed analysis of a new unexplored population of this species, applying conventional and advanced techniques of molecular cytogenetics, that is, Fluorescence in Situ Hybridization (FISH) using six multigenic families (18S and 5S rDNA and U1, U2, U4 and U6 snDNA) and telomeric sequences (TTAGGG)n as probes
The cells in the primary cultures of H. multifasciatus attained confluence in 2–6 days (Figure 1a), and after subcultivation, they occupied the entire surface of the flasks in 2–4 days (Figure 1b)

Summary

Introduction

Characidae (Teleostei, Characiformes) is the most diverse family of Neotropical fishes, with approximately 1,200 smallbodied species (Fricke et al, 2020; Malabarba and Malabarba, 2020). The technology of cell culture, very widespread in several areas of Biology, is still little explored in fish cytogenetics, despite being an excellent alternative to obtain quality chromosomal preparations. This little use is mainly related to the difficulty in standardizing the isolation and maintenance techniques of these cell cultures (Amemiya et al, 1984; Bejar et al, 1997; Zhang et al, 1998; Wang et al, 2012; Paim et al, 2018). We performed a detailed analysis of a new unexplored population of this species, applying conventional and advanced techniques of molecular cytogenetics, that is, Fluorescence in Situ Hybridization (FISH) using six multigenic families (18S and 5S rDNA and U1, U2, U4 and U6 snDNA) and telomeric sequences (TTAGGG)n as probes

Material and Methods

Results and Discussion

Conflict of Interest