Hibridação natural entre duas espécies de lambaris na cabeceira do Rio Grande e suas consequências na gametogênese e morfologia gonadal dos híbridos

Abstract

Hybridization is a relevant process for ecology, conservation and evolution of species, but its occurrence and consequences on the biology of hybrids are still poorly studied. In addition, although it is an event that frequently leads to gonadal changes in the hybrids, the morphology of their gonads and the cellular pathways involved in these changes have been poorly investigated. In this sense, the first aim of this study was to test the hypothesis that the two lambaris species A. fasciatus and A. paranae, living in sympatry at the headwaters of Grande river, are able to interbreed in the natural environment and generate hybrids. For this, we used a multidisciplinary approach that involved methods of morphometric and meristic analysis of fish, study of the reproductive biology and gonadal morphology, as well as sequencing of fragments of mitochondrial (COI) and nuclear (S7) genes. The anatomical analysis showed intermediate characteristics in the putative hybrids, while the parental species showed exclusive characters. The study of reproductive biology and gametogenesis showed that A. fasciatus and A. paranae present overlap of reproductive period, similarity in the morphology of gonads and gametes, as well as reproductive strategies. Genetic analyzes supported the hybridization hypothesis, since they showed the existence of two distinct species and an intermediate group, with mitochondrial DNA from both species, despite having a single morphotype and nuclear DNA with double peaks (coinciding with those of species A. fasciatus and A. paranae) in the sequence chromatogram, indicating heterozygosity. Then, the aim was to explore the gonadal morphology of the hybrids and this analysis showed several changes such as absence of spermatids and spermatozoa in males and previtellogenic and vitellogenic oocytes in females, vacuolization and degeneration of germ cells, increased interstitial tissue and inflammatory infiltrate, loss of the organization of germ cells in cysts or nests; all these data indicated the infertility of the hybrids. Finally, the last aim was to test the hypothesis that autophagy, apoptosis and extracellular matrix proteins are involved in the germ cell degeneration process and tissue remodeling in the gonads of hybrids. For this, we immunolocalized the proteins Beclin1, CathepsinD, Caspase3, Fibronectin and Collagen type I. The quantification of all analyzed proteins showed that gonads of hybrids exhibit a larger marked area for these molecules compared to the parental species. These results indicate that the morphological changes in the gonads of the infertile hybrids culminate in autophagy and catpsinD-mediated lisossomal degradation, followed by apoptosis and tissue remodeling with fibrosis.