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Preliminary study on testicular germ cell isolation and transplantation in an endangered endemic species Brycon orbignyanus (Characiformes: Characidae)

  • Diógenes Henrique de Siqueira-SilvaEmail author
  • Amanda Pereira dos Santos Silva
  • Raphael da Silva Costa
  • José Augusto Senhorini
  • Alexandre Ninhaus-Silveira
  • Rosicleire Veríssimo-Silveira
Article

Abstract

We aimed to develop a simplified protocol for transplantation of Brycon orbignyanus spermatogonial stem cells (SSCs) into Astyanax altiparanae testes. Brycon orbignyanus testes were enzymatically digested and SSC purified by a discontinuous density gradient. Endogenous spermatogenesis was suppressed in A. altiparanae using busulfan or by incubation at 35 °C water, and SSCs from B. orbignyanus labeled with PKH26 were injected into their testes via the urogenital papilla. Twenty-two hours post-transplantation, labeled spermatogonia were observed in A. altiparanae tubular lumen. After 7 days, spermatogonia proliferated in the epithelium, and 21 days post-transplantation, sperm was observed in the lumen. Of surviving host fish, nearly 67% of those treated with busulfan and 85% of those held in warm water showed labeled cells in host germinal epithelium. The present study standardized, by a simple and accessible method, germ cell transplantation between sexually mature Characiformes fish species. This is the first report of xenogenic SSC transplantation in this fish order.

Keywords

Fish chimera Germ cell technology Spermatogonial proliferation Surrogate production 

Notes

Acknowledgments

The authors are deeply grateful to Douglas de Castro Ribeiro for help constructing the aquarium system and to Usina Hidrelétrica Engenheiro Souza Dias (Jupiá-CESP) and Centro Nacional de Pesquisa e Conservação da Biota Aquática Continental (CEPTA) for providing the fish used in this study.

Funding information

This study was supported by Fundação de Apoio à Pesquisa de São Paulo (FAPESP; Process number: 2013/24218-5) and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq; grant number: 160-380/2011).

Compliance with ethical standards

Experimental procedures were conducted in accordance with the guidance for care and use of animals in the laboratory of Universidade Estadual Paulista (UNESP-Ilha Solteira). The Committee of Ethics and Research of UNESP approved the protocol 006/2012/CONCEA.

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Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  • Diógenes Henrique de Siqueira-Silva
    • 1
    • 2
    • 3
    Email author
  • Amanda Pereira dos Santos Silva
    • 2
  • Raphael da Silva Costa
    • 2
  • José Augusto Senhorini
    • 4
  • Alexandre Ninhaus-Silveira
    • 1
    • 2
  • Rosicleire Veríssimo-Silveira
    • 1
    • 2
  1. 1.Campus de Ilha Solteira, Departament of Biology and Animal Science, L.I.NEO – Laboratório de Ictiologia NeotropicalUNESP – Univ. Estadual PaulistaIlha SolteiraBrazil
  2. 2.Campus de São José do Rio Preto, Post-Graduation Program in Animal BiologyUNESP – Univ. Estadual PaulistaSão José do Rio PretoBrazil
  3. 3.Campus de Ilha Solteira, Departamento de Biologia e Zootecnia, L.I.NEO - Laboratório de Ictiologia NeotropicalUNESP –Univ. Estadual PaulistaSao PauloBrazil
  4. 4.CEPTA-ICMBIO - National Center for Research and Conservation of Continental FishChico Mendes Institute of Biodiversity ConservationPirassunungaBrazil

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