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

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.

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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

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).

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Correspondence to Diógenes Henrique de Siqueira-Silva.

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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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de Siqueira-Silva, D.H., dos Santos Silva, A.P., da Silva Costa, R. et al. Preliminary study on testicular germ cell isolation and transplantation in an endangered endemic species Brycon orbignyanus (Characiformes: Characidae). Fish Physiol Biochem (2019). https://doi.org/10.1007/s10695-019-00631-8

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Keywords

  • Fish chimera
  • Germ cell technology
  • Spermatogonial proliferation
  • Surrogate production