Investigation of the Guinea fowl and domestic fowl hybrids as potential surrogate hosts for avian cryopreservation programmes

Mariann Molnár,Krisztina Liptói,Bence Lázár,Nikoletta Sztán,Eszter Patakiné Várkonyi,Michael J Mcgrew,Barbara Végi,Miklós Marosán,Árpád Drobnyák,Sunil Nandi,Roland Tóth,Elen Gócza

doi:10.1038/s41598-019-50763-3

Abstract

In the last decade, avian gene preservation research has focused on the use of the early precursors of the reproductive cells, the primordial germ cells (PGCs). This is because avian PGCs have a unique migration route through the vascular system which offers easy accessibility. Furthermore, culturing of the cells in vitro, freezing/thawing, reintegration into a recipient embryo and the development of the germ cells can be carried out in well-defined laboratory circumstances. The efficient recovery of the donor genotype and the frequency of germline transmission from the surrogate host animals are still areas which need further development. Thus, the aim of the present study was to investigate an infertile interspecific hybrid (recipient) as an appropriate host for primordial germ cells from native poultry breeds. Guinea fowl × chicken hybrids were produced, the crossing was repeated inversely. The phenotype, the hatching time, the hatching rate, the sex ratio, the presence of own germ cells, the fertility and the phenotype of viable hybrids and the incidence of chromosomal abnormalities of dead hybrid embryos were described. 6.65% viable offspring was obtained with crossing of Guinea fowl females with domestic fowl males. Crossing of domestic fowl hens with Guinea fowl male resulted in lower fertility, 0.14% viable offspring. Based on the investigations, the observed offspring from the successful crossing were sterile male hybrids, thus an extreme form of Haldane’s rule was manifested. The sterile hybrid male embryos were tested by injecting fluorescently labeled chicken PGCs. The integration rate of labeled PGCs was measured in 7.5-day, 14.5-day and 18.5-day old embryonic gonads. 50%, 5.3% and 2.4% of the injected hybrid embryos survived and 40%, 5.3% and 2.4% of the examined gonads contained fluorescent labeled donor PGCs. Therefore, these sterile hybrid males may be suitable recipients for male PGCs and possibly for female PGCs although with lower efficiency. This research work shows that the sterility of hybrids can be used in gene conservation to be a universal host for PGCs of different avian species.

Highlights

According to the International Union for Conservation of Nature and Natural Resources 14% of avian species are listed as threatened with extinction[1]
primordial germ cells (PGCs) have unique characteristics and accessibility. They migrate through the vascular system to colonize the developing gonads; they can be isolated from and reintegrated into the embryonic circulatory system[6] with the goal of production of germline chimeras. These germline chimeras, ‘surrogate hosts’, can be bred and will give rise to a progeny carrying the genome of the donor PGCs
Three different phenotypes were observed in the hybrids: a dark brown (Fig. 2d), a light brown (Fig. 2e) and a white spotted type (Fig. 2f), but none of the hybrids had helmet, crest or facial wattles which distinguished them from pure Guinea fowl

Summary

Introduction

According to the International Union for Conservation of Nature and Natural Resources 14% of avian species are listed as threatened with extinction[1]. They migrate through the vascular system to colonize the developing gonads; they can be isolated from and reintegrated into the embryonic circulatory system[6] with the goal of production of germline chimeras. The first successful experiments aiming this were based on UV7 or χ-radiation[10,11,12,13,14] Using these techniques, more donor-derived PGCs colonized the recipient gonads, but the development of the treated embryos fell behind the non-treated control embryos[10]. Researchers developed a technique during which the busulfan is delivered directly to the embryo by injection of an emulsion[20] The effectiveness of this method was a significant improvement; donor cells colonized the recipient gonads with 99.5% success and germline transmission of donor PGCs was improved[21]. As previously described in numerous studies, hybrids between different species are often sterile dependent on the phylogenetic distance between them

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