A Bird's-Eye View of Endangered Species Conservation: Avian Genomics and Stem Cell Approaches for Green Peafowl (Pavo muticus).

Aves ranks among the top two classes for the highest number of endangered and extinct species in the kingdom Animalia. Notably, the IUCN Red List classified the green peafowl as endangered. This highlights promising strategies using genetics and reproductive technologies for avian wildlife conservation. These platforms provide the capacity to predict population trends and enable the practical breeding of such species. The conservation of endangered avian species is facilitated through the application of genomic data storage and analysis. Storing the sequence is a form of biobanking. An analysis of sequence can identify genetically distinct individuals for breeding. Here, we reviewed avian genomics and stem cell approaches which not only offer hope for saving endangered species, such as the green peafowl but also for other birds threatened with extinction.

Dosage compensation of Z sex chromosome genes in avian fibroblast cells.

In birds, sex is genetically determined; however, the molecular mechanism is not well-understood. The avian Z sex chromosome (chrZ) lacks whole chromosome inactivation, in contrast to the mammalian chrX. To investigate chrZ dosage compensation and its role in sex specification, we use a highly quantitative method and analyze transcriptional activities of male and female fibroblast cells from seven bird species. Our data indicate that three fourths of chrZ genes are strictly compensated across Aves, similar to mammalian chrX. We also present a complete list of non-compensated chrZ genes and identify Ribosomal Protein S6 (RPS6) as a conserved sex-dimorphic gene in birds.

Establishment of fishing cat cell biobanking for sustainable conservation.

The fishing cat (Prionailurus viverrinus) is a vulnerable wild felid that is currently under threat from habitat destruction and other human activities. The zoo provides insurance to ensure the survival of the fishing cat population. Creating a biobank of fishing cats is a critical component of recent zoo strategies for securely stocking cell samples for long-term survival. Here, our goal was to compare cell biobanking techniques (tissue collection, primary culture, and reprogramming) and tissue sources (ear skin, abdominal skin, testis) from captive (n = 6)/natural (n = 6) vs. living (n = 8)/postmortem (n = 4) fishing cats. First, we show that dermal fibroblasts from the medial border of the helix of the ear pinna and abdominal tissues of living fishing cats can be obtained, whereas postmortem animals provided far fewer fibroblasts from the ears than from the testes. Furthermore, we can extract putative adult spermatogonial stem cells from the postmortem fishing cat's testes. The main barrier to expanding adult fibroblasts was early senescence, which can be overcome by overexpressing reprogramming factors through felid-specific transfection programs, though we demonstrated that reaching iPSC state from adult fibroblasts of fishing cats was ineffective with current virus-free mammal-based induction approaches. Taken together, the success of isolating and expanding primary cells is dependent on a number of factors, including tissue sources, tissue handling, and nature of limited replicative lifespan of the adult fibroblasts. This study provides recommendations for tissue collection and culture procedures for zoological research to facilitate the preservation of cells from both postmortem and living felids.

Evolutionary origin of vertebrate OCT4/POU5 functions in supporting pluripotency.

The support of pluripotent cells over time is an essential feature of development. In eutherian embryos, pluripotency is maintained from naïve states in peri-implantation to primed pluripotency at gastrulation. To understand how these states emerged, we reconstruct the evolutionary trajectory of the Pou5 gene family, which contains the central pluripotency factor OCT4. By coupling evolutionary sequence analysis with functional studies in mouse embryonic stem cells, we find that the ability of POU5 proteins to support pluripotency originated in the gnathostome lineage, prior to the generation of two paralogues, Pou5f1 and Pou5f3 via gene duplication. In osteichthyans, retaining both genes, the paralogues differ in their support of naïve and primed pluripotency. The specialization of these duplicates enables the diversification of function in self-renewal and differentiation. By integrating sequence evolution, cell phenotypes, developmental contexts and structural modelling, we pinpoint OCT4 regions sufficient for naïve pluripotency and describe their adaptation over evolutionary time.

Avian Embryonic Culture: A Perspective of In Ovo to Ex Ovo and In Vitro Studies.

The avian embryos growing outside the natural eggshell (ex ovo) were observed since the early 19th century, and since then chick embryonic structures have revealed reaching an in-depth view of external and internal anatomy, enabling us to understand conserved vertebrate development. However, the internal environment within an eggshell (in ovo) would still be the ideal place to perform various experiments to understand the nature of avian development and to apply other biotechnology techniques. With the advent of genetic manipulation and cell culture techniques, avian embryonic parts were dissected for explant culture to eventually generate expandable cell lines (in vitro cell culture). The expansion of embryonic cells allowed us to unravel the transcriptional network for understanding pluripotency and differentiation mechanism in the embryos and in combination with stem cell technology facilitated the applications of avian culture to the next levels in transgenesis and wildlife conservation. In this review, we provide a panoramic view of the relationship among different cultivation platforms from in ovo studies to ex ovo as well as in vitro culture of cell lines with recent advances in the stem cell fields.

Reproductive biology and biotechnologies in wild felids.

Conservation strategies in natural habitats as well as in breeding centers are necessary for maintaining and reinforcing viable populations of wild felids. Among the fundamental knowledge that is required for conservation breeding, a solid understanding of reproductive biology is critical for improving natural breeding and enhance genetic diversity. Additionally, it offers the opportunity to develop assisted reproductive technologies (ARTs) in threatened and endangered species. Conservation breeding and reproductive biotechnologies of wild felids have advanced in the past decade. It has been clearly shown that female felids have species and individual patterns of reproductive cycles and respond differently to exogenous hormones. In males, several species still have poor semen quality often due to the loss of genetic diversity in small populations. To overcome the challenges of natural breeding (incompatibility between individuals or suboptimal environment) and mitigate inbreeding, artificial insemination, embryo production and embryo transfer have been further developed in 24 wild cat species. Major factors limiting ART success are inconsistent responses to ovarian stimulation, variable quality of gametes and embryos, and preparation of recipient females. Additional approaches including stem cell technologies have been explored for future medical applications. However, there still is a critical need for better knowledge of feline reproductive biology and improvement of ARTs efficiency to increase the genetic diversity and create sustainable populations of wild felids.

Discovery of Neonrosellavitiata (Darwin) and Newmanellaspinosus Chan & Cheang (Balanomorpha, Tetraclitidae) from the Andaman Sea, eastern Indian Ocean.

In this present study, distantly related acorn barnacle species in the subfamily Newmanellinae (Cirripedia, Thoracica, Tetraclitidae), including Neonrosellavitiata (Darwin, 1854) and Newmanellaspinosus Chan & Cheang, 2016, were discovered in the Andaman Sea of Thailand. Neo.vitiata can be readily distinguished from other newmanellids by shell plate and operculum morphology (external shell, tergum geometry, and pattern of parietal tube) and arthropodal characters (presence of basi-dorsal point at base of penis and triangular spines on cirri, setal type, and mouth parts). Both species were found to share overlapping territories on rocks at the rockweed zone, an area submerged under seawater most of the time throughout the year. This study highlights the first discovery of Neonrosella in the eastern Indian Ocean, whose ultrastructure compared to Newmanella is redescribed and illustrated here based on scanning electron microscopy.

A conserved Oct4/POUV-dependent network links adhesion and migration to progenitor maintenance.

BACKGROUND: The class V POU domain transcription factor Oct4 (Pou5f1) is a pivotal regulator of embryonic stem cell (ESC) self-renewal and reprogramming of somatic cells to induced pluripotent stem (iPS) cells. Oct4 is also an important evolutionarily conserved regulator of progenitor cell differentiation during embryonic development. RESULTS: Here we examine the function of Oct4 homologs in Xenopus embryos and compare this to the role of Oct4 in maintaining mammalian embryo-derived stem cells. Based on a combination of expression profiling of Oct4/POUV-depleted Xenopus embryos and in silico analysis of existing mammalian Oct4 target data sets, we defined a set of evolutionary-conserved Oct4/POUV targets. Most of these targets were regulators of cell adhesion. This is consistent with Oct4/POUV phenotypes observed in the adherens junctions in Xenopus ectoderm, mouse embryonic, and epiblast stem cells. A number of these targets could rescue both Oct4/POUV phenotypes in cellular adhesion and multipotent progenitor cell maintenance, whereas expression of cadherins on their own could only transiently support adhesion and block differentiation in both ESC and Xenopus embryos. CONCLUSIONS: Currently, the list of Oct4 transcriptional targets contains thousands of genes. Using evolutionary conservation, we identified a core set of functionally relevant factors that linked the maintenance of adhesion to Oct4/POUV. We found that the regulation of adhesion by the Oct4/POUV network occurred at both transcriptional and posttranslational levels and was required for pluripotency.