Robert Kenneth Browne

Intergenerational justice is the core principle supporting the legacy of benefit toward future generations, including the perpetuation of species and their genetic diversity, as a key component of biospheric sustainability. Thirty percent of Earth’s terrestrial habitats are now undergoing protection, biodiversity hotspots are being targeted, and there is increasing community awareness and engagement in conservation. However, the impending sixth mass extinction threatens to drive many species to extinction in the wild, irrespective of these interventions. Earth’s biosphere is now undergoing terraforming through ecosystem destruction and modification, urbanization, and agriculture. Therefore, transformative cultural, political, and economic incentives are needed to maximize the legacy of the Earth’s biodiversity and biospheric sustainability toward future generations. Reproduction and advanced biotechnologies can perpetuate species and their genetic diversity while also contributing to human and animal health and agricultural production. Advanced reproduction biotechnologies, including genetic engineering and synthetic biology, provide a new horizon for biospheric management, through the de-extinction of ancient species, restoring recently lost species, and maintaining the genetic diversity of extant species through wildlife biobanking. More extensive and inclusive conservation breeding programs and wildlife biobanking resources/facilities are desperately needed to perpetuate more than 3,000 Critically Endangered terrestrial/freshwater species; a goal fully attainable for amphibians and smaller fishes through global inclusion of stakeholders including private caregivers, plausible for freshwater mussels and crayfish, in development for reptiles and birds, and applicable for many mammals. As this capacity develops, many otherwise neglected species that are losing their natural habitat can be perpetuated solely in biobanks, thus enabling the more efficient utilization of resources toward meaningful field conservation primarily through habitat protection. The full potential of reproduction and advanced biotechnologies includes the development of artificial wombs to address the human population crisis and to avoid surrogacy mismatching during species restoration or de-extinction. The use of advanced reproduction biotechnologies for direct human benefit, for species management, and for biospheric sustainability, are subject to evolving ethical and legal frameworks, particularly regarding genetic engineering, transhumanism, and the de-extinction of ancient species.

Primary themes in intergenerational justice are a healthy environment, the perpetuation of Earth’s biodiversity, and the sustainable management of the biosphere. However, the current rate of species declines globally, ecosystem collapses driven by accelerating and catastrophic global heating, and a plethora of other threats preclude the ability of habitat protection alone to prevent a cascade of amphibian and other species mass extinctions. Reproduction and advanced biotechnologies, biobanking of germplasm and somatic cells, and conservation breeding programs (RBCs) offer a transformative change in biodiversity management. This change can economically and reliably perpetuate species irrespective of environmental targets and extend to satisfy humanity’s future needs as the biosphere expands into space. Currently applied RBCs include the hormonal stimulation of reproduction, the collection and refrigerated storage of sperm and oocytes, sperm cryopreservation, in vitro fertilization, and biobanking of germplasm and somatic cells. The benefits of advanced biotechnologies in development, such as assisted evolution and cloning for species adaptation or restoration, have yet to be fully realized. We broaden our discussion to include genetic management, political and cultural engagement, and future applications, including the extension of the biosphere through humanity’s interplanetary and interstellar colonization. The development and application of RBCs raise intriguing ethical, theological, and philosophical issues. We address these themes with amphibian models to introduce the Multidisciplinary Digital Publishing Institute Special Issue, The Sixth Mass Extinction and Species Sustainability through Reproduction Biotechnologies, Biobanking, and Conservation Breeding Programs.

Primary themes in intergenerational justice are a healthy environment, the perpetuation of Earth’s biodiversity, and the sustainable management of the biosphere. However, the current rate of species declines globally, ecosystem collapses driven by accelerating and catastrophic global heating and a plethora of other threats, preclude the ability of habitat protection alone to prevent a cascade of amphibian and other species mass extinctions. Reproduction biotechnologies, biobanking, and conservation breeding programs (RBCs), enabling species restoration, offer a critical transformative change that can economically and reliably perpetuate species irrespective of environmental targets. These capacities extend indefinitely into the future and satisfy humanities needs as the biosphere expands into space. Currently applied RBCs include the hormonal stimulation of reproduction, the collection and refrigerated storage of sperm and oocytes, sperm cryopreservation, in vitro fertilisation, along biobanking. Advanced biotechnologies such as assisted evolution and cloning for species adaptation or restoration have yet to be fully realised. We broaden our discussion to include genetic management, broad political and cultural engagement, and future applications. The development and application of RBCs, and particularly assisted evolution and species restoration, raise intriguing ethical, theological, and philosophical issues. We address these themes with amphibian models to introduce the MDPI Special Issue, The Sixth Mass Extinction and Species Sustainability through Reproduction Biotechnologies, Biobanking, and Conservation Breeding Programs.

Intergenerational justice entitles the maximum retention of Earth’s biodiversity. The 2022 United Nations COP 15, “Ecological Civilisation: Building a Shared Future for All Life on Earth”, is committed to protecting 30% of Earth’s terrestrial environments and, through COP 28, to mitigate the effects of the climate catastrophe on the biosphere. We focused this review on three core themes: the need and potential of reproduction biotechnologies, biobanks, and conservation breeding programs (RBCs) to satisfy sustainability goals; the technical state and current application of RBCs; and how to achieve the future potentials of RBCs in a rapidly evolving environmental and cultural landscape. RBCs include the hormonal stimulation of reproduction, the collection and storage of sperm and oocytes, and artificial fertilisation. Emerging technologies promise the perpetuation of species solely from biobanked biomaterials stored for perpetuity. Despite significant global declines and extinctions of amphibians, and predictions of a disastrous future for most biodiversity, practical support for amphibian RBCs remains limited mainly to a few limited projects in wealthy Western countries. We discuss the potential of amphibian RBCs to perpetuate amphibian diversity and prevent extinctions within multipolar geopolitical, cultural, and economic frameworks. We argue that a democratic, globally inclusive organisation is needed to focus RBCs on regions with the highest amphibian diversity. Prioritisation should include regional and international collaborations, community engagement, and support for RBC facilities ranging from zoos and other institutions to those of private carers. We tabulate a standard terminology for field programs associated with RBCs for publication and media consistency.

The term ‘mitochondrial vesicle’ was first used in 2003 in a description of anuran sperm and persists to this day throughout the literature on assisted reproductive technologies (ART) for amphibians. In the present paper, we argue that the term is inappropriate because the widely accepted definition of a ‘vesicle’ refers to an integral structure with an enclosing lipid bilayer/membrane. Moreover, there are no electron micrographs that show a vesicular structure encapsulating mitochondria on amphibian sperm heads in the literature. In fact, in 1993, the mitochondria in the anuran sperm head had been described as positioned in ‘mitochondrial collars’ or ‘mitochondrial sheaths’ surrounded by the plasma membrane of the sperm head. On the other hand, mitochondrial-derived vesicles are defined as vesicles shed from mitochondria surfaces, potentially creating confusion. Therefore, our view is that the term ‘mitochondrial vesicle’ should be avoided in describing the positioning of mitochondria on sperm.

Cloning as it relates to the animal kingdom generally refers to the production of genetically identical individuals. Because cloning is increasingly the subject of renewed attention as a tool for rescuing endangered or extinct species, it seems timely to dissect the role of the numerous reproductive techniques encompassed by this term in animal species conservation. Although cloning is typically associated with somatic cell nuclear transfer, the recent advent of additional techniques that allow genome replication without genetic recombination demands that the use of induced pluripotent stem cells to generate gametes or embryos, as well as older methods such as embryo splitting, all be included in this discussion. Additionally, the phenomenon of natural cloning (e.g., a subset of fish, birds, invertebrates, and reptilian species that reproduce via parthenogenesis) must also be pointed out. Beyond the biology of these techniques are practical considerations and the ethics of using cloning and associated procedures in endangered or extinct species. All of these must be examined in concert to determine whether cloning has a place in species conservation. Therefore, we synthesize progress in cloning and associated techniques and dissect the practical and ethical aspects of these methods as they pertain to endangered species conservation.

Of the amphibian orders, the Gymnophiona (caecilians) have the lowest number of species and are the least known. We report new information on the morphology, biology, range, and distribution of Ichthyophis kodaguensis, a striped ichthyophiid caecilian from the Western Ghats, India that shows the first evidence of possible sexual-dimorphism in this species. Based on the clutch size, limited range, relatively low fecundity, and agricultural practices in their habitats, we consider that I. kodaguensis is highly threatened when compared to other striped ichthyophiids from the Western Ghats, a biodiversity hotspot.

Intergenerational Justice entitles the maximum retention of Earth’s Biodiversity. The United Nations COP 15 committed to the protection of 30% of Earth’s terrestrial environments to provide Biospheric Sustainability. However, proactive amphibian sustainability needs support from amphibian Reproduction Technologies, Biobanking, and Conservation Breeding Programs [RBCs]. These technologies include the reliable practice of stimulation of reproduction and the collection of sperm and oocytes, refrigerated and cryopreserved sperm storage, in vitro fertilization. Emerging technologies promise the perpetuation of species solely from biobanked biomaterials. However, although exemplary national programs for amphibian RBCs are now established in the USA and Australia, these potentials are not fully realised resulting in unacceptable biodiversity loss. To fully realise these potentials an independent and democratic global organisation is proposed to, 1) focus on the bioregions with the highest amphibian biodiversity, 2) maximise regional inclusion in international initiatives, and 3) provide for maximum community engagement and financial resources. We present the first overall multidisciplinary review of amphibian RBCs, including their geopolitical, cultural, and scientific background, along with their development and application to economically, efficiently, and reliably, perpetuate amphibian biodiversity. We provide genetic models, the potential for diploid cell cryopreservation to perpetuate species, along with significant new information on methodologies and techniques, and standardise nomenclature for specificity in publications and media consistency.

We review the use of reproduction technologies (RTs) to support the sustainable management of threatened Caudata (salamanders) and Gymnophiona (caecilian) biodiversity in conservation breeding programs (CBPs) or through biobanking alone. The Caudata include ∼760 species with ∼55% threatened, the Gymnophiona include ∼215 species with an undetermined but substantial number threatened, with 80% of Caudata and 65% of Gymnophiona habitat unprotected. Reproduction technologies include: (1) the exogenous hormonal induction of spermatozoa, eggs, or mating, (2) in vitro fertilisation, (3) intracytoplasmic sperm injection (ICSI), (4) the refrigerated storage of spermatozoa, (5) the cryopreservation of sperm, cell or tissues, (6) cloning, and (7) gonadal tissue or cell transplantation into living amphibians to eventually produce gametes and then individuals. Exogenous hormone regimens have been applied to 11 Caudata species to stimulate mating and to 14 species to enable the collection of spermatozoa or eggs. In vitro fertilisation has been successful in eight species, spermatozoa have been cryopreserved in seven species, and in two species in vitro fertilisation with cryopreserved spermatozoa has resulted in mature reproductive adults. However, the application of RTs to Caudata needs research and development over a broader range of species. Reproduction technologies are only now being developed for Gymnophiona, with many discoveries and pioneering achievement to be made. Species with the potential for repopulation are the focus of the few currently available amphibian CBPs. As Caudata and Gymnophiona eggs or larvae cannot be cryopreserved, and the capacity of CBPs is limited, the perpetuation of the biodiversity of an increasing number of species depends on the development of RTs to recover female individuals from cryopreserved and biobanked cells or tissues.

Semen of high to moderate quality was collected following the hormonal induction of North American giant salamanders Cryptobranchus alleganiensis. Oocytes from one female yielded the first C. alleganiensis produced while maintained in aquaria under human care and the first externally fertilising salamander produced with cryopreserved spermatozoa and IVF. Further research is needed with North American giant salamanders to establish reliable techniques to produce large numbers of viable offspring, along with the application of cryopreserved spermatozoa.