The Sixth Mass Extinction and Amphibian Species Sustainability Through Reproduction and Advanced Biotechnologies, Biobanking of Germplasm and Somatic Cells, and Conservation Breeding Programs (RBCs).

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.

The Effect of Gentamicin on the Motility of Hormonally Induced Spermatozoa of Toad Bufo bufo during Storage at 4°C.

We studied the effect of antibiotic gentamicin at concentrations of 0.05, 0.1, 0.

The case for considering the term 'mitochondrial vesicle' as a misnomer in publications about assisted reproductive technologies (ART) for amphibians.

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.

Reproduction technologies for the sustainable management of Caudata (salamander) and Gymnophiona (caecilian) biodiversity.

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.

Influence of Xenon on Survival of Sperm of Common Frog Rana temporaria during Slow Freezing.

We studied the effect of xenon on the survival rate of the spermatozoa of the common frog Rana temporaria during slow freezing with saturation of the suspension with xenon at a pressure of up to 1.2 bar. The cryoprotective properties of xenon were analyzed in comparison with nitrogen.

Influence of helium, xenon, and other noble gases on cryopreservation of Hela and l929 cell lines.

Any biological material contains dissolved gases that affect physical and biological processes associated with cooling and freezing. However, in the cryobiology literature, little attention has been paid to the effect of gasses on cryopreservation. We studied the influence of helium, neon, krypton, xenon, argon, nitrogen, and sulfur hexafluoride on the survivability of HeLa and L929 cell lines during cryopreservation.

Cryopreservation of spermatozoa obtained postmortem from the European common frog .

Cryopreserved spermatozoa offers a reliable, efficient and cost-effective means to perpetuate the genetic variation of endangered amphibian species in concert with conservation breeding programs. Here we describe successful cryopreservation of testicular spermatozoa of the common frog Rana temporaria , preliminarily stored in the carcasses of decapitated animals at +4°C for 0, 1 and 4 days. The motility, membrane integrity and fertilisation capability of fresh testicular spermatozoa treated with cryoprotective medium supplemented with 15% dimethylformamide (DMF) or 15% dimethylsulfoxide (DMSO) were examined.

Sperm collection and storage for the sustainable management of amphibian biodiversity.

Current rates of biodiversity loss pose an unprecedented challenge to the conservation community, particularly with amphibians and freshwater fish as the most threatened vertebrates. An increasing number of environmental challenges, including habitat loss, pathogens, and global warming, demand a global response toward the sustainable management of ecosystems and their biodiversity. Conservation Breeding Programs (CBPs) are needed for the sustainable management of amphibian species threatened with extinction.

Refrigerated storage of European common frog Rana temporaria oocytes.

Reproduction technologies (RTs) for the storage and use of amphibian gametes have rapidly developed since the recognition of the amphibian conservation crisis in the late 20th Century. Of these RTs, the refrigerated storage of oocytes and sperm can help to achieve reliable pair-matching when unexpected deaths could lead to critical gaps in studbook programs, and also to enable gamete transport between facilities or when sampled from field populations. Viable sperm can be reliably stored in vitro in testes, as suspensions in refrigerators for weeks and in situ in refrigerated carcasses for days.

Sperm motility of externally fertilizing fish and amphibians.

We review the phylogeny, sperm competition, morphology, physiology, and fertilization environments of the sperm of externally fertilizing fish and amphibians. Increased sperm competition in both fish and anurans generally increases sperm numbers, sperm length, and energy reserves. The difference between the internal osmolarity and iconicity of sperm cells and those of the aquatic medium control the activation, longevity, and velocity of sperm motility.

The quality and fertility of sperm collected from European common frog (Rana temporaria) carcasses refrigerated for up to 7 days.

There is a catastrophic decrease in the biodiversity of amphibians coupled with the loss of genetic variation. The perpetuation of amphibian biodiversity demands a multifaceted approach, including the use of reproduction technologies (RTs), to enable efficient reproduction in captivity and to prevent the loss of genetic variation. Reproduction technologies for the storage of amphibian sperm for days to weeks, when refrigerated at 4°C, or for millennia when cryopreserved have recently undergone rapid development.

Hormonal induction of spermatozoa from amphibians with Rana temporaria and Bufo bufo as anuran models.

The use of hormonally induced spermatozoa expressed in urine (HISu) is a valuable component of reproduction technologies for amphibians. Five protocols for sampling HISu from the European common frog (Rana temporaria) were compared: (1) pituitary extracts, (2) 0.12 µg g⁻¹ luteinising hormone-releasing hormone analogue (LHRHa), (3) 1.

Cryopreservation of hormonally induced sperm for the conservation of threatened amphibians with Rana temporaria as a model research species.

The survival of hundreds of threatened amphibian species is increasingly dependent on conservation breeding programs (CBPs). However, there is an ongoing loss of genetic variation in CBPs for most amphibians, reptiles, birds, and mammals. Low genetic variation results in the failure of CBPs to provide genetically competent individuals for release in supplementation or rehabitation programs.

[Fluorescent analysis of cryopreserved totipotent cells of amphibian embryos].

The cryotolerance of totipotent cells from dissociated embryos of amphibian (grass frog Rana temporaria and grey toad Bufo bufo) was studied. Cell integrity and preservation of the cell barrier function were evaluated by fluorescent analysis. It was shown that the best cryopreservation of the cells was achieved by using the cryoprotective agent 10% dimethyl sulfoxide and 10% saccharose.