Habitat loss and fragmentation are major drivers of biodiversity decline, reducing connectivity among populations and leading to genetic isolation, loss of diversity, increased inbreeding, and reduced fitness. Translocations that promote gene flow by introducing genetically distinct individuals-a process known as genetic rescue-can mitigate these effects by increasing genetic diversity, alleviating inbreeding, and improving adaptive capacity. However, a limited understanding of a population's demographic history, genetic differentiation, and connectivity can hinder the effective application of genetic rescue. We used the Stephens' kangaroo rat (Dipodomys stephensi), a species threatened by habitat loss and fragmentation in southern California, as a model for developing range-wide genetic management strategies. We analyzed mitochondrial DNA and microsatellite data to investigate genetic structure and estimate both historical and recent demographic patterns, and we used landscape resistance modeling to assess the impacts of natural and anthropogenic barriers on gene flow. Genetic analyses suggest a relatively recent diversification of Stephens' kangaroo rat populations, with higher allelic diversity concentrated in central populations and reduced diversity in isolated northern and southern populations. Although natural geographic features explain much of the genetic structure, landscape resistance models showed that anthropogenic barriers (e.g., roads, development) play a key role in current genetic isolation and are expected to continue driving population differentiation. To guide management, we used population viability simulations to test translocation strategies aimed at reversing genetic erosion. Repeated translocations were far more effective than single events at boosting heterozygosity and population persistence. The frequency and size of translocations were less important than their continued implementation. For very small populations, concurrent habitat restoration to increase carrying capacity was essential to prevent extirpation. Our findings highlight the value of integrating genetic, demographic, and landscape data into conservation planning. This approach is broadly applicable to other species experiencing habitat fragmentation and population isolation.

The distribution, frequency, and severity of genetic load (the accumulation of deleterious alleles, levels of homozygosity, and heterozygosity) are widely used as metrics for fitness and viability in small populations. Higher allelic diversity may enable populations to persist and evolve in unpredictable environments, suggesting that individuals with higher homozygosity, lower heterozygosity, and higher inbreeding coefficients may exhibit reduced fitness. Under a climate change scenario, drastic temperature fluctuations might be particularly challenging to thrive and survive due to their deleterious physiological effects. Our goal is to test whether performance of primary fibroblasts under heat stress correlates with genetic load parameters in the threatened Southern White Rhino ( Ceratotherium simum simum ). Published data on homozygosity, heterozygosity, and inbreeding scores will be correlated with cellular and mitochondrial function, assessed through multiple physiological markers. To obtain this data, primary fibroblasts (n = 15) were exposed to either 41°C (heat stress) or 37°C (control) for 24 hours. Afterwards, mitochondrial function was measured via a mitochondrial stress test (Agilent Seahorse Analyzer), cellular morphology (nuclear size and cytoskeletal organization, mitochondrial distribution) was evaluated through immunofluorescence (IF) staining, along with senescence markers (β-galactosidase presence), oxidative stress (reactive oxygen species presence), and RNA transcription (SYTO RNA Select stain). Heat shock protein (HSP70) abundance will be quantified using western blot analysis. The IF data were analyzed with CellProfiler™ software or with IF spectrophotometer. Preliminary results show that acute exposure to heat result in a series deleterious effects on SWR fibroblasts performance, including significant reduction of mitochondrial function (basal respiration: t = 3.7, df = 9, p < 0.01; maximum respiration: t = 3.0, df = 9, p < 0.02; ATP production: t = 5.8, df = 9, p < 0.001; coupling efficiency: t = 7.6, df = 9, p < 0.001) and ROS production (t = 3.35, df = 7, p = 0.01). Heat stress also increases nuclear size (area: t = 6.8, df = 9, p < 0.001) without altering RNA transcription (p = 0.63), and increase the presence of the senescence marker (W = 104.0, n = 15, p < 0.001). Future analyses will test the if genetic load parameters can explain individual cellular thermosensitivity. This study aims to provide insights into the connection between genetic markers and organismal performance under global warming scenario, as cellular and mitochondrial function plays a fundamental role in an organism's ability to adjust and thrive under environmental change. San Diego Zoo Wildlife Alliance This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

The most dynamic and repetitive regions of great ape genomes have traditionally been excluded from comparative studies 1–3 . Consequently, our understanding of the evolution of our species is incomplete. Here we present haplotype-resolved reference genomes and comparative analyses of six ape species: chimpanzee, bonobo, gorilla, Bornean orangutan, Sumatran orangutan and siamang. We achieve chromosome-level contiguity with substantial sequence accuracy (<1 error in 2.7 megabases) and completely sequence 215 gapless chromosomes telomere-to-telomere. We resolve challenging regions, such as the major histocompatibility complex and immunoglobulin loci, to provide in-depth evolutionary insights. Comparative analyses enabled investigations of the evolution and diversity of regions previously uncharacterized or incompletely studied without bias from mapping to the human reference genome. Such regions include newly minted gene families in lineage-specific segmental duplications, centromeric DNA, acrocentric chromosomes and subterminal heterochromatin. This resource serves as a comprehensive baseline for future evolutionary studies of humans and our closest living ape relatives.

With dramatic advancements in biological data generation, genetic rescue and reproductive technologies, and inter-institutional coordination of care across entire animal populations, zoos, aquariums, and their collaborators are uniquely positioned to lead population-wide research benefiting animal wellbeing and species survival. However, procedural and inter-institutional barriers make it exceedingly difficult to access existing zoological biospecimens and data at scale. To address this, the Zoonomics Working Group, representing diverse roles across three zoological associations (AZA, EAZA, WAZA), proposes a biodiversity biobank alliance that develops and delivers shared resources to support the collection, storage, and sharing of biological samples and associated data across the zoological and conservation community. By biobank alliance, we mean a community-guided effort that develops shared resources, standards, ethos, and practices for collecting, storing, and sharing biological samples and associated data voluntarily through transparent processes, consistent with professional accreditation standards and international best practices. While initially focused on addressing the needs and regulatory landscape of U.S. institutions, the alliance is designed to create frameworks that are adaptable and adoptable for international expansion. Such a framework would help the zoological community navigate the ethical, legal, and practical challenges of managing biospecimen collections, making access more efficient, reliable, and robust. Achieving this vision requires collective agreement on ethical principles such as reciprocity, transparency, and data stewardship, ensuring that research is both feasible and proactively supported. Such coordination will drive advances in fundamental biology and accelerate progress in animal health, welfare, management, and biodiversity conservation.

We present haplotype-resolved reference genomes and comparative analyses of six ape species, namely: chimpanzee, bonobo, gorilla, Bornean orangutan, Sumatran orangutan, and siamang. We achieve chromosome-level contiguity with unparalleled sequence accuracy (<1 error in 500,000 base pairs), completely sequencing 215 gapless chromosomes telomere-to-telomere. We resolve challenging regions, such as the major histocompatibility complex and immunoglobulin loci, providing more in-depth evolutionary insights. Comparative analyses, including human, allow us to investigate the evolution and diversity of regions previously uncharacterized or incompletely studied without bias from mapping to the human reference. This includes newly minted gene families within lineage-specific segmental duplications, centromeric DNA, acrocentric chromosomes, and subterminal heterochromatin. This resource should serve as a definitive baseline for all future evolutionary studies of humans and our closest living ape relatives.

Abstract As biodiversity loss outpaces recovery, conservationists are increasingly turning to novel tools for preventing extinction, including cloning and in vitro gametogenesis of biobanked cells. However, restoration of populations can be hindered by low genetic diversity and deleterious genetic load. The persistence of the northern white rhino (Ceratotherium simum cottoni) now depends on the cryopreserved cells of 12 individuals. These banked genomes have higher genetic diversity than southern white rhinos (C. s. simum), a sister subspecies that successfully recovered from a severe bottleneck, but the potential impact of genetic load is unknown. We estimated how demographic history has shaped genome‐wide genetic load in nine northern and 13 southern white rhinos. The bottleneck left southern white rhinos with more fixed and homozygous deleterious alleles and longer runs of homozygosity, whereas northern white rhinos retained more deleterious alleles masked in heterozygosity. To gauge the impact of genetic load on the fitness of a northern white rhino population restored from biobanked cells, we simulated recovery using fitness of southern white rhinos as a benchmark for a viable population. Unlike traditional restoration, cell‐derived founders can be reintroduced in subsequent generations to boost lost genetic diversity and relieve inbreeding. In simulations with repeated reintroduction of founders into a restored population, the fitness cost of genetic load remained lower than that borne by southern white rhinos. Without reintroductions, rapid growth of the restored population (>20–30% per generation) would be needed to maintain comparable fitness. Our results suggest that inbreeding depression from genetic load is not necessarily a barrier to recovery of the northern white rhino and demonstrate how restoration from biobanked cells relieves some constraints of conventional restoration from a limited founder pool. Established conservation methods that protect healthy populations will remain paramount, but emerging technologies hold promise to bolster these tools to combat the extinction crisis.

Conservation breeding programs have long been an important management tool for species recovery. Recently, breeding programs have begun to integrate next generation sequencing data into pedigree-based management strategies designed to maximize retention of genetic diversity and minimize inbreeding. In 2005, an ex situ breeding program for two of the three extant, geographically separate populations of the endangered southern mountain yellow-legged frog ( Rana muscosa ) was established at the San Diego Zoo Wildlife Alliance. To provide improved breeding recommendations for the species, we conducted molecular population genetic assessments using double digest restriction site-associated DNA sequencing data . We first studied genetic diversity, population differentiation, and genetic structure of wild frogs to validate the status of management units in R. muscosa . Genetic diversity was relatively low (H O = 0.166–0.245), population differentiation moderate (F ST = 0.29), and patterns of genetic structure and admixture supported the evolutionary divergence of frogs by mountain ranges with limited gene flow. In the breeding program, relatedness estimates identified close relatives amongst the founders, and molecularly-derived kinship values were used to determine and rank suitable breeders to minimize population mean kinship. Ex situ management recommendations highlight the need for importing additional founders from the wild to enhance genetic diversity in the breeding program, as an effective source for genetic restoration. Alternatively, the use of advanced reproductive technologies to capture wild diversity without removal of individuals from the wild seem promising.

Climate change is associated with increasing global mean temperatures and extreme heat events. These drastic changes in temperature can have significant deleterious physiological effects, particularly on large strict homeothermic mammals such as the threatened Southern White Rhino (SWR - Ceratotherium simum simum). Developing alternative strategies to understand how climatic events impact species that are logistically challenging to study are crucial. Dermal fibroblasts have proven to be effective in vitro models for studying species resilience to climate since they can reflect an organism's thermosensitivity. These cells can provide an integrative and mechanistic explanation on how cellular function can inform organismal responses to environmental changes. We investigated the effects of heat on mitochondrial function and cellular biology by exposing SWR primary dermal fibroblasts (n = 10 biological replicates) to 24 hours of heat (41°C) or 37°C as a control in culture. Then, we evaluated mitochondrial function using mitochondrial stress test, glycolytic rate test and immunofluorescence (IF) to track nucleus size and mitochondrial distribution in the cytoplasm. Metabolic function was assessed using Agilent Seahorse analyzer and IF images were analyzed on CellProfilerTM software. Our results indicate that compared to the control (37°C), heat exposure decreases mitochondrial respiration (basal: t = 3.7, df = 9, p < 0.01; maximum: t = 3.0, df = 9, p < 0.02), ATP production (t = 5.8, df = 9, p < 0.001), and coupling effciency (t = 7.6, df = 9, p < 0.001). No changes were observed in spare respiratory capacity (t = 0.7, df = 9, p = 0.5), proton leak (t = 0.7, df = 9, p = 0.5) and glycolysis (basal: t = 0.3, df = 4, p = 0.4; compensatory: t = 0.6, df = 4, p = 0.6). Furthermore, heat exposure increases nucleus size and mitochondrial distribution area (Nucleus area: t = 6.8, d.f. = 9, P < 0.001, Mitochondrial coverage area: t = 5.3, d.f. = 9, p < 0.001) and decreases fluorescence intensity of the mitochondrial dye and solidity area covered by mitochondria (Fluorescence: t = 2.3, d.f. = 9, p = 0.04, Mitochondrial area solidity: t = 5.6, d.f. = 9, p < 0.001). These findings show that acute heatwaves trigger a series of complex and interconnected responses in SWR fibroblasts resulting in mitochondrial disfunction, possibly mitophagy and/or mitochondria disintegration, and larger dispersion through the cytoplasm. The thermosensitive phenotype exhibited by SWR fibroblasts provides insights into the likely consequences of global warming on the performance of this species, as cellular and mitochondrial functioning plays a fundamental role in an organism's ability to adjust and thrive under environmental change. Funded by NSF and San Diego Zoo Wildlife Alliance. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Evolutionary constraint and acceleration are powerful, cell-type agnostic measures of functional importance. Previous studies in mammals were limited by species number and reliance on human-referenced alignments. We explore the evolution of placental mammals, including humans, through reference-free whole-genome alignment of 240 species and protein-coding alignments for 428 species. We estimate 10.7% of the human genome is evolutionarily constrained. We resolve constraint to single nucleotides, pinpointing functional positions, and refine and expand by over seven-fold the catalog of ultraconserved elements. Overall, 48.5% of constrained bases are as yet unannotated, suggesting yet-to-be-discovered functional importance. Using species-level phenotypes and an updated phylogeny, we associate coding and regulatory variation with olfaction and hibernation. Focusing on biodiversity conservation, we identify genomic metrics that predict species at risk of extinction.

Species persistence can be influenced by the amount, type, and distribution of diversity across the genome, suggesting a potential relationship between historical demography and resilience. In this study, we surveyed genetic variation across single genomes of 240 mammals that compose the Zoonomia alignment to evaluate how historical effective population size ( N e ) affects heterozygosity and deleterious genetic load and how these factors may contribute to extinction risk. We find that species with smaller historical N e carry a proportionally larger burden of deleterious alleles owing to long-term accumulation and fixation of genetic load and have a higher risk of extinction. This suggests that historical demography can inform contemporary resilience. Models that included genomic data were predictive of species’ conservation status, suggesting that, in the absence of adequate census or ecological data, genomic information may provide an initial risk assessment.

Zoonomia is the largest comparative genomics resource for mammals produced to date. By aligning genomes for 240 species, we identify bases that, when mutated, are likely to affect fitness and alter disease risk. At least 332 million bases (~10.7%) in the human genome are unusually conserved across species (evolutionarily constrained) relative to neutrally evolving repeats, and 4552 ultraconserved elements are nearly perfectly conserved. Of 101 million significantly constrained single bases, 80% are outside protein-coding exons and half have no functional annotations in the Encyclopedia of DNA Elements (ENCODE) resource. Changes in genes and regulatory elements are associated with exceptional mammalian traits, such as hibernation, that could inform therapeutic development. Earth’s vast and imperiled biodiversity offers distinctive power for identifying genetic variants that affect genome function and organismal phenotypes.

Bakker et al. use Robinson et al.'s reconstruction of three species of vulture to illustrate how incorrect generation time estimates can yield inaccurate results, underscoring the importance of generation time specification for genetically based reconstructions, especially for comparisons and species of conservation concern.

High-quality reference genomes are fundamental tools for understanding population history, and can provide estimates of genetic and demographic parameters relevant to the conservation of biodiversity. The federally endangered Pacific pocket mouse (PPM), which persists in three small, isolated populations in southern California, is a promising model for studying how demographic history shapes genetic diversity, and how diversity in turn may influence extinction risk. To facilitate these studies in PPM, we combined PacBio HiFi long reads with Omni-C and Hi-C data to generate a de novo genome assembly, and annotated the genome using RNAseq. The assembly comprised 28 chromosome-length scaffolds (N50 = 72.6 MB) and the complete mitochondrial genome, and included a long heterochromatic region on chromosome 18 not represented in the previously available short-read assembly. Heterozygosity was highly variable across the genome of the reference individual, with 18% of windows falling in runs of homozygosity (ROH) >1 MB, and nearly 9% in tracts spanning >5 MB. Yet outside of ROH, heterozygosity was relatively high (0.0027), and historical N-e estimates were large. These patterns of genetic variation suggest recent inbreeding in a formerly large population. Currently the most contiguous assembly for a heteromyid rodent, this reference genome provides insight into the past and recent demographic history of the population, and will be a critical tool for management and future studies of outbreeding depression, inbreeding depression, and genetic load.

Decrypting the rearrangements that drive mammalian chromosome evolution is critical to understanding the molecular bases of speciation, adaptation, and disease susceptibility. Using 8 scaffolded and 26 chromosome-scale genome assemblies representing 23/26 mammal orders, we computationally reconstructed ancestral karyotypes and syntenic relationships at 16 nodes along the mammalian phylogeny. Three different reference genomes (human, sloth, and cattle) representing phylogenetically distinct mammalian superorders were used to assess reference bias in the reconstructed ancestral karyotypes and to expand the number of clades with reconstructed genomes. The mammalian ancestor likely had 19 pairs of autosomes, with nine of the smallest chromosomes shared with the common ancestor of all amniotes (three still conserved in extant mammals), demonstrating a striking conservation of synteny for ∼320 My of vertebrate evolution. The numbers and types of chromosome rearrangements were classified for transitions between the ancestral mammalian karyotype, descendent ancestors, and extant species. For example, 94 inversions, 16 fissions, and 14 fusions that occurred over 53 My differentiated the therian from the descendent eutherian ancestor. The highest breakpoint rate was observed between the mammalian and therian ancestors (3.9 breakpoints/My). Reconstructed mammalian ancestor chromosomes were found to have distinct evolutionary histories reflected in their rates and types of rearrangements. The distributions of genes, repetitive elements, topologically associating domains, and actively transcribed regions in multispecies homologous synteny blocks and evolutionary breakpoint regions indicate that purifying selection acted over millions of years of vertebrate evolution to maintain syntenic relationships of developmentally important genes and regulatory landscapes of gene-dense chromosomes.

Robinson and colleagues respond to the points raised about their paper by Bakker et al.

Small populations are often exposed to high inbreeding and mutational load that can increase the risk of extinction. The Sumatran rhinoceros was widespread in Southeast Asia, but is now restricted to small and isolated populations on Sumatra and Borneo, and most likely extinct on the Malay Peninsula. Here, we analyse 5 historical and 16 modern genomes from these populations to investigate the genomic consequences of the recent decline, such as increased inbreeding and mutational load. We find that the Malay Peninsula population experienced increased inbreeding shortly before extirpation, which possibly was accompanied by purging. The populations on Sumatra and Borneo instead show low inbreeding, but high mutational load. The currently small population sizes may thus in the near future lead to inbreeding depression. Moreover, we find little evidence for differences in local adaptation among populations, suggesting that future inbreeding depression could potentially be mitigated by assisted gene flow among populations.

Parthenogenesis is a relatively rare event in birds, documented in unfertilized eggs from columbid, galliform, and passerine females with no access to males. In the critically endangered California condor, parentage analysis conducted utilizing polymorphic microsatellite loci has identified two instances of parthenogenetic development from the eggs of two females in the captive breeding program, each continuously housed with a reproductively capable male with whom they had produced offspring. Paternal genetic contribution to the two chicks was excluded. Both parthenotes possessed the expected male ZZ sex chromosomes and were homozygous for all evaluated markers inherited from their dams. These findings represent the first molecular marker-based identification of facultative parthenogenesis in an avian species, notably of females in regular contact with fertile males, and add to the phylogenetic breadth of vertebrate taxa documented to have reproduced via asexual reproduction.

Bitter taste receptors serve as a vital component in the defense system against toxin intake by animals, and the family of genes encoding these receptors has been demonstrated, usually by family size variance, to correlate with dietary preference. However, few systematic studies of specific Tas2R to unveil their functional evolution have been conducted. Here, we surveyed Tas2R16 across all major clades of primates and reported a rare case of a convergent change to increase sensitivity to β-glucopyranosides in human and a New World monkey, the white-faced saki. Combining analyses at multiple levels, we demonstrate that a parallel amino acid substitution (K172N) shared by these two species is responsible for this functional convergence of Tas2R16. Considering the specialized feeding preference of the white-faced saki, the K172N change likely played an important adaptive role in its early evolution to avoid potentially toxic cyanogenic glycosides, as suggested for the human TAS2R16 gene.

Maintaining the existing biodiversity of endangered species is a goal of conservation management programs, and a major component of many collaborative efforts undertaken by zoos, field biologists, and conservation scientists. Over the past 3 decades, the San Diego Zoo has performed long-term genetic studies in support of the recovery program for the critically endangered California Condor (Gymnogyps californianus). This work has included sex determination of hatchlings and parentage confirmation using microsatellite genotyping. This paper describes the genetic work associated with the California Condor recovery program, which aims to provide a highly accurate pedigree for making informed captive pairing and release recommendations. Initial genotyping began after reintroduced California Condors started reproducing, and the focus was on birds hatched from their wild-laid eggs. However, genetic analysis showed discrepancies relative to behavioral observations of wild birds and led to the species-wide testing of all available samples. This genetic study has resolved instances of individual misidentifications and parental misassignments, verified the first cases of extra-pair paternity in this species, identified parentage where chicks were observed being raised by trios, and found 2 apparent de novo mutations in the captive condor population. Correcting the California Condor pedigree according to genetic parental analysis has produced more accurate estimates of mean kinship values among living birds, ranking potential breeders according to their actual breeding value and helping managers to make informed decisions about captive pairing and release of condors in the wild.

Only five species of the once-diverse Rhinocerotidae remain, making the reconstruction of their evolutionary history a challenge to biologists since Darwin. We sequenced genomes from five rhinoceros species (three extinct and two living), which we compared to existing data from the remaining three living species and a range of outgroups. We identify an early divergence between extant African and Eurasian lineages, resolving a key debate regarding the phylogeny of extant rhinoceroses. This early Miocene (∼16 million years ago [mya]) split post-dates the land bridge formation between the Afro-Arabian and Eurasian landmasses. Our analyses also show that while rhinoceros genomes in general exhibit low levels of genome-wide diversity, heterozygosity is lowest and inbreeding is highest in the modern species. These results suggest that while low genetic diversity is a long-term feature of the family, it has been particularly exacerbated recently, likely reflecting recent anthropogenic-driven population declines.