Give and take: Effects of genetic admixture on mutation load in endangered Florida panthers.

Genetic admixture is a biological event inherent to genetic rescue programs aimed at the long-term conservation of endangered wildlife. Although the success of such programs can be measured by the increase in genetic diversity and fitness of subsequent admixed individuals, predictions supporting admixture costs to fitness due to the introduction of novel deleterious alleles are necessary. Here, we analyzed nonsynonymous variation from conserved genes to quantify and compare levels of mutation load (i.e. proportion of deleterious alleles and genotypes carrying these alleles) among endangered Florida panthers and non-endangered Texas pumas. Specifically, we used canonical (i.e. non-admixed) Florida panthers, Texas pumas, and F1 (canonical Florida × Texas) panthers dating from a genetic rescue program and Everglades National Park panthers with Central American ancestry resulting from an earlier admixture event. We found neither genetic drift nor selection significantly reduced overall proportions of deleterious alleles in the severely bottlenecked canonical Florida panthers. Nevertheless, the deleterious alleles identified were distributed into a disproportionately high number of homozygous genotypes due to close inbreeding in this group. Conversely, admixed Florida panthers (either with Texas or Central American ancestry) presented reduced levels of homozygous genotypes carrying deleterious alleles but increased levels of heterozygous genotypes carrying these variants relative to canonical Florida panthers. Although admixture is likely to alleviate the load of standing deleterious variation present in homozygous genotypes, our results suggest that introduced novel deleterious alleles (temporarily present in heterozygous state) in genetically rescued populations could potentially be expressed in subsequent generations if their effective sizes remain small.

PSEUDORABIES (AUJESZKY'S DISEASE) IS AN UNDERDIAGNOSED CAUSE OF DEATH IN THE FLORIDA PANTHER (PUMA CONCOLOR CORYI).

Feral swine (Sus scrofa), an important prey species for the endangered Florida panther (Puma concolor coryi), is the natural host for pseudorabies virus (PRV). Prior to this study, PRV had been detected in just three panthers. To determine the effect of PRV on the panther population, we prospectively necropsied 199 panthers and retrospectively reviewed necropsy and laboratory findings, reexamined histology, and tested archived tissues using real-time PCR from 46 undiagnosed panther mortalities. Seven additional infections (two prospective, five retrospective) were detected for a total of 10 confirmed panther mortalities due to PRV. To further evaluate the effect of PRV, we categorized radio-collared (n=168) and uncollared panther mortalities (n=367) sampled from 1981 to 2018 based on the likelihood of PRV infection as confirmed, probable, suspected, possible, or unlikely/negative. Of 168 radio-collared panthers necropsied, PRV was the cause of death for between eight (confirmed; 4.8%) and 32 (combined confirmed, probable, suspected, and possible categories; 19.0%) panthers. The number of radio-collared panther mortalities due to PRV was estimated to be 15 (95% empirical limits: 12-19), representing 8.9% (confidence interval: 4.6-13.2%) of mortalities. Gross necropsy findings in 10 confirmed cases were nonspecific. Microscopic changes included slight to mild perivascular cuffing and gliosis (primarily in the brain stem), lymphoplasmacytic meningoencephalitis (cerebral cortex), and intranuclear inclusion bodies (adrenal medulla). The PRV glycoprotein C gene sequences from three positive panthers grouped with the sequence from a Florida feral swine. Our findings indicate that PRV may be an important and underdiagnosed cause of death in Florida panthers.

Author Correction: Puma genomes from North and South America provide insights into the genomic consequences of inbreeding.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Puma genomes from North and South America provide insights into the genomic consequences of inbreeding.

Pumas are the most widely distributed felid in the Western Hemisphere. Increasingly, however, human persecution and habitat loss are isolating puma populations. To explore the genomic consequences of this isolation, we assemble a draft puma genome and a geographically broad panel of resequenced individuals. We estimate that the lineage leading to present-day North American pumas diverged from South American lineages 300-100 thousand years ago. We find signatures of close inbreeding in geographically isolated North American populations, but also that tracts of homozygosity are rarely shared among these populations, suggesting that assisted gene flow would restore local genetic diversity. The genome of a Florida panther descended from translocated Central American individuals has long tracts of homozygosity despite recent outbreeding. This suggests that while translocations may introduce diversity, sustaining diversity in small and isolated populations will require either repeated translocations or restoration of landscape connectivity. Our approach provides a framework for genome-wide analyses that can be applied to the management of similarly small and isolated populations.

De Novo Assembly and Annotation from Parental and F1 Puma Genomes of the Florida Panther Genetic Restoration Program.

In the mid-1990s, the population size of Florida panthers became so small that many individuals manifested traits associated with inbreeding depression (e.g., heart defects, cryptorchidism, high pathogen-parasite load). To mitigate these effects, pumas from Texas were introduced into South Florida to augment genetic variation in Florida panthers. In this study, we report a de novo puma genome assembly and annotation after resequencing 10 individual genomes from partial Florida-Texas-F1 trios. The final genome assembly consisted of ∼2.6 Gb and 20,561 functionally annotated protein-coding genes. Foremost, expanded gene families were associated with neuronal and embryological development, whereas contracted gene families were associated with olfactory receptors. Despite the latter, we characterized 17 positively selected genes related to the refinement of multiple sensory perceptions, most notably to visual capabilities. Furthermore, genes under positive selection were enriched for the targeting of proteins to the endoplasmic reticulum, degradation of mRNAs, and transcription of viral genomes. Nearly half (48.5%) of ∼6.2 million SNPs analyzed in the total sample set contained putative unique Texas alleles. Most of these alleles were likely inherited to subsequent F1 Florida panthers, as these individuals manifested a threefold increase in observed heterozygosity with respect to their immediate, canonical Florida panther predecessors. Demographic simulations were consistent with a recent colonization event in North America by a small number of founders from South America during the last glacial period. In conclusion, we provide an extensive set of genomic resources for pumas and elucidate the genomic effects of genetic rescue on this iconic conservation success story.

Evolutionary and Functional Mitogenomics Associated With the Genetic Restoration of the Florida Panther.

Florida panthers are endangered pumas that currently persist in reduced patches of habitat in South Florida, USA. We performed mitogenome reference-based assemblies for most parental lines of the admixed Florida panthers that resulted from the introduction of female Texas pumas into South Florida in 1995. With the addition of 2 puma mitogenomes, we characterized 174 single nucleotide polymorphisms (SNPs) across 12 individuals. We defined 5 haplotypes (Pco1-Pco5), one of which (Pco1) had a geographic origin exclusive to Costa Rica and Panama and was possibly introduced into the Everglades National Park, Florida, prior to 1995. Haplotype Pco2 was native to Florida. Haplotypes Pco3 and Pco4 were exclusive to Texas, whereas haplotype Pco5 had an undetermined geographic origin. Phylogenetic inference suggests that haplotypes Pco1-Pco4 diverged ~202000 (95% HPDI = 83000-345000) years ago and that haplotypes Pco2-Pco4 diverged ~61000 (95% HPDI = 9000-127000) years ago. These results are congruent with a south-to-north continental expansion and with a recent North American colonization by pumas. Furthermore, pumas may have migrated from Texas to Florida no earlier than ~44000 (95% HPDI = 2000-98000) years ago. Synonymous mutations presented a greater mean substitution rate than other mitochondrial functional regions: nonsynonymous mutations, tRNAs, rRNAs, and control region. Similarly, all protein-coding genes were under predominant negative selection constraints. We directly and indirectly assessed the presence of potential deleterious SNPs in the ND2 and ND5 genes in Florida panthers prior to and as a consequence of the introduction of Texas pumas. Screenings for such variants are recommended in extant Florida panthers.

Hidden semi-Markov models reveal multiphasic movement of the endangered Florida panther.

Animals must move to find food and mates, and to avoid predators; movement thus influences survival and reproduction, and ultimately determines fitness. Precise description of movement and understanding of spatial and temporal patterns as well as relationships with intrinsic and extrinsic factors is important both for theoretical and applied reasons. We applied hidden semi-Markov models (HSMM) to hourly geographic positioning system (GPS) location data to understand movement patterns of the endangered Florida panther (Puma concolor coryi) and to discern factors influencing these patterns. Three distinct movement modes were identified: (1) Resting mode, characterized by short step lengths and turning angles around 180(o); (2) Moderately active (or intermediate) mode characterized by intermediate step lengths and variable turning angles, and (3) Traveling mode, characterized by long step lengths and turning angles around 0(o). Males and females, and females with and without kittens, exhibited distinctly different movement patterns. Using the Viterbi algorithm, we show that differences in movement patterns of male and female Florida panthers were a consequence of sex-specific differences in diurnal patterns of state occupancy and sex-specific differences in state-specific movement parameters, whereas the differences between females with and without dependent kittens were caused solely by variation in state occupancy. Our study demonstrates the use of HSMM methodology to precisely describe movement and to dissect differences in movement patterns according to sex, and reproductive status.

A cat's tale: the impact of genetic restoration on Florida panther population dynamics and persistence.

1. Genetic restoration has been suggested as a management tool for mitigating detrimental effects of inbreeding depression in small, inbred populations, but the demographic mechanisms underlying population-level responses to genetic restoration remain poorly understood. 2. We studied the dynamics and persistence of the endangered Florida panther Puma concolor coryi population and evaluated the potential influence of genetic restoration on population growth and persistence parameters. As part of the genetic restoration programme, eight female Texas pumas P. c. stanleyana were released into Florida panther habitat in southern Florida in 1995. 3. The overall asymptotic population growth rate (λ) was 1.04 (5th and 95th percentiles: 0.95-1.14), suggesting an increase in the panther population of approximately 4% per year. Considering the effects of environmental and demographic stochasticities and density-dependence, the probability that the population will fall below 10 panthers within 100 years was 0.072 (0-0.606). 4. Our results suggest that the population would have declined at 5% per year (λ = 0.95; 0.83-1.08) in the absence of genetic restoration. Retrospective life table response experiment analysis revealed that the positive effect of genetic restoration on survival of kittens was primarily responsible for the substantial growth of the panther population that would otherwise have been declining. 5. For comparative purposes, we also estimated probability of quasi-extinction under two scenarios - implementation of genetic restoration and no genetic restoration initiative - using the estimated abundance of panthers in 1995, the year genetic restoration was initiated. Assuming no density-dependence, the probability that the panther population would fall below 10 panthers by 2010 was 0.098 (0.002-0.332) for the restoration scenario and 0.445 (0.032-0.944) for the no restoration scenario, providing further evidence that the panther population would have faced a substantially higher risk of extinction if the genetic restoration initiative had not been implemented. 6. Our results, along with those reporting increases in population size and improvements in biomedical correlates of inbreeding depression, provide strong evidence that genetic restoration substantially contributed to the observed increases in the Florida panther population.

Does genetic introgression improve female reproductive performance? A test on the endangered Florida panther.

Genetic introgression has been suggested as a management tool for mitigating detrimental effects of inbreeding depression, but the role of introgression in species conservation has been controversial, partly because population-level impacts of genetic introgressions are not well understood. Concerns about potential inbreeding depression in the endangered Florida panther (Puma concolor coryi) led to the release of eight female Texas pumas (P. c. stanleyana) into the Florida panther population in 1995. We used long-term reproductive data (1995-2008) collected from 61 female Florida panthers to estimate and model reproduction probability (probability of producing a litter) and litter size, and to investigate the influence of intentional genetic introgression on these parameters. Overall, 6-month probability of reproduction (±1SE) was 0.232 ± 0.021 and average litter size was 2.60 ± 0.09. Although F(1) admixed females had a lower reproduction probability than females with other ancestries, this was most likely because kittens born to F(1) females survive better; consequently, these females are unavailable for breeding until kittens are independent. There was no evidence for the effect of ancestry on litter size or of heterozygosity on probability of reproduction or litter size. In contrast, earlier studies have shown that genetic introgression positively affected Florida panther survival. Our results, along with those of earlier studies, clearly suggest that genetic introgression can have differential effects on components of fitness and highlight the importance of examining multiple demographic parameters when evaluating the effects of management actions.

Intentional genetic introgression influences survival of adults and subadults in a small, inbred felid population.

1. Inbreeding and low genetic diversity can cause reductions in individual fitness and increase extinction risk in animal populations. Intentional introgression, achieved by releasing genetically diverse individuals into inbred populations, has been used as a conservation tool to improve demographic performance in endangered populations. 2. By the 1980s, Florida panthers (Puma concolor coryi) had been reduced to a small, inbred population that appeared to be on the brink of extinction. In 1995, female pumas from Texas (P. c. stanleyana) were released in occupied panther range as part of an intentional introgression programme to restore genetic variability and improve demographic performance of panthers. 3. We used 25 years (1981-2006) of continuous radiotelemetry and genetic data to estimate and model subadult and adult panther survival and cause-specific mortality to provide rigorous sex and age class-specific survival estimates and evaluate the effect of the introgression programme on these parameters. 4. Genetic ancestry influenced annual survival of subadults and adults after introgression, as F(1) generation admixed panthers ( = 0·98) survived better than pre-introgression type panthers ( = 0·77) and other admixed individuals ( = 0·82). Furthermore, heterozygosity was higher for admixed panthers relative to pre-introgression type panthers and positively influenced survival. 5. Our results are consistent with hybrid vigour; however, extrinsic factors such as low density of males in some areas of panther range may also have contributed to higher survival of F(1) panthers. Regardless, improved survival of F(1) subadults and adults likely contributed to the numerical increase in panthers following introgression, and our results indicate that intentional admixture, achieved here by releasing individuals from another population, appears to have been successful in improving demographic performance in this highly endangered population.

Genetic Introgression and the Survival of Florida Panther Kittens.

Estimates of survival for the young of a species are critical for population models. These models can often be improved by determining the effects of management actions and population abundance on this demographic parameter. We used multiple sources of data collected during 1982-2008 and a live recapture-dead recovery modeling framework to estimate and model survival of Florida panther (Puma concolor coryi) kittens (age 0 - 1 year). Overall, annual survival of Florida panther kittens was 0.323 ± 0.071 (SE), which was lower than estimates used in previous population models. In 1995, female pumas from Texas (P. c. stanleyana) were released into occupied panther range as part of an intentional introgression program to restore genetic variability. We found that kitten survival generally increased with degree of admixture: F(1) admixed and backcrossed to Texas kittens survived better than canonical Florida panther and backcrossed to canonical kittens. Average heterozygosity positively influenced kitten and older panther survival, whereas index of panther abundance negatively influenced kitten survival. Our results provide strong evidence for the positive population-level impact of genetic introgression on Florida panthers. Our approach to integrate data from multiple sources was effective at improving robustness as well as precision of estimates of Florida panther kitten survival, and can be useful in estimating vital rates for other elusive species with sparse data.

Genetic restoration of the Florida panther.

The rediscovery of remnant Florida panthers (Puma concolor coryi) in southern Florida swamplands prompted a program to protect and stabilize the population. In 1995, conservation managers translocated eight female pumas (P. c. stanleyana) from Texas to increase depleted genetic diversity, improve population numbers, and reverse indications of inbreeding depression. We have assessed the demographic, population-genetic, and biomedical consequences of this restoration experiment and show that panther numbers increased threefold, genetic heterozygosity doubled, survival and fitness measures improved, and inbreeding correlates declined significantly. Although these results are encouraging, continued habitat loss, persistent inbreeding, infectious agents, and possible habitat saturation pose new dilemmas. This intensive management program illustrates the challenges of maintaining populations of large predators worldwide.

Weight change affects serum leptin and corticosterone in the collared lemming.

The aim of this study was to determine whether photoperiod-induced changes in body and fat mass are accompanied by changes in leptin and corticosterone concentrations in collared lemmings. At weaning, eighty male lemmings were maintained in either long photoperiod (LD, n = 40 ) or short photoperiod (SD, n = 40 ). Ten weeks post-weaning lemmings were weighed and assigned to a secondary photoperiod that consisted of either remaining in the same photoperiod or being transferred to the alternate photoperiod (LD-LD, LD-SD, SD-SD, and SD-LD; n = 20 per group). Ten days post-switch, half the animals from each group were sacrificed. The remaining animals were sacrificed on day 21 post-switch. Blood was collected for determination of serum leptin and corticosterone, and carcasses were analyzed for body composition. LD-SD lemmings gained the most weight, whilst SD-LD lemmings lost weight. SD-LD lemmings had significantly lower leptin levels relative to fat mass than SD-SD lemmings. Corticosterone levels were higher in the SD-LD than SD-SD lemmings on both days 10 and 21. Levels were not significantly different between LD-LD and LD-SD lemmings; however by day 21 the levels were slightly lower in the LD-SD lemmings. Discussion. Lemmings showed seasonally appropriate body weight changes in response to the changing photoperiod. Weight loss was associated with higher corticosterone, and lower leptin levels adjusted for fat mass. Neither relative leptin levels, nor corticosterone levels changed significantly with weight gain. Our results indicate that corticosterone and leptin are associated more with seasonal weight loss than weight gain in lemmings.