Genetic diversity in wild and reared populations of the Japanese bitterling <Emphasis Type="Italic">Tanakia tanago</Emphasis> (Cyprinidae)

 The Japanese bitterling Tanakia tanago is an endangered cyprinid species; thus, captive breeding programs are being conducted in various facilities as ex situ conservation. To examine the genetic diversity in one wild and three reared populations, and its changes during the process of captive breeding, sequences of the mitochondrial cytochrome b gene and control region were determined. The wild population, collected in 1993, was monomorphic. Although the reared population that originated from the wild population was almost monomorphic, a rare haplotype, distinct from all others by a relatively large sequence divergence, was also observed. In the other reared populations, some degree of genetic diversity had been maintained. A reared hybrid population, which originated from a mixture of three distinct populations, showed the greatest genetic diversity. These results suggest considerable genetic diversity within and among populations of T. tanago in the past. Although a loss of genetic diversity was observed in some year-classes of reared populations, there was no tendency for genetic diversity to decrease as a result of captive breeding, probably because offspring were obtained from multi-year-class parents in the captive breeding program. Accordingly, this breeding method should be appropriate for conserving the genetic diversity of T. tanago. Received: June 12, 2002 / Revised: December 3, 2002 / Accepted: December 16, 2002     

Genetic population structure and management units of the endangered Tokyo bitterling, Tanakia tanago (Cyprinidae)

The Tokyo bitterling Tanakia tanago (Cyprinidae) was once found throughout the Kanto Plain, central Japan, but most of their habitats have been lost due to human activities such as urbanization and improvement of paddy fields. Subsequently, conservation efforts, including captive breeding and reintroduction, have been ongoing. However, the genetic relationships among populations of this species including captive and remnant wild populations have been uncertain and thus management units for this species have been unidentified. We examined the population differentiation among 12 populations, including four wild and eight captive populations, and their relative genetic diversities to assist in conservation management decisions. Phylogeographic analyses based on partial mitochondrial cytochrome b gene sequences and microsatellite polymorphisms revealed four geographically associated genetic groups in the populations. Northern Tochigi populations have diverged from other populations (0.77% of d _A ), likely stemming from allopatric fragmentation following a change in the route of the Naka River, which occurred during the middle of the Pleistocene epoch. Microsatellite analysis has revealed that the genetic diversity of each population is generally low, and that most of the populations have experienced genetic bottlenecks. For future in- and ex-situ conservation programs to succeed, the population structure and genetic variability of remnant populations need to be taken into consideration.     

Effective population size is strongly correlated with breeding pond size in the endangered California tiger salamander, <Emphasis Type="Italic">Ambystoma californiense</Emphasis>

Maintaining genetic diversity and population viability in endangered and threatened species is a primary concern of conservation biology. Genetic diversity depends on population connectivity and effective population size (N_e), both of which are often compromised in endangered taxa. While the importance of population connectivity and gene flow has been well studied, investigating effective population sizes in natural systems has received far less attention. However, N_e plays a prominent role in the maintenance of genetic diversity, the prevention of inbreeding depression, and in determining the probability of population persistence. In this study, we examined the relationship between breeding pond characteristics and N_e in the endangered California tiger salamander, Ambystoma californiense. We sampled 203 individuals from 10 breeding ponds on a local landscape, and used 11 polymorphic microsatellite loci to quantify genetic structure, gene flow, and effective population sizes. We also measured the areas of each pond using satellite imagery and classified ponds as either hydrologically-modified perennial ponds or naturally occurring vernal pools, the latter of which constitute the natural breeding habitat for A. californiense. We found no correlation between pond area and heterozygosity or allelic diversity, but we identified a strong positive relationship between breeding pond area and N_e, particularly for vernal pools. Our results provide some of the first empirical evidence that variation in breeding habitat can be associated with differences in N_e and suggest that a more complete understanding of the environmental features that influence N_e is an important component of conservation genetics and management.     

How many came home? Evaluating ex situ conservation of green turtles in the Cayman Islands

Ex situ management is an important conservation tool that allows the preservation of biological diversity outside natural habitats while supporting survival in the wild. Captive breeding followed by re‐introduction is a possible approach for endangered species conservation and preservation of genetic variability. The Cayman Turtle Centre Ltd was established in 1968 to market green turtle (Chelonia mydas) meat and other products and replenish wild populations, thought to be locally extirpated, through captive breeding. We evaluated the effects of this re‐introduction programmme using molecular markers (13 microsatellites, 800‐bp D‐loop and simple tandem repeat mitochondrial DNA sequences) from captive breeders (N = 257) and wild nesting females (N = 57) (sampling period: 2013–2015). We divided the captive breeders into three groups: founders (from the original stock), and then two subdivisions of F₁ individuals corresponding to two different management strategies, cohort 1995 (“C1995”) and multicohort F₁ (“MCF1”). Loss of genetic variability and increased relatedness was observed in the captive stock over time. We found no significant differences in diversity among captive and wild groups, and similar or higher levels of haplotype variability when compared to other natural populations. Using parentage and sibship assignment, we determined that 90% of the wild individuals were related to the captive stock. Our results suggest a strong impact of the re‐introduction programmme on the present recovery of the wild green turtle population nesting in the Cayman Islands. Moreover, genetic relatedness analyses of captive populations are necessary to improve future management actions to maintain genetic diversity in the long term and avoid inbreeding depression.     

Patterns of genetic diversity of mitochondrial DNA within captive populations of the endangered itasenpara bitterling: implications for a reintroduction program

In this study, the level of genetic diversity of captive populations of the itasenpara bitterling (Acheilognathus longipinnis) was assessed to obtain information useful for successful captive breeding and reintroduction; this analysis was performed using mitochondrial DNA (mtDNA) sequence data. Comparison of the captive and wild populations showed low levels of genetic diversity within the captive population and significant genetic differentiation among the captive populations and also between the wild and captive populations, suggesting at chance effect during the founding process for the captive population and a subsequent genetic drift. Therefore, for successful reintroduction, it is important that the reintroduced population reflects all the genetic diversity available from the captive populations, and that releasing a large number of individuals that consist of all captive populations.     

Genetic wealth,population health: Major histocompatibility complex variation in captive and wild ring‐tailed lemurs (Lemur catta)

Across species, diversity at the major histocompatibility complex (MHC) is critical to individual disease resistance and, hence, to population health; however, MHC diversity can be reduced in small, fragmented, or isolated populations. Given the need for comparative studies of functional genetic diversity, we investigated whether MHC diversity differs between populations which are open, that is experiencing gene flow, versus populations which are closed, that is isolated from other populations. Using the endangered ring‐tailed lemur (Lemur catta) as a model, we compared two populations under long‐term study: a relatively “open,” wild population (n = 180) derived from Bezà Mahafaly Special Reserve, Madagascar (2003–2013) and a “closed,” captive population (n = 121) derived from the Duke Lemur Center (DLC, 1980–2013) and from the Indianapolis and Cincinnati Zoos (2012). For all animals, we assessed MHC‐DRB diversity and, across populations, we compared the number of unique MHC‐DRB alleles and their distributions. Wild individuals possessed more MHC‐DRB alleles than did captive individuals, and overall, the wild population had more unique MHC‐DRB alleles that were more evenly distributed than did the captive population. Despite management efforts to maintain or increase genetic diversity in the DLC population, MHC diversity remained static from 1980 to 2010. Since 2010, however, captive‐breeding efforts resulted in the MHC diversity of offspring increasing to a level commensurate with that found in wild individuals. Therefore, loss of genetic diversity in lemurs, owing to small founder populations or reduced gene flow, can be mitigated by managed breeding efforts. Quantifying MHC diversity within individuals and between populations is the necessary first step to identifying potential improvements to captive management and conservation plans.     

Contemporary effective population size and predicted maintenance of genetic diversity in the endangered kea (Nestor notabilis)

Population size and the potential for maintenance of genetic diversity are critical information for the monitoring of species of conservation concern. However, direct estimates of population size are not always feasible, making indirect genetic approaches a valuable alternative. We estimated contemporary effective population size (N_e) in the endangered kea (Nestor notabilis) using three different methods. We then inferred the census size (N_C) using published N_e/N_C ratios and modelled the future maintenance of genetic diversity assuming a number of demographic parameters. Short-term N_e was small with a range-wide N_e?N_C was within the range of the current estimate (c. 1000–5000). Forward simulations showed low probability of retaining 90% of rare alleles without immigration. However, the probability of maintaining genetic diversity was high with immigration, juvenile survival of?≥?30%, and an initial sex ratio of c. 0.5–0.6. Despite the low N_e in kea, predator control and/or artificial immigration might be sufficient to maintain the present genetic diversity.     

Evaluating ex situ conservation projects: Genetic structure of the captive population of the Arabian sand cat

Ex situ conservation plays an increasingly important role in the conservation of endangered species. Molecular genetic markers can be helpful to assess the status of captive breeding programmes. We present the first molecular genetic analysis of the captive population of the Arabian sand cat (Felis margarita harrisoni) using microsatellites. Our data indicates that the captive population of F. m. harrisoni comprises three genetic clusters, which are based on different founder lineages. Genetic diversity was relatively high, the effective population size even exceeded the number of founders. This was presumably caused by subsequently integrating unrelated, genetically diverse founders into the captive population and a careful management based on minimizing kinship. However, we detected an error in the studbook records, which might have led to incestuous matings and underlines the usefulness of molecular evaluations in captive breeding programmes for endangered species.     

Genome‐wide linkage disequilibrium and past effective population size in three Korean cattle breeds

The routine collection and use of genomic data are useful for effectively managing breeding programs for endangered populations. Linkage disequilibrium (LD) using high‐density DNA markers has been widely used to determine population structures and predict the genomic regions that are associated with economic traits in beef cattle. The extent of LD also provides information about historical events, including past effective population size (N_e), and it allows inferences on the genetic diversity of breeds. The objective of this study was to estimate the LD and N_e in three Korean cattle breeds that are genetically similar but have different coat colors (Brown, Brindle and Jeju Black Hanwoo). Brindle and Jeju Black are endangered breeds with small populations, whereas Brown Hanwoo is the main breeding population in Korea. DNA samples from these cattle breeds were genotyped using the Illumina BovineSNP50 Bead Chip. We examined 13 cattle breeds, including European taurines, African taurines and indicines, and hybrids to compare their LD values. Brown Hanwoo consistently had the lowest mean LD compared to Jeju Black, Brindle and the other 13 cattle breeds (0.13, 0.19, 0.21 and 0.15–0.22 respectively). The high LD values of Brindle and Jeju Black contributed to small N_e values (53 and 60 respectively), which were distinct from that of Brown Hanwoo (531) for 11 generations ago. The differences in LD and N_e for each breed reflect the breeding strategy applied. The N_e for these endangered cattle breeds remain low; thus, effort is needed to bring them back to a sustainable tract.     

Breeding without breeding: minimum fingerprinting effort with respect to the effective population size

We present a probabilistic model to minimize the fingerprinting effort associated with the implementation of the “breeding without breeding” scheme under partial pedigree reconstruction. Our approach is directed at achieving a declared target population’s minimum effective population size (N _e ), following the pedigree reconstruction and genotypic selection and is based on the graph theory algorithm. The primary advantage of the proposed method is to reduce the cost associated with fingerprinting before the implementation of the pedigree reconstruction for seed parent–offspring derived from breeding arboreta and production or natural populations. Stochastic simulation was conducted to test the method’s efficiency assuming a simple polygenic model and a single trait. Hypothetical population consisted of 30 parental trees that were paired at random (selfing excluded), resulting in 600 individuals (potential candidates for forwards selection). The male parentage was assumed initially unknown. The model was used to estimate the minimum genotyping sample size needed to reaching the prescribed N _e . Results were compared with the known pedigree data. The model was successful in revealing the true relationship pattern over the whole range of N _e . Two to three offspring entered genotyping to meet the N _e  = 2 while 41 to 43 were required to satisfy the N _e  = 14. Importantly, genetic gain was affected at the lower limits of the genotyping effort. Doubling the number of parents resulted in considerable reduction of the genotyping effort at higher N _e values.     

Conservation genetics and population history of the threatened European mink Mustela lutreola, with an emphasis on the west European population

In species of great conservation concern, special attention must be paid to their phylogeography, in particular the origin of animals for captive breeding and reintroduction. The endangered European mink lives now in at least three well-separated populations in northeast, southeast and west Europe. Our aim is to assess the genetic structure of these populations to identify 'distinct population segments' (DPS) and advise captive breeding programmes. First, the mtDNA control region was completely sequenced in 176 minks and 10 polecats. The analysis revealed that the western population is characterized by a single mtDNA haplotype that is closely related to those in eastern regions but nevertheless, not found there to date. The northeast European animals are much more variable (pi = 0.012, h = 0.939), with the southeast samples intermediate (pi = 0.0012, h = 0.469). Second, 155 European mink were genotyped using six microsatellites. The latter display the same trends of genetic diversity among regions as mtDNA [gene diversity and allelic richness highest in northeast Europe (H(E) = 0.539, R(S) = 3.76), lowest in west Europe (H(E) = 0.379, R(S) = 2.12)], and provide evidences that the southeast and possibly the west populations have undergone a recent bottleneck. Our results indicate that the western population derives from a few animals which recently colonized this region, possibly after a human introduction. Microsatellite data also reveal that isolation by distance occurs in the western population, causing some inbreeding because related individuals mate. As genetic data indicate that the three populations have not undergone independent evolutionary histories for long (no phylogeographical structure), they should not be considered as distinct DPS. In conclusion, the captive breeding programme should use animals from different parts of the species' present distribution area.     

Genetic structure analysis of a highly inbred captive population of the African antelope Addax nasomaculatus. Conservation and management implications

The African antelope Addax nasomaculatus is a rare mammal at high risk of extinction, with no more than 300 individuals in the wild and 1,700 captive animals distributed in zoos around the world. In this work, we combine genetic data and genealogical information to assess the structure and genetic diversity of a captive population located at Parque Lecocq Zoo (N=27), originated from only two founders. We amplified 39 microsatellites previously described in other Artiodactyls but new to this species. Seventeen markers were polymorphic, with 2–4 alleles per locus (mean=2.71). Mean expected heterozygosity (He) per locus was between 0.050 (marker ETH3) and 0.650 (marker D5S2), with a global He of 0.43. The mean inbreeding coefficient of the population computed from pedigree records of all registered individuals (N=53) was 0.222. The mean coancestry of the population was 0.298 and F_IS index was ?0.108. These results reflect the importance of an adequate breeding management on a severely bottlenecked captive population, which would benefit by the incorporation of unrelated individuals. Thanks to the successful amplification of a large number of microsatellites commonly used in domestic bovids, this study will provide useful information for the management of this population and serve as future reference for similar studies in other captive populations of this species. Zoo Biol 30:399–411, 2011. © 2010 Wiley‐Liss, Inc.     

Effective population size in the captive breeding program of the Lake Victoria cichlid Paralabidochromis chilotes

Microsatellite DNA markers were used to monitor levels of genetic variation in 3 generations of the American Zoo and Aquarium Association [AZA] Species Survival Plan [SSP] captive breeding program for the Lake Victoria cichlid Paralabidochromis chilotes. Temporal changes in the frequency of 15 alleles, across four polymorphic loci, were used to estimate effective population size (N_e). The upper limit of the 95% confidence interval for N_e never exceeded eight individuals, with all of the corresponding N_e/N ratios falling below 0.15. A test of the proportion of expected heterozygous individuals between the F₁ and F₃ generations indicated a significant decline in expected heterozygosity of 5% per generation. Alternative husbandry protocols, including subdividing the captive population, are addressed to reduce the further loss of genetic variation. Zoo Biol 18:215–222, 1999. © 1999 Wiley‐Liss, Inc.     

Genetic evaluation of the Association of Zoos and Aquariums Matschie's tree kangaroo (Dendrolagus matschiei) captive breeding program

Matschie's tree kangaroo (Dendrolagus matschiei) is an endangered species that has been bred in captivity since the 1970s. In 1992, the Tree Kangaroo Species Survival Plan^® (TKSSP) was established to coordinate the captive management of Association of Zoos and Aquariums (AZA) D. matschiei. The TKSSP makes annual breeding recommendations primarily based on the mean kinship (MK) strategy. Captive breeding programs often use the MK strategy to preserve genetic diversity in small populations—to avoid the negative consequences of inbreeding and retain their adaptive potential. The ability of a captive breeding program to retain the population's genetic diversity over time can be evaluated by comparing the genetic diversity of the captive population to wild populations. We analyzed DNA extracted from blood and fecal samples from AZA (n = 71), captive (n = 28), and wild (n = 22) D. matschiei using eight microsatellite markers and sequenced the partial mitochondrial DNA control region gene. AZA D. matschiei had a similar expected heterozygosity (H_e = 0.595±0.184) compared with wild D. matschiei (H_e = 0.628±0.143), but they had different allelic frequencies (F_ST = 0.126; P<0.001). AZA D. matschiei haplotype diversity was almost two times lower than wild D. matschiei (). These data will assist management of AZA D. matschiei and serve as a baseline for AZA and wild D. matschiei genetic diversity values that could be used to monitor future changes in their genetic diversity. Zoo Biol 30:636–646, 2011. © 2010 Wiley Periodicals, Inc.     

A comparison of pedigree‐ and DNA‐based measures for identifying inbreeding depression in the critically endangered Attwater's Prairie‐chicken

The primary goal of captive breeding programmes for endangered species is to prevent extinction, a component of which includes the preservation of genetic diversity and avoidance of inbreeding. This is typically accomplished by minimizing mean kinship in the population, thereby maintaining equal representation of the genetic founders used to initiate the captive population. If errors in the pedigree do exist, such an approach becomes less effective for minimizing inbreeding depression. In this study, both pedigree‐ and DNA‐based methods were used to assess whether inbreeding depression existed in the captive population of the critically endangered Attwater's Prairie‐chicken (Tympanuchus cupido attwateri), a subspecies of prairie grouse that has experienced a significant decline in abundance and concurrent reduction in neutral genetic diversity. When examining the captive population for signs of inbreeding, variation in pedigree‐based inbreeding coefficients (f_pedigree) was less than that obtained from DNA‐based methods (f_DNA). Mortality of chicks and adults in captivity were also positively correlated with parental relatedness (r_DNA) and f_DNA, respectively, while no correlation was observed with pedigree‐based measures when controlling for additional variables such as age, breeding facility, gender and captive/release status. Further, individual homozygosity by loci (HL) and parental r_DNA values were positively correlated with adult mortality in captivity and the occurrence of a lethal congenital defect in chicks, respectively, suggesting that inbreeding may be a contributing factor increasing the frequency of this condition among Attwater's Prairie‐chickens. This study highlights the importance of using DNA‐based methods to better inform management decisions when pedigrees are incomplete or errors may exist due to uncertainty in pairings.     

Sexual selection has minimal impact on effective population sizes in species with high rates of random offspring mortality: An empirical demonstration using fitness distributions

The effective population size (N_e) is a fundamental parameter in population genetics that influences the rate of loss of genetic diversity. Sexual selection has the potential to reduce N_e by causing the sex‐specific distributions of individuals that successfully reproduce to diverge. To empirically estimate the effect of sexual selection on N_e, we obtained fitness distributions for males and females from an outbred, laboratory‐adapted population of Drosophila melanogaster. We observed strong sexual selection in this population (the variance in male reproductive success was ～14 times higher than that for females), but found that sexual selection had only a modest effect on N_e, which was 75% of the census size. This occurs because the substantial random offspring mortality in this population diminishes the effects of sexual selection on N_e, a result that necessarily applies to other high fecundity species. The inclusion of this random offspring mortality creates a scaling effect that reduces the variance/mean ratios for male and female reproductive success and causes them to converge. Our results demonstrate that measuring reproductive success without considering offspring mortality can underestimate N_e and overestimate the genetic consequences of sexual selection. Similarly, comparing genetic diversity among different genomic components may fail to detect strong sexual selection.     

Population viability analysis predicts decreasing genetic diversity in <Emphasis Type="Italic">ex situ</Emphasis> populations of the Itasenpara bitterling <Emphasis Type="Italic">Acheilognathus longipinnis</Emphasis> from the Kiso River,Japan

To conserve endangered species, the maintenance of ex situ captive populations with sustainable genetic diversity is often required, in combination with population viability analysis (PVA). Since 2010, the threatened Itasenpara bitterling Acheilognathus longipinnis lineages in the Kiso region, Japan, have been maintained in ex situ rearing facilities to allow for conservation efforts. In this study, we obtained microsatellite data from DNA extracted from these captive populations to elucidate their genetic diversity and effective population size. The populations of several initial generations indicated a deviation from Hardy–Weinberg equilibrium, probably due to the limited number of extracted founder individuals analyzed. The effective population size of the captive population tended to increase over the course of generations, although the degree of genetic diversity tended to decrease highlighting the concern for the progression of inbreeding. Our prediction based on the PVA suggests that the maintenance of the captive population under the current conditions could lead to extinction of the Itasenpara bitterling in 50 years. In contrast, simultaneously increasing the carrying capacity and individual exchange among populations appears to enhance the effective management of captive Itasenpara bitterling populations.     

Genetic diversity of North American captive‐born gorillas (Gorilla gorilla gorilla)

Western lowland gorillas (Gorilla gorilla gorilla) are designated as critically endangered and wild populations are dramatically declining as a result of habitat destruction, fragmentation, diseases (e.g., Ebola) and the illegal bushmeat trade. As wild populations continue to decline, the genetic management of the North American captive western lowland gorilla population will be an important component of the long‐term conservation of the species. We genotyped 26 individuals from the North American captive gorilla collection at 11 autosomal microsatellite loci in order to compare levels of genetic diversity to wild populations, investigate genetic signatures of a population bottleneck and identify the genetic structure of the captive‐born population. Captive gorillas had significantly higher levels of allelic diversity (t₇ = 4.49, P = 0.002) and heterozygosity (t₇ = 4.15, P = 0.004) than comparative wild populations, yet the population has lost significant allelic diversity while in captivity when compared to founders (t₇ = 2.44, P = 0.04). Analyses suggested no genetic evidence for a population bottleneck of the captive population. Genetic structure results supported the management of North American captive gorillas as a single population. Our results highlight the utility of genetic management approaches for endangered nonhuman primate species.     

Considerations for monitoring population trends of colonial waterbirds using the effective number of breeders and census estimates

Detecting trends in population size fluctuations is a major focus in ecology, evolution, and conservation biology. Populations of colonial waterbirds have been monitored using demographic approaches to determine annual census size (N_a). We propose the addition of genetic estimates of the effective number of breeders (N_b) as indirect measures of the risk of loss of genetic diversity to improve the evaluation of demographics and increase the accuracy of trend estimates in breeding colonies. Here, we investigated which methods of the estimation of N_b are more precise under conditions of moderate genetic diversity, limited sample sizes and few microsatellite loci, as often occurs with natural populations. We used the wood stork as a model species and we offered a workflow that researchers can follow for monitoring bird breeding colonies. Our approach started with simulations using five estimators of N_b and the theoretical results were validated with empirical data collected from breeding colonies settled in the Brazilian Pantanal wetland. In parallel, we estimated census size using a corrected method based on counting active nests. Both in simulations and in natural populations, the approximate Bayesian computation (ABC) and sibship assignment (SA) methods yielded more precise estimates than the linkage disequilibrium, heterozygosity excess, and molecular coancestry methods. In particular, the ABC method performed best with few loci and small sample sizes, while the other estimators required larger sample sizes and at least 13 loci to not underestimate N_b. Moreover, according to our N_b/N_a estimates (values were often ≤0.1), the wood stork colonies evaluated could be facing the loss of genetic diversity. We demonstrate that the combination of genetic and census estimates is a useful approach for monitoring natural breeding bird populations. This methodology has been recommended for populations of rare species or with a known history of population decline to support conservation efforts.     

Using molecular markers for pedigree reconstruction of the greater amberjack (Seriola dumerili) in the absence of parental information

Ensuring appropriate levels of genetic diversity in captive populations is essential to avoid inbreeding and loss of rare alleles by genetic drift. Pedigree reconstruction and parentage analysis in the absence of parental genotypes can be a challenging task that relies in the assignment of sibship relationships among the offspring. Here, we used eight highly variable microsatellite markers and three different assignment methods to reconstruct the most likely genotypes of a parental group of wild Seriola dumerili fish based on the genotypes of six cohorts of their offspring, to assess their relative contributions to the offspring. We found that a combination of the four most variable microsatellites was enough to identify the number of parents and their contribution to the offspring, suggesting that the variability of the markers can be more critical than the number of markers. Estimated effective population sizes were lower than the number of breeders and variable among years. The results suggest unequal parental contribution that should be accounted for breeding programs in the future.