Small effective population sizes of two remnant ocelot populations (<Emphasis Type="Italic">Leopardus pardalis albescens</Emphasis>) in the United States

Threatened populations are vulnerable to the effects of genetic drift and inbreeding, particularly when gene flow is low and the effective population size is small. Estimates of effective population size (N _e ) provide important information on the status of endangered populations that have experienced severe fragmentation and serve as indicators of genetic viability. Genetic data from microsatellite loci were used to estimate N _e for the 2 remaining populations of the endangered ocelot (Leopardus pardalis albescens) occurring in the United States. Several methods were used to calculate N _e , resulting in estimates ranging from N _e  = 8.0 (95% CI: 3.2–23.1) to 13.9 (95% CI: 7.7–25.1) for the population located at the Laguna Atascosa Wildlife Refuge in Cameron County, Texas. The ocelot population in Willacy County, Texas, had N _e estimates of 2.9 (95% CI: 1.7–5.6) and 3.1 (95% CI: 1.9–13.5), respectively. Estimates of N _e in both populations were below the critical value recommended for short-term viability.     

Population genetic structure and distribution of introduced American mink (<Emphasis Type="Italic">Mustela vison</Emphasis>) in Spain,based on microsatellite variation

The population genetic structure of an invasive species in Spain, the American mink (Mustela vison), was investigated using microsatellite DNA markers. This semi-aquatic carnivore, originating from North America, was imported into Europe for fur farming since the beginning of the 20th century. Due to massive escapes, farm damages, deliberate releases and/or accidents, feral mink populations were established in the aquatic ecosystems of many European countries, including Spain. We genotyped 155 American mink originating from the Spanish regions Basque Country, Catalonia, Castilla-Leon, Aragon, Valencia and Galicia using 10 polymorphic microsatellite loci to highlight population genetic structure, distribution and dispersal. M. vison populations in Spain appear differentiated and not yet connected by gene flow. Bayesian clustering analyses and spatial analyses of molecular variance detected four inferred clusters, overall coinciding with the sampled geographical localities. Preliminary testing shows moderate to large estimated effective population sizes. Molecular analyses result useful to provide baseline data for further research on the evolution of invasive mink populations, as well as support local management strategies and indirectly benefit the conservation of threatened species in Spain, such as the endangered European mink (Mustela lutreola), and the polecat (Mustela putorius), which share the habitat with the American mink. This paper is dedicated to the memory of Xavier Domingo-Roura.     

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.     

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.     

Effects of population characteristics and structure on estimates of effective population size in a house sparrow metapopulation

Effective population size (N_e) is a key parameter to understand evolutionary processes and the viability of endangered populations as it determines the rate of genetic drift and inbreeding. Low N_e can lead to inbreeding depression and reduced population adaptability. In this study, we estimated contemporary N_e using genetic estimators (LDNE, ONeSAMP, MLNE and CoNe) as well as a demographic estimator in a natural insular house sparrow metapopulation. We investigated whether population characteristics (population size, sex ratio, immigration rate, variance in population size and population growth rate) explained variation within and among populations in the ratio of effective to census population size (N_e/N_c). In general, N_e/N_c ratios increased with immigration rates. Genetic N_e was much larger than demographic N_e, probably due to a greater effect of immigration on genetic than demographic processes in local populations. Moreover, although estimates of genetic N_e seemed to track N_c quite well, the genetic N_e‐estimates were often larger than N_c within populations. Estimates of genetic N_e for the metapopulation were however within the expected range (<N_c). Our results suggest that in fragmented populations, even low levels of gene flow may have important consequences for the interpretation of genetic estimates of N_e. Consequently, further studies are needed to understand how N_e estimated in local populations or the total metapopulation relates to actual rates of genetic drift and inbreeding.     

Genetic Variability and Structuring of Arctic Charr (Salvelinus alpinus) Populations in Northern Fennoscandia

Variation in presumably neutral genetic markers can inform us about evolvability, historical effective population sizes and phylogeographic history of contemporary populations. We studied genetic variability in 15 microsatellite loci in six native landlocked Arctic charr (Salvelinus alpinus) populations in northern Fennoscandia, where this species is considered near threatened. We discovered that all populations were genetically highly (mean F _ST ≈ 0.26) differentiated and isolated from each other. Evidence was found for historical, but not for recent population size bottlenecks. Estimates of contemporary effective population size (N _e) ranged from seven to 228 and were significantly correlated with those of historical N _e but not with lake size. A census size (N _C) was estimated to be approximately 300 individuals in a pond (0.14 ha), which exhibited the smallest N _e (i.e. N _e/N _C = 0.02). Genetic variability in this pond and a connected lake is severely reduced, and both genetic and empirical estimates of migration rates indicate a lack of gene flow between them. Hence, albeit currently thriving, some northern Fennoscandian populations appear to be vulnerable to further loss of genetic variability and are likely to have limited capacity to adapt if selection pressures change.     

Naturally rare versus newly rare: demographic inferences on two timescales inform conservation of Galápagos giant tortoises

Long‐term population history can influence the genetic effects of recent bottlenecks. Therefore, for threatened or endangered species, an understanding of the past is relevant when formulating conservation strategies. Levels of variation at neutral markers have been useful for estimating local effective population sizes (N_e) and inferring whether population sizes increased or decreased over time. Furthermore, analyses of genotypic, allelic frequency, and phylogenetic information can potentially be used to separate historical from recent demographic changes. For 15 populations of Galápagos giant tortoises (Chelonoidis sp.), we used 12 microsatellite loci and DNA sequences from the mitochondrial control region and a nuclear intron, to reconstruct demographic history on shallow (past ~100 generations, ~2500 years) and deep (pre‐Holocene, >10 thousand years ago) timescales. At the deep timescale, three populations showed strong signals of growth, but with different magnitudes and timing, indicating different underlying causes. Furthermore, estimated historical N_e of populations across the archipelago showed no correlation with island age or size, underscoring the complexity of predicting demographic history a priori. At the shallow timescale, all populations carried some signature of a genetic bottleneck, and for 12 populations, point estimates of contemporary N_e were very small (i.e., < 50). On the basis of the comparison of these genetic estimates with published census size data, N_e generally represented ~0.16 of the census size. However, the variance in this ratio across populations was considerable. Overall, our data suggest that idiosyncratic and geographically localized forces shaped the demographic history of tortoise populations. Furthermore, from a conservation perspective, the separation of demographic events occurring on shallow versus deep timescales permits the identification of naturally rare versus newly rare populations; this distinction should facilitate prioritization of management action.     

Extremely Low Levels of Allozyme Variation in Southern Korean Populations of the Two Rare and Endangered Lithophytic or Epiphytic <Emphasis Type="Italic">Bulbophyllum Drymoglossum</Emphasis> and <Emphasis Type="Italic">Sarcanthus Scolopendrifolius</Emphasis> (Orchidaceae): Implications for Conservation

Allozyme variation was investigated in two local populations of Bulbophyllum drymoglossum and three populations Sarcanthus scolopendrifolius, two rare and endangered lithophytes and epiphytes from South Korea. Genetic diversity was extremely low within populations (mean H _e = 0.011 for B. drymoglossum; 0.002 for S. scolopendrifolius). Among the putative screened 21 loci, we found only one polymorphic locus for each species. Only one polymorphic locus, detected just one population of each species, revealed significantly high degree of population differentiation between and among populations (F _ST = 0.253 for B. drymoglossum and F _ST = 0.899 for S. scolopendrifolius). These results suggest that genetic drift (consequence of a very small effective population size), coupled with a limited gene flow would play a major role in shaping population genetics of these species in South Korea. The current status of both species (small population sizes, spatially isolated populations, and highly localized habitats) in addition to the extremely low levels of genetic diversity and reckless collection of endangered orchids by plant sellers, significantly threaten the long-term survival of these species in Korea. Conservation of the two species requires both in situ strategies, by introducing of genets to increase effective population sizes by minimizing adverse effects (e.g., outbreeding depression and genetic swamping by non-native genotypes), and ex situ strategies, such as collection of genets from clonal ramets.     

Reconciling Genetic and Demographic Estimators of Effective Population Size in the Anuran Amphibian Bufo Calamita

We used genetic and demographic methods to estimate the variance effective population sizes (N _e) of three populations of natterjack toads Bufo calamita in Britain. This amphibian breeds in temporary pools where survival rates can vary among families. Census population sizes (N) were derived from spawn string counts. Point and coalescent-based maximum likelihood estimates of N _e based on microsatellite allele distributions were similar. N _e/N ratios based on genetic estimates of N _e ranged between 0.02 and 0.20. Mean demographic estimates of N _e were consistently higher (2.7–8.0-fold) than genetic estimates for all three populations when variance in breeding success was evaluated at the point where females no longer influence their progeny. However, discrepancies between genetic and demographic estimators could be removed by using a model that included extra variance in survivorship (above to Poisson expectations) among families. The implications of these results for the estimation of N _e in wild populations are discussed.     

Genetic variation and its relationship to population size in reintroduced populations of pink sand verbena, Abronia umbellata subsp. breviflora (Nyctaginaceae)

Genetic monitoring of reintroduced plantpopulations can allow assessment of the successin establishing new populations thatgenetically resemble native populations. Weused a PCR-based method (Intersimple SequenceRepeats) to quantify genetic variation in fourreintroduced populations of Abroniaumbellata ssp. breviflora, an annualforb native to the Pacific Coast that isstate-listed endangered in Oregon. Thereintroduced populations ranged in size from 18to 4,111 individuals in the year they weresampled. Genetic variation was also quantifiedin the natural population that served as theseed source for the reintroduction efforts. Atotal of 77 loci (bands) was observed using twoISSR primers, providing 65 polymorphic loci. Asignificant, positive regression was observedbetween the log of population size for the fivepopulations and genetic variation when measuredas percent polymorphic loci (P), expectedheterozygosity (H_e> ), and with adissimilarity index (1 – S_xy) based on bandsharing. Two of the reintroduced populationsmaintained approximately 90% of the geneticvariation we observed in the source population. Based on these results, we predict thatreintroduced populations of A. u. ssp.breviflora that have at least 1,000individuals should maintain 90% of the geneticvariation of the source population.     

Genetic variation in relation to demography of peripheral pool frog populations (Rana lessonae)

Summary Genetic variation in an isolated northern metapopulation of the pool frog (Rana lessonae) in Sweden was compared to that of Central European populations using enzyme electrophoresis and literature data. Of the 31 loci scored, two (EST-2 andIDH-2) were polymorphic while no variation occurred in seven of the eight loci which are polymorphic in Central European populations.The heterozygosity level of the Swedish pool frogs is very low compared to that of other anuran populations, but their mean proportion of fertilized eggs within egg masses (97.5%) was not lower than in more heterozygous species, and their body size-specific fecundity did not differ from that of Polish conspecifics. The low genetic variability of the Swedish pool frogs is discussed in relation to features of the local populations such as size (N), calculated effective size (N _e ) reproductive success and probable history. It is concluded that long-term strong fluctuations in population size caused by reporductive failure in cold years have contributed more to the low genetic variability than could a single founder event due to a recent introduction by man.     

Tandem repeat markers for population genetic studies of the protected California tiger salamander (<Emphasis Type="Italic">Ambystoma californiense</Emphasis>)

Habitat loss is the single greatest threat to persistence of the critically threatened California tiger salamander (Ambystoma californiense). To aid management plans that designate critical habitat for this species, I developed and characterized 21 tetranucleotide microsatellite markers using two native populations in Santa Barbara and Alameda Counties. Allelic variation and average heterozygosities were lower in the endangered Santa Barbara population (allele range 1–4, mean 2.4; H _O = 0.308 H _E = 0.288) compared with the threatened Alameda population (allele range 2–10, mean 6.7; H _O = 0.712, H _E = 0.722). In-depth population studies using these markers will provide vital information for plans to assign critical habitat that optimize gene flow among breeding populations, as well as for identifying non-native hybrid genotypes that threaten native A. californiense stocks. Beyond the conservation goals for A. californiense, the close phylogenetic relationships within the tiger salamander complex also suggest a broad utility for population studies using these markers.     

INCREASED PROBABILITY OF EXTINCTION DUE TO DECREASED GENETIC EFFECTIVE POPULATION SIZE: EXPERIMENTAL POPULATIONS OF CLARKIA PULCHELLA

We established replicated experimental populations of the annual plant Clarkia pulchella to evaluate the existence of a causal relationship between loss of genetic variation and population survival probability. Two treatments differing in the relatedness of the founders, and thus in the genetic effective population size (N_e), were maintained as isolated populations in a natural environment. After three generations, the low N_e treatment had significantly lower germination and survival rates than did the high N_e treatment. These lower germination and survival rates led to decreased mean fitness in the low N_e populations: estimated mean fitness in the low N_e populations was only 21% of the estimated mean fitness in the high N_e populations. This inbreeding depression led to a reduction in population survival: at the conclusion of the experiment, 75% of the high N_e populations were still extant, whereas only 31% of the low N_e populations had survived. Decreased genetic effective population size, which leads to both inbreeding and the loss of alleles by genetic drift, increased the probability of population extinction over that expected from demographic and environmental stochasticity alone. This demonstrates that the genetic effective population size can strongly affect the probability of population persistence.     

Multiple estimates of effective population size for monitoring a long‐lived vertebrate: an application to Yellowstone grizzly bears

Effective population size (N_e) is a key parameter for monitoring the genetic health of threatened populations because it reflects a population's evolutionary potential and risk of extinction due to genetic stochasticity. However, its application to wildlife monitoring has been limited because it is difficult to measure in natural populations. The isolated and well‐studied population of grizzly bears (Ursus arctos) in the Greater Yellowstone Ecosystem provides a rare opportunity to examine the usefulness of different N_e estimators for monitoring. We genotyped 729 Yellowstone grizzly bears using 20 microsatellites and applied three single‐sample estimators to examine contemporary trends in generation interval (GI), effective number of breeders (N_b) and N_e during 1982–2007. We also used multisample methods to estimate variance (N_eV) and inbreeding N_e (N_eI). Single‐sample estimates revealed positive trajectories, with over a fourfold increase in N_e (≈100 to 450) and near doubling of the GI (≈8 to 14) from the 1980s to 2000s. N_eV (240–319) and N_eI (256) were comparable with the harmonic mean single‐sample N_e (213) over the time period. Reanalysing historical data, we found N_eV increased from ≈80 in the 1910s–1960s to ≈280 in the contemporary population. The estimated ratio of effective to total census size (N_e/N_c) was stable and high (0.42–0.66) compared to previous brown bear studies. These results support independent demographic evidence for Yellowstone grizzly bear population growth since the 1980s. They further demonstrate how genetic monitoring of N_e can complement demographic‐based monitoring of N_c and vital rates, providing a valuable tool for wildlife managers.     

Spatial and temporal genetic differentiation and effective population size of brown trout (<Emphasis Type="Italic">Salmo trutta</Emphasis>, L.) in small Danish rivers

The spatial and temporal genetic structure of brown trout populations from three small tributaries of Lake Hald, Denmark, was studied using analysis of variation at eight microsatellite loci. From two of the populations temporal samples were available, separated by up to 13 years (3.7 generations). Significant genetic differentiation was observed among all samples, however, hierarchical analysis of molecular variance (AMOVA) showed that differentiation among populations accounted for a non-significant amount of the genetic differentiation, whereas differentiation among temporal samples within populations was highly significant (0.0244, P<0.001). Estimates of effective population size (N _e) using a maximum-likelihood based implementation of the temporal method, yielded small values (N _e ranging from 33 to 79). When a model was applied that allows for migration among populations, N _e estimates were even lower (24–54), and migration rates were suggested to be high (0.13–0.36). All samples displayed a clear signal of a recent bottleneck, probably stemming from a period of unfavourable conditions due to organic pollution in the 1970–1980’s. By comparison to other estimates of N _e in brown trout, Lake Hald trout represent a system of small populations linked by extensive gene flow, whereas other populations in larger rivers exhibit much higher N _e values and experience lower levels of immigration. We suggest that management considerations for systems like Lake Hald brown trout should focus both on a regional scale and at the level of individual populations, as the future persistence of populations depends both on maintaining individual populations and ensuring sufficient migration links among these populations.     

Estimation of census and effective population sizes: the increasing usefulness of DNA-based approaches

Population census size (N _C) and effective population sizes (N _e) are two crucial parameters that influence population viability, wildlife management decisions, and conservation planning. Genetic estimators of both N _C and N _e are increasingly widely used because molecular markers are increasingly available, statistical methods are improving rapidly, and genetic estimators complement or improve upon traditional demographic estimators. We review the kinds and applications of estimators of both N _C and N _e, and the often undervalued and misunderstood ratio of effective-to-census size (N _e /N _C). We focus on recently improved and well evaluated methods that are most likely to facilitate conservation. Finally, we outline areas of future research to improve N _e and N _C estimation in wild populations.     

Demographic bottlenecks and low gene flow in remnant populations of the critically endangered <Emphasis Type="Italic">Berchemiella wilsonii</Emphasis> var<Emphasis Type="Italic">. pubipetiolata</Emphasis> (Rhamnaceae) inferred from microsatellite markers

Berchemiella wilsonii var. pubipetiolata (Rhamnaceae) is an endangered plant with only four remnant populations in eastern China. Population genetic information is essential for understanding population history and formulating conservation strategies for this species. Thirteen microsatellite loci were used to investigate genetic variation and population structure of the four remnant populations. Moderate levels of expected heterozygosity (H _E = 0.466–0.543) and low allelic diversity (A = 3.1–3.6 and A _R = 2.2–2.4, respectively) were observed within populations. Bottleneck tests found three out of four populations to deviate from mutation-drift equilibrium under the two-phase model (TPM), suggesting a recent population decline, which is congruent with known demographic history. The evolutionary history of the species seems dominated by genetic drift rather than gene flow. Low historical gene flow was inferred from several different approaches and N _m ranged from 0.582 by the private allele method to 0.783 by the coalescent method. Contemporary gene flow was also found to be even lower for only one first generation migrant was detected with individual-based assignment analysis. Restricted pollen and seed dispersal as well as a recent decline in population size associated with habitat fragmentation may have contributed to low levels of historical and contemporary gene flow, and resulted in a high genetic differentiation. Under this scenario, Berchemiella wilsonii var. pubipetiolata populations are expected to display more pronounced population genetic structure in the future as a result of increased inbreeding and genetic drift.     

Trapped within the city: integrating demography,time since isolation and population‐specific traits to assess the genetic effects of urbanization

Urbanization is a severe form of habitat fragmentation that can cause many species to be locally extirpated and many others to become trapped and isolated within an urban matrix. The role of drift in reducing genetic diversity and increasing genetic differentiation is well recognized in urban populations. However, explicit incorporation and analysis of the demographic and temporal factors promoting drift in urban environments are poorly studied. Here, we genotyped 15 microsatellites in 320 fire salamanders from the historical city of Oviedo (Est. 8th century) to assess the effects of time since isolation, demographic history (historical effective population size; N_e) and patch size on genetic diversity, population structure and contemporary N_e. Our results indicate that urban populations of fire salamanders are highly differentiated, most likely due to the recent N_e declines, as calculated in coalescence analyses, concomitant with the urban development of Oviedo. However, urbanization only caused a small loss of genetic diversity. Regression modelling showed that patch size was positively associated with contemporary N_e, while we found only moderate support for the effects of demographic history when excluding populations with unresolved history. This highlights the interplay between different factors in determining current genetic diversity and structure. Overall, the results of our study on urban populations of fire salamanders provide some of the very first insights into the mechanisms affecting changes in genetic diversity and population differentiation via drift in urban environments, a crucial subject in a world where increasing urbanization is forecasted.     

Temporal genetic samples indicate small effective population size of the endangered yellow-eyed penguin

There is an increasing awareness that the long-term viability of endemic island populations is negatively affected by genetic factors associated with population bottlenecks and/or persistence at small population size. Here we use contemporary samples and historic museum specimens (collected 1888–1938) to estimate the effective population size (N _e) for the endangered yellow-eyed penguin (Megadyptes antipodes) in South Island, New Zealand, and evaluate the genetic concern for this iconic species. The South Island population of M. antipodes—constituting almost half of the species’ census size—is thought to be descended from a small number of founders that reached New Zealand just a few hundred years ago. Despite intensive conservation measures, this population has shown dramatic fluctuations in size over recent decades. We compare estimates of the harmonic mean N _e for this population, obtained using one moment and three likelihood based-temporal methods, including one method that simultaneously estimates migration rate. Evaluation of the N _e estimates reveals a harmonic mean N _e in the low hundreds. Additionally, the inferred low immigration rates (m = 0.003) agree well with contemporary migration rate estimates between the South Island and subantarctic populations of M. antipodes. The low N _e of South Island M. antipodes is likely affected by strong fluctuations in population size, and high variance in reproductive success. These results show that genetic concerns for this population are valid and that the long-term viability of this species may be compromised by reduced adaptive potential.     

Genetic diversity through time and space: diversity and demographic history from natural history specimens and serially sampled contemporary populations of the threatened Gouldian finch (<Emphasis Type="Italic">Erythrura gouldiae</Emphasis>)

Declines in population size can compromise the viability of populations by reducing the effective population size (N_e), which may result in loss of genetic diversity and inbreeding. Temporal population genetic data can be a powerful tool for testing the presence and severity of reductions in N_e. The Gouldian finch (Erythrura gouldiae) is a flagship for conservation of Australian monsoonal savanna species. This species underwent severe population declines in the twentieth century due to land use changes associated with European colonization. Microsatellite and mitochondrial genetic data from Gouldian finch samples sourced from natural history collections prior to land use changes were compared with contemporary samples to estimate the severity of decline in effective population size and to detect changes in gene flow. These data show that Gouldian finch decline was not as severe as some sources suggest, and that population genetic connectivity has not changed following land use changes in the twentieth century. Multiple estimators of current N_e using genetic data from consecutive years suggest the Gouldian finch N_e is likely between a few hundred and a few thousand individuals, with some estimates within the range considered of conservation concern. This work has identified the need to genetically characterize populations in Queensland, and to understand critical demographic parameters (e.g. lifespan) in the Gouldian finch. Understanding these factors is vital to further improve genetic estimates of population size, key to the formation of appropriate conservation management of this species.