When the shoe doesn’t fit: applying conservation unit concepts to western painted turtles at their northern periphery

As biodiversity continues to be lost at an alarming rate, strategies for prioritizing populations for conservation have become increasingly important. Maintaining intraspecific genetic diversity is of particular importance for preserving evolutionary history and the potential for future adaptation. In order to effectively protect this diversity, units below the species level need to be defined. However, delineation of such units is subject to many challenges, with no one strategy applying universally across taxa. In this study we carried out the first genetic assessment of the western painted turtle (Chrysemys picta bellii) at its northern periphery in British Columbia (BC), Canada, using mitochondrial DNA haplotypic and microsatellite genotypic data to examine population structure and demographic history. We compared the application of evolutionarily significant unit and management unit criteria with Canadian designatable unit guidelines to determine appropriate conservation units. Our results show that BC western painted turtles form a single evolutionarily significant unit, with each occupied site constituting a separate management unit. In contrast, there is evidence for six discrete designatable units. Patterns of genetic variation in BC western painted turtles indicate that the conservation of each region is important to maintaining regional diversity and evolutionary novelty in this widespread species.     

Applications of mitochondrial DNA analysis in conservation: a critical review

Patterns of variation in mitochondrial DNA (mtDNA) increasingly are being investigated in threatened or managed species, but not always with clearly defined goals for conservation. In this review I identify uses of mtDNA analysis which fall into two different areas: (i) 'gene conservation' - the identification and management of genetic diversity, and (ii) 'molecular ecology' - the use of mtDNA variation to guide and assist demographic studies of populations. These two classes of application have different conceptual bases, conservation goals and time-frames. Gene conservation makes extensive use of phylogenetic information and is, in general, most relevant to long-term planning. Appropriate uses here include identification of Evolutionarily Significant Units and assessment of conservation priority of taxa or areas from an evolutionary perspective. Less appropriate are inferences about fitness from within-population diversity and about species boundaries. Molecular ecology makes more use of allele frequencies and provides information useful for short-term management of populations. Powerful applications are to identify Management Units and to define and use naturally occurring genetic tags. Estimating demographic parameters, e.g migration rate and population size, from patterns of mtDNA diversity is fraught with difficulty, particularly where populations are fluctuating, and is unlikely to produce quantitative estimates sufficiently accurate to be useful for practical management of contemporary populations. However, through comparative studies, mtDNA analysis can provide qualitative signals of population changes, allowing efficient targeting of resource-intensive ecological studies. Thus, there are some relatively straightforward uses of mtDNA, preferably in conjunction with assays of nuclear variation, that can make a significant contribution to the long-term planning and short-term execution of species recovery plans.     

华木莲居群遗传结构与保护单元

华木莲(Sinomanglietia glauca)仅分布于江西宜春和湖南永顺, 是我国一级重点保护植物。前人采用RAPD、ISSR以及叶绿体SSCP(single-stranded conformation polymorphism)标记对华木莲进行了居群遗传学研究, 但未包括后发现的湖南居群或未检出居群内遗传变异。为了全面检测华木莲遗传多样性及其空间分布格局, 并据此确定保护单元, 本研究采用细胞核微卫星标记对华木莲所有4个居群共77个个体进行了居群遗传学分析。结果表明, 华木莲具有较低的遗传多样性(平均等位基因数A = 2.604, 平均期望杂合度H_E = 0.423)和较高的遗传分化(F_ST = 0.425)。STRUCTURE和主成分分析(Principal Coordinated Analysis, PCA)将4个居群首先分为江西、湖南两组, 江西的2个居群实际上是同一个自然繁育居群, 而湖南的2个居群则为2个分化明显的自然繁育居群。研究还发现湖南居群存在明显的杂合子过剩现象, 可能是小居群内随机因素造成的。研究结果表明华木莲可能在近期历史上遭受过强烈的瓶颈效应, 导致种群缩小、遗传多样性丧失和居群分化加剧, 需要加强对其进化潜力的保护。在制定保护措施时, 需要考虑其较高的遗传分化水平, 根据遗传结构可以将其划分为3个保护单元, 即湖南居群和江西居群分别为2个进化显著单元, 湖南居群进一步划分为2个管理单元(分别为朗溪乡云盘村和小溪乡鲁家村居群)。     

Catchments as conservation units for riverine biodiversity

The geological structure and longitudinal nature of river systems provide a possible barrier to the dispersal of lotic organisms. This has the potential to drive evolutionary processes such as genetic differentiation and subsequent allopatric speciation. In the conservation of lotic ecosystems population and evolutionary processes have largely been ignored. The traditional approach to river conservation has been focussed toward maintaining the physical habitat that provides the template for the biota. With a shift toward recognising the catchment as the primary operational unit for the conservation and management of lotic systems an examination of the biological relevance of these catchment units is required. This paper examines the structural influence of catchment units over population structure in lotic organisms and the extent to which catchments reflect populations and represent evolutionarily significant units. These being unique assemblages representing segments of biological diversity that share a common evolutionary lineage and contain the potential for a unique evolutionary future. This is of particular importance given the increased use of inter-basin water transfers to move water between historically isolated catchments and recognition of the catchment as the primary unit for the conservation and management of river ecosystems.     

Conservation implications of significant population differentiation in an endangered estuarine seahorse

The spatial distribution of a species’ genetic diversity can provide insights into underlying evolutionary, ecological and environmental processes, and can contribute information towards the delineation of conservation units. The Knysna seahorse, Hippocampus capensis, is endangered and occurs in only three estuaries on the warm-temperate south coast of South Africa: Knsyna, Keurbooms and Swartvlei. Population sizes in the latter two estuaries have been very small for a prolonged period of time, and the populations residing in them may thus benefit from translocations as a means of increasing population sizes and possibly also genetic diversity. However, information on whether these three estuaries constitute distinct conservation units that warrant separate management is presently lacking. Here, we used genetic information from mitochondrial (control region) and nuclear microsatellite loci to assess the genetic diversity and spatial structure across the three estuaries, and also whether translocations should be included in the management plan for the Knysna seahorse. Although each population had a unique combination of alleles, and clustering methods identified the Swartvlei Estuary as being distinct from the others, levels of genetic admixture were high, and there was no evidence for reciprocal monophyly that would indicate that each estuary has a unique demographic history. On these grounds, we suggest recognising the three populations as a single evolutionarily significant unit (ESU), and encourage translocations between them to ensure the species’ long-term survival.     

Disjunct,highly divergent genetic lineages within two rare Eremophila (Scrophulariaceae: Myoporeae) species in a biodiversity hotspot: implications for taxonomy and conservation

Effective conservation management should target appropriate conservation units, but evolutionarily and genetically divergent lineages within nominal taxa are often unrecognized. The south‐western Australian biodiversity hotspot may harbour many cryptic taxa, as it contains many plant species with naturally fragmented population distributions. Using microsatellite markers, we tested the hypothesis that disjunct population groups in the rare species Eremophila microtheca and E. rostrata (Scrophulariaceae: Myoporeae) are highly genetically divergent and represent separate evolutionarily significant units (ESUs). Chromosome counts indicated that all individuals assessed were diploid (2n = 36). Genetic differentiation among disjunct population groups was highly significant (P < 0.001) for both E. microtheca (F_ST = 0.301–0.383; D_est = 0.756–0.774) and E. rostrata (F_ST = 0.325–0.346; D_est = 0.628–0.660), and was similar to their differentiation from allied species. These results, including high incidences of private alleles, suggest historical divergence among cryptic taxa within E. microtheca and E. rostrata. Population groups in E. rostrata have recently been taxonomically recognized as two subspecies. Our study suggests that E. microtheca should also be reassessed as two taxa or considered as two ESUs, and the southern occurrence should be listed as Critically Endangered. We suggest a precautionary approach for flora in this and similar landscapes, whereby historically wide geographical disjunctions are assumed to indicate separate units for conservation. © 2014 State of Western Australia. Botanical Journal of the Linnean Society © 2014 The Linnean Society of London, 2015, 177 , 96–111.     

Identifying evolutionarily significant units for conservation of the endangered Malus sieversii using genome‐wide RADseq data

Malus sieversii, a wild progenitor of the domesticated apple, is an endangered species and is assigned second conservation priority by the China Plant Red Data Book. It is urgent to carry out in situ conservation of this species, but previous studies have not identified evolutionarily significant units (ESUs) for conservation management. In this study, we investigated the genetic diversity and relationships of six M. sieversii populations from China using integrated analysis of microsatellite (nSSR) data, genome‐wide SNPs and previous results in order to propose a reasonable conservation management. The results showed that levels of genetic diversity were inconsistently reflected by our nSSR and previous studies, suggesting that indices of genetic diversity are not effective to identify priority conservation areas for M. sieversii. Based on the selection criteria of ESUs for endangered species conservation, ESUs should reflect lineage divergence, geographical separation and different adaptive variation. Our phylogenetic tree based on genome‐wide SNPs yielded a clear relationship of divergent lineages among M. sieversii populations, leading to new different from those of previous studies. Three independent lineages, including the pairs of populations Huocheng‐Yining, Gongliu‐Xinyuan and Tuoli‐Emin, were identified. The geographic distances between populations among the different phylogenetic lineages were much greater than those within the same phylogenetic lineage. A cluster analysis on environmental variables showed that the three independent lineages inhabit different environmental conditions, suggesting that they may have adapted to different environments. Based on the results, we propose that three independent ESUs should be recognized as conservation units for M. sieversii in China.     

The subspecies concept in butterflies: has its application in taxonomy and conservation biology outlived its usefulness?

Subspecies lie at the interface between systematics and population genetics, and represent a unit of biological organization in zoology that is widely used in the disciplines of taxonomy and conservation biology. In this review, we explore the utility of subspecies in relation to their application in systematics and biodiversity conservation, and briefly summarize species concepts and criteria for their diagnosis, particularly from an invertebrate perspective. The subspecies concept was originally conceived as a formal means of documenting geographical variation within species based on morphological characters; however, the utility of subspecies is hampered by inconsistencies by which they are defined conceptually, a lack of objective criteria or properties that serve to delimit their boundaries, and their frequent failure to reflect distinct evolutionary units according to population genetic structure. Moreover, the concept has been applied to populations largely comprising different components of genetic diversity reflecting contrasting evolutionary processes. We recommend that, under the general lineage (unified) species concept, the definition of subspecies be restricted to extant animal groups that comprise evolving populations representing partially isolated lineages of a species that are allopatric, phenotypically distinct, and have at least one fixed diagnosable character state, and that these character differences are (or are assumed to be) correlated with evolutionary independence according to population genetic structure. Phenotypic character types include colour pattern, morphology, and behaviour or ecology. Under these criteria, allopatric subspecies are a type of evolutionarily significant unit within species in that they show both neutral divergence through the effects of genetic drift and adaptive divergence under natural selection, and provide an historical context for identifying biodiversity units for conservation. Conservation of the adaptedness and adaptability of gene pools, however, may require additional approaches. Recent studies of Australian butterflies exemplify these points. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ?? , ??–??.     

Genetic structure in a montane ranid frog: restricted gene flow and nuclear-mitochondrial discordance

There is substantial debate over the criteria that should be used to group populations of a species into distinct units for conservation (e.g. evolutionarily significant units, management units, distinct population segments). However, in practice molecular genetic differentiation is often the only or main criterion used to identify such units. Most genetic studies attempting to define conservation units in animals use a single molecular marker, most often mitochondrial, and use samples from a limited number of populations throughout the species' range. Although there are many benefits to using mtDNA, certain features can cause it to show patterns of differentiation among populations that do not reflect the history of differentiation at the nuclear genome where loci controlling traits of adaptive significance presumably occur. Here we illustrate an example of such mitochondrial-nuclear discordance in a ranid frog, and show how using mtDNA or nuclear loci alone could have led to very different conservation recommendations. We also found very high genetic differentiation among populations on a local scale, and discuss the conservation implications of our results.     

Conservation genetics of South American aquatic mammals: an overview of gene diversity,population structure,phylogeography, non‐invasive methods and forensics

• 1 Most aquatic mammals have high dispersal potential, and there are often severe conservation concerns related to their legal or illegal harvesting. Therefore, economic, social and forensic factors often arise in decisions relating to their population management. Molecular markers are essential tools in modern conservation genetics, revealing previously unknown aspects of aquatic mammal behaviour, natural history, population structure and demography. Molecular markers also have been used to define management units, to recognize taxonomic units, to conduct forensic analyses and to control illegal wildlife trade, providing valuable information for decision‐making in wildlife conservation and management.
• 2 We review studies published in peer‐reviewed journals between 1993 and 2010, in which genetic approaches have been applied to conservation‐related issues involving natural populations of 25 species of aquatic mammals in South America. These studies cover just 34% of the 70 aquatic mammal species recorded in South America.
• 3 Most of the studies are related to population structure, phylogeography, gene flow and dispersal movements. In addition, recent findings relate to evolutionarily significant units, management units, forensics and conservation policy.
• 4 Finally, we look to the future and, based on numbers of studies and conservation concerns, suggest which species, geographic areas and genetic studies should be prioritized. Moreover, we discuss constraints on research and suggest collaborative works that would provide critical information towards the effective conservation and management of aquatic mammals in South America.

     

High genetic diversity of the endangered Iberian three‐spined stickleback (Gasterosteus aculeatus) at the Mediterranean edge of its range

1. The three‐spined stickleback (Gasterosteus aculeatus) on the Iberian Peninsula is only distributed in freshwater habitats and has completely disappeared from most of its range, mainly as a consequence of habitat degradation and invasive fish introductions. Genetic investigations have shown that Mediterranean‐Adriatic sticklebacks constitute an evolutionarily significant unit. Here, we present the first genetic data for Iberian populations living on the southern edge of the stickleback’s range. We used microsatellite markers to study gene diversity, population structure and genetic demography of stickleback populations. 2. High genetic differentiation among collections yielded a model of four genetically homogeneous units related to geography. The observed pattern of isolation by distance resulted mainly from the hydrographical pattern and limited gene flow among rivers. Moreover, low levels of gene diversity, high isolation and recent bottleneck events, which have led to small or even critical effective population sizes in several locations, could be explained by additional recent anthropogenic fragmentation. 3. We defined at least four evolutionarily significant units threatened by habitat fragmentation in north‐eastern Iberian sticklebacks. Because they retain long evolutionary histories, these populations should be considered of high conservation priority, and urgent management measures should be implemented.     

Considering evolutionary processes in conservation biology

Conservation biologists assign population distinctiveness by classifying populations as evolutionarily significant units (ESUs). Historically, this classification has included ecological and genetic data. However, recent ESU concepts, coupled with increasing availability of data on neutral genetic variation, have led to criteria based exclusively on molecular phylogenies. We argue that the earlier definitions of ESUs, which incorporated ecological data and genetic variation of adaptive significance, are more relevant for conservation. Furthermore, this dichotomous summary (ESU or not) of a continuum of population differentiation is not adequate for determining appropriate management actions. We argue for a broader categorization of population distinctiveness based on concepts of ecological and genetic exchangeability (sensu Templeton).     

Assessing natural and disturbed population structure in European grayling Thymallus thymallus: melding phylogeographic,population genetic and jurisdictional perspectives for conservation planning

Natural and human‐mediated population structure of European grayling Thymallus thymallus was assessed in primarily Austrian reaches of the Danube River catchment. Data on phylogeographic structure based on mitochondrial DNA (mtDNA) were combined with variation found across 13 microsatellite loci to assess introgression stemming from stocking activities. Populations north of the Alps generally showed no signs of long‐term introgression from allochthonous stocks, although one population comprised two distinct genotypic groups, one of which may stem from recently stocked material. South of the Alps, levels of introgression with stocks from the north were extensive and only one of six populations is believed to represent a reference state. Allelic diversity and expected heterozygosity were positively correlated with mtDNA admixture supporting more recent introgression and that there is little or no reproductive barrier between these two major lineages north and south of the Alps. A third unrelated mtDNA lineage is described from the Lafnitz, a tributary of the Raab drainage. The high lineage diversity in the upper Austrian Danube is not concordant with an existing model of T. thymallus evolutionarily significant units in Europe. Caveats in naming such units or following a strict hierarchical conservation unit structure for broadly distributed species with complex phylogeographic distributions stretching over various jurisdictions are discussed. The necessity of using both phylogeographic and population genetic approaches in evaluating the history and conservation value of populations in a conservation context is additionally highlighted.     

Perturbation,Restoration and Seeking Ecological Sustainability in Australian Flowing Waters

It is now accepted that most of the rivers and streams of Australia have been degraded to varying extents by European settlement. The scale and level of this degradation have been documented by State and Federal government authorities. To halt and reverse the degradation in the next two decades, it is crucial that the conservation and restoration of streams, rivers, riparian zones, and catchments become paramount in land and water management. Effective stream restoration requires a coordinated effort at the catchment level rather than at the level of many individual local sites. Monitoring, including the gathering of before-restoration data, and the setting of feasibly-attainable goals are key components of effective restoration. In planning projects and setting goals, it needs to be recognized that some goals may only be attained in the long-term. Large-scale restoration projects will require partnerships to be formed between resource managers and scientists, with other stakeholders possibly involved. Selected projects could be adaptively managed with the emphasis on gaining scientific knowledge on the effects of management interventions.     

Contrasting coarse and fine scale genetic structure among isolated relic populations of Kmeria septentrionalis

Trees of the Magnoliaceae family are of scientific, cultural and socio-economic importance. Kmeria septentrionalis Dandy (Magnoliaceae) is a dioecious tree, found in small, isolated, relic populations in Southern China, and is subject to extensive protection due to its rarity and high economic values. To improve conservation outcomes and in particular, germplasm collection guidelines, information on spatial genetic structure of the species is required. In this study, we investigated the spatial genetic structure and genetic diversity of 161 individuals of K. septentrionalis collected from five natural populations using AFLP molecular markers. Within-population genetic variation was measured, with percentage of polymorphic bands (PPB) ranged from 63% to 87%, while H _S (genetic diversity within population) varied from 0.185 to 0.244 with a mean of 0.215 ± 0.025. Significant genetic differentiations were revealed between pairwise populations, indicating each population existing as an independent evolutionarily significant unit. Mantel test results showed no pattern of isolation-by-distance among populations separated by large distance. Fine scale spatial patterns of genetic variation suggested significant effects of isolation-by-distance within population at distances of 22 m. The results of contrasting genetic structure at coarse and fine scale in K. septentrionalis may indicate restricted pollen flow and seed dispersal at fine scales, and separated evolution in isolated populations over long period of time at coarser scales. Finally, we make several suggestions for improved management practices that may assist in the conservation of this species.     

Genetic structure of the critically endangered Red‐headed Wood Pigeon Columba janthina nitens and its implications for the management of threatened island populations

The Red‐headed Wood Pigeon Columba janthina nitens is endemic to the Ogasawara Islands, an oceanic island chain located 1000 km south of the main islands of Japan. The subspecies is at high risk of extinction because of its small population size and restricted habitat range. We undertook genetic analyses of this pigeon using sequences of a portion of the mitochondrial control region and five microsatellite markers to estimate the genetic characteristics of two wild populations from the Bonin and Volcano Islands, as well as one captive breeding population. The genetic diversity of the wild individuals was exceptionally low in both the mitochondria (nucleotide diversity = 0.00105) and at the microsatellite (3.2 alleles per locus and H_E = 0.12) loci. Higher numbers of microsatellite genotypes were observed in the Volcano Islands population than in the Bonin Islands population, which may be because of the relatively low impact of human disturbance. The most common mitochondrial haplotypes and microsatellite alleles observed in the two wild populations were completely fixed in the captive population. Our results suggest that the genetic diversity of the captive population needs to be increased. However, introduction of a wild individual into a captive population can lead to a decreased genetic diversity in the wild population and therefore should be done with caution. The genetic differentiation between the Bonin and the Volcano island groups was low, and the populations of the two island groups should be regarded as a single evolutionarily significant unit. However, special consideration is required for habitat conservation in the Volcano Islands, which may be functioning as a sanctuary for the Red‐headed Wood Pigeon. For the long‐term conservation of threatened bird species that live on remote oceanic islands, determination of management units considering gene flow caused by their flying capacity and maintenance of genetically suitable wild and captive populations are essential.     

Multilocus assignment analyses reveal multiple units and rare migration events in the recently expanded yellow-eyed penguin (Megadyptes antipodes)

The identification of demographically independent populations and the recognition of management units have been greatly facilitated by the continuing advances in genetic tools. Managements units now play a key role in short-term conservation management programmes of declining species, but their importance in expanding populations receives comparatively little attention. The endangered yellow-eyed penguin ( Megadyptes antipodes ) expanded its range from the subantarctic to New Zealand's South Island a few hundred years ago and this new population now represents almost half of the species' total census size. This dramatic expansion attests to M. antipodes' high dispersal abilities and suggests the species is likely to constitute a single demographic population. Here we test this hypothesis of panmixia by investigating genetic differentiation and levels of gene flow among penguin breeding areas using 12 autosomal microsatellite loci along with mitochondrial control region sequence analyses for 350 individuals. Contrary to our hypothesis, however, the analyses reveal two genetically and geographically distinct assemblages: South Island vs. subantarctic populations. Using assignment tests, we recognize just two first-generation migrants between these populations (corresponding to a migration rate of < 2%), indicating that ongoing levels of long-distance migration are low. Furthermore, the South Island population has low genetic variability compared to the subantarctic population. These results suggest that the South Island population was founded by only a small number of individuals, and that subsequent levels of gene flow have remained low. The demographic independence of the two populations warrants their designation as distinct management units and conservation efforts should be adjusted accordingly to protect both populations.     

Conservation status of the White-Bellied Sea-Eagle <Emphasis Type="Italic">Haliaeetus leucogaster</Emphasis> in Australia determined using mtDNA control region sequence data

Considered to have a declining world population, concern has been expressed in recent years over the conservation status of the White-bellied Sea-Eagle Haliaeetus leucogaster (Gmelin, 1788) within Australia. We used mitochondrial (mtDNA) control region sequence data to investigate the current distribution of genetic variation in this species at the continental level and within and between specified regional units. We were specifically interested in identifying breaks in genetic connectivity between the west and east of the continent and between Tasmania and the Australian mainland. We also investigated the likelihood of a bottleneck at the time of colonisation, and propose hypotheses regarding colonisation history. Sequence data were obtained from 128 individuals describing 15 haplotypes. Overall, diversity was low and AMOVA results failed to provide any significant level of genetic subdivision between regions. We suggest that the population expanded from a bottleneck approximately 160,000 years ago during the late Pleistocene, and spread throughout the continent through a contiguous range expansion. There is insufficient evidence to suggest division of the population into different units for conservation management purposes based on the theoretical definition of the ‘evolutionary significant unit’. It is clear from the analysis that there are signatures of both historical and contemporary processes affecting the current distribution. Additional sampling and confirmation of the perceived pattern of population structure using a nuclear marker is recommended to validate conservation monitoring and management at a continental scale.     

Genetic differentiation across eastern Pacific oceanographic barriers in the threatened seahorse <Emphasis Type="Italic">Hippocampus ingens</Emphasis>

Understanding the population structure and evolutionary history of the eastern Pacific seahorse Hippocampus ingens is critical for the effective management of this threatened species. Life history characteristics of H. ingens (site fidelity and brooding of young) may limit gene flow and lead to population differentiation. A recent study analyzing conserved fragments of the mitochondrial cyt b and control region found no population structure. We re-assess this conclusion with a phylogeographic analysis of relationships among 115 individuals of H. ingens over a broader geographic range (San Diego Bay in California, Gulf of California, Central America, Ecuador, and Peru) based on a more variable 428 base pair fragment of the control region. This expanded analysis affirms low overall nucleotide diversity relative to other seahorses (θ_π = 0.004), and shows evidence of a recent bottleneck and population expansion since the middle Pleistocene. AMOVA analysis shows moderate overall population structure (Φ_ST = 0.10, P val = 0.00), and pairwise Φ_ST estimates indicate structure between the Gulf of California and all Pacific coast localities. Knowledge of population structure in H. ingens may improve conservation efforts by identifying evolutionarily important management units, and could determine source regions in the continuing trade of seahorses for traditional Chinese medicine. The level of genetic divergence observed between the Gulf of California and all other localities sampled may distinguish the Gulf as a separate management unit. Additional phylogeographic research with more quickly evolving genetic markers and targeted sampling at the mouth of the Gulf of California is warranted to inform strategies for conservation of this threatened seahorse.     

Subspecies of the Central American Squirrel Monkey (Saimiri oerstedii) as Units for Conservation

The accurate diagnosis of conservation units now typically includes recognition of genetic diversity and unique evolutionary lineages and is necessary to inform the conservation management of endangered species. We evaluated whether the two currently recognized subspecies of the endangered Central American squirrel monkey (Saimiri oerstedii) in Costa Rica are evolutionarily significant units (ESUs) that should be managed separately in conservation efforts. We used previously published sequences of 50 individuals of Saimiri oerstedii for 880 bp of the mtDNA d-loop and genotypes of 244 individuals for 16 microsatellites and conducted novel analyses to characterize genetic differentiation between subspecies of Saimiri oerstedii. We measured sequence differentiation and inferred an intraspecific molecular phylogeny and a haplotype network, and found consistent results supporting statistically significant divergence and reciprocal monophyly between subspecies. A population aggregation analysis also supported Saimiri oerstedii citrinellus and S. o. oerstedii as diagnosably distinct units. These results confirm previous genetic studies with smaller sample sizes and are consistent with other factors including differences in pelage and morphology and divergence at nuclear markers. Conservation managers should manage these subspecies separately to prevent the loss of genetic diversity via artificially induced outbreeding. High levels of genetic diversity may buffer populations against outside extinction pressures, to which Saimiri oerstedii are vulnerable because of their dwindling habitat and small population size.