Using a multi-gene approach to infer the complicated phylogeny and evolutionary history of lorises (Order Primates: Family Lorisidae).

Extensive phylogenetic studies have found robust phylogenies are modeled by using a multi-gene approach and sampling from the majority of the taxa of interest. Yet, molecular studies focused on the lorises, a cryptic primate family, have often relied on one gene, or just mitochondrial DNA, and many were unable to include all four genera in the analyses, resulting in inconclusive phylogenies. Past phylogenetic loris studies resulted in lorises being monophyletic, paraphyletic, or an unresolvable trichotomy with the closely related galagos. The purpose of our study is to improve our understanding of loris phylogeny and evolutionary history by using a multi-gene approach. We used the mitochondrial genes cytochrome b, and cytochrome c oxidase subunit 1, along with a nuclear intron (recombination activating gene 2) and nuclear exon (the melanocortin 1 receptor). Maximum Likelihood and Bayesian phylogenetic analyses were conducted based on data from each locus, as well as on the concatenated sequences. The robust, concatenated results found lorises to be a monophyletic family (Lorisidae) (PP ≥ 0.99) with two distinct subfamilies: the African Perodictinae (PP ≥ 0.99) and the Asian Lorisinae (PP ≥ 0.99). Additionally, from these analyses all four genera were all recovered as monophyletic (PP ≥ 0.99). Some of our single-gene analyses recovered monophyly, but many had discordances, with some showing paraphyly or a deep-trichotomy. Bayesian partitioned analyses inferred the most recent common ancestors of lorises emerged ∼42 ±â€¯6 million years ago (mya), the Asian Lorisinae separated ∼30 ±â€¯9 mya, and Perodictinae arose ∼26 ±â€¯10 mya. These times fit well with known historical tectonic shifts of the area, as well as with the sparse loris fossil record. Additionally, our results agree with previous multi-gene studies on Lorisidae which found lorises to be monophyletic and arising ∼40 mya (Perelman et al., 2011; Pozzi et al., 2014). By taking a multi-gene approach, we were able to recover a well-supported, monophyletic loris phylogeny and inferred the evolutionary history of this cryptic family.

Spatial genetic structure in American black bears (Ursus americanus): female philopatry is variable and related to population history.

Previously, American black bears (Ursus americanus) were thought to follow the pattern of female philopatry and male-biased dispersal. However, recent studies have identified deviations from this pattern. Such flexibility in dispersal patterns can allow individuals greater ability to acclimate to changing environments. We explored dispersal and spatial genetic relatedness patterns across ten black bear populations-including long established (historic), with known reproduction >50 years ago, and newly established (recent) populations, with reproduction recorded <50 years ago-in the Interior Highlands and Southern Appalachian Mountains, United States. We used spatially explicit, individual-based genetic simulations to model gene flow under scenarios with varying levels of population density, genetic diversity, and female philopatry. Using measures of genetic distance and spatial autocorrelation, we compared metrics between sexes, between population types (historic and recent), and among simulated scenarios which varied in density, genetic diversity, and sex-biased philopatry. In empirical populations, females in recent populations exhibited stronger patterns of isolation-by-distance (IBD) than females and males in historic populations. In simulated populations, low-density populations had a stronger indication of IBD than medium- to high-density populations; however, this effect varied in empirical populations. Condition-dependent dispersal strategies may permit species to cope with novel conditions and rapidly expand populations. Pattern-process modeling can provide qualitative and quantitative means to explore variable dispersal patterns, and could be employed in other species, particularly to anticipate range shifts in response to changing climate and habitat conditions.

Phylogeographic Analyses of American Black Bears (Ursus americanus) Suggest Four Glacial Refugia and Complex Patterns of Postglacial Admixture.

Studies of species with continental distributions continue to identify intraspecific lineages despite continuous habitat. Lineages may form due to isolation by distance, adaptation, divergence across barriers, or genetic drift following range expansion. We investigated lineage diversification and admixture within American black bears (Ursus americanus) across their range using 22 k single nucleotide polymorphisms and mitochondrial DNA sequences. We identified three subcontinental nuclear clusters which we further divided into nine geographic regions: Alaskan (Alaska-East), eastern (Central Interior Highlands, Great Lakes, Northeast, Southeast), and western (Alaska-West, West, Pacific Coast, Southwest). We estimated that the western cluster diverged 67 ka, before eastern and Alaskan divergence 31 ka; these divergence dates contrasted with those from the mitochondrial genome where clades A and B diverged 1.07 Ma, and clades A-east and A-west diverged 169 ka. We combined estimates of divergence timing with hindcast species distribution models to infer glacial refugia for the species in Beringia, Pacific Northwest, Southwest, and Southeast. Our results show a complex arrangement of admixture due to expansion out of multiple refugia. The delineation of the genomic population clusters was inconsistent with the ranges for 16 previously described subspecies. Ranges for U. a. pugnax and U. a. cinnamomum were concordant with admixed clusters, calling into question how to order taxa below the species level. Additionally, our finding that U. a. floridanus has not diverged from U. a. americanus also suggests that morphology and genetics should be reanalyzed to assess taxonomic designations relevant to the conservation management of the species.

Ecological resistance surfaces predict fine-scale genetic differentiation in a terrestrial woodland salamander.

Landscape genetics has seen tremendous advances since its introduction, but parameterization and optimization of resistance surfaces still poses significant challenges. Despite increased availability and resolution of spatial data, few studies have integrated empirical data to directly represent ecological processes as genetic resistance surfaces. In our study, we determine the landscape and ecological factors affecting gene flow in the western slimy salamander (Plethodon albagula). We used field data to derive resistance surfaces representing salamander abundance and rate of water loss through combinations of canopy cover, topographic wetness, topographic position, solar exposure and distance from ravine. These ecologically explicit composite surfaces directly represent an ecological process or physiological limitation of our organism. Using generalized linear mixed-effects models, we optimized resistance surfaces using a nonlinear optimization algorithm to minimize model AIC. We found clear support for the resistance surface representing the rate of water loss experienced by adult salamanders in the summer. Resistance was lowest at intermediate levels of water loss and higher when the rate of water loss was predicted to be low or high. This pattern may arise from the compensatory movement behaviour of salamanders through suboptimal habitat, but also reflects the physiological limitations of salamanders and their sensitivity to extreme environmental conditions. Our study demonstrates that composite representations of ecologically explicit processes can provide novel insight and can better explain genetic differentiation than ecologically implicit landscape resistance surfaces. Additionally, our study underscores the fact that spatial estimates of habitat suitability or abundance may not serve as adequate proxies for describing gene flow, as predicted abundance was a poor predictor of genetic differentiation.

Influence of drift and admixture on population structure of American black bears (Ursus americanus) in the Central Interior Highlands, USA, 50 years after translocation.

Bottlenecks, founder events, and genetic drift often result in decreased genetic diversity and increased population differentiation. These events may follow abundance declines due to natural or anthropogenic perturbations, where translocations may be an effective conservation strategy to increase population size. American black bears (Ursus americanus) were nearly extirpated from the Central Interior Highlands, USA by 1920. In an effort to restore bears, 254 individuals were translocated from Minnesota, USA, and Manitoba, Canada, into the Ouachita and Ozark Mountains from 1958 to 1968. Using 15 microsatellites and mitochondrial haplotypes, we observed contemporary genetic diversity and differentiation between the source and supplemented populations. We inferred four genetic clusters: Source, Ouachitas, Ozarks, and a cluster in Missouri where no individuals were translocated. Coalescent models using approximate Bayesian computation identified an admixture model as having the highest posterior probability (0.942) over models where the translocation was unsuccessful or acted as a founder event. Nuclear genetic diversity was highest in the source (AR = 9.11) and significantly lower in the translocated populations (AR = 7.07-7.34; P = 0.004). The Missouri cluster had the lowest genetic diversity (AR = 5.48) and served as a natural experiment showing the utility of translocations to increase genetic diversity following demographic bottlenecks. Differentiation was greater between the two admixed populations than either compared to the source, suggesting that genetic drift acted strongly over the eight generations since the translocation. The Ouachitas and Missouri were previously hypothesized to be remnant lineages. We observed a pretranslocation remnant signature in Missouri but not in the Ouachitas.

Effects of a hydropower project on a high-value Asian elephant population.

Habitat loss and fragmentation are leading contributors to the endangered status of species. In 2006, the Nakai Plateau contained the largest known Asian elephant (Elephas maximus) population in the Lao People's Democratic Republic (Lao PDR), and the population was among those with the highest genetic diversity reported for Asian elephants. In 2008, completion of the Nam Theun 2 hydroelectric dam inundated much of the Plateau, resulting in the loss of 40% of elephant habitat. We studied elephant presence, movements, and the incidence of human-elephant conflict (HEC) on the Nakai Plateau and surrounding areas from 2004 to 2020, before and for 12 years after dam completion. To examine contemporary population dynamics in the Nakai elephants, we used genetic sampling to compare minimum population numbers, demography, and levels of genetic diversity from the wet and dry seasons in 2018/2019, 10 years after dam completion, with those reported in a pre-dam-completion genetic survey. After dam completion, we found a major increase in HEC locally and the creation of new, serious, and persistent HEC problems as far as 100 km away. While we were unable to compare estimated population sizes before and after dam completion, our data revealed a decrease in genetic diversity, a male-biased sex ratio, and evidence of dispersal from the Plateau by breeding-age females. Our results raise concerns about the long-term viability of this important population as well as that of other species in this region. Given that hydropower projects are of economic importance throughout Laos and elsewhere in southeast Asia, this study has important implications for understanding and mitigating their impact.

Resolving the evolution of extant and extinct ruminants with high-throughput phylogenomics.

The Pecorans (higher ruminants) are believed to have rapidly speciated in the Mid-Eocene, resulting in five distinct extant families: Antilocapridae, Giraffidae, Moschidae, Cervidae, and Bovidae. Due to the rapid radiation, the Pecoran phylogeny has proven difficult to resolve, and 11 of the 15 possible rooted phylogenies describing ancestral relationships among the Antilocapridae, Giraffidae, Cervidae, and Bovidae have each been argued as representations of the true phylogeny. Here we demonstrate that a genome-wide single nucleotide polymorphism (SNP) genotyping platform designed for one species can be used to genotype ancient DNA from an extinct species and DNA from species diverged up to 29 million years ago and that the produced genotypes can be used to resolve the phylogeny for this rapidly radiated infraorder. We used a high-throughput assay with 54,693 SNP loci developed for Bos taurus taurus to rapidly genotype 678 individuals representing 61 Pecoran species. We produced a highly resolved phylogeny for this diverse group based upon 40,843 genome-wide SNP, which is five times as many informative characters as have previously been analyzed. We also establish a method to amplify and screen genomic information from extinct species, and place Bison priscus within the Bovidae. The quality of genotype calls and the placement of samples within a well-supported phylogeny may provide an important test for validating the fidelity and integrity of ancient samples. Finally, we constructed a phylogenomic network to accurately describe the relationships between 48 cattle breeds and facilitate inferences concerning the history of domestication and breed formation.

Genomic divergence, local adaptation, and complex demographic history may inform management of a popular sportfish species complex.

The Neosho Bass (Micropterus velox), a former subspecies of the keystone top-predator and globally popular Smallmouth Bass (M. dolomieu), is endemic and narrowly restricted to small, clear streams of the Arkansas River Basin in the Central Interior Highlands (CIH) ecoregion, USA. Previous studies have detected some morphological, genetic, and genomic differentiation between the Neosho and Smallmouth Basses; however, the extent of neutral and adaptive divergence and patterns of intraspecific diversity are poorly understood. Furthermore, lineage diversification has likely been impacted by gene flow in some Neosho populations, which may be due to a combination of natural biogeographic processes and anthropogenic introductions. We assessed: (1) lineage divergence, (2) local directional selection (adaptive divergence), and (3) demographic history among Smallmouth Bass populations in the CIH using population genomic analyses of 50,828 single-nucleotide polymorphisms (SNPs) obtained through ddRAD-seq. Neosho and Smallmouth Bass formed monophyletic clades with 100% bootstrap support. We identified two major lineages within each species. We discovered six Neosho Bass populations (two nonadmixed and four admixed) and three nonadmixed Smallmouth Bass populations. We detected 29 SNPs putatively under directional selection in the Neosho range, suggesting populations may be locally adapted. Two populations were admixed via recent asymmetric secondary contact, perhaps after anthropogenic introduction. Two other populations were likely admixed via combinations of ancient and recent processes. These species comprise independently evolving lineages, some having experienced historical and natural admixture. These results may be critical for management of Neosho Bass as a distinct species and may aid in the conservation of other species with complex biogeographic histories.

Genetic variability and viral seroconversion in an outcrossing vertebrate population.

Inverse correlations between genetic variability and parasitism are important concerns for conservation biologists. We examined correlations between neutral genetic variability and the presence of antibodies to canine distemper virus (CDV) and feline parvovirus (FPV) in a free-ranging population of raccoons. Over 3 years there was a strong relationship between age and seroprevalence rates. Most young animals were seronegative to CDV and FPV, but the oldest age class was greater than 80 per cent seropositive to both viruses. CDV-seropositive animals had greater heterozygosity and lower measures of inbreeding compared with CDV-seronegative animals. This relationship was strongest among the youngest animals and did not occur during a 1 year CDV epidemic. In contrast, FPV-seropositive animals only had significantly lower measures of inbreeding in 1 year, perhaps because FPV-associated mortality is relatively low or primarily occurs among very young individuals that were under-represented in our sampling. These results suggest that even in large outcrossing populations, animals with lower heterozygosity and higher measures of inbreeding are less likely to successfully mount an immune response when challenged by highly pathogenic parasites.

Examining the molecular basis of coat color in a nocturnal primate family (Lorisidae).

Organisms use color for camouflage, sexual signaling, or as a warning sign of danger. Primates are one of the most vibrantly colored Orders of mammals. However, the genetics underlying their coat color are poorly known, limiting our ability to study molecular aspects of its evolution. The role of the melanocortin 1 receptor (MC1R) in color evolution has been implicated in studies on rocket pocket mice (Chaetodipus intermediusi), toucans (Ramphastidae), and many domesticated animals. From these studies, we know that changes in MC1R result in a yellow/red or a brown/black morphology. Here, we investigate the evolution of MC1R in Lorisidae, a monophyletic nocturnal primate family, with some genera displaying high contrast variation in color patterns and other genera being monochromatic. Even more unique, the Lorisidae family has the only venomous primate: the slow loris (Nycticebus). Research has suggested that the contrasting coat patterns of slow lorises are aposematic signals for their venom. If so, we predict the MC1R in slow lorises will be under positive selection. In our study, we found that Lorisidae MC1R is under purifying selection (ω = 0.0912). In Lorisidae MC1R, there were a total of 75 variable nucleotides, 18 of which were nonsynonymous. Six of these nonsynonymous substitutions were found on the Perodicticus branch, which our reconstructions found to be the only member of Lorisidae that has predominantly lighter coat color; no substitutions were associated with Nycticebus. Our findings generate new insight into the genetics of pelage color and evolution among a unique group of nocturnal mammals and suggest putative underpinnings of monochromatic color evolution in the Perodicticus lineage.

Effects of diet, habitat, and phylogeny on the fecal microbiome of wild African savanna (Loxodonta africana) and forest elephants (L. cyclotis).

The gut microbiome, or the community of microorganisms inhabiting the digestive tract, is often unique to its symbiont and, in many animal taxa, is highly influenced by host phylogeny and diet. In this study, we characterized the gut microbiome of the African savanna elephant (Loxodonta africana) and the African forest elephant (Loxodonta cyclotis), sister taxa separated by 2.6-5.6 million years of independent evolution. We examined the effect of host phylogeny on microbiome composition. Additionally, we examined the influence of habitat types (forest versus savanna) and diet types (crop-raiding versus noncrop-raiding) on the microbiome within L. africana. We found 58 bacterial orders, representing 16 phyla, across all African elephant samples. The most common phyla were Firmicutes, Proteobacteria, and Bacteroidetes. The microbiome of L. africana was dominated by Firmicutes, similar to other hindgut fermenters, while the microbiome of L. cyclotis was dominated by Proteobacteria, similar to more frugivorous species. Alpha diversity did not differ across species, habitat type, or diet, but beta diversity indicated that microbial communities differed significantly among species, diet types, and habitat types. Based on predicted KEGG metabolic pathways, we also found significant differences between species, but not habitat or diet, in amino acid metabolism, energy metabolism, and metabolism of terpenoids and polyketides. Understanding the digestive capabilities of these elephant species could aid in their captive management and ultimately their conservation.

Evolution of microsatellite loci in the adaptive radiation of Hawaiian honeycreepers.

Previous studies have examined germ-line mutations to infer the processes that generate and maintain variability in microsatellite loci. Few studies, however, have examined patterns to infer processes that act on microsatellite loci over evolutionary time. Here, we examine changes in 8 dinucleotide loci across the adaptive radiation of Hawaiian honeycreepers. The loci were found to be highly variable across the radiation, and we did not detect ascertainment bias with respect to allelic diversity or allele size ranges. In examining patterns at the sequence level, we found that changes in flanking regions, repeat motifs, or repeat interruptions were often shared between closely related species and may be phylogenetically informative. Genetic distance measures based on microsatellites were strongly correlated with those based on mitochondrial DNA (mtDNA) sequences as well as with divergence time up to 3 My. Phylogenetic inferences based on microsatellite genetic distances consistently recovered 2 of the 4 honeycreeper clades observed in a tree based on mtDNA sequences but differed from the mtDNA tree in the relationships among clades. Our results confirm that microsatellite loci may be conserved over evolutionary time, making them useful in population-level studies of species that diverged from the species in which they were characterized as long as 5 Ma. Despite this, we found that their use in phylogenetic inference was limited to closely related honeycreeper species.

Post-Pleistocene differentiation in a Central Interior Highlands endemic salamander.

AIM: For many endemic species with limited dispersal capacities, the relationship between landscape changes and species distributions is still unclear. We characterized the population structure of the endemic ringed salamander (Ambystoma annulatum) across its distribution in the Central Interior Highlands (CIH) of North America, an area of high species endemism, to infer the ecological and evolutionary history of the species. METHODS: We sampled 498 individuals across the species distribution and characterized the population genetic structure using nuclear microsatellite and mitochondrial DNA (mtDNA) markers. RESULTS: Ambystoma annulatum exist in two strongly supported nuclear genetic clusters across the CIH that correspond to a northern cluster that includes the Missouri Ozark populations and a southern cluster that includes the Arkansas and Oklahoma Ozarks and the Ouachita Mountains. Our demographic models estimated that these populations diverged approximately 2,700 years ago. Pairwise estimates of genetic differentiation at microsatellite and mtDNA markers indicated limited contemporary gene flow and suggest that genetic differentiation was primarily influenced by changes in the post-Pleistocene landscape of the CIH. MAIN CONCLUSIONS: Both the geologic history and post-European settlement history of the CIH have influenced the population genetic structure of A. annulatum. The low mtDNA diversity suggests a retraction into and expansion out of refugial areas in the south-central Ozarks, during temperature fluctuations of the Pleistocene and Holocene epochs. Similarly, the estimated divergence time for the two nuclear clusters corresponds to changes in the post-Pleistocene landscape. More recently, decreased A. annulatum gene flow may be a result of increased habitat fragmentation and alteration post-European settlement.

Genetic structure along an elevational gradient in Hawaiian honeycreepers reveals contrasting evolutionary responses to avian malaria.

BACKGROUND: The Hawaiian honeycreepers (Drepanidinae) are one of the best-known examples of an adaptive radiation, but their persistence today is threatened by the introduction of exotic pathogens and their vector, the mosquito Culex quinquefasciatus. Historically, species such as the amakihi (Hemignathus virens), the apapane (Himatione sanguinea), and the iiwi (Vestiaria coccinea) were found from the coastal lowlands to the high elevation forests, but by the late 1800's they had become extremely rare in habitats below 900 m. Recently, however, populations of amakihi and apapane have been observed in low elevation habitats. We used twelve polymorphic microsatellite loci to investigate patterns of genetic structure, and to infer responses of these species to introduced avian malaria along an elevational gradient on the eastern flanks of Mauna Loa and Kilauea volcanoes on the island of Hawaii. RESULTS: Our results indicate that amakihi have genetically distinct, spatially structured populations that correspond with altitude. We detected very few apapane and no iiwi in low-elevation habitats, and genetic results reveal only minimal differentiation between populations at different altitudes in either of these species. CONCLUSION: Our results suggest that amakihi populations in low elevation habitats have not been recolonized by individuals from mid or high elevation refuges. After generations of strong selection for pathogen resistance, these populations have rebounded and amakihi have become common in regions in which they were previously rare or absent.

Spatial genetic patterns indicate mechanism and consequences of large carnivore cohabitation within development.

Patterns of human development are shifting from concentrated housing toward sprawled housing intermixed with natural land cover, and wildlife species increasingly persist in close proximity to housing, roads, and other anthropogenic features. These associations can alter population dynamics and evolutionary trajectories. Large carnivores increasingly occupy urban peripheries, yet the ecological consequences for populations established entirely within urban and exurban landscapes are largely unknown. We applied a spatial and landscape genetics approach, using noninvasively collected genetic data, to identify differences in black bear spatial genetic patterns across a rural-to-urban gradient and quantify how development affects spatial genetic processes. We quantified differences in black bear dispersal, spatial genetic structure, and migration between differing levels of development within a population primarily occupying areas with >6 houses/km2 in western Connecticut. Increased development disrupted spatial genetic structure, and we found an association between increased housing densities and longer dispersal. We also found evidence that roads limited gene flow among bears in more rural areas, yet had no effect among bears in more developed ones. These results suggest dispersal behavior is condition-dependent and indicate the potential for landscapes intermixing development and natural land cover to facilitate shifts toward increased dispersal. These changes can affect patterns of range expansion and the phenotypic and genetic composition of surrounding populations. We found evidence that subpopulations occupying more developed landscapes may be sustained by male-biased immigration, creating potentially detrimental demographic shifts.

Genetic polymorphism of Baylisascaris procyonis in host infrapopulations and component populations in the Central USA.

Baylisascaris procyonis is a nematode of significant concern to public and domestic animal health as well as wildlife management. The population genetics of B. procyonis is poorly understood. To gain insights into patterns of genetic diversity within (infrapopulation level) and among (component population level) raccoon (Procyon lotor) hosts, and specifically to assess the relative importance of indirect and direct transmission of the parasite for explaining observed population structure, we collected 69 B. procyonis from 17 wild raccoons inhabiting five counties in Missouri and Arkansas, USA. Informative regions of mitochondrial (CO1, CO2) and nuclear (28S, ITS2) genes were amplified and the distribution and genetic variability of these genes were assessed within and across raccoons. Concatenation of the CO1 and CO2 mtDNA sequences resulted in 5 unique haplotypes, with haplotype diversity 0.456 ±â€¯0.068. The most common haplotype occurred in 94% of raccoons and 72.5% of B. procyonis. Sequences for 28S rDNA revealed four unique nuclear genotypes, the most common found in 100% of raccoons and 82.6% of B. procyonis. ITS2 genotypes were assessed using fragment analysis, and there was a 1:1 correspondence between 28S and ITS-2 genotypes. Infrapopulation variation in haplotypes and genotypes was high and virtually all hosts infected with multiple sequenced nematodes also harbored multiple haplotypes and genotypes. There was a positive relationship between the size of the analyzed infrapopulation (i.e., the number of nematodes analyzed) and the number of haplotypes identified in an individual. Collectively this work emphasizes the importance of indirect transmission in the lifecycle to this parasite.

The influence of breeding phenology on the genetic structure of four pond-breeding salamanders.

Understanding metapopulation dynamics requires knowledge about local population dynamics and movement in both space and time. Most genetic metapopulation studies use one or two study species across the same landscape to infer population dynamics; however, using multiple co-occurring species allows for testing of hypotheses related to different life history strategies. We used genetic data to study dispersal, as measured by gene flow, in three ambystomatid salamanders (Ambystoma annulatum, A. maculatum, and A. opacum) and the Central Newt (Notophthalmus viridescens louisianensis) on the same landscape in Missouri, USA. While all four salamander species are forest dependent organisms that require fishless ponds to reproduce, they differ in breeding phenology and spatial distribution on the landscape. We use these differences in life history and distribution to address the following questions: (1) Are there species-level differences in the observed patterns of genetic diversity and genetic structure? and (2) Is dispersal influenced by landscape resistance? We detected two genetic clusters in A. annulatum and A. opacum on our landscape; both species breed in the fall and larvae overwinter in ponds. In contrast, no structure was evident in A. maculatum and N. v. louisianensis, species that breed during the spring. Tests for isolation by distance were significant for the three ambystomatids but not for N. v. louisianensis. Landscape resistance also contributed to genetic differentiation for all four species. Our results suggest species-level differences in dispersal ability and breeding phenology are driving observed patterns of genetic differentiation. From an evolutionary standpoint, the observed differences in dispersal distances and genetic structure between fall breeding and spring breeding species may be a result of the trade-off between larval period length and size at metamorphosis which in turn may influence the long-term viability of the metapopulation. Thus, it is important to consider life history differences among closely related and ecologically similar species when making management decisions.

Reducing bias in population and landscape genetic inferences: the effects of sampling related individuals and multiple life stages.

In population or landscape genetics studies, an unbiased sampling scheme is essential for generating accurate results, but logistics may lead to deviations from the sample design. Such deviations may come in the form of sampling multiple life stages. Presently, it is largely unknown what effect sampling different life stages can have on population or landscape genetic inference, or how mixing life stages can affect the parameters being measured. Additionally, the removal of siblings from a data set is considered best-practice, but direct comparisons of inferences made with and without siblings are limited. In this study, we sampled embryos, larvae, and adult Ambystoma maculatum from five ponds in Missouri, and analyzed them at 15 microsatellite loci. We calculated allelic richness, heterozygosity and effective population sizes for each life stage at each pond and tested for genetic differentiation (F ST and D C ) and isolation-by-distance (IBD) among ponds. We tested for differences in each of these measures between life stages, and in a pooled population of all life stages. All calculations were done with and without sibling pairs to assess the effect of sibling removal. We also assessed the effect of reducing the number of microsatellites used to make inference. No statistically significant differences were found among ponds or life stages for any of the population genetic measures, but patterns of IBD differed among life stages. There was significant IBD when using adult samples, but tests using embryos, larvae, or a combination of the three life stages were not significant. We found that increasing the ratio of larval or embryo samples in the analysis of genetic distance weakened the IBD relationship, and when using D C , the IBD was no longer significant when larvae and embryos exceeded 60% of the population sample. Further, power to detect an IBD relationship was reduced when fewer microsatellites were used in the analysis.

Nucleic acid isolation from ecological samples--animal scat and other associated materials.

Noninvasive sampling is very attractive to field biologists and has tremendous potential for studying secretive species and being a cost-effective method of increasing sample sizes in studies of large, dangerous animals. Extracting DNA from noninvasively collected samples can be challenging, and the methods have been developed mainly through modification of previously developed protocols for other sample types. We present the most commonly used methods along with modifications used by some researchers to deal with the problem of coextraction of polymerase chain reaction (PCR) inhibitors. Although it is difficult to generalize about which methods should be used on particular sample types, we discuss the success of the methods in studies to date. We close with general suggestions for dealing with potential problems associated with the analysis of DNA obtained from noninvasively collected samples.

The evolution and phylogeography of the African elephant inferred from mitochondrial DNA sequence and nuclear microsatellite markers.

Recent genetic results support the recognition of two African elephant species: Loxodonta africana, the savannah elephant, and Loxodonta cyclotis, the forest elephant. The study, however, did not include the populations of West Africa, where the taxonomic affinities of elephants have been much debated. We examined mitochondrial cytochrome b control region sequences and four microsatellite loci to investigate the genetic differences between the forest and savannah elephants of West and Central Africa. We then combined our data with published control region sequences from across Africa to examine patterns at the continental level. Our analysis reveals several deeply divergent lineages that do not correspond with the currently recognized taxonomy: (i) the forest elephants of Central Africa; the forest and savannah elephants of West Africa; and (iii) the savannah elephants of eastern, southern and Central Africa. We propose that the complex phylogeographic patterns we detect in African elephants result from repeated continental-scale climatic changes over their five-to-six million year evolutionary history. Until there is consensus on the taxonomy, we suggest that the genetic and ecological distinctness of these lineages should be an important factor in conservation management planning.