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Hooper’s deer mouse, Peromyscus hooperi, is the sole member of the Peromyscus hooperi species group. This species is endemic to México where it is restricted to the grassland transition zone in the states of Coahuila, Zacatecas, and San Luis Potosí. Previous studies using mitochondrial and nuclear genes (Cytb, Adh1-I2, Fgb-I7 and Rbp3) did not resolve the phylogenetic relationships of this relatively poorly known species. It was hypothesized that P. hooperi is sister to P. crinitus, and these two taxa are related to P. melanotis, P. polionotus, P. maniculatus, P. keeni, P. leucopus, P. gossypinus, P. eremicus, P. californicus, and Osgoodomys banderanus. Based on morphological characters, karyotypes, and allozymes, P. hooperi does not align with either subgenera Haplomylomys or Peromyscus. However, its unique characteristics (e. g., phallus, karyotype) have been recognized, and therefore it has been retained as its own species group. To better resolve the phylogenetic placement of P. hooperi, we performed target-enrichment and high-throughput sequencing and obtained several thousand nuclear ultraconserved elements and a complete mitogenome from a specimen collected in 1896 by Nelson and Goldman in Coahuila, México. We compared these data with 21 other species of neotomines using genome-wide data. Contrary to previous studies, we found high nodal support for the placement of P. hooperi as sister to a clade that includes Podomys floridanus, Neotomodon alstoni, Habromys simulatus, H. ixtlani, Peromyscus mexicanus, P. megalops, P. melanophrys, P. perfulvus, P. aztecus, P. attwateri, P. pectoralis, and P. boylii. We dated a Pliocene divergence of P. hooperi from its sister group at approximately 3.98 mya, and after the split of P. crinitus at ca. 4.31 mya from other peromyscines. We demonstrated that genome-wide data improve the phylogenetic signal, independently of taxon sampling, for a phylogenetically problematic species such as P. hooperi. We recommend that future genomic studies expand taxon sampling, including members of the subgenus Haplomylomys, to confirm the phylogenetic relationships of P. hooperi and the genetic status of its populations.

While museum voucher specimens continue to be the standard for species identifications, biodiversity data are increasingly represented by photographic records from camera traps and amateur naturalists. Some species are easily recognized in these pictures, others are impossible to distinguish. Here we quantify the extent to which 335 terrestrial nonvolant North American mammals can be identified in typical photographs, with and without considering species range maps. We evaluated all pairwise comparisons of species and judged, based on professional opinion, whether they are visually distinguishable in typical pictures from camera traps or the iNaturalist crowdsourced platform on a 4-point scale: (1) always, (2) usually, (3) rarely, or (4) never. Most (96.5%) of the 55,944 pairwise comparisons were ranked as always or usually distinguishable in a photograph, leaving exactly 2,000 pairs of species that can rarely or never be distinguished from typical pictures, primarily within clades such as shrews and small-bodied rodents. Accounting for a species geographic range eliminates many problematic comparisons, such that the average number of difficult or impossible-to-distinguish species pairs from any location was 7.3 when considering all species, or 0.37 when considering only those typically surveyed with camera traps. The greatest diversity of difficult-to-distinguish species was in Arizona and New Mexico, with 57 difficult pairs of species, suggesting the problem scales with overall species diversity. Our results show which species are most readily differentiated by photographic data and which taxa should be identified only to higher taxonomic levels (e.g., genus). Our results are relevant to ecologists, as well as those using artificial intelligence to identify species in photographs, but also serve as a reminder that continued study of mammals through museum vouchers is critical since it is the only way to accurately identify many smaller species, provides a wealth of data unattainable from photographs, and constrains photographic records via accurate range maps. Ongoing specimen voucher collection, in addition to photographs, will become even more important as species ranges change, and photographic evidence alone will not be sufficient to document these dynamics for many species.

The San Bernardino flying squirrel (Glaucomys oregonensis californicus) is thought to be the southernmost population of the Humboldt’s flying squirrel (G. oregonensis) and is restricted to the San Bernardino and San Jacinto Mountains in California. However, recent surveys indicate they have been extirpated from the latter locality. In this study, we characterized the intraspecific genetic structure and diversity of G. oregonensis. We amplified and sequenced at least 300 bp of the mitochondrial cytochrome-b gene and 11 nuclear microsatellites to provide baseline genetic data for this species and subspecies, investigate G. oregonensis genetic structure with a focus on the subspecies distributed in California, and finally evaluate the genetic diversity within G. o. californicus. Population and phylogenetic analyses were performed on a total of 147 samples (tissue, hair, and museum specimen) using the Illumina high-throughput sequencing (HTS) platform, thereby bioinformatically coding microsatellite alleles using established pipelines. Our results indicate reduced genetic diversity in G. o. californicus and highlight genetic distinctiveness of the San Jacinto population. In addition, the differences in genetic diversity between the mitochondrial and nuclear markers suggest sex-biased dispersal and historic separation of the species in multiple refugia followed by more recent gene flow among subpopulations in northern California. This study contributes valuable information toward understanding the genetic variation within G. oregonensis, provides information for future conservation decisions for G. o. californicus, and has novel implications for HTS genotyping-by-sequencing.

Historical DNA obtained from voucher specimens housed in natural history museums worldwide have allowed the study of elusive, rare or even extinct species that in many cases are solely represented by museum holdings. This has resulted in the increase of taxonomic representation of many taxa, has led to the discovery of new species, and has yielded stunning novel insights into the evolutionary history of cryptic or even undescribed species. Peromyscus mekisturus, is a critically endangered cricetid rodent endemic to Mexico and is only known from two museum specimens collected in 1898 and 1947. Intensive field work efforts to attempt to determine if viable populations still exist have failed, suggesting that this species is extinct or is nearing extinction. In addition, a recent study using mitogenomes demonstrated that P. mekisturus forms a well-supported clade outside the genus Peromyscus and hypothesized that this taxon is the sister group of the genus Reithrodontomys. Here, we used target enrichment and high-throughput sequencing of several thousand nuclear ultraconserved elements and mitogenomes to reconstruct dated phylogenies to test the previous phylogenetic hypothesis. We analyzed the holotype and the only other known specimen of P. mekisturus and museum samples from other peromyscine rodents to test the phylogenetic position of the species. Our results confirm that the only two specimens known to science of P. mekisturus belong to the same species and support the hypothesis that this species belongs to an undescribed genus of cricetid rodents that is sister to the genus Reithrodontomys. We dated the origin of P. mekisturus together with other speciation events in peromyscines during the late Pliocene – early Pleistocene and related these events with the Pleistocene climatic cycles. In light of our results, we recommend a taxonomic re-evaluation of this enigmatic species to properly recognize its taxonomic status as a new genus. We also acknowledge the relevance of generating genomic data from type specimens and highlight the need and importance of continuing to build the scientific heritage of the collections to study and better understand past, present, and future biodiversity.

The use of museum specimens held in natural history repositories for population and conservation genetic research is increasing in tandem with the use of massively parallel sequencing technologies. Short Tandem Repeats (STRs), or microsatellite loci, are commonly used genetic markers in wildlife and population genetic studies. However, they traditionally suffered from a host of issues including length homoplasy, high costs, low throughput, and difficulties in reproducibility across laboratories. Massively parallel sequencing technologies can address these problems, but the incorporation of museum specimen derived DNA suffers from significant fragmentation and exogenous DNA contamination. Combatting these issues requires extra measures of stringency in the lab and during data analysis, yet there have not been any high-throughput sequencing studies evaluating microsatellite allelic dropout from museum specimen extracted DNA. In this study, we evaluate genotyping errors derived from mammalian museum skin DNA extracts for previously characterized microsatellites across PCR replicates utilizing high-throughput sequencing. We found it useful to classify samples based on DNA concentration, which determined the rate by which genotypes were accurately recovered. Longer microsatellites performed worse in all museum specimens. Allelic dropout rates across loci were dependent on sample quantity, with high concentration museum specimens performing as well and recovering quality metrics nearly as high as the frozen tissue sample. Based on our results, we provide a set of best practices for quality assurance and incorporation of reliable genotypes from museum specimens.

The extraction of nucleic acids is one of the most routine procedures used in molecular biology laboratories, yet kit performance may influence the downstream processing of samples, particularly for samples which are degraded, and in low concentrations. Here we tested several commercial kits for specific use on commonly sampled mammalian museum specimens to evaluate the yield, size distribution, and endogenous content. Samples were weighed and had approximately equal input material for each extraction. These sample types are typical of natural history repositories ranged from 53 to 130 years old. The tested protocols spanned spin-column based extractions, magnetic bead purification, phenol/chloroform isolation, and specific modifications for ancient DNA. Diverse types of mammalian specimens were tested including adherent osteological material, bone and teeth, skin, and baleen. The concentration of DNA was quantified via fluorometry, and the size distributions of extracts visualized on an Agilent TapeStation. Overall, when DNA isolation was successful, all methods had quantifiable concentrations, albeit with variation across extracts. The length distributions varied based on the extraction protocol used. Shotgun sequencing was performed to evaluate if the extraction methods influenced the amount of endogenous versus exogenous content. The DNA content was similar across extraction methods indicating no obvious biases for DNA derived from different sources. Qiagen kits and phenol/chloroform isolation outperformed the Zymo magnetic bead isolations in these types of samples. Statistical analyses revealed that extraction method only explained 5% of the observed variation, and that specimen age explained variation (29%) more effectively.

Animal specimens in natural history collections are invaluable resources in examining the historical context of pathogen dynamics in wildlife and spillovers to humans. For example, natural history specimens may reveal new associations between bat species and coronaviruses. However, RNA viruses are difficult to study in historical specimens because protocols for extracting RNA from these specimens have not been optimized. Advances have been made in our ability to recover nucleic acids from formalin-fixed paraffin-embedded samples (FFPE) commonly used in human clinical studies, yet other types of formalin preserved samples have received less attention. Here, we optimize the recovery of RNA from formalin-fixed ethanol-preserved museum specimens in order to improve the usability of these specimens in surveys for zoonotic diseases. We provide RNA quality and quantity measures for replicate tissues subsamples of 22 bat specimens from five bat genera (Rhinolophus, Hipposideros, Megareops, Cynopterus, and Nyctalus) collected in China and Myanmar from 1886 to 2003. As tissues from a single bat specimen were preserved in a variety of ways, including formalin-fixed (8 bats), ethanol-preserved and frozen (13 bats), and flash frozen (2 bats), we were able to compare RNA quality and yield across different preservation methods. RNA extracted from historical museum specimens is highly fragmented, but usable for short-read sequencing and targeted amplification. Incubation of formalin-fixed samples with Proteinase-K following thorough homogenization improves RNA yield. This optimized protocol extends the types of data that can be derived from existing museum specimens and facilitates future examinations of host and pathogen RNA from specimens.

Eutherian dentition has been the focus of a great deal of studies in the areas of evolution, development, and genomics. The development of molar teeth is regulated by an antero-to-posterior cascade mechanism of activators and inhibitors molecules, where the relative sizes of the second (M2) and third (M3) molars are dependent of the inhibitory influence of the first molar (M1). Higher activator/inhibitor ratios will result in higher M2/M1 or M3/M1. Pax9 has been shown to play a key role in tooth development. We have previously shown that a G-quadruplex in the first intron of Pax9 can modulate the splicing efficiency. Using a sliding window approach with we analyzed the association of the folding energy (Mfe) of the Pax9 first intron with the relative molar sizes in 42 mammalian species, representing 9 orders. The Mfe of two regions located in the first intron of Pax9 were shown to be significantly associated with the M2/M1 and M3/M1 areas and mesiodistal lengths. The first region is located at the intron beginning and can fold into a stable G4 structure, whereas the second is downstream the G4 and 265 bp from intron start. Across species, the first intron of Pax9 varied in G-quadruplex structural stability. The correlations were further increased when the Mfe of the two sequences were added. Our results indicate that this region has a role in the evolution of the mammalian dental pattern by influencing the relative size of the molars.

The most comprehensive phylogenomic reconstruction to date was generated on all nominal taxa within the lemur genus Propithecus. Over 200 wild-caught individuals were included in this study to evaluate the intra and interspecific relationships across this genus. Ultraconserved Elements (UCEs) resulted in well-supported phylogenomic trees. Complete mitochondrial genomes (CMGs) largely agreed with the UCEs, except where a mitochondrial introgression was detected between one clade of the diademed sifaka (Propithecus diadema) and the Milne-Edwards sifaka (P. edwardsi). Additionally, the crowned (P. coronatus) and Von der Decken’s (P. deckeni) sifakas belonged to a single admixed lineage from UCEs. Further sampling across these two species is warranted to determine if our sampling represents a hybrid zone. P. diadema recovered two well-supported clades, which were dated and estimated as being ancient as the split between the Perrier’s (P. perrierii) and silky (P. candidus) sifakas. The reconstructed demographic history of the two clades also varied over time. We then modeled the modern ecological niches of the two cryptic P. diadema clades and found that they were significantly diverged (p < 0.01). These ecological differences result in a very limited zone of geographic overlap for the P. diadema clades (

The hyperdiverse shrew genus Crocidura is one of few small mammal genera distributed across Sundaland and all of its boundaries. This represents a rare opportunity to study the geological history of this region through the evolutionary history of these shrews. We generate a phylogeny of all recognized species of Sundaland Crocidura and show that most speciation events took place during the Pleistocene, prior to the inundation of the Sunda Shelf around 400 000 years ago. We find east–west differentiation within two separate lineages on Borneo, and that the current taxonomy of its two endemic species does not reflect evolutionary history, but ecophenotypic variation of plastic traits related to elevation. Sulawesi shrews are monophyletic, with a single notable exception: the black-footed shrew (C. nigripes). We show that the black-footed shrew diverged from its relatives on Borneo recently, suggesting a human-assisted breach of Wallace’s line. Overall, the number of Crocidura species, especially on Borneo, probably remains an underestimate.

This article is protected by copyright. All rights reserved. With the accelerating pace of global change, it is imperative that we obtain rapid inventories of the status and distribution of wildlife for ecological inferences and conservation planning. To address this challenge, we launched the SNAPSHOT USA project, a collaborative survey of terrestrial wildlife populations using camera traps across the United States. For our first annual survey, we compiled data across all 50 states during a 14-week period (17 August - 24 November of 2019). We sampled wildlife at 1509 camera trap sites from 110 camera trap arrays covering 12 different ecoregions across four development zones. This effort resulted in 166,036 unique detections of 83 species of mammals and 17 species of birds. All images were processed through the Smithsonian's eMammal camera trap data repository and included an expert review phase to ensure taxonomic accuracy of data, resulting in each picture being reviewed at least twice. The results represent a timely and standardized camera trap survey of the USA. All of the 2019 survey data are made available herein. We are currently repeating surveys in fall 2020, opening up the opportunity to other institutions and cooperators to expand coverage of all the urban-wild gradients and ecophysiographic regions of the country. Future data will be available as the database is updated at eMammal.si.edu/snapshot-usa, as well as future data paper submissions. These data will be useful for local and macroecological research including the examination of community assembly, effects of environmental and anthropogenic landscape variables, effects of fragmentation and extinction debt dynamics, as well as species-specific population dynamics and conservation action plans. There are no copyright restrictions; please cite this paper when using the data for publication. Publisher PDF

A surprising amount of hidden phylogenetic diversity exists in the small to medium size, drab colored squirrels of the genus Sundasciurus. This genus is endemic to Sundaland and the Philippines, where it is widespread. An earlier revision of this genus found that the high elevation ‘populations’ of the widespread, lowland slender squirrel (S. tenuis) were different species. Previous phylogenies based on mitochondrial cytochrome b sequences also suggested that the widespread, lowland Low’s squirrel (S. lowii) and the narrow endemic Fraternal squirrel (S. fraterculus) are not reciprocally monophyletic. Additionally, deep divergences have been identified between lineages within Low’s squirrel that date to the early Pliocene. Here we focus on evaluating the relationships and differences within and between populations of these two nominal species using whole mitochondrial genome sequences, nuclear intron sequences, and morphology. We reassess the taxonomy of this group, revalidate the species status of Robinson’s squirrel (Sundasciurus robinsoni Bonhote, 1903) support the species level recognition of the Natuna squirrel (Sundasciurus natunensis Thomas, 1895) and identify three other lineages that require further study. We estimate times of divergence and integrate geologic history to find that most of the divergences are pre-Pleistocene, and thus predate the Pleistocene flooding of Sundaland. Biogeographic, and ecological factors may have played a more important role than climatic factors in generating these patterns. While divergence in allopatry seems to be the main process driving speciation in lowland Sundaland squirrels (Sundasciurus), ecomorphological and behavioral adaptations in this clade suggest an important role of niche divergence., AH was supported by an Ernst Mayr Travel grant and AMNH Collection Study Grant during museum data collection. This research received support from the SYNTHESYS Project financed by European Community Research Infrastructure Action under the FP7 “Capacities” Program. AH was also supported by a Spanish Ministry of Economy contract CGL2014-58793-P. The Spanish Ministry of Science and Innovation grant CGL2017-86068-P to JL supported this study. MH was supported by discretionary research funding from Humboldt State University.

The use of museum specimens held in natural history repositories for population and conservation genetic research is increasing in tandem with the use of next generation sequencing technologies. Short Tandem Repeats (STRs), or microsatellite loci, are commonly used genetic markers in population and conservation genetic studies. However, they traditionally suffered from a host of issues: fragment size homology, high costs, and low throughput as a result of capillary electrophoresis genotyping and difficulty in reproducibility across laboratories. Next generation sequencing technologies can address these problems, but the incorporation of DNA derived from museum specimens suffers from significant fragmentation and contamination with exogenous DNA. Combatting these issues requires extra measures of stringency in the lab and during data analysis, yet there have not been any studies evaluating microsatellite allelic dropout from museum specimen extracted DNA. In this study, we explore a high throughput sequencing method to evaluate the amount of variation found within museum specimen DNA extracts for previously characterized microsatellites across PCR replicates. We found it useful to classify samples based on quality after replicated PCRs, which determined the rate by which genotypes were accurately recovered. We also found that longer microsatellites performed worse in all museum specimens, so when designing a study invoking museum specimens, short markers (under 250 bp) should be preferentially selected. Allelic dropout rates across loci were dependent on sample quality. The high quality museum specimens performed as well, and recovered nearly as high quality metrics as our tissue sample. Mitochondrial DNA sequences were not predictive of nuclear DNA presence, as all samples recovered cytochrome b fragments yet many lacked microsatellite genotypes, particularly in samples deemed low quality. Based on our results, we have provided a set of best practices for screening, quality assurance, and incorporation of reliable genotypes from museum specimens.

Janzen's influential "mountain passes are higher in the tropics" hypothesis predicts restricted gene flow and genetic isolation among populations spanning elevational gradients in the tropics. Few studies have tested this prediction, and studies that focus on population genetic structure in Southeast Asia are particularly underrepresented in the literature. Here, we test the hypothesis that mountain treeshrews (Tupaia montana) exhibit limited dispersal across their broad elevational range which spans ~2,300 m on two peaks in Kinabalu National Park (KNP) in Borneo: Mt Tambuyukon (MT) and Mt Kinabalu (MK). We sampled 83 individuals across elevations on both peaks and performed population genomics analyses on mitogenomes and single nucleotide polymorphisms from 4,106 ultraconserved element loci. We detected weak genetic structure and infer gene flow both across elevations and between peaks. We found higher genetic differentiation on MT than MK despite its lower elevation and associated environmental variation. This implies that, contrary to our hypothesis, genetic structure in this system is not primarily shaped by elevation. We propose that this pattern may instead be the result of historical processes and limited upslope gene flow on MT. Importantly, our results serve as a foundational estimate of genetic diversity and population structure from which to track potential future effects of climate change on mountain treeshrews in KNP, an important conservation stronghold for the mountain treeshrew and other montane species.

Previous studies found that aye-ayes (Daubentonia madagascariensis) consume a variety of non-invertebrate resources, supporting the hypothesis that aye-ayes are too large-bodied to focus feeding efforts on insects. However, these conclusions were based on introduced populations, with little known about aye-aye behaviour and ecology in its natural habitat. This study investigates activity budgets, diet, and home range/territories of 2 male aye-ayes in the Kianjavato Classified Forest, a disturbed forest in south-eastern Madagascar, from October 2013 to October 2014. We used radiotelemetry and focal-animal sampling methods to collect behavioural data. We recorded GPS coordinates of the focal animal every 20 min, calculating the home range size for each individual. Results showed that male aye-ayes spent most of their time feeding and travelling. Their feeding time primarily focussed on larvae and adult insects from various substrates, and to a lesser extent Canarium spp. seeds. Home range sizes of the 2 males varied between individuals, overlapped greatly, and appeared to be related to the location of females. Our findings indicate that aye-aye behaviours are strongly indicative of insect-centric feeding ecology.

Ectoparasites frequently vector pathogens from often unknown pathogen reservoirs to both human and animal populations. Simultaneous identification of the ectoparasite species, the wildlife host that provided their most recent blood meal(s), and their pathogen load would greatly facilitate the understanding of the complex transmission dynamics of vector-borne diseases. Currently, these identifications are principally performed using multiple polymerase chain reaction (PCR) assays. We developed an assay (EctoBaits) based on in-solution capture paired with high-throughput sequencing to simultaneously identify ectoparasites, host blood meals and pathogens. We validated our in-solution capture results using double-blind PCR assays, morphology and collection data. The EctoBaits assay effectively and efficiently identifies ectoparasites, blood meals, and pathogens in a single capture experiment, allowing for high-resolution taxonomic identification while preserving the DNA sample for future analyses.

Background The greater bamboo lemur (Prolemur simus) is a member of the Family Lemuridae that is unique in their dependency on bamboo as a primary food source. This Critically Endangered species lives in small forest patches in eastern Madagascar, occupying a fraction of its historical range. Here we sequence the genome of the greater bamboo lemur for the first time, and provide genome resources for future studies of this species that can be applied across its distribution. Results Following whole genome sequencing of five individuals we identified over 152,000 polymorphic single nucleotide variants (SNVs), and evaluated geographic structuring across nearly 19 k SNVs. We characterized a stronger signal associated with a north-south divide than across elevations for our limited samples. We also evaluated the demographic history of this species, and infer a dramatic population crash. This species had the largest effective population size (estimated between ~ 900,000 to one million individuals) between approximately 60,000–90,000 years before present (ybp), during a time in which global climate change affected terrestrial mammals worldwide. We also note the single sample from the northern portion of the extant range had the largest effective population size around 35,000 ybp. Conclusions From our whole genome sequencing we recovered an average genomic heterozygosity of 0.0037%, comparable to other lemurs. Our demographic history reconstructions recovered a probable climate-related decline (60–90,000 ybp), followed by a second population decrease following human colonization, which has reduced the species to a census size of approximately 1000 individuals. The historical distribution was likely a vast portion of Madagascar, minimally estimated at 44,259 km2, while the contemporary distribution is only ~ 1700 km2. The decline in effective population size of 89–99.9% corresponded to a vast range retraction. Conservation management of this species is crucial to retain genetic diversity across the remaining isolated populations. Electronic supplementary material The online version of this article (10.1186/s12864-018-4841-4) contains supplementary material, which is available to authorized users.

Here, we present a set of RNA-based probes for whole mitochondrial genome in-solution enrichment, targeting a diversity of mammalian mitogenomes. This probes set was designed from seven mammalian orders and tested to determine the utility for enriching degraded DNA. We generated 63 mitogenomes representing five orders and 22 genera of mammals that yielded varying coverage ranging from 0 to >5400X. Based on a threshold of 70% mitogenome recovery and at least 10× average coverage, 32 individuals or 51% of samples were considered successful. The estimated sequence divergence of samples from the probe sequences used to construct the array ranged up to nearly 20%. Sample type was more predictive of mitogenome recovery than sample age. The proportion of reads from each individual in multiplexed enrichments was highly skewed, with each pool having one sample that yielded a majority of the reads. Recovery across each mitochondrial gene varied with most samples exhibiting regions with gaps or ambiguous sites. We estimated the ability of the probes to capture mitogenomes from a diversity of mammalian taxa not included here by performing a clustering analysis of published sequences for 100 taxa representing most mammalian orders. Our study demonstrates that a general array can be cost and time effective when there is a need to screen a modest number of individuals from a variety of taxa. We also address the practical concerns for using such a tool, with regard to pooling samples, generating high quality mitogenomes and detail a pipeline to remove chimeric molecules.

Biodiversity across elevational gradients generally follows patterns, the evolutionary origins of which are debated. We trapped small non-volant mammals across an elevational gradient on Mount (Mt.) Kinabalu (4,101 m) and Mt. Tambuyukon (2,579 m), two neighboring mountains in Borneo, Malaysia. We also included visual records and camera trap data from Mt. Tambuyukon. On Mt. Tambuyukon we trapped a total of 299 individuals from 23 species in 6,187 trap nights (4.8% success rate). For Mt. Kinabalu we trapped a total 213 animals from 19 species, in 2,044 trap nights, a 10.4% success rate. We documented the highest diversity in the low elevations for both mountains, unlike previous less complete surveys which supported a mid-elevation diversity bulge on Mt. Kinabalu. Species richness decreased gradually towards the highlands to a more even community with different species (high turnover), less rich but with the highest levels of endemism. These patterns suggest that an interplay of topography and climatic history of the region were drivers of the diversity gradient, in addition to standing climatic and spatial hypothesis.

Mountains offer replicated units with large biotic and abiotic gradients in a reduced spatial scale. This transforms them into well-suited scenarios to evaluate biogeographic theories. Mountain biogeography is a hot topic of research and many theories have been proposed to describe the changes in biodiversity with elevation. Geometric constraints, which predict the highest diversity to occur in mid-elevations, have been a focal part of this discussion. Despite this, there is no general theory to explain these patterns, probably because of the interaction among different predictors with the local effects of historical factors. We characterize the diversity of small non-volant mammals across the elevational gradient on Mount (Mt.) Kinabalu (4,095 m) and Mt. Tambuyukon (2,579 m), two neighboring mountains in Borneo, Malaysia. We documented a decrease in species richness with elevation which deviates from expectations of the geometric constraints and suggests that spatial factors (e.g., larger diversity in larger areas) are important. The lowland small mammal community was replaced in higher elevations (from above ~1,900 m) with montane communities consisting mainly of high elevation Borneo endemics. The positive correlation we find between elevation and endemism is concordant with a hypothesis that predicts higher endemism with topographical isolation. This supports lineage history and geographic history could be important drivers of species diversity in this region.

[Aim] The genetics of organisms currently isolated in refugia has received little attention compared to post-glacial expansions. We study the population history and connectivity of a rat endemic to montane habitat in Borneo to better understand the history and potential of populations in interglacial mountain refugia., [Location] Sabah, Borneo, Malaysia., [Methods] We performed a field survey of the summit rat (Rattus baluensis) on two mountains, Mt. Kinabalu and Mt. Tambuyukon, its entire known distribution. We sequenced mitogenomes and 27 introns (19 of which were polymorphic) in 49 individuals from both populations. We analysed their current genetic structure and diversity, and inferred their demographic history with approximate Bayesian computation., [Results] Summit rats were tightly associated with mountain mossy forest and scrubland above 2,000 m, facilitating the prediction of their past and future distributions. The genetic analysis supports a Holocene fragmentation of a larger population into smaller ones that are now isolated in interglacial refugia on mountaintops. These findings are consistent with climatic reconstructions and the retreat of upland forest to higher elevations after the Last Glacial Maximum (LGM), ~21 kya., [Main conclusions] The two isolated populations of summit rats formed through the upland shift of their habitat after the LGM. The current trend of global warming will likely lead to diminishing suitable upland habitat and result in the extinction of the population on Mt. Tambuyukon. The population on Mt. Kinabalu, the higher peak, could persist at higher elevations, highlighting the singular value of high tropical mountains as reservoirs of biodiversity during past and ongoing climate change., Logistical support was provided by Laboratorio de Ecología Molecular, Estación Biológica de Doñana, CSIC (LEM‐EBD). Digital elevation models for maps was obtained from the U.S. The Spanish Ministry of Science and Innovation grants CGL2010‐21524 and CGL2014‐58793‐P supported this work. MCS is supported by the Spanish Ministry of Science and Innovation Predoctoral Fellowship BES‐2011‐049186, and part of his fieldwork was also funded by EEBB‐I‐12‐05317.

The taxonomy of American deer has been established almost entirely on the basis of morphological data and without the use of explicit phylogenetic methods; hence, phylogenetic analyses including data for all of the currently recognized species, even if based on a single gene, might improve current understanding of their taxonomy. We tested the monophyly of the morphology-defined genera and species of New World deer (Odocoileini) with phylogenetic analyses of mitochondrial DNA sequences. This is the first such test conducted using extensive geographic and taxonomic sampling. Our results do not support the monophyly of Mazama, Odocoileus, Pudu, M. americana, M. nemorivaga, Od. hemionus, and Od. virginianus. Mazama contains species that belong to other genera. We found a novel sister-taxon relationship between “Mazama” pandora and a clade formed by Od. hemionus columbianus and Od. h. sitkensis, and transfer pandora to Odocoileus. The clade formed by Od. h. columbianus and Od. h. sitkensis may represent a valid species, whereas the remaining subspecies of Od. hemionus appear closer to Od. virginianus. Pudu (Pudu) puda was not found sister to Pudu (Pudella) mephistophiles. If confirmed, this result will prompt the recognition of the monotypic Pudella as a distinct genus. We provide evidence for the existence of an undescribed species now confused with Mazama americana, and identify other instances of cryptic, taxonomically unrecognized species-level diversity among populations here regarded as Mazama temama, “Mazama” nemorivaga, and Hippocamelus antisensis. Noteworthy records that substantially extend the known distributions of M. temama and “M.” gouazoubira are provided, and we unveil a surprising ambiguity regarding the distribution of “M.” nemorivaga, as it is described in the literature. The study of deer of the tribe Odocoileini has been hampered by the paucity of information regarding voucher specimens and the provenance of sequences deposited in GenBank. We pinpoint priorities for future systematic research on the tribe Odocoileini.

Some primate populations include both trichromatic and dichromatic (red–green colour blind) individuals due to allelic variation at the X-linked opsin locus. This polymorphic trichromacy is well described in day-active New World monkeys. Less is known about colour vision in Malagasy lemurs, but, unlike New World monkeys, only some day-active lemurs are polymorphic, while others are dichromatic. The evolutionary pressures underlying these differences in lemurs are unknown, but aspects of species ecology, including variation in activity pattern, are hypothesized to play a role. Limited data on X-linked opsin variation in lemurs make such hypotheses difficult to evaluate. We provide the first detailed examination of X-linked opsin variation across a lemur clade (Indriidae). We sequenced the X-linked opsin in the most strictly diurnal and largest extant lemur,Indri indri, and nine species of smaller, generally diurnal indriids (Propithecus). Although nocturnalAvahi(sister taxon toPropithecus) lacks a polymorphism, at least eight species of diurnal indriids have two or more X-linked opsin alleles. Four rainforest-living taxa—I. indriand the three largestPropithecusspecies—have alleles not previously documented in lemurs. Moreover, we identified at least three opsin alleles inIndriwith peak spectral sensitivities similar to some New World monkeys.

Background The African wild dog (Lycaon pictus) is an endangered African canid threatened by severe habitat fragmentation, human-wildlife conflict, and infectious disease. A highly specialized carnivore, it is distinguished by its social structure, dental morphology, absence of dewclaws, and colorful pelage. Results We sequenced the genomes of two individuals from populations representing two distinct ecological histories (Laikipia County, Kenya and KwaZulu-Natal Province, South Africa). We reconstructed population demographic histories for the two individuals and scanned the genomes for evidence of selection. Conclusions We show that the African wild dog has undergone at least two effective population size reductions in the last 1,000,000 years. We found evidence of Lycaon individual-specific regions of low diversity, suggestive of inbreeding or population-specific selection. Further research is needed to clarify whether these population reductions and low diversity regions are characteristic of the species as a whole. We documented positive selection on the Lycaon mitochondrial genome. Finally, we identified several candidate genes (ASIP, MITF, MLPH, PMEL) that may play a role in the characteristic Lycaon pelage. Electronic supplementary material The online version of this article (doi:10.1186/s12864-016-3368-9) contains supplementary material, which is available to authorized users.

The family Lepilemuridae includes 26 species of sportive lemurs, most of which were recently described. The cryptic morphological differences confounded taxonomy until recent molecular studies; however, some species’ boundaries remain uncertain. To better understand the genus Lepilemur, we analyzed 35 complete mitochondrial genomes representing all recognized 26 sportive lemur taxa and estimated divergence dates. With our dataset we recovered 25 reciprocally monophyletic lineages, as well as an admixed clade containing Lepilemur mittermeieri and Lepilemur dorsalis. Using modern distribution data, an ancestral area reconstruction and an ecological vicariance analysis were performed to trace the history of diversification and to test biogeographic hypotheses. We estimated the initial split between the eastern and western Lepilemur clades to have occurred in the Miocene. Divergence of most species occurred from the Pliocene to the Pleistocene. The biogeographic patterns recovered in this study were better addressed with a combinatorial approach including climate, watersheds, and rivers. Generally, current climate and watershed hypotheses performed better for western and eastern clades, while speciation of northern clades was not adequately supported using the ecological factors incorporated in this study. Thus, multiple mechanisms likely contributed to the speciation and distribution patterns in Lepilemur.

The plain long-nosed squirrels, genus Dremomys, are high elevation species in East and Southeast Asia. Here we present a complete molecular phylogeny for the genus based on nuclear and mitochondrial DNA sequences. Concatenated mitochondrial and nuclear gene trees were constructed to determine the tree topology, and date the tree. All speciation events within the plain-long nosed squirrels (genus Dremomys) were ancient (dated to the Pliocene or Miocene), and averaged older than many speciation events in the related Sunda squirrels, genus Sundasciurus. Within the plain long-nosed squirrels, the most recent interspecific split occurred 2.9 million years ago, older than some splits within Sunda squirrels, which dated to the Pleistocene. Our results demonstrate that the plain long-nosed squirrels are not monophyletic. The single species with a distinct distribution, the Bornean mountain ground squirrel (Dremomys everetti), which is endemic to the high mountains of Borneo, is nested within the Sunda squirrels with high support. This species diverged from its sister taxa in the Sunda squirrels 6.62 million years ago, and other plain long-nosed squirrels over 11 million years ago. Our analyses of morphological traits in these related genera support the re-classification of the Bornean mountain ground squirrel, Dremomys everetti, to the genus Sundasciurus, which changes its name to Sundasciurus everetti. Past inclusion in the plain long-nosed squirrels (Dremomys) reflects convergent evolution between these high elevation species., Funding was provided by the Spanish Government (CGL2010-21524).

Background: The Indonesian island of Sulawesi has a complex geological history. It is composed of several landmasses that have arrived at a near modern configuration only in the past few million years. It is the largest island in the biodiversity hotspot of Wallacea—an area demarcated by the biogeographic breaks between Wallace’s and Lydekker’s lines. The mammal fauna of Sulawesi is transitional between Asian and Australian faunas. Sulawesi’s three genera of squirrels, all endemic (subfamily Nannosciurinae: Hyosciurus, Rubrisciurus and Prosciurillus), are of Asian origin and have evolved a variety of phenotypes that allow a range of ecological niche specializations. Here we present a molecular phylogeny of this radiation using data from museum specimens. High throughput sequencing technology was used to generate whole mitochondrial genomes and a panel of nuclear ultraconserved elements providing a large genome-wide dataset for inferring phylogenetic relationships. Results: Our analysis confirmed monophyly of the Sulawesi taxa with deep divergences between the three endemic genera, which predate the amalgamation of the current island of Sulawesi. This suggests lineages may have evolved in allopatry after crossing Wallace’s line. Nuclear and mitochondrial analyses were largely congruent and well supported, except for the placement of Prosciurillus murinus. Mitochondrial analysis revealed paraphyly for Prosciurillus, with P. murinus between or outside of Hyosciurus and Rubrisciurus, separate from other species of Prosciurillus. A deep but monophyletic history for the four included species of Prosciurillus was recovered with the nuclear data. Conclusions: The divergence of the Sulawesi squirrels from their closest relatives dated to ~9.7–12.5 million years ago (MYA), pushing back the age estimate of this ancient adaptive radiation prior to the formation of the current conformation of Sulawesi. Generic level diversification took place around 9.7 MYA, opening the possibility that the genera represent allopatric lineages that evolved in isolation in an ancient proto-Sulawesian archipelago. We propose that incongruence between phylogenies based on nuclear and mitochondrial sequences may have resulted from biogeographic discordance, when two allopatric lineages come into secondary contact, with complete replacement of the mitochondria in one species.

Time calibration derived from the fossil record is essential for molecular phylogenetic and evolutionary studies. Fossil mice and rats, discovered in the Siwalik Group of Pakistan, have served as one of the best-known fossil calibration points in molecular phylogenic studies. Although these fossils have been widely used as the 12 Ma date for the Mus/Rattus split or a more basal split, conclusive paleontological evidence for the nodal assignments has been absent. This study analyzes newly recognized characters that demonstrate lineage separation in the fossil record of Siwalik murines and examines the most reasonable nodal placement of the diverging lineages in a molecular phylogenetic tree by ancestral state reconstruction. Our specimen-based approach strongly indicates that Siwalik murines of the Karnimata clade are fossil members of the Arvicanthini-Otomyini-Millardini clade, which excludes Rattus and its relatives. Combining the new interpretation with the widely accepted hypothesis that the Progonomys clade includes Mus, the lineage separation event in the Siwalik fossil record represents the Mus/Arvicanthis split. Our test analysis on Bayesian age estimates shows that this new calibration point provides more accurate estimates of murine divergence than previous applications. Thus, we define this fossil calibration point and refine two other fossil-based points for molecular dating.

We studied the taxonomy and biogeography of Mazama bricenii, a brocket deer classified as Vulnerable by the IUCN, drawing on qualitative and quantitative morphology and sequences of the mitochondrial cytochrome-b gene. We used Ecological Niche Modeling (ENM) to evaluate the hypothesis that M. bricenii of the Venezuelan Cordillera de Merida (CM) might have become isolated from populations of its putative sister species, Mazama rufina, in the Colombian Cordillera Oriental (CO). This hypothesis assumes that warm, dry climatic conditions in the Tachira Depression were unsuitable for the species. Our analyses did not reveal morphological differences between specimens geographically attributable to M. bricenii and M. rufina, and phylogenetic analyses of molecular data recovered M. bricenii nested within the diversity of M. rufina. These results indicate that M. bricenii should be regarded as a junior synonym of M. rufina. ENM analyses revealed the existence of suitable climatic conditions for M. rufina in the Tachira Depression during the last glacial maximum and even at present, suggesting that gene flow between populations in the CO and CM may have occurred until at least the beginning of the current interglacial period and may continue today. Because this pattern might characterize other mammals currently considered endemic to the CM, we examined which of these species match two criteria that we propose herein to estimate if they can be regarded as endemic to the CM with confidence: (1) that morphological or molecular evidence exists indicating that the putative endemic taxon is distinctive from congeneric populations in the CO; and (2) that the putative endemic taxon is restricted to either cloud forest or paramo, or both. Only Aepeomys reigi, Cryptotis meridensis, and Nasuella meridensis matched both criteria; hence, additional research is necessary to assess the true taxonomic status and distribution of the remaining species thought to be CM endemics.

Aim: Pleistocene environmental fluctuations had well-characterized impacts on the patterns of within-species divergences and diversity in temperate habitats. Here we examine the impact the Pleistocene had on widely distributed forest vertebrates in a tropical system where the distribution of the habitat was affected by those fluctuations. Location: Sundaland, tropical Southeast Asia. Methods: We conducted a comparative phylogeographical analysis of 28 non-migratory, forest-dependent vertebrates, for which we constructed rooted, intraspecifc phylogenies based on mitochondrial DNA sequences of individuals from at least the three major landmasses in the area (Borneo, Sumatra and the Malay Peninsula) and compared them to hypothetical phylogenies based on independent geological data and climate models regarding connections and relationships between the major landmasses of Sundaland. Java was included where possible. We dated the phylogenies to determine whether patterns of differentiation were concordant across species. Results: In most species, populations on the Malay Peninsula and Sumatra were most closely related, and sister to those from Borneo. The dates of these divergences, however, varied extensively between species. Borneo harbours multiple deeply divergent lineages of many species compared to the diversity within those species. Javan populations of several birds were most divergent relative to those from the rest of the Sunda Shelf. Main conclusions: These results suggest a dynamic history, including recurrent population extinctions and replacements and a strong priority effect for local populations. The close relationship between populations in Sumatra and the Malay Peninsula supports the existence of forest on the exposed shelf during the Pleistocene at many different times, and suggests that proximity was more important than the presence of palaeorivers for dispersal of forest taxa between landmasses.

Island endemics are typically differentiated from their mainland progenitors in behavior, morphology, and genetics, often resulting from long-term evolutionary change. To examine mechanisms for the origins of island endemism, we present a phylogeographic analysis of whole mitochondrial genomes from the endangered island fox (Urocyon littoralis), endemic to California’s Channel Islands, and mainland gray foxes (U. cinereoargenteus). Previous genetic studies suggested that foxes first appeared on the islands >16,000 years ago, before human arrival (~13,000 cal BP), while archaeological and paleontological data supported a colonization >7000 cal BP. Our results are consistent with initial fox colonization of the northern islands probably by rafting or human introduction ~9200–7100 years ago, followed quickly by human translocation of foxes from the northern to southern Channel Islands. Mitogenomes indicate that island foxes are monophyletic and most closely related to gray foxes from northern California that likely experienced a Holocene climate-induced range shift. Our data document rapid morphological evolution of island foxes (in ~2000 years or less). Despite evidence for bottlenecks, island foxes have generated and maintained multiple mitochondrial haplotypes. This study highlights the intertwined evolutionary history of island foxes and humans, and illustrates a new approach for investigating the evolutionary histories of other island endemics.