Multilocus coalescent analyses reveal the demographic history and speciation patterns of mouse lemur sister species

BackgroundDebate continues as to whether allopatric speciation or peripatric speciation through a founder effect is the predominant force driving evolution in vertebrates. The mouse lemurs of Madagascar are a system in which evolution has generated a large number of species over a relatively recent time frame. Here, we examine speciation patterns in a pair of sister species of mouse lemur, Microcebus murinus and M. griseorufus. These two species have ranges that are disparately proportioned in size, with M. murinus showing a much more extensive range that marginally overlaps that of M. griseorufus. Given that these two species are sister taxa, the asymmetric but overlapping geographic ranges are consistent with a model of peripatric speciation. To test this hypothesis, we analyze DNA sequence data from four molecular markers using coalescent methods. If the peripatric speciation model is supported, we predict substantially greater genetic diversity in M. murinus, relative to M. griseorufus. Further, we expect a larger effective population size in M. murinus and in the common ancestor of the two species than in M. griseorufus, with a concomitant decrease in gene tree/species tree incongruence in the latter and weak signs of demographic expansion in M. murinus.ResultsOur results reject a model of peripatric divergence. Coalescent effective population size estimates were similar for both extant species and larger than that estimated for their most recent common ancestor. Gene tree results show similar levels of incomplete lineage sorting within species with respect to the species tree, and locus-specific estimates of genetic diversity are concordant for both species. Multilocus demographic analyses suggest range expansions for M. murinus, with this species also experiencing more recent population declines over the past 160 thousand years.ConclusionsResults suggest that speciation occurred in allopatry from a common ancestor narrowly distributed throughout southwest Madagascar, with subsequent range expansion for M. murinus. Population decline in M. murinus is likely related to patterns of climate change in Madagascar throughout the Pleistocene, potentially exacerbated by continual anthropogenic perturbation. Genome-level data are needed to quantify the role of niche specialization and adaptation in shaping the current ranges of these species.

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title: First indications of a highland specialist among mouse lemurs (Microcebus spp.) and evidence for a new mouse lemur species from eastern Madagascar
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title: Adaptive radiation and behaviour of the Malagasy lemurs.
abstract: The systematic distribution of behavioural characters in lemurs can be analysed using the same techniques as for anatomical characters, without considering physiological mechanisms. Behaviour and structure are usually interdependent (functional morphology), so it follows that behavioural features probably evolve hand-in-hand with morphology. Behavioural and morphological characters generally exhibit the same patterns of systematic distribution, though it is not yet clear whether evolution typically operates through selection of inherited behaviour patterns, or through indirect canalization of behaviour which is dependent upon particular structures. The extant Malagasy lemurs and their recent subfossil relatives must be considered together as an integrated lemur fauna, which has undergone great reduction over the last few thousand years. 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Although most authors agree that emigration from Africa has provided the main basis for biological invasion of Madagascar, there has been some controversy about the pattern of spatial relationships between Madagascar and Africa over time. Some authors (notably Simpson (1943) and Millot (1952)) have favoured a 'stable continents' hypothesis, and this has led to a concentration of interest on the Northern continents as the seat of Primate evolution. One outcome of this has been the suggestion that lemurs and lorises are separately derived from Northern European Adapinae. New geophysical evidence indicates that the 'stable continents' hypothesis is virtually untenable, and that continental drift theory provides the only coherent explanation of terrestrial evolution. This shifts the emphasis on Primate evolution to the Southern continents (notably Africa), and it seems likely that the lemurs and lorises had a common ancestry in Africa during the early Tertiary (for which no fossil evidence is available). One further consequence of drift theory is the observation that the Mozambique Channel has probably increased in width throughout the Tertiary, and that emigration of mammals to Madagascar from Africa has become increasingly improbable. Having established that Madagascar was probably invaded by a very small number of ancestral lemur species, which subsequently underwent adaptive radiation within the island, the systematic distribution of behavioural characters among living forms is examined. Attention is given to annual and daily patterns of activity, nesting patterns, diet (and some correlated dental features), locomotion (and some skeletal features), reproduction and social behaviour. In each case, it is shown that the Mouse Lemur group (Cheirogaleinae) and the Indri group (Indriidae) are internally cohesive in their characteristic behaviour patterns. The True Lemur group (Lemurinae) exhibits a wide range of behavioural adaptation, which is paralleled by equivalent morphological diversity. Behaviourally, the Aye-aye (Daubentonia) is as distinct as it is in morphological terms. By a process of induction, it is established that the behaviour of the ancestral lemurs was probably quite similar to that now exhibited by some Cheirogaleinae (particularly Microcebus), although living species in this group exhibit a number of probable specializations away from the ancestral condition. This conclusion is not surprising, since the Cheirogaleinae are also the least specialized of the lemurs in morphological terms. However, it is significant that the same ancestral pattern can be deduced for the loris/bush-baby group. Thus, the common ancestor of the Southern Strepsirhini (lemurs + lorises) was probably a small omnivorous form feeding primarily on insects, fruit and sap. The sap would have been gathered with the 'tooth-scraper' in the lower anterior dentition. There was probably a weakly developed spatial system of social organization, with central males of a population nucleus having access to females (a small number to each male), and peripheral males living on the fringe of each population. Competition between males would have provided a basis for selective mating and migration of peripheral males between population nuclei would have ensured exogamy. Extension of Walker's (1967) exemplary study of prosimian locomotion shows that the ancestral lemur/loris probably exhibited hindlimb dominated locomotion based on a grasping function of the extremities (primarily developed in the pes). The ancestral lemur/loris was probably nest-living, giving birth to - and caring for - a small number of well-developed infants after a relatively long period of gestation. There is some evidence that this ancestral form was nocturnal in habits, and it seems likely that the ancestral species which invaded Madagascar would have had a well-developed seasonal pattern of activity. Arboreal adaptation, the attachment to a nest, the small body size, and the ability to survive an adverse period of poor food supply (e.g. on the basis of fat reserves) would have fitted the early lemurs for a period of chance emigration across the Mozambique Channel on natural rafts of vegetation. Such rafts could have been formed from trees and other vegetation torn from forests lining rivers (e.g. the River Zambesi) on the east coast of Africa. Since the common ancestor of the lemurs and lorises was not very far removed from the ancestral Primate stock, many of the characters listed above must have been to some extent developed in the earliest Primates. This provides further evidence for the hypothesis that tree-shrews, anagalids and plesiadapids are quite separately derived from the ancestral Eutherian mammal stock, and that these three groups have no specific relationship to the Order Primates.

title: Population Genetic Analysis of Myzopoda (Chiroptera: Myzopodidae) in Madagascar
abstract: The chiropteran family Myzopodidae is endemic to Madagascar and is characterized by several unique morphologies, such as sessile adhesive discs on the thumb and sole. A new species, Myzopoda schliemanni, was recently described from western Madagascar that is morphologically distinct and geographically disjunct from the eastern species, M. aurita, the only other member of this family. Geographic variation within Myzopoda has only recently been studied at the morphological level and has never been addressed at the genetic level. We used a combination of phylogenetic, coalescent, and population genetic analyses to characterize the speciation history of Myzopoda and to clarify current and former patterns of gene flow within and between Myzopoda. Mitochondrial DNA sequences were used to determine whether genetic data support the morphologically distinct species M. schliemanni, to infer the distribution of the common ancestor of extant Myzopoda, to estimate effective population sizes (Ne) and levels of migration between species, and to determine patterns of population structure within species. Phylogenetic and network analyses revealed the existence of 4 well-supported clades in Myzopoda, but could not resolve relationships among those clades. Divergent haplotypes within species may result from either recent gene flow between the 2 species or more likely from incomplete lineage sorting. Multiple coalescent-based methodologies produced concordant estimates of Ne for Myzopoda, but conflicting signals for migration between the species, probably reflecting differences in the underlying models used by the methods. We found significant genetic structure within M. aurita, but no correlation with geography. This pattern may result from recent gene flow facilitated by expansion of Ravenala stands, an important day-roost tree for Myzopoda, associated with anthropogenic deforestation and the opening up of new habitat for members of this genus.

title: Biogeographic Evolution of Madagascar's Microendemic Biota
abstract: The endemic species richness on Madagascar, relative to landmass area, is unparalleled in the world. Many organisms on the island have restricted geographical ranges. A comprehensive hypothesis explaining the evolution of this microendemism has yet to be developed. Using an analysis of watersheds in the context of Quaternary climatic shifts, we provide a new mechanistic model to explain the process of explosive speciation on the island. River catchments with sources at relatively low elevations were zones of isolation and hence led to the speciation of locally endemic taxa, whereas those at higher elevations were zones of retreat and dispersion and hence contain proportionately lower levels of microendemism. These results provide a framework for biogeographic and phylogeographic studies, as well as a basis for prioritizing conservation actions of the remaining natural forest habitats on the island.

title: Testing species-level diversification hypotheses in Madagascar: the case of microendemic Brookesia leaf chameleons.
abstract: Madagascar's flora and fauna are remarkable both for their diversity and supraspecific endemism. Moreover, many taxa contain large numbers of species with limited distributions. Several hypotheses have been proposed to explain this high level of microendemism, including 1) riverine barrier, 2) mountain refuge, and 3) watershed contraction hypotheses, the latter 2 of which center on fragmentation due to climatic shifts associated with Pliocene/Pleistocene glaciations. The Malagasy leaf chameleon genus Brookesia is a speciose group with a high proportion of microendemic taxa, thus making it an excellent candidate to test these vicariance scenarios. We used mitochondrial and nuclear sequence data to construct a Brookesia phylogeny, and temporal concordance with Pliocene/Pleistocene speciation scenarios was tested by estimating divergence dates using a relaxed-clock Bayesian method. We strongly reject a role for Pliocene/Pleistocene climatic fluctuations in species-level diversification of Brookesia. We also used simulations to test the spatial predictions of the watershed contraction model in a phylogenetic context, independent of its temporal component, and found no statistical support for this model. The riverine barrier model is likewise a qualitatively poor fit to our data, but some relationships support a more ancient mountain refuge effect. We assessed support for the 3 hypotheses in a nonphylogenetic context by examining altitude and species richness and found a significant positive correlation between these variables. This is consistent with a mountain refuge effect but does not support the watershed contraction or riverine barrier models. Finally, we find repeated higher level east-west divergence patterns 1) between the 2 sister clades comprising the Brookesia minima group and 2) within the clade of larger leaf chameleons, which shows a basal divergence between western and eastern/northern sister clades. Our results highlight the central role of phylogeny in any meaningful tests of species-level diversification theories.

title: Madagascar as a model region of species diversification.
abstract: Tropical biotas provide excellent settings in which to explore mechanisms of evolutionary diversification, yet these processes remain poorly understood. Pioneering work on biodiversity patterns and diversification processes in other tropical regions has recently been complemented by studies in Madagascar. Here we review diversity models and diversification mechanisms proposed for the fauna of this island and the perspectives for testing them. Madagascar has a diverse biota that has evolved in isolation, and is characterised by regionally pronounced and locally steep environmental gradients, common patterns of microendemism across taxa and numerous evolutionary radiations. These characteristics establish Madagascar as a promising system for the study of pattern and process in species diversification.

title: Remarkable species diversity in Malagasy mouse lemurs (primates, Microcebus).
abstract: Phylogenetic analysis of mtDNA sequence data confirms the observation that species diversity in the world's smallest living primate (genus Microcebus) has been greatly underestimated. The description of three species new to science, and the resurrection of two others from synonymy, has been justified on morphological grounds and is supported by evidence of reproductive isolation in sympatry. This taxonomic revision doubles the number of recognized mouse lemur species. The molecular data and phylogenetic analyses presented here verify the revision and add a historical framework for understanding mouse lemur species diversity. Phylogenetic analysis revises established hypotheses of ecogeographic constraint for the maintenance of species boundaries in these endemic Malagasy primates. Mouse lemur clades also show conspicuous patterns of regional endemism, thereby emphasizing the threat of local deforestation to Madagascar's unique biodiversity.

title: Mouse Lemur Phylogeography Revises a Model of Ecogeographic Constraint in Madagascar
abstract: Mouse lemurs (genus Microcebus) are small nocturnal primates that are ubiquitously distributed throughout Madagascar. Until the past decade or so, it was believed that there were only two species, one that occupied the eastern regions of Madagascar (M. rufus) and one that occupied the western regions of Madagascar (M. murinus). Intensive field studies, accompanied by genetic analysis, have revealed that the two species taxonomy vastly underestimates the actual species diversity, however, with eight species now recognized. There are numerous indicators that even the eight species taxonomy is an insufficient representation of their actual evolutionary diversity. Our chapter reviews some of the evidence both for the presently acknowledged species diversity, and clarifies the evidence for supposing that there are other species yet to be identified. Primarily, the chapter focuses on the unexpected phylogeographic patterns revealed by mitochondrial DNA analysis. These data show that mouse lemur species do not form western and eastern clades, as ecogeographic evidence might suggest. Rather, there appears to be a historical separation of species into northern and southern clades. We emphasize the point, however, that this latter pattern is based on incomplete species and geographic sampling. Only complete sampling of populations from all regions of Madagascar will reveal the true historical patterns of mouse lemur evolution.

title: The ever-increasing diversity in mouse lemurs: three new species in north and northwestern Madagascar.
abstract: Mouse lemurs (Microcebus spp.) are the world's smallest primates and endemic to Madagascar. Several recent taxonomic revisions resulted in an extraordinary increase of recognized species. What still was considered as being two species at the beginning of the 20th century is currently recognized as 12 taxa. Based on fur coloration pattern, they can be divided into grayish and reddish forms. Two major models have been proposed to explain the extensive speciation events in the Malagasy fauna. The best known suggests that major rivers and mountains combine to act as effective barriers to gene flow and thereby facilitate allopatric speciation processes. A more recent model used an analysis of watersheds in the context of Quaternary climatic shifts to explain the process of explosive speciation on the island. We tested these two models by covering the areas between all major rivers (n=8) in northwestern and northern Madagascar. Mouse lemurs were systematically caught, sampled and morphometrically characterized in 25 sites (with 2-49 individuals per site and species). A complete phylogeny was constructed on the basis of the sequences of three mitochondrial loci (in total 1296 bp). The phylogenetic data revealed a previously unknown biodiversity with three new mouse lemur species among the reddish forms, each having a very small distribution, i.e., being restricted to only one Inter-River-System (IRS). Morphometric analyses underlined their distinctiveness and a brief formal species description is provided. In contrast to the reddish forms, grayish forms have a very low species diversity coupled with broad distributions that cover more than one IRS. These differences among the species are discussed as outcome of divergent colonization scenarios. Elements of both biogeographic models are combined in a new hypothesis that aims to explain the speciation process leading to the present distribution of mouse lemurs in Madagascar.

title: Molecular Evidence of Reproductive Isolation in Sympatric Sibling Species of Mouse Lemurs
abstract: Recent morphological and molecular phylogenetic studies of mouse lemurs (Microcebus) living in the western and southern regions of Madagascar have shown that specific diversity had been considerably underestimated. In large part, this underestimate was due to the lack of sufficient specimens from given localities to assess properly the level of phenotypic variation within and between populations. The accurate delineation of specific boundaries has no doubt been confounded by the diminutive size, nocturnal habits, and subtle morphological variation characteristic of mouse lemurs, which can make field identification of individuals problematic. We illustrate the use of molecular phylogenetic analysis to reveal reproductive isolation in two sympatric mouse lemur species, Microcebus murinus and M. griseorufus. Their documentation in the Berenty Private Reserve in the extreme south of Madagascar verifies the historically-broad distribution of Microcebus griseorufus, a species recently resurrected from synonomy.

title: Hybridization between mouse lemurs in an ecological transition zone in southern Madagascar
abstract: Hybrid zones in ecotones can be useful model systems for the study of evolutionary processes that shape the distribution and discreteness of species. Such studies could be important for an improved understanding of the complex biogeography of Madagascar, which is renowned for its outstanding degree of small‐scale endemism. Certain forest remnants in central Madagascar indicate that transitional corridors across the island could have connected microendemics in different forest types in the past. Evolutionary processes in such corridors are difficult to study because most of these corridors have disappeared due to deforestation in central Madagascar. We studied a hybrid zone in one of the few remaining ecotonal corridors between dry and humid forests in Madagascar, which connects two species of mouse lemurs, Microcebus griseorufus in dry spiny forest and Microcebus murinus in humid littoral forest. We sampled 162 mouse lemurs at nine sites across this boundary. Morphometric analyses revealed intermediate morphotypes of many individuals in transitional habitat. Bayesian clustering of microsatellite genotypes and assignment tests yielded evidence for a mixed ancestry of mouse lemurs in the ecotone, where we also observed significant linkage disequilibria and heterozygote deficiency. In contrast to these observations, mitochondrial haplotypes displayed a sharply delimited boundary at the eastern edge of spiny forest, which was noncoincident with the signals from microsatellite data. Among several alternative scenarios, we propose asymmetric nuclear introgression due to male‐biased dispersal, divergent environmental selection, and an expansion of dry spiny forest in the course of aridification as a probable explanation of our observations.

title: Hybridization of mouse lemurs: different patterns under different ecological conditions
abstract: BackgroundSeveral mechanistic models aim to explain the diversification of the multitude of endemic species on Madagascar. The island's biogeographic history probably offered numerous opportunities for secondary contact and subsequent hybridization. Existing diversification models do not consider a possible role of these processes. One key question for a better understanding of their potential importance is how they are influenced by different environmental settings. Here, we characterized a contact zone between two species of mouse lemurs, Microcebus griseorufus and M. murinus, in dry spiny bush and mesic gallery forest that border each other sharply without intermediate habitats between them. We performed population genetic analyses based on mtDNA sequences and nine nuclear microsatellites and compared the results to a known hybrid zone of the same species in a nearby wide gradient from dry spiny bush over transitional forest to humid littoral forest.ResultsIn the spiny-gallery system, Microcebus griseorufus is restricted to the spiny bush; Microcebus murinus occurs in gallery forest and locally invades the dryer habitat of its congener. We found evidence for bidirectional introgressive hybridization, which is closely linked to increased spatial overlap within the spiny bush. Within 159 individuals, we observed 18 hybrids with mitochondrial haplotypes of both species. Analyses of simulated microsatellite data indicate that we identified hybrids with great accuracy and that we probably underestimated their true number. We discuss short-term climatic fluctuations as potential trigger for the dynamic of invasion and subsequent hybridization. In the gradient hybrid zone in turn, long-term aridification could have favored unidirectional nuclear introgression from Microcebus griseorufus into M. murinus in transitional forest.ConclusionsMadagascar's southeastern transitional zone harbors two very different hybrid zones of mouse lemurs in different environmental settings. This sheds light on the multitude of opportunities for the formation of hybrid zones and indicates an important influence of environmental factors on secondary contact and hybridization. Our findings suggest that hybridization could enhance the adaptability of mouse lemurs without necessarily leading to a loss of distinctiveness. They point to a potential role of hybridization in Madagascar's diversification history that requires further investigation.

title: Origins, Diversity and Relationships of Lemurs
abstract: AbstractI review new evidence on origins and adaptive radiation of Malagasy lemurs, a remarkably diverse group containing 13% of living primate species. The number of recognized lemur species has increased significantly, partly due to research revealing specific subdivisions within known populations but mainly because of discovery of new populations through fieldwork. Some species feared to be extinct have also been rediscovered. Specific numbers have increased particularly in small-bodied, cryptic genera for which continued research will surely reveal even more species.Adaptative radiation of lemurs has been essentially confined to Madagascar. The high density of lemur species on that island, associated with very small geographical ranges, has major implications both for their evolutionary divergence and for conservation. Reconstructions of phylogenetic relationships among primates have been considerably enhanced by DNA sequence data. Sufficient data are now available from both nuclear and mitochondrial sequences to examine relationships among and within the major groups of living primates. Most studies have confirmed that lemurs constitute a monophyletic sister-group of the lorisiform clade and all exclude a specific relationship between cheirogaleids and lorisiforms repeatedly inferred from morphological evidence. However, some analyses indicate that the aye-aye may have branched away before the divergence between other lemurs and lorisiforms. DNA sequence analyses have also yielded a broad consensus for relationships between Eulemur, Hapalemur, Lemur and Varecia: Varecia branched away first, while Lemur is more closely related to Hapalemur than to Eulemur.
As debate about phylogenetic relationships among lemurs and other primates seems to have been settled in favor of lemur monophyly (possibly excluding the aye-aye), only a single invasion of Madagascar is required; but it must still be explained how ancestral lemurs could have migrated there at an appropriate time. Separation between Madagascar and Africa was apparently complete by about 120 Ma, too far in the past for direct overland migration. A recent hypothesis suggested that uplifted land in the Mozambique Channel assisted colonization of Madagascar 26-45 Ma, seemingly agreeing with an estimated date of about 40 Ma for divergence of lemurs from other primates. However, mounting evidence suggests that divergence occurred significantly earlier. Because the earliest known fossil representatives of several modern orders of placental mammals (including primates) are dated no earlier than the early Tertiary, it is widely accepted that their divergence took place after the Cretaceous/Tertiary mass extinction. Yet the known fossil record can only yield minimum divergence times; if sampling is poor and/or biased there may be a considerable discrepancy between minimum and actual dates. There is, for example, virtually no known fossil record for lemurs in Madagascar and the earliest known representatives are subfossil lemurs, so in this case a direct reading of the fossil record would indicate that the lemurs first originated just a few thousand years ago! Examination of underestimation of times of origin because of poor sampling in the fossil record has confirmed previous suggestions that primates originated considerably earlier than generally believed. Several recent phylogenetic reconstructions based on DNA sequence data and using calibration dates derived from groups other than primates provide independent support for this inference. Overall, it now seems that primates originated at around 90 Ma rather than the 55 Ma indicated by direct reading of the known fossil record. Hence, colonization of Madagascar by lemurs would have taken place at about 80 Ma, double the date usually accepted, and should be interpreted in terms of contemporary continental relationships.

title: Multiple nuclear loci reveal patterns of incomplete lineage sorting and complex species history within western mouse lemurs (Microcebus).
abstract: Mouse lemurs (genus Microcebus) are nocturnal primates endemic to the island of Madagascar. Until recently, they were classified as two species, one from eastern and one from western Madagascar. Previously published analyses of morphometric and mitochondrial DNA data show strong support for the recognition of more than eight species, however. Here, we test the eight-species hypothesis with DNA data derived from four independently segregating nuclear loci. We find many areas of congruence between the mitochondrial and nuclear data, but incomplete lineage sorting and low mutation rates limit the phylogenetic resolution of the nuclear data. Even so, the nuclear loci unanimously find evidence for three deeply diverged lineages within the mouse lemur radiation: one that is congruent with the mtDNA "southern clade", another that is congruent with the mtDNA "northern clade", and one monospecific branch comprised of the species Microcebus ravelobensis. The latter result in particular emphasizes the need for careful biological study of this species.

title: Delimiting Species without Nuclear Monophyly in Madagascar's Mouse Lemurs
abstract: Speciation begins when populations become genetically separated through a substantial reduction in gene flow, and it is at this point that a genetically cohesive set of populations attain the sole property of species: the independent evolution of a population-level lineage. The comprehensive delimitation of species within biodiversity hotspots, regardless of their level of divergence, is important for understanding the factors that drive the diversification of biota and for identifying them as targets for conservation. However, delimiting recently diverged species is challenging due to insufficient time for the differential evolution of characters—including morphological differences, reproductive isolation, and gene tree monophyly—that are typically used as evidence for separately evolving lineages.  In this study, we assembled multiple lines of evidence from the analysis of mtDNA and nDNA sequence data for the delimitation of a high diversity of cryptically diverged population-level mouse lemur lineages across the island of Madagascar. Our study uses a multi-faceted approach that applies phylogenetic, population genetic, and genealogical analysis for recognizing lineage diversity and presents the most thoroughly sampled species delimitation of mouse lemur ever performed.  The resolution of a large number of geographically defined clades in the mtDNA gene tree provides strong initial evidence for recognizing a high diversity of population-level lineages in mouse lemurs. We find additional support for lineage recognition in the striking concordance between mtDNA clades and patterns of nuclear population structure. Lineages identified using these two sources of evidence also exhibit patterns of population divergence according to genealogical exclusivity estimates. Mouse lemur lineage diversity is reflected in both a geographically fine-scaled pattern of population divergence within established and geographically widespread taxa, as well as newly resolved patterns of micro-endemism revealed through expanded field sampling into previously poorly and well-sampled regions.

title: Incongruence between genetic and morphological diversity in Microcebus griseorufus of Beza Mahafaly
abstract: BackgroundThe past decade has seen a remarkable increase in the number of recognized mouse lemur species (genus Microcebus). As recently as 1994, only two species of mouse lemur were recognized according to the rules of zoological nomenclature. That number has now climbed to as many as fifteen proposed species. Indeed, increases in recognized species diversity have also characterized other nocturnal primates – galagos, sportive lemurs, and tarsiers. Presumably, the movement relates more to a previous lack of information than it does to any recent proclivity for taxonomic splitting. Due to their nocturnal habits, one can hypothesize that mouse lemurs will show only minimal variation in pelage coloration as such variation should be inconsequential for the purposes of mate and/or species recognition. Even so, current species descriptions for nocturnal strepsirrhines place a good deal of emphasis on relatively fine distinctions in pelage coloration.ResultsHere, we report results from a multi-year study of mouse lemur populations from Beza Mahafaly in southern Madagascar. On the basis of morphological and pelage variation, we initially hypothesized the presence of up to three species of mouse lemurs occurring sympatrically at this locality, one of which appeared to be undescribed. Genetic analysis reveals definitively, however, that all three color morphs belong to a single recognized species, Microcebus griseorufus. Indeed, in some cases, the three color morphs can be characterized by identical mitochondrial haplotypes.ConclusionGiven these results, we conclude that investigators should always proceed with caution when using a single data source to identify novel species. A synthetic approach that combines morphological, genetic, geographic, and ecological data is most likely to reveal the true nature of species diversity.

title: Geneious Basic: An integrated and extendable desktop software platform for the organization and analysis of sequence data
abstract: The two main functions of bioinformatics are the organization and analysis of biological data using computational resources. Geneious Basic has been designed to be an easy-to-use and flexible desktop software application framework for the organization and analysis of biological data, with a focus on molecular sequences and related data types. It integrates numerous industry-standard discovery analysis tools, with interactive visualizations to generate publication-ready images. One key contribution to researchers in the life sciences is the Geneious public application programming interface (API) that affords the ability to leverage the existing framework of the Geneious Basic software platform for virtually unlimited extension and customization. The result is an increase in the speed and quality of development of computation tools for the life sciences, due to the functionality and graphical user interface available to the developer through the public API. Geneious Basic represents an ideal platform for the bioinformatics community to leverage existing components and to integrate their own specific requirements for the discovery, analysis and visualization of biological data.  Binaries and public API freely available for download at http://www.geneious.com/basic, implemented in Java and supported on Linux, Apple OSX and MS Windows. The software is also available from the Bio-Linux package repository at http://nebc.nerc.ac.uk/news/geneiousonbl.

title: MAFFT: a novel method for rapid multiple sequence alignment based on fast Fourier transform.
abstract: A multiple sequence alignment program, MAFFT, has been developed. The CPU time is drastically reduced as compared with existing methods. MAFFT includes two novel techniques. (i) Homo logous regions are rapidly identified by the fast Fourier transform (FFT), in which an amino acid sequence is converted to a sequence composed of volume and polarity values of each amino acid residue. (ii) We propose a simplified scoring system that performs well for reducing CPU time and increasing the accuracy of alignments even for sequences having large insertions or extensions as well as distantly related sequences of similar length. Two different heuristics, the progressive method (FFT-NS-2) and the iterative refinement method (FFT-NS-i), are implemented in MAFFT. The performances of FFT-NS-2 and FFT-NS-i were compared with other methods by computer simulations and benchmark tests; the CPU time of FFT-NS-2 is drastically reduced as compared with CLUSTALW with comparable accuracy. FFT-NS-i is over 100 times faster than T-COFFEE, when the number of input sequences exceeds 60, without sacrificing the accuracy.

title: RDP3: a flexible and fast computer program for analyzing recombination
abstract: RDP3 is a new version of the RDP program for characterizing recombination events in DNA-sequence alignments. Among other novelties, this version includes four new recombination analysis methods (3SEQ, VISRD, PHYLRO and LDHAT), new tests for recombination hot-spots, a range of matrix methods for visualizing over-all patterns of recombination within datasets and recombination-aware ancestral sequence reconstruction. Complementary to a high degree of analysis flow automation, RDP3 also has a highly interactive and detailed graphical user interface that enables more focused hands-on cross-checking of results with a wide variety of newly implemented phylogenetic tree construction and matrix-based recombination signal visualization methods. The new RDP3 can accommodate large datasets and is capable of analyzing alignments ranging in size from 1000×10 kilobase sequences to 20×2 megabase sequences within 48 h on a desktop PC. Availability: RDP3 is available for free from its web site http://darwin.uvigo.es/rdp/rdp.html Contact: darrenpatrickmartin@gmail.com Supplementary information: The RDP3 program manual contains detailed descriptions of the various methods it implements and a step-by-step guide describing how best to use these.

title: Possible emergence of new geminiviruses by frequent recombination.
abstract: Although exchange of genetic information by recombination plays a role in the evolution of viruses, the extent to which it generates diversity is not clear. We analyzed genomes of geminiviruses for recombination using a new statistical procedure developed to detect gene conversions. Geminiviruses (family, Geminiviridae) are a group of plant viruses characterized by a genome of circular single-stranded DNA (approximately 2700 nucleotides in length) encapsidated in twinned quasi-isometric particles. Complete nucleotide sequences of geminiviruses were aligned, and recombination events were detected by searching pairs of viruses for sequences that are significantly more similar than expected based on random distribution of polymorphic sites. The analyses revealed that recombination is very frequent and occurs between species and within and across genera. Tests identified 420 statistically significant recombinant fragments distributed across the genome. The results suggest that recombination is a significant contributor to geminivirus evolution. The high rate of recombination may be contributing to the recent emergence of new geminivirus diseases.

title: Analyzing the mosaic structure of genes
abstract: SummarySome genes in prokaryotes consist of a mosaic of regions derived from different ancestors by horizontal gene transfer. A method is described for demonstrating the statistical significance of such mosaic structure and for locating the crossover points separating different regions.

title: RDP: detection of recombination amongst aligned sequences
abstract: Recombination Detection Program (RDP) is a program that applies a pairwise scanning approach to the detection of recombination amongst a group of aligned DNA sequences. The software runs under Windows95 and combines highly automated screening of large numbers of sequences with a highly interactive interface for examining the results of the analyses.

title: DnaSP v5: A software for comprehensive analysis of DNA polymorphism data
abstract: DnaSP is a software package for a comprehensive analysis of DNA polymorphism data. Version 5 implements a number of new features and analytical methods allowing extensive DNA polymorphism analyses on large datasets. Among other features, the newly implemented methods allow for: (i) analyses on multiple data files; (ii) haplotype phasing; (iii) analyses on insertion/deletion polymorphism data; (iv) visualizing sliding window results integrated with available genome annotations in the UCSC browser.  Freely available to academic users from: http://www.ub.edu/dnasp

title: Development and application of a phylogenomic toolkit: resolving the evolutionary history of Madagascar's lemurs.
abstract: Lemurs and the other strepsirrhine primates are of great interest to the primate genomics community due to their phylogenetic placement as the sister lineage to all other primates. Previous attempts to resolve the phylogeny of lemurs employed limited mitochondrial or small nuclear data sets, with many relationships poorly supported or entirely unresolved. We used genomic resources to develop 11 novel markers from nine chromosomes, representing approximately 9 kb of nuclear sequence data. In combination with previously published nuclear and mitochondrial loci, this yields a data set of more than 16 kb and adds approximately 275 kb of DNA sequence to current databases. Our phylogenetic analyses confirm hypotheses of lemuriform monophyly and provide robust resolution of the phylogenetic relationships among the five lemuriform families. We verify that the genus Daubentonia is the sister lineage to all other lemurs. The Cheirogaleidae and Lepilemuridae are sister taxa and together form the sister lineage to the Indriidae; this clade is the sister lineage to the Lemuridae. Divergence time estimates indicate that lemurs are an ancient group, with their initial diversification occurring around the Cretaceous-Tertiary boundary. Given the power of this data set to resolve branches in a notoriously problematic area of primate phylogeny, we anticipate that our phylogenomic toolkit will be of value to other studies of primate phylogeny and diversification. Moreover, the methods applied will be broadly applicable to other taxonomic groups where phylogenetic relationships have been notoriously difficult to resolve.

title: Delimiting species without monophyletic gene trees.
abstract: Genetic data are frequently used to delimit species, where species status is determined on the basis of an exclusivity criterium, such as reciprocal monophyly. Not only are there numerous empirical examples of incongruence between the boundaries inferred from such data compared to other sources like morphology -- especially with recently derived species, but population genetic theory also clearly shows that an inevitable bias in species status results because genetic thresholds do not explicitly take into account how the timing of speciation influences patterns of genetic differentiation. This study represents a fundamental shift in how genetic data might be used to delimit species. Rather than equating gene trees with a species tree or basing species status on some genetic threshold, the relationship between the gene trees and the species history is modeled probabilistically. Here we show that the same theory that is used to calculate the probability of reciprocal monophyly can also be used to delimit species despite widespread incomplete lineage sorting. The results from a preliminary simulation study suggest that very recently derived species can be accurately identified long before the requisite time for reciprocal monophyly to be achieved following speciation. The study also indicates the importance of sampling, both with regards to loci and individuals. Withstanding a thorough investigation into the conditions under which the coalescent-based approach will be effective, namely how the timing of divergence relative to the effective population size of species affects accurate species delimitation, the results are nevertheless consistent with other recent studies (aimed at inferring species relationships), showing that despite the lack of monophyletic gene trees, a signal of species divergence persists and can be extracted. Using an explicit model-based approach also avoids two primary problems with species delimitation that result when genetic thresholds are applied with genetic data -- the inherent biases in species detection arising from when and how speciation occurred, and failure to take into account the high stochastic variance of genetic processes. Both the utility and sensitivities of the coalescent-based approach outlined here are discussed; most notably, a model-based approach is essential for determining whether incompletely sorted gene lineages are (or are not) consistent with separate species lineages, and such inferences require accurate model parameterization (i.e., a range of realistic effective population sizes relative to potential times of divergence for the purported species). It is the goal (and motivation of this study) that genetic data might be used effectively as a source of complementation to other sources of data for diagnosing species, as opposed to the exclusion of other evidence for species delimitation, which will require an explicit consideration of the effects of the temporal dynamic of lineage splitting on genetic data.

title: Bayesian inference of population size history from multiple loci
abstract: BackgroundEffective population size (Ne) is related to genetic variability and is a basic parameter in many models of population genetics. A number of methods for inferring current and past population sizes from genetic data have been developed since JFC Kingman introduced the n-coalescent in 1982. Here we present the Extended Bayesian Skyline Plot, a non-parametric Bayesian Markov chain Monte Carlo algorithm that extends a previous coalescent-based method in several ways, including the ability to analyze multiple loci.ResultsThrough extensive simulations we show the accuracy and limitations of inferring population size as a function of the amount of data, including recovering information about evolutionary bottlenecks. We also analyzed two real data sets to demonstrate the behavior of the new method; a single gene Hepatitis C virus data set sampled from Egypt and a 10 locus Drosophila ananassae data set representing 16 different populations.ConclusionThe results demonstrate the essential role of multiple loci in recovering population size dynamics. Multi-locus data from a small number of individuals can precisely recover past bottlenecks in population size which can not be characterized by analysis of a single locus. We also demonstrate that sequence data quality is important because even moderate levels of sequencing errors result in a considerable decrease in estimation accuracy for realistic levels of population genetic variability.

title: Bayesian Phylogenetics with BEAUti and the BEAST 1.7
abstract: Computational evolutionary biology, statistical phylogenetics and coalescent-based population genetics are becoming increasingly central to the analysis and understanding of molecular sequence data. We present the Bayesian Evolutionary Analysis by Sampling Trees (BEAST) software package version 1.7, which implements a family of Markov chain Monte Carlo (MCMC) algorithms for Bayesian phylogenetic inference, divergence time dating, coalescent analysis, phylogeography and related molecular evolutionary analyses. This package includes an enhanced graphical user interface program called Bayesian Evolutionary Analysis Utility (BEAUti) that enables access to advanced models for molecular sequence and phenotypic trait evolution that were previously available to developers only. The package also provides new tools for visualizing and summarizing multispecies coalescent and phylogeographic analyses. BEAUti and BEAST 1.7 are open source under the GNU lesser general public license and available at http://beast-mcmc.googlecode.com and http://beast.bio.ed.ac.uk

title: jModelTest 2: more models, new heuristics and parallel computing
abstract: jModelTest 2: more models, new heuristics and parallel computing Diego Darriba, Guillermo L. Taboada, Ramón Doallo and David Posada Supplementary Table 1. New features in jModelTest 2 Supplementary Table 2. Model selection accuracy Supplementary Table 3. Mean square errors for model averaged estimates Supplementary Note 1. Hill-climbing hierarchical clustering algorithm Supplementary Note 2. Heuristic filtering Supplementary Note 3. Simulations from prior distributions Supplementary Note 4. Speed-up benchmark on real and simulated datasets

title: Divergence dates for Malagasy lemurs estimated from multiple gene loci: geological and evolutionary context
abstract: The lemurs of Madagascar are a unique radiation of primates that show an extraordinary diversity of lifestyles, morphologies and behaviours. However, very little is known about the relative antiquity of lemuriform clades due to the lack of terrestrial fossils for the Tertiary of Madagascar. Here, we employ a Bayesian method to estimate divergence dates within the lemuriform radiation using several unlinked gene loci and multiple fossil calibrations outside the lemuriform clade. Two mitochondrial genes (cytochrome oxidase II and cytochrome b), two nuclear introns (transthyretin intron 1 and von Willebrand factor gene intron 11) and one nuclear exon (interphotoreceptor retinoid binding protein, exon 1) are used in separate and combined analyses. The genes differ in taxon sampling and evolutionary characteristics but produce congruent date estimates. Credibility intervals narrow considerably in combined analyses relative to separate analyses due to the increased amount of data. We also test the relative effects of multiple vs. single calibration points, finding that, when only single calibration points are employed, divergence dates are systematically underestimated. For the mitochondrial DNA data set, we investigate the effects of sampling density within the mouse lemur radiation (genus Microcebus). When only two representative species are included, estimated dates throughout the phylogeny are more recent than with the complete‐species sample, with basal nodes less affected than recent nodes. The difference appears to be due to the manner in which priors on node ages are constructed in the two analyses. In nearly all analyses, the age of the lemuriform clade is estimated to be approximately 62–65 Ma, with initial radiation of mouse lemurs and true lemurs (genus Eulemur) occurring approximately 8–12 Ma. The antiquity of the mouse lemur radiation is surprising given the near uniform morphology among species. Moreover, the observation that mouse lemurs and true lemurs are of similar ages suggests discrepancies in rates of morphological, behavioural and physiological evolution in the two clades, particularly with regard to characteristics of sexual signalling. These differences appear to correlate with the nocturnal vs. diurnal lifestyles, respectively, of these two primate groups.

title: Bayesian Inference of Species Trees from Multilocus Data
abstract: Until recently, it has been common practice for a phylogenetic analysis to use a single gene sequence from a single individual organism as a proxy for an entire species. With technological advances, it is now becoming more common to collect data sets containing multiple gene loci and multiple individuals per species. These data sets often reveal the need to directly model intraspecies polymorphism and incomplete lineage sorting in phylogenetic estimation procedures. For a single species, coalescent theory is widely used in contemporary population genetics to model intraspecific gene trees. Here, we present a Bayesian Markov chain Monte Carlo method for the multispecies coalescent. Our method coestimates multiple gene trees embedded in a shared species tree along with the effective population size of both extant and ancestral species. The inference is made possible by multilocus data from multiple individuals per species. Using a multiindividual data set and a series of simulations of rapid species radiations, we demonstrate the efficacy of our new method. These simulations give some insight into the behavior of the method as a function of sampled individuals, sampled loci, and sequence length. Finally, we compare our new method to both an existing method (BEST 2.2) with similar goals and the supermatrix (concatenation) method. We demonstrate that both BEST and our method have much better estimation accuracy for species tree topology than concatenation, and our method outperforms BEST in divergence time and population size estimation.

title: Relaxed Phylogenetics and Dating with Confidence
abstract: In phylogenetics, the unrooted model of phylogeny and the strict molecular clock model are two extremes of a continuum. Despite their dominance in phylogenetic inference, it is evident that both are biologically unrealistic and that the real evolutionary process lies between these two extremes. Fortunately, intermediate models employing relaxed molecular clocks have been described. These models open the gate to a new field of “relaxed phylogenetics.” Here we introduce a new approach to performing relaxed phylogenetic analysis. We describe how it can be used to estimate phylogenies and divergence times in the face of uncertainty in evolutionary rates and calibration times. Our approach also provides a means for measuring the clocklikeness of datasets and comparing this measure between different genes and phylogenies. We find no significant rate autocorrelation among branches in three large datasets, suggesting that autocorrelated models are not necessarily suitable for these data. In addition, we place these datasets on the continuum of clocklikeness between a strict molecular clock and the alternative unrooted extreme. Finally, we present analyses of 102 bacterial, 106 yeast, 61 plant, 99 metazoan, and 500 primate alignments. From these we conclude that our method is phylogenetically more accurate and precise than the traditional unrooted model while adding the ability to infer a timescale to evolution.

title: RAxML-VI-HPC: maximum likelihood-based phylogenetic analyses with thousands of taxa and mixed models
abstract: RAxML-VI-HPC (randomized axelerated maximum likelihood for high performance computing) is a sequential and parallel program for inference of large phylogenies with maximum likelihood (ML). Low-level technical optimizations, a modification of the search algorithm, and the use of the GTR+CAT approximation as replacement for GTR+Gamma yield a program that is between 2.7 and 52 times faster than the previous version of RAxML. A large-scale performance comparison with GARLI, PHYML, IQPNNI and MrBayes on real data containing 1000 up to 6722 taxa shows that RAxML requires at least 5.6 times less main memory and yields better trees in similar times than the best competing program (GARLI) on datasets up to 2500 taxa. On datasets > or =4000 taxa it also runs 2-3 times faster than GARLI. RAxML has been parallelized with MPI to conduct parallel multiple bootstraps and inferences on distinct starting trees. The program has been used to compute ML trees on two of the largest alignments to date containing 25,057 (1463 bp) and 2182 (51,089 bp) taxa, respectively.


AVAILABILITY
icwww.epfl.ch/~stamatak

title: Speciation in little: the role of range and body size in the diversification of Malagasy mantellid frogs
abstract: BackgroundThe rate and mode of lineage diversification might be shaped by clade-specific traits. In Madagascar, many groups of organisms are characterized by tiny distribution ranges and small body sizes, and this high degree of microendemism and miniaturization parallels a high species diversity in some of these groups. We here investigate the geographic patterns characterizing the radiation of the frog family Mantellidae that is virtually endemic to Madagascar. We integrate a newly reconstructed near-complete species-level timetree of the Mantellidae with georeferenced distribution records and maximum male body size data to infer the influence of these life-history traits on each other and on mantellid diversification.ResultsWe reconstructed a molecular phylogeny based on nuclear and mitochondrial DNA for 257 species and candidate species of the mantellid frog radiation. Based on this phylogeny we identified 53 well-supported pairs of sister species that we used for phylogenetic comparative analyses, along with whole tree-based phylogenetic comparative methods. Sister species within the Mantellidae diverged at 0.2-14.4 million years ago and more recently diverged sister species had geographical range centroids more proximate to each other, independently of their current sympatric or allopatric occurrence. The largest number of sister species pairs had non-overlapping ranges, but several examples of young microendemic sister species occurring in full sympatry suggest the possibility of non-allopatric speciation. Range sizes of species included in the sister species comparisons increased with evolutionary age, as did range size differences between sister species, which rejects peripatric speciation. For the majority of mantellid sister species and the whole mantellid radiation, range and body sizes were associated with each other and small body sizes were linked to higher mitochondrial nucleotide substitution rates and higher clade diversity. In contrast, small range sizes were unexpectedly associated with a slow-down of mitochondrial substitution rates.ConclusionsBased on these results we define a testable hypothesis under which small body sizes result in limited dispersal capabilities and low physiological tolerances, causing smaller and more strongly fragmented ranges. This can be thought to facilitate reproductive isolation and thus favor speciation. Contrary to the expectation of the faster speciation of such microendemic phenotype species, we only found small body sizes of mantellid frogs to be linked to higher diversification and substitution rates, but not small range sizes. A joint analysis of various species-rich regional anuran radiations might provide enough species with all combinations of range and body sizes for a more conclusive test of this hypothesis.

title: Signals of recent spatial expansions in the grey mouse lemur (Microcebus murinus)
abstract: BackgroundPleistocene events have shaped the phylogeography of many taxa worldwide. Their genetic signatures in tropical species have been much less explored than in those living in temperate regions. We analysed the genetic structure of a Malagasy primate species, a mouse lemur with a wide distribution (M. murinus), in order to investigate such phylogeographic processes on a large tropical island. We also evaluated the effects of anthropogenic pressures (fragmentation/deforestation) and natural features (geographic distance, rivers) on genetic structure in order to complement our understanding of past and present processes of genetic differentiation.ResultsThe analysis of the mitochondrial D-loop sequences of 195 samples from 15 study sites (10 from a continuous forest and five from isolated forest fragments) from two adjacent Inter-River-Systems (IRSs) revealed that forest fragmentation and the river restrict gene flow, thereby leading to an increased genetic differentiation between populations beyond the effect of isolation-by-distance. Demographic simulations detected signals of two successive spatial expansions that could be preliminarily dated to the late Pleistocene and early Holocene. The haplotype network revealed geographic structure and showed deep molecular divergences within and between the IRSs that would be congruent with a two-step colonization scenario.ConclusionsThis study supports the hypothesis of a relatively recent spatial expansion of the grey mouse lemur in northwestern Madagascar, which may also explain why this taxon, in contrast to its congeners, has not yet undergone allopatric speciation in the studied area and possibly across its presently wide range.

title: Latitude drives diversification in Madagascar's endemic dry forest rodent Eliurus myoxinus (subfamily Nesomyinae)
abstract: Numerous hypotheses have been proposed for the historical processes governing the rich endemism of Madagas- car's biodiversity. The 'watershed model' suggests that drier climates in the recent geological past have resulted in the contraction of forests around major watersheds, thereby defining areas of endemism. We test whether this hypothesis explains phylogeographical patterns in a dry forest-dependent rodent, Eliurus myoxinus, an endemic species widely distributed through western Madagascar. We sequenced the mitochondrial cytochrome b locus and nuclear introns of the β-fibrinogen and the growth hormone receptor genes for E. myoxinus. Using a parametric bootstrapping approach, we tested whether the mitochondrial gene tree data fit expectations of local differentiation given the watershed model. We additionally estimated population differentiation and historical demographic parameters, and reconstructed the spatial history of E. myoxinus to highlight spatial and temporal patterns of differentiation. The data do not support the watershed model as a clear explanation for the genetic patterns of diversity within extant E. myoxinus populations. We find striking patterns of latitudinal genetic structure within western Madagascar, and indicate possible roles for environmental and ecological gradients along this axis in generating phylogeographical diversity.

title: A chronology for late prehistoric Madagascar.
abstract: A database has been assembled with 278 age determinations for Madagascar.  14C dated include pretreated sediments and plant macrofossils from cores and excavations throughout the island, and bones, teeth, or eggshells of most of the extinct megafaunal taxa, including the giant lemurs, hippopotami, and ratites. Additional measurements come from uranium-series dates on speleothems and thermoluminescence dating of pottery. Changes documented include late Pleistocene climatic events and, in the late Holocene, the apparently human-caused transformation of the environment. Multiple lines of evidence point to the earliest human presence at ca. 2300 14C yr BP (350 cal yr BC). A decline in megafauna, inferred from a drastic decrease in spores of the coprophilous fungus Sporormiella spp. in sediments at 1720+/-40 14C yr BP (230-410 cal yr AD), is followed by large increases in charcoal particles in sediment cores, beginning in the SW part of the island, and spreading to other coasts and the interior over the next millennium. The record of human occupation is initially sparse, but shows large human populations throughout the island by the beginning of the Second Millennium AD. Dating of the "subfossil" megafauna, including pygmy hippos, elephant birds, giant tortoises, and large lemurs, demonstrates that most if not all the extinct taxa were still present on the island when humans arrived. Many taxa overlapped chronologically with humans for a millennium or more. The extinct lemurs Hadropithecus stenognathus, Pachylemur insignis, Mesopropithecus pithecoides, and Daubentonia robusta, and the elephant birds Aepyornis spp. and Mullerornis spp., were still present near the end of the First Millennium AD. Palaeopropithecus ingens, Megaladapis edwardsi, and Archaeolemur sp. (cf. edwardsi) may have survived until the middle of the Second Millennium A.D. One specimen of Hippopotamus of unknown provenance dates to the period of European colonization.

title: Rates, Patterns, and Processes of Landscape Transformation and Extinction in Madagascar
abstract: Inferring cause and effect from the fossil record is not a wholly satisfying enterprise. The evidence is stale. Many useful details are missing, perhaps never to be found. Sequential events may be collapsed together, inverted, or mixed with evidence from other times. Few relevant parameters can be measured directly. Paleoecologists must forge ahead despite these obstacles, however, as extinction and environmental change are subjects too important to ignore.

title: Distribution and Morphological Variation of Microcebus spp. Along an Environmental Gradient in Southeastern Madagascar
abstract: The lemurs of Madagascar are known for their extraordinary levels of speciation. However, the mechanisms and environmental conditions that led to this diversity remain obscure. We used 3 species of Microcebus (M. griseorufus, M. murinus, M. rufus) occurring along an environmental gradient as a model to investigate 1) how the different species are distributed in relation to variation in environmental conditions and ecotones; 2) whether or not the morphology of a given species varies in relation to environmental conditions; and 3) whether or not there is evidence for morphological character displacement to reduce congeneric competition in sympatry vs. allopatry. The 3 species of Microcebus show clear associations with specific habitat types. Distributions overlap at ecotones. Nevertheless, the ecotone between dry spiny and gallery forest represents a species boundary between Microcebus griseorufus and M. murinus while the ecotone between dry spiny forest and evergreen humid forest represents the species boundary between M. murinus and M. rufus. Different ambient conditions are not reflected in changes in body measurements of Microcebus murinus living in different vegetation formations. There is no indication for character displacement in sympatry vs. allopatry. Thus, differences in body mass or other morphological characteristics do not contribute to species separation between Microcebus griseorufus and M. murinus. The results confirm the importance of ecotones as species boundaries as a condition postulated for the radiation of lemur and other species on Madagascar. They also demonstrate different habitat affinities of seemingly very similar lemur species and thus illustrate our very limited understanding of the actual selection pressures, adaptations of lemurs to their environments, and their possible response to interspecific competition.

title: Habitat Separation of Sympatric Microcebus spp. in the Dry Spiny Forest of South-Eastern Madagascar
abstract: We investigated whether or not habitat structure contributes to the separation of two sister species of lemurs and their hybrids. For this, we studied Microcebusmurinus and M. griseorufus along a continuous vegetation gradient where populations of the two species occur in sympatry or in allopatry. In allopatry, the two species are generalists without any sign of microhabitat selectivity. In sympatry, both species differed significantly and discriminated against certain habitat structures: M. murinus was found in microhabitats with larger trees than average while M. griseorufus utilized microhabitats with smaller trees. Hybrids between the two species did not show any significant discrimination for or against microhabitat structure and did not differ in their habitat utilization from either parent species. Both species can go into torpor and hibernation. M. griseorufus is seen more frequently during the cool dry season than M. murinus. We assume that M. murinus goes into extended torpor or hibernation more frequently than M. griseorufus. We interpret the different occurrence of large-sized trees in microhabitats of M. murinus as a prerequisite for M. murinus to be able to spend extended periods of time in tree holes that are isolated and allow hibernation at reduced temperature levels.

title: Patterns of species change in anthropogenically disturbed forests of Madagascar
abstract: Five main conclusions arise from this review of the responses of species to anthropogenic disturbance in Madagascar: First, species’ reactions to anthropogenic disturbance are generally negative, but remain poorly known. Our knowledge is patchy among and within higher taxonomic groups; we are still largely gathering case studies. Second, taxonomic groups vary considerably in which proximate factors are most important. Third, several groups show differing responses within different ecoregions. Whether these differences are consistent across groups requires further testing. Fourth, related species often have divergent reactions to disturbance, even within lower taxonomic groupings (families or genera). Thus, we cannot rely on phylogenetic relatedness or even ecological similarity to infer similarity in responses. Finally, disturbance typically reduces species diversity (especially of native and/or endemic species), but also causes species turnover, typically with forest specialists replaced by grassland generalists, and endemics replaced by non-endemics (including invasives). Given these knowledge gaps, we stress the urgency of applied studies that assess species’ ecology, behaviour and health across disturbance gradients, including purely anthropogenic landscapes. Remaining natural vegetation and protected areas will be unable to preserve Madagascar’s biodiversity under the impact of climatic change; we must understand responses of plants and animals to disturbance in order to create buffer zones and corridors combining secondary, degraded and natural habitats.