BACKGROUND
"wave after wave of immigration came in from asia, recruiting the fauna at each successive stage, but leaving little opportunity for new types to arise here.
even those genera which seem to be of native origin, might prove to be immigrants, if all their history were known ."
the diversification of the mustelidae  is a striking example of adaptive radiation, the evolution of ecological and phenotypic diversity from a common ancestor  <cit> .
mustelids exhibit both locomotor and dietary diversity, with taxa that are fossorial , semi-arboreal  and semi-aquatic , and diets that vary from specialization on rodents  to piscivory .
ecomorphological diversity in the family is thus correspondingly high, reflecting the adaptation of different species of mustelids to different habits and habitats.
as with other cases of adaptive radiation  <cit> , resolving relationships within the mustelidae, especially among genera, has been challenging.
many taxonomic schemes proposed for mustelids within the last century were based on morphology and classified genera into various numbers of subfamilies  <cit>  whose boundaries were largely determined by ecomorphological similarity.
at one extreme, pocock  <cit>  divided extant mustelids into  <dig>  subfamilies based on descriptive analyses of external characters .
at the other end, the system proposed by simpson  <cit>  cast mustelids into five subfamilies based on both phylogeny and 'similarity in adaptiveness' of constituent genera: lutrinae , melinae , mellivorinae , mephitinae  and mustelinae .
while such a scheme may indeed reflect the true phylogeny in some instances, morphological similarity does not necessarily imply phylogenetic affinity, as has been well demonstrated in certain groups that exemplify adaptive radiation .
moreover, such criteria can lead to recognition of polyphyletic grades rather than monophyletic groups.
nonetheless, simpson's subfamilial classification of the mustelidae has been followed for many years, although the latest classification provisionally places all mustelids  into two subfamilies, lutrinae and mustelinae  <cit> .
this latter scheme was proposed in recognition of demonstrated paraphyly of the traditional subfamilies by more recent morphological-based phylogenetic studies  <cit> .
during the last decade, dna sequence-based studies have begun to challenge the validity of the five-subfamily scheme and even monophyly of the family itself.
studies using both mitochondrial and nuclear sequences have consistently demonstrated that skunks and stink badgers  descend from a common ancestor and together form a lineage  that diverged prior to the split between mustelidae and procyonidae  <cit> .
these studies and those more focused on mustelids  <cit>  have also suggested that:  the lutrinae is monophyletic; but that  both melinae and mustelinae are polyphyletic.
however, phylogenetic relationships within the family remain uncertain or unknown because taxon sampling, especially for genera, has been incomplete.
further, adequate character sampling is also an important issue because adaptive radiations are often composed of lineages that have rapidly diverged  <cit> .
short stem lengths of topologies revealed in recent studies, albeit with incomplete taxon sampling  <cit>  suggests that deeper lineages of mustelids may have radiated within a short span of time.
adequate character sampling is therefore critical in achieving an accurate, well-resolved and robust phylogenetic hypothesis.
fossil evidence indicates that the biogeographic history of mustelids is characterized by numerous intercontinental dispersals, primarily originating from eurasia where the earliest fossil remains of the family  are found  <cit> .
for instance, a large majority of mustelid diversity in north  america is considered to have originated from lineages that repeatedly dispersed from eurasia via the bering land bridge  <cit> .
the earliest immigrants to north america arrived in the early miocene and included a paraphyletic group of stem taxa referred to as 'paleomustelids', whose affinities to crown group mustelids  remains ambiguous, as well as genera that belonged to the extinct subfamily leptarctinae  <cit> .
the first appearance of various mustelid genera in north america is used to help define the beginning of biostratigraphic boundaries of north american land mammal ages  during the neogene  <cit> .
for example, the first appearance of extinct genera trogonictis and sminthosinis and extant genera lutra and mustela help mark the latest hemphilian nalma )  <cit> .
there is still uncertainty, however, about the exact number of intercontinental dispersal events underlying the biogeographic distribution of extant genera and species and, therefore, how much of mustelid continental diversity is a result of in situ versus ex situ evolution.
further, the sequence of dispersal events has been difficult to decipher, stemming from incompleteness of the fossil record.
clarifying the biogeographic history of mustelids has implications for understanding the community ecology of mustelids.
multiple species of mustelid are often found in the same community and consequently they have been the subject of important studies on character displacement and resource partitioning  <cit> .
however, many of these studies lacked a historical perspective , which has been shown to exert a strong influence on community assembly and structure  <cit> .
here, we present a nearly complete generic-level phylogeny of the mustelidae using ~ <dig>  base pairs  of mitochondrial and nuclear dna data obtained from  <dig> gene segments.
we use this phylogeny to address three objectives.
first, we compare our phylogenetic hypothesis with previous hypotheses generated with morphological or molecular data.
second, we estimate relative divergence times using new bayesian relaxed molecular clock methods to understand the temporal pattern of diversification in the family.
do mustelids exhibit one or more bursts of rapid cladogenesis characteristic of many adaptive radiations  <cit> ?
moreover, correlation of divergence times with paleoenvironmental changes can provide insight into the mechanisms that might have been responsible for bursts of diversification  <cit> .
finally, we assess the biogeographic history of the mustelidae, especially with regard to understanding dispersal history between continents of the old world  and those of the new world .
RESULTS
phylogenetic relationships
phylogenetic analyses using maximum parsimony , two different methods of maximum likelihood  and bayesian inference  all recovered the same hypothesis for intergeneric relationships, which resolves mustelidae into seven primary divisions that include four major clades and three monotypic lineages .
otters  form a clade  that is sister to a clade comprising mink and true weasels .
these clades, in turn, are sister to a clade that includes weasel-like species with aposematically colored pelage .
next, ferret-badgers  are a monotypic lineage  that is sister to these three combined clades.
the fifth major clade  comprises two subclades, one containing hog-nosed and eurasian badgers  and the other containing tayra, wolverine and martens .
finally, as the earliest divergences in the tree, the american badger  and honey badger  form successive monotypic lineages sister to all other mustelid genera.
intergeneric relationships observed are largely congruent with those recovered in recent analyses of mitochondrial and nuclear dna sequences  <cit> .
in contrast, our topology is highly incongruent with a cladistic analysis based on morphology  <cit> .
for example, bryant et al  <cit>  found that melogale was reconstructed as sister to all remaining mustelids and that eira, gulo and martes were polyphyletic.
except for the monophyly of otters, skunks , and arctonyx and meles, all other nodes in the bryant et al  <cit>  tree had low bootstrap support values .
furthermore, alternative topologies were recovered in the bryant et al study when certain characters were weighted differentially.
regarding relationships within the four major clades, otters are resolved into three primary lineages, whose relationships are congruent with previous analyses based on fewer dna sequence data  <cit> : one containing old world river otters and the sea otter , a second containing new world river otters  and a third containing the monotypic giant otter .
relationships of enhydra and hydrictis were unresolved in the bayesian consensus tree , but were resolved differently in mp and ml analyses.
with mp, enhydra and hydrictis were resolved as sister taxa, whereas with ml, they were resolved as successive sister lineages to remaining old world otters, with hydrictis forming the first basal split.
however, neither of these relationships was well supported .
two clades comprise the true weasel and mink clade , with the new world american mink  and long-tailed weasel  in one clade  and all other sampled mustela species in the second clade .
within the latter clade, m. nudipes and m. strigidorsa are sister to a clade comprising species largely distributed in temperate regions of the northern hemisphere.
despite the similar ecology, american mink  and european mink  are distantly related, as found in previous studies  <cit> .
species of true weasels and mink have been divided into five  <cit>  or nine subgenera  <cit>  based on morphological criteria.
given the taxa we have sampled, our phylogeny suggests that only one proposed subgenus constitutes a natural group, putorius, containing the steppe polecat , european polecat  and the north american black-footed ferret .
further, mustela is paraphyletic with respect to neovison and suggests that the placement of the american mink in the separate genus neovison may not be warranted despite the observed differences in karyotype and morphology between this taxon and other species of mustela <cit> .
in the third major clade, grisons of central and south america  are sister to a clade containing the marbled polecat  of asia and three african species, the libyan striped weasel , zorilla  and striped weasel .
monophyly of these genera is of interest because all exhibit an aposematically colored pelage combined with defense behaviors that include threat displays and excretion of pungent musk from enlarged anal glands  <cit> .
placement of vormela within this clade is congruent with a recent cytb study  <cit> .
interestingly, there is a north to south progression in branching order from vormela to i. striatus and p. albinucha of sub-saharan africa, which renders ictonyx paraphyletic.
this suggests that p. albinucha should be placed into the genus ictonyx kaup,  <dig>  given the priority of the latter name over poecilogale thomas,  <dig>  as for the clade containing eira, gulo and martes , the last genus is clearly paraphyletic, in agreement with other recent studies  <cit> .
two new world species, the tayra  and fisher  either comprise a clade or form successive lineages sister to a clade containing wolverine  and the remaining species of martes.
our phylogeny indicates that true martens  are monophyletic  and sister to yellow-throated marten .
within the subgenus martes, mp and ml analyses resulted in different phylogenetic placements of m. americana and m. melampus relative to the  clade.
in the mp tree, m. americana was sister to the clade ) whereas m. americana and m. melampus were joined as sister taxa in the ml tree.
relationships of m. americana, m. melampus and  were unresolved in the bayesian tree, resulting in a trichotomy .
although more data is required to establish the precise branching order of the fisher, the strong support for this taxon being paraphyletic with the remaining species of martes strongly suggests placement of the fisher in its own genus, pekania.
nodal support
concatenation of  <dig>  bp, with indels coded as missing, includes  <dig>   variable characters and  <dig>   parsimony-informative characters.
when indels are coded for information content, the concatenation is reduced to  <dig>  bp, with  <dig>   variable and  <dig>   parsimony-informative characters.
the majority of internodes in our phylogeny have >90% bootstrap support  and  <dig>  posterior probabilities , see figure  <dig> and table  <dig>  indicating that our phylogenetic hypothesis, given our data set, is robustly supported.
moreover,  <dig> internodes are supported by one or more synapomorphic indels .
three clades, however, have low support in mp, ml and/or bi analyses: nodes  <dig>   <dig> and  <dig>  internodes associated with these clades have very short branch lengths , suggesting that these splits occurred rapidly.
alternatively, speciation may have occurred relatively recently, also resulting in inadequate phylogenetic signal, as among m. eversmanni, m. putorius and m. nigripes  as well as m. americana, m. melampus and .
many nuclear segments show no differences among species that comprise these groups, suggesting that there has been insufficient time for sequence differences to accumulate at these more slowly evolving loci.
indeed, the four species of martes have been described as a superspecies complex of closely related, yet largely allopatrically distributed taxa  <cit> .
regardless of the exact cause, short branch lengths of these internodes can result in a high number of anomalous gene trees , leading to an incorrectly inferred species tree, especially when data from multiple data partitions is concatenated  <cit> .
additional analyses, including population genetic-level sampling, will be needed to confidently resolve relationships among these recently evolved species, as in reference  <cit>  for example.
even so, internodes with low support may be stable or unstable, depending on the relative stability or instability of their constituent  taxa or leaves  <cit> .
further, unstable taxa can influence nodal support in other parts of a phylogenetic tree  <cit> .
we measured leaf stability of all taxa using bootstrap trees from mp analyses and the program radcon  <cit> .
for three different measures of leaf stability, arctonyx collaris and meles meles were the least stable taxa .
we repeated mp and hill-climbing ml analyses  along with bootstrapping after excluding these two taxa, thereby reducing the data set to  <dig> taxa.
most bootstrap values in these analyses showed little or no change compared with the  <dig> taxa data set, but interestingly, support for two stem clades  increased from over 70% to more than 90%, as did average phylogenetic stability .
the lower stability of a. collaris and m. meles and their influence on bootstrap support at internodes  <dig>   <dig> and  <dig> may be caused by character conflict among different gene segments associated with these two taxa  <cit> .
however, we note that these nodes all have high or maximal posterior probabilities in bi analyses .
with regards to subfamilial classification, our phylogeny clearly indicates that the mustelinae, as both traditionally  <cit>  and recently  <cit>  conceived, is polyphyletic, as suggested by previous studies  <cit> .
our results also indicate that various genera of badgers , most of which are often placed in the melinae  <cit> , are also polyphyletic, supporting earlier conclusions based on morphology that these taxa are not closely related  <cit> .
instead, badgers with specific adaptations for fossoriality  constitute a basal grade of lineages that are best recognized as distinct subfamilies.
overall, our phylogeny  is consistent with the subfamilial classification scheme recently proposed by sato et al  <cit>  and fulton and strobeck  <cit> , which redefines the boundaries of traditional subfamilies  as well as resurrects other subfamilies such as helictidinae and mellivorinae, thereby reflecting actual phyletic lines that have been difficult to resolve based on morphology alone.
divergence times
divergence times across the mustelid phylogeny were estimated using the uncorrelated relaxed lognormal molecular clock model calibrated simultaneously by eight fossil constraints.
use of this molecular clock model provides a measure of rate heterogeneity among lineages or how well data conform or deviate from a strict molecular clock  <cit> .
for the root age and crown age prior combination of  <dig>  mya and  <dig> mya, the coefficient of variation  averaged across three independent runs was  <dig>  :  <dig> – <dig> ), which suggests that the data show rate heterogeneity among lineages .
we found greater rate heterogeneity among lineages when older root age and crown age prior combinations were used; for example, for  <dig>  mya and  <dig>  mya, averaged across three runs, σr =  <dig>  .
furthermore, a likelihood ratio test significantly rejects the ml phylogeny with an enforced molecular clock versus one without a molecular clock  thereby providing additional support that our data departs from clocklike behavior.
in general, mean posterior estimates as well as 95% hpd intervals of node ages were highly consistent among different combinations of root age and crown age prior combinations .
the root and node  <dig>  were most sensitive to alterations in priors, because they showed the largest shift in divergence times  and little overlap in 95% hpd intervals in runs using older versus younger combinations of root age/crown age priors .
nonetheless, sampling of the joint prior distribution by performing markov chain monte carlo  analyses without any data  suggested that the eight fossil calibration point priors  did not have a strong influence on their estimated posterior distributions  and thus, estimated divergence times.
how do our estimates of divergence times compare with those based on previous molecular studies of mustelids  <cit> ?
at nodes shared among the different studies, divergence times in previous studies are either a mix of younger, older and overlapping dates relative to those in our study  <cit>  or they tend to be older in general  <cit> .
for example, the estimated divergence time for the split between neovison vison and the remaining taxa of mustela was dated at  <dig> – <dig>  mya,  <dig> – <dig>  mya and 10– <dig> mya in sato et al  <cit> , marmi et al  <cit>  and hosoda et al  <cit> , respectively.
in our study, this split , which also includes m. frenata as sister to n. vison, is dated at ~ <dig>  mya .
however, earlier studies differ in several important respects from our study.
first, these studies were based on a smaller number of taxa and loci.
for example, in sato et al  <cit>  and hosoda et al  <cit> , taxon sampling largely consisted of species from martes and mustela and was based on one and two loci, respectively.
undersampling of taxa and characters can bias divergence time estimates  <cit>  and reduces the power to detect rate variation among lineages  <cit> .
second, previous studies all employed a single root age fossil constraint, treated as a hard bound, precluding an accurate estimation of error associated with this calibration point.
in contrast, we employed multiple fossil constraints simultaneously and placed soft-bound priors on these constraints to account for the uncertainty associated with the fossil record, thereby making it possible to evaluate errors associated with divergence time estimates  <cit> .
third, the previous studies each estimated a substitution rate that was assumed to be constant across their respective phylogenies.
however, our results indicate that substitution rates vary across different lineages of the mustelid phylogeny , consistent with recent evidence from different groups that rates of substitution vary across lineages, even among closely related species  <cit> .
tempo and mode of mustelid diversification
using the root age and crown age prior combination of  <dig>  mya and  <dig> mya as a reference, our dating analyses indicate that, following the initial divergence of taxidea in the early miocene , mustelids underwent two main bursts of diversification .
the first burst occurred during a ~ <dig>  million year interval from the middle to late miocene  and gave rise to most of the extant primary clades and lineages .
the second and larger burst occurred during the pliocene epoch  in which as many as  <dig> generic-level or specific-level lineages originated within a  <dig>  million year span of time .
furthermore, results of our biogeographic analyses show that the majority of cladogenetic events associated with these bursts of diversification occurred in the old world, largely in eurasia .
paleoenvironmental and biotic changes driven by changes in climate during the latter half of the neogene may have promoted the two bursts of cladogenesis within mustelids.
following the mid-miocene climatic optimum and onset of modern oceanic circulation , there is a marked cooling of the global climate near the end of the middle miocene that continues through to the holocene  <cit> .
this period of cooling coincides with formation of a permanent antarctic ice sheet in the mid to late miocene and an arctic ice sheet in the pliocene  <cit> .
in addition, several major sea-level lowstands occurred during the late miocene and pliocene, including the serravallian sea-lowering event near the beginning of the late miocene, 11- <dig> mya  <cit> .
these changes in climate and sea level increased overall terrestrial aridity and seasonality, which in turn promoted a shift from closed vegetation habitats  to more open vegetation habitats   <cit> .
by the early late miocene, plant and animal fossil evidence indicates that the eurasian continent was a mosaic of vegetation types and generally more heterogeneous in vegetation structure relative to that in the early to middle miocene  <cit> .
these changes in vegetation had a concomitant impact on faunal communities and may have fostered diversification in a variety of lineages, including mustelids, via geographic isolation, divergent selection among different habitats, and/or ecological opportunity through the creation of new niches or the reorganization of former niches.
interestingly, the initial radiation of the primary clades and lineages of mustelids coincides with a major faunal turnover in western europe  that affected many groups of mammals, including the carnivora  <cit> .
in fact, nearly half of carnivoran species that went extinct during this turnover were mustelids  <cit>  and turnover in mustelids remained high throughout the late miocene in western eurasia  <cit> .
although evidence for faunal change in other parts of eurasia is less clear, changes in habitat and extinction of earlier lineages of mustelids may have created ecological opportunities that fostered the initial burst of diversification of modern mustelids.
further cooling and drying during the pliocene, coincident with onset of high latitude glacial cycles  <cit> , caused a dramatic expansion of low-biomass vegetation, including grasslands and steppe at midlatitudes and development of taiga at high latitudes of eurasia and north america  <cit> .
coupled with these changes was diversification of prey species such as rodents  and passerine birds that exploited these new habitats, which in turn provided new niches for predators  <cit> .
part of the pliocene burst of diversification of mustelids may have been promoted by this diversification of prey species, as species in genera such martes  and especially mustela  became specialized in hunting small prey such as rodents.
such a scenario is consistent with king's  <cit>  hypothesis that evolution of small body size in mustela was partly driven by adaptation to exploit abundant resources presented by rodent diversification during the pliocene.
we also note that diversification of four of the five species within the subgenus martes , which are all closely associated with taiga forest habitat  <cit> , coincides with expansion of this type of habitat across the holarctic during the plio-pleistocene  <cit> .
this finding is consistent with fossil evidence that indicates that taxa ancestral to these living species primarily evolved in forested habitats  <cit> .
in africa, the split between ictonyx libyca and the clade containing i. striatus and poecilogale albinucha  dated at  <dig> – <dig>  mya , marginally overlaps with a major increase in african aridity and climate variability that occurred  <dig> - <dig>  mya, according to paleoclimatic and faunal evidence  <cit> .
moreover, this interval was accompanied by rapid radiation in several mammalian lineages such as bovids and hominids  <cit> .
our results suggest that divergence of i. libyca, which occurs in north africa along margins of the sahara desert, and the clade containing i. striatus and p. albinucha, which both occur south of the sahara, may have also been caused by this shift to greater aridity, especially considering that extensive desert conditions in the sahara did not occur until the late pliocene, around  <dig>  mya  <cit> .
if mustelid diversification was promoted by climatically driven paleoenvironmental changes, then synchronous patterns and tempos of diversification should also be recorded in the evolutionary histories of other, unrelated groups.
a number of recent molecular phylogenetic studies on various groups of mammals and other vertebrates employing multiple loci and relaxed molecular clock methods provide corroboration for this hypothesis.
for example, squirrels , cats , rabbits and hares , deer  and woodpeckers  each show one or more episodes of rapid diversification that are roughly contemporaneous with one or both of those observed in mustelids .
taking cats as an example, seven of the eight primary lineages of felids radiated in the early part of the late miocene  whereas genera and species that comprise the eight lineages largely radiated during the pliocene  <cit> .
we also note that divergence events from a number of independent lineages within the mustelidae are synchronous as well.
for example, using the root age and crown age prior combination of  <dig>  mya and  <dig> mya as reference, nodes  <dig>   <dig>   <dig>   <dig>   <dig> and  <dig> all codiversify around  <dig>  mya .
similarly, nodes  <dig>   <dig> and  <dig> overlap in their divergence time estimates of around  <dig>  mya .
while ecological circumstances obviously differ from group to group, such congruence in patterns and tempos of diversification supports the idea that a common cause, namely, large-scale changes in past environments, has shaped the phylogenetic histories among disparate groups of organisms as well as independent lineages of mustelids.
our results have important implications for theories about the mode and tempo of adaptive radiations.
first, the scenario of diversification across different trophic groups we have outlined above is consistent with the model of cascading radiation proposed by stanley  <cit> , in which diversification of one group in a trophic cascade  promotes diversification in a second group , which in turn promotes diversification in a third group .
this model could be tested more rigorously using dated phylogenies of multiple groups representing different trophic levels.
second, our finding that the two primary bursts of cladogenesis in the evolutionary history of extant mustelids  coincided with periods of climatically mediated environmental changes provides support for models that show that evolution may be greatly accelerated in temporally and spatially changing environments  <cit> .
during these times, new niches are created or former niches are reorganized, providing new ecological opportunities that may foster rapid speciation and thus, diversification  <cit>  .
the fact that multiple groups show contemporaneous periods of rapid cladogenesis  suggests that mainland environments undergoing environmental changes may function like newly colonized island archipelagoes in promoting diversification.
historical biogeography
the two-state and four-state biogeographical reconstructions indicate that the vast majority of the modern diversification of mustelids has occurred in the old world , specifically in eurasia .
in fact, eurasia was unambiguously reconstructed as the ancestral area for nearly every ancestral node in the four-state analysis .
these results are consistent with two other observations that suggest eurasia has been the center of mustelid diversification:  eurasia contains the majority of extant species, with  <dig> of the  <dig> known species either exclusively endemic to or having part of their distribution on this continent; and  the earliest fossils of extant lineages or those associated with the ancestors of extant lineages are often found in eurasia  <cit> .
nodes located near the base of the tree tend to be those where likelihood ratios are not significant , such as the root node and the node joining taxidea as sister to the remaining taxa of mustelids in the two-state analysis .
such results are not uncommon in likelihood or other model-based approaches to ancestral state reconstruction because the degree of uncertainty associated with reconstruction increases with time  <cit> .
nonetheless, proportional likelihood values for these nodes still favor the old world as the ancestral area in the two-state analysis .
ancestral reconstructions were robust when enhydra lutris, gulo gulo and mellivora capensis were coded for alternative states .
in contrast to the extensive in situ diversification that has taken place in eurasia, mustelid faunas of africa and the new world are largely comprised of genera or species that repeatedly colonized these regions from eurasia.
the mustelid fauna of africa contains eight species, seven of which are included in our taxon sampling .
of these seven species, five are derived from separate colonizations from eurasia  whereas two are derived from in situ speciation events ; see figure  <dig>  similarly, our reconstructions show that nine separate dispersal events from eurasia and only one in situ speciation event accounts for the diversity of mustelids that are either endemic to north america  or have part of their distribution there .
genera and species of mustelids found in south america today are largely descended from north american immigrants that arrived as part of the great american interchange following the rise of the panamanian isthmus,  <dig> - <dig>  mya  <cit> .
such a relationship is clearly indicated for the clade of new world otters in which l. canadensis is sister to l. felina + l. longicaudis, with the latter two species found in central and/or south america .
moreover, this split  is estimated to have occurred  <dig> – <dig>  mya , which overlaps well with timing of the formation of the panamanian land bridge.
the long-tailed weasel, m. frenata, ranges from north america to northern south america  <cit>  and two species of weasels  are endemic to south america.
fossil evidence clearly indicates that mustela colonized south america from the north, apparently well after the panamanian isthmus was in place  <cit> .
our results show that pteronura, galictis and eira dispersed separately into south america, with eurasia reconstructed as the continent of origin for each genus .
these results are anomalous because these genera have been allied with extinct taxa from north america, suggesting a more proximate origin for these lineages  <cit> .
for example, pteronura may be related to the extinct genus satherium from the pliocene of north america  <cit> .
however, paleontological studies suggest that the ultimate ancestry of these extinct taxa lies in eurasia  <cit> .
a combination of evidence from the fossil record and divergence times from our phylogeny indicates that the mustelid faunas of africa, north america and south america have been assembled gradually over time.
for example, fossil evidence suggests mustelids colonized the new world across beringia during different intervals when the land bridge between eurasia and north america was open.
multiple genera of mustelids entered north america during the late miocene , prior to the first opening of the bering strait  <dig> – <dig>  mya, which severed the route across beringia  <cit> .
many genera that colonized north america during the late miocene or earliest pliocene became extinct  <cit> .
nonetheless, among the genera that may have been included in this wave of dispersal were the earliest representatives of lutra  and mustela, both of which are first recorded in north america from the late miocene to early pliocene .
these taxa may have been the forerunners of modern species of lontra and mustela/neovison found in north and south america today.
the late miocene divergence time of the splits leading to lontra and neovison + m. frenata  are consistent with this possibility.
two extinct genera of american badgers, chamitataxus and pliotaxidea, are recorded from the late miocene, around  <dig>  mya and  <dig>  mya, respectively  <cit> .
pliotaxidea has been shown to be sister group to taxidea based on morphological evidence  <cit> , thereby suggesting that the lineage leading to taxidea arrived in north america before the opening of the bering strait.
meline badgers  are presently found only in the old world.
however, recent discovery of late miocene to early pliocene fossils of meline badgers at two different sites in north america  <cit>  indicates that this lineage had also immigrated into north america and was a component of the new world mustelid fauna.
following these earlier dispersal events, fossil evidence indicates that mustela erminea, m. nigripes, m. nivalis and martes americana later entered north america during the pleistocene  <cit> .
the molecular divergence time for m. nigripes , around  <dig>  mya , for example, supports a pleistocene dispersal scenario for this species.
although fossil records of mustelids in africa and south america are less well known than those of the northern continents, first appearance datums of both extant and extinct genera of mustelids nevertheless suggest that these regions also were colonized through successive dispersal events  <cit> .
among the extinct genera known from late miocene deposits in east africa that may have arisen from eurasian immigrants are the gigantic and cat-like ekorus, the largest mustelid discovered thus far, and an otter, vishnuonyx  <cit> .
in contrast, the earliest known remains of ictonyx striatus are from the pleistocene  <cit> , suggesting a later immigration into africa.
implications for mustelid community ecology
most studies of modern-day mustelid communities  have focused on recent ecological factors, such as partitioning of resources  via competition, to explain coexistence of species within these communities  <cit> .
as these studies have operated on an ecological timeframe, they implicitly assume that ecological differences among species have evolved relatively recently.
however, several observations derived from our biogeographical and phylogenetic results suggest that history  also has been an important component in the structure of mustelid assemblages on different continents.
first, these assemblages are largely composed of species belonging to different clades or lineages that differ significantly in their ecomorphology with regards to diet and locomotor mode  <cit> .
for example, seven species of mustelids are sympatric on the british isles in eurasia, with three species of weasels,  and one species each of mink , marten , badger  and otter   <cit> .
weasels, martens, badgers and otters obviously comprise a phylogenetically and ecologically heterogeneous set of species, yet these fundamental differences may indeed facilitate coexistence of these species within this community.
even the three species of weasels found in this community  are not closely related , suggesting that resource partitioning among these species may also be in part determined by historical causes.
second, studies of other vertebrate communities  have demonstrated that competition and divergence in ecological traits varies according to the degree of phylogenetic relatedness among species, such that, for example, competition is expected to be low among distantly related species but high among closely related species  <cit> .
divergence in ecological traits should therefore be greatest between closely related species, especially sister species occupying the same area.
within mustelids, there are several instances where sister taxa that are broadly sympatric within a region often exhibit pronounced ecological differences.
in africa, poecilogale albinucha preys almost exclusively on rodents whereas ictonyx striatus has a more generalized diet that includes invertebrate prey  <cit> .
similarly, mustela frenata and neovison vison overlap extensively in north america, but differ significantly in several aspects of their ecologies  <cit> .
the sister taxa aonyx cinerea and lutrogale perspicillata co-occur in parts of south and southeast eurasia with the former species feeding largely on freshwater crabs and the latter feeding mostly on fish  <cit> .
third, the fossil record along with molecular dating results indicate that continental assemblages of mustelids have been built up gradually over time .
this suggests that species interactions with these communities are temporally mosaic, with some interactions being ancient while others are more recent.
we certainly do not suggest that present-day ecological factors  operating over ecological time scales have not been important in determining the structure of modern-day mustelid communities.
instead, we simply suggest that historical factors should also be considered and that such a perspective has been lacking in most previous studies of mustelid community ecology.
as other researchers have pointed out, both historical factors and more recent ecological processes contribute to the structure of modern day communities  <cit> .
CONCLUSIONS
we have reconstructed a nearly complete generic-level phylogeny of the mustelidae based on a supermatrix of  <dig> gene segments.
using a variety of phylogenetic reconstruction methods, we have shown that mustelids are consistently resolved into four primary clades and three monotypic lineages and that nearly all nodes for this topology are well supported.
furthermore, by applying bayesian dating techniques we have shown two bursts of diversification, first during the miocene, which gave rise to the primary extant clades and lineages, and another during the pliocene, which gave rise to a large proportion of the species diversity observed today.
these bursts of diversification coincided with major paleoenvironmental and biotic changes that occurred during the neogene and are broadly contemporaneous with periods of rapid cladogenesis in other vertebrate groups.
such concordance in pattern and tempo of diversification suggests that global-scale changes have shaped the histories of many diverse groups of taxa  <cit> .
we used ancestral reconstruction of biogeographic areas to show that most of the extant diversity of mustelids originated in eurasia.
further, the mustelid fauna of africa, north america and south america are composed of taxa from nearly all major clades and lineages, suggesting that in situ speciation has been a relatively minor component in the assembly of these faunas.
finally, divergence times estimated from the molecular data combined with the fossil record suggests that different lineages of mustelids dispersed to africa, north america and south america in successive waves, which has implications for understanding the structure of mustelid communities.

 "wave after wave of immigration came in from asia, recruiting the fauna at each successive stage, but leaving little opportunity for new types to arise here. even those genera which seem to be of native origin, might prove to be immigrants, if all their history were known." the diversification of the mustelidae is a striking example of adaptive radiation, the evolution of ecological and phenotypic diversity from a common ancestor. mustelids exhibit both locomotor and dietary diversity, with taxa that are fossorial, semi-arboreal and semi-aquatic, and diets that vary from specialization on rodents to piscivory. ecomorphological diversity in the family is thus correspondingly high, reflecting the adaptation of different species of mustelids to different habits and habitats. as with other cases of adaptive radiation, resolving relationships within the mustelidae, especially among genera, has been challenging. many taxonomic schemes proposed for mustelids within the last century were based on morphology and classified genera into various numbers of subfamilies whose boundaries were largely determined by ecomorphological similarity. at one extreme, pocock divided extant mustelids into subfamilies based on descriptive analyses of external characters. at the other end, the system proposed by simpson cast mustelids into five subfamilies based on both phylogeny and'similarity in adaptiveness' of constituent genera: lutrinae, melinae, mellivorinae, mephitinae and mustelinae. while such a scheme may indeed reflect the true phylogeny in some instances, morphological similarity does not necessarily imply phylogenetic affinity, as has been well demonstrated in certain groups that exemplify adaptive radiation. moreover, such criteria can lead to recognition of polyphyletic grades rather than monophyletic groups. nonetheless, simpson's subfamilial classification of the mustelidae has been followed for many years, although the latest classification provisionally places all mustelids into two subfamilies, lutrinae and mustelinae. this latter scheme was proposed in recognition of demonstrated paraphyly of the traditional subfamilies by more recent morphological-based phylogenetic studies. during the last decade, dna sequence-based studies have begun to challenge the validity of the five-subfamily scheme and even monophyly of the family itself. studies using both mitochondrial and nuclear sequences have consistently demonstrated that skunks and stink badgers descend from a common ancestor and together form a lineage that diverged prior to the split between mustelidae and procyonidae. these studies and those more focused on mustelids have also suggested that: the lutrinae is monophyletic; but that both melinae and mustelinae are polyphyletic. however, phylogenetic relationships within the family remain uncertain or unknown because taxon sampling, especially for genera, has been incomplete. further, adequate character sampling is also an important issue because adaptive radiations are often composed of lineages that have rapidly diverged. short stem lengths of topologies revealed in recent studies, albeit with incomplete taxon sampling suggests that deeper lineages of mustelids may have radiated within a short span of time. adequate character sampling is therefore critical in achieving an accurate, well-resolved and robust phylogenetic hypothesis. fossil evidence indicates that the biogeographic history of mustelids is characterized by numerous intercontinental dispersals, primarily originating from eurasia where the earliest fossil remains of the family are found. for instance, a large majority of mustelid diversity in north america is considered to have originated from lineages that repeatedly dispersed from eurasia via the bering land bridge. the earliest immigrants to north america arrived in the early miocene and included a paraphyletic group of stem taxa referred to as'paleomustelids', whose affinities to crown group mustelids remains ambiguous, as well as genera that belonged to the extinct subfamily leptarctinae. the first appearance of various mustelid genera in north america is used to help define the beginning of biostratigraphic boundaries of north american land mammal ages during the neogene. for example, the first appearance of extinct genera trogonictis and sminthosinis and extant genera lutra and mustela help mark the latest hemphilian nalma ). there is still uncertainty, however, about the exact number of intercontinental dispersal events underlying the biogeographic distribution of extant genera and species and, therefore, how much of mustelid continental diversity is a result of in situ versus ex situ evolution. further, the sequence of dispersal events has been difficult to decipher, stemming from incompleteness of the fossil record. clarifying the biogeographic history of mustelids has implications for understanding the community ecology of mustelids. multiple species of mustelid are often found in the same community and consequently they have been the subject of important studies on character displacement and resource partitioning. however, many of these studies lacked a historical perspective, which has been shown to exert a strong influence on community assembly and structure. here, we present a nearly complete generic-level phylogeny of the mustelidae using ~ base pairs of mitochondrial and nuclear dna data obtained from gene segments. we use this phylogeny to address three objectives. first, we compare our phylogenetic hypothesis with previous hypotheses generated with morphological or molecular data. second, we estimate relative divergence times using new bayesian relaxed molecular clock methods to understand the temporal pattern of diversification in the family. do mustelids exhibit one or more bursts of rapid cladogenesis characteristic of many adaptive radiations? moreover, correlation of divergence times with paleoenvironmental changes can provide insight into the mechanisms that might have been responsible for bursts of diversification. finally, we assess the biogeographic history of the mustelidae, especially with regard to understanding dispersal history between continents of the old world and those of the new world.  phylogenetic relationships phylogenetic analyses using maximum parsimony, two different methods of maximum likelihood and bayesian inference all recovered the same hypothesis for intergeneric relationships, which resolves mustelidae into seven primary divisions that include four major clades and three monotypic lineages. otters form a clade that is sister to a clade comprising mink and true weasels. these clades, in turn, are sister to a clade that includes weasel-like species with aposematically colored pelage. next, ferret-badgers are a monotypic lineage that is sister to these three combined clades. the fifth major clade comprises two subclades, one containing hog-nosed and eurasian badgers and the other containing tayra, wolverine and martens. finally, as the earliest divergences in the tree, the american badger and honey badger form successive monotypic lineages sister to all other mustelid genera. intergeneric relationships observed are largely congruent with those recovered in recent analyses of mitochondrial and nuclear dna sequences. in contrast, our topology is highly incongruent with a cladistic analysis based on morphology. for example, bryant et al found that melogale was reconstructed as sister to all remaining mustelids and that eira, gulo and martes were polyphyletic. except for the monophyly of otters, skunks, and arctonyx and meles, all other nodes in the bryant et al tree had low bootstrap support values. furthermore, alternative topologies were recovered in the bryant et al study when certain characters were weighted differentially. regarding relationships within the four major clades, otters are resolved into three primary lineages, whose relationships are congruent with previous analyses based on fewer dna sequence data: one containing old world river otters and the sea otter, a second containing new world river otters and a third containing the monotypic giant otter. relationships of enhydra and hydrictis were unresolved in the bayesian consensus tree, but were resolved differently in mp and ml analyses. with mp, enhydra and hydrictis were resolved as sister taxa, whereas with ml, they were resolved as successive sister lineages to remaining old world otters, with hydrictis forming the first basal split. however, neither of these relationships was well supported. two clades comprise the true weasel and mink clade, with the new world american mink and long-tailed weasel in one clade and all other sampled mustela species in the second clade. within the latter clade, m. nudipes and m. strigidorsa are sister to a clade comprising species largely distributed in temperate regions of the northern hemisphere. despite the similar ecology, american mink and european mink are distantly related, as found in previous studies. species of true weasels and mink have been divided into five or nine subgenera based on morphological criteria. given the taxa we have sampled, our phylogeny suggests that only one proposed subgenus constitutes a natural group, putorius, containing the steppe polecat, european polecat and the north american black-footed ferret. further, mustela is paraphyletic with respect to neovison and suggests that the placement of the american mink in the separate genus neovison may not be warranted despite the observed differences in karyotype and morphology between this taxon and other species of mustela. in the third major clade, grisons of central and south america are sister to a clade containing the marbled polecat of asia and three african species, the libyan striped weasel, zorilla and striped weasel. monophyly of these genera is of interest because all exhibit an aposematically colored pelage combined with defense behaviors that include threat displays and excretion of pungent musk from enlarged anal glands. placement of vormela within this clade is congruent with a recent cytb study. interestingly, there is a north to south progression in branching order from vormela to i. striatus and p. albinucha of sub-saharan africa, which renders ictonyx paraphyletic. this suggests that p. albinucha should be placed into the genus ictonyx kaup, given the priority of the latter name over poecilogale thomas, as for the clade containing eira, gulo and martes, the last genus is clearly paraphyletic, in agreement with other recent studies. two new world species, the tayra and fisher either comprise a clade or form successive lineages sister to a clade containing wolverine and the remaining species of martes. our phylogeny indicates that true martens are monophyletic and sister to yellow-throated marten. within the subgenus martes, mp and ml analyses resulted in different phylogenetic placements of m. americana and m. melampus relative to the clade. in the mp tree, m. americana was sister to the clade ) whereas m. americana and m. melampus were joined as sister taxa in the ml tree. relationships of m. americana, m. melampus and were unresolved in the bayesian tree, resulting in a trichotomy. although more data is required to establish the precise branching order of the fisher, the strong support for this taxon being paraphyletic with the remaining species of martes strongly suggests placement of the fisher in its own genus, pekania. nodal support concatenation of bp, with indels coded as missing, includes variable characters and parsimony-informative characters. when indels are coded for information content, the concatenation is reduced to bp, with variable and parsimony-informative characters. the majority of internodes in our phylogeny have >90% bootstrap support and posterior probabilities, see figure and table indicating that our phylogenetic hypothesis, given our data set, is robustly supported. moreover, internodes are supported by one or more synapomorphic indels. three clades, however, have low support in mp, ml and/or bi analyses: nodes and internodes associated with these clades have very short branch lengths, suggesting that these splits occurred rapidly. alternatively, speciation may have occurred relatively recently, also resulting in inadequate phylogenetic signal, as among m. eversmanni, m. putorius and m. nigripes as well as m. americana, m. melampus and. many nuclear segments show no differences among species that comprise these groups, suggesting that there has been insufficient time for sequence differences to accumulate at these more slowly evolving loci. indeed, the four species of martes have been described as a superspecies complex of closely related, yet largely allopatrically distributed taxa. regardless of the exact cause, short branch lengths of these internodes can result in a high number of anomalous gene trees, leading to an incorrectly inferred species tree, especially when data from multiple data partitions is concatenated. additional analyses, including population genetic-level sampling, will be needed to confidently resolve relationships among these recently evolved species, as in reference for example. even so, internodes with low support may be stable or unstable, depending on the relative stability or instability of their constituent taxa or leaves. further, unstable taxa can influence nodal support in other parts of a phylogenetic tree. we measured leaf stability of all taxa using bootstrap trees from mp analyses and the program radcon. for three different measures of leaf stability, arctonyx collaris and meles meles were the least stable taxa. we repeated mp and hill-climbing ml analyses along with bootstrapping after excluding these two taxa, thereby reducing the data set to taxa. most bootstrap values in these analyses showed little or no change compared with the taxa data set, but interestingly, support for two stem clades increased from over 70% to more than 90%, as did average phylogenetic stability. the lower stability of a. collaris and m. meles and their influence on bootstrap support at internodes and may be caused by character conflict among different gene segments associated with these two taxa. however, we note that these nodes all have high or maximal posterior probabilities in bi analyses. with regards to subfamilial classification, our phylogeny clearly indicates that the mustelinae, as both traditionally and recently conceived, is polyphyletic, as suggested by previous studies. our results also indicate that various genera of badgers, most of which are often placed in the melinae, are also polyphyletic, supporting earlier conclusions based on morphology that these taxa are not closely related. instead, badgers with specific adaptations for fossoriality constitute a basal grade of lineages that are best recognized as distinct subfamilies. overall, our phylogeny is consistent with the subfamilial classification scheme recently proposed by sato et al and fulton and strobeck, which redefines the boundaries of traditional subfamilies as well as resurrects other subfamilies such as helictidinae and mellivorinae, thereby reflecting actual phyletic lines that have been difficult to resolve based on morphology alone. divergence times divergence times across the mustelid phylogeny were estimated using the uncorrelated relaxed lognormal molecular clock model calibrated simultaneously by eight fossil constraints. use of this molecular clock model provides a measure of rate heterogeneity among lineages or how well data conform or deviate from a strict molecular clock. for the root age and crown age prior combination of mya and mya, the coefficient of variation averaged across three independent runs was: – ), which suggests that the data show rate heterogeneity among lineages. we found greater rate heterogeneity among lineages when older root age and crown age prior combinations were used; for example, for mya and mya, averaged across three runs, σr =. furthermore, a likelihood ratio test significantly rejects the ml phylogeny with an enforced molecular clock versus one without a molecular clock thereby providing additional support that our data departs from clocklike behavior. in general, mean posterior estimates as well as 95% hpd intervals of node ages were highly consistent among different combinations of root age and crown age prior combinations. the root and node were most sensitive to alterations in priors, because they showed the largest shift in divergence times and little overlap in 95% hpd intervals in runs using older versus younger combinations of root age/crown age priors. nonetheless, sampling of the joint prior distribution by performing markov chain monte carlo analyses without any data suggested that the eight fossil calibration point priors did not have a strong influence on their estimated posterior distributions and thus, estimated divergence times. how do our estimates of divergence times compare with those based on previous molecular studies of mustelids? at nodes shared among the different studies, divergence times in previous studies are either a mix of younger, older and overlapping dates relative to those in our study or they tend to be older in general. for example, the estimated divergence time for the split between neovison vison and the remaining taxa of mustela was dated at – mya, – mya and 10– mya in sato et al, marmi et al and hosoda et al, respectively. in our study, this split, which also includes m. frenata as sister to n. vison, is dated at ~ mya. however, earlier studies differ in several important respects from our study. first, these studies were based on a smaller number of taxa and loci. for example, in sato et al and hosoda et al, taxon sampling largely consisted of species from martes and mustela and was based on one and two loci, respectively. undersampling of taxa and characters can bias divergence time estimates and reduces the power to detect rate variation among lineages. second, previous studies all employed a single root age fossil constraint, treated as a hard bound, precluding an accurate estimation of error associated with this calibration point. in contrast, we employed multiple fossil constraints simultaneously and placed soft-bound priors on these constraints to account for the uncertainty associated with the fossil record, thereby making it possible to evaluate errors associated with divergence time estimates. third, the previous studies each estimated a substitution rate that was assumed to be constant across their respective phylogenies. however, our results indicate that substitution rates vary across different lineages of the mustelid phylogeny, consistent with recent evidence from different groups that rates of substitution vary across lineages, even among closely related species. tempo and mode of mustelid diversification using the root age and crown age prior combination of mya and mya as a reference, our dating analyses indicate that, following the initial divergence of taxidea in the early miocene, mustelids underwent two main bursts of diversification. the first burst occurred during a ~ million year interval from the middle to late miocene and gave rise to most of the extant primary clades and lineages. the second and larger burst occurred during the pliocene epoch in which as many as generic-level or specific-level lineages originated within a million year span of time. furthermore, results of our biogeographic analyses show that the majority of cladogenetic events associated with these bursts of diversification occurred in the old world, largely in eurasia. paleoenvironmental and biotic changes driven by changes in climate during the latter half of the neogene may have promoted the two bursts of cladogenesis within mustelids. following the mid-miocene climatic optimum and onset of modern oceanic circulation, there is a marked cooling of the global climate near the end of the middle miocene that continues through to the holocene. this period of cooling coincides with formation of a permanent antarctic ice sheet in the mid to late miocene and an arctic ice sheet in the pliocene. in addition, several major sea-level lowstands occurred during the late miocene and pliocene, including the serravallian sea-lowering event near the beginning of the late miocene, 11- mya. these changes in climate and sea level increased overall terrestrial aridity and seasonality, which in turn promoted a shift from closed vegetation habitats to more open vegetation habitats. by the early late miocene, plant and animal fossil evidence indicates that the eurasian continent was a mosaic of vegetation types and generally more heterogeneous in vegetation structure relative to that in the early to middle miocene. these changes in vegetation had a concomitant impact on faunal communities and may have fostered diversification in a variety of lineages, including mustelids, via geographic isolation, divergent selection among different habitats, and/or ecological opportunity through the creation of new niches or the reorganization of former niches. interestingly, the initial radiation of the primary clades and lineages of mustelids coincides with a major faunal turnover in western europe that affected many groups of mammals, including the carnivora. in fact, nearly half of carnivoran species that went extinct during this turnover were mustelids and turnover in mustelids remained high throughout the late miocene in western eurasia. although evidence for faunal change in other parts of eurasia is less clear, changes in habitat and extinction of earlier lineages of mustelids may have created ecological opportunities that fostered the initial burst of diversification of modern mustelids. further cooling and drying during the pliocene, coincident with onset of high latitude glacial cycles, caused a dramatic expansion of low-biomass vegetation, including grasslands and steppe at midlatitudes and development of taiga at high latitudes of eurasia and north america. coupled with these changes was diversification of prey species such as rodents and passerine birds that exploited these new habitats, which in turn provided new niches for predators. part of the pliocene burst of diversification of mustelids may have been promoted by this diversification of prey species, as species in genera such martes and especially mustela became specialized in hunting small prey such as rodents. such a scenario is consistent with king's hypothesis that evolution of small body size in mustela was partly driven by adaptation to exploit abundant resources presented by rodent diversification during the pliocene. we also note that diversification of four of the five species within the subgenus martes, which are all closely associated with taiga forest habitat, coincides with expansion of this type of habitat across the holarctic during the plio-pleistocene. this finding is consistent with fossil evidence that indicates that taxa ancestral to these living species primarily evolved in forested habitats. in africa, the split between ictonyx libyca and the clade containing i. striatus and poecilogale albinucha dated at – mya, marginally overlaps with a major increase in african aridity and climate variability that occurred - mya, according to paleoclimatic and faunal evidence. moreover, this interval was accompanied by rapid radiation in several mammalian lineages such as bovids and hominids. our results suggest that divergence of i. libyca, which occurs in north africa along margins of the sahara desert, and the clade containing i. striatus and p. albinucha, which both occur south of the sahara, may have also been caused by this shift to greater aridity, especially considering that extensive desert conditions in the sahara did not occur until the late pliocene, around mya. if mustelid diversification was promoted by climatically driven paleoenvironmental changes, then synchronous patterns and tempos of diversification should also be recorded in the evolutionary histories of other, unrelated groups. a number of recent molecular phylogenetic studies on various groups of mammals and other vertebrates employing multiple loci and relaxed molecular clock methods provide corroboration for this hypothesis. for example, squirrels, cats, rabbits and hares, deer and woodpeckers each show one or more episodes of rapid diversification that are roughly contemporaneous with one or both of those observed in mustelids. taking cats as an example, seven of the eight primary lineages of felids radiated in the early part of the late miocene whereas genera and species that comprise the eight lineages largely radiated during the pliocene. we also note that divergence events from a number of independent lineages within the mustelidae are synchronous as well. for example, using the root age and crown age prior combination of mya and mya as reference, nodes and all codiversify around mya. similarly, nodes and overlap in their divergence time estimates of around mya. while ecological circumstances obviously differ from group to group, such congruence in patterns and tempos of diversification supports the idea that a common cause, namely, large-scale changes in past environments, has shaped the phylogenetic histories among disparate groups of organisms as well as independent lineages of mustelids. our results have important implications for theories about the mode and tempo of adaptive radiations. first, the scenario of diversification across different trophic groups we have outlined above is consistent with the model of cascading radiation proposed by stanley, in which diversification of one group in a trophic cascade promotes diversification in a second group, which in turn promotes diversification in a third group. this model could be tested more rigorously using dated phylogenies of multiple groups representing different trophic levels. second, our finding that the two primary bursts of cladogenesis in the evolutionary history of extant mustelids coincided with periods of climatically mediated environmental changes provides support for models that show that evolution may be greatly accelerated in temporally and spatially changing environments. during these times, new niches are created or former niches are reorganized, providing new ecological opportunities that may foster rapid speciation and thus, diversification. the fact that multiple groups show contemporaneous periods of rapid cladogenesis suggests that mainland environments undergoing environmental changes may function like newly colonized island archipelagoes in promoting diversification. historical biogeography the two-state and four-state biogeographical reconstructions indicate that the vast majority of the modern diversification of mustelids has occurred in the old world, specifically in eurasia. in fact, eurasia was unambiguously reconstructed as the ancestral area for nearly every ancestral node in the four-state analysis. these results are consistent with two other observations that suggest eurasia has been the center of mustelid diversification: eurasia contains the majority of extant species, with of the known species either exclusively endemic to or having part of their distribution on this continent; and the earliest fossils of extant lineages or those associated with the ancestors of extant lineages are often found in eurasia. nodes located near the base of the tree tend to be those where likelihood ratios are not significant, such as the root node and the node joining taxidea as sister to the remaining taxa of mustelids in the two-state analysis. such results are not uncommon in likelihood or other model-based approaches to ancestral state reconstruction because the degree of uncertainty associated with reconstruction increases with time. nonetheless, proportional likelihood values for these nodes still favor the old world as the ancestral area in the two-state analysis. ancestral reconstructions were robust when enhydra lutris, gulo gulo and mellivora capensis were coded for alternative states. in contrast to the extensive in situ diversification that has taken place in eurasia, mustelid faunas of africa and the new world are largely comprised of genera or species that repeatedly colonized these regions from eurasia. the mustelid fauna of africa contains eight species, seven of which are included in our taxon sampling. of these seven species, five are derived from separate colonizations from eurasia whereas two are derived from in situ speciation events; see figure similarly, our reconstructions show that nine separate dispersal events from eurasia and only one in situ speciation event accounts for the diversity of mustelids that are either endemic to north america or have part of their distribution there. genera and species of mustelids found in south america today are largely descended from north american immigrants that arrived as part of the great american interchange following the rise of the panamanian isthmus, - mya. such a relationship is clearly indicated for the clade of new world otters in which l. canadensis is sister to l. felina + l. longicaudis, with the latter two species found in central and/or south america. moreover, this split is estimated to have occurred – mya, which overlaps well with timing of the formation of the panamanian land bridge. the long-tailed weasel, m. frenata, ranges from north america to northern south america and two species of weasels are endemic to south america. fossil evidence clearly indicates that mustela colonized south america from the north, apparently well after the panamanian isthmus was in place. our results show that pteronura, galictis and eira dispersed separately into south america, with eurasia reconstructed as the continent of origin for each genus. these results are anomalous because these genera have been allied with extinct taxa from north america, suggesting a more proximate origin for these lineages. for example, pteronura may be related to the extinct genus satherium from the pliocene of north america. however, paleontological studies suggest that the ultimate ancestry of these extinct taxa lies in eurasia. a combination of evidence from the fossil record and divergence times from our phylogeny indicates that the mustelid faunas of africa, north america and south america have been assembled gradually over time. for example, fossil evidence suggests mustelids colonized the new world across beringia during different intervals when the land bridge between eurasia and north america was open. multiple genera of mustelids entered north america during the late miocene, prior to the first opening of the bering strait – mya, which severed the route across beringia. many genera that colonized north america during the late miocene or earliest pliocene became extinct. nonetheless, among the genera that may have been included in this wave of dispersal were the earliest representatives of lutra and mustela, both of which are first recorded in north america from the late miocene to early pliocene. these taxa may have been the forerunners of modern species of lontra and mustela/neovison found in north and south america today. the late miocene divergence time of the splits leading to lontra and neovison + m. frenata are consistent with this possibility. two extinct genera of american badgers, chamitataxus and pliotaxidea, are recorded from the late miocene, around mya and mya, respectively. pliotaxidea has been shown to be sister group to taxidea based on morphological evidence, thereby suggesting that the lineage leading to taxidea arrived in north america before the opening of the bering strait. meline badgers are presently found only in the old world. however, recent discovery of late miocene to early pliocene fossils of meline badgers at two different sites in north america indicates that this lineage had also immigrated into north america and was a component of the new world mustelid fauna. following these earlier dispersal events, fossil evidence indicates that mustela erminea, m. nigripes, m. nivalis and martes americana later entered north america during the pleistocene. the molecular divergence time for m. nigripes, around mya, for example, supports a pleistocene dispersal scenario for this species. although fossil records of mustelids in africa and south america are less well known than those of the northern continents, first appearance datums of both extant and extinct genera of mustelids nevertheless suggest that these regions also were colonized through successive dispersal events. among the extinct genera known from late miocene deposits in east africa that may have arisen from eurasian immigrants are the gigantic and cat-like ekorus, the largest mustelid discovered thus far, and an otter, vishnuonyx. in contrast, the earliest known remains of ictonyx striatus are from the pleistocene, suggesting a later immigration into africa. implications for mustelid community ecology most studies of modern-day mustelid communities have focused on recent ecological factors, such as partitioning of resources via competition, to explain coexistence of species within these communities. as these studies have operated on an ecological timeframe, they implicitly assume that ecological differences among species have evolved relatively recently. however, several observations derived from our biogeographical and phylogenetic results suggest that history also has been an important component in the structure of mustelid assemblages on different continents. first, these assemblages are largely composed of species belonging to different clades or lineages that differ significantly in their ecomorphology with regards to diet and locomotor mode. for example, seven species of mustelids are sympatric on the british isles in eurasia, with three species of weasels, and one species each of mink, marten, badger and otter. weasels, martens, badgers and otters obviously comprise a phylogenetically and ecologically heterogeneous set of species, yet these fundamental differences may indeed facilitate coexistence of these species within this community. even the three species of weasels found in this community are not closely related, suggesting that resource partitioning among these species may also be in part determined by historical causes. second, studies of other vertebrate communities have demonstrated that competition and divergence in ecological traits varies according to the degree of phylogenetic relatedness among species, such that, for example, competition is expected to be low among distantly related species but high among closely related species. divergence in ecological traits should therefore be greatest between closely related species, especially sister species occupying the same area. within mustelids, there are several instances where sister taxa that are broadly sympatric within a region often exhibit pronounced ecological differences. in africa, poecilogale albinucha preys almost exclusively on rodents whereas ictonyx striatus has a more generalized diet that includes invertebrate prey. similarly, mustela frenata and neovison vison overlap extensively in north america, but differ significantly in several aspects of their ecologies. the sister taxa aonyx cinerea and lutrogale perspicillata co-occur in parts of south and southeast eurasia with the former species feeding largely on freshwater crabs and the latter feeding mostly on fish. third, the fossil record along with molecular dating results indicate that continental assemblages of mustelids have been built up gradually over time. this suggests that species interactions with these communities are temporally mosaic, with some interactions being ancient while others are more recent. we certainly do not suggest that present-day ecological factors operating over ecological time scales have not been important in determining the structure of modern-day mustelid communities. instead, we simply suggest that historical factors should also be considered and that such a perspective has been lacking in most previous studies of mustelid community ecology. as other researchers have pointed out, both historical factors and more recent ecological processes contribute to the structure of modern day communities.  we have reconstructed a nearly complete generic-level phylogeny of the mustelidae based on a supermatrix of gene segments. using a variety of phylogenetic reconstruction methods, we have shown that mustelids are consistently resolved into four primary clades and three monotypic lineages and that nearly all nodes for this topology are well supported. furthermore, by applying bayesian dating techniques we have shown two bursts of diversification, first during the miocene, which gave rise to the primary extant clades and lineages, and another during the pliocene, which gave rise to a large proportion of the species diversity observed today. these bursts of diversification coincided with major paleoenvironmental and biotic changes that occurred during the neogene and are broadly contemporaneous with periods of rapid cladogenesis in other vertebrate groups. such concordance in pattern and tempo of diversification suggests that global-scale changes have shaped the histories of many diverse groups of taxa. we used ancestral reconstruction of biogeographic areas to show that most of the extant diversity of mustelids originated in eurasia. further, the mustelid fauna of africa, north america and south america are composed of taxa from nearly all major clades and lineages, suggesting that in situ speciation has been a relatively minor component in the assembly of these faunas. finally, divergence times estimated from the molecular data combined with the fossil record suggests that different lineages of mustelids dispersed to africa, north america and south america in successive waves, which has implications for understanding the structure of mustelid communities. 
adaptive radiation, the evolution of ecological and phenotypic diversity from a common ancestor, is a central concept in evolutionary biology and characterizes the evolutionary histories of many groups of organisms. our biogeographic analyses show eurasia as the center of origin of mustelid diversity and that mustelids in africa, north america and south america have been assembled over time largely via dispersal, which has important implications for understanding the ecology of mustelid communities. biogeographical analyses indicate that most of the extant diversity of mustelids originated in eurasia and mustelids have colonized africa, north america and south america on multiple occasions. using bayesian dating techniques, we provide evidence that mustelids underwent two bursts of diversification that coincide with major paleoenvironmental and biotic changes that occurred during the neogene and correspond with similar bursts of cladogenesis in other vertebrate groups. we constructed a nearly complete generic-level phylogeny of the mustelidae using a data matrix comprising gene segments analyzed with maximum parsimony, maximum likelihood and bayesian inference methods. mustelids are also widely distributed, with multiple genera found on different continents. we show that mustelids are consistently resolved with high nodal support into four major clades and three monotypic lineages. as with other groups that have undergone adaptive radiation, resolving the phylogenetic history of mustelids presents a number of challenges because ecomorphological convergence may potentially confound morphologically based phylogenetic inferences, and because adaptive radiations often include one or more periods of rapid cladogenesis that require a large amount of data to resolve. extant mustelids display extensive ecomorphological diversity, with different lineages having evolved into an array of adaptive zones, from fossorial badgers to semi-aquatic otters. combined with information from the fossil record, our phylogenetic and dating analyses suggest that mustelid diversification may have been spurred by a combination of faunal turnover events and diversification at lower trophic levels, ultimately caused by climatically driven environmental changes.
 adaptive radiation, the evolution of ecological and phenotypic diversity from a common ancestor, is a central concept in evolutionary biology and characterizes the evolutionary histories of many groups of organisms. one such group is the mustelidae, the most species-rich family within the mammalian order carnivora, encompassing species classified into genera. extant mustelids display extensive ecomorphological diversity, with different lineages having evolved into an array of adaptive zones, from fossorial badgers to semi-aquatic otters. mustelids are also widely distributed, with multiple genera found on different continents. as with other groups that have undergone adaptive radiation, resolving the phylogenetic history of mustelids presents a number of challenges because ecomorphological convergence may potentially confound morphologically based phylogenetic inferences, and because adaptive radiations often include one or more periods of rapid cladogenesis that require a large amount of data to resolve.  we constructed a nearly complete generic-level phylogeny of the mustelidae using a data matrix comprising gene segments analyzed with maximum parsimony, maximum likelihood and bayesian inference methods. we show that mustelids are consistently resolved with high nodal support into four major clades and three monotypic lineages. using bayesian dating techniques, we provide evidence that mustelids underwent two bursts of diversification that coincide with major paleoenvironmental and biotic changes that occurred during the neogene and correspond with similar bursts of cladogenesis in other vertebrate groups. biogeographical analyses indicate that most of the extant diversity of mustelids originated in eurasia and mustelids have colonized africa, north america and south america on multiple occasions.  combined with information from the fossil record, our phylogenetic and dating analyses suggest that mustelid diversification may have been spurred by a combination of faunal turnover events and diversification at lower trophic levels, ultimately caused by climatically driven environmental changes. our biogeographic analyses show eurasia as the center of origin of mustelid diversity and that mustelids in africa, north america and south america have been assembled over time largely via dispersal, which has important implications for understanding the ecology of mustelid communities. 