Asparagus acutifolius L. is a dioecious and native plant species, widely distributed in the Mediterranean Basin. It is known for its fine flavour and could represent an important resource for cultivation programs in desert areas. Few molecular studies have been performed on this species. In the present paper, the ISSR technique was employed to study genetic diversity in Italian A. acutifolius.ST (0.4561) and Theta B (0.4776) values indicate a wide genetic variation among the samples examined. The distance UPGMA tree grouped together the genotypes strictly according to their geographical origin, showing that each sample is genetically structured and can be considered a distinct population. AMOVA analysis further confirmed genetic structuring of the populations. Population-specific fragments were also detected.Twenty-three primers produced a total of 228 polymorphic fragments used to evaluate genetic variation. FA. acutifolius according to geographical origin, and confirm the importance of genetic studies for designing germplasm conservation strategies.The results suggest that ISSR markers are useful in distinguishing the populations of  The availability of a variety of DNA markers, such as restriction fragment length polymorphism (RFLP), amplified fragment length polymorphism (AFLP), random amplified polymorphic DNA (RAPD), simple sequence repeat (SSR) and intersimple sequence repeat (ISSR) has enabled researchers to investigate genetic diversity among various plant species across natural populations -5. Amongin situ and ex situ of genetic diversity in natural populations is imperative to guarantee sustainable development [Local populations of traditional cultivars provide a valuable resource for plant breeding as well as for the preservation of genetic diversity . The expelopment .Asparagus acutifolius L. (Liliaceae) is a native, perennial plant species widely distributed throughout the Mediterranean areas, whose flowers are classified as dioecious and are mainly bee-pollinated; it generally does not reproduce by self-pollination. It grows in bushy and semi-dry places, sunny or semi-shade, mainly on limestone.A. officinalis and does not require rich soils for cultivation; for these reasons, it could be an economically important resource for the recovery of arid rural areas where controlled introduced programs could be achieved.This species is known for its strong taste compared to the cultivated A. officinalis, for which many molecular markers have been characterized  [A. acutifolius are drawn from RAPD analyses [Viola pubescens [Oryza [To date, there is little information available on the genetic variability of this species. At present, the most widely studied species is P, AFLP) . The fewanalyses  and the analyses . The ISSanalyses . They haanalyses , Viola pubescens , potato ubescens , and Orys [Oryza .A. acutifolius collecting samples in eight different scattered rural areas: six continental and one each from the Italian islands of Sardinia and Sicily.In this study we used ISSR markers to analyse the genetic diversity of Italian A. acutifolius was collected and their characteristics.Figure A. acutifolius is summarized in Table Among the 42 primers tested, 23 proved useful to characterize the samples  was exhibited in the Sassari and the lowest (35.09%) in the Recco samples.Genetic structuring was evident due to the detection of specific bands in each sample examined. Spoleto and Caserta samples showed one and three fixed specific fragments, respectively, found to be statistically significant (P < 0.0001). For the other samples, 27 ISSR specific polymorphic fragments were detected, with a varying degree of statistical significance ranging from P < 0.0400 to P < 0.0001.Genetic distances  were exaSamples collected at different geographic site grouped together, as shown in the UPGMA tree Fig. , and theThe values of gene diversity are summarized in Table T) was 0.2618 ± 0.0240 and the average variation within samples (average HS) was 0.1424 ± 0.0084. The mean diversity among the samples (DST) was 0.1194. The fixation index FST = (HT-HS)/HT was 0.4561, indicating a reduction of genetic diversity of about 45%. The Theta-B value obtained by Hickory analysis is an estimate of FST under a random-effects model of population sampling. Its mean value is 0.4766 ± 0.0173; the HT and HS values are, respectively, 0.2859 ± 0.0115 and 0.1619 ± 0.0026 showing that there is a general agreement between the results obtained using the two different approaches.As summarized in Table A. acutifolius in an Italian population. ISSRs also revealed diversity within each sub-population. The results obtained are in accordance with the principle that the number of individuals used to estimate average heterozygosity can be very small if a large number of loci is studied [ISSR markers can be used in population genetic studies of plant species as they effectively detect very low levels of genetic variation . They al studied .The gene diversity values Table  ranged fST and the Theta-B value (0.4766) demonstrated a very great genetic differentiation among samples, possibly caused by random genetic drift. Further statistical support to the genetic structuring of the samples examined comes from the AMOVA analysis. Despite the continuous distribution of A. acutifolius, the eight samples represent genetically differentiated populations.The fixation index is 0.4561, which indicates a substantial reduction of genetic diversity (about 45%), probably due to the high genetic isolation of samples analysed. The FA. acutifolius populations.In each population it was possible to identify ISSR specific fragments. As shown in Table A. acutifolius throughout the Italian peninsula, there is poor gene flow through the isolates. The high genetic differentiation of the A. acutifolius populations examined might be attributed to the kind of pollinators (mainly bees) that can act at short distances, preventing the gene flow, and to the effects of anthropogenic habitat fragmentation.The high degree of genetic differentiation is confirmed by the UPGMA tree topology, in which all accessions from the same population grouped together Fig. . The popA. acutifolius populations [The results obtained using ISSR markers are in agreement with the RAPD analysis that also identified population-specific fragments in different Italian ulations .A. acutifolius has well-differentiated populations, despite their morphological low variability. These results show that to maintain genetic diversity within A. acutifolius it is necessary to conserve many populations.Information about the spatial organization of genetic variability is essential for the conservation of genetic resources . Our resA. acutifolius collected in the eight different locations in Italy  and 1X Taq DNA polymerase buffer, in a total volume of 25 μL. The amplification programme was 1.5 min at 94°C; 35 × 40 s at 94°C, 45 s at the primer annealing optimal temperature  and negative and positive controls were loaded and run at constant voltage (150 V) for 2 hours. After running, the gels were UV visualised and recorded using a Kodak Digital Science dS1D DC40/DC120 Camera. To verify the repeatability of the results, each DNA extraction, PCR amplification, and gel running was repeated twice.Unequivocally scorable and consistently reproducible amplified DNA fragments were transformed into binary character matrix .T), heterozygosity within population (HS), diversity among populations (DST), fixation index (FST), and genetic distance [Genetic variation within and among sub-populations was analysed on the basis of the banding profile using various parameters such as percentage polymorphism (P), total heterozygosity (Hdistance ,22-24, udistance . Since Idistance  based onAMOVA analysis, implemented in Arlequin , was conThe Mantel test of genetic and geographic distances was carried out to evaluate the correlation between the two data matrices.The UPGMA tree was generated using the PAUP*4.0 software , and BooMS carried out part of the sample collection, designed ISSR primers and carried out ISSR work; GG did part of DNA extraction; SM carried out part of the sample collection and DNA extraction; LG participated in the manuscript preparation and revision; SA conceived the study, carried out data analysis, co-ordination and interpretation of the results.


Grewia flava, a common, fleshy-fruited shrub species in the southern Kalahari. Understanding the population dynamics of Grewia flava is a crucial task, because it is widely involved in the shrub/bush encroachment process, a major concern for rangeland management due to its adverse effect on livestock carrying capacity and biodiversity.In Southern Africa savannas climate change has been proposed to alter rainfall, the most important environmental driver for woody plants. Woody plants are a major component of savanna vegetation determining rangeland condition and biodiversity. In this study we use a spatially explicit, stochastic computer model to assess the impact of climate change on the population dynamics of z of the time-shrub density relationship to quantify the population trend. For each climate change scenario we then compared the departure of z from typical stable population dynamics under current climatic conditions. Based on the simulation experiments we observed a positive population trend for scenario (1) and a negative trend for scenario (2). In terms of the projected rates of precipitation change for scenario (3) and (4) population dynamics were found to be relatively stable. However, for a larger increase in inter-annual variation or in temporal auto-correlation of rainfall population trends were negative, because favorable rainfall years had a limited positive impact due to the limited shrub carrying capacity.For our study we consider four climate change scenarios that have been proposed for the southern Kalahari for the coming decades: (1) an increase in annual precipitation by 30–40%, (2) a decrease by 5–15%, (3) an increase in variation of extreme rainfall years by 10–20%, (4) and increase in temporal auto-correlation, i.e. increasing length and variation of periodic rainfall oscillations related to El Niño/La Niña phenomena. We evaluate the slope We conclude that a possible increase in precipitation will strongly facilitate shrub encroachment threatening savanna rangeland conditions and regional biodiversity. Furthermore, the negative effects found for positive auto-correlated rainfall support current ecological theory stating that periodically fluctuating environments can reduce population viability because species suffer disproportionately from poor environmental conditions. In order to assess biodiversity response under climate change, Hannah et al.  recentlyGrewia flava DC, a common, fleshy-fruited, woody plant species of South African savannas. Understanding the population dynamics of Grewia flava is a crucial task, because it is widely involved in the shrub/bush encroachment process . Shr. ShrGrewss e.g. -13) and  and Grewss (e.g. ). In the)s Figure . Thus, wnsidered .Grewia flava population trend. Above this threshold frequent droughts kill the offspring and inhibit emergence. Ecological theory generally suggest that population viability decreases with increase in environmental variability or stochasticity [An increase in variability of more than 100% revealed a significant threshold behavior with a negative asticity , becauseasticity . HoweverICV, i.e. intra-cycle variation and PL, i.e. period length to evaluate the impact of climate change related increase in positive auto-correlation of rainfall. In our study region, Kruger [PL value of 22 years between 1955 and 1991. The ICV values used in our simulation experiments refer to an increase in the probability of an extreme rainfall category and are thus difficult to compare with the cyclic variability of actual total rainfall. However, we compared the ICV values from our model with departures of annual rainfall from the inter-annual mean of a typical rainfall oscillation provided by Kruger [ICV value above 60% resembles a realistic rainfall variation between favorable and unfavorable periods of a 22-year oscillation. Although we did not find a negative impact on Grewia flava population dynamics for this scenario, a slight increase in ICV combined with an increasing length of El Niño/La Niña phases might have a significant negative impact on the population viability of Grewia flava  outweigh positive effects in wet years (i.e. increased reproduction). This is noteworthy, as mean annual rainfall and inter-annual variation was kept constant. Until now, this has been primarily shown in solely theoretical studies with under-compensating population dynamics . ,32. 31,3cs (e.g. ) or indics (e.g. ). To ourSGM it is important to discuss the inference of actual precipitation amounts on probability values. Threshold values for extreme rainfall years are based on expert knowledge and may vary depending on the plant species considered. Moreover, percentage changes in extreme rainfall years, as applied in our model, are not necessarily equivalent to percentages changes in mean annual rainfall. For example, an increase in 'high' rainfall years by 10% represents an increase in mean annual rainfall of approximately 10%, depending on actual precipitation amounts in each 'high' rainfall year. Thus, it is important to derive principal population trends from the SGM model results rather than absolute predictions. Another issue regards the temporal dimension: long-term SGM simulation periods deviate from short-term predictions of climate change. However, population trends applied for only a few decades may yield biased results due to the cyclicity of Grewia population dynamics (see [In order to assess the predictive power of ics (see ). Finallics (see ).Grewia flava is relatively long-lived, fire and drought-resistant. Intuitively, this would suggest a low climate change impact on Grewia flava population dynamics. However, we found that despite Grewia flava's capability to survive in a harsh environment, it may be strongly affected when rainfall decreases as predicted, or increases in periodical fluctuations. This may also include possible range shifts in regional distribution which have not been studied here. Based on the model and the climate change scenarios analysed, it would be inappropiate to forecasts changes in the geographical distribution of Grewia flava, because SGM parameters have been estimated and validated for populations in the Kimberley region of the southern Kalahari. Moreover, current geographical distribution suggests that buffer mechanisms may facilitate survival in more arid parts of the southern Kalahari .In this study we have shown that climate change may have severe and sometimes unexpected implications at a regional species level. A woody plant like Grewia flava encroachment in the southern Kalahari. However, increase in rainfall will likely enhance this process. A shift from typical open grassland with solitary trees towards wide spread, homogeneous Grewia flava thickets implies negative cascade effects for other species, resulting in the loss of biodiversity [Decrease in rainfall may reduce the severity of iversity . As a fuSGM and previous results have been presented elsewhere . SGM. SGMSGM Grewia flava typically grows beneath the dominant tree species Acacia erioloba [Grewia flava. Under high cattle grazing a substantial proportion of seeds may be distributed into the open matrix vegetation, since cattle feed on the foliage and fleshy fruits of Grewia flava [Grewia cover, particularly around boreholes . Grewiater fire  and low ter fire . Size clter fire .SGM  (i.e. occupied by Acacia erioloba) or matrix type (M) (i.e. grassland vegetation). However, it switched from T to M status when a tree died after a mean life span of 200 years. The converse occurred where a new tree establishes. Initial tree distribution and spatial recruitment pattern was random with a constant density of 5 trees ha-1 for the entire simulation period. In each time step, SGM simulated important life history stages, environmental conditions and the key ecological processes of Grewia flava, the majority of which are governed by annual rainfall patterns  repr reprSGM ari (see ), an ini-1. Rain falls almost exclusively during summer (November to April) with an inter-annual mean of 417 mm for the period 1940 – 2000 . For this period we defined a threshold value of 150 mm below and above the long-term mean to classify years into 'low', 'average' and 'high' rainfall years  . Similar classifications have been applied in other studies, e.g. in a spatial simulation model of Acacia raddiana in the Negev Desert [Annual rainfall in the southern Kalahari is highly variable, with precipitation ranging between 200 and 700 mm yrly) (see ). Accordv Desert .Grewia size classes and may vary depending on the annual rainfall . Seed dia flava  and reprcal data , the assGrewia flava seeds [M and T. However, depending on annual rainfall conditions, the probability of emergence in the model may vary between 1–2% in average and 2–4% in high rainfall years . As survival of Grewia flava seeds in the soil is very low, seeds that do not emerge are assumed to die.Even though microsite types differ markedly in micro-environmental conditions, they do not differ in emergence rates of va seeds  suggestiSGM, fire was simulated probabilistically for the entire grid and may only occur in high and average rainfall years. Average frequency of 7.9 years in the model is supported by fire intervals reported from similar savannah types [T cells, fire mortality probability is 0% for adults and 75% for seedlings. These model assumptions are realistic, since grass fuel accumulation and fire severity beneath trees is less than in tree inter-spaces . In . In SGM,ption in ). Annualption in  and Schuption in . For seeption in ). All seGrewia flava individuals into categories of small , medium-sized  and large plants  as well as seedlings . We assumed a maximum age for each size class member with MaxAgeS = 5 years for small and MaxAgeM = 25 years for medium-sized shrubs. These estimates are based on annual shoot growth rates and aerial photograph analysis of the study area  = 0.2 for small and Ptransition(M) = 0.1 for medium-sized shrubs. No growth occurs in low rainfall years.In the model, each shrub was assigned a size class with an average canopy cover. For definition of shrub size classes we used the canopy volume to group rea (see ). In eacK = 25 m2 as maximum total cell shrub cover. If shrub cover exceeded K, individuals in the cell died. Density-dependent mortality was simulated annually by removing the smallest individuals first, i.e. in descending order of size class, until shrub cover <K . Inter-rix (see ). For thz-value, i.e. the slope of the year – shrub density relationship. This is a reasonable approach for analyzing time-abundance relationships . When pBased on the standard scenario we performed simulation experiments for each of the four different scenarios of climate change with variation in inter-annual mean of precipitation, coefficient of inter-annual variation, and temporal auto-correlation . Probability of low rainfall years is constant whereas frequency of average years is reduced, accordingly.(1) increase in precipitation, i.e. stepwise increase of Plow resulting in a higher probability of low rainfall years (increase in drought events),(2) decrease in precipitation, i.e. stepwise increase of Plow and Phigh resulting in a higher probability of low and high rainfall years at the cost of average years ,(3) increase in inter-annual variation of precipitation, i.e. stepwise increase of P(4) increase in temporal auto-correlation of rainfall with 'good' and 'poor' phases.For the scenario (4) we varied two parameters:PL, i.e. longer phases of favorable and unfavorable rain conditions and,(4a) introduction of rain cycles with increasing period length ICV, i.e. increasing variation between alternating favorable and unfavorable periods within one rain cycle.(4b) increase in intra-cycle variation PL we applied a period length of 10, 20, 50, 100 and 250 years, respectively, with each period subdivided into a similarly long 'good' and 'poor' phase. For example, a 10-year period length was subdivided into a 5-year period of superior and a 5-year period of poor conditions. For an increase in ICV we alternately increased and decreased high and low rainfall probabilities in each period, respectively (see 4a and 4b in Table ICV value of 100% Plow equaled 0.00 and Phigh 0.26 in a 'good' phase, whereas values were 0.30 and 0.00, respectively for a 'poor' phase. Through this procedure inter-annual mean and variation of rainfall probabilities were identical for each scenario and the default set of rainfall probabilities. Combined increase in PL and ICV resulted in an increase of positive auto-correlation. Near-decadal epochs of above- and below-normal rainfall have been identified for the period 1955–1991 [ICV under predicted climate change.For SGM simulation model, conceived the study and drafted the manuscript. FJ supervised the study and the manuscript writing. Both authors read and approved the final manuscript.JT developed the


Scientists continue to argue the extent that human activities drive global warming, but few would argue that it exists. The International Panel on Climate Change predicts that greenhouse gases will increase global temperatures by 3.6 degrees F by 2100—a rise unprecedented over the past 10,000 years. What might the world look like as we approach that point? Wetlands will disappear. Floods, hurricanes, and droughts will become progressively more severe. Infectious diseases will increase in virulence and range. Montana's famed glaciers may all but disappear within 30 years. A quarter of species may vanish by 2050.While the effects of climate change on species' geographic ranges and population dynamics have been studied to some extent, scientists know little about how species respond to climate change at the genetic level. In this issue, Elizabeth Hadly and colleagues analyze three different dynamic processes—environmental change, population response, and gene diversity fluctuations—and present evidence that climate change influences variation in genetic diversity.Focusing on two mammal species—the Montane vole and the northern pocket gopher—Hadly et al. asked how the two species responded to historical climate-induced habitat alterations in northwestern Wyoming. They gathered fossils from Yellowstone National Park's Lamar Cave, which contains a treasure trove of carbon-dated deposits that mirror the community of mammals in the area today. Comparing genetic material extracted from fossil samples from different time points over the past 3,000 years to genetic data taken from contemporary animals, Hadly's team tracked genetic changes in populations of the two species and used this information  to estimate changes in effective population size over time.  The genetic and demographic data were then combined with environmental records to analyze the relationship between the factors.Studying these populations in space and time—an approach the authors call “phylochronology”—offers an opportunity to analyze the genetic diversity of a species against the backdrop of environmental fluctuation within an evolutionary time frame. It also suggests how microevolutionary forces—factors that affect genetic variation in populations over successive generations—shape genetic responses to climate change. Such evolutionary forces include mutation, genetic drift , and gene flow (changes in the gene frequency of a population caused by migration).The past 3,000 years includes two periods marked by dramatic climate change—the Medieval Warm Period and the Little Ice Age—that had different effects on local mammal populations depending on their habitat preferences. Habitat specialists, the vole and pocket gopher live in the wet mountain regions of western North America. Though both showed population increases during wetter climates and declines during warmer periods, Hadly et al. predicted the gene diversity fluctuations of the two species would differ based on their different ecological behaviors. And that's what they found: genetic response is tied to population size. Pocket gophers have low population densities, stick close to home, and are fiercely territorial, while voles live in high-density populations and range more widely. For the gophers, population declines resulted in reduced gene diversity; for the voles—which have a larger effective population size and greater dispersal between populations—population declines resulted in increased gene diversity. But what forces underlie these differences in genetic variation?A recent study suggests that migration (a primary agent of gene flow) is most common in and between low-density patches in vole populations, which implicates gene flow as the driver of gene diversity patterns. But the authors don't rule out selection as a possibility, and suggest how to go about resolving the question. Hadly et al. show that phylochronology opens a unique window onto the relatively recent evolutionary past and offers “the potential to separate cause from effect.” They also conclude that “differences in species demography can produce differential genetic response to climate change, even when ecological response is similar.” With a 3-degree temperature increase in just the past 50 years in the American West, conservation of biodiversity may well depend on such insights.


We use ancient DNA to compare two independent estimates of population size  and corroborate our results with gene diversity and serial coalescent simulations. Our data and analyses indicate that, with population size decreasing at times of climatic change, some species will exhibit declining gene diversity as expected from simple population genetic models, whereas others will not. While our results could be consistent with selection, independent lines of evidence implicate differences in gene flow, which depends on the life history strategy of species.Understanding how climatic change impacts biological diversity is critical to conservation. Yet despite demonstrated effects of climatic perturbation on geographic ranges and population persistence, surprisingly little is known of the genetic response of species. Even less is known over ecologically long time scales pertinent to understanding the interplay between microevolution and environmental change. Here, we present a study of population variation by directly tracking genetic change and population size in two geographically widespread mammal species  A novel analysis of paleoclimatic data, fossil abundance and contemporary and ancient DNA from different time periods introduces "phylochronology" and reveals how species respond genetically to climate change Phylogeography has advanced our understanding of the spatial distribution of genetic diversity within and between species . HoweverOur study takes advantage of a continuous, well-sampled mammalian fossil sequence spanning the last 3,000 years . Lamar Cave has an extraordinarily complete representation of the local species in the vicinity, with over 10,000 identified mammalian specimens representing over 80% of the mammal species in the local habitat . Late-HoMicrotus montanus (montane vole) and Thomomys talpoides (northern pocket gopher), species that presently are widespread in mountain habitats of western North America. Due to their preferences for wetter habitats, both responded demographically by increasing in relative abundance during wetter climates and declining during warmer climates. M. montanus showed an increase in abundance relative to other common rodents during periods of wet, cool climate in Yellowstone  techniques allows us to investigate directly the impacts of these environmental perturbations on neutral genetic diversity concurrent with these species population responses. We obtained ancient and modern mitochondrial DNA sequences from M. montanus and used previously published data for T. talpoides. Although these two species are broadly similar in body size  and are principally herbivorous, they differ in their natural history. T. talpoides is characterized by low population densities (1–62 gophers/hectare [ha]), a fossorial mode of life, maximum dispersal distances of a few hundred meters, and fiercely territorial behavior  despite the absence of obvious migration barriers.Although the population responses of behavior . Populatcalities . This spM. montanus achieves higher average population densities (60–186 voles/ha)  have found little evidence for population subdivision over the scale of hundreds of kilometers   than T. lometers , as expelometers . Such deplotypes .The primary advantage of a phylochronologic approach as opposed to a single time slice for understanding mammalian response is the ability to reveal changes in genetic variation through time. This is in contrast to modern genetic studies that seek to reconstruct demographic history based on inferences from past climate or geologic records e.g., . Our stuM. montanus and T. talpoides fossil specimens and modern population densities. (2) We calculated gene diversity over this 3,000-year period to determine the impact of environmental perturbations on genetic effective population size and gene diversity in M. montanus and T. talpoides. Unlike previous aDNA work  might influence the record of gene diversity over time. Thus, we combined four methods of estimating population size, determining statistical significance, and assessing gene diversity through time. (1) We derived independent ecological estimates of population size through time from abundances of DNA work , we usedDNA work . Our appT. talpoides, we predicted that changes in genetic variation through time would be dominated by drift, as suggested by the modern life history characteristics of small effective population size, low dispersal, and high amounts of population substructure. Therefore, as the ecological effective population size of T. talpoides declined with warmer climates, we expected genetic variation to decline. For M. montanus, we predicted that changes in genetic variation through time may be influenced more by migration, as suggested by large effective population sizes, high rates of dispersal, and low amounts of population substructure. As a result, past declines in ecological estimates of population size of M. montanus would not necessarily have resulted in a decrease in genetic variation.Despite similar population-level responses to climatic change of the late Holocene, we expected differences in gene diversity change for the two species. For e_ecol(N >> e_gen)N  . Howeveralpoides A and M. montanus B. For M.M. montanus suggests that the sequences we obtained for this study are target mtDNA. Of the 312 bp we sequenced for M. montanus, 96.5% of all the mutations were third-position codon changes, with first- and second-position mutations accounting for 3.5% and 0%, respectively. These ratios of variation are concordant with expectations for within-species variation and small overall sequence divergences . Most of the mutations are synonymous (97.7%); the transition-to-transversion ratio of the entire data set was 4.1 to 1, which is consistent with expectations for mammalian cytochrome b and evolution in other Microtus species  and translated successfully. Together with the frequency distribution of our pairwise differences, the prevalence of silent and third-position codon changes, and the standard of obtaining both forward and reverse fragments of overlapping sequence regions, these data permit us to conclude that the genetic diversity we have sampled represents authentic mitochondrial population variation and is unlikely to be from nuclear copies or pseudogenes.The genetic evidence we have assembled from ergences . Nucleot species , with an species . Fossil R2 = 0.004, not significant). All but one (out of 121) of the extraction controls were negative. When sequenced, this extraction blank BLASTed similar to Montanus townsendii, a taxon we had never worked on in the facility; this sequence has not since been amplified in the lab, and that extraction was not used further. Out of 87 successfully amplified samples and one sequence obtained from GenBank (AF119280), we identified 17 haplotypes within four haplogroups (A–D) of M. montanus  fall within haplogroups A and D, with 84% of the samples within one substitution of the locally ancestral haplotype A  and a deficit of guanine (11.4%). All mutations were synonymous; the transition-to-transversion ratio of the entire data set was 4.0 to 1.Protocols for M. montanus and T. talpoides T. talpoides (N exhibit the same trend as the genetic estimates e-tt_gen),(N are lower than those derived from genetic data e-mm_gen) for all time points except for 2,525 ybp, where observed gene diversity was significantly lower than could have been calculated given our sample size (n = 4), suggesting that we do not have sufficient power to detect processes at this time period.For ple size . Since tT. talpoides is not different given expectations from a closed population, we attempted to determine the statistical limitations of these data. Investigation of the effects of sampling for T. talpoides revealed that given the smaller number of base pairs (64 bp), we do not have enough statistical power to reject the null hypothesis. For every sampling time point, the Ewens distribution predicted that only one haplotype would be present in the genetic samples (unpublished data). As a result, the predicted gene diversity was zero for all time points. Because the observed gene diversity for T. talpoides was higher than predicted we also simulated five combinations of mutation rate and abundance, which could result in a higher predicted diversity  for 166 ybp (n = 34). Results for all five combinations predicted presence of a single haplotype. Since we do not have adequate statistical power given the genetic data for T. talpoides, we did not conduct significance tests for this species. However, the observed values of gene diversity are not unexpected from dynamics within a closed population.Although the gene diversity for M. montanus gene diversity throughout the entire time series   based on the observed change in 166 ybp) . The expM. montanus prior to 2,525 ybp. We modeled population reduction to 104,577 (0.75 × e-mm_ecol2525ybpN), 69,718 (0.5 × e-mm_ecol2525ybpN), and 34,859 (0.25 × e-mm_ecol2525ybpN) prior to 2,525 ybp. For a 75% bottleneck prior to 2,525 ybp, we were no longer able to reject the null hypothesis of a closed population. These results indicate that an extreme bottleneck where population size was reduced to 75% or more might result in the observed change in gene diversity. Our simulations reveal that unless an extreme bottleneck happened prior to 2,525 ybp, we can be confident that the observed change in gene diversity is not likely to be from events that occurred immediately prior to our historic data, and thus is due to migration or selection.Our results may depend on our assumptions of equilibrium population size prior to 2,525 ybp. We investigated sensitivity to this assumption by modeling a population bottleneck in M. montanus. Historic genetic data allow us to discriminate between population processes over millennia much better than do modern data alone.Coalescent simulations used to investigate the significance of the observed gene diversity value for the modern samples demonstrated that the null hypothesis of past population size change could be rejected at only two of the nine mutation rate and density combinations. These results reveal that given data from only the modern samples, it was not possible to reject the null hypothesis of past population size change in M. montanus populations. Estimates of β-diversity (used here to measure haplotypic turnover) from haplotypic data reveal that turnover was highest between 2,525 and 845 ybp . Closer examination of the haplotypic distributions demonstrates that five novel haplotypes appeared by 845 ybp, three of which are ≥3.2% different from the most common haplogroup (A)  A at 2,52T. talpoides and M. montanus between 1,438 and 470 ybp, a period spanning the Medieval Warm Period  results in an increase in gene diversity. We attempted to statistically validate our results by the use of serial coalescent simulations to demonstrate that the change in gene diversity of M. montanus between 2,525 and 845 ybp is significantly different from that expected based on the decrease in ecological estimates of population size. Taken together, these results indicate a departure from conditions of equilibrium (closed population without selection) for M. montanus. Our results have the following possible explanations: (1) the sampling area for fossils changed, (2) the local population size expanded, (3) selection occurred, and/or (4) gene flow occurred.Our results demonstrate different genetic responses by two species of small mammals to changes in population size driven by climatic change. Fossil abundance data reveal population decline for both m Period . For T. Microtus oeconomus, indicate that migration occurs most frequently in and between low-density patches  and between populations (gene flow). Discrepancy between the ecological and genetic estimates of population size and significant changes in haplotypic diversity prior to the Medieval Warm Period implicate increased gene flow into the Lamar Cave M. montanus population. Additionally, the observed haplotypic turnover in the Yellowstone population during this period suggests that as abundance of M. montanus declined through the last 845 years, relatively more individuals carried newly introduced haplotypes.Here we demonstrate, using a phylochronologic approach, that it is possible to distinguish the dynamic processes that govern gene diversity over relatively short time scales (hundreds to thousands of years). We have documented environmental change, population response, genetic diversity change, and the correlations between the three. Without serial data, we would capture just a single record of these historic processes: modern genetic diversity. Although it is possible to hypothesize about historic events using modern data, phylochronology affords a unique look into the past and the potential ability to separate cause from effect. In particular, we show that M. montanus was maintained. In contrast, gene flow has not played a significant role in the recent genetic history of T. talpoides. This species, instead, responded more as a closed population over this time. The disparate nature of population response to climatic change of these two species is likely due to differences in demographic dispersal patterns between their populations. Such differences in species demography have resulted in differential genetic response to climatic change, even when ecological response is similar. Thus, genetic response to environmental change can be viewed as “individualistic,” similar to unique adjustments of species ranges  declined while others expanded. Our data show that even with a prolonged ecological population size decline, the genetic diversity of s ranges . Life hin = 8,589]) and are concordant with expectations of taxonomic diversity in montane mammal communities of western North America .Lamar Cave contains well-stratified, thoroughly radiocarbon-dated deposits, which display high fidelity to the local mammalian community , 1999. T America . The cumMicrotus samples encompass 15 of 16 radiocarbon-dated stratigraphic levels from Lamar Cave N and e-mm_ecol (M. montanus).NIn order to compare ecological estimates of effective population size to genetic estimates, we estimated mitochondrial effective population size assuming that Microtus species are cryptic with respect to these teeth. Genetic diagnosis indicated that these 78 fossil samples are derived from multiple arvicoline species, with a total of 47 M. montanus specimens (see Microtus molariform teeth) ranged from 3.5 to 15.3 mg . We used two previously described extraction methods on the fossil teeth (b from ancient samples (n = 47) using the following primers: forward primers (5′–3′), CLETH 37 TAY AAY ATA ATY GAA ACH TGA A , CLETH 37L AYG GMT CTT AYA ACA TAA TCG AAA CAT G , MMONT 1 CAG TAA TTA CAA AYC TWC TAT CA , and MMONT 3 AGT GAA TCT GAG GGG GCT TCT CAG TAG A ; reverse primers (5′–3′), ARVIC 08 CAG ATY CAY TCY ACT AGT GTT G , ARVIC 08L CTC AGA TTC ACT CTA CTA GTG TTG TG , MMONT 4 TTR TTT GAT CCT GTT TCG TGT AGG AAT A , and MMONT 2L TTG ACT GTG TAG TAA GGG TGA AAT GGG A . Attempts were made to amplify the region in two overlapping fragments using CLETH37/ARVIC 08, CLETH37L/ARVIC 08L, and MMONT 1/MMONT 2L. However, the low rate of success for MMONT 1/MMONT 2L (40%) in the fossil samples necessitated the breaking of the second fragment into two overlapping fragments using MMONT 1/MMONT 4 and MMONT 3/MMONT 2L.From the fossil material, we used the upper first molar (from only one side of the jaw per level) to avoid sampling the same individual multiple times. Some mens see . Fossil il teeth . We obtaAncient DNA samples were run on an ABI PRISM 310 Genetic Analyzer in the post-PCR lab, and modern DNA samples were run on an ABI PRISM 377 Sequencer in a separate sequencing facility . Fragments were sequenced in both directions, primer regions were overlapped, and sequences with any ambiguous sites were rerun until completely resolved in order to provide additional corroboration and eliminate ambiguity.We adhered to strict extraction and amplification protocols . The pro and 12 teeth  from the vicinity (within 10 km) of the fossil site. In addition, DNA was successfully extracted from 51 modern specimens (liver tissue and museum skins) collected within a 400-km radius of Lamar Cave. Of these 51 samples, 22 were derived from museum skins  and 29 from liver tissue  of specimens trapped in the field. We followed the animal tissue protocol using the Qiagen Dneasy Tissue Kit  on 6.5 mg of liver samples and 7.5 mg of tooth samples (n = 3). For museum samples, DNA was extracted from the ventral skin incision . We amplified the entire cytochrome b gene  for some modern skins (n = 13) and liver tissue (n = 29) with the following primers: forward primers (5′–3′), MVZ 05 CGA AGC TTG ATA TGA AAA ACC ATC GTT  and ARVIC 07 AAA GCC ACC CTC ACA CGA TT ; reverse primers (5′–3′) MICRO 06 GGA TTA TTT GAT CCT GTT TCG T  and VOLE 14 TTT CAT TAC TGG TTT ACA AGA C .Modern  genetic sampling was obtained from a variety of sources including modern skins, modern liver tissue, museum skins, and teeth derived from modern raptor pellets. DNA was successfully extracted from 16 modern skins Microtus samples, and 79 of those yielded successful amplification. Of these 79 samples, 48 were positively identified as M. montanus. Of the M. montanus samples, 41 specimens were successfully haplotyped. Sequences have been deposited in GenBank  , which is not found elsewhere in a 400-km radius around Lamar Cave. This constancy in the genetic lineage of T. talpoides within a single locality persists in spite of climatic changes and concurrent significant population and body size changes (We have built upon the previously published data set  with add= 3) see . T. talp changes .e) was used to estimate e. θSN was preferred over other estimators of θ since Hθ is biased for single locus estimates, kθ does not incorporate sequence information, and πθ has higher variance. These estimates are determined entirely from genetic data; thus we label them e-tt_gen N and e-mm_gen (M. montanus).N We assumed a range of μ   using Cody's index (β-diversity each haplotype was treated as a “species.”Arlequin v. 2.000  was used per bp) . Additio's index . For β-dN, a gene with mutation rate μ, and an infinite alleles model given a sample size n. Here we use the Ewens distribution to ascertain the ability of our data to detect variation in values of gene diversity. In addition, this distribution allows us to (1) predict the distribution of expected gene diversity at each point in time (independently) given estimates of e_ecol,N a moderate mutation rate of 4% per million years, typed sequence length , and sample size  and (2) predict change in the expected gene diversity distribution for a large sample size of 100. A modified version of MONTE CARLO , and gene diversity was calculated for each configuration. Simulations were repeated 1,000 times for each sampling time point, resulting in the 95% confidence intervals for the distribution of predicted gene diversities given a closed, selectively neutral population through time.Given the general limitations of obtaining aDNA sequence data, sample sizes will always present challenges to ancient population genetic studies. To investigate whether our samples are adequate for addressing temporal gene diversity in both species, we used a neutral population model to evaluate expected diversity. TE CARLO , 1996 waM. montanus evolution during the past 2,525 years. Simulations were repeated 1,000 times to generate genetic data and changes in gene diversity for the null hypothesis. The significance of the observed changes was then inferred by comparing it to the generated null values (we thus used a Monte Carlo significance test). Estimates of e_ecolN were used to set up a null hypothesis specifying population size change through time in a closed population. We assumed that past changes in population size between intervals were due to exponential growth or decline. The effective population size at two time points and the time between the points was used to calculate a growth rate. The estimated growth rates were 1438–2525ybpr = −0.000178, 845–1438ybpr = 0.000732, 470–845ybpr = 0.0004385, and 166–470ybpr = 0.0003212. The null hypothesis thus corresponds to effective population sizes at particular points in the past and exponential growth or decline between those intervals, and represents an ecologically realistic description of the past 2,525 years. We assumed a constant population size prior to 2,525 ybp, as we have no data before this point in time. The coalescent program SIMCOAL , we investigated the significance of observed changes in gene diversity at all nine parameter combinations over the entire time span  and between 2,525 and 845 ybp .We assessed the significance of the observed changes in gene diversity between time points using simulations. The serial coalescent was used as a framework to model  SIMCOAL  was modialescent  has beenAdditionally, we also investigated the relevance of temporal data to our ability to reject the null hypothesis. Significance tests were repeated assuming the observed data were from only the most recent genetic samples. Again, simulations were repeated for all nine combinations of mutation rate and density estimates.M. montanus specimens described in Sequences for the successfully haplotyped


Why does a population in the wild grow or shrink, or remain the same over generations? Genes would seem an obvious factor, but in fact there is remarkably little evidence that genetic effects influence year-to-year population dynamics, beyond the well-recognized negative effect from inbreeding on very small populations.This lack of evidence for a genetic effect partly reflects the difficulty of choosing which gene to study, as only a few genes may be under strong selection at any given time, and fewer still will affect population dynamics. It also partly reflects an important truth about natural selection—it may determine who survives to reproduce, but not necessarily the total number of survivors. In many cases, ecological factors such as resource abundance and natural enemies may overwhelm any slight genetic effect on population dynamics.Finding a genetic effect, then, would require both knowing which pin in the genetic haystack to look for and having detailed knowledge of the complex and powerful environmental factors against which this effect plays out. In a new study, Ilkka Hanski and Ilik Saccheri show that variants of a sugar-metabolizing gene do indeed influence population growth in a species of butterfly, but in a complex and habitat-dependent way.(Pgi), a key enzyme in the breakdown of sugar. ThePgi gene occurs in several forms, or alleles, whose proteins differ in their kinetic properties and thermal stability. These alleles have been previously linked to differences in flight metabolic rate and fecundity in this butterfly, making the gene a good candidate for observing a population effect, if there is one. Thef andd alleles ofPgi are the most common, and previous work has shown that butterflies with either anff or anfd genotype have a higher flight metabolic rate and are more fecund than those with add genotype.The authors studied the Glanville fritillary butterfly on the Åland Islands in Finland, where its population dynamics are well studied, and its habitat—patches of dry meadows spread across the landscape—is well mapped. They focused on the gene phosphoglucose isomeraseff orfd genotypes predominated, but in larger meadows, the opposite was true—these genotypes predicted a decline in numbers instead of a rise, whiledd was favored. The effect appeared to be specific toPgi, as there was no correlation with genotype for any of the six other genes.By analyzing genotypes, population growth, and habitat area simultaneously among more than 130 small butterfly populations, the authors showed that, in small meadows, growth was highest when thef andd alleles. Those withf alleles mature quickly and lay more eggs early on, just the strategy for exploiting a small patch, from which many butterflies risk drifting away rather quickly in their life. Those withd alleles mature later but also die later, allowing them to exploit a larger habitat more thoroughly. However, the authors note that this may not be the only, or even the main, reason for the genotype-habitat area effect, sincePgi is likely to influence many different aspects of life history.The likely explanation for this effect, according to the authors, is related to the differences in maturation and egg laying between females bearingThe results of this study confirm that, under the right circumstances, intraspecific genetic variation can influence population growth. But they also make an important point about fitness. A major goal of evolutionary physiology is to understand the selective advantage of the traits found in a population, and it is often tempting in this pursuit to assume there is a single “best” genotype. This study provides a strong counterargument against such one-size-fits-all models of evolution, pointing out that in the ecological theater, which script plays best is a function of exactly which stage you are on.


About 14,000 years ago—a few hundred thousand years after our putative modern forebears spread out from Africa—descendants of archaic humans crossed the Bering land bridge from Siberia to North America. Several lines of evidence support this model, but that's where the consensus ends. The details remain hotly debated, focusing mostly on which Asian population migrated, when they did it, and whether they did it more than once.Part of the challenge in reconstructing this history stems from the dynamic nature of human populations—which experience unpredictable changes in size, composition, density, and mating patterns—and the difficulty in interpreting genetic history. To get a better picture of the range of possible scenarios, scientists are using new statistical approaches that require computer simulations. Jody Hey now extends this approach in a novel method for the study of the origins of New World populations. Along with DNA analysis and computer simulations, Hey adds a new twist to an old model to reveal how the sizes of the first New World populations have changed since they were founded. His results fall in line with archeological, genetic, and linguistic evidence, pointing to a relatively recent colonization of the Americas. But they go a step further by showing that the New World was colonized by a small population with an effective size of only about 70 individuals. Hey's approach addresses shortcomings in population genetic studies that rely on just one gene and that assume that population sizes have been constant over time. Studying levels of DNA sequence variation at a single genomic region, or loci, can offer insight into the history of that gene, but the stochastic nature of gene evolution means that different genes have different histories. And as simplified versions of a very complex reality, population genetics models, such as the “isolation with migration” (IM) model, that aim to capture the population dynamics during the early stages of divergence or speciation have necessary limitations. The widely used IM model, for example, assumes that a founding population splits into two descendant populations that may interbreed, and incorporates a large number of parameters. But until now the IM model has required the assumption that all of the populations were constant in size, and therefore it has not been useful for assessing how descendant populations arose or changed in size.Hey analyzed DNA sequences from nine loci, so that the population genetic history could be found despite the variation among genes. He also added an additional parameter to the standard IM model to incorporate changes in the size of the ancestral population and of each founder population through time. The genetic data included DNA sequences from Asian and American Indian individuals. Hey varied the parameters in his model, which included founding population size, changes in population size, time of population formation (and splitting), and gene exchange between the populations, to work out the demographic scenario that best fit the available genetic data.His analysis suggests that only about 70 individuals left their ancestral Asian population, estimated at about 9,000 individuals, to reach America 7,000 to 14,000 years ago. Archeological evidence places the earliest American inhabitants in the New World at around 14,000 years ago. Though Hey's estimates are more recent, they also indicate a high probability at this time period. Hey did not include genetic data from Eskimo-Aleut and Na Déne speakers, so the number of migrations was not addressed. But with this new approach, researchers will be able to explore this and many other questions to fill in the details of the first American immigration.


Ambystoma tigrinum) for evidence of population-level changes in body size and paedomorphosis to climatic change over the last 3000 years.Amphibians are sensitive indicators of environmental conditions and show measurable responses, such as changes in phenology, abundance and range limits to local changes in precipitation and temperature regimes. Amphibians offer unique opportunities to study the important ecological and evolutionary implications of responses in life history characteristics to climatic change. We analyzed a late-Holocene fossil record of the Tiger Salamander (We found a significant difference in body size index between paedomorphic and metamorphic individuals during the time interval dominated by the Medieval Warm Period. There is a consistent ratio of paedomorphic to metamorphic specimens through the entire 3000 years, demonstrating that not all life history characteristics of the population were significantly altered by changes in climate on this timescale.Ambystoma tigrinum we used spans an ecologically relevant timescale appropriate for understanding population and community response to projected climatic change. The population-level responses we documented are concordant with expectations based on modern environmental studies, and yield insight into population-level patterns across hundreds of generations, especially the independence of different life history characteristics. These conclusions lead us to offer general predictions about the future response of this species based on likely scenarios of climatic warming in the Rocky Mountain region.The fossil record of  The global mean temperature has risen 0.6°C in the last 100 years and is projected to increase between 1.4°C and 5.8°C over the next 100 years -4. SeverAmphibians are often considered sensitive indicators of environmental conditions -16. ManyLamar Cave is an exceptionally rich paleontological site located in Yellowstone National Park (YNP), Wyoming, USA . The fosAmbystoma tigrinum) is an excellent species with which to study population level responses to climatic change. Many studies show that Ambystoma demonstrates developmental growth plasticity in response to environmental conditions : fos: fosAmbynditions ,28-31. Atic form ,33 of poA. tigrinum depends on various environmental stimuli [Paedomorphosis in modern  stimuli ,34. Seve stimuli ,31,35,36 stimuli . Populat stimuli ,29,37,38A. tigrinum, we used a late-Holocene fossil record to track these traits through the last 3000 years. We analyzed trends within the context of known climatic change, and attempted to distinguish patterns of response that represent specific climatic periods through this time.Given the environmental forces that drive body size and metamorphic changes in Ambystoma specimens from the excavation was 2850. The total minimum number of individuals (MNI) from the excavation was determined to be 99. The abundance, represented by MNI, is relatively low in Intervals A, B, D, and E. Interval C, which corresponds to the bulk of the MWP, contains 66% of the Ambystoma fossils, while the intervals before (D and E) and after the MWP (A and B) all contain lower A. tigrinum abundance based on MNI  measurements for the paedomorphs and the terrestrial adults do not show significant changes through time within an age class . However, the data do show variability through time that may be a measure of changes in the population, even if it is not a statistically significant deviation. To identify potentially biologically relevant trends between age class BSI's and known climatic periods, the terrestrial adult BSI was compared to the paedomorph BSI. There is a significant difference between the paedomorphic and terrestrial adults when analyzed by interval . Comparisons within intervals indicate a significant difference in BSI between paedomorphic and terrestrial adults within interval C  within the sampled population over a relatively long ecological time period, it is likely that the two distinct life history strategies were maintained during short-term climatic fluctuations of the late Holocene. Thus, we conclude according to this case study that although change in frequency of paedomorphosis in a population occurs in relation to different environmental conditions over one generation, on the time scale of 100 to 1000s of years, if there is short-term climate variability little overall change would be expected in a population, even with coarsely distinguished climatic events.An assumption made in interpreting the fossil record is that the specimens are representative of the sample population. Another caveat is that the common characteristics of the population also will occur in the fossil record if there are no effective sampling biases . With thA. tigrinum responded to the largest climatic anomaly in the Yellowstone region over the last 3000 years, the Medieval Warm Period (MWP) [A. tigrinum fossil abundance over the last 3,000 years shows a marked increase during Interval C, which corresponds to the MWP. Due to the deposition of this fossil assembly, fossil abundances reflect an overall increase in salamander abundance, an increase in successful predation and collection, or a combination of both factors. However, prior study on taphonomic biases potentially at work in this cave accumulation find no evidence for a change in predators, a change in collection radius, or an adjustment in collection vectors [Our results provide an opportunity to determine how the local population of od (MWP) . The MWPod (MWP)  and occuod (MWP) . These c vectors .A. tigrinum populations during the warming based on modern studies [The periods of higher effective moisture may provide deeper, more permanent ponds that would offer greater protection from aerial predators, but we cannot conclude that such a sampling bias is the only factor affecting fossil abundance. The significant difference in BSI between paedomorphic and terrestrial adults during interval C, along with the increase in abundance, is evidence for warm climatic conditions that allows indeterminate growth of a terrestrial ectotherm to continue. This agrees with expected changes in  studies ,42.A. tigrinum can show changes in average body size without changes in the percentage of different morphologies. In the oldest intervals, E and D, paedomorphs appear to have been smaller (based on BSI) than in other intervals, while the percent paedomorphic remained relatively constant. This suggests a favorable terrestrial environment where metamorphosing individuals continue to grow following the transition from the aquatic environment. This terrestrial growth will be most facilitated by warm conditions and abundant food. The available climate records show these relatively long time periods as variable, with periods of high and low effective moisture [Our results also demonstrate that a population of moisture -45.14C chronology, exacerbated time-averaging [Ambystoma metamorphosis, yet we cannot dismiss the role of certain biotic elements. Specifically, density-dependent effects are correlated with body size and metamorphosis, which may skew our interpretations. Larval density will impact resource competition, with higher competition driving metamorphosis. Patterns in larval individuals have not been documented here because of small sample sizes. Finally, likelihood of paedomorphosis may have a significant genetic control, which we do not discount here. Our results support the maintenance of alternate, but coexisting, life history strategies suited to selection pressures in variable environments.There are a variety of confounding factors that complicate our interpretations. First, although the excavation was remarkably rich and the number of identified salamander specimens quite large, our conservative use of MNI reduced our sample size. Our use of time intervals (A-E), while justified based on our own veraging . In addiAmbystoma fossils from Lamar Cave provide a unique opportunity to examine the important ecological and evolutionary implications of changes in life history characteristics in response to climatic change. We present a record of change in a population of tiger salamanders over the last 3000 years. Significant perturbation in the environment is possible under future climate change scenarios, and the role of local populations in the terrestrial or aquatic system may be altered. Warmer and drier climate scenarios as predicted for the Yellowstone region [Ambystoma food sources. Such changes to the ecological system could result in unexpected biological feedbacks. Also, higher percentages and rates of metamorphosis would increase gene flow between populations. This, combined with probable decreases in sizes of populations, has the potential to alter the overall genetic diversity in the meta-population over time, perhaps reducing the ability of the species to respond to further perturbation of the system.The late Holocene e region  would liThe Greater Yellowstone Ecosystem (GYE) is often considered one of the last intact, temperate ecosystems in the world. This ecosystem contains all native mammals and few exotics, and is thought to be functioning in a relatively natural state . The GYEA. tigrinum fossils used in this analysis were excavated from Lamar Cave, a paleontological site in YNP. The details of the excavation and stratigraphy are described elsewhere [A. tigrinum. The A. tigrinum samples are most likely from predation in these ponds and surrounding lands. The current study analyzes fossils obtained from 15 of the 16 stratigraphic levels from the excavation (level 11 did not contain any Ambystoma specimens). For the analyses the levels were pooled into five intervals, labeled A-E (youngest to oldest). This aggregation was based on 95% confidence limits around the radiocarbon dating of the intervals [The lsewhere . Isotopilsewhere . Within ntervals .A. tigrinum fossils include femora, humeri, vertebrae, and various skull bones. We used the fossil vertebrae because of their abundance (N = 2850) and because they record metamorphic state. All vertebrae were identified, but for the purposes of this study only the first cervical and sacral vertebrae were used. Because these particular vertebrae are unique to every skeleton, they are useful in determining the minimum number of individuals from a locality [Easily identified locality . The foslocality ,33. SpecAmbystoma skeleton contains only one of each of these elements. The abundance levels were then standardized by dividing by the MNI of the wood rat, Neotoma cinerea. Unlike other common small mammals found at this site, wood rats show a constant relative abundance [Abundance was determined by a standardized minimum number of individuals (MNI) . The MNIbundance . This pabundance , which ibundance .Because plasticity in growth rate cannot be directly measured in the fossil record, it is inferred from body size in different age classes. The centrum length and anterior width of each specimen were measured with electronic calipers. A body size index (BSI) was created for each specimen by dividing the centrum length by the anterior centrum width .Percent paedomorphosis by time interval was determined by dividing the standardized MNI of paedomorphic vertebrae by the standardized MNI of all adult vertebrae, defined as Old Larval, Young terrestrial, and Old Terrestrial morphs. Thus, we calculate abundance, mean body size, and percent paedomorphosis, each as potentially independent responses of the salamander population to the abiotic environment around Lamar Cave.EAH designed the study and collected the specimens. WL collected data. JEB analyzed data.


We show that the allelic composition of the glycolytic enzyme phosphoglucose isomerase(Pgi) has a significant effect on the growth of local populations, consistent with previously reported effects of allelic variation on flight metabolic performance and fecundity in the Glanville fritillary andColias butterflies. The strength and the sign of the molecular effect on population growth are sensitive to the ecological context , which affects genotype-specific gene flow and the influence of migration on the dynamics of local populations. The biological significance of the results forPgi is underscored by lack of any association between population growth and allelic variation at six other loci typed in the same material. In demonstrating, to our knowledge for the first time, that molecular variation in a candidate gene affects population growth, this study challenges the perception that differential performance of individual genotypes, leading to differential fitness, is irrelevant to population dynamics. These results also demonstrate that the spatial configuration of habitat and spatial dynamics of populations contribute to maintenance ofPgi polymorphism in this species.The dynamics of natural populations are thought to be dominated by demographic and environmental processes with little influence of intraspecific genetic variation and natural selection, apart from inbreeding depression possibly reducing population growth in small populations. Here we analyse hundreds of well-characterised local populations in a large metapopulation of the Glanville fritillary butterfly Pgi locus (an enzyme linked with flight metabolic performance) and local population growth within a metapopulation of butterflies.Researchers observe a link between the genetic composition of the The notion that a population's genetic composition influences its ecological dynamics is conceptually deeply rooted in population biology –3, but For gene, which encodes the cGMP-dependent protein kinase, influence foraging behaviour inDrosophila melanogaster larvae and adults. The analogous gene in honeybees influences the regulation of division of labour between forager and nurse bees  for populations with at least six copies ofPgi typed in the autumn 1995 sample and hence a minimum sample size of three larvae from three larval families (hence minimumtN was 3). The material included 139 such populations, but we lacked information for eight populations in 1996 and hence these populations had to be omitted. We added “1” to population size to allow the inclusion of populations that went extinct by 1996 (n = 34). Regional change in population sizes in the surroundings of the focal populationi is defined asi,trendN =i,tS1+/i,tS [i,*S is given byProportional change in population size  was calculated as/Si,t , whereSPgi-f allele in small and large habitat patches using the following results and assumptions. Among the 139 populations in 1995, the average sizes of “small” and “large” populations were 0.10 and 0.89 ha, respectively, using the limit ofA = 0.3 ha between the two size classes  gave the following daily probabilities of laying subsequent clutches, and the average clutch sizes: et al. ; hence wf and non-f females):f females before and after day 14, 0.05 and 0.3; non-f females before and after day 14, 0.05 and 0.15.In the experiment described by Hanski et al. , daily mL =f 0.2,Sf = 0.4, non-L =f 0.05, non-Sf = 0.4. The justifications are as follows. Hanski et al. [f females than old populations [f and non-f females, but it is likely that all butterflies have high emigration rate from very small patches (see [Proclossiana eunomia).Butterflies were assumed to stay in the natal patch until they became mated and had laid their first clutch, after which they emigrated with the following daily probabilities: et al.  found dalations . We do nes , which Table S1Click here for additional data file.Table S2Click here for additional data file.Table S3Click here for additional data file.


Humans' capacity to alter the environment has grown exponentially over the past century, challenging other species to adapt to a transfigured landscape. Over some 15,000 plants and animals face extinction, the overwhelming majority threatened by increasingly fragmented, polluted, and deteriorating habitats. These forces are most acute in Africa and Asia, where 25% of all primate species are threatened. Chimpanzees, bonobos, orang-utans, and gorillas could vanish from the wild completely within a matter of decades.In Africa, deforestation, illegal hunting, and disease outbreaks are decimating chimp, bonobo, and gorilla populations. Orang-utans, the only non-African great ape, face similar threats, compounded by agricultural conversion of logged forests. But because orang-utans spend most of their time in trees and avoid open spaces, they may suffer most from deforestation. A new study by Benoit Goossens, Lounès Chikhi, Michael Bruford, and their colleagues shows that the case for orang-utans is in fact even more desperate than previously believed. Using computer simulations and the largest genetic dataset ever collected from wild orangutans, the authors build up a picture of alarming—and very recent—declines of orang-utan populations of over 95%.Orang-utans, Malaysian for “man of the forest,” once ranged from Indonesia to southern China and northeastern India, but now struggle to survive in isolated forest patches in northern Sumatra and Borneo. Over 66% of the Sumatra orang-utan population was killed between 1993 and 2000; 33% of the Borneo population may have succumbed to drought and fire between 1996 and 1997 alone. When a 1987 study estimated that over 20,000 orang-utans persisted in Sabah, Malaysia, in the 1980s, it was challenged as too optimistic.Developing effective conservation and recovery programs depends on determining when the decline began, its trajectory, and the original population size. To shed light on these questions, Goossens and Chikhi used three statistical methods to mine the genetic material of wild orang-utans in Eastern Sabah, scarred by over 50 years of large-scale commercial logging and agriculture.For the genetic analysis, the authors and their local assistants collected hair from tree nests—some nearly 100 feet above the ground—and feces found under nests or near orang-utans encountered along the Kinabatangan River. Two hundred orang-utans were identified using genetic markers called microsatellites, tandem repeats of short DNA motifs that are sometimes used as genetic fingerprints. The authors used 14 microsatellites from each animal in an analysis that compares observed genetic diversity with that expected for a stable population, based on the number of alleles  and expected heterozygosity .When population size drops significantly, genetic diversity decreases as well, but in a very specific way: rare alleles are lost whereas heterozygosity remains relatively little affected. And it is this signature that the authors were looking for. The authors first simulated what the expected heterozygosity would be, based on the number of alleles and sample size (nine forest blocks were sampled) for each population and microsatellite locus, and compared these values to those seen in the collected genetic data. Three different mutation models were used to simulate variations of microsatellite repeat number under different mutational processes and demographic histories to detect evidence of a decline. Even the most conservative model produced the same “strong and significant signal for a population bottleneck.”They next used two computer-intensive statistical approaches to characterize the extent and timing of the decline, focusing on the two largest populations. The first statistical model  makes specific demographic and mutation assumptions that can quantify the relative change in population size. Again, this model yielded the same result: orang-utan populations have declined by at least a factor of 50 (with 95% probability) or 100 (with 90% probability).The second model (the Storz and Beaumont method) allowed the authors to estimate past and present population sizes and timing of the collapse. They found strong genetic evidence of a collapse for both Sabah populations, and a likely time frame within the past decades—coinciding with accelerated deforestation in the region in the 1950s and 1970s. This result also supports the 1987 estimate of 20,000 and the current estimate of 11,000 Sabah orang-utans—which means the decline was far more recent, and far more drastic, than previously assumed.Normally, the genetic effects of such recent events would be obscured by ancient demographic events, such as those following major climatic changes, or the arrival of the first hunter-gatherers or farmers. But humans have so devastated orang-utan populations that the genetic signature of the recent bottleneck may have overshadowed any previous population fluctuations. These results demonstrate that genetic data can provide independent and extremely detailed information by detecting and quantifying the consequences of human impacts on endangered species. They also underscore the need to act now to protect the long-term survival of the species. The animals show enough genetic diversity to stabilize, the authors argue, if immediate steps are taken to reconnect remnant forest patches and halt further deforestation. Otherwise, humans will have restricted the fabled forest man to the realm of memory—or a life behind bars in a zoo.


Understanding the role of seascape in shaping genetic and demographic population structure is highly challenging for marine pelagic species such as cetaceans for which there is generally little evidence of what could effectively restrict their dispersal. In the present work, we applied a combination of recent individual-based landscape genetic approaches to investigate the population genetic structure of a highly mobile extensive range cetacean, the harbour porpoise in the eastern North Atlantic, with regards to oceanographic characteristics that could constrain its dispersal.Analyses of 10 microsatellite loci for 752 individuals revealed that most of the sampled range in the eastern North Atlantic behaves as a 'continuous' population that widely extends over thousands of kilometres with significant isolation by distance (IBD). However, strong barriers to gene flow were detected in the south-eastern part of the range. These barriers coincided with profound changes in environmental characteristics and isolated, on a relatively small scale, porpoises from Iberian waters and on a larger scale porpoises from the Black Sea.The presence of these barriers to gene flow that coincide with profound changes in oceanographic features, together with the spatial variation in IBD strength, provide for the first time strong evidence that physical processes have a major impact on the demographic and genetic structure of a cetacean. This genetic pattern further suggests habitat-related fragmentation of the porpoise range that is likely to intensify with predicted surface ocean warming. In the marine realm, pelagic species that have large geographic range and high dispersal capabilities represent a serious challenge to the idea of allopatric divergence  and to speciation processes in a seemingly continuous environment . The higCetaceans are good examples of this kind of species. Despite their broad range and their high dispersal capabilities, many cetaceans often show substantial genetic structure at regional or even fine scale, although the extent varies among species . It is gPhocoena phocoena (L. 1758), with regards to seascape characteristics. Harbour porpoises occur fairly continuously throughout cold coastal waters of the North Pacific and the North Atlantic, with a relict population in the Black Sea separated from the Atlantic range by the Mediterranean Sea where porpoises are nowadays absent . We conducted a series of independent runs with different proposals for K, testing all values from 1 to 10. Each runs used 106 iterations after a burn-in of length 4 × 104, testing different models: (a) with or without admixture, and (b) correlated or uncorrelated allele frequencies. To check for convergence of the Markov chain Monte Carlo (MCMC), we performed 10 replicates for each value of K and then checked the consistency of results. The estimated number of clusters (K) was taken to be the value of K with the highest Pr(X|K)  are below the diagonal and the significance level of the exact tests for population differentiation [ntiation  are abovClick here for file


Speciation often occurs in complex or uncertain temporal and spatial contexts. Processes such as reinforcement, allopatric divergence, and assortative mating can proceed at different rates and with different strengths as populations diverge. The Central American Midas cichlid fish species complex is an important case study for understanding the processes of speciation. Previous analyses have demonstrated that allopatric processes led to species formation among the lakes of Nicaragua as well as sympatric speciation that is occurring within at least one crater lake. However, since speciation is an ongoing process and sampling genetic diversity of such lineages can be biased by collection scheme or random factors, it is important to evaluate the robustness of conclusions drawn on individual time samples.In order to assess the validity and reliability of inferences based on different genetic samples, we have analyzed fish from several lakes in Nicaragua sampled at three different times over 16 years. In addition, this time series allows us to analyze the population genetic changes that have occurred between lakes, where allopatric speciation has operated, as well as between different species within lakes, some of which have originated by sympatric speciation. Focusing on commonly used genetic markers, we have analyzed both DNA sequences from the complete mitochondrial control region as well as nuclear DNA variation at ten microsatellite loci from these populations, sampled thrice in a 16 year time period, to develop a robust estimate of the population genetic history of these diversifying lineages.The conclusions from previous work are well supported by our comprehensive analysis. In particular, we find that the genetic diversity of derived crater lake populations is lower than that of the source population regardless of when and how each population was sampled. Furthermore, changes in various estimates of genetic diversity within lakes are minimal and provide no evidence for drastic changes during the last 20 years, supporting the hypothesis that the processes which have resulted in rapid speciation are primarily historical. In contrast, there is some evidence for ongoing evolution, particularly selection, in all lakes except crater Lake Masaya, perhaps reflecting the persistence of speciational processes. Importantly, we find that the crater Lake Apoyo population, for which strong evidence of sympatric speciation has been demonstrated, has lower genetic diversity than other crater lakes and the strongest evidence for ongoing selection. Over the past several years, largely due to the development of sensitive molecular markers, our knowledge of the factors involved in speciation has grown tremendously. Research into the life history, ecological, and genetic characteristics of speciating lineages has enriched the understanding of the underlying processes that generate diversity ,2. NeverAmphilophus citrinellus, A. labiatus, and A. zaliosus, are distributed in the lakes of western Nicaragua . This mrinellus -18.Oreochromis niloticus), was introduced into Lake Apoyo in the late 1980's as a foodstuff .EF219198–EF219272]. The 145 sequences from 2001 represent 48 unique haplotypes . The 327 sequences from 2003 represent 91 haplotypes . Ten unlinked , the , the 5]]Testing population genetic hypotheses using multiple sampling periods should increase statistical power through multiple independent tests, as well as allow the testing of the hypothesis that the samples being measured are representative of ongoing evolutionary processes ,43. BecaIt should be noted that despite significant differences in haplotype diversity and nucleotide diversity discovered in Lake Apoyo in the three time samples, no trends were apparent that indicate the introduction of tilapia has had an impact on genetic diversity. Allelic richness also stayed consistent for more than 10 years since the introduction of tilapia into this small crater lake. This may be indicative of low or no competition between species  or tEvidence for ongoing evolutionary change within lakes is indicated by the values of Fu and Li's  D* and FLake Apoyo shows the strongest trends associated with both selection and low genetic diversity. It is perhaps not unexpected, then, that the endemic cichlids of this lake also present one of the strongest cases for sympatric speciation yet detected in animals ,18,46. RA. zaliosus from a founding population of A. citrinellus from Lake Nicaragua may be the clear result of strong diversifying pressures including selection and drift, during the short existence of the population [Wilson et al.  speculatpulation . Though pulation ,24, may pulation .F-statistics demonstrating more divergence of the Lake Apoyo and Lake Xiloa populations than of those in Lake Masaya represent either the divergence associated with the uniqueness of the founding individuals, different mechanisms of gene flow between the lakes , or environmental factors such as variation in the frequency and magnitude of volcanic activity, that is unique to each of the crater lakes.The colonization of new habitats by small, genetically homogenous founding populations is likely to promote speciation through both sympatric and allopatric mechanisms . A corolIt is difficult to distinguish between demographic factors, such as population size and migration rate, from selective pressures that result in evolutionary change. But it is the combination of these factors that is most relevant to studies of speciation as they can influence the strength and direction of each other in striking ways. These results do not detect any meaningful trends in genetic diversity during a 16-year period. As a conclusion, the evolutionary forces driving speciation in places such as Lake Apoyo are persistent and not likely to be heavily influenced by demographic changes over a few (~20) generations. This persistence of evolutionary forces is apparent despite ongoing anthropogenic pressures on the lakes, including increased fishing and the introduction of non-native competitors , as wellImportantly for researchers employing population genetic analyses for the inference of evolutionary process, these results indicate that variation in sample collection design do not necessarily bias the conclusions drawn on limited data. Further investigation of what aspects of sampling design and field collection of data, including sampling the individuals as well as the particular genetic markers used, will help clarify the best collection strategy for guaranteeing robust inferences of evolutionary processes from population genetic data.PMEB helped collect the molecular data, designed and performed the analyses, and drafted the manuscript. MB participated in sample collection and helped collect the molecular data. AM conceived of the study, participated in sample collection, and helped draft the manuscript. All authors have read and approved of the final manuscript.


GM and KM immunoglobulin allotypes in the Galician population and, thus, provides further insights into the extent of genetic diversity in populations settled in the geographic extremes of the Cantabrian region of northern Spain. Furthermore, the genetic relationships of Galicians with other European populations have been investigated.The current genetic structure of Iberian populations has presumably been affected by the complex orography of its territory, the different people and civilizations that settled there, its ancient and complex history, the diverse and persistent sociocultural patterns in its different regions, and also by the effects of the Iberian Peninsula representing a refugium area after the last glacial maximum. This paper presents the first data on GM haplotypes that is defined by the high presence of the European Mediterranean GM*3 23 5* haplotype, and the relatively high incidence of the African marker GM*1,17 23' 5*. Data based on comparisons between Galician and other Spanish populations (mainly from the north of the peninsula) reveal a poor correlation between geographic and genetic distances , a noticeable but variable genetic distances between Galician and Basque subpopulations, and a rather close genetic affinity between Galicia and Valencia, populations which are geographically separated by a long distance and have quite dissimilar cultures and histories. Interestingly, Galicia occupies a central position in the European genetic map, despite being geographically placed at one extreme of the European continent, while displaying a close genetic proximity to Portugal, a finding that is consistent with their shared histories over centuries.Galician population shows a genetic profile for These findings suggest that the population of Galicia is the result of a relatively balanced mixture of European populations or of the ancestral populations that gave rise to them. This would support the importance of the migratory movements that have taken place in Europe over the course of recent human history and their effects on the European genetic landscape. It forms part of the Cantabrian mountain range, which extends along the coast of the Cantabrian Sea from Cape Finisterre to the western limit of the Pyrenees. The territory is bordered to the north and west by the Atlantic Ocean, to the south by Portugal and to the east by the regions of Asturias and Leon , as well as a new political organization and restructuring of the population through two important cities, Braga  and Lugo . It should be noted that the main route of communication in the north of the Peninsula and in an east-west direction occurred through the flat areas of the central plateau and not through the mountainous costal areas, which were accessed by branches at various points leading off the main route. Thus, this communication system maintained a certain degree of isolation between nearby coastal populations.Romanization was an important process in the history of Europe, particularly that of the Iberian Peninsula. The populations in the north of the peninsula avoided Roman domination until the campaign of Augustus in the 1st century BC and maintained some aspects of their indigenous cultures. Roman domination of Hispania. Both Swabians and Visigoths maintained the sociocultural organization of the Roman Empire.At the beginning of 5th century, when the Roman Empire fell, the Swabians, a people of German origin, settled for a period of time in Galicia. They lived alongside the native population and at the end of 6th century the Swabian kingdom was incorporated into the Visigothic As occurred with the process of Romanization, the impact of the Muslim invasion of the Iberian Peninsula in the year 711 was less in both Galicia and other northern territories than in other regions. As a consequence, the regions of Galicia, northern Portugal, and the area to the north of the River Duero acted as important refuges from the Muslim invasion during the 9th and 10th centuries. This led to a process of repopulation that was most rapid in the western part of Galicia and Portugal.Roman Galicias on either side of the border.Following the Christian reconquest, the kingdom of Asturias and Leon appeared, including part of Galicia. This kingdom was later integrated as part of the Crown of Castile. Following a series of changes, Portugal was recognized as an independent kingdom in 1143. The process of the Christian reconquest also led to significant migration from the north to the south of the peninsula to repopulate the conquered territories, while east-west movement was limited by the existence of different Christian kingdoms. The border between Castile and Portugal caused a significant reduction in the previously unrestricted movement between the old vias of the Antoninus Itinerary, known since then as the Camino de Santiago.Pilgrimages to Santiago de Compostela, one of the main Christian destinations alongside Jerusalem and Rome, have allowed contact between Galicia and numerous peoples of Europe who have continued to arrive since the Middle Ages, largely following the old Geography, relief, and climate, as well as history, would have contributed to endowing a certain similarity between the populations of the north of Spain and to distinguishing them from other populations of the peninsula. This observation is supported by analysis of inbreeding patterns. The high rates of marriages between close relatives (uncle-niece/aunt-nephew: M12 and first cousins: M22) and the levels of inbreeding distinguish northern Iberians, namely Galicians, Asturians, Cantabrians and Basques -7 from oAlu insertions . Certainly, the four Basque subpopulations from Spain represent a variable degree of differentiation with Galician and other Iberian populations, Guipúzcoa representing the Basque province with the most genetic distinctiveness, a finding that would agree with the Basque linguistic map .The close genetic similarity between Galicia, an Atlantic population, and Valencia, a Mediterranean one, is also unexpected as they are geographically separated by a long distance and represent quite dissimilar cultures with different histories. One of the main routes of human dispersion from Europe towards Iberia from the late Palaeolithic has been the corridor of the western Pyrenees. The logical direction of those migratory movements would have involved three main axes: one of them westward, along the Cantabrian cornice; another towards the Castilian central plateau, in a southward and westward direction crossing all over the region of Castile-Leon until reaching Galicia and northern Portugal; and a third route of gene flow following the Ebro valley towards the Mediterranean coast of Spain. Those continuous population movements are likely to have produced interesting processes of admixture among rather distant Iberian populations located within the geographic area of influence of that corridor. Taking as a reference a critical border area between Spain and France (in the Basque province of Guipúzcoa), and thus located inside the main passageway of communication of the western Pyrenees, the geographic distance of Galicia and Valencia to that point would be quite similar. Nevertheless, it will be of interest to examine how Iberian genetic population diversities correlate with geographic distances to that main Pyrenean entrance point for gene flow, when further genetic data from different genomic regions were available, despite the continued presence of the Basque distortion. Such an analysis would obviously need to be accompanied by more extensive sampling to cover the main, critical geographic regions of Spain and Portugal.When Galicia is compared with other European populations by principal component analysis, our study sample is seen to be integrated within the central cluster, even though its geographic position in the European territory is extreme. This finding suggests that the population of Galicia is the result of a relatively balanced mixture of European populations or of the ancestral populations that gave rise to them. This would support the importance of the migratory movements that have taken place in Europe over the course of recent human history and their effects on the European genetic landscape.GM and KM allotypes show a poor correlation between geographic and genetic distances in Spanish populations. The Basque subpopulations from Spain exhibit a variable degree of differentiation in relation to Galicia, the Basques from Guipúzcoa being the most genetically distant. The close genetic similarity observed between samples from Galicia and Valencia, despite the geographical distance between them and their cultural and historical distinctiveness, may be explained by their being located at similar distances from the western end of the Pyrenees, the main passageway for communication between Spain and the rest of Europe, through which most Iberian gene flow would have passed. Likewise, the relatively high incidence of the African marker GM* 1, 17 23' 5* in Galicia may be the result of ancient admixtures as well as the recent introduction of genes from mixed black populations from America.The Galician population is genetically close to the population of Portugal, its geographic neighbor, but also to that of France, which is further away. The central position of Galicia in the European genetic landscape, despite its extreme geographical location in the Iberian Peninsula, would support the decisive role of historic migratory movements in Europe.2), in 1900 it was 1 980 525 and in 2000 the population reached 2 731 900 (93 inhabitants per km2). The proportion of the Spanish population accounted for by Galicia has been reduced by half over the last two centuries  1/2, where ijD is the distance between points i and j. Otherwise, UTM coordinates were transformed into geodesic coordinates to take into account the curvature of the earth. These calculations were performed with specific software [Geographic distances were calculated from Universal Transverse Mercator (UTM) coordinates, taking the corresponding capital town as a reference when samples referred to a province. In the case of Galicia, as the sampling mimics the population of the whole region, the UTM coordinates of the weighted centre of the present Galician population was used. When both populations were located within the same UTM zone, their geographic distance was calculated as a common Euclidean distance software .GM haplotype frequencies in European populations and made interesting suggestions while preparing the manuscript. AG-M and BA worked together in the genetic data analysis. All authors contributing to this study read and agreed the final manuscript.RC planned and took responsibility of writing the manuscript. TAV, RL and JF designed the sampling process and collected the blood samples in the Galician region. EG carried out the characterization of immunoglobulin markers in the Galician sample, JMD supervised the allotypic genetic data, prepared the DataBase on