Genetic structure of North American wolverine (Gulo gulo) populations

Wolverines (Gulo gulo) are found in low densities throughout their circumpolar distribution. They are also potentially susceptible to human-caused population fragmentation (development, recreation and fur harvesting). The combination of these factors has contributed to this species being listed as having either vulnerable or endangered status across much of its current range. The effects of inherently low densities and anthropogenic pressures on the genetic structure and variation of wolverine populations are, as yet, unknown. In this study, 461 individuals were typed at 12 microsatellite loci to investigate the population genetic structure of wolverines from north-western Alaska to eastern Manitoba. Levels of gene flow and population differentiation among the sampled regions were estimated via a genotype assignment test, pairwise F(ST), and two genetic distance measures. Our results suggest that wolverine populations from southernmost regions, in which anthropogenic factors are strongest, revealed more genetic structuring than did northern populations. Furthermore, these results suggest that reductions in this species' range may have led to population fragmentation in the extreme reaches of its southern distribution. The continued reduction of suitable habitat for this species may lead to more populations becoming isolated remnants of a larger distribution of northern wolverines, as documented in other North American carnivore species.     

Capture and medetomidine-ketamine anesthesia of free-ranging wolverines (Gulo gulo)

Capture and anesthesia with medetomidine-ketamine were evaluated in free-ranging wolverines (Gulo gulo) immobilized for marking with radiocollars or intraperitoneal radiotransmitters in Norrbotten, Sweden, during early June 2004 and 2005. Twelve juvenile wolverines were captured by hand and injected with 0.14 +/- 0.03 mg/kg (mean +/- SD) medetomidine and 7.5 +/- 2.0 mg/kg ketamine. Twelve adult wolverines were darted from a helicopter or the ground, or captured by hand. Adults received 0.37 +/- 0.06 mg/kg medetomidine and 9.4 +/- 1.4 mg/kg ketamine. Arterial blood samples were collected between 15 min and 30 min and between 45 min and 60 min after drug administration and immediately analyzed for selected hematologic and plasma variables. Hyperthermia was recorded initially in one juvenile wolverine and 11 adults. Rectal temperature, heart rate, and lactate decreased significantly during anesthesia, whereas hemoglobin oxygen saturation, pH, partial pressure of arterial carbon dioxide, and base excess increased. Adult wolverines darted from a helicopter had a significantly higher rectal temperature, higher glucose and hematocrit values, and a lower heart rate than juveniles captured by hand. Impaired arterial oxygenation was evident in all wolverines. This study provides baseline data on physiologic variables in adult and juvenile wolverines captured with different methods and anesthetized with medetomidine-ketamine.     

The diet of breeding female wolverines (Gulo gulo) in two areas of Finland

The wolverine (Gulo gulo) hunts and scavenges for food. We examined how the presence of a semi-domesticated reindeer (Rangifer tarandus tarandus) influences the diet of breeding female wolverines in Finland. We compared the dietary composition of wolverines breeding in the reindeer management area (northern Finland) with that of wolverines breeding outside this area (eastern Finland) by analyzing 421 scats collected from six active dens in northern Finland and four dens in eastern Finland during 2004–2006. The occurrence of food sources in the diet was assessed for both areas using logistic mixed-effects models. Diet breadth per study area was also calculated. As expected, semi-domesticated reindeer was the most important food source (66 %) in northern Finland. Mountain hares (Lepus timidus) were the second most important food item (16 %). Wolverines also utilized grouse (7 %), small rodents (6 %), and moose (Alces alces; 4 %) in their diet. In eastern Finland, the most utilized prey species was moose (55 %), while the diet also comprised mountain hares (13 %), domestic pigs (Sus scrofa; 13 %), grouse (11 %), and small rodents (2 %). The diet of wolverines in eastern Finland included more moose and less semi-domesticated reindeers and small rodents than in northern Finland. The niche breadth did not differ between the study areas. In eastern Finland, the diet of wolverines shifted to 55 % moose and 0 % semi-domesticated reindeer; compared to 4 % moose and 66 % semi-domesticated reindeer in northern Finland. Our study highlights the important role of ungulates in the diet of breeding female wolverines. In areas with a low density of medium-sized ungulates, scavenging for wolf- and human-killed moose and carcasses on feeding sites plays an essential role in food acquisition by wolverines.     

Behavioral and endocrine correlates of reproductive failure in social aggregations of captive wolverines (Gulo gulo)

Sociality in mammals is often viewed as a dichotomy, with sociality contrasted against solitariness. However, variation within these broad categories may have strong effects on individual fitness. For example, reproductive suppression of social subordinates is generally associated with group living, but suppression may also occur in solitary species if the behavioral and physiological processes involved can be modulated by the demographic environment. To investigate whether behavioral and physiological traits that normally are associated with group living might be latent even in a solitary species, we explored the level of sociality and investigated causes and mechanisms of reproductive failure in female wolverines Gulo gulo that experienced a highly aggregated social environment in captivity. Behaviorally, females showed low levels of aggression and intermediate levels of social interactions. Reproductive failure seemed to have been related to low social rank and to have occurred between ovulation and implantation in 13 out of 15 breeding attempts. However, three of eight females observed to mate produced offspring, indicating that no individual female fully managed to monopolize breeding. Reproductive failure was not related to elevated levels of glucocorticoid stress hormones. Rather, elevated glucocorticoid levels during the mating season were associated with successful reproduction. We suggest that social tendencies and physiological mechanisms mediating reproductive suppression may be viewed as reaction norms to the social environment. We further suggest that the social flexibility of solitary carnivores might be greater than is commonly observed, due to ecological constraints that may limit aggregation.     

Two new species of Sarcocystis (Apicomplexa: Sarcocystidae) infecting the wolverine (Gulo gulo) from Nunavut, Canada

Infection with Sarcocystis species is common in many species of animals, but it has not yet been reported in wolverines (Gulo gulo). Histological sections of tongues from 41 wolverines in the Kitikmeot Region, Nunavut, Canada, were examined for sarcocysts. Sarcocysts were found in 33 (80.4%) wolverines. Two structurally distinct types of sarcocysts were found. Type A sarcocysts were thin (<1 μm thick) walled. Ultrastructurally, the parasitophorous vacuolar membrane (Pvm) had minute undulations, but it lacked villar protrusions and was not invaginated into the granular layer. The bradyzoites were slender, about 5 × 1 μm in size. Structurally, these sarcocysts were distinct from known species of Sarcocystis and possessed a novel 18S and ITS-1 sequence, sharing 98% and 78% sequence similarity with Sarcocystis canis . A new species name, Sarcocystis kalvikus, is proposed for type A sarcocysts. In contrast, type B sarcocysts had relatively thicker (about 2 μm) cyst walls and larger bradyzoites, each about 10 × 2-3 μm. Ultrastructurally, the Pvm on the sarcocyst wall had villar protrusions that were either mushroom-like or sloping. Molecular analysis identified a unique 18S and ITS-1 sequence that placed them in a clade within the Sarcocystidae. Based on histology, TEM, and genetic data, the new name, Sarcocystis kitikmeotensis, is proposed. Sarcocystis kalvikus was found in 14 (34.1%), S. kitikmeotensis was found in 7 (17%), and both species were found in 12 (29.2%) of 41 wolverines.     

Genetic diversity and population structure of wolverine (Gulo gulo) populations at the southern edge of their current distribution in North America with implications for genetic viability

The current range of wolverines (Gulo gulo) within the lower 48 states includes small, remnant populations in Idaho, Washington, Wyoming and Montana. The size and trend of each of these populations and connectivity to adjacent populations in the contiguous United States and Canada are poorly understood. In this study, levels of genetic diversity and population genetic structure were examined in three states (Idaho, Wyoming, and Montana) and two Canadian provinces (Alberta and British Columbia) using both mitochondrial (mtDNA) and nuclear microsatellite DNA. Restricted levels of gene flow were detected among these populations with mitochondrial and nuclear DNA and our observations suggest a pattern of male-mediated gene flow. Populations in the United States appear to be receiving migrants from Canada, however, substantial genetic differentiation suggests that gene flow may not be high enough to prevent genetic drift. Our analyses suggest that at least 400 breeding pairs or 1–2 effective migrants per generation would be needed to ensure genetic viability in the long-term for each of the populations in the United States. Significant matrilineal structuring and restricted female gene flow indicates that demographic viability will depend upon the movement of female wolverines into new territories. Results from this study provide guidelines for conservation and management and indicate the need for more ecological data.     

Analyzing the Spatial Structure of Broughtonia cubensis (Orchidaceae) Populations in the Dry Forests of Guanahacabibes,Cuba

A current key issue in ecology is the role of spatial effects on population and community dynamics. In this paper, we test several hypotheses related to spatial structures and coexistence of epiphytic tropical orchid species with special emphasis on the endemic species Broughtonia cubensis. More specifically, we explored the spatial structure of orchid–host plant communities at three different levels of organization (occupied vs. nonoccupied host trees, trees with B. cubensis vs. other orchids, and reproductive vs. nonreproductive B. cubensis plants). We mapped all potential host trees and orchids at three 20 × 20 m plots and applied techniques of spatial point pattern analysis such as mark connection and mark correlation functions to evaluate departures from randomized communities. We found spatial aggregation of trees with epiphytic orchids and segregation between trees with and without epiphytic orchids, and that there was an intraspecific spatial aggregation of B. cubensis in relation to the other seven epiphytic orchid species. Furthermore, we found spatial aggregation of reproductive B. cubensis individuals and segregation between reproductive and nonreproductive individuals on their phorophytes. Thus, orchid–host plant communities show hierarchical spatial structuring with aggregation and segregation at different levels of organization. Our results point to an enhancement of local species in the coexistence of tropical epiphytic orchid communities, by reducing competition through niche differentiation.     

Contrasting Genetic Structure among Populations of Two Amphidromous Fish Species (Sicydiinae) in the Central West Pacific

Both present-day and past processes can shape connectivity of populations. Pleistocene vicariant events and dispersal have shaped the present distribution and connectivity patterns of aquatic species in the Indo-Pacific region. In particular, the processes that have shaped distribution of amphidromous goby species still remain unknown. Previous studies show that phylogeographic breaks are observed between populations in the Indian and Pacific Oceans where the shallow Sunda shelf constituted a geographical barrier to dispersal, or that the large spans of open ocean that isolate the Hawaiian or Polynesian Islands are also barriers for amphidromous species even though they have great dispersal capacity. Here we assess past and present genetic structure of populations of two amphidromous fish (gobies of the Sicydiinae) that are widely distributed in the Central West Pacific and which have similar pelagic larval durations. We analysed sections of mitochondrial COI, Cytb and nuclear Rhodospine genes in individuals sampled from different locations across their entire known range. Similar to other Sicydiinae fish, intraspecific mtDNA genetic diversity was high for all species (haplotype diversity between 0.9–0.96). Spatial analyses of genetic variation in Sicyopus zosterophorum demonstrated strong isolation across the Torres Strait, which was a geologically intermittent land barrier linking Australia to Papua New Guinea. There was a clear genetic break between the northwestern and the southwestern clusters in Si. zosterophorum (φ_ST = 0.67502 for COI) and coalescent analyses revealed that the two populations split at 306 Kyr BP (95% HPD 79–625 Kyr BP), which is consistent with a Pleistocene separation caused by the Torres Strait barrier. However, this geographical barrier did not seem to affect Sm. fehlmanni. Historical and demographic hypotheses are raised to explain the different patterns of population structure and distribution between these species. Strategies aiming to conserve amphidromous fish should consider the presence of cryptic evolutionary lineages to prevent stock depletion.     

Allozyme Variation among Populations of Rana pirica (Amphibia: Anura)

Genetic variation of Rana pirica , an east Asian brown frog of the R. temporaria group, was elucidated by analysing 140 specimens from 11 populations from Hokkaido and Sakhalin, both locating near the eastern coasts of the Asian continent, and 12 specimens of R. ornativentris from Honshu, Japan mainland, as an outgroup, through horizontal starch gel electrophoresis. Rana pirica shows relatively low genetic differentiation, and high genetic affinities are found between Hokkaido and Sakhalin populations. Populations from these islands are morphologically somewhat differentiated but should be regarded as conspecific. Degree of genetic divergence among populations of amphibian species from Hokkaido, including R. pirica , is not so extensive as that reported for species from Japan mainland and relatively recent formation of amphibian fauna in Hokkaido is suggested.     

Spatial Heterogeneity of Bacterial Populations in Monomictic Lake Estanya (Huesca,Spain)

Bacterial population changes were investigated in the monomictic Lake Estanya by combining microscopic analysis and two molecular methods involving the amplification of 16S rDNA genes using primers for the domain Bacteria and subsequent restriction fragment length polymorphism (PCR–RFLP) and denaturing gradient gel electrophoresis (PCR–DGGE). Both approaches revealed the vertical distribution of predominant microbial morphotypes and phylotypes in both holomictic and stratified periods, respectively, and showed that variations in structure and composition of bacterial populations are occurring in this lake as a function of depth and time. Through principal component analysis (PCA), these shifts could be related to different physicochemical parameters with temperature, oxygen concentration, and the incident light being of paramount importance as structuring variables. Comparison of RFLP and DGGE profiles by scoring similarities using the Jaccard coefficient and then building a multidimensional scaling map (MDS) showed equivalent results. Both techniques revealed that bacterial populations, present in the whole water column in the holomictic period, showed a high similarity with those located in the deeper part of the lake in the stratified period, evidencing that other factors, both biotic and abiotic, should also be considered as a force driving change in the composition of the bacterial community. Furthermore, DGGE analysis showed that sequences from prominent bands were affiliated to members of four major phyla of the domain Bacteria: Cyanobacteria, Bacteroidetes, Proteobacteria, and Actinobacteria, most of which corresponded to heterotrophic bacterial populations involved in carbon, sulfide, and nitrogen biogeochemical cycles, which were indistinguishable under the light microscope.     

Community Structure (Niche Limitation and Guild Proportionality) in Relation to the Effect of Spatial Scale, in a

A Nothofagus-dominated rainforest in eastern Fiordland, New Zealand, was sampled by shoot frequency in contiguous 1 x 1 m quadrats, along a topologically-circular transect. The data were analysed at five scales up to 5 x 1 m, to search for assembly rules, i.e., generalised restrictions on species co- occurrences. There was no evidence of niche limitation in terms of the whole community, at any scale examined. Rather, variance in species richness was greater than expected from a null model, suggesting environmental heterogeneity. This conclusion was confirmed by using a patch-model. Guild structure was examined in terms of synusiae, based on vertical stratification. The proportion of species in a quadrat that were from the Herb guild was significantly more constant than expected under the null model. However, using a patch-model this effect was reduced, and no longer significant. This suggests pools of species adapted to different micro- environments, but with parallel guild composition. Liane guild proportions were more variable than predicted by the null model; although this effect was not significant, it parallels reports from other South Island forests. The work confirms conclusions from previous work, that plant assembly rules are to be found only at small spatial scales. Processes leading to high and low variance in species richness and in guild proportions are discussed. Selection of a uniform site in the present work, and use of a patch-model, make it less likely that effects are due to environmental or historical effects. The use of guild proportionality as the criterion eliminates the effect of limitation on the number of individuals that can co-occur locally.     

Continental-Scale Footprint of Balancing and Positive Selection in a Small Rodent (Microtus arvalis)

Genetic adaptation to different environmental conditions is expected to lead to large differences between populations at selected loci, thus providing a signature of positive selection. Whereas balancing selection can maintain polymorphisms over long evolutionary periods and even geographic scale, thus leads to low levels of divergence between populations at selected loci. However, little is known about the relative importance of these two selective forces in shaping genomic diversity, partly due to difficulties in recognizing balancing selection in species showing low levels of differentiation. Here we address this problem by studying genomic diversity in the European common vole (Microtus arvalis) presenting high levels of differentiation between populations (average F _ST = 0.31). We studied 3,839 Amplified Fragment Length Polymorphism (AFLP) markers genotyped in 444 individuals from 21 populations distributed across the European continent and hence over different environmental conditions. Our statistical approach to detect markers under selection is based on a Bayesian method specifically developed for AFLP markers, which treats AFLPs as a nearly codominant marker system, and therefore has increased power to detect selection. The high number of screened populations allowed us to detect the signature of balancing selection across a large geographic area. We detected 33 markers potentially under balancing selection, hence strong evidence of stabilizing selection in 21 populations across Europe. However, our analyses identified four-times more markers (138) being under positive selection, and geographical patterns suggest that some of these markers are probably associated with alpine regions, which seem to have environmental conditions that favour adaptation. We conclude that despite favourable conditions in this study for the detection of balancing selection, this evolutionary force seems to play a relatively minor role in shaping the genomic diversity of the common vole, which is more influenced by positive selection and neutral processes like drift and demographic history.     

Genetic Structure of Earthworm Populations at a Regional Scale: Inferences from Mitochondrial and Microsatellite Molecular Markers in Aporrectodea icterica (Savigny 1826)

Despite the fundamental role that soil invertebrates (e.g. earthworms) play in soil ecosystems, the magnitude of their spatial genetic variation is still largely unknown and only a few studies have investigated the population genetic structure of these organisms. Here, we investigated the genetic structure of seven populations of a common endogeic earthworm (Aporrectodea icterica) sampled in northern France to explore how historical species range changes, microevolutionary processes and human activities interact in shaping genetic variation at a regional scale. Because combining markers with distinct modes of inheritance can provide extra, complementary information on gene flow, we compared the patterns of genetic structure revealed using nuclear (7 microsatellite loci) and mitochondrial markers (COI). Both types of markers indicated low genetic polymorphism compared to other earthworm species, a result that can be attributed to ancient bottlenecks, for instance due to species isolation in southern refugia during the ice ages with subsequent expansion toward northern Europe. Historical events can also be responsible for the existence of two divergent, but randomly interbreeding mitochondrial lineages within all study populations. In addition, the comparison of observed heterozygosity among microsatellite loci and heterozygosity expected under mutation-drift equilibrium suggested a recent decrease in effective size in some populations that could be due to contemporary events such as habitat fragmentation. The absence of relationship between geographic and genetic distances estimated from microsatellite allele frequency data also suggested that dispersal is haphazard and that human activities favour passive dispersal among geographically distant populations.     

Essential Oil Variation among Natural Populations of Lavandula multifida L. (Lamiaceae)

Volatiles from twelve wild Tunisian populations of Lavandula multifida L. growing in different bioclimatic zones were assessed by GC (RI) and GC/MS. Thirty‐six constituents, representing 83.48% of the total oil were identified. The major components at the species level were carvacrol (31.81%), β‐bisabolene (14.89%), and acrylic acid dodecyl ester (11.43%). These volatiles, together with α‐pinene, were also the main compounds discriminating the populations. According to these dominant compounds, one chemotype was revealed, a carvacrol/β‐bisabolene/acrylic acid dodecyl ester chemotype. However, a significant variation among the populations was observed for the majority of the constituents. A high chemical‐population structure, estimated both by principal component analysis (PCA) and unweighted pair group method with averaging (UPGMA) cluster analysis based on Euclidean distances, was observed. Both methods allowed separation of the populations in three groups defined rather by minor than by major compounds. The population groups were not strictly concordant with their bioclimatic or geographic location. Conservation strategies should concern all populations, because of their low size and their high level of destruction. Populations exhibiting particular compounds other than the major ones should be protected first.     

Allozyme Diversity and Population Structure in Korean Populations of Cymbidium goeringii (Orchidaceae)

Cymbidium goeringii (Orchidaceae). Mean observed population heterozygosity (H _op=0.181), expected heterozygosity (H _eP=0.240), and total genetic diversity (H _T =0.351) were all higher than average values for species with similar life-history characteristics. A considerable deficit of heterozygotes relative to Hardy–Weinberg expectations was detected (77% of fixation indices were positive) with a mean F _IS of 0.278. On average, 90% of the total genetic diversity was found within populations (mean G _ST = 0.098). An indirect estimate of the number of migrants per generation (Nm=2.30, calculated from G _ST , Nm=8.48, calculated from the frequencies of unique alleles) indicated that gene flow has been extensive in C. goeringii. Results of a spatial autocorrelation analysis based on allele frequencies of 16 populations revealed a trend with respect to the distance classes (0<63 km, six significant positive values; beyond that distance, 12 significant negative values). It is highly probable that C. goeringii has a history of relative large, continuous populations that had greater chance for gene movement among adjacent populations via large numbers of small seeds, following the last Ice Age. Factors contributing to the high levels of genetic diversity found within populations of C. goeringii include its large and continuous populations, its long-lived perennial habit, its widespread geographical distribution, and its ability for relatively long distance seed movement by wind. Received 18 June 1998/ Accepted in revised form 18 November 1998     

Genetic Structure of Natural Populations of Wild Soybean (Glycine soja) in Beijing Region

In order to exploit the genetic resources of wild soybean (Glycine soja) which is the progenitor of cultivated soybean (Glycine max), the genic frequencies of Ti (coding trypsin inhibitors) and Sp1 (coding β-amylase isozymes) for 13 populations of wild soybean in Beijing region were determined. There are 2 alleles (Tia and Tib) in Ti locus of Beijing populations. Calculation of heterozygosity indicates Sp1 is polymorphic, while this monomorphic within a population. Based on the vatiation (from 0 to 50%) for heterozygosity of Sp1 among populations, with special reference to the values of genetic distances among populatious, and no heterozygote has been found in 1300 plants which would be heterozygotes if they were outbreeder, we suggested that wild soybean in natural populations is absolute inbreeder. The frequencies of Ti and Sp1 alleles vary from place to place extremely, however, no correlation exists between allozyme frequencies and ecological factors. Field investigation has shown that there is a threat from the reduction in available habitats, caused by building irrigation works .and urbanization. Finally, sampling strategy for conservation of genetic resources of wild soybean was discussed and some suggestions were made.