Integrating ecological and genetic structure to define management units for caribou in Eastern Canada

Genetic diversity is a key parameter to delineate management units, but many organisms also display ecological characteristics that may reflect potential local adaptations. Here, we used ecological and genetic information to delineate management units for a complex system involving several ecotypes of caribou (Rangifer tarandus) from Québec and Labrador, eastern Canada. We genotyped 560 caribou at 16 microsatellite loci and used three Bayesian clustering methods to spatially delineate and characterize genetic structure across the landscape. The different approaches employed did not converge on the same solution, and differed in the number of inferred genetic clusters that best fit the dataset but also in the spatial distribution of genetic variation. We reconciled variability among the methods using a synthetic approach that considers the sum of the partitions obtained by each of them and retrieved six genetically distinct groups that differ in their spatial extent across the range of caribou in the study area. These genetic groups are not consistent with the presently defined ecological designations for this species. Combining both genetic and ecological criteria, we distinguished eight independent management units. Overall, the management units we propose should be the focus of conservation and management actions aimed to maximize genetic and ecological diversity and ensure the persistence of caribou populations inhabiting increasingly disturbed landscapes.     

Rodent foraging is affected by indirect, but not by direct, cues of predation risk

We used foraging trays to determine whether oldfield mice, Peromyscuspolionotus, altered foraging in response to direct cues of predationrisk (urine of native and nonnative predators) and indirectcues of predation risk (foraging microhabitat, precipitation,and moon illumination). The proportion of seeds remaining ineach tray (a measure of the giving-up density [GUD]) was usedto measure risk perceived by mice. Mice did not alter theirGUD when presented with cues of native predators (bobcats, Lynxrufus, and red foxes, Vulpes vulpes), recently introduced predators(coyotes, Canis latrans), nonnative predators (ocelots, Leoparduspardalis), a native herbivore (white-tailed deer, Odocoileusvirginianus), or a water control. Rather, GUD was related tomicrohabitat: rodents removed more seeds from foraging trayssheltered beneath vegetative cover compared with exposed traysoutside of cover. Rodents also removed more seeds during nightswith precipitation and when moon illumination was low. Our resultssuggest that P. polionotus used indirect cues rather than directcues to assess risk of vertebrate predation. Indirect cues maybe more reliable than are direct scent cues for estimating riskfrom multiple vertebrate predators that present the most riskin open environments.     

Landscape genetics of a specialized grasshopper inhabiting highly fragmented habitats: a role for spatial scale

Aim The study of geographical discontinuities in the distribution of genetic variability in natural populations is a central topic in both evolutionary and conservation research. In this study, we aimed to analyse (1) the factors associated with genetic diversity at the landscape spatial scale in the highly specialized grasshopper Mioscirtus wagneri and (2) to identify the relative contribution of alternative factors to the observed patterns of genetic structure in this species. Location La Mancha region, Central Spain. Methods We sampled 28 populations of the grasshopper M. wagneri and genotyped 648 individuals at seven microsatellite loci. We employed a causal modelling approach to identify the most influential variables associated with genetic differentiation within a multiple hypothesis‐testing framework. Results We found that genetic diversity differs among populations located in different river basins and decreases with population isolation. Causal modelling analyses showed variability in the relative influence of the studied landscape features across different spatial scales. When a highly isolated population is considered, the analyses suggested that geographical distance is the only factor explaining the genetic differentiation between populations. When that population is excluded, the causal modelling analysis revealed that elevation and river basins are also relevant factors contributing to explaining genetic differentiation between the studied populations. Main conclusions These results indicate that the spatial scale considered and the inclusion of outlier populations may have important consequences on the inferred contribution of alternative landscape factors on the patterns of genetic differentiation even when all populations are expected to similarly respond to landscape structure. Thus, a multiscale perspective should also be incorporated into the landscape genetics framework to avoid biased conclusions derived from the spatial scale analysed and/or the geographical distribution of the studied populations.     

Microbiological quality of blue mussels (Mytilus edulis) in Nunavik, Quebec: a pilot study

This pilot study was aimed at documenting the presence of fecal indicators and enteric pathogens in blue mussels (Mytilus edulis) from 6 communities in Nunavik, Quebec. One to four 2?kg samples of mussels were collected at low tide in each community. Samples were investigated by enumeration methods for the fecal indicators enterococci, Escherichia coli, F-specific coliphages, Clostridium perfringens, and by molecular identification for the pathogens norovirus, Salmonella spp., Campylobacter jejuni, Campylobacter coli, and Campylobacter lari, verocytotoxin-producing E.?coli (particularly serovar O157:H7), Shigella spp., and Yersinia enterocolitica. In 5 communities, the presence of Giardia duodenalis and Cryptosporidium spp. was also tested by microscopy and molecular methods and that of Toxoplasma gondii was tested by molecular methods. Apart from small quantities of Clostridium perfringens in 2 samples, no bacterial or viral pathogens were detected in the mussels. Toxoplasma gondii was also not detected. However, G.?duodenalis and Cryptosporidium spp. were present in 18% and 73% of the samples investigated for these pathogens, respectively. When considering the indicators and the viral and bacterial pathogens investigated, the mussels examined were of good microbiological quality, but considering the presence of potentially zoonotic protozoa, it should be recommended that consumers cook the molluscs well before eating them.     

Computational Finishing of Large Sequence Contigs Reveals Interspersed Nested Repeats and Gene Islands in the rf1-Associated Region of Maize

The architecture of grass genomes varies on multiple levels. Large long terminal repeat retrotransposon clusters occupy significant portions of the intergenic regions, and islands of protein-encoding genes are interspersed among the repeat clusters. Hence, advanced assembly techniques are required to obtain completely finished genomes as well as to investigate gene and transposable element distributions. To characterize the organization and distribution of repeat clusters and gene islands across large grass genomes, we present 961- and 594-kb contiguous sequence contigs associated with the rf1 (for restorer of fertility1) locus in the near-centromeric region of maize (Zea mays) chromosome 3. We present two methods for computational finishing of highly repetitive bacterial artificial chromosome clones that have proved successful to close all sequence gaps caused by transposable element insertions. Sixteen repeat clusters were observed, ranging in length from 23 to 155 kb. These repeat clusters are almost exclusively long terminal repeat retrotransposons, of which the paleontology of insertion varies throughout the cluster. Gene islands contain from one to four predicted genes, resulting in a gene density of one gene per 16 kb in gene islands and one gene per 111 kb over the entire sequenced region. The two sequence contigs, when compared with the rice (Oryza sativa) and sorghum (Sorghum bicolor) genomes, retain gene colinearity of 50% and 71%, respectively, and 70% and 100%, respectively, for high-confidence gene models. Collinear genes on single gene islands show that while most expansion of the maize genome has occurred in the repeat clusters, gene islands are not immune and have experienced growth in both intragene and intergene locations.Genome sequencing of the maize (Zea mays) genome is nearing completion (Bennetzen et al., 2001; Chandler and Brendel, 2002; Wessler, 2006); it is the largest and most difficult-to-assemble plant genome sequenced to date. Maize is an important economic, agricultural, industrial, and research crop; however, with a genome close to the size of the human genome (2.8 Gb) and its high percentage of repetitive elements, acquiring the maize genome seemed a daunting task. Approximately 67% of the genome is made up of transposable elements (TEs; Haberer et al., 2005; Kronmiller and Wise, 2008), increasing the difficulty of assembly (Rabinowicz and Bennetzen, 2006). Much exploratory work has gone into isolating and sequencing just the gene areas and ignoring the repetitive regions, both by methylation filtration (Rabinowicz et al., 1999; Palmer et al., 2003; Whitelaw et al., 2003) and high-C₀t (Whitelaw et al., 2003; Yuan et al., 2003) systems, which have assisted researchers with selecting only genic regions to sequence. These methods have captured a majority of the maize genic sequence (Fu et al., 2005), but they still have the potential to miss important regions. The current genome-sequencing project aims to capture the entire gene set of maize, including regulatory regions. However, the current strategy will not provide a fully assembled genome but rather assembled bacterial artificial chromosome (BAC) contigs ordered and orientated to provide complete gene regions that are adjacent to potentially incomplete TE clusters.The landscape of the maize genome provides an interesting challenge for both sequencing and subsequent annotation. A high density of long terminal repeat (LTR) retrotransposons has had a direct effect on the genome size of many plant genomes, including maize (SanMiguel et al., 1996; Bennetzen et al., 2005; Hawkins et al., 2006; Piegu et al., 2006). Besides expanding genome size, LTR retrotransposons can have an impact on evolution of the species (Kidwell and Lisch, 2000). LTR retrotransposon insertions tend to form nested clusters (SanMiguel and Bennetzen, 1998), which are separated by small regions of several genes. Large nested repeat clusters consist of TE insertions inside TE sequences, expanding the repeat cluster and breaking up the sequence of the TEs found within, hindering repeat and gene annotation and increasing the difficulty of assembly. However, full sequence completion of the repetitive regions can be of great benefit to understanding the evolutionary history of the maize genome. LTR retrotransposons can provide an estimated time since insertion by calculating the divergence of their LTRs (Kimura, 1980; Ma and Bennetzen, 2004), and carefully sequenced assemblies of nested repeat clusters can help to illustrate their expansion, proliferation, and evolution across the genome (Kronmiller and Wise, 2008).Previous studies of large contiguous regions of maize have provided a general view of the landscape of the genome. Unfinished sequence totaling 7.8 Mb from chromosome 1 and 6.6 Mb from chromosome 9 shows a gene density of one gene per 33 and 27 kb, respectively (Bruggmann et al., 2006). BAC contigs ranging in size from 126 to 405 kb show a gene density of one gene per 19 kb and genes found in small groups between large repeat clusters (Brunner et al., 2005). Genome-wide analysis of maize BACs has painted a different picture: while gene density of 100 random BACs at one gene per 44 kb was similar to the above results, genes were not observed in tight clusters (Haberer et al., 2005). When investigating gene-specific areas of maize, this dichotomy of gene density is also seen. Analysis of gene-rich regions such as the 22-kd α-zein gene family on maize chromosome 4 reveals a high density of genes, with one gene observed per 10 kb over 346 kb (Song et al., 2001). The Adh1 locus on maize chromosome 1 contains two genes across 280 kb, or one gene per 140 kb. Perhaps the only message learned here is that the gene density across the maize genome varies to a great degree, and large contiguous sequenced regions can begin to capture the true diversity of maize chromosome architecture.In order to characterize large contiguous regions of maize sequence, we identified and sequenced two B73 BAC contigs from the centromeric region of chromosome 3. These contigs of 961 and 594 kb correspond to contigs 117 and 119, respectively, on maize WebFPC (Wei et al., 2007) and span regions associated with the rf1 (for restorer of fertility1) locus for Texas (T) cytoplasmic male sterility (cmsT; Duvick et al., 1961; Wise et al., 1996). As a foundation for the isolation of the Rf1 locus, four rf1 male-sterile mutants were recovered from a screen of 123,500 flowering plants (Wise et al., 1996). A 5.5-kb Mu1-hybridizing EcoRI restriction fragment was identified that cosegregated with the rf1-m3207 allele. Sequences from this fragment were hybridized to a Rf1 cDNA library, and probes designed from the identified cDNA, p6140-1 (Wise et al., 1999), were found to cosegregate with the rf1 locus in a recombinant population selected from over 10,000 progeny.Using probes designed off the 5.5-kb cosegregating restriction fragment and the p6140-1 cDNA, we have identified two BAC contigs spanning the rf1 locus. Sixteen BACs were sequenced to completion to provide high-quality finished sequence. Here, we present two methods for computational finishing of highly repetitive grass genomes, which were successfully utilized to close 11 TE-induced gaps. Sixteen nested repeat clusters were found, each spanning as much as 155 kb and containing a variety of LTR retrotransposon types and ages of insertion. Genes are found tightly clustered, showing a density rate of one gene per 16 kb within gene islands. Finally, comparative analysis with rice (Oryza sativa) and sorghum (Sorghum bicolor) shows that while many genes are retained across all three species, genes have both been lost and translocated across the genomes.     

Rapid method for measuring ScFv thermal stability by yeast surface display

We have characterized a simplified method to determine the relative thermal stability of single-chain antibodies by following the irreversible denaturation of scFv fusions on the surface of yeast by flow cytometry. The method was highly reproducible and correlated well with other methods used to monitor thermal denaturation of the soluble proteins. We found a range of thermal stabilities for wild-type single-chain antibodies with half-maximum denaturation temperatures between 43 and 61 degrees C. The ability to quantitate thermal stability of antibodies or other proteins that are immobilized on the surface of yeast allows rapid comparisons of primary structural information with stability. Thermal denaturation could be a useful parameter to consider in the choice of scFv fragments for various applications.     

Mutations in the S6 Gate Isolate a Late Step in the Activation Pathway and Reduce 4-AP Sensitivity in Shaker Kv Channel

K_v channels detect changes in the membrane potential via their voltage-sensing domains (VSDs) that control the status of the S6 bundle crossing (BC) gate. The movement of the VSDs results in a transfer of the S4 gating charges across the cell membrane but only the last 10–20% of the total gating charge movement is associated with BC gate opening, which involves cooperative transition(s) in the subunits. Substituting the proline residue P475 in the S6 of the Shaker channel by a glycine or alanine causes a considerable shift in the voltage-dependence of the cooperative transition(s) of BC gate opening, effectively isolating the late gating charge component from the other gating charge that originates from earlier VSD movements. Interestingly, both mutations also abolished Shaker’s sensitivity to 4-aminopyridine, which is a pharmacological tool to isolate the late gating charge component. The alanine substitution (that would promote a α-helical configuration compared to proline) resulted in the largest separation of both gating charge components; therefore, BC gate flexibility appears to be important for enabling the late cooperative step of channel opening.     

The tempo and mode of gopher mound production in a tallgrass prairie remnant

To gain better insight into how small-scale disturbances might affect ecological processes, such as the maintenance of plant species diversity, we conducted a two-year study characterizing spatio-temporal patterns of gopher mound production on a tallgrass prairie remnant located at Anderson Prairie, Iowa. USA. The locations of all newly produced gopher mounds were mapped on two 80 × 80 m permanent plots. We used these data to characterize spatio-temporal patterns of mound production across a range of scales. We found that mound production was highly clustered at scales of < 8 m over short periods of time (< 2 weeks), but shifted in location over a 3–4 weeks time period, resulting in a clustered pattern at scales of < 20 m over longer time periods (up to the 2 yr of the study). We also found that patterns of mound production at intermediate spatial scales (> 20 m) remained fairly static over time, although they differed significantly from site to site. The results of this study suggest that small-scale patterns of variability in mound production may increase habitat variability over very short spatial scales, possibly providing a mechanism that can enhance the development and maintenance of species diversity.