GBSX: a toolkit for experimental design and demultiplexing genotyping by sequencing experiments

Background

Massive parallel sequencing is a powerful tool for variant discovery and genotyping. To reduce costs, sequencing of restriction enzyme based reduced representation libraries can be utilized. This technology is generally referred to as Genotyping By Sequencing (GBS). To deal with GBS experimental design and initial processing specific bioinformatic tools are needed.

Results

GBSX is a package that assists in selecting the appropriate enzyme and the design of compatible in-line barcodes. Post sequencing, it performs optimized demultiplexing using these barcodes to create fastq files per barcode which can easily be plugged into existing variant analysis pipelines. Here we demonstrate the usability of the GBSX toolkit and demonstrate improved in-line barcode demultiplexing and trimming performance compared to existing tools.

Conclusions

GBSX provides an easy to use suite of tools for designing and demultiplexing of GBS experiments.     

Habitat Interference by Axis Deer on White-Tailed Deer

Abstract: Many studies of interactions between exotic and native ungulates have not had temporal and spatial controls nor have they considered the types of competitive interactions that would allow coexistence. For exotic axis deer (Axis axis) and native white-tailed deer (Odocoileus virginianus) to coexist one species should be superior at interference competition and the other species should be superior at exploitative competition. We generated and tested predictions, based on body size and diet breadth, about habitat selection by white-tailed deer in the presence and absence of axis deer, dominance relationships, and time at sites provisioned with high quality forage. We conducted our study in treatment (axis and white-tailed deer) and control (white-tailed deer only) areas when both species were present and after axis deer were removed. We conducted vehicle surveys to determine habitat use of both species. At provisioned feeding sites we recorded aggressive behaviors and amount of time species spent at feeding sites alone and together. In the treatment area white-tailed deer selection for wooded habitat increased 2.1 times after axis deer were removed, whereas habitat selection by white-tailed deer was constant in the control area over the same time. At feeding sites axis deer were dominant to white-tailed deer; both species spent a significantly greater amount of time alone than at feeders together, but amount of time that individuals of each species spent at feeders did not differ. Axis deer were superior at interference competition, but white-tailed deer were not superior at exploitative competition; thus, species coexistence is unlikely. Whether white-tailed deer are negatively impacted by axis deer at spatial scales larger than our experiment probably depends on abundance of axis deer at larger spatial scales. Experiments of species interactions with temporal and spatial controls that consider types of competitive interactions increase a manager's understanding of when and how native ungulates may be negatively impacted by exotic ungulates.     

<Emphasis Type="Italic">Chlamydia pneumoniae</Emphasis> induces aponecrosis in human aortic smooth muscle cells

Background  

The intracellular bacterium Chlamydia pneumoniae is suspected to play a role in formation and progression of atherosclerosis. Many studies investigated cell death initiation versus inhibition by Chlamydia pneumoniae in established cell lines but nothing is known in primary human aortic smooth muscle cells, a cell type among others known to be involved in the formation of the atherosclerotic plaque. Type of cell death was analyzed by various methods in primary aortic smooth muscle cells after infection with Chlamydia pneumoniae to investigate a possible pathogenic link in atherosclerosis.     

Refinements of and commentary on the silver staining techniques of Fernández-Galiano

The original ammoniacal silver carbonate staining technique and subsequent modification developed by Fernández-Galiano are useful for investigating ciliate protozoan systematics and/or ciliate cortical structure and morphogenesis. The technique is complicated, however, by both uncertainties arising from the need to count drops of reagents and subjective control of the staining intensity. I have resolved these complications by defining volumes of reagents rather than using drops and by defining a range of staining times. I also comment on various steps of the techniques. My techniques are simplified and refined to produce consistent, high quality staining results.     

Using Genetic Tools to Track Desert Bighorn Sheep Colonizations

ABSTRACT Understanding colonization is vital for managing fragmented populations. We employed mitochondrial DNA haplotypes and 14 microsatellite (nuclear DNA) markers to infer the origins of newly established populations of desert bighorn sheep (Ovis canadensis nelsoni) and to assess loss of genetic diversity during natural colonizations. We used haplotype distribution, F-statistics, Bayesian population clustering, and assignment tests to infer source populations for 3 recent colonies and identified a previously undetected colonization from multiple source populations. Allelic richness declined in 3 of 4 colonies in comparison to the primary source populations, but not as much as has been reported for translocated populations. Heterozygosity declined in only one colony. We also demonstrated that both native and translocated desert bighorn sheep have naturally recolonized empty habitats and suggest that colonization may partially offset population extinction in the region as long as connectivity is maintained. Genetic techniques and mitochondrial DNA haplotypes we described will allow managers to determine the origins of future colonizations by bighorn sheep in California, USA, and prioritize protection of linkages between known sources and colonies.     

Sublethal damage during cryopreservation of rainbow trout sperm

Although cellular damage during cryopreservation of freshwater fish spermatozoa has been reported in several studies, there is a lack of correlation between this damage and the fertility rates of eggs using postthawed milt. The apparent lack of such correlation may be due to other undetected sublethal cryodamage, which could affect the cell functionality and viability. This may be extremely important for freshwater fish spermatozoa whose ability to fertilize the egg requires dilution in water or hypoosmotic solutions, an hazardous environment for the cells. This study tested the change in cell permeability during cryopreservation, using Hoechst 33258 to assess cell permeability. The permeability of spermatozoa at different times after dilution in several hypoosmotic media were investigated. In the first trial, fresh semen, sperm diluted in freezing media (CPT), and freeze/thawed semen were studied. Three CPT were tested (Me2SO, DMA, and methanol). In the second trial, the addition of egg yolk as a membrane stabilizer was investigated. Samples were frozen at -20 degreesC/min in a programmable cooler and thawed in a 25 degreesC water bath. Dilution in the CPTs slightly increased the susceptibility of cells to damage but freezing/thawing caused a dramatic increase in the fragility of cells, which were killed in a few seconds after their contact with the hypoosmotic solutions. Egg yolk provided a significant protection to the membrane, allowing the cells a greater and more prolonged survival in the fertilization media. Samples frozen with Me2SO displayed the best results. These results are consistent with the achieved fertility rates that demonstrated sublethal cryodamage in the function of the sperm membrane that was not detected by standard procedures. Copyright 1998 Academic Press.