Desiccation induces varied responses within a soil bacterial genus.

Desiccation impacts a suite of physiological processes in microbes by elevating levels of damaging reactive oxygen species and inducing DNA strand breaks. In response to desiccation-induced stress, microbes have evolved specialized mechanisms to help them survive. Here, we performed a 128-day lab desiccation experiment on nine strains from three clades of an abundant soil bacterium, Curtobacterium. We sequenced RNA from each strain at three time points to investigate their response. Curtobacterium was highly resistant to desiccation, outlasting both Escherichia coli and a famously DNA damage-resistant bacterium, Deinococcus radiodurans. However, within the genus, there were also 10-fold differences in survival rates among strains. Transcriptomic profiling revealed responses shared within the genus including up-regulation of genes involved in DNA damage repair, osmolyte production, and efflux pumps, but also up-regulation of pathways and genes unique to the three clades. For example, trehalose synthesis gene otsB, the chaperone groEL, and the oxygen scavenger katA were all found in either one or two clades but not the third. Here, we provide evidence of considerable variation in closely related strains, and further elucidation of the phylogenetic conservation of desiccation tolerance remains an important goal for microbial ecologists.

Differential Response of Bacterial Microdiversity to Simulated Global Change.

Global change experiments often observe shifts in bacterial community composition based on 16S rRNA gene sequences. However, this genetic region can mask a large amount of genetic and phenotypic variation among bacterial strains sharing even identical 16S regions. As such, it remains largely unknown whether variation at the sub-16S level, sometimes termed microdiversity, responds to environmental perturbations and whether such changes are relevant to ecosystem processes. Here, we investigated microdiversity within Curtobacterium, the dominant bacterium found in the leaf litter layer of soil, to simulated drought and nitrogen addition in a field experiment. We first developed and validated Curtobacterium-specific primers of the groEL gene to assess microdiversity within this lineage. We then tracked the response of this microdiversity to simulated global change in two adjacent plant communities, grassland and coastal sage scrub (CSS). Curtobacterium microdiversity responded to drought but not nitrogen addition, indicating variation within the genus of drought tolerance but not nitrogen response. Further, the response of microdiversity to drought depended on the ecosystem, suggesting that litter substrate selects for a distinct composition of microdiversity that is constrained in its response, perhaps related to tradeoffs in resource acquisition traits. Supporting this interpretation, a metagenomic analysis revealed that the composition of Curtobacterium-encoded carbohydrate-active enzymes (CAZymes) varied distinctly across the two ecosystems. Identifying the degree to which relevant traits are phylogenetically conserved may help to predict when the aggregated response of a 16S-defined taxon masks differential responses of finer-scale bacterial diversity to global change. IMPORTANCE Microbial communities play an integral role in global biogeochemical cycling, but our understanding of how global change will affect microbial community structure and functioning remains limited. Microbiome analyses typically aggregate large amounts of genetic diversity which may obscure finer variation in traits. This study found that fine-scale diversity (or microdiversity) within the bacterial genus Curtobacterium was affected by simulated global changes. However, the degree to which this was true depended on the type of global change, as the composition of Curtobacterium microdiversity was affected by drought, but not by nitrogen addition. Further, these changes were associated with variation in carbon degradation traits. Future work might improve predictions of microbial community responses to global change by considering microdiversity.

Modeling wall effects in capillary electrochromatography.

A volume averaging technique for modeling electroosmotic and pressure driven flow in microchannels is applied to a packed capillary electrochromatography column. This model can be applied to fluid flow in both porous and open channels and can account for porosity variation in the column due to packing and zeta potential mismatches between the wall and the packing material. Numerical results are presented and compared with experimental results from the literature. Several different porosity models are shown to produce similar concentration profiles and allow the model to reproduce wall effects observed in experimental columns.

Dose enhancement effects to the nucleus and mitochondria from gold nanoparticles in the cytosol.

Gold nanoparticles (GNPs) have shown potential as dose enhancers for radiation therapy. Since damage to the genome affects the viability of a cell, it is generally assumed that GNPs have to localise within the cell nucleus. In practice, however, GNPs tend to localise in the cytoplasm yet still appear to have a dose enhancing effect on the cell. Whether this effect can be attributed to stress-induced biological mechanisms or to physical damage to extra-nuclear cellular targets is still unclear. There is however growing evidence to suggest that the cellular response to radiation can also be influenced by indirect processes induced when the nucleus is not directly targeted by radiation. The mitochondrion in particular may be an effective extra-nuclear radiation target given its many important functional roles in the cell. To more accurately predict the physical effect of radiation within different cell organelles, we measured the full chemical composition of a whole human lymphocytic JURKAT cell as well as two separate organelles; the cell nucleus and the mitochondrion. The experimental measurements found that all three biological materials had similar ionisation energies ∼70 eV, substantially lower than that of liquid water ∼78 eV. Monte Carlo simulations for 10-50 keV incident photons showed higher energy deposition and ionisation numbers in the cell and organelle materials compared to liquid water. Adding a 1% mass fraction of gold to each material increased the energy deposition by a factor of ∼1.8 when averaged over all incident photon energies. Simulations of a realistic compartmentalised cell show that the presence of gold in the cytosol increases the energy deposition in the mitochondrial volume more than within the nuclear volume. We find this is due to sub-micron delocalisation of energy by photoelectrons, making the mitochondria a potentially viable indirect radiation target for GNPs that localise to the cytosol.

Quadriceps and hamstrings peak torque ratio changes in persons with chronic anterior cruciate ligament deficiency.

STUDY DESIGN: Cross-sectional analytical study in which subjects served as their own controls. OBJECTIVE: To assess the concentric and eccentric peak torque in the hamstrings and quadriceps muscles, hamstrings/quadriceps amplitude ratios, and lean thigh volume differences in the involved and uninvolved limb of subjects with anterior cruciate ligament (ACL) deficiency. BACKGROUND: [corrected] Although the hamstrings/quadriceps ratios for concentric and eccentric activity have been studied, the more functional eccentric hamstings/concentric quadriceps ratio has not been previously described in chronic ACL deficient individuals. METHODS AND MEASURES: Eighteen subjects (36 +/- 11 years; 12 men, 6 women) with unilateral chronic ACL deficiency were recruited. Changes in activities of daily living, lean thigh volume (LTV) and isokinetic peak torque and total work capacity of both the quadriceps and hamstring muscles were investigated. The uninvolved limb served as control. RESULTS: Eccentric quadriceps peak torque in the ACL deficient limb was reduced by 38% compared with the 16% reduction of concentric quadriceps peak torque. Eccentric hamstrings peak torque was reduced by 15% compared with an 8% reduction in concentric hamstrings peak torque. LTV in the ACL deficient limb was 11% smaller than the uninvolved limb (3,541 +/- 899 vs 3,161 +/- 742 cc, uninvolved versus ACL deficient limb). The relation between LTV and eccentric peak torque was stronger in the uninvolved (r = 0.82) compared with the ACL deficient limb (r = 0.66). The eccentric hamstrings/quadriceps ratio was significantly higher in the ACL deficient (0.80 +/- 0.26) compared with the uninvolved limb (0.55 +/- 0.13). However, the eccentric hamstrings/concentric quadriceps ratio was similar between ACL deficient (0.75 +/- 0.17) and uninvolved (0.77 +/- 0.20) limbs, while the concentric hamstrings/eccentric quadriceps ratio was significantly higher in the ACL deficient (0.79 +/- 0.26) compared with the uninvolved (0.50 +/- 0.14) limb. CONCLUSIONS: Quadriceps and hamstrings peak torque values were significantly decreased in ACL deficient compared with the uninvolved limb. Eccentric muscle activity was affected to a greater degree than concentric muscle activity in the quadriceps muscle after ACL injury. The eccentric hamstrings/concentric quadriceps ratios were similar in ACL deficient and normal limbs, indicating that muscular co-ordination strategies may have been altered to maintain normal limb activity despite the strength losses particularly evident in quadriceps muscle function.

Automated image segmentation for breast analysis using infrared images.

In order to realize a fully automated thermogram analysis package for breast cancer detection, it is necessary to identify the region of interest in the thermal image prior to analysis. A nearly fully automated approach is outlined that is able to successfully locate the breast regions in most of the images analyzed. The approach consists of a sequence of Canny edge detectors to determine the body boundaries and to isolate the most likely candidates for the bottom breast boundary. Three different strategies for identifying the bottom breast boundary are investigated: a variation of the Hough transform to identify the curved edges in the image, an algorithm used to detect the longest connected edges that are not part of the body boundary, and a third approach involving the density of detected edges in the breast region. The last two methods show great promise in successfully segmenting the breasts.

Generation of transgenic porcine chimeras using primordial germ cell-derived colonies.

In mice, two pluripotent cell lines, embryonic stem (ES) cells and embryonic germ (EG) cells, have been identified. We present here results indicating that porcine EG cell lines can be isolated, genetically transformed, and utilized to make transgenic chimeras. Briefly, primordial germ cells (PGCs) were isolated from Day 25-27 fetuses and plated on STO feeder cells in Dulbecco's modified Eagle's medium:Ham's F-10 medium supplemented with 0.01 mM nonessential amino acids, 2 mM glutamine, 15% fetal bovine serum, 0.1 mM 2-mercaptoethanol, 40 ng/ml human stem cell factor, 20 ng/ml human basic fibroblast growth factor, and 20 ng/ml human leukemia inhibitory factor. For genetic transformation, cells were electroporated with a construct containing the green fluorescent protein under control of the cytomegalovirus promoter. After electroporation, cells were plated and later examined under fluorescein isothiocyanate excitation. Fluorescent colonies were selected for chimera generation. Blastocysts collected from gilts on Day 5 were injected with 10-15 transgenic PGC-derived cells and transferred into recipient gilts. Gilts were hysterectomized on Day 25, and fetal tissues were analyzed by Southern blotting. Three chimeras out of 20 fetuses analyzed were transgenic. Additionally, when one recipient gilt was allowed to go to term, one piglet with transgenic contribution was identified.