The genetic basis of energy conservation in the sulfate-reducing bacterium Desulfovibrio alaskensis G20.

Sulfate-reducing bacteria play major roles in the global carbon and sulfur cycles, but it remains unclear how reducing sulfate yields energy. To determine the genetic basis of energy conservation, we measured the fitness of thousands of pooled mutants of Desulfovibrio alaskensis G20 during growth in 12 different combinations of electron donors and acceptors. We show that ion pumping by the ferredoxin:NADH oxidoreductase Rnf is required whenever substrate-level phosphorylation is not possible. The uncharacterized complex Hdr/flox-1 (Dde_1207:13) is sometimes important alongside Rnf and may perform an electron bifurcation to generate more reduced ferredoxin from NADH to allow further ion pumping. Similarly, during the oxidation of malate or fumarate, the electron-bifurcating transhydrogenase NfnAB-2 (Dde_1250:1) is important and may generate reduced ferredoxin to allow additional ion pumping by Rnf. During formate oxidation, the periplasmic [NiFeSe] hydrogenase HysAB is required, which suggests that hydrogen forms in the periplasm, diffuses to the cytoplasm, and is used to reduce ferredoxin, thus providing a substrate for Rnf. During hydrogen utilization, the transmembrane electron transport complex Tmc is important and may move electrons from the periplasm into the cytoplasmic sulfite reduction pathway. Finally, mutants of many other putative electron carriers have no clear phenotype, which suggests that they are not important under our growth conditions, although we cannot rule out genetic redundancy

Functional genomics with a comprehensive library of transposon mutants for the sulfate-reducing bacterium Desulfovibrio alaskensis G20.

UnlabelledThe genomes of sulfate-reducing bacteria remain poorly characterized, largely due to a paucity of experimental data and genetic tools. To meet this challenge, we generated an archived library of 15,477 mapped transposon insertion mutants in the sulfate-reducing bacterium Desulfovibrio alaskensis G20. To demonstrate the utility of the individual mutants, we profiled gene expression in mutants of six regulatory genes and used these data, together with 1,313 high-confidence transcription start sites identified by tiling microarrays and transcriptome sequencing (5' RNA-Seq), to update the regulons of Fur and Rex and to confirm the predicted regulons of LysX, PhnF, PerR, and Dde_3000, a histidine kinase. In addition to enabling single mutant investigations, the D. alaskensis G20 transposon mutants also contain DNA bar codes, which enables the pooling and analysis of mutant fitness for thousands of strains simultaneously. Using two pools of mutants that represent insertions in 2,369 unique protein-coding genes, we demonstrate that the hypothetical gene Dde_3007 is required for methionine biosynthesis. Using comparative genomics, we propose that Dde_3007 performs a missing step in methionine biosynthesis by transferring a sulfur group to O-phosphohomoserine to form homocysteine. Additionally, we show that the entire choline utilization cluster is important for fitness in choline sulfate medium, which confirms that a functional microcompartment is required for choline oxidation. Finally, we demonstrate that Dde_3291, a MerR-like transcription factor, is a choline-dependent activator of the choline utilization cluster. Taken together, our data set and genetic resources provide a foundation for systems-level investigation of a poorly studied group of bacteria of environmental and industrial importance.ImportanceSulfate-reducing bacteria contribute to global nutrient cycles and are a nuisance for the petroleum industry. Despite their environmental and industrial significance, the genomes of sulfate-reducing bacteria remain poorly characterized. Here, we describe a genetic approach to fill gaps in our knowledge of sulfate-reducing bacteria. We generated a large collection of archived, transposon mutants in Desulfovibrio alaskensis G20 and used the phenotypes of these mutant strains to infer the function of genes involved in gene regulation, methionine biosynthesis, and choline utilization. Our findings and mutant resources will enable systematic investigations into gene function, energy generation, stress response, and metabolism for this important group of bacteria

Effect of temperature on the rate of ageing : an experimental study of the Blowfly Calliphora stygia

All organisms age, the rate of which can be measured by demographic analysis of mortality rates. The rate of ageing is thermally sensitive in ectothermic invertebrates and we examined the effects of temperature on both demographic rates of ageing and on cellular senescence in the blowfly, Calliphora stygia. The short lifespan of these flies is advantageous for demographic measurements while their large body size permits individual-based biochemical characterisation. Blowflies maintained at temperatures from 12°C to 34°C had a five to six-fold decrease in maximum and average longevity, respectively. Mortality rates were best described by a two-phase Gompertz relation, which revealed the first-phase of ageing to be much more temperature sensitive than the second stage. Flies held at low temperatures had both a slower first-phase rate of ageing and a delayed onset of second-phase ageing, which significantly extended their longevity compared with those at high temperatures. Blowflies that were transferred from 29°C to 15°C had higher first-phase mortality rates than those of flies held at constant 15°C, but their onset of second-phase ageing was deferred beyond that of flies held constantly at this temperature. The accumulation of fluorescent AGE pigment, a measure of cellular oxidative damage, increased steadily over time in all blowflies, irrespective of the temporal pattern of mortality. Pigment accumulated steadily during periods of ‘negligible senescence’, as measured by minimal rate of mortality, and the rate of accumulation was significantly affected by temperature. Thus accumulation of AGE pigment is more representative of chronological age than a reflection of biological age or a cause of mortality

CELLO: A fast algorithm for Covariance Estimation

We present CELLO (Covariance Estimation and Learning through Likelihood Optimization), an algorithm for predicting the covariances of measurements based on any available informative features. This algorithm is intended to improve the accuracy and reliability of on-line state estimation by providing a principled way to extend the conventional fixed-covariance Gaussian measurement model. We show that in experiments, CELLO learns to predict measurement covariances that agree with empirical covariances obtained by manually annotating sensor regimes. We also show that using the learned covariances during filtering provides substantial quantitative improvement to the overall state estimate. © 2013 IEEE.United States. National Aeronautics and Space AdministrationSiemens Corporate ResearchUnited States. Office of Naval Research. Multidisciplinary University Research InitiativeMicro Autonomous Consortium Systems and Technolog

Dissecting a complex chemical stress: chemogenomic profiling of plant hydrolysates.

The efficient production of biofuels from cellulosic feedstocks will require the efficient fermentation of the sugars in hydrolyzed plant material. Unfortunately, plant hydrolysates also contain many compounds that inhibit microbial growth and fermentation. We used DNA-barcoded mutant libraries to identify genes that are important for hydrolysate tolerance in both Zymomonas mobilis (44 genes) and Saccharomyces cerevisiae (99 genes). Overexpression of a Z. mobilis tolerance gene of unknown function (ZMO1875) improved its specific ethanol productivity 2.4-fold in the presence of miscanthus hydrolysate. However, a mixture of 37 hydrolysate-derived inhibitors was not sufficient to explain the fitness profile of plant hydrolysate. To deconstruct the fitness profile of hydrolysate, we profiled the 37 inhibitors against a library of Z. mobilis mutants and we modeled fitness in hydrolysate as a mixture of fitness in its components. By examining outliers in this model, we identified methylglyoxal as a previously unknown component of hydrolysate. Our work provides a general strategy to dissect how microbes respond to a complex chemical stress and should enable further engineering of hydrolysate tolerance

Hip Strength Disparities in College Soccer Players: Implications for Injury Risk and Conditioning Practices

The Y-balance Test (YBT) has been used to identify potential risk factors associated with lower extremity injuries (LEI), while mobility and strength assessments are often used to monitor progress following an injury. PURPOSE: This study aimed to explore differences in YBT scores along with lower body mobility and strength in Division I male soccer players who have reported a LEI in the last 12 months (LEI-Y) compared those without reported injuries (LEI-N). METHODS: Twenty-eight male soccer players (age: 20.7 ± 1.8 y; height: 183.8 ± 7.1 cm; mass: 78.5 ± 6.7 kg) completed pre-season evaluations which included YBT, hip strength, hamstring flexibility, and hip and ankle range of motion (ROM). All players were cleared for testing by the head athletic trainer and team physician and had no current LEI precluding them from participating. However, players self-reported if they had experienced a lower extremity injury or surgery in the prior twelve months. Hip adduction and abduction strength was evaluated using a dynamometer and calculated relative to body weight. Hip and ankle ROM were measured using a goniometer, and hamstring flexibility was measured using the sit-and-reach test. The YBT scores and leg length were used to determine a composite score for each leg. Data was presented as means ± standard deviations. RESULTS: For LEI-N, there were no notable imbalances in right to left side comparisons for hip adduction strength (44.8 ± 6.8 vs 43.5 ± 7.5 %) or hip abduction strength (49.2 ± 5.3 vs 49.0 ± 9.1 %). In the LEI-Y group, there was a discernible difference between the right and left side for hip adduction strength (43.7 ± 7.8 vs 41.4 ± 6.1 %) and hip abduction strength (48.4 ± 8.3 vs 45.1 ± 6.3 %). Relative hip strength and ankle ROM scores were also lower overall in the LEI-Y group compared to the LEI-N group, but no such trends were observed for YBT composite scores, hip ROM, or hamstring flexibility. CONCLUSION: The findings of this study indicate that the LEI-Y group had greater bilateral imbalance in lower limb strength, lower hip strength, and lower ankle ROM scores compared with those without injury history. Future research is recommended to evaluate the impact of injury on these markers and examine their associations with future injury risk

Beyond a warming fingerprint: individualistic biogeographic responses to heterogeneous climate change in California.

Understanding recent biogeographic responses to climate change is fundamental for improving our predictions of likely future responses and guiding conservation planning at both local and global scales. Studies of observed biogeographic responses to 20th century climate change have principally examined effects related to ubiquitous increases in temperature - collectively termed a warming fingerprint. Although the importance of changes in other aspects of climate - particularly precipitation and water availability - is widely acknowledged from a theoretical standpoint and supported by paleontological evidence, we lack a practical understanding of how these changes interact with temperature to drive biogeographic responses. Further complicating matters, differences in life history and ecological attributes may lead species to respond differently to the same changes in climate. Here, we examine whether recent biogeographic patterns across California are consistent with a warming fingerprint. We describe how various components of climate have changed regionally in California during the 20th century and review empirical evidence of biogeographic responses to these changes, particularly elevational range shifts. Many responses to climate change do not appear to be consistent with a warming fingerprint, with downslope shifts in elevation being as common as upslope shifts across a number of taxa and many demographic and community responses being inconsistent with upslope shifts. We identify a number of potential direct and indirect mechanisms for these responses, including the influence of aspects of climate change other than temperature (e.g., the shifting seasonal balance of energy and water availability), differences in each taxon's sensitivity to climate change, trophic interactions, and land-use change. Finally, we highlight the need to move beyond a warming fingerprint in studies of biogeographic responses by considering a more multifaceted view of climate, emphasizing local-scale effects, and including a priori knowledge of relevant natural history for the taxa and regions under study

Novel actions of next-generation taxanes benefit advanced stages of prostate cancer.

PURPOSE: To improve the outcomes of patients with castration-resistant prostate cancer (CRPC), there is an urgent need for more effective therapies and approaches that individualize specific treatments for patients with CRPC. These studies compared the novel taxane cabazitaxel with the previous generation docetaxel, and aimed to determine which tumors are most likely to respond. EXPERIMENTAL DESIGN: Cabazitaxel and docetaxel were compared via in vitro modeling to determine the molecular mechanism, biochemical and cell biologic impact, and cell proliferation, which was further assessed ex vivo in human tumor explants. Isogenic pairs of RB knockdown and control cells were interrogated in vitro and in xenograft tumors for cabazitaxel response. RESULTS: The data herein show that (i) cabazitaxel exerts stronger cytostatic and cytotoxic response compared with docetaxel, especially in CRPC; (ii) cabazitaxel induces aberrant mitosis, leading to pyknotic and multinucleated cells; (iii) taxanes do not act through the androgen receptor (AR); (iv) gene-expression profiling reveals distinct molecular actions for cabazitaxel; and (v) tumors that have progressed to castration resistance via loss of RB show enhanced sensitivity to cabazitaxel. CONCLUSIONS: Cabazitaxel not only induces improved cytostatic and cytotoxic effects, but also affects distinct molecular pathways, compared with docetaxel, which could underlie its efficacy after docetaxel treatment has failed in patients with CRPC. Finally, RB is identified as the first potential biomarker that could define the therapeutic response to taxanes in metastatic CRPC. This would suggest that loss of RB function induces sensitization to taxanes, which could benefit up to 50% of CRPC cases

3D-Image Analysis of the Impact of Toothpaste Abrasivity on the Progression of Simulated Non-Carious Cervical Lesions

Objectives To investigate the effect of toothpaste abrasive level on the progression of non-carious cervical lesions (NCCLs) using 3D-image subtraction. Methods Upper first premolars were allocated into seven groups (n = 16) of toothpaste/abrasive slurries: A-Zeodent113/5%, B-Zeodent124/10%, C-Zeodent103/15%, D-Sensodyne Pronamel, E-Crest Cavity-Protection, F-Crest Pro-Health-Whitening, and G-Deionized water (DIW). Teeth were mounted on acrylic blocks, and their root surfaces covered with acrylic resin, except for 2-mm near the cemento-enamel junction that was exposed to toothbrushing. Specimens were brushed with the slurries for 5000-, 15,000-, 35,000- and 65,000-strokes. Impressions were taken at baseline and after each brushing time, and then scanned by a 3D optical profilometer. Dentine volume loss was calculated by image subtraction software and subjected to mixed-model ANOVA and multiple comparison tests (α = 0.05). Results No significant differences among slurries were observed at 5000 and 15,000. At 35,000, F showed higher loss than all other groups except C, which did not differ from the others. At 65,000, F (4.19 ± 3.29 mm3) showed the highest loss, followed by C (2.33 ± 1.47 mm3), which differed from all the other groups except B (1.85 ± 0.91 mm3). Groups B, A (1.35 ± 0.65 mm3), D (1.17 ± 0.48 mm3), E (1.40 ± 0.68 mm3) and G (1.12 ± 0.73 mm3) did not differ from each other. Groups F and C showed significant increase of volume loss starting at 35,000, while B, A, D and E only at 65,000; no increase loss was observed for G. Conclusions 3D-image subtraction was able to quantify and differentiate tooth loss, but only at advanced stages. The progression of NCCLs was more evident and faster for highly abrasive slurries. Clinical significance Upon root dentin exposure, brushing with lower abrasive dentifrices is advisable to reduce the risk for NCCLs development