Water temperature affects the biotransformation and accumulation of a psychoactive pharmaceutical and its metabolite in aquatic organisms.

Pharmaceutically active compounds (PhACs) have been shown to accumulate in aquatic and riparian food-webs. Yet, our understanding of how temperature, a key environmental factor in nature, affects uptake, biotransformation, and the subsequent accumulation of PhACs in aquatic organisms is limited. In this study, we tested to what extent bioconcentration of an anxiolytic drugs (temazepam and oxazepam) is affected by two temperature regimes (10 and 20 °C) and how the temperature affects the temazepam biotransformation and subsequent accumulation of its metabolite (oxazepam) in aquatic organisms. We used European perch (Perca fluviatilis) and dragonfly larvae (Sympetrum sp.), which represent predator and prey species of high ecological relevance in food chains of boreal and temperate aquatic ecosystems. Experimental organisms were exposed to target pharmaceuticals at a range of concentrations (0.2-6 µg L-1) to study concentration dependent differences in bioconcentration and biotransformation. We found that the bioconcentration of temazepam in perch was significantly reduced at higher temperatures. Also, temperature had a strong effect on temazepam biotransformation in the fish, with the production and subsequent accumulation of its metabolite (oxazepam) being two-fold higher at 20 °C compared to 10 °C. In contrast, we found no temperature dependency for temazepam bioconcentration in dragonfly larvae and no detectable biotransformation of the parent compound that would result in measurable concentrations of oxazepam in this organism. Our results highlight that while organisms may share the same aquatic ecosystem, their exposure to PhACs may change differently across temperature gradients in the environment.

The cost-effectiveness of banning highly hazardous pesticides to prevent suicides due to pesticide self-ingestion across 14 countries: an economic modelling study.

BACKGROUND: Reducing suicides is a key Sustainable Development Goal target for improving global health. Highly hazardous pesticides are among the leading causes of death by suicide in low-income and middle-income countries. National bans of acutely toxic highly hazardous pesticides have led to substantial reductions in pesticide-attributable suicides across several countries. This study evaluated the cost-effectiveness of implementing national bans of highly hazardous pesticides to reduce the burden of pesticide suicides. METHODS: A Markov model was developed to examine the costs and health effects of implementing a national ban of highly hazardous pesticides to prevent suicides due to pesticide self-poisoning, compared with a null comparator. We used WHO cost-effectiveness and strategic planning (WHO-CHOICE) methods to estimate pesticide-attributable suicide rates for 100 years from 2017. Country-specific costs were obtained from the WHO-CHOICE database and denominated in 2017 international dollars (I$), discounted at a 3% annual rate, and health effects were measured in healthy life-years gained (HLYGs). We used a demographic projection model beginning with the country population in the baseline year (2017), split by 1-year age group and sex. Country-specific data on overall suicide rates were obtained for 2017 by age and sex from the Global Burden of Disease Study 2017 Data Resources. The analysis involved 14 countries spanning low-income to high-income settings, and cost-effectiveness ratios were analysed at the country-specific level and aggregated according to country income group and the proportion of suicides due to pesticides. FINDINGS: Banning highly hazardous pesticides across the 14 countries studied could result in about 28 000 (95% uncertainty interval [UI] 24 000-32 000) fewer suicide deaths each year at an annual cost of I$0·007 per capita (95% UI 0·006-0·008). In the population-standardised results for the base case analysis, national bans produced cost-effectiveness ratios of $94 per HLYG (95% UI 73-123) across low-income and lower-middle-income countries and $237 per HLYG (95% UI 191-303) across upper-middle-income and high-income countries. Bans were more cost-effective in countries where a high proportion of suicides are attributable to pesticide self-poisoning, reaching a cost-effectiveness ratio of $75 per HLYG (95% UI 58-99) in two countries with proportions of more than 30%. INTERPRETATION: National bans of highly hazardous pesticides are a potentially cost-effective and affordable intervention for reducing suicide deaths in countries with a high burden of suicides attributable to pesticides. However, our study findings are limited by imperfect data and assumptions that could be improved upon by future studies. FUNDING: WHO.

Voluntary intake and digestibility by sheep of alfalfa ensiled at different moisture concentrations following fertilization with dairy slurry.

Dairy slurry is used commonly as an animal-sourced fertilizer in agronomic production. However, residual effects of slurry application on intake and digestibility of alfalfa (Medicago sativa L.) silage from subsequent harvests are not well known. The objective of this study was to determine if moisture concentration of alfalfa silage and timing of dairy slurry application relative to subsequent harvest affected intake and digestibility by sheep. Katahdin crossbred ewes (n = 18; 48 ± 5.3 kg) in mid-gestation were stratified by BW and allocated randomly in each of two periods to one of six treatments arranged in a two × three factorial arrangement. Treatments consisted of recommended (RM; 46.8%) or low (LM; 39.7%) moisture at baling after either no slurry application (NS), slurry application to stubble immediately after removal of the previous cutting (S0), or slurry application 14 d after removal of the previous cutting (S14). Silages were chopped through a commercial straw chopper, packed into plastic trash cans, and then offered to ewes within 4 d of chopping. Period 1 of the intake and digestion study consisted of a 14-d adaptation followed by a 7-d fecal collection period. Period 2 followed period 1 after a 4-d rest and consisted of an 11-d adaptation followed by 7 d of fecal collection. Ewes were housed individually in 1.4 × 4.3-m pens equipped with rubber mat flooring. Feces were swept from the floor twice daily, weighed, and dried at 50 °C. Ewes had ad libitum access to water and were offered chopped silage for a minimum of 10% refusal (DM). Blood samples were collected immediately prior to feeding, and 4 and 8 h after feeding on the day prior to the end of each period. Organic matter intake (g/kg BW) and OM digestibility tended (P < 0.10) to be, and digestible OM intake (g/kg BW) was reduced by slurry application. Lymphocytes (% of total white blood cells) were greater (P < 0.05) from LM vs. RM and from NS vs. S0 and S14. Red blood cell concentrations were greater (P < 0.05) from S14 vs. S0 and from S0 and S14 vs. NS. Serum urea N concentrations did not differ (P > 0.17) across treatments. Therefore, moisture concentration of alfalfa silage within the range used in this study may not affect voluntary intake or digestibility, but slurry application may have an effect on digestible OM intake. Also, moisture concentration of alfalfa silage and time of dairy slurry application may affect specific blood hemograms.

Genetic diversity and comparison of diagnostic tests for characterization of foot-and-mouth disease virus strains from Pakistan 2008-2012.

We report the laboratory analysis of 125 clinical samples from suspected cases of foot-and-mouth disease (FMD) in cattle and Asian buffalo collected in Pakistan between 2008 and 2012. Of these samples, 89 were found to contain viral RNA by rRT-PCR, of which 88 were also found to contain infectious FMD virus (FMDV) by virus isolation (VI), with strong correlation between these tests (κ = 0.96). Samples that were VI-positive were serotyped by antigen detection ELISA (Ag-ELISA) and VP1 sequence acquisition and analysis. Sequence data identified FMDV serotypes A (n = 13), O (n = 36) and Asia-1 (n = 41), including three samples from which both serotypes Asia-1 and O were detected. Serotype A viruses were classified within three different Iran-05 sublineages: HER-10, FAR-11 and ESF-10. All serotype Asia-1 were within Group VII (Sindh-08 lineage), in a genetic clade that differs from viruses isolated prior to 2010. All serotypes O were classified as PanAsia-2 within two different sublineages: ANT-10 and BAL-09. Using VP1 sequencing as the gold standard for serotype determination, the overall sensitivity of Ag-ELISA to correctly determine serotype was 74%, and serotype-specific sensitivity was 8% for serotype A, 88% for Asia-1 and 89% for O. Serotype-specific specificity was 100% for serotype A, 93% for Asia-1 and 94% for O. Interestingly, 12 of 13 serotype A viruses were not detected by Ag-ELISA. This study confirms earlier accounts of regional genetic diversity of FMDV in Pakistan and highlights the importance of continued validation of diagnostic tests for rapidly evolving pathogens such as FMDV.

[Prolonged weaning during early neurological and neurosurgical rehabilitation : S2k guideline published by the Weaning Committee of the German Neurorehabilitation Society (DGNR)]. / Prolongiertes Weaning in der neurologisch-neurochirurgischen Frührehabilitation : S2k-Leitlinie herausgegeben von der Weaning-Kommission der Deutschen Gesellschaft für Neurorehabilitation e. V. (DGNR).

Prolonged weaning of patients with neurological or neurosurgery disorders is associated with specific characteristics, which are taken into account by the German Society for Neurorehabilitation (DGNR) in its own guideline. The current S2k guideline of the German Society for Pneumology and Respiratory Medicine is referred to explicitly with regard to definitions (e.g., weaning and weaning failure), weaning categories, pathophysiology of weaning failure, and general weaning strategies. In early neurological and neurosurgery rehabilitation, patients with central of respiratory regulation disturbances (e.g., cerebral stem lesions), swallowing disturbances (neurogenic dysphagia), neuromuscular problems (e.g., critical illness polyneuropathy, Guillain-Barre syndrome, paraplegia, Myasthenia gravis) and/or cognitive disturbances (e.g., disturbed consciousness and vigilance disorders, severe communication disorders), whose care during the weaning of ventilation requires, in addition to intensive medical competence, neurological or neurosurgical and neurorehabilitation expertise. In Germany, this competence is present in centers of early neurological and neurosurgery rehabilitation, as a hospital treatment. The guideline is based on a systematic search of guideline databases and MEDLINE. Consensus was established by means of a nominal group process and Delphi procedure moderated by the Association of the Scientific Medical Societies in Germany (AWMF). In the present guideline of the DGNR, the special structural and substantive characteristics of early neurological and neurosurgery rehabilitation and existing studies on weaning in early rehabilitation facilities are examined.Addressees of the guideline are neurologists, neurosurgeons, anesthesiologists, palliative physicians, speech therapists, intensive care staff, ergotherapists, physiotherapists, and neuropsychologists. In addition, this guideline is intended to provide information to specialists for physical medicine and rehabilitation (PMR), pneumologists, internists, respiratory therapists, the German Medical Service of Health Insurance Funds (MDK) and the German Association of Health Insurance Funds (MDS). The main goal of this guideline is to convey the current knowledge on the subject of "Prolonged weaning in early neurological and neurosurgery rehabilitation".

Outcome of neurological early rehabilitation patients carrying multi-drug resistant bacteria: results from a German multi-center study.

BACKGROUND: Colonization or infection with multi-drug resistant (MDR) bacteria is considered detrimental to the outcome of neurological and neurosurgical early rehabilitation patients. METHODS: In a German multi-center study, 754 neurological early rehabilitation patients were enrolled and and reviewed in respect to MDR status, length of stay (LOS) and the following outcome variables: Barthel Index (BI), Early Rehabilitation Index (ERI), Glasgow Outcome Score Extended (GOSE), Coma Remission Scale (CRS), Functional Ambulation Categories (FAC). RESULTS: The mean age of the study population was 68.0 ± 14.8 years. Upon admission, the following prevalence for MDRs was observed: MRSA (methicillin resistant staphylococcus aureus) 7.0% (53/754), ESBL- (extended spectrum beta-lactamase) producing bacteria strains 12.6% (95/754), VRE (vancomycin resistant enterococci) 2.8% (21/754). Patients colonized or infected with MDR bacteria (MDR+) were significantly more frequently diagnosed with a critical illness polyneuropathy - CIP - than non-colonized (MDR-) patients: 29.0% vs. 14.8%. In addition, they were more frequently mechanically ventilated (MDR+: 55/138, 39.9%; MDR- 137/616, 22.2%). MDR+ patients were referred to rehabilitation earlier, had a longer LOS in early rehabilitation, lower BI on admission and at discharge, lower ERI on admission and lower CRS at discharge than MDR- patients. There was a highly significant correlation of the BI upon admission with the BI at discharge (rs = 0.492, p < 0.001). GOSE at discharge differed significantly between both groups (χ 2-test, p < 0.01). Perhaps of greatest importance, mortality among MDR+ was higher in comparison to MDR- (18.1% vs. 7.6%). CONCLUSIONS: The outcome of neurological early rehabilitation patients colonized or infected with MDR bacteria including MRSA or ESBL producing strains is significantly poorer than by non-colonized patients. There is some evidence that the poor outcome could be related to the higher morbidity and lower functional status upon admission.

[Efficacy of early neurological and neurosurgical rehabilitation : Evidence-based treatment, outcome and prognostic factors]. / Wirksamkeit der neurologisch-neurochirurgischen Frührehabilitation : Evidenzbasierte Therapieformen, Outcome und Prognosefaktoren.

BACKGROUND: Early neurological and neurosurgical rehabilitation (ENNR) as a complex post-acute form of treatment for patients with severe neurological diseases and continued need for intensive care is well established in Germany. OBJECTIVE: To assess the efficacy of ENNR from the perspective of evidence-based medicine as well as to present data on the outcome of ENNR patients including the analysis of prognostic factors. MATERIAL AND METHODS: A search was carried out in PubMed databases to identify early rehabilitation treatment forms evaluated by randomized controlled trials and with respect to large multicenter surveys of outcome and prognostic factors. RESULTS: For ENNR as a complex treatment concept, effectiveness not has been shown with regard to evidence-based medicine but it includes individually effective treatment forms. In two large multicenter evaluations the average duration of treatment was between 51 and 57 days and mortality was between 6 % and 10 %, increasing with the proportion of mechanically ventilated patients. Lower need for nursing support on admission indicated better outcome, whereas mechanical ventilation was more likely to be associated with poor outcome. Long-term outcome was negatively influenced by mechanical ventilation as well as severe neurogenic dysphagia with and without the need for a tracheal cannula and/or percutaneous endoscopic gastrostomy (PEG) and also by severely impaired communication at the end of ENNR. DISCUSSION: These prognostic factors indicate the primary aims of ENNR, which are to reduce the need for nursing support and to establish the capability for rehabilitation. If these aims are achieved, favorable functional and long-term outcome can be expected for ENNR patients. The presented studies verify the sustained efficacy of ENNR as an essential part of the overall treatment concept for severely neurologically impaired patients.

Criterion validity and sensitivity to change of the Early Rehabilitation Index (ERI): results from a German multi-center study.

BACKGROUND: Evaluation of functional status is difficult in neurological and neurosurgical early rehabilitation patients. The Early Rehabilitation Index (ERI) was introduced in Germany over 20 years ago, but since then validation studies are lacking. The ERI (range -325 to 0 points) includes highly relevant items including the necessity of intermittent mechanical ventilation or tracheostomy. METHODS: The present paper analyzed data from a German multi-center study, enrolling 754 neurological early rehabilitation patients. Together with ERI, Barthel Index (BI), Glasgow Coma Scale (GCS), Glasgow Outcome Score Extended, Coma Remission Scale (CRS), Functional Ambulation Categories and length of stay were obtained. RESULTS: ERI showed significant improvements from admission to discharge (p < 0.001). In addition, there were significant correlations of the ERI upon admission and at discharge with BI, CRS and GCS. CONCLUSIONS: Evaluation of our study data suggest that the ERI may be used as a valid assessment instrument for neurological and neurosurgical early rehabilitation patients.

[Course of rehabilitation in early neurological/neurosurgical rehabilitation. Results of a 2014 multi-center evaluation in Germany]. / Rehabilitationsverlauf von Patienten in der neurologisch-neurochirurgischen Frührehabilitation.

BACKGROUND: In Germany, neurological-neurosurgical early rehabilitation is well established in the treatment of severe neurological diseases. To develop quality standards, knowledge of the current rehabilitation course is required. PATIENTS AND METHODS: A retrospective analysis was performed on the course of rehabilitation from patients in an early neurological/neurosurgical rehabilitation program in 16 centers from 10 German states. The odds for a good or poor outcome were investigated using a multivariate logistic regression model. RESULTS: Seven hundred and fifty-four patients were included in the study. The average age of the patients was 68 ± 15 years. Of the patients studied, 26 % were on mechanical ventilation commencing their neurological rehabilitation. The average duration of stay was 56 ± 51 days. Weaning rate from mechanical ventilation was 65 % and the rate of weaning from tracheal cannula was 54 %. Mean improvement in the Barthel Index of 17 points, significant reduction of dysphagia (from 62 to 30 %) and depended walking (from 99 to 82 %), and the achievement of phase C (the next stage of rehabilitation) in 38 % can still be counted as signs of successful rehabilitation. During their course of stay, near 10 % of the patients died. Of these, 67 % received solely palliative care. In the multivariate logistic models, the absence of the factor "necessity for mechanical ventilation on admission" (odds ratio 0.61; 95 % confidence interval (CI): 0.42 … 0.89) increased the chance for good outcome and the presence of this factor the risk of dying with an odds ratio of 8.07 (95 % CI: 4.54-14.34). DISCUSSION: In spite of the severity of neurological deficits, significant functional progress has been made. These results could be interpret as positive proof of the efficacy of neurological/neurosurgical early rehabilitation programs.

Cyanobacterial Alkanes Modulate Photosynthetic Cyclic Electron Flow to Assist Growth under Cold Stress.

All cyanobacterial membranes contain diesel-range C15-C19 hydrocarbons at concentrations similar to chlorophyll. Recently, two universal but mutually exclusive hydrocarbon production pathways in cyanobacteria were discovered. We engineered a mutant of Synechocystis sp. PCC 6803 that produces no alkanes, which grew poorly at low temperatures. We analyzed this defect by assessing the redox kinetics of PSI. The mutant exhibited enhanced cyclic electron flow (CEF), especially at low temperature. CEF raises the ATP:NADPH ratio from photosynthesis and balances reductant requirements of biosynthesis with maintaining the redox poise of the electron transport chain. We conducted in silico flux balance analysis and showed that growth rate reaches a distinct maximum for an intermediate value of CEF equivalent to recycling 1 electron in 4 from PSI to the plastoquinone pool. Based on this analysis, we conclude that the lack of membrane alkanes causes higher CEF, perhaps for maintenance of redox poise. In turn, increased CEF reduces growth by forcing the cell to use less energy-efficient pathways, lowering the quantum efficiency of photosynthesis. This study highlights the unique and universal role of medium-chain hydrocarbons in cyanobacterial thylakoid membranes: they regulate redox balance and reductant partitioning in these oxygenic photosynthetic cells under stress.

Fine-Tuning of Photoautotrophic Protein Production by Combining Promoters and Neutral Sites in the Cyanobacterium Synechocystis sp. Strain PCC 6803.

Cyanobacteria are photosynthetic cell factories that use solar energy to convert CO2 into useful products. Despite this attractive feature, the development of tools for engineering cyanobacterial chassis has lagged behind that for heterotrophs such as Escherichia coli or Saccharomyces cerevisiae. Heterologous genes in cyanobacteria are often integrated at presumptively "neutral" chromosomal sites, with unknown effects. We used transcriptome sequencing (RNA-seq) data for the model cyanobacterium Synechocystis sp. strain PCC 6803 to identify neutral sites from which no transcripts are expressed. We characterized the two largest such sites on the chromosome, a site on an endogenous plasmid, and a shuttle vector by integrating an enhanced yellow fluorescent protein (EYFP) expression cassette expressed from either the Pcpc560 or the Ptrc1O promoter into each locus. Expression from the endogenous plasmid was as much as 14-fold higher than that from the chromosome, with intermediate expression from the shuttle vector. The expression characteristics of each locus correlated predictably with the promoters used. These findings provide novel, characterized tools for synthetic biology and metabolic engineering in cyanobacteria.

"Like walking into an empty room": effects of eurythmy therapy on stress perception in comparison with a sports intervention from the subjects' perspective-a qualitative study.

Background. Stress and health-related quality of life are important constructs used in treatment evaluation today. This study is based on a randomised controlled trial examining the stress-reducing effect of eurythmy therapy in comparison with step aerobics in 106 healthy but stressed subjects. The aim of the analysis was to characterise changes in the subjective perceptions of the participants. Methods. Interviews were conducted with 76 healthy adults, 36 (f = 31/m = 5) from the eurythmy group and 40 (f = 28/m = 12) from the step aerobics group both analysed by content analysis and phenomenologically. Results. The following categories were identified for the eurythmy therapy group: enabling a productive therapeutic response, emergence of a new perceptual space, reevaluation of the accustomed perception, and emergence of new options for action. Step aerobics places increased physical and intellectual demands. These are perceived differently as pleasant and relaxing, insufficiently challenging and/or boring, and too challenging and thus experienced as stress-enhancing. Conclusion. The qualitative results provided revealing insights into the profound effects of and subjective assignments of meaning to external and internal stress factors. Processes of mental reinterpretation leading to stress reduction can be stimulated by physical procedures such as eurythmy therapy.

Effects of the self-myocardial retroperfusion with aortic-coronary sinus shunt on cardiac output and ischemic events in high-risk patients undergoing OPCAB surgery.

AIM: Despite controversies, off-pump coronary artery bypass (OPCAB) surgery has become a routine procedure. Obvious advantages have been demonstrated in high-risk patients. However, OPCAB surgery has limitations in specific high-risk situations with hazards of operative deleterious events. We describe an innovative procedure of self-myocardial retroperfusion (SMR) with an aortic-coronary sinus shunt (ACSS). We prospectively evaluated the protective effects and benefits of SMR in high-risk coronary patients with impaired LVEF. METHODS: Eighteen consecutive high-risk (ES>10) coronary patients (mean age: 65.94 years; range: 34-85; mean ES: 26.97%) with LVEF≤35% who were not eligible for IABP were assigned for OPCAB surgery. Following sternotomy, the cardiac indexes (CI) were measured before, during SMR and after completion of coronary artery bypasses. Operative events with and without SMR were accurately collected, and postoperative cardiac Troponin T release was measured. RESULTS: OPCAB procedures were performed in all patients. Intraoperative use of SMR significantly increased CI (P=3.1041810.10-8) and reversed deleterious operative events (ECG changes/low cardiac output). Hospital mortality was 0%. Incidence of transient atrial fibrillation was 33.33%. Neither stroke nor renal insufficiency was observed. The mean graft number/patient was 2.05. Mean postoperative cardiac Troponin T value was 0.79 µg/L. Beating heart preservation optimized by SMR contributed to reduce ischemia-reperfusion injury, as validated by an immediate increase of CI after completion of coronary bypasses (P=3.35009.10-9). CONCLUSION: The concept of SMR with an ACSS during OPCAB procedures definitely improved CI and reversed ischemic features in high-risk patients and should be considered as an operative temporary myocardial assistance.

Effects of dairy slurry on silage fermentation characteristics and nutritive value of alfalfa.

Dairy producers frequently ask questions about the risks associated with applying dairy slurry to growing alfalfa (Medicago sativa L.). Our objectives were to determine the effects of applying dairy slurry on the subsequent nutritive value and fermentation characteristics of alfalfa balage. Dairy slurry was applied to 0.17-ha plots of alfalfa; applications were made to the second (HARV1) and third (HARV2) cuttings during June and July of 2012, respectively, at mean rates of 42,400 ± 5271 and 41,700 ± 2397 L/ha, respectively. Application strategies included (1) no slurry, (2) slurry applied directly to stubble immediately after the preceding harvest, (3) slurry applied after 1 wk of post-ensiled regrowth, or (4) slurry applied after 2 wk of regrowth. All harvested forage was packaged in large, rectangular bales that were ensiled as wrapped balage. Yields of DM harvested from HARV1 (2,477 kg/ha) and HARV2 (781 kg/ha) were not affected by slurry application treatment. By May 2013, all silages appeared to be well preserved, with no indication of undesirable odors characteristic of clostridial fermentations. Clostridium tyrobutyricum, which is known to negatively affect cheese production, was not detected in any forage on either a pre- or post-ensiled basis. On a pre-ensiled basis, counts for Clostridium cluster 1 were greater for slurry-applied plots than for those receiving no slurry, and this response was consistent for HARV1 (4.44 vs. 3.29 log10 genomic copies/g) and HARV2 (4.99 vs. 3.88 log10 genomic copies/g). Similar responses were observed on a post-ensiled basis; however, post-ensiled counts also were greater for HARV1 (5.51 vs. 5.17 log10 genomic copies/g) and HARV2 (5.84 vs. 5.28 log10 genomic copies/g) when slurry was applied to regrowth compared with stubble. For HARV2, counts also were greater following a 2-wk application delay compared with a 1-wk delay (6.23 vs. 5.45 log10 genomic copies/g). These results suggest that the risk of clostridial fermentations in alfalfa silages is greater following applications of slurry. Based on pre- and post-ensiled clostridial counts, applications of dairy slurry on stubble are preferred (and less risky) compared with delayed applications on growing alfalfa.

Fall-grown oat to extend the fall grazing season for replacement dairy heifers.

Our objective was to assess the pasture productivity and forage characteristics of 2 fall-grown oat (Avena sativa L.) cultivars, specifically for extending the grazing season and reducing reliance on harvested forages by replacement dairy heifers. A total of 160 gravid Holstein heifers (80 heifers/yr) were stratified by weight, and assigned to 1 of 10 identical research pens (8 heifers/pen). Initial body weights were 480 ± 43.5 kg in 2011 and 509 ± 39.4 kg in 2012. During both years of the trial, four 1.0-ha pasture replicates were seeded in August with Ogle oat (Schumitsch Seed Inc., Antigo, WI), and 4 separate, but similarly configured, pasture replicates were seeded with Forage Plus oat (Kratz Farms, Slinger, WI). Heifer groups were maintained as units, assigned to specific pastures, and then allowed to graze fall-oat pastures for 6h daily before returning to the barn, where they were offered a forage-based basal total mixed ration. Two heifer groups were retained in confinement (without grazing) as controls and offered the identical total mixed ration as pasture groups. During 2011, available forage mass increased with strong linear and quadratic effects for both cultivars, peaking at almost 9 Mg/ha on October 31. In contrast, forage mass was not affected by evaluation date in 2012, remaining ≤ 2,639 kg/ha across all dates because of droughty climatic conditions. During 2012, Ogle exhibited greater forage mass than Forage Plus across all sampling dates (2,678 vs. 1,856 kg/ha), largely because of its more rapid maturation rate and greater canopy height. Estimates of energy density for oat forage ranged from 59.6 to 69.1% during 2011, and ranged narrowly from 68.4 to 70.4% during 2012. For 2011, responses for both cultivars had strong quadratic character, in which the most energy-dense forages occurred in mid November, largely due to accumulation of water-soluble carbohydrates that reached maximum concentrations of 18.2 and 15.1% for Forage Plus and Ogle, respectively. Across the 2-yr trial, average daily gain for grazing heifer groups tended to be greater than heifers remaining in confinement (0.85 vs. 0.74 kg/d), but both management strategies produced weight gains within reasonable proximity to normal targets for heifers in this weight range. Fall-grown oat should be managed as stockpiled forage for deferred grazing, and good utilization of fall-oat forage can be accomplished by a one-time removal of standing forage, facilitated by a single lead wire advanced daily to prevent waste.

Rapid construction of metabolic models for a family of Cyanobacteria using a multiple source annotation workflow.

BACKGROUND: Cyanobacteria are photoautotrophic prokaryotes that exhibit robust growth under diverse environmental conditions with minimal nutritional requirements. They can use solar energy to convert CO2 and other reduced carbon sources into biofuels and chemical products. The genus Cyanothece includes unicellular nitrogen-fixing cyanobacteria that have been shown to offer high levels of hydrogen production and nitrogen fixation. The reconstruction of quality genome-scale metabolic models for organisms with limited annotation resources remains a challenging task. RESULTS: Here we reconstruct and subsequently analyze and compare the metabolism of five Cyanothece strains, namely Cyanothece sp. PCC 7424, 7425, 7822, 8801 and 8802, as the genome-scale metabolic reconstructions iCyc792, iCyn731, iCyj826, iCyp752, and iCyh755 respectively. We compare these phylogenetically related Cyanothece strains to assess their bio-production potential. A systematic workflow is introduced for integrating and prioritizing annotation information from the Universal Protein Resource (Uniprot), NCBI Protein Clusters, and the Rapid Annotations using Subsystems Technology (RAST) method. The genome-scale metabolic models include fully traced photosynthesis reactions and respiratory chains, as well as balanced reactions and GPR associations. Metabolic differences between the organisms are highlighted such as the non-fermentative pathway for alcohol production found in only Cyanothece 7424, 8801, and 8802. CONCLUSIONS: Our development workflow provides a path for constructing models using information from curated models of related organisms and reviewed gene annotations. This effort lays the foundation for the expedient construction of curated metabolic models for organisms that, while not being the target of comprehensive research, have a sequenced genome and are related to an organism with a curated metabolic model. Organism-specific models, such as the five presented in this paper, can be used to identify optimal genetic manipulations for targeted metabolite overproduction as well as to investigate the biology of diverse organisms.

Synthetic biology of cyanobacteria: unique challenges and opportunities.

Photosynthetic organisms, and especially cyanobacteria, hold great promise as sources of renewably-produced fuels, bulk and specialty chemicals, and nutritional products. Synthetic biology tools can help unlock cyanobacteria's potential for these functions, but unfortunately tool development for these organisms has lagged behind that for S. cerevisiae and E. coli. While these organisms may in many cases be more difficult to work with as "chassis" strains for synthetic biology than certain heterotrophs, the unique advantages of autotrophs in biotechnology applications as well as the scientific importance of improved understanding of photosynthesis warrant the development of these systems into something akin to a "green E. coli." In this review, we highlight unique challenges and opportunities for development of synthetic biology approaches in cyanobacteria. We review classical and recently developed methods for constructing targeted mutants in various cyanobacterial strains, and offer perspective on what genetic tools might most greatly expand the ability to engineer new functions in such strains. Similarly, we review what genetic parts are most needed for the development of cyanobacterial synthetic biology. Finally, we highlight recent methods to construct genome-scale models of cyanobacterial metabolism and to use those models to measure properties of autotrophic metabolism. Throughout this paper, we discuss some of the unique challenges of a diurnal, autotrophic lifestyle along with how the development of synthetic biology and biotechnology in cyanobacteria must fit within those constraints.

Unique interrelationships between fiber composition, water-soluble carbohydrates, and in vitro gas production for fall-grown oat forages.

Sixty samples of 'ForagePlus' oat were selected from a previous plot study for analysis of in vitro gas production (IVGP) on the basis of 2 factors: (1) high (n=29) or low (n=31) neutral detergent fiber (NDF; 62.7±2.61 and 45.1±3.91%, respectively); and (2) the range of water-soluble carbohydrates (WSC) within the high- and low-NDF groups. For the WSC selection factor, concentrations ranged from 4.7 to 13.4% (mean=7.9±2.06%) and from 3.5 to 19.4% (mean=9.7±4.57%) within high- and low-NDF forages, respectively. Our objectives were to assess the relationships between IVGP and various agronomic or nutritional characteristics for high- and low-NDF fall-oat forages. Cumulative IVGP was fitted to a single-pool nonlinear regression model: Y=MAX × (1 - e ([-)(K)(× (t - lag)])), where Y=cumulative gas produced (mL), MAX=maximum cumulative gas produced with infinite incubation time (mL), K=rate constant, t=incubation time (h), and lag=discrete lag time (h). Generally, cumulative IVGP after 12, 24, 36, or 48h within high-NDF fall-oat forages was negatively correlated with NDF, hemicellulose, lignin, and ash, but positively correlated with WSC, nonfiber carbohydrate (NFC), and total digestible nutrients (TDN). For low-NDF fall-grown oat forages, IVGP was positively correlated with growth stage, canopy height, WSC, NFC, and TDN; negative correlations were observed with ash and crude protein (CP) but not generally with fiber components. These responses were also reflected in multiple regression analysis for high- and low-NDF forages. After 12, 24, or 36h of incubation, cumulative IVGP within high-NDF fall-oat forages was explained by complex regression equations utilizing (lignin:NDF)(2), lignin:NDF, hemicellulose, lignin, and TDN(2) as independent variables (R(2)≥0.43). Within low-NDF fall-grown oat forages, cumulative IVGP at these incubation intervals was explained by positive linear relationships with NFC that also exhibited high coefficients of determination (R(2)≥0.75). Gas production was accelerated at early incubation times within low-NDF forages, specifically in response to large pools of WSC that were most likely to be present as forages approached boot stage by late-fall.