Molprobity's ultimate rotamer‐library distributions for model validation

Here we describe the updated MolProbity rotamer‐library distributions derived from an order‐of‐magnitude larger and more stringently quality‐filtered dataset of about 8000 (vs. 500) protein chains, and we explain the resulting changes and improvements to model validation as seen by users. To include only side‐chains with satisfactory justification for their given conformation, we added residue‐specific filters for electron‐density value and model‐to‐density fit. The combined new protocol retains a million residues of data, while cleaning up false‐positive noise in the multi‐ datapoint distributions. It enables unambiguous characterization of conformational clusters nearly 1000‐fold less frequent than the most common ones. We describe examples of local interactions that favor these rare conformations, including the role of authentic covalent bond‐angle deviations in enabling presumably strained side‐chain conformations. Further, along with favored and outlier, an allowed category (0.3–2.0% occurrence in reference data) has been added, analogous to Ramachandran validation categories. The new rotamer distributions are used for current rotamer validation in MolProbity and PHENIX, and for rotamer choice in PHENIX model‐building and refinement. The multi‐dimensional distributions and Top8000 reference dataset are freely available on GitHub. These rotamers are termed “ultimate” because data sampling and quality are now fully adequate for this task, and also because we believe the future of conformational validation should integrate side‐chain with backbone criteria. Proteins 2016; 84:1177–1189. © 2016 Wiley Periodicals, Inc.     

Collaborative research partnerships inform monitoring and management of aquatic ecosystems by Indigenous rangers

Aquatic ecosystems are critical to the long-term viability and vibrancy of communities and economies across northern Australia. In a region that supports significant cultural and ecological water values, partnerships between Indigenous and non-Indigenous stakeholders can benefit aquatic ecosystem management. We present, as a case study from the Kimberley region of Western Australia, a collaborative research program that successfully documented Indigenous and Western Scientific knowledge of remote wetlands, using a variety of field-based activities, questionnaires, interviews and workshops. The sharing of knowledge between Indigenous and non-Indigenous research partners facilitated a comprehensive understanding of ecosystem values, threats, processes, management priorities and aspirations. These formed the basis of a management plan and monitoring tools, designed to build the capacity of an Indigenous ranger group to engage in research, monitoring and management of wetlands. The project provides a useful example of the benefits of collaborations in the context of remote-area management where local communities are responsible for environmental management and monitoring, such as is the case in northern Australia and presumably other areas of the world.     

The Genetic Basis of Host Preference and Resting Behavior in the Major African Malaria Vector,Anopheles arabiensis

Malaria transmission is dependent on the propensity of Anopheles mosquitoes to bite humans (anthropophily) instead of other dead end hosts. Recent increases in the usage of Long Lasting Insecticide Treated Nets (LLINs) in Africa have been associated with reductions in highly anthropophilic and endophilic vectors such as Anopheles gambiae s.s., leaving species with a broader host range, such as Anopheles arabiensis, as the most prominent remaining source of transmission in many settings. An. arabiensis appears to be more of a generalist in terms of its host choice and resting behavior, which may be due to phenotypic plasticity and/or segregating allelic variation. To investigate the genetic basis of host choice and resting behavior in An. arabiensis we sequenced the genomes of 23 human-fed and 25 cattle-fed mosquitoes collected both in-doors and out-doors in the Kilombero Valley, Tanzania. We identified a total of 4,820,851 SNPs, which were used to conduct the first genome-wide estimates of “SNP heritability” for host choice and resting behavior in this species. A genetic component was detected for host choice (human vs cow fed; permuted P = 0.002), but there was no evidence of a genetic component for resting behavior (indoors versus outside; permuted P = 0.465). A principal component analysis (PCA) segregated individuals based on genomic variation into three groups which were characterized by differences at the 2Rb and/or 3Ra paracentromeric chromosome inversions. There was a non-random distribution of cattle-fed mosquitoes between the PCA clusters, suggesting that alleles linked to the 2Rb and/or 3Ra inversions may influence host choice. Using a novel inversion genotyping assay, we detected a significant enrichment of the standard arrangement (non-inverted) of 3Ra among cattle-fed mosquitoes (N = 129) versus all non-cattle-fed individuals (N = 234; χ², p = 0.007). Thus, tracking the frequency of the 3Ra in An. arabiensis populations may be of use to infer selection on host choice behavior within these vector populations; possibly in response to vector control. Controlled host-choice assays are needed to discern whether the observed genetic component has a direct relationship with innate host preference. A better understanding of the genetic basis for host feeding behavior in An. arabiensis may also open avenues for novel vector control strategies based on driving genes for zoophily into wild mosquito populations.     

Inactivation of Prions and Amyloid Seeds with Hypochlorous Acid

Hypochlorous acid (HOCl) is produced naturally by neutrophils and other cells to kill conventional microbes in vivo. Synthetic preparations containing HOCl can also be effective as microbial disinfectants. Here we have tested whether HOCl can also inactivate prions and other self-propagating protein amyloid seeds. Prions are deadly pathogens that are notoriously difficult to inactivate, and standard microbial disinfection protocols are often inadequate. Recommended treatments for prion decontamination include strongly basic (pH ≥~12) sodium hypochlorite bleach, ≥1 N sodium hydroxide, and/or prolonged autoclaving. These treatments are damaging and/or unsuitable for many clinical, agricultural and environmental applications. We have tested the anti-prion activity of a weakly acidic aqueous formulation of HOCl (BrioHOCl) that poses no apparent hazard to either users or many surfaces. For example, BrioHOCl can be applied directly to skin and mucous membranes and has been aerosolized to treat entire rooms without apparent deleterious effects. Here, we demonstrate that immersion in BrioHOCl can inactivate not only a range of target microbes, including spores of Bacillus subtilis, but also prions in tissue suspensions and on stainless steel. Real-time quaking-induced conversion (RT-QuIC) assays showed that BrioHOCl treatments eliminated all detectable prion seeding activity of human Creutzfeldt-Jakob disease, bovine spongiform encephalopathy, cervine chronic wasting disease, sheep scrapie and hamster scrapie; these findings indicated reductions of ≥10³- to 10⁶-fold. Transgenic mouse bioassays showed that all detectable hamster-adapted scrapie infectivity in brain homogenates or on steel wires was eliminated, representing reductions of ≥~10^5.75-fold and >10⁴-fold, respectively. Inactivation of RT-QuIC seeding activity correlated with free chlorine concentration and higher order aggregation or destruction of proteins generally, including prion protein. BrioHOCl treatments had similar effects on amyloids composed of human α-synuclein and a fragment of human tau. These results indicate that HOCl can block the self-propagating activity of prions and other amyloids.     

AMPK Signalling and Defective Energy Metabolism in Amyotrophic Lateral Sclerosis

Amyotrophic lateral sclerosis (ALS) is caused by selective loss of upper and lower motor neurons by complex mechanisms that are incompletely understood. Motor neurons are large, highly polarised and excitable cells with unusually high energetic demands to maintain resting membrane potential and propagate action potentials. This leads to higher ATP consumption and mitochondrial metabolism in motor neurons relative to other cells. Here, we review increasing evidence that defective energy metabolism and homeostasis contributes to selective vulnerability and degeneration of motor neurons in ALS. Firstly, we provide a brief overview of major energetic pathways in the CNS, including glycolysis, oxidative phosphorylation and the AMP-activated protein kinase (AMPK) signalling pathway, while highlighting critical metabolic interactions between neurons and astrocytes. Next, we review evidence from ALS patients and transgenic mutant SOD1 mice for weight loss, hypermetabolism, hyperlipidemia and mitochondrial dysfunction in disease onset and progression. Genetic and therapeutic modifiers of energy metabolism in mutant SOD1 mice will also be summarised. We also present evidence that additional ALS-linked proteins, TDP-43 and FUS, lead to energy disruption and mitochondrial defects in motor neurons. Lastly, we review emerging evidence including our own that dysregulation of the AMPK signalling cascade in motor neurons is an early and common event in ALS pathogenesis. We suggest that an imbalance in energy metabolism should be considered an important factor in both progression and potential treatment of ALS.     

Regional and seasonal variation in airborne grass pollen levels between cities of Australia and New Zealand

Although grass pollen is widely regarded as the major outdoor aeroallergen source in Australia and New Zealand (NZ), no assemblage of airborne pollen data for the region has been previously compiled. Grass pollen count data collected at 14 urban sites in Australia and NZ over periods ranging from 1 to 17 years were acquired, assembled and compared, revealing considerable spatiotemporal variability. Although direct comparison between these data is problematic due to methodological differences between monitoring sites, the following patterns are apparent. Grass pollen seasons tended to have more than one peak from tropics to latitudes of 37°S and single peaks at sites south of this latitude. A longer grass pollen season was therefore found at sites below 37°S, driven by later seasonal end dates for grass growth and flowering. Daily pollen counts increased with latitude; subtropical regions had seasons of both high intensity and long duration. At higher latitude sites, the single springtime grass pollen peak is potentially due to a cooler growing season and a predominance of pollen from C₃ grasses. The multiple peaks at lower latitude sites may be due to a warmer season and the predominance of pollen from C₄ grasses. Prevalence and duration of seasonal allergies may reflect the differing pollen seasons across Australia and NZ. It must be emphasized that these findings are tentative due to limitations in the available data, reinforcing the need to implement standardized pollen-monitoring methods across Australasia. Furthermore, spatiotemporal differences in grass pollen counts indicate that local, current, standardized pollen monitoring would assist with the management of pollen allergen exposure for patients at risk of allergic rhinitis and asthma.     

Integrating new practices: a qualitative study of how hospital innovations become routine

Background

Hospital quality improvement efforts absorb substantial time and resources, but many innovations fail to integrate into organizational routines, undermining the potential to sustain the new practices. Despite a well-developed literature on the initial implementation of new practices, we have limited knowledge about the mechanisms by which integration occurs.

Methods

We conducted a qualitative study using a purposive sample of hospitals that participated in the State Action on Avoidable Rehospitalizations (STAAR) initiative, a collaborative to reduce hospital readmissions that encouraged members to adopt new practices. We selected hospitals where risk-standardized readmission rates (RSRR) had improved (n?=?7) or deteriorated (n?=?3) over the course of the first 2 years of the STAAR initiative (2010–2011 to 2011–2012) and interviewed a range of staff at each site (90 total). We recruited hospitals until reaching theoretical saturation. The constant comparative method was used to conduct coding and identification of key themes.

Results

When innovations were successfully integrated, participants consistently reported that a small number of key staff held the innovation in place for as long as a year while more permanent integrating mechanisms began to work. Depending on characteristics of the innovation, one of three categories of integrating mechanisms eventually took over the role of holding new practices in place. Innovations that proved intrinsically rewarding to the staff, by making their jobs easier or more gratifying, became integrated through shifts in attitudes and norms over time. Innovations for which the staff did not perceive benefits to themselves were integrated through revised performance standards if the innovation involved complex tasks and through automation if the innovation involved simple tasks.

Conclusions

Hospitals have an opportunity to promote the integration of new practices by planning for the extended effort required to hold a new practice in place while integration mechanisms take hold. By understanding how integrating mechanisms correspond to innovation characteristics, hospitals may be able to foster integrating mechanisms most likely to work for particular innovations.

     

Impact of Contextual Factors on the Effect of Interventions to Improve Health Worker Performance in Sub-Saharan Africa: Review of Randomised Clinical Trials

Background

Africa bears 24% of the global burden of disease but has only 3% of the world’s health workers. Substantial variation in health worker performance adds to the negative impact of this significant shortfall. We therefore sought to identify interventions implemented in sub-Saharan African aiming to improve health worker performance and the contextual factors likely to influence local effectiveness.

Methods and Findings

A systematic search for randomised controlled trials of interventions to improve health worker performance undertaken in sub-Saharan Africa identified 41 eligible trials. Data were extracted to define the interventions’ components, calculate the absolute improvement in performance achieved, and document the likelihood of bias. Within-study variability in effect was extracted where reported. Statements about contextual factors likely to have modified effect were subjected to thematic analysis. Interventions to improve health worker performance can be very effective. Two of the three trials assessing mortality impact showed significant reductions in death rates (age<5 case fatality 5% versus 10%, p<0.01; maternal in-hospital mortality 6.8/1000 versus 10.3/1000; p<0.05). Eight of twelve trials focusing on prescribing had a statistically significant positive effect, achieving an absolute improvement varying from 9% to 48%. However, reported range of improvement between centres within trials varied substantially, in many cases exceeding the mean effect. Nine contextual themes were identified as modifiers of intervention effect across studies; most frequently cited were supply-line failures, inadequate supervision or management, and failure to follow-up training interventions with ongoing support, in addition to staff turnover.

Conclusions

Interventions to improve performance of existing staff and service quality have the potential to improve patient care in underserved settings. But in order to implement interventions effectively, policy makers need to understand and address the contextual factors which can contribute to differences in local effect. Researchers therefore must recognise the importance of reporting how context may modify effect size.