Microfluidics: From Crystallization to Serial Time-Resolved Crystallography

Capturing protein structural dynamics in real-time has tremendous potential in elucidating biological functions and providing information for structure-based drug design. While time-resolved structure determination has long been considered inaccessible for a vast majority of protein targets, serial methods for crystallography have remarkable potential in facilitating such analyses. Here, we review the impact of microfluidic technologies on protein crystal growth and X-ray diffraction analysis. In particular, we focus on applications of microfluidics for use in serial crystallography experiments for the time-resolved determination of protein structural dynamics

The impact of biological bedforms on near-bed and subsurface flow: a laboratory evaluated numerical study of flow in the vicinity of pits and mounds

The complex surface topography of river substrates controls near-bed hydraulics and drives the exchange of subsurface and surface flow. In rivers, the topographic structures that are studied are usually formed by the flow but, it is known that many animals also create biogenic bedforms, such as pits and mounds. Here, a Large-Eddy Simulation (LES) model of flow over a pit and a mound is evaluated with flume experiments. The model includes actual bedform topography, and the topographic complexity of the surrounding bed surface. Subsurface grains are organized in a body-centered cubic packing arrangement. Model evaluation showed strong agreement between experimental and modelling results for velocity (R2 > 0.8) and good agreement for Reynolds stresses (R2 > 0.7), which is comparable to other similar studies. Simulation of the pit shows that the length of the downwelling region is smaller than the upwelling region and that the velocity magnitude is higher in the downwelling region. Simulation of the mound reveals that the flow is forced into the bed upstream of the mound and re-emerges near the top of the mound. The recirculation zone is limited at the leeside of the mound. With increasing Reynolds number, the depth of the upwelling region at the leeside of the mound increases. The analysis of shear stress indicates that sediments on the upstream edge of the pit and on the downstream face of the mound are relatively unstable. These results demonstrate the effect of biogenic structures on the near-bed flow field, hyporheic exchange, and sediment stability

Temporary interruption of baricitinib: characterization of interruptions and effect on clinical outcomes in patients with rheumatoid arthritis

Background In clinical practice, temporary interruption of rheumatoid arthritis (RA) therapy is common for various reasons including side effects, non-compliance, or necessity for surgery. To characterize temporary interruptions of baricitinib and placebo-matched tablets in phase 3 studies of patients with moderate-to-severe rheumatoid arthritis (RA) and describe their impact on efficacy and safety. Methods During 4 baricitinib phase 3 studies, investigators documented timing, reason, and duration of investigator-initiated temporary interruptions of study drug. In 2 studies, patients recorded RA symptoms in daily diaries for 12 weeks. Post hoc analyses investigated changes in symptom scores during interruptions and resumption of treatment. Interruptions were evaluated for reoccurrence of adverse events or laboratory abnormalities after retreatment. Results Across the placebo-controlled studies, interruptions occurred in larger proportions of baricitinib- (2 mg, 18%; 4 mg, 18%) vs placebo-treated (9%) patients in only one study (bDMARD-inadequate responder patients, RA-BEACON). In the active comparator-controlled studies, the lowest rates of interruption were in the baricitinib monotherapy arm (9%) of RA-BEGIN (vs methotrexate monotherapy or combination therapy), and proportions were similar for baricitinib (10%) and adalimumab (9%) in RA-BEAM. Adverse events were the most common reason for interruption, but their reoccurrence after drug restart was infrequent. Most interruptions lasted ≤ 2 weeks. Daily diaries indicated modest symptom increases during interruption with return to pre-interruption levels or better after resumption. Interruptions had no impact on long-term efficacy outcomes. Conclusions Consistent with its pharmacologic properties, brief interruptions of baricitinib during phase 3 studies were associated with minor increases in RA symptoms that resolved following retreatment. This analysis provides useful information for clinicians, as temporary interruption of antirheumatic therapy is common in the care of patients with RA

Correction to: Temporary interruption of baricitinib: characterization of interruptions and effect on clinical outcomes in patients with rheumatoid arthritis

Background In clinical practice, temporary interruption of rheumatoid arthritis (RA) therapy is common for various reasons including side effects, non-compliance, or necessity for surgery. To characterize temporary interruptions of baricitinib and placebo-matched tablets in phase 3 studies of patients with moderate-to-severe rheumatoid arthritis (RA) and describe their impact on efficacy and safety. Methods During 4 baricitinib phase 3 studies, investigators documented timing, reason, and duration of investigator-initiated temporary interruptions of study drug. In 2 studies, patients recorded RA symptoms in daily diaries for 12 weeks. Post hoc analyses investigated changes in symptom scores during interruptions and resumption of treatment. Interruptions were evaluated for reoccurrence of adverse events or laboratory abnormalities after retreatment. Results Across the placebo-controlled studies, interruptions occurred in larger proportions of baricitinib- (2 mg, 18%; 4 mg, 18%) vs placebo-treated (9%) patients in only one study (bDMARD-inadequate responder patients, RA-BEACON). In the active comparator-controlled studies, the lowest rates of interruption were in the baricitinib monotherapy arm (9%) of RA-BEGIN (vs methotrexate monotherapy or combination therapy), and proportions were similar for baricitinib (10%) and adalimumab (9%) in RA-BEAM. Adverse events were the most common reason for interruption, but their reoccurrence after drug restart was infrequent. Most interruptions lasted ≤ 2 weeks. Daily diaries indicated modest symptom increases during interruption with return to pre-interruption levels or better after resumption. Interruptions had no impact on long-term efficacy outcomes. Conclusions Consistent with its pharmacologic properties, brief interruptions of baricitinib during phase 3 studies were associated with minor increases in RA symptoms that resolved following retreatment. This analysis provides useful information for clinicians, as temporary interruption of antirheumatic therapy is common in the care of patients with RA

Children with language delay referred to Dutch speech and hearing centres: caseload characteristics

Background: Early detection and remediation of language disorders are important in helping children to establish appropriate communicative and social behaviour and acquire additional information about the world through the use of language. In the Netherlands, children with (a suspicion of) language disorders are referred to speech and hearing centres for multidisciplinary assessment. Reliable data are needed on the nature of language disorders, as well as the age and source of referral, and the effects of cultural and socioeconomic profiles of the population served in order to plan speech and language therapy service provision. Aims: To provide a detailed description of caseload characteristics of children referred with a possible language disorder by generating more understanding of factors that might influence early identification. Methods & Procedures: A database of 11,450 children was analysed consisting of data on children, aged 2–7 years (70% boys, 30% girls), visiting Dutch speech and hearing centres. The factors analysed were age of referral, ratio of boys to girls, mono- and bilingualism, nature of the language delay, and language profile of the children. Outcomes & Results: Results revealed an age bias in the referral of children with language disorders. On average, boys were referred 5 months earlier than girls, and monolingual children were referred 3 months earlier than bilingual children. In addition, bilingual children seemed to have more complex problems at referral than monolingual children. They more often had both a disorder in both receptive and expressive language, and a language disorder with additional (developmental) problems. Conclusions & Implications: This study revealed a bias in age of referral of young children with language disorders. The results implicate the need for objective language screening instruments and the need to increase the awareness of staff in primary child healthcare of red flags in language development of girls and multilingual children aiming at earlier identification of language disorders in these children. What this paper adds What is already known on the subject Identifying language disorders before children enter school can foster the initiation of early interventions before these problems interfere with formal education and behavioural adjustment. Information on caseload characteristics is important to plan speech and language therapy service provision. There are only a few studies on the caseload characteristics of children at first referral for language assessment. What this paper adds to existing knowle

Regional variation in life history traits and plastic responses to temperature of the major malaria vector Nyssorhynchus darlingi in Brazil

The primary Brazilian malaria vector, Nyssorhynchus darlingi (formerly Anopheles darlingi), ranges from 0°S-23°S across three biomes (Amazonia, Cerrado, Mata Atlântica). Rising temperatures will increase mosquito developmental rates, and models predict future malaria transmission by Ny. darlingi in Brazil will shift southward. We reared F1 Ny. darlingi (progeny of field-collected females from 4 state populations across Brazil) at three temperatures (20, 24, 28 °C) and measured key life-history traits. Our results reveal geographic variation due to both genetic differences among localities and plastic responses to temperature differences. Temperature significantly altered all traits: faster larval development, shorter adult life and overall lifespan, and smaller body sizes were seen at 28 °C versus 20 °C. Low-latitude Amazonia mosquitoes had the fastest larval development at all temperatures, but at 28 °C, average development rate of high-latitude Mata Atlântica mosquitoes was accelerated and equivalent to low-latitude Amazonia. Body size of adult mosquitoes from the Mata Atlântica remained larger at all temperatures. We detected genetic variation in the plastic responses among mosquitoes from different localities, with implications for malaria transmission under climate change. Faster development combined with larger body size, without a tradeoff in adult longevity, suggests vectorial capacities of some Mata Atlântica populations may significantly increase under warming climates.NIH-NIAID [1R01AI110112]; Biodefense and Emerging Infectious Disease training fellowship [T32AI05532901]Open access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Sand as Maxwell's demon

We consider a dilute gas of granular material inside a box, kept in a stationary state by shaking. A wall separates the box into two identical compartments, save for a small hole at some finite height $h$. As the gas is cooled, a second order phase transition occurs, in which the particles preferentially occupy one side of the box. We develop a quantitative theory of this clustering phenomenon and find good agreement with numerical simulations

UV-induced ligand exchange in MHC class I protein crystals

High-throughput structure determination of protein−ligand complexes is central in drug development and structural proteomics. To facilitate such high-throughput structure determination we designed an induced replacement strategy. Crystals of a protein complex bound to a photosensitive ligand are exposed to UV light, inducing the departure of the bound ligand, allowing a new ligand to soak in. We exemplify the approach for a class of protein complexes that is especially recalcitrant to high-throughput strategies: the MHC class I proteins. We developed a UV-sensitive, “conditional”, peptide ligand whose UV-induced cleavage in the crystals leads to the exchange of the low-affinity lytic fragments for full-length peptides introduced in the crystallant solution. This “in crystallo” exchange is monitored by the loss of seleno-methionine anomalous diffraction signal of the conditional peptide compared to the signal of labeled MHC β2m subunit. This method has the potential to facilitate high-throughput crystallography in various protein families

Cryptotomography: reconstructing 3D Fourier intensities from randomly oriented single-shot diffraction patterns

We reconstructed the 3D Fourier intensity distribution of mono-disperse prolate nano-particles using single-shot 2D coherent diffraction patterns collected at DESY's FLASH facility when a bright, coherent, ultrafast X-ray pulse intercepted individual particles of random, unmeasured orientations. This first experimental demonstration of cryptotomography extended the Expansion-Maximization-Compression (EMC) framework to accommodate unmeasured fluctuations in photon fluence and loss of data due to saturation or background scatter. This work is an important step towards realizing single-shot diffraction imaging of single biomolecules.Comment: 4 pages, 4 figure

Ecological divergence and evolutionary transition of resprouting types in Banksia attenuata

Resprouting is a key functional trait that allows plants to survive diverse disturbances. The fitness benefits associated with resprouting include a rapid return to adult growth, early flowering, and setting seed. The resprouting responses observed following fire are varied, as are the ecological outcomes. Understanding the ecological divergence and evolutionary pathways of different resprouting types and how the environment and genetics interact to drive such morphological evolution represents an important, but under-studied, topic. In the present study, microsatellite markers and microevolutionary approaches were used to better understand: (1) whether genetic differentiation is related to morphological divergence among resprouting types and if so, whether there are any specific genetic variations associated with morphological divergence and (2) the evolutionary pathway of the transitions between two resprouting types in Banksia attenuata (epicormic resprouting from aerial stems or branch; resprouting from a underground lignotuber). The results revealed an association between population genetic differentiation and the morphological divergence of postfire resprouting types in B. attenuata. A microsatellite allele has been shown to be associated with epicormic populations. Approximate Bayesian Computation analysis revealed a likely evolutionary transition from epicormic to lignotuberous resprouting in B. attenuata. It is concluded that the postfire resprouting type in B. attenuata is likely determined by the fire's characteristics. The differentiated expression of postfire resprouting types in different environments is likely a consequence of local genetic adaptation. The capacity to shift the postfire resprouting type to adapt to diverse fire regimes is most likely the key factor explaining why B. attenuata is the most widespread member of the Banksia genus