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The presence of even small amounts of liquid between particles dramatically changes the dynamics of collisions. This study considers granular collisions between two particles coated with a thin layer of viscous liquid, such that the capillary number is high and viscous forces dominate. High-speed particle tracking velocimetry was used to experimentally study the collisions of two smooth spheres with and without liquid coatings. We then use these experiments to examine four theoretical models that describe the collisions. A key challenge when modelling viscous forces is that the force which is predicted as particles approach each other scales with the inverse of the distance, i.e. tends to infinity. Most existing models impose a limit to the viscous force. One recent model instead assumes a hard sphere collision. These fundamentally different approaches produce different rebound outcomes. A fair match between experiments and simulations was obtained when using the hard sphere collision model, but only if an empirically-fitted glass transition pressure model was used to describe the minimum approach distance.

Granular dilatancy has been previously characterised through a simple linear relationship between the packing fraction and dimensionless shear rate. However, this relationship was developed for granular flows in a simple shear cell geometry. Here we examine inertial volume changes in a shear cell with gravity, a vertical chute, and a pseudo-2D hopper. In so doing, we show that the packing fraction displays both a local and non-local response, analogous to what is typically observed for the stress ratio µ.

To gain fundamental insight into granular flow behaviour, quantitative measurements of velocity and solid fraction are required. The aim of this study is to measure the solid fraction and velocity of 3D granular flows quantitively using a recently developed MRI method. Time-averaged spatial maps of the solid fraction and velocity are obtained for hoppers with wall angles of 30°, 60°, and 90°. From these maps, the mass flow rate of the material was calculated along the height of the hoppers. Excellent agreement was observed between the MRI and gravimetric mass flow rate measurements, confirming the quantitative nature of the measurements. The resulting solid fraction and velocity measurements provide insight into the dynamics of granular flow.

Abstract Background Community-based screening for TB, combined with HIV and syphilis testing, faces a number of barriers. One significant barrier is the value that target communities place on such screening. Methods Integrated testing for TB, HIV, and syphilis was performed in neighborhoods identified using geographic information systems-based disease mapping. TB testing included skin testing and interferon gamma release assays. Subjects completed a survey describing disease risk factors, healthcare access, healthcare utilization, and willingness to pay for integrated testing. Results Behavioral and social risk factors among the 113 subjects were prevalent (71% prior incarceration, 27% prior or current crack cocaine use, 35% homelessness), and only 38% had a regular healthcare provider. The initial 24 subjects reported that they would be willing to pay a median $20 (IQR: 0-100) for HIV testing and $10 (IQR: 0-100) for TB testing when the question was asked in an open-ended fashion, but when the question was changed to a multiple-choice format, the next 89 subjects reported that they would pay a median $5 for testing, and 23% reported that they would either not pay anything to get tested or would need to be paid $5 to get tested for TB, HIV, or syphilis. Among persons who received tuberculin skin testing, only 14/78 (18%) participants returned to have their skin tests read. Only 14/109 (13%) persons who underwent HIV testing returned to receive their HIV results. Conclusion The relatively high-risk persons screened in this community outreach study placed low value on testing. Reported willingness to pay for such testing, while low, likely overestimated the true willingness to pay. Successful TB, HIV, and syphilis integrated testing programs in high risk populations will likely require one-visit diagnostic testing and incentives.

Abstract Background Chemotherapy may improve survival in patients undergoing resection of colorectal liver metastases (CLM). Neoadjuvant chemotherapy may help identify patients with occult extrahepatic disease (averting unnecessary metastasectomy), and it provides in vivo chemosensitivity data. Methods A phase II trial was initiated in which patients with resectable CLM received CPT-11, 5-FU and LV for 12 weeks. Metastasectomy was performed unless extrahepatic disease appeared. Postoperatively, patients with stable or responsive disease received the same regimen for 12 weeks. Patients with progressive disease received either second-line chemotherapy or best supportive care. The primary endpoint was disease-free survival (DFS); secondary endpoints included overall survival (OS) and safety. Results 35 patients were accrued. During preoperative chemotherapy, 16 patients (46%) had grade 3/4 toxicities. Resection was not possible in 5 patients. One patient died of arrhythmia following surgery, and 1 patient had transient liver failure. During the postoperative treatment phase, 12 patients (55%) had grade 3/4 toxicities. Deep venous thrombosis (DVT) occurred in 11 patients (34%) at various times during treatment. Of those who underwent resection, median DFS was 23.0 mo. and median OS has not been reached. The overall survival from time of diagnosis of liver metastases was 51.6 mo for the entire cohort. Conclusion A short course of chemotherapy prior to hepatic metastasectomy may serve to select candidates best suited for resection and it may also direct postoperative systemic treatment. Given the significant incidence of DVT, alternative systemic neoadjuvant regimens should be investigated, particularly those that avoid the use of a central venous line. Trial Registration ClinicalTrials.gov NCT00168155.

Abstract Background Although previous research has demonstrated that referral to pre-dialysis clinics is associated with favourable objective outcomes, the benefit of a pre-dialysis clinic from the perspective of patient-perceived subjective outcomes, such as quality of life (QOL), is less well defined. Methods A retrospective incident cohort study was conducted to determine if pre-dialysis clinic attendance was a predictor of better QOL scores measured within the first six months of hemodialysis (HD) initiation. Inclusion criteria were HD initiation from January 1 1998 to January 1 2000, diagnosis of chronic renal failure, and completion of the QOL questionnaire within six months of HD initiation. Patients receiving HD for less than four weeks were excluded. An incident cohort of 120 dialysis patients was identified, including 74 patients who attended at least one pre-dialysis clinic and 46 patients who did not. QOL was measured using the SF 36-Item Health Survey. Independent variables included age, sex, diabetes, pre-dialysis clinic attendance and length of attendance, history of ischemic heart disease, stroke, peripheral vascular disease, heart failure, malignancy, and chronic lung disease, residual creatinine clearance at dialysis initiation, and kt/v, albumin and hemoglobin at the time of QOL assessment. Bivariate and multivariate linear regression analyses were used to identify predictors of QOL scores. Results Multivariate analysis suggested that pre-dialysis clinic attendance was an independent predictor of higher QOL scores in four of eight health domains (physical function, p < 0.01; emotional role limitation, p = 0.01; social function, p = 0.01; and general health, p = 0.03), even after statistical adjustment for age, sex, residual renal function, kt/v, albumin, and co-morbid disease. Pre-dialysis clinic attendance was also an independent predictor of the physical component summary score (p = 0.03). Conclusions We conclude that pre-dialysis clinic attendance favourably influences patient-perceived quality of life within six months of dialysis initiation.

The rise of mRNA as a novel vaccination strategy presents new opportunities to confront global disease. Double-stranded RNA (dsRNA) is an impurity byproduct of the in vitro transcription reaction used to manufacture mRNA that may affect the potency and safety of the mRNA vaccine in patients. Careful quantitation of dsRNA during manufacturing is critical to ensure that residual dsRNA is minimized in purified mRNA drug substances. In this work, we describe the development and implementation of a sandwich Enzyme-Linked Immunosorbent Assay (ELISA) to quantitate nanogram quantities of residual dsRNA contaminants in mRNA process intermediates using readily available commercial reagents. This sandwich ELISA developed in this study follows a standard protocol and can be easily adapted to most research laboratory environments. Additionally, a liquid handler coupled with an automated robotics system was utilized to increase assay throughput, improve precision, and reduce the analyst time requirement. The final automated sandwich ELISA was able to measure <10 ng/mL of dsRNA with a specificity for dsRNA over 2000-fold higher than mRNA, a variability of <15%, and a throughput of 72 samples per day.

Introducing new recombinant protein antigens to existing pediatric combination vaccines is important in improving coverage and affordability, especially in low- and middle-income countries (LMICs). This case-study highlights the analytical and formulation challenges encountered with three recombinant non-replicating rotavirus vaccine (NRRV) antigens (t-NRRV formulated with Alhydrogel ® adjuvant, AH) combined with a mock multidose formulation of a pediatric pentavalent vaccine used in LMICs. This complex formulation contained (1) vaccine antigens (i.e., whole-cell pertussis (wP), diphtheria (D), tetanus (T), Haemophilus influenza (Hib), and hepatitis B (HepB), (2) a mixture of aluminum-salt adjuvants (AH and Adju-Phos ® , AP), and (3) a preservative (thimerosal, TH). Selective, stability-indicating competitive immunoassays were developed to monitor binding of specific mAbs to each antigen, except wP which required the setup of a mouse immunogenicity assay. Simple mixing led to the desorption of t-NRRV antigens from AH and increased degradation during storage. These deleterious effects were caused by specific antigens, AP, and TH. An AH-only pentavalent formulation mitigated t-NRRV antigen desorption; however, the Hib antigen displayed previously reported AH-induced instability. The same rank-ordering of t-NRRV antigen stability (P[8] > P[4] > P[6]) was observed in mock pentavalent formulations and with various preservatives. The lessons learned are discussed to enable future multidose, combination vaccine formulation development with new vaccine candidates.

In a companion paper, a two-step developability assessment is presented to rapidly evaluate low-cost formulations (multi-dose, aluminum-adjuvanted) for new subunit vaccine candidates. As a case study, a non-replicating rotavirus (NRRV) recombinant protein antigen P[4] was found to be destabilized by the vaccine preservative thimerosal, and this effect was mitigated by modification of the free cysteine (C173S). In this work, the mechanism(s) of thimerosal-P[4] protein interactions, along with subsequent effects on the P[4] protein's structural integrity, are determined. Reversible complexation of ethylmercury, a thimerosal degradation byproduct, with the single cysteine residue of P[4] protein is demonstrated by intact protein mass analysis and biophysical studies. A working mechanism involving a reversible S-Hg coordinate bond is presented based on the literature. This reaction increased the local backbone flexibility of P[4] within the helical region surrounding the cysteine residue and then caused more global destabilization, both as detected by HX-MS. These effects correlate with changes in antibody-P[4] binding parameters and alterations in P[4] conformational stability due to C173S modification. Epitope mapping by HX-MS demonstrated involvement of the same cysteine-containing helical region of P[4] in antibody-antigen binding. Future formulation challenges to develop low-cost, multi-dose formulations for new recombinant protein vaccine candidates are discussed., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

A two-step developability assessment workflow is described to screen variants of recombinant protein antigens under various formulation conditions to rapidly identify stable, aluminum-adjuvanted, multi-dose vaccine candidates. For proof-of-concept, a series of sequence variants of the recombinant non-replicating rotavirus (NRRV) P[8] protein antigen (produced in Komagataella phaffii) were compared in terms of primary structure, post-translational modifications, antibody binding, conformational stability, relative solubility and preservative compatibility. Based on these results, promising P[8] variants were down-selected and the impact of key formulation conditions on storage stability was examined (e.g., presence or absence of the aluminum-adjuvant Alhydrogel and the preservative thimerosal) as measured by differential scanning calorimetry (DSC) and antibody binding assays. Good correlations between rapidly-generated developability screening data and storage stability profiles (12 weeks at various temperatures) were observed for aluminum-adsorbed P[8] antigens. These findings were extended and confirmed using variants of a second NRRV antigen, P[4]. These case-study results with P[8] and P[4] NRRV variants are discussed in terms of using this vaccine formulation developability workflow to better inform and optimize formulation design with a wide variety of recombinant protein antigens, with the long-term goal of rapidly and cost-efficiently identifying low-cost vaccine formulations for use in low and middle income countries., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Mycosporine-like amino acids (MAAs) are secondary metabolites of a variety of marine organisms including cyanobacteria and macroalgae. These compounds have strong ultraviolet (UV) absorption maxima between 310 and 362 nm and are biological sunscreens for counteracting the damaging effects of UV radiation in nature. The common MAA shinorine has been used as one key active ingredient of environmentally friendly sunscreen creams. Commercially used shinorine is isolated from one red algae that is generally harvested from the wild. Here, we describe the use of Synechocystis sp. PCC6803 as a host for the heterologous production of shinorine. We mined a shinorine gene cluster from the filamentous cyanobacterium Fischerella sp. PCC9339. When expressing the cluster in Synechocystis sp. PCC6803, we observed the production of shinorine using LC-MS analysis, but its productivity was three times lower than the native producer. Integrated transcriptional and metabolic profiling identified rate-limiting steps in the heterologous production of shinorine. The use of multiple promoters led to a 10-fold increase of its yield to 2.37 ± 0.21 mg/g dry biomass weight, comparable to commercially used shinorine producer. The UV protection of shinorine was further confirmed using the engineered Synechocystis sp. PCC6803. This work was the first time to demonstrate the photosynthetic overproduction of MAA. The results suggest that Synechocystis sp. PCC6803 can have broad applications as the synthetic biology chassis to produce other cyanobacterial natural products, expediting the translation of genomes into chemicals.

Three-photon excitation fluorescence correlation spectroscopy was used to detect oligomerization equilibria of rat liver phosphofructokinase. The fluorescence intensity produced by the three-photon excitation of tryptophan was collected using the DIVER microscope. In this home-built upright microscope, a large area photomultiplier, placed directly below the sample, is used as the detector. The lack of optical elements in the microscope detection path results in a significantly improved detection efficiency in the UV region down to about 300 nm, which encompasses the fluorescence emission from tryptophan. The three-photon excitation autocorrelation decays obtained for phosphofructokinase in the presence of F6P showed the presence of large oligomers. Substitution of F6P with ATP in the buffer medium results in dissociation of the large oligomers, which is reported by the decreased autocorrelation amplitude. The three-photon excitation process was verified from the slope of the log-log plot of intensity against laser power.

Future long-duration spaceflights are now being planned to the Moon and Mars as a part of the "Vision for Space Exploration" program initiated by NASA in 2004. This report describes the design reference missions for the International Space Station, Lunar Base, and eventually a Mars Expedition. There is a need to develop more stringent preflight medical screening for crewmembers to minimize risk factors for diseases which cannot be effectively treated in flight. Since funding for space life sciences research and development has been eliminated to fund program development, these missions will be enabled by countermeasures much like those currently in use aboard the International Space Station. Artificial gravity using centrifugation in a rotating spacecraft has been suggested repeatedly as a "universal countermeasure" against deconditioning in microgravity and could be an option if other countermeasures are found to be ineffective. However, the greatest medical unknown in interplanetary flight may be the effects of radiation exposure. In addition, a Mars expedition would lead to a far greater level of isolation and psychological stress than any space mission attempted previously; because of this, psychiatric decompensation remains a risk. Historically, mortality and morbidity related to illness and injury have accounted for more failures and delays in new exploration than have defective transportation systems. The medical care system on a future Mars expedition will need to be autonomous and self-sufficient due to the extremely long separation from definitive medical care. This capability could be expanded by the presence of a physician in the crew and including simple, low-technology surgical capability.

Introduction: During approach to a remote island location, a U.S. Air Force heavy-airlift aircraft was flown into an aerodynamic stall, resulting in the loss of more than 4000 ft of altitude, with the crew recovering the aircraft just before impact would have occurred., Methods: An analysis of the mishap was conducted through a review of non-privileged USAF mishap data, cockpit voice recordings, flight data records, and interviews of the aircrew involved. A thorough examination of fatigue-related factors was conducted, including computerized fatigue modeling., Results: The crew traveled over 11,000 mi in a westward direction over a 6-d period. They had been on duty for nearly 21 h on the day of the mishap, with minimal in-flight rest. The pilots were late beginning their descent for landing, and a minor aircraft malfunction distracted the crew, contributing to channelized attention and degraded situational awareness. A breakdown in crew communication and failure to adequately monitor and interpret true aircraft state culminated in loss of aircraft control. Analysis of the crew's work/rest schedule confirmed that multiple elements of fatigue were present during this mishap, including acute and cumulative fatigue, circadian disruptions, and sleep inertia. Additionally, reduced situational awareness and spatial disorientation, exacerbated by the underlying fatigue, were causal in this mishap., Discussion: This mishap highlights the importance of maintaining a high degree of situational awareness during long-haul flights with a continuing need to address issues regarding spatial disorientation, proper application of human engineering principles in modern cockpits, and mitigation of aircrew fatigue factors.

Introduction: Visual perception task, complex motor flight task, and psychomotor vigilance task performances were evaluated in U.S. Air Force pilots navigating a high-fidelity fixed wing jet simulator over 26.5 h of continuous wakefulness., Methods: Eight military pilots on flight status performed the primary task of flying a simulated 12.5-h overnight mission in an Air Refueling Part Task Trainer (ARPTT): Response omission to presentation of single- and double-light stimuli displayed in random sequence across the cockpit instrument panel was the metric used to assess choice visual perception task (CVPT) performance. Deviation from an established azimuth heading in the ARPTT during the CVPT was the flight metric used to assess complex motor performance. Speed, lapse, false start, and anticipation were the metrics used to assess psychomotor vigilance task (PVT) performance during crew rest periods., Results: Significant visual perceptual, complex motor, and psychomotor vigilance (speed and lapse) impairments occurred at 19 h awake in the eight-subject group. CVPT response omissions significantly correlated with ARPTT azimuth deviations at r = 0.97, and with PVT speed at r = -0.92 and lapses at r = 0.90. ARPTT azimuth deviations significantly correlated with PVT speed at r = -0.92 and lapses at r = 0.91., Conclusions: Acute sleep deprivation degrades visual perceptual, complex motor, and simple motor performance. Complex motor impairments strongly correlate with visual perceptual impairments. This research provides support for the use of visual perceptual measures as surrogates of complex motor performance in operational situations where the primary cognitive inputs are through the visual system.