Development of a capillary zone electrophoresis method to monitor magnesium ion consumption during in vitro transcription for mRNA production.

Messenger RNA (mRNA) vaccines represent a landmark in vaccinology, especially with their success in COVID-19 vaccines, which have shown great promise for future vaccine development and disease prevention. As a platform technology, synthetic mRNA can be produced with high fidelity using in vitro transcription (IVT). Magnesium plays a vital role in the IVT process, facilitating the phosphodiester bond formation between adjacent nucleotides and ensuring accurate transcription to produce high-quality mRNA. The development of the IVT process has prompted key inquiries about in-process characterization of magnesium ion (Mg++) consumption, relating to the RNA polymerase (RNAP) activation, fed-batch mode production yield, and mRNA quality. Hence, it becomes crucial to monitor the free Mg++ concentration throughout the IVT process. However, no free Mg++ analysis method has been reported for complex IVT reactions. Here we report a robust capillary zone electrophoresis (CZE) method with indirect UV detection. The assay allows accurate quantitation of free Mg++ for the complex IVT reaction where it is essential to preserve IVT samples in their native-like state during analysis to avoid dissociation of bound Mg complexes. By applying this CZE method, the relationships between free Mg++ concentration, the mRNA yield, and dsRNA impurity level were investigated. Such mechanistic understanding facilitates informed decisions regarding the quantity and timing of feeding starting materials to increase the yield. Furthermore, this approach can serve as a platform method for analyzing the free Mg++ in complex sample matrices where preserving the native-like state of Mg++ binding is key for accurate quantitation.

Square Root Statistics of Density Matrices and Their Applications.

To estimate the degree of quantum entanglement of random pure states, it is crucial to understand the statistical behavior of entanglement indicators such as the von Neumann entropy, quantum purity, and entanglement capacity. These entanglement metrics are functions of the spectrum of density matrices, and their statistical behavior over different generic state ensembles have been intensively studied in the literature. As an alternative metric, in this work, we study the sum of the square root spectrum of density matrices, which is relevant to negativity and fidelity in quantum information processing. In particular, we derive the finite-size mean and variance formulas of the sum of the square root spectrum over the Bures-Hall ensemble, extending known results obtained recently over the Hilbert-Schmidt ensemble.

Continuous daytime and nighttime forecast of atmospheric optical turbulence from numerical weather prediction models.

Future satellite-to-ground optical communication systems will benefit from accurate forecasts of atmospheric optical turbulence; namely for site selection, for the routing and the operation of optical links, and for the design of optical communication terminals. This work presents a numerical approach based on the Weather Research and Forecasting software that enables continuous forecast of the refractive index structure parameter, C n2, vertical profiles. Two different C n2 models are presented and compared. One is based on monitoring the turbulent kinetic energy, while the other is a hybrid model using the Tatarskii equation to depict the free atmosphere region, and the Monin-Obukhov similarity theory for describing the boundary layer. The validity of both models is assessed by using thermosonde measurements from the Terrain-induced Rotor Experiment campaign, and from day and night measurements of the coherence length collected during a six-day campaign at Paranal observatory by a Shack-Hartmann Image Motion Monitor. The novelty of this work is the ability of the presented approach to continuously predict optical turbulence both during daytime and nighttime, and its validation with measurements in day and night conditions.

Demonstrating 24-hour continuous vertical monitoring of atmospheric optical turbulence.

We report what is believed to be the first example of fully continuous, 24-hour vertical monitoring of atmospheric optical turbulence. This is achieved using a novel instrument, the 24-hour Shack-Hartmann Image Motion Monitor (24hSHIMM). Optical turbulence is a fundamental limitation for applications such as free-space optical communications, where it limits the achievable bandwidth, and ground-based optical astronomy, restricting the observational precision. Knowledge of the turbulence enables us to select the best sites, design optical instrumentation and optimise the operation of ground-based optical systems. The 24hSHIMM estimates the vertical optical turbulence coherence length, time, angle and Rytov variance from the measurement of a four-layer vertical turbulence profile and a wind speed profile retrieved from meteorological forecasts. To illustrate our advance we show the values of these parameters recorded during a 36-hour, continuous demonstration of the instrument. Due to its portability and ability to work in stronger turbulence, the 24hSHIMM can also operate in urban locations, providing the field with a truly continuous, versatile turbulence monitor for all but the most demanding of applications.

Draft Genome Sequence of Nereida sp. Strain MMG025, Isolated from Giant Kelp.

Here, we report the draft genome sequence of Nereida sp. strain MMG025, isolated from the surface of giant kelp and assembled and analyzed by undergraduate students participating in a marine microbial genomics course. A genomic comparison suggests that MMG025 is a novel species, providing a resource for future microbiology and biotechnology investigations.

First stellar photons for an integrated optics discrete beam combiner at the William Herschel Telescope.

We present the first on-sky results of a four-telescope integrated optics discrete beam combiner (DBC) tested at the 4.2 m William Herschel Telescope. The device consists of a four-input pupil remapper followed by a DBC and a 23-output reformatter. The whole device was written monolithically in a single alumino-borosilicate substrate using ultrafast laser inscription. The device was operated at astronomical H-band (1.6 µm), and a deformable mirror along with a microlens array was used to inject stellar photons into the device. We report the measured visibility amplitudes and closure phases obtained on Vega and Altair that are retrieved using the calibrated transfer matrix of the device. While the coherence function can be reconstructed, the on-sky results show significant dispersion from the expected values. Based on the analysis of comparable simulations, we find that such dispersion is largely caused by the limited signal-to-noise ratio of our observations. This constitutes a first step toward an improved validation of the DBC as a possible beam combination scheme for long-baseline interferometry.

Flow virometry for process monitoring of live virus vaccines-lessons learned from ERVEBO.

Direct at line monitoring of live virus particles in commercial manufacturing of vaccines is challenging due to their small size. Detection of malformed or damaged virions with reduced potency is rate-limited by release potency assays with long turnaround times. Thus, preempting batch failures caused by out of specification potency results is almost impossible. Much needed are in-process tools that can monitor and detect compromised viral particles in live-virus vaccines (LVVs) manufacturing based on changes in their biophysical properties to provide timely measures to rectify process stresses leading to such damage. Using ERVEBO, MSD's Ebola virus vaccine as an example, here we describe a flow virometry assay that can quickly detect damaged virus particles and provide mechanistic insight into process parameters contributing to the damage. Furthermore, we describe a 24-h high throughput infectivity assay that can be used to correlate damaged particles directly to loss in viral infectivity (potency) in-process. Collectively, we provide a set of innovative tools to enable rapid process development, process monitoring, and control strategy implementation in large scale LVV manufacturing.

Adaptive Optics pre-compensated laser uplink to LEO and GEO.

We present the results from a Monte Carlo computer simulation of adaptive optics (AO) pre-compensated laser uplink propagation through the Earth's atmospheric turbulence from the ground to orbiting satellites. The simulation includes the so-called point-ahead angle and tests several potential AO mitigation modes such as tip/tilt or full AO from the downlink beam, and a laser guide star at the point ahead angle. The performance of these modes, as measured by metrics relevant for free-space optical communication, are compared with no correction and perfect correction. The aim of the study is to investigate fundamental limitations of free-space optical communications with AO pre-compensation and a point-ahead angle, therefore the results represent an upper bound of AO corrected performance, demonstrating the potential of pre-compensation technology. Performance is assessed with varying launch aperture size, wavelength, launch geometry, ground layer turbulence strength (i.e. day/night), elevation angle and satellite orbit (Low-Earth and Geostationary). By exploring this large parameter space we are able examine trends on performance with the aim of informing the design of future optical ground stations and demonstrating and quantifying the potential upper bounds of adaptive optics performance in free-space optical communications.

Metastatic MSSA infection of the spine and extremities.

Here, we present a rare case of metastatic methicillin sensitive Staphylococcus aureus (MSSA) infection arising from an unknown focus and spreading throughout the lumbar spine with associated pyomyositis of the paraspinal musculature, and septic arthritis of the knee, ankle and sternoclavicular joint. This case highlights the potential for missed aspects and delay in diagnosis in the care of metastatic S. aureus and the need for multispecialty intervention. Treatment of S. aureus infections requires a high index of suspicion and careful examination of multiple organ systems to identify the full extent of the disease. A discussion on metastatic S. aureus infection follows the report.

Radiographic Analysis of Cervical and Spinal Alignment in Multilevel ACDF with Lordotic Interbody Device.

BACKGROUND: Restoration and maintenance of cervical lordosis is an important clinical parameter in spine surgery. The purpose of this study was to determine the extent to which a multilevel anterior cervical discectomy and fusion (ACDF: greater than 3 levels) procedure restores cervical lordosis and the affect of increasing lordosis on sagittal vertical axis. METHODS: We performed a retrospective radiographic analysis of 69 patients who underwent multilevel ACDF by 2 surgeons between 2013 and 2014. We measured the global and segmental sagittal alignment of the cervical spine using the cobb method at 4 time intervals (preop, post op 4wks, 10wks and 6 months) as well as the sagittal vertical axis (SVA) using both a C1-S1 and C7-S1 plumb line methods at 2 time intervals (preop and post op 4wks). Radiographs were measured by three reviewers. RESULTS: Interrater reliability was good to excellent for all measurements. Cervical lordosis significantly increased from preop 10.26° to 4 weeks postop 19.44° and was maintained up to 6 months 19.34 (p<0.0005). Segmental cervical lordosis was also significantly increased from preop 8.22° to post op at 4 weeks (20.26°) and was maintained at post op 10weeks 20.30° and post op 6 months 19.56° (p<0.0005). C7-S1 SVA and C1-S1 SVA also significantly increased from 12.04mm preop to 27.49mm post op 4 wks (p<0.0005) and -1.93mm preop to 8.67mm post op (p<0.0005) respectively. A change in C2-C7 lordosis positively correlated with a change in C7-SVA and C1-SVA (r=0.37, P<0.005, and r=0.312, p<0.05 respectively). CONCLUSIONS: Multilevel ACDF significantly increases and maintains both segmental and global cervical lordosis up to 6 months after surgery. Increasing C2-C7 global lordosis is correlated with increasing positive sagittal vertical axis. Level of evidence: IV.

Comparative Study of Neural Network Frameworks for the Next Generation of Adaptive Optics Systems.

Many of the next generation of adaptive optics systems on large and extremely large telescopes require tomographic techniques in order to correct for atmospheric turbulence over a large field of view. Multi-object adaptive optics is one such technique. In this paper, different implementations of a tomographic reconstructor based on a machine learning architecture named "CARMEN" are presented. Basic concepts of adaptive optics are introduced first, with a short explanation of three different control systems used on real telescopes and the sensors utilised. The operation of the reconstructor, along with the three neural network frameworks used, and the developed CUDA code are detailed. Changes to the size of the reconstructor influence the training and execution time of the neural network. The native CUDA code turns out to be the best choice for all the systems, although some of the other frameworks offer good performance under certain circumstances.

Rare cause of neck pain: tumours of the posterior elements of the cervical spine.

Here we present two cases of primary bone tumours of the cervical spine in patients who had persistent neck pain-in one case, lasting 8 years. In each case, there was a delay in diagnosis and referral to a spine specialist was prolonged. Primary bone tumours of the spine are rare, which is in contrast to the wide prevalence of cervical neck pain. Many primary care providers may go an entire career without encountering a symptomatic primary cervical spine tumour. In this paper, we discuss the clinical course and treatment of each patient and review the current literature on primary bone tumours of the spine. Owing to the subtle roentgenographic findings of primary cervical tumours, we highlight the importance of advanced imaging in the clinical work-up of simple axial neck pain lasting >6 weeks to avoid misdiagnosis of serious pathology.

The epidemiology of thoracolumbar trauma: A meta-analysis.

PURPOSE: To describe the epidemiology of thoracolumbar fractures and associated injuries in blunt trauma patients. METHODS: A systematic review and metaanalysis was performed based on a MEDLINE database search using MeSH terms for studies matching our inclusion criteria. The search yielded 21 full-length articles, each sub-grouped according to content. Data extraction and multiple analyses were performed on descriptive data. RESULTS: The rate of thoracolumbar fracture in blunt trauma patients was 6.90% (±3.77, 95% CI). The rate of spinal cord injury was 26.56% (±10.70), and non-contiguous cervical spine fracture occurred in 10.49% (±4.17). Associated injury was as follows: abdominal trauma 7.63% (±9.74), thoracic trauma 22.64% (±13.94), pelvic trauma 9.39% (±6.45), extremity trauma 18.26% (±5.95), and head trauma 12.96% (±2.01). Studies that included cervical spine fracture with thoracolumbar fracture had the following rates of associated trauma: 3.78% (±5.94) abdominal trauma, 21.65% (±16.79) thoracic trauma, 3.62% (±1.07) pelvic trauma, 18.36% (±4.94) extremity trauma, and 15.45% (±11.70) head trauma. A subgroup of flexion distraction injuries showed an associated intra-abdominal injury rate of 38.70% (±13.30). The most common vertebra injured was L1 at a rate of 34.40% (±15.90). T7 was the most common non-junctional vertebra injured at 3.90% (±1.09). Burst/AO type A3 fractures were the most common morphology 39.50% (±16.30) followed by 33.60% (±15.10) compression/AO type A1, 14.20% (±8.08) fracture dislocation/AO type C, and 6.96% (±3.50) flexion distraction/AO type B. The most common etiology for a thoracolumbar fracture was motor vehicle collision 36.70% (±5.35), followed by high-energy fall 31.70% (±6.70). CONCLUSIONS: Here we report the incidence of thoracolumbar fracture in blunt trauma and the spectrum of associated injuries. To our knowledge, this paper provides the first epidemiological road map for blunt trauma thoracolumbar injuries.

Subacute hepatic necrosis mimicking veno-occlusive disease in a patient with HFE H63D homozygosity after allogeneic hematopoietic cell transplantation with busulfan conditioning.

Busulfan is a commonly used chemotherapeutic agent in myeloablative conditioning regimens for allogeneic hematopoietic cell transplantation (allo-HCT). It has been associated with sinusoidal-obstructive syndrome(SOS) as a life-threatening complication of myeloablative allo-HCT, yet it has not been found to cause severe hepatocellular injury, even in cases of significant accidental overdose.We report the case of a 31-year-old male with a history of high-risk myelodysplastic syndrome transitioning to acute myeloid leukemia, who in complete remission underwent allo-HCT using myeloablative busulfan­fludarabine conditioning, and who developed hepatic failure. While he met clinical criteria for SOS and was treated with defibrotide,liver biopsy demonstrated severe subacute hepatic necrosis and lacked characteristics of SOS. Further evaluation revealed that the patient was homozygous for the HFE H63D gene mutation, associated with hereditary hemochromatosis.Both Busulfan and iron overload related to HFE H63D homozygosity can cause oxidative stress resulting in cellular injury, and the cumulative effects of these risk factors are possibly responsible for the severe hepatocellular injury in this case, making our patient the first-known case of subacute hepatic necrosis related to busulfan administration.

An ANN-based smart tomographic reconstructor in a dynamic environment.

In astronomy, the light emitted by an object travels through the vacuum of space and then the turbulent atmosphere before arriving at a ground based telescope. By passing through the atmosphere a series of turbulent layers modify the light's wave-front in such a way that Adaptive Optics reconstruction techniques are needed to improve the image quality. A novel reconstruction technique based in Artificial Neural Networks (ANN) is proposed. The network is designed to use the local tilts of the wave-front measured by a Shack Hartmann Wave-front Sensor (SHWFS) as inputs and estimate the turbulence in terms of Zernike coefficients. The ANN used is a Multi-Layer Perceptron (MLP) trained with simulated data with one turbulent layer changing in altitude. The reconstructor was tested using three different atmospheric profiles and compared with two existing reconstruction techniques: Least Squares type Matrix Vector Multiplication (LS) and Learn and Apply (L + A).

Comparison of vibration mitigation controllers for adaptive optics systems.

Vibrations are detrimental to the performance of modern adaptive optics (AO) systems. In this paper, we describe new methods tested to mitigate the vibrations encountered in some of the instruments of the Gemini South telescope. By implementing a spectral analysis of the slope measurements from several wavefront sensors and an imager, we can determine the frequencies and magnitude of these vibrations. We found a persistent vibration at 55 Hz with others occurring occasionally at 14 and 100 Hz. Two types of AO controllers were designed and implemented, Kalman and H∞, in the multiconjugate AO tip-tilt loop. The first results show a similar performance for these advanced controllers and a clear improvement in vibration rejection and overall performance over the classical integrator scheme. It is shown that the reduction in the standard deviation of the residual slopes (as measured by wavefront sensors) is highly dependent on turbulence, wind speed, and vibration conditions, ranging--in terms of slopes RMS value--from an almost negligible reduction for high speed wind to a factor of 5 for a combination of low wind and strong vibrations.

Using artificial neural networks for open-loop tomography.

Modern adaptive optics (AO) systems for large telescopes require tomographic techniques to reconstruct the phase aberrations induced by the turbulent atmosphere along a line of sight to a target which is angularly separated from the guide sources that are used to sample the atmosphere. Multi-object adaptive optics (MOAO) is one such technique. Here, we present a method which uses an artificial neural network (ANN) to reconstruct the target phase given off-axis references sources. We compare our ANN method with a standard least squares type matrix multiplication method and to the learn and apply method developed for the CANARY MOAO instrument. The ANN is trained with a large range of possible turbulent layer positions and therefore does not require any input of the optical turbulence profile. It is therefore less susceptible to changing conditions than some existing methods. We also exploit the non-linear response of the ANN to make it more robust to noisy centroid measurements than other linear techniques.

ApoE: the link between Alzheimer's-related glucose hypometabolism and Aß deposition?

Alzheimer's disease (AD) is a complex, multifactorial progressive neurodegenerative disease. Pathologically, AD is characterized by extracellular deposits of amyloid beta (Aß) protein and intracellular accumulation of neurofibrillary tangles (NFTs) of tau. The central role of Aß protein in the AD etiology is well-established, and its increased deposition in AD brain is attributed to its decreased clearance from the brain. It is noteworthy that apolipoprotein E (ApoE), the most significant risk factor for late-onset AD, has been shown to play a vital role in brain Aß clearance and the ability of ApoE to do this depends mainly upon its lipidation status. Thus, lower ApoE lipidation status leading to decreased Aß clearance may underlie the increased Aß deposition observed in AD brain. In addition to the pathophysiological Aß deposits, AD is also characterized by certain metabolic changes. Among them, decreased cerebral glucose metabolism is one of the distinct characteristics of AD brain and is also observed in patients with Mild Cognitive Impairment (MCI) who subsequently develop AD. Thus, decreased cerebral glucose metabolism is an early event in AD pathology and may precede the neuropathological Aß deposition associated with AD. In this context, we hypothesize here that the decreased glucose metabolism in pre-AD and early AD stages, may lead to lower ApoE lipidation status, which in turn may lead to decreased clearance and hence, increased deposition of Aß protein in AD brain.