Virus Purification Framework And Enhancement In Aqueous Two-Phase System

Viral infections regularly pose detrimental health risks to humans. Preventing viral infections through global immunization requires the production of large doses of vaccines. The increasing demand for vaccines, especially during pandemics such as COVID-19, has challenged current manufacturing strategy to develop advanced unit operations with high throughput capability. Over the decade, the upstream processing responsible for synthesizing viral products in cell cultures has shown significant success in yielding high titers of viruses and virus-like particles. The progress in the upstream stage has now shifted the bottleneck to the downstream processing (DSP). Overlooked for decades, the DSP responsible for viral product purification from the cell culture contaminants requires a makeover with the development of new purification strategies and an upgrade in the traditional unit operations. The current DSP train employing chromatography and filtration methods have been suboptimal in efficiently processing comparatively complex and fragile viral particles. Thus, the lack of platform technology for viral vaccine and biotherapeutic DSP has led to a search for alternative and innovative methods that have not only high-throughput capabilities but also have potential for continuous operation. In the pool of potential technologies, aqueous two-phase system (ATPS) has shown to be a promising candidate with the numerous advantages over conventional methods. However, an unambiguous and complex biomolecule partitioning mechanism has required a large experimental setup for optimizing virus purification. This work focused on a framework utilizing a phase diagram of a rationalized polyethylene glycol-citrate system to optimize virus purification. The partitioning behavior of two non-enveloped viruses, porcine parvovirus (PPV) and human rhinovirus-14 (HRV), were studied in various system compositions. A tie-line length framework was utilized to define the systems and relate the partitioning behavior of viruses with different surface physicochemical characteristics

Performance Evaluation of 802.11P Protocol for Emergency Application

Vehicle manufacturers have adopted this technology to assist the drivers drive their car like alarming the driver if he is close to some object while driving or safely parking the car to name a few. However, the main difference in an Adhoc network that involves a vehicle is the mobility factor. A vehicle is constantly in motion varying its location with time which means constant change in its position and surroundings. With the increase in the number of vehicles running in the road the probability of road accidents increases. The aim of the vehicle manufacturers and government agencies is to provide safe driving experience to save lives and assets. Sensor nodes has gone a long way from being just a technology to being an integral part of our life and its no surprise that vehicle manufacturers have incorporated them to create VANET that has a dedicated 802.11p protocol. In this essay based on my thesis, we show that 802.11p protocol is performing as per expected and all the nodes receive packet as should be the case. Simulation is done using an open source, discrete event driven tool called NS2 where a node generates and transmits a packet which is passed on by the other nodes. The simulation is performed considering node distance, retrans-mission attempt, and contention window. The results were analysed on the basis of con-figuration parameter mentioned. We evaluate the performance of 802.11p protocol by studying the output based on the node distance, retransmission attempt, and contention window. The graph generated from the simulation showed that all nodes receives a mes-sage which shows that 802.11p protocol is in fact working as intended. The most surprising result was that all of the nodes received the packet generated by the first node

Data Governance in Data Mesh Infrastructures: The Saxo Bank Case Study

Data governance (DG) is the management of data in a manner that the value of data is maximised and data related risks are minimised. Three aspects of DG are data catalogue, data quality, and data ownership and these aim to provide transparency, foster trust, and manage access and control the data. DG solution involves change management and alignment of incentives and mere technology is not enough to address this. In this paper we aim to provide a holistic view of data governance that is a synthesis of academic and practitioner viewpoints, and conclude by giving an example of a pilot case study (Saxo Bank) where authors worked on tech and cultural interventions to address the data governance challenges

Outcomes of patients treated with minimally invasive plate osteosynthesis in the management of distal tibial fractures

Background: Distal tibial fractures are a common injury, often caused by high-energy trauma, and can result in significant morbidity. Traditional surgical management with open reduction internal fixation (ORIF) is associated with complications including infection and soft tissue injury. Minimally invasive plate osteosynthesis (MIPO) has emerged as a potential alternative, with advantages including less soft tissue injury and a faster return to function.  The study aimed to evaluate the outcomes of MIPO for distal tibial fractures. Method: Sample size of 38 patients between 18-80 years of age with distal tibial fractures treated with MIPO was included. Patient demographics, fracture characteristics, surgical details and postop outcomes collected. Results: A total of 38 patients with distal tibia fractures with a mean age of 44.36 years were included in the study. The mean duration of hospital stay was 12.71 days, and the time required for a union was 18.61 weeks. The AOFAS score improved significantly from 3 to 6 months after surgery. However, 15 patients (34.88%) reported complications, with deep infection and skin necrosis being the most common. Out of 43 patients, 11 underwent culture and sensitivity testing, with no growth in 5 cases, and the remaining cases showed various bacterial infections. Seven patients underwent implant removal due to complications. Conclusions: The study provides valuable information about the use of MIPO in treating distal tibia fractures, as well as the risk of complications associated with this approach

Single-particle characterization of SARS-CoV-2 isoelectric point and comparison to variants of interest

SARS-CoV-2, the cause of COVID-19, is a new, highly pathogenic coronavirus, which is the third coronavirus to emerge in the past 2 decades and the first to become a global pandemic. The virus has demonstrated itself to be extremely transmissible and deadly. Recent data suggest that a targeted approach is key to mitigating infectivity. Due to the proliferation of cataloged protein and nucleic acid sequences in databases, the function of the nucleic acid, and genetic encoded proteins, we make predictions by simply aligning sequences and exploring their homology. Thus, similar amino acid sequences in a protein usually confer similar biochemical function, even from distal or unrelated organisms. To understand viral transmission and adhesion, it is key to elucidate the structural, surface, and functional properties of each viral protein. This is typically first modeled in highly pathogenic species by exploring folding, hydrophobicity, and isoelectric point (IEP). Recent evidence from viral RNA sequence modeling and protein crystals have been inadequate, which prevent full understanding of the IEP and other viral properties of SARS-CoV-2. We have thus experimentally determined the IEP of SARS-CoV-2. Our findings suggest that for enveloped viruses, such as SARS-CoV-2, estimates of IEP by the amino acid sequence alone may be unreliable. We compared the experimental IEP of SARS-CoV-2 to variants of interest (VOIs) using their amino acid sequence, thus providing a qualitative comparison of the IEP of VOIs

Design Rules for Sequestration of Viruses into Polypeptide Complex Coacervates

Encapsulation is a strategy that has been used to facilitate the delivery and increase the stability of proteins and viruses. Here, we investigate the encapsulation of viruses via complex coacervation, which is a liquid–liquid phase separation resulting from the complexation of oppositely charged polymers. In particular, we utilized polypeptide-based coacervates and explored the effects of peptide chemistry, chain length, charge patterning, and hydrophobicity to better understand the effects of the coacervating polypeptides on virus incorporation. Our study utilized two nonenveloped viruses, porcine parvovirus (PPV) and human rhinovirus (HRV). PPV has a higher charge density than HRV, and they both appear to be relatively hydrophobic. These viruses were compared to characterize how the charge, hydrophobicity, and patterning of chemistry on the surface of the virus capsid affects encapsulation. Consistent with the electrostatic nature of complex coacervation, our results suggest that electrostatic effects associated with the net charge of both the virus and polypeptide dominated the potential for incorporating the virus into a coacervate, with clustering of charges also playing a significant role. Additionally, the hydrophobicity of a virus appears to determine the degree to which increasing the hydrophobicity of the coacervating peptides can enhance virus uptake. Nonintuitive trends in uptake were observed with regard to both charge patterning and polypeptide chain length, with these parameters having a significant effect on the range of coacervate compositions over which virus incorporation was observed. These results provide insights into biophysical mechanisms, where sequence effects can control the uptake of proteins or viruses into biological condensates and provide insights for use in formulation strategies

Thermostabilization of viruses via complex coacervation

Widespread vaccine coverage for viral diseases could save the lives of millions of people each year. For viral vaccines to be effective, they must be transported and stored in a narrow temperature range of 2–8 °C. If temperatures are not maintained, the vaccine may lose its potency and would no longer be effective in fighting disease; this is called the cold storage problem. Finding a way to thermally stabilize a virus and end the need to transport and store vaccines at refrigeration temperatures will increase access to life-saving vaccines. We explore the use of polymer-rich complex coacervates to stabilize viruses. We have developed a method of encapsulating virus particles in liquid complex coacervates that relies on the electrostatic interaction of viruses with polypeptides. In particular, we tested the incorporation of two model viruses; a non-enveloped porcine parvovirus (PPV) and an enveloped bovine viral diarrhea virus (BVDV) into coacervates formed from poly(lysine) and poly(glutamate). We identified optimal conditions (i.e., the relative amount of the two polypeptides) for virus encapsulation, and trends in this composition matched differences in the isoelectric point of the two viruses. Furthermore, we were able to achieve a ∼103–104-fold concentration of virus into the coacervate phase, such that the level of virus remaining in the bulk solution approached our limit of detection. Lastly, we demonstrated a significant enhancement of the stability of non-enveloped PPV during an accelerated aging study at 60 °C over the course of a week. Our results suggest the potential for using coacervation to aid in the purification and formulation of both enveloped and non-enveloped viruses, and that coacervate-based formulations could help limit the need for cold storage throughout the transportation and storage of vaccines based on non-enveloped viruses

Thermostabilization of Viruses via Complex Coacervation

Widespread vaccine coverage for viral diseases could save the lives of millions of people each year. For viral vaccines to be effective, they must be transported and stored in a narrow temperature range of 2-8°C. If temperatures are not maintained, the vaccine may lose its potency and would no longer be effective in fighting disease; this is called the cold storage problem. Finding a way to thermally stabilize a virus and end the need to transport and store vaccines at refrigeration temperatures will increase access to life-saving vaccines. We explore the use of polymer-rich complex coacervates to stabilize viruses. We have developed a method of encapsulating virus particles in liquid complex coacervates that relies on the electrostatic interaction of viruses with polypeptides. In particular, we tested the incorporation of two model viruses; a non-enveloped porcine parvovirus (PPV) and an enveloped bovine viral diarrhea virus (BVDV) into coacervates formed from poly(lysine) and poly(glutamate). We identified optimal conditions (i.e., the relative amount of the two polypeptides) for virus encapsulation, and trends in this composition matched differences in the isoelectric point of the two viruses. Furthermore, we were able to achieve a ~103 – 104-fold concentration of virus into the coacervate phase, such that the level of virus remaining in the bulk solution approached our limit of detection. Lastly, we demonstrated a significant enhancement of the stability of non-enveloped PPV during an accelerated aging study at 60°C over the course of a week. Our results suggest the potential for using coacervation to aid in the purification and formulation of both enveloped and non-enveloped viruses, and that coacervate-based formulations could help limit the need for cold storage throughout the transportation and storage of vaccines based on non-enveloped viruses