Single-particle chemical force microscopy to characterize virus surface chemistry

Two important viral surface characteristics are the hydrophobicity and surface charge, which determine the viral colloidal behavior and mobility. Chemical force microscopy allows the detection of viral surface chemistry in liquid samples with small amounts of virus sample. This single-particle method requires the functionalization of an atomic force microscope (AFM) probe and covalent bonding of viruses to a surface. A hydrophobic methyl-modified AFM probe was used to study the viral surface hydrophobicity, and an AFM probe terminated with either negatively charged carboxyl acid or positively charged quaternary amine was used to study the viral surface charge. With an understanding of viral surface properties, the way in which viruses interact with the environment can be better predicted

Inclusion of Youths with Disabilities in 4-H: A Scoping Literature Review

The Journal of Extension serves as a conduit for the dissemination of current research and practices within Extension and 4-H. We conducted a review of Journal of Extension articles published since passage of the Americans with Disabilities Act of 1990. Our purpose was to determine what practices, programs, and studies have occurred regarding inclusion in 4-H of youths with disabilities or special health care needs. The review resulted in detailed examination of 16 articles and revealed information about Extension professionals\u27 attitudes toward inclusion, strategies and program approaches related to inclusion, and specific areas that need to be addressed further to increase inclusion

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

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Adsorption of a non-enveloped mammalian virus to functionalized nanofibers

In the pursuit of finding superior methods to remove pathogens from drinking water, this study examines the adsorption of a non-enveloped, mammalian virus to highly charged nanofibers. N-[(2-Hydroxyl-3-trimethylammonium) propyl] chitosan (HTCC) nanofibers were synthesized by the addition of a quaternary amine to chitosan. HTCC was blended with polyvinyl alcohol (PVA) to produce nanofibers by electrospinning. The nanofibers were stabilized against water by crosslinking with glutaraldehyde. When studied in the range of 100-200. nm in diameter, larger fibers were able to adsorb about 90% more virus than smaller fibers. The kinetics of the adsorption was modeled with pseudo-first order kinetics and equilibrium was achieved in as little as 10. min. Equilibrium adsorption was modeled with the Freundlich isotherm with a Freundlich constant of 1.4. When the Freundlich constant deviates from 1, this demonstrates that there is heterogeneity at the adsorption surface. The heterogeneity likely occurs at the nanofiber surface since a polymeric blend of two polymers was used to electrospin the nanofibers. The model mammalian virus, porcine parvovirus (PPV), has a fairly homogeneous, icosahedral protein capsid available for adsorption. The fast adsorption kinetics and high capacity of the nanofibers make HTCC/PVA a potential filter material for the removal of pathogens from drinking water. © 2014 Elsevier B.V

Virus adsorption of water-stable quaternized chitosan nanofibers

The burden of unsafe drinking water is responsible for millions of deaths each year. To relieve this burden, we are in search of an inexpensive material that can adsorb pathogens from drinking water. In this pursuit, we have studied the natural carbohydrate, chitosan. To impart virus removal features, chitosan has been functionalized with a quaternary amine to form quaternized chitosan N-[(2-hydroxyl-3-trimethylammonium) propyl] chitosan (HTCC). HTCC can be electrospun into nanofibers with the non-ionogenic polyvinyl alcohol (PVA), creating a high surface area mat. High surface area is a major requirement for effective adsorption processes. HTCC is antiviral and antimicrobial, making it a good material for water purification. However, HTCC dissolves in water. We have explored the parameters to crosslink the nanofibers with glutaraldehyde. We have imparted water stability so there is a maximum of 30% swelling of the fibers after 6 h in water. The water stable fibers retain their ability to adsorb virus, as shown for an enveloped and nonenveloped virus. HTCC now has the potential to be incorporated into a microfiltration membrane that can remove viruses. This could create an inexpensive, low pressure filtration membrane for drinking water purification. © 2014 Elsevier Ltd. All rights reserved

Virus and chlorine adsorption onto guanidine modified cellulose nanofibers using covalent and hydrogen bonding

Unsafe drinking water leads to millions of human deaths each year, while contaminated wastewater discharges are a significant threat to aquatic life. To relieve the burden of unsafe water, we are in search of an inexpensive material that can adsorb pathogenic viruses from drinking water and adsorb toxic residual chlorine from wastewater. To impart virus and chlorine removal abilities to cellulosic materials, we modified the primary hydroxyl group with a positively charged guanidine group, to yield guanidine modified cellulose derivatives. Microcrystalline cellulose (MC) bearing covalently bonded guanidine hydrochloride (MC-GC) and hydrogen-bonded guanidine hydrochloride (MC-GH) were synthesized, and electrospun into nanofibers after blending with the non-ionogenic polyvinyl alcohol (PVA), to produce large pore sized, high surface area membranes. The MC-GC/PVA and MC-GH/PVA nanofibers were stabilized against water dissolution by crosslinking with glutaraldehyde vapor. The water-stable MC-GC/PVA mats were able to remove more than 4 logs of non-enveloped porcine parvovirus (PPV) and enveloped Sindbis virus and reached 58% of chlorine removal. The MC-GC/PVA nanofibers demonstrated better performance for pathogen removal and dechlorination than MC-GH/PVA nanofibers. This first study of MC-GC/PVA electrospun mats for virus removal shows they are highly effective and merit additional research for virus removal

Experimental and computational surface hydrophobicity analysis of a non-enveloped virus and proteins

© 2017 Elsevier B.V. The physical characteristics of viruses needs to be understood in order to manipulate the interaction of viruses with host cells, as well as to create specific molecular recognition techniques to detect, purify, and remove viruses. Viruses are generally believed to be positively charged at physiological pH, but there are few other defining characteristics. Here, we have experimentally and computationally demonstrated that a non-enveloped virus is more hydrophobic than a panel of model proteins. Reverse-phase and hydrophobic interaction chromatography and ANS fluorescence determined the experimental hydrophobic strength of each entity. Computational surface hydrophobicity was calculated by the solvent exposed surface area of the protein weighted by the hydrophobicity of each amino acid. The results obtained indicate a strong correlation between the computational surface hydrophobicity and experimentally determined hydrophobicity using reverse-phase chromatography and ANS fluorescence. The surface hydrophobicity did not compare strongly to the weighted average of the amino acid sequence hydrophobicity. This demonstrates that our simple method of calculating the surface hydrophobicity gives general hydrophobicity information about proteins and viruses with crystal structures. In the process, this method demonstrated that porcine parvovirus (PPV) is more hydrophobic than the model proteins used in this study. This adds an additional dimension to currently known virus characteristics and can improve our manipulation of viruses for gene therapy targeting, surface adsorption and general understanding of virus interactions