Measurement of Cerenkov radiation induced by the gamma-rays of Co-60 therapy units using wavelength shifting fiber.

In this study, a wavelength shifting fiber that shifts ultra-violet and blue light to green light was employed as a sensor probe of a fiber-optic Cerenkov radiation sensor. In order to characterize Cerenkov radiation generated in the developed wavelength shifting fiber and a plastic optical fiber, spectra and intensities of Cerenkov radiation were measured with a spectrometer. The spectral peaks of light outputs from the wavelength shifting fiber and the plastic optical fiber were measured at wavelengths of 500 and 510 nm, respectively, and the intensity of transmitted light output of the wavelength shifting fiber was 22.2 times higher than that of the plastic optical fiber. Also, electron fluxes and total energy depositions of gamma-ray beams generated from a Co-60 therapy unit were calculated according to water depths using the Monte Carlo N-particle transport code. The relationship between the fluxes of electrons over the Cerenkov threshold energy and the energy depositions of gamma-ray beams from the Co-60 unit is a near-identity function. Finally, percentage depth doses for the gamma-ray beams were obtained using the fiber-optic Cerenkov radiation sensor, and the results were compared with those obtained by an ionization chamber. The average dose difference between the results of the fiber-optic Cerenkov radiation sensor and those of the ionization chamber was about 2.09%.

Comparison of Nasal Swabs, Nasopharyngeal Swabs, and Saliva Samples for the Detection of SARS-CoV-2 and other Respiratory Virus Infections.

Background: Nasal swabs and saliva samples are being considered alternatives to nasopharyngeal swabs (NPSs) for detecting severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2); however, few studies have compared the usefulness of nasal swabs, NPSs, and saliva samples for detecting SARS-CoV-2 and other respiratory virus infections. We compared the positivity rates and concentrations of viruses detected in nasal swabs, NPSs, and saliva samples using cycle threshold (Ct) values from real-time PCR tests for respiratory viruses. Methods: In total, 236 samples (48 five-rub and 10 10-rub nasal swabs, 96 NPSs collected using two different products, 48 saliva swabs, and 34 undiluted saliva samples) from 48 patients (34 patients with SARS-CoV-2 and 14 with other respiratory virus infections) and 40 samples from eight healthy controls were obtained. The PCR positivity and Ct values were compared using Allplex Respiratory Panels 1/2/3 and Allplex SARS-CoV-2 real-time PCR. Results: NPSs showed the lowest Ct values (indicating the highest virus concentrations); however, nasal and saliva samples yielded positive results for SARS-CoV-2 and other respiratory viruses. The median Ct value for SARS-CoV-2 E gene PCR using nasal swab samples collected with 10 rubs was significantly different from that obtained using nasal swabs collected with five rubs (Ct=24.3 vs. 28.9; P=0.002), but not from that obtained using NPSs. Conclusions: Our results confirm that the NPS is the best sample type for detecting respiratory viruses, but nasal swabs and saliva samples can be alternatives to NPSs. Vigorously and sufficiently rubbed nasal swabs can provide SARS-CoV-2 concentrations similar to those obtained with NPSs.

Development of a Potent Stabilizer for Long-Term Storage of Foot-and-Mouth Disease Vaccine Antigens.

A local virus isolate, O/SKR/JC/2014 (O JC), has been considered as a candidate vaccine strain in the development of a domestic foot-and-mouth disease (FMD) vaccine in Korea. However, producing and preserving a sufficient quantity of intact vaccine antigens from the O JC strain was difficult owing to its distinctive structural instability compared to other candidate vaccine strains. Based on this feature, the O JC strain was adopted as a model virus for the stabilization study to determine the optimal stabilizer composition, which enables long-term storage of the FMD vaccine antigen in both aqueous and frozen phases. In contrast to O JC vaccine antigens stored in routinely used Tris-buffered or phosphate-buffered saline, those stored in Tris-KCl buffer showed extended shelf-life at both 4 °C and -70 °C. Additionally, the combined application of 10% sucrose and 5% lactalbumin hydrolysate could protect O JC 146S particles from massive structural breakdown in an aqueous state for up to one year. The stabilizer composition was also effective for other FMDV strains, including serotypes A and Asia 1. With this stabilizer composition, FMD vaccine antigens could be flexibly preserved during the general production process, pending status under refrigeration and banking under ultrafreezing.

Foot-and-Mouth Disease Virus Evades Innate Immune Response by 3C-Targeting of MDA5.

Foot-and-mouth disease (FMD) is a highly contagious disease caused by FMD virus (FMDV) in cloven-hoofed animals. Retinoic acid-inducible gene I (RIG-I) and melanoma differentiation-associated gene 5 (MDA5) are representative receptors in the cytoplasm for the detection of viral RNA and trigger antiviral responses, leading to the production of type I interferon. Although MDA5 is a crucial receptor for sensing picornavirus RNA, the interplay between MDA5 and FMDV is relatively unknown compared to the interplay between RIG-I and FMDV. Here, we observed that the FMDV infection inhibits MDA5 protein expression. Of the non-structural proteins, the Lb and 3C proteinases (Lbpro and 3Cpro) were identified to be primarily responsible for this inhibition. However, the inhibition by 3Cpro was independent of proteasome, lysosome and caspase-dependent pathway and was by 3C protease activity. A direct interaction between 3Cpro and MDA5 protein was observed. In conclusion, this is the first report that 3Cpro inhibits MDA5 protein expression as a mechanism to evade the innate immune response during FMDV infection. These results elucidate the pathogenesis of FMDV and provide fundamental insights for the development of a novel vaccine or therapeutic agent.

Scale-Up Production of Type O and A Foot-and-Mouth Disease Bivalent Vaccine and Its Protective Efficacy in Pigs.

South Korea has experienced FMD outbreaks almost every year since 2014. Therefore, a novel local vaccine that can cover various topotypes of viruses is required. Two virus strains, O/Boeun/SKR/2017 and A/Yeoncheon/SKR/2017, were cultured up to the pilot scale based on the optimized conditions set up on the flask scale. FMDV particles (146S) of 2 µg/mL or more were obtained from the virus culture supernatant using a 100 L bioreactor. The viruses were fully inactivated using binary ethylenimine within 16 h through two inactivation cycles and mixed with an adjuvant into a bivalent vaccine (types O and A) consisting of 15 µg viruses per strain. The experimental bivalent vaccine showed a broad spectrum of high neutralizing antibody titers against heterologous viruses, including type O Cathay strain and type A Asia topotypes, except for GVII. The 50% protective dose was determined as 12.5 for O/Boeun/SKR/2017 and 15.6 for A/Yeoncheon/SKR/2017. Collectively, we expect that the bivalent vaccine could protect against FMDV types O and A circulating in South Korea and neighboring countries. To our knowledge, this is the first report demonstrating that the vaccine strains could be successfully scaled-up to a 100 L bioreactor, with the determination of its protective efficacy in pigs.

Efficient Removal of Non-Structural Protein Using Chloroform for Foot-and-Mouth Disease Vaccine Production.

To differentiate foot-and-mouth disease (FMD)-infected animals from vaccinated livestock, non-structural proteins (NSPs) must be removed during the FMD vaccine manufacturing process. Currently, NSPs cannot be selectively removed from FMD virus (FMDV) culture supernatant. Therefore, polyethylene glycol (PEG) is utilized to partially separate FMDV from NSPs. However, some NSPs remain in the FMD vaccine, which after repeated immunization, may elicit NSP antibodies in some livestock. To address this drawback, chloroform at a concentration of more than 2% (v/v) was found to remove NSP efficiently without damaging the FMDV particles. Contrary to the PEG-treated vaccine that showed positive NSP antibody responses after the third immunization in goats, the chloroform-treated vaccine did not induce NSP antibodies. In addition to this enhanced vaccine purity, this new method using chloroform could maximize antigen recovery and the vaccine production time could be shortened by two days due to omission of the PEG processing phase. To our knowledge, this is the first report to remove NSPs from FMDV culture supernatant by chemical addition. This novel method could revolutionize the conventional processes of FMD vaccine production.

Application of Heparin Affinity Chromatography to Produce a Differential Vaccine without Eliciting Antibodies against the Nonstructural Proteins of the Serotype O Foot-and-Mouth Disease Viruses.

Although polyethylene glycol (PEG) application is the most widely used method in removing nonstructural proteins (NSPs) for foot-and-mouth disease (FMD) vaccine production, some NSPs remaining in the antigen could elicit antibodies against these proteins after repeated vaccinations in livestock. Therefore, the purpose of this study was to purify the FMD virus (FMDV) via affinity chromatography using a heparin ligand to remove most proteins, including NSPs. Chromatography showed an intact virus (146S) particle recovery of 70% or more for three different strains of serotype O FMDV (two locally isolated strains and one genetically modified strain). The experimental vaccine made with antigens eluted via heparin affinity chromatography elicited virus-neutralizing antibodies against homologous viruses but did not induce antibodies against NSPs even after five immunizations in goats; this indicated that the NSPs were effectively removed from the vaccine antigen. This method can then be used to produce a higher-quality vaccine compared with PEG application in terms of the purity of the FMD vaccine. Therefore, this result would be an important groundwork for advanced FMD vaccine manufacturing in the near future.

Recombinant vesicular stomatitis virus glycoprotein carrying a foot-and-mouth disease virus epitope as a vaccine candidate.

Foot-and-mouth disease (FMD) is one of the most highly contagious animal diseases. In an effort to overcome the drawbacks of the currently used inactivated foot-and-mouth disease virus vaccine, a novel recombinant protein carrying foot-and-mouth disease virus VP1 GH loop epitope linked to vesicular stomatitis virus glycoprotein was expressed in a baculovirus system. Its antigenicity was confirmed with ELISA using monoclonal antibody against foot-and-mouth disease virus. Twice immunizations one month apart in field pigs resulted in a significant antibody increase compared to the glutathione S-transferase carrier containing the same epitope and the commercial vaccine. To my knowledge, this is the first report that the recombinant protein vaccine was superior to the current vaccine. Although further studies are required to examine their immunogenicity in a large number of animals, this study sheds light on the development of a novel recombinant protein vaccine that could be easily produced in a general laboratory as an alternative to the current FMD vaccine, which requires a biosafety level 3 containment facility for vaccine production.

Determination of the optimal method for the concentration and purification of 146S particles for foot-and-mouth disease vaccine production.

After the severe outbreak of foot-and-mouth disease (FMD) in South Korea in 2010, the Korean government implemented a vaccination policy and set out to develop an FMD vaccine using a local FMD virus (FMDV) strain. As a part of the basic research for domestic FMD vaccine development, three methods commonly used for the concentration and purification of FMDV to produce FMD vaccine antigens were compared. Among common concentration methods, including polyethylene glycol (PEG) precipitation, ammonium sulfate precipitation, and ultrafiltration, the most effective method both for concentrating 146S particles and eliminating non-structural proteins (NSPs) was found to be PEG precipitation. Classical PEG precipitation showed the highest recovery of 146S particles (85.4%) with removing 99.8% of the other proteins, including NSPs. To the author's knowledge, this is the first study to compare the current three methods with regard to quantifying intact virus particles (146S). These findings may provide important insights for the development of new FMD vaccines using a local FMDV strain in the near future.

A simple and rapid assay to evaluate purity of foot-and-mouth disease vaccine before animal experimentation.

Currently, foot-and-mouth disease (FMD) vaccine purity is tested in cattle to detect antibodies against the non-structural protein (NSP) after repeated immunization with the final vaccine product. In case of vaccine failure, the manufacturing company would suffer significant economic loss. To prevent such unfortunate losses with the final vaccine product, in vitro testing is required to quantitate an NSP antigen during the manufacturing process prior to animal experiments. A novel lateral-flow assay device was developed using a monoclonal antibody (MAb) against the 3B NSP. To determine the minimal amount of NSP required to elicit antibodies in livestock, goats were immunized several times with various concentrations of either the recombinant 3AB (rec.3AB) protein or FMD virus culture supernatant. Antibodies against 3AB were elicited after a second immunization with 10.6 ng to 42.5 ng of rec.3AB and a third immunization with a 10-fold diluted FMD virus culture supernatant in goats. The lateral-flow assay device detected the minimal amount of rec.3AB and native NSP in FMD virus culture supernatant required to induce NSP antibodies in goats. The in vitro assay device is simple and economical, provides rapid results, and should be useful for FMD vaccine-manufacturing companies prior to conducting animal experiments to test the vaccine purity.

Discarded Egg Yolk as an Alternate Source of Poly(3-Hydroxybutyrate-co-3-hydroxyhexanoate).

Many poultry eggs are discarded worldwide because of infection (i.e., avian flu) or presence of high levels of pesticides. The possibility of adopting egg yolk as a source material to produce polyhydroxyalkanoate (PHA) biopolymer was examined in this study. Cupriavidus necator Re2133/pCB81 was used for the production of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) or poly(3HHx), a polymer that would normally require long-chain fatty acids as carbon feedstocks for the incorporation of 3HHx monomers. The optimal medium contained 5% egg yolk oil and ammonium nitrate as a nitrogen source, with a carbon/nitrogen (C/N) ratio of 20. Time course monitoring using the optimized medium was conducted for 5 days. Biomass production was 13.1 g/l, with 43.7% co-polymer content. Comparison with other studies using plant oils and the current study using egg yolk oil revealed similar polymer yields. Thus, discarded egg yolks could be a potential source of PHA.

Production of glutaric acid from 5-aminovaleric acid by robust whole-cell immobilized with polyvinyl alcohol and polyethylene glycol.

Glutaric acid is an attractive C5 dicarboxylic acid with wide applications in the biochemical industry. Glutaric acid can be produced by fermentation and bioconversion, and several of its biosynthesis pathways have been well characterized, especially the simple pathway involving glutaric acid from l-lysine using 5-aminovaleric acid. We previously reported the production of glutaric acid using 5-aminovaleric acid and α-ketoglutaric acid by a whole-cell reaction, resulting in a high conversion yield. In this study, we sought to enhance the stability and reusability of this whole-cell system for realizing the efficient production of glutaric acid under harsh reaction conditions. To this end, various matrices were screened to immobilize Escherichia coli whole-cell overexpressing 4-aminobutyrate aminotransferase (GabT), succinate semi-aldehyde dehydrogenase (GabD), and NAD(P)H oxidase (NOX). We ultimately selected a PVA-PEG gel (LentiKats®) for cell entrapment, and several factors of the reaction were optimized. The optimal temperature and pH were 35 °C and 8.5, respectively. Treatment with Tween 80 as a surfactant, as well as additional NOX, was found to be effective. Under the optimized conditions, an immobilized cell retained 55% of its initial activity even after the eighth cycle, achieving 995.2 mM accumulated glutaric acid, whereas free cell lost most of their activity after only two cycles. This optimized whole-cell system can be used in the large-scale production of glutaric acid.

Determination of optimal age for single vaccination of growing pigs with foot-and-mouth disease bivalent vaccine in South Korea.

In South Korea, pigs were vaccinated once between 8 and 12 weeks of age because of the injection-site granulomas. Therefore this study was performed to determine the optimal age for single vaccination of growing pigs with the currently used type O FMD vaccine. With 498 pigs divided into four groups, seroprevalence of the antibody was analyzed with enzyme-linked immunosorbent assay. Although double vaccination is necessary to completely protect growing pigs from FMD virus infection with the current vaccine, the age of 8 weeks can be considered as the optimal age for piglet vaccination if the booster injection is unavailable.

Effect of powdered activated carbon on the performance of an aerobic membrane bioreactor: comparison between cross-flow and submerged membrane systems.

The effects of powdered activated carbon (PAC) on the performance of an aerobic membrane bioreactor were investigated under two different filtration modes: cross-flow and submerged filtration. Under a cross-flow microfiltration mode, floc breakage resulting from sludge recirculation caused a rapid decrease in the microbial floc size and the release of colloidal and soluble components including extracellular polymeric substances (EPS). The released components caused rapid loss of membrane permeability by the formation of a dense cake layer on the surface of the membrane. Biological activated carbon (BAC) sludge formed by the addition of PAC released lower amounts of fine colloids and EPS. Although the pattern of floc breakage of the BAC sludge by pumping shear was similar to that of conventional activated sludge, PAC in the BAC microbial floc adsorbed or entrapped some microfloc components into the floc and reduced permeability loss by approximately 35% compared with conventional activated sludge. Under a submerged microfiltration mode, the effect of PAC addition was more pronounced. For the BAC sludge, the increase in transmembrane pressure was more sluggish and, thus, the operating interval could be extended up to 3 times that for the submerged membrane bioreactor with normal activated sludge.

Serological responses after vaccination of growing pigs with foot-and-mouth disease trivalent (type O, A and Asia1) vaccine.

Korea experienced its fifth Foot-and-Mouth Disease (FMD) outbreak (type O) since 1934 from November 2010 to April 2011. The Korean government initiated emergency vaccination for all FMD-susceptible domestic animals in December 2010 using type O FMD vaccines. Starting in September 2011, trivalent FMD vaccines (types O, A, and Asia1) were used for the vaccination of all animals. This study was performed to identify the appropriate time for FMD vaccination in growing pigs when vaccination is applied only once (at either 8 weeks or 12 weeks of age). Seroprevalences from growing pigs under different vaccination regimens (once or twice) were also studied. A total of 526 growing pigs on 7 farms were used in this study. This study showed that the vaccination of growing pigs at both 8 and 12 weeks of age resulted in higher seroprevalences (97.5% in type O, 92.3% in type A and 99.4% in type Asia1) than did a single vaccination at 8 weeks of age (86.7% in type O, 88.0% in type A and 93.0% in type Asia1) (P<0.05). Pigs vaccinated once at 8 weeks of age showed much higher seroprevalences than pigs vaccinated once at 12 weeks of age (60.9% in type O, 62.8% in type A and 77.6% in Asia1, respectively) (P<0.05).

Tissue reactions to suture materials in the oral mucosa of beagle dogs.

PURPOSE: The objective of this study was to compare and evaluate the inflammatory responses of three widely used suture materials in the keratinized gingiva and buccal mucosa of beagle dogs. METHODS: Silk, polyglycolic acid, and nylon sutures were placed within the mandibular keratinized gingiva and maxillary buccal mucosa of four male beagle dogs. Biopsies were taken 3, 7, and 14 days after suturing. Specimens were prepared with hematoxylin-eosin stain for evaluation under a light microscope. RESULTS: The suture materials placed in the oral mucosa elicited more inflammatory reactions than did those placed in the keratinized gingiva. The multifilament suture materials caused more inflammatory tissue reactions than did the monofilament suture materials in the oral mucosa. CONCLUSIONS: If oral hygiene is well maintained and suture materials are placed in the keratinized gingiva, silk, nylon, and polyglycolic acid are considered to be proper suture materials for oral surgery. However, it is advisable to use monofilament suture materials if the suture site is within the oral mucosa.