Background: Coronavirus disease 2019 (COVID-19) outbreaks occur in hospitals in many parts of the world. In hospital settings, the possibility of airborne transmission needs to be investigated thoroughly. Materials and Methods: There was a nosocomial outbreak of COVID-19 in a hematologic ward in a tertiary hospital, Seoul, Korea. We found 11 patients and guardians with COVID-19 through vigorous contact tracing and closed-circuit television monitoring. We found one patient who probably had acquired COVID-19 through airborne-transmission. We performed airflow investigation with simulation software, whole-genome sequencing of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Results: Of the nine individuals with COVID-19 who had been in the hematologic ward, six stayed in one multi-patient room (Room 36), and other three stayed in different rooms (Room 1, 34, 35). Guardian in room 35 was close contact to cases in room 36, and patient in room 34 used the shared bathroom for teeth brushing 40 minutes after index used.Airflow simulation revealed that air was spread from the bathroom to the adjacent room 1 while patient in room 1 did not used the shared bathroom. Airflow was associated with poor ventilation in shared bathroom due to dysfunctioning air-exhaust, grill on the door of shared bathroom and the unintended negative pressure of adjacent room. Conclusion Transmission of SARS-CoV-2 in the hematologic ward occurred rapidly in the multi-patient room and shared bathroom settings. In addition, there was a case of possible airborne transmission due to unexpected airflow.

Background: Remdesivir is widely used for the treatment of coronavirus disease 2019 (COVID-19), but controversies regarding its efficacy still remain. Methods: A retrospective cohort study was conducted to evaluate the effect of remdesivir on clinical and virologic outcomes of severe COVID-19 patients from June to July 2020. Primary clinical endpoints included clinical recovery, additional mechanical ventilator (MV) support, and duration of oxygen or MV support. Viral load reduction by hospital day (HD) 15 was evaluated by calculating changes in cycle threshold (Ct) values. Results: A total of 86 severe COVID-19 patients were evaluated including 48 remdesivirtreated patients. Baseline characteristics were not significantly different between the two groups. Remdesivir was administered an average of 7.42 days from symptom onset. The proportions of clinical recovery of the remdesivir and supportive care group at HD 14 (56.3% and 39.5%) and HD 28 (87.5% and 78.9%) were not statistically different. The proportion of patients requiring MV support by HD 28 was significantly lower in the remdesivir group than in the supportive care group (22.9% vs. 44.7%, P = 0.032), and MV duration was significantly shorter in the remdesivir group (average, 1.97 vs. 5.37 days; P = 0.017). Analysis of upper respiratory tract specimens demonstrated that increases of Ct value from HD 1–5 to 11–15 were significantly greater in the remdesivir group than the supportive care group (average, 10.19 vs. 5.36; P = 0.007), and the slope of the Ct value increase was also significantly steeper in the remdesivir group (average, 5.10 vs. 2.68; P = 0.007). Conclusion The remdesivir group showed clinical and virologic benefit in terms of MV requirement and viral load reduction, supporting remdesivir treatment for severe COVID-19.

Background: Coronavirus disease 2019 (COVID-19) outbreaks occur in hospitals in many parts of the world. In hospital settings, the possibility of airborne transmission needs to be investigated thoroughly. Materials and Methods: There was a nosocomial outbreak of COVID-19 in a hematologic ward in a tertiary hospital, Seoul, Korea. We found 11 patients and guardians with COVID-19 through vigorous contact tracing and closed-circuit television monitoring. We found one patient who probably had acquired COVID-19 through airborne-transmission. We performed airflow investigation with simulation software, whole-genome sequencing of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Results: Of the nine individuals with COVID-19 who had been in the hematologic ward, six stayed in one multi-patient room (Room 36), and other three stayed in different rooms (Room 1, 34, 35). Guardian in room 35 was close contact to cases in room 36, and patient in room 34 used the shared bathroom for teeth brushing 40 minutes after index used.Airflow simulation revealed that air was spread from the bathroom to the adjacent room 1 while patient in room 1 did not used the shared bathroom. Airflow was associated with poor ventilation in shared bathroom due to dysfunctioning air-exhaust, grill on the door of shared bathroom and the unintended negative pressure of adjacent room. Conclusion Transmission of SARS-CoV-2 in the hematologic ward occurred rapidly in the multi-patient room and shared bathroom settings. In addition, there was a case of possible airborne transmission due to unexpected airflow.

Background: Remdesivir is widely used for the treatment of coronavirus disease 2019 (COVID-19), but controversies regarding its efficacy still remain. Methods: A retrospective cohort study was conducted to evaluate the effect of remdesivir on clinical and virologic outcomes of severe COVID-19 patients from June to July 2020. Primary clinical endpoints included clinical recovery, additional mechanical ventilator (MV) support, and duration of oxygen or MV support. Viral load reduction by hospital day (HD) 15 was evaluated by calculating changes in cycle threshold (Ct) values. Results: A total of 86 severe COVID-19 patients were evaluated including 48 remdesivirtreated patients. Baseline characteristics were not significantly different between the two groups. Remdesivir was administered an average of 7.42 days from symptom onset. The proportions of clinical recovery of the remdesivir and supportive care group at HD 14 (56.3% and 39.5%) and HD 28 (87.5% and 78.9%) were not statistically different. The proportion of patients requiring MV support by HD 28 was significantly lower in the remdesivir group than in the supportive care group (22.9% vs. 44.7%, P = 0.032), and MV duration was significantly shorter in the remdesivir group (average, 1.97 vs. 5.37 days; P = 0.017). Analysis of upper respiratory tract specimens demonstrated that increases of Ct value from HD 1–5 to 11–15 were significantly greater in the remdesivir group than the supportive care group (average, 10.19 vs. 5.36; P = 0.007), and the slope of the Ct value increase was also significantly steeper in the remdesivir group (average, 5.10 vs. 2.68; P = 0.007). Conclusion The remdesivir group showed clinical and virologic benefit in terms of MV requirement and viral load reduction, supporting remdesivir treatment for severe COVID-19.

The detailed kinetics of the cytomegalovirus (CMV)-specific T cell response in hematopoietic stem cell transplant (HCT) recipients have not yet been fully assessed. We evaluated these kinetics of CMV-specific T cell response and factors associated with high CMV-specific T cell responses 1 year after HCT. In HCT recipients, CMV pp65 and IE1-specific ELISPOT assay were performed before HCT (D0), and at 30 (D30), 90 (D90), 180 (D180), and 360 (D360) days after HCT. Of the 51 HCT recipients with donor-positive (D+)/recipient-positive (R+) serology, 26 (51%) developed CMV infections after HCT. The patterns of post-transplantation reconstitution for CMV-specific T cell response were classified into 4 types: 1) an initial decrease at D30 followed by gradual T cell reconstitution without CMV infection (35%), 2) an initial decrease at D30 followed by gradual T cell reconstitution preceded by CMV infection (35%), 3) failure of gradual or constant T cell reconstitution (26%), and 4) no significant T cell reconstitution (4%). There was no significant difference between ELISPOT counts of D360 and those of D0. High CMV-specific T cell responses at D360 were not associated with high CMV-specific T cell response at D0, CMV infection, ganciclovir therapy, graft versus host disease (GVHD), and immunosuppressant use. In conclusion, there are 4 distinct patterns of reconstitution of the CMV-specific T cell response after HCT. In addition, reconstituted donor-origin CMV-specific T cell responses appeared to be constant until day 360 after HCT, regardless of the level of the pre-transplant CMV-specific T cell response, CMV infection, and immunosuppressant use.
Cytomegalovirus ; Enzyme-Linked Immunospot Assay ; Follow-Up Studies* ; Ganciclovir ; Graft vs Host Disease ; Hematopoietic Stem Cell Transplantation* ; Hematopoietic Stem Cells* ; Kinetics* ; Theophylline

The argon plasma jet (Ar-PJ) is widely used in medical fields such as dermatology and dentistry, and it is considered a promising tool for cancer therapy. However, the in vivo effects of Ar-PJ for medical uses have not yet been investigated, and there are no biological tools to determine the appropriate clinical dosages of Ar-PJ. In this study, we used the caudal fin and embryo of zebrafish as novel in vivo tools to evaluate the biosafety of Ar-PJ. Typically, Ar-PJ is known to induce cell death in two-dimensional (2D) cell culture systems. By contrast, no detrimental effects of Ar-PJ were shown in our 3D zebrafish systems composed of 2D cells. The Ar-PJ-treated caudal fins grew by an average length of 0.7 mm, similar to the length of the normally regenerating fins. Remarkably, Ar-PJ did not affect the expression patterns of Wnt8a and β-Catenin, which play important roles in fin regeneration. In the embryo system, 85% of the Ar-PJ-treated embryos hatched, and the lateral length of these embryos was ~3.3 mm, which are equivalent to the lengths of normal embryos. In particular, vasculogenesis, which is the main cellular process during tissue regeneration and embryogenesis, occurred normally under the Ar-PJ dose used in this study. Therefore, our biosafety evaluation tools that use living model systems can be used to provide an experimental guideline to determine the clinically safe dosage of Ar-PJ.
Argon* ; Cell Culture Techniques ; Cell Death ; Dentistry ; Dermatology ; Embryonic Development* ; Embryonic Structures ; Female ; Plasma* ; Pregnancy ; Regeneration* ; Zebrafish*

Amyotrophic lateral sclerosis (ALS) is a late-onset progressive neurodegenerative disease characterized by the loss of motor neurons in the spinal cord and brain. Mutations in Cu/Zn superoxide dismutase 1 (SOD1) are known to induce ALS. Although many research models have been developed, the exact pathological mechanism of ALS remains unknown. The recently developed induced pluripotent stem (iPS) cell technology is expected to illuminate the pathological mechanisms and new means of treatment for neurodegenerative diseases. To determine the pathological mechanism of ALS, we generated mouse iPS (miPS) cells from experimental ALS transgenic mice and control mice and characterized the cells using molecular biological methods. The generated miPS cells expressed many pluripotent genes and differentiated into three germ layers in vitro and in vivo. Motor neurons derived from ALS-related miPS cells recapitulated the pathological features of ALS. The ALS-model motor neurons showed SOD1 aggregates, as well as decreased cell survival rate and neurite length compared with wild-type motor neurons. Our study will be helpful in revealing the mechanism of motor neuronal cell death in ALS.
Amyotrophic Lateral Sclerosis ; Animals ; Brain ; Cell Death ; Cell Survival ; Germ Layers ; In Vitro Techniques ; Induced Pluripotent Stem Cells* ; Mice* ; Mice, Transgenic ; Motor Neurons* ; Neurites ; Neurodegenerative Diseases ; Spinal Cord ; Superoxide Dismutase

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease caused by the degeneration of motor neurons. Mutations in Cu/Zn superoxide dismutase (SOD1), including G93A, were reportedly linked to familial ALS. SOD1 is a key antioxidant enzyme, and is also one of the major targets for oxidative damage in the brains of patients suffering from Alzheimer's disease (AD). Several lines of evidence suggest that intracellular amyloid beta (Abeta) is associated with the pathogenesis of AD. In this report we demonstrate that intracellular Abeta directly interacts with SOD1, and that this interaction decreases the enzymatic activity of the enzyme. We observed Abeta-SOD1 aggregates in the perinuclear region of H4 cells, and mapped the SOD1 binding region to Abeta amino acids 26-42. Interestingly, intracellular Abeta binds to the SOD1 G93A mutant with greater affinity than to wild-type SOD1. This resulted in considerably less mutant enzymatic activity. Our study implicates a potential role for Abeta in the development of ALS by interacting with the SOD1 G93A mutant.
Amino Acid Sequence ; Amyloid beta-Protein/chemistry/*metabolism ; Amyotrophic Lateral Sclerosis/*enzymology ; Apoptosis ; Cell Line ; Cell Line, Tumor ; Humans ; Molecular Sequence Data ; Point Mutation ; Protein Binding ; Protein Interaction Domains and Motifs ; Superoxide Dismutase/genetics/*metabolism

Serine protease activity of high temperature requrement 2 (HtrA2) is essential for promoting cell death, as well as for protecting against cellular stresses. An X-ray crystallographic study described the formation of a pyramid shaped homotrimer that is a proteolytically competent form of HtrA2; however, little is known about effects of the trimeric structure of HtrA2 on the natural substrates. In this study, we generated the HtrA2 protein that has a single point mutation at the homotrimerization motif to assess relationship between structure and the proteolytic activity of HtrA2 on its substrates. Using gel filtration, a native gel electrophoresis system, and a co-precipitation assay, we confirm that phenylalanine 149 in HtrA2 is a crucial determinant for the formation of the HtrA2 homotrimeric structure. Moreover, we described that the HtrA2 monomeric form abolished not only autoproteolytic activity, but also the proteolytic activity against XIAP (X-linked inhibitor of apoptosis protein) known as the HtrA2 substrate. Taken together, the results indicate that the homotrimeric structure of HtrA2 is required for executing its serine protease activity.
Alanine/metabolism ; Amino Acid Motifs ; Amino Acid Sequence ; Amino Acid Substitution ; Cell Line ; Chromatography, Gel ; Crystallography, X-Ray ; Escherichia coli/genetics ; Glutathione Transferase/metabolism ; Hydrolysis ; Molecular Sequence Data ; Phenylalanine/metabolism ; Point Mutation ; Precipitin Tests ; Protein Structure, Tertiary ; Recombinant Fusion Proteins/metabolism ; Research Support, Non-U.S. Gov't ; Sequence Homology, Amino Acid ; Serine Endopeptidases/*chemistry/genetics/isolation & purification/*metabolism ; Structure-Activity Relationship ; Transfection

BACKGROUND: The resurgence of tuberculosis and outbreaks of multidrug resistant (MDR) tuberculosis have increased the emphasis for the development of new susceptibility testing of the Mycobacterium tuberculosis for the effective treatment and control of the disease. Conventional drug susceptibility testings, such as those using egg-based or agar-based media have some limits, such as the time required and difficulties in determining critical inhibitory concentrations, but these are still being used in many diagnostic laboratories because of no better alternatives, considering cost and accuracy. To overcome these limits, a rapid and simple method for new susceptibility testing, using live and dead assays, was applied for a bacterial cell viability assay to distinguish dead from live bacterial cells based on two-color fluorescence. MATERIALS AND METHODS STRAINS: Forty strains were used in this study, 20 susceptible to all antituberculosis drugs and the other 20 resistant to the four first line antituberculosis drugs isoniazid, rifampicin, streptomycin and ethambutol. ANTIBIOTICS: The four antibiotics were dissolved in 7H9 broth to make the following solutions: 0.1micro gram isoniazid(INH)/ml, 0.4micro gram rifampicin(RMP)/ml, 4.0micro gram streptomycin(SM)/ml and 4.0micro gram ethambutol(EMB)/ml. RESULTS: Live and dead Mycobacterium tuberculosis cells fluoresced green and red with the acridin (Syto 9) and propidium treatments, respectively. These results are very well accorded with conventional drug susceptibility testing by proportional method on Lowensen-Jensen media (L-J) containing 4 drugs (INH, RMP, EMB and SM), showing a 93.7 % accordance rate in susceptible strains and 95% in resistant strains. CONCLUSION: The results of the drug susceptibility testing using the live and dead bacterial cell assay showed high accordance rates compared with the conventional proportion method on L-J. This finding suggests that the live and dead bacterial cell assay can be used as an alternative to conventional drug susceptibility testing for M. tuberculosis strains.
Anti-Bacterial Agents ; Cell Differentiation* ; Cell Survival ; Disease Outbreaks ; Ethambutol ; Fluorescence ; Isoniazid ; Mycobacterium tuberculosis* ; Mycobacterium* ; Propidium ; Rifampin ; Streptomycin ; Tuberculosis