Thermophysiological and Perceptual Responses of Amateur Healthcare Workers: Impacts of Ambient Condition, Inner-Garment Insulation and Personal Cooling Strategy.

While personal protective equipment (PPE) protects healthcare workers from viruses, it also increases the risk of heat stress. In this study, the effects of environmental heat stress, the insulation of the PPE inner-garment layer, and the personal cooling strategy on the physiological and perceptual responses of PPE-clad young college students were evaluated. Three levels of wet bulb globe temperatures (WBGT = 15 °C, 28 °C, and 32 °C) and two types of inner garments (0.37 clo and 0.75 clo) were chosen for this study. In an uncompensable heat stress environment (WBGT = 32 °C), the effects of two commercially available personal cooling systems, including a ventilation cooling system (VCS) and an ice pack cooling system (ICS) on the heat strain mitigation of PPE-clad participants were also assessed. At WBGT = 15 °C with 0.75 clo inner garments, mean skin temperatures were stabilized at 31.2 °C, Hskin was 60-65%, and HR was about 75.5 bpm, indicating that the working scenario was on the cooler side. At WBGT = 28 °C, Tskin plateaued at approximately 34.7 °C, and the participants reported "hot" thermal sensations. The insulation reduction in inner garments from 0.75 clo to 0.37 clo did not significantly improve the physiological thermal comfort of the participants. At WBGT = 32 °C, Tskin was maintained at 35.2-35.7 °C, Hskin was nearly 90% RH, Tcore exceeded 37.1 °C, and the mean HR was 91.9 bpm. These conditions indicated that such a working scenario was uncompensable, and personal cooling to mitigate heat stress was required. Relative to that in NCS (no cooling), the mean skin temperatures in ICS and VCS were reduced by 0.61 °C and 0.22 °C, respectively, and the heart rates were decreased by 10.7 and 8.5 bpm, respectively. Perceptual responses in ICS and VCS improved significantly throughout the entire field trials, with VCS outperforming ICS in the individual cooling effect.

Asthma mortality attributable to ambient temperatures: A case-crossover study in China.

BACKGROUND: Whether ambient temperature exposure contributes to death from asthma remains unknown to date. We therefore conducted a case-crossover study in China to quantitatively evaluate the association and burden of ambient temperature exposure on asthma mortality. METHODS: Using data from the National Mortality Surveillance System in China, we conducted a time-stratified case-crossover study of 15 888 individuals who lived in Hubei and Jiangsu province, China and died from asthma as the underlying cause in 2015-2019. Individual-level exposures to air temperature and apparent temperature on the date of death and 21 days prior were assessed based on each subject's residential address. Distributed lag nonlinear models based on conditional logistic regression were used to quantify exposure-response associations and calculate fraction and number of deaths attributable to non-optimum ambient temperatures. RESULTS: We observed a reverse J-shaped association between air temperature and risk of asthma mortality, with a minimum mortality temperature of 21.3 °C. Non-optimum ambient temperature is responsible for substantial excess mortality from asthma. In total, 26.3% of asthma mortality were attributable to non-optimum temperatures, with moderate cold, moderate hot, extreme cold and extreme hot responsible for 21.7%, 2.4%, 2.1% and 0.9% of asthma mortality, respectively. The total attributable fraction and number was significantly higher among adults aged less than 80 years in hot temperature. CONCLUSIONS: Exposure to non-optimum ambient temperature, especially moderate cold temperature, was responsible for substantial excess mortality from asthma. These findings have important implications for planning of public-health interventions to minimize the adverse respiratory damage from non-optimum ambient temperature.

The impact of temperature on the transmissibility potential and virulence of COVID-19 in Tokyo, Japan.

Assessing the impact of temperature on COVID-19 epidemiology is critical for implementing non-pharmaceutical interventions. However, few studies have accounted for the nature of contagious diseases, i.e., their dependent happenings. We aimed to quantify the impact of temperature on the transmissibility and virulence of COVID-19 in Tokyo, Japan, employing two epidemiological measurements of transmissibility and severity: the effective reproduction number ([Formula: see text]) and case fatality risk (CFR). We estimated the [Formula: see text] and time-delay adjusted CFR and to subsequently assess the nonlinear and delayed effect of temperature on [Formula: see text] and time-delay adjusted CFR. For [Formula: see text] at low temperatures, the cumulative relative risk (RR) at the first temperature percentile (3.3 °C) was 1.3 (95% confidence interval (CI): 1.1-1.7). As for the virulence to humans, moderate cold temperatures were associated with higher CFR, and CFR also increased as the temperature rose. The cumulative RR at the 10th and 99th percentiles of temperature (5.8 °C and 30.8 °C) for CFR were 3.5 (95% CI: 1.3-10.0) and 6.4 (95% CI: 4.1-10.1). Our results suggest the importance to take precautions to avoid infection in both cold and warm seasons to avoid severe cases of COVID-19. The results and our proposed approach will also help in assessing the possible seasonal course of COVID-19 in the future.

Estimating the cause-specific relative risks of non-optimal temperature on daily mortality: a two-part modelling approach applied to the Global Burden of Disease Study.

BACKGROUND: Associations between high and low temperatures and increases in mortality and morbidity have been previously reported, yet no comprehensive assessment of disease burden has been done. Therefore, we aimed to estimate the global and regional burden due to non-optimal temperature exposure. METHODS: In part 1 of this study, we linked deaths to daily temperature estimates from the ERA5 reanalysis dataset. We modelled the cause-specific relative risks for 176 individual causes of death along daily temperature and 23 mean temperature zones using a two-dimensional spline within a Bayesian meta-regression framework. We then calculated the cause-specific and total temperature-attributable burden for the countries for which daily mortality data were available. In part 2, we applied cause-specific relative risks from part 1 to all locations globally. We combined exposure-response curves with daily gridded temperature and calculated the cause-specific burden based on the underlying burden of disease from the Global Burden of Diseases, Injuries, and Risk Factors Study, for the years 1990-2019. Uncertainty from all components of the modelling chain, including risks, temperature exposure, and theoretical minimum risk exposure levels, defined as the temperature of minimum mortality across all included causes, was propagated using posterior simulation of 1000 draws. FINDINGS: We included 64·9 million individual International Classification of Diseases-coded deaths from nine different countries, occurring between Jan 1, 1980, and Dec 31, 2016. 17 causes of death met the inclusion criteria. Ischaemic heart disease, stroke, cardiomyopathy and myocarditis, hypertensive heart disease, diabetes, chronic kidney disease, lower respiratory infection, and chronic obstructive pulmonary disease showed J-shaped relationships with daily temperature, whereas the risk of external causes (eg, homicide, suicide, drowning, and related to disasters, mechanical, transport, and other unintentional injuries) increased monotonically with temperature. The theoretical minimum risk exposure levels varied by location and year as a function of the underlying cause of death composition. Estimates for non-optimal temperature ranged from 7·98 deaths (95% uncertainty interval 7·10-8·85) per 100 000 and a population attributable fraction (PAF) of 1·2% (1·1-1·4) in Brazil to 35·1 deaths (29·9-40·3) per 100 000 and a PAF of 4·7% (4·3-5·1) in China. In 2019, the average cold-attributable mortality exceeded heat-attributable mortality in all countries for which data were available. Cold effects were most pronounced in China with PAFs of 4·3% (3·9-4·7) and attributable rates of 32·0 deaths (27·2-36·8) per 100 000 and in New Zealand with 3·4% (2·9-3·9) and 26·4 deaths (22·1-30·2). Heat effects were most pronounced in China with PAFs of 0·4% (0·3-0·6) and attributable rates of 3·25 deaths (2·39-4·24) per 100 000 and in Brazil with 0·4% (0·3-0·5) and 2·71 deaths (2·15-3·37). When applying our framework to all countries globally, we estimated that 1·69 million (1·52-1·83) deaths were attributable to non-optimal temperature globally in 2019. The highest heat-attributable burdens were observed in south and southeast Asia, sub-Saharan Africa, and North Africa and the Middle East, and the highest cold-attributable burdens in eastern and central Europe, and central Asia. INTERPRETATION: Acute heat and cold exposure can increase or decrease the risk of mortality for a diverse set of causes of death. Although in most regions cold effects dominate, locations with high prevailing temperatures can exhibit substantial heat effects far exceeding cold-attributable burden. Particularly, a high burden of external causes of death contributed to strong heat impacts, but cardiorespiratory diseases and metabolic diseases could also be substantial contributors. Changes in both exposures and the composition of causes of death drove changes in risk over time. Steady increases in exposure to the risk of high temperature are of increasing concern for health. FUNDING: Bill & Melinda Gates Foundation.

Association between growth of the SARS-CoV-2 epidemic and increased coughing rate in cold temperatures: surveillance in Hokkaido, Japan.

OBJECTIVE: Although the transmissibility of SARS-CoV-2 in winter is thought to increase through viral droplets when coughing, current epidemiological data in this regard are limited. SETTING: Using the national epidemiological surveillance data in the autumn and winter seasons in Hokkaido, Japan, between February 2020 and February 2021, we analysed the relationship between case increase ratio and prevalence rate of coughing in patients with PCR-confirmed SARS-CoV-2 in two age groups (0-40s and 50-100s) with concomitant air temperature and humidity. PARTICIPANTS: The 7893 cases of symptomatic PCR-positive patients consisted of 5361 cases in the young age group and 2532 cases in the older age group. PRIMARY AND SECONDARY OUTCOME MEASURES: Pearson's correlation analysis and regression models were used to assess the relationships. Sex-adjusted OR of having cough in the young and old age groups in the autumn and winter seasons was calculated using logistic regression analysis. RESULTS: The monthly prevalence rate of coughing in the young age group was negatively correlated with temperature (r=-0.77, p<0.05), and in the old age group it was negatively correlated with humidity (r=-0.71, p<0.05). Quadratic regression models were fitted for the relationship between cold temperatures and rate of coughing rate in the young age group and case increase ratios. The sex-adjusted OR of having a cough in the young age group in winter was 1.18 (95% CI 1.05 to 1.31) as compared with autumn. CONCLUSIONS: The results suggest increased rate of coughing contributes to the epidemic of SARS-CoV-2 in the winter. An effective control with a focus on these trends should be considered.

The Impact of Protective Gloves on Manual Dexterity in Cold Environments-A Pilot Study.

Our research aimed to determine the impact of two types of protective gloves. The research tested the glove performance on men exposed to a range of temperatures reflecting the working conditions in fruit and vegetable processing. The gloves were assessed for performance within the time required to complete a specific manual task and for performance relative to the subjective thermal sensations in the male subjects. Six males participated in a total of 3 study variants: at +5 °C (with double gloves and single glove), at -1 °C (with double gloves and single glove) and in reference conditions +20 °C (without gloves), in which they performed manual tasks. The measurement of manual task performance time was used to assess manual dexterity. Subjective thermal sensations were determined. Differences in the time required to complete specific tasks were observed between the variants with gloves (both at a temperature of +5 °C and -1 °C), and without gloves (p < 0.05). The type of protective gloves had an impact on the time needed to complete manual tasks and therefore may affect manual dexterity.

On the footsteps of Hippocrates, Sanctorius and Harvey to better understand the influence of cold on the occurrence of COVID-19 in European countries in 2020.

COVID-19 pandemic has been characterized by a pattern of consecutive declines and regrowth in European countries in 2020. After being partially regressed during the summer, the reappearance of the infection during fall 2020 in many temperate countries strongly suggests that temperature and cold may play a role in influencing the infectivity and virulence of SARS-CoV-2. While promoting medicine as an art, Hippocrates interpreted with logical reasoning the occurrence of diseases such as epidemics, as a consequence of environmental factors, in particular climatic variations. During the Renaissance, Sanctorius was one of the first to perform quantitative measurements, and Harvey discovered the circulation of blood by performing experimental procedures in animals. We think that a reasoning mixing various observations, measurements and experiments is fundamental to understand how cold increases infectivity and virulence of SARS-CoV-2. By this review, we provide evidence linking cold, angiotensin-II, vasoconstriction, hypoxia and aerobic glycolysis (the Warburg effect) to explain how cold affects the epidemiology of COVID-19. Also, a low humidity increases virus transmissibility, while a warm atmosphere, a moderate airway humidity, and the production of vasodilator angiotensin 1-7 by ACE2 are less favorable to the virus entry and/or its development. The meteorological and environmental parameters impacting COVID-19 pandemic should be reintegrated into a whole perspective by taking into account the different factors influencing transmissibility, infectivity and virulence of SARS-CoV-2. To understand the modern enigma represented by COVID-19, an interdisciplinary approach is surely essential.

Detection of SARS-CoV-2 by real-time PCR under challenging pre-analytical conditions reveals independence of swab media and cooling chain.

With global demand for SARS-CoV-2 testing ever rising, shortages in commercially available viral transport media pose a serious problem for laboratories and health care providers. For reliable diagnosis of SARS-CoV-2 and other respiratory viruses, executed by Real-time PCR, the quality of respiratory specimens, predominantly determined by transport and storage conditions, is crucial. Therefore, our aim was to explore the reliability of minimal transport media, comprising saline or the CDC recommended Viral Transport Media (HBSS VTM), for the diagnosis of SARS-CoV-2 and other respiratory viruses (influenza A, respiratory syncytial virus, adenovirus, rhinovirus and human metapneumovirus) compared to commercial products, such as the Universal Transport Media (UTM). We question the assumptions, that the choice of medium and temperature for storage and transport affect the accuracy of viral detection by RT-PCR. Both alternatives to the commercial transport medium (UTM), HBSS VTM or saline, allow adequate detection of SARS-CoV-2 and other respiratory viruses, regardless of storage temperatures up to 28 °C and storage times up to 28 days. Our study revealed the high resilience of SARS-CoV-2 and other respiratory viruses, enabling proper detection in clinical specimens even after long-time storage at high temperatures, independent of the transport medium's composition.

Cold and dry winter conditions are associated with greater SARS-CoV-2 transmission at regional level in western countries during the first epidemic wave.

Higher transmissibility of SARS-CoV-2 in cold and dry weather conditions has been hypothesized since the onset of the COVID-19 pandemic but the level of epidemiological evidence remains low. During the first wave of the pandemic, Spain, Italy, France, Portugal, Canada and USA presented an early spread, a heavy COVID-19 burden, and low initial public health response until lockdowns. In a context when testing was limited, we calculated the basic reproduction number (R0) in 63 regions from the growth in regional death counts. After adjusting for population density, early spread of the epidemic, and age structure, temperature and humidity were negatively associated with SARS-CoV-2 transmissibility. A reduction of mean absolute humidity by 1 g/m3 was associated with a 0.15-unit increase of R0. Below 10 °C, a temperature reduction of 1 °C was associated with a 0.16-unit increase of R0. Our results confirm a dependency of SARS-CoV-2 transmissibility to weather conditions in the absence of control measures during the first wave. The transition from summer to winter, corresponding to drop in temperature associated with an overall decrease in absolute humidity, likely contributed to the intensification of the second wave in north-west hemisphere countries. Non-pharmaceutical interventions must be adjusted to account for increased transmissibility in winter conditions.

Ubiquitous convalescent plasma: An artificial universal plasma for COVID-19 patients.

OBJECTIVES AND BACKGROUND: In December 2019, the first case of COVID-19 was reported in Wuhan, China. Its causative virus, is a novel strain of RNA viruses with high mortality rate. There is no definitive treatment, but among available approaches the use of recovered patients' plasma containing specific antibodies can enhance the immune response against coronavirus. However, the dearth of eligible donors and also ABO incompatibility in plasma transfusion, have limited this therapeutic method. Therefore, it is highly desirable to introduce a simple procedure that allows efficient reduction or even removal of natural ABO antibodies. Accordingly, we aimed to evaluate a RBC-mediated adsorption technique that reduces the titer of the mentioned antibodies in plasma. METHODS/MATERIALS: This experimental study was conducted in Kerman University of Medical Sciences, Kerman, Iran. The pre- and post-incubation antibody titers of 168 plasma samples were determined. For incubation, each plasma sample was exposed (60 min) to different percentages of RBCs at room temperature or 4 °C. RESULTS: The results evidenced that both the concentration of RBCs and temperature had significant decreasing effects on antibody titer (P < 0.001) and all concentrations significantly reduced titer. Compared to RT, 4 °C further reduced the antibody titer. Overall, the best incubation condition for reducing antibody titer in all blood groups was 4 °C and 2% RBCs concentration. CONCLUSION: The presented adsorption procedure is able to produce universal plasma (we call it Ubiquitous Convalescent Plasma) with a non-immunogenic level of ABO mismatch antibodies which can be used for COVID-19 patients with any type of blood group with desirable simplicity, feasibility, and efficacy.

Temperature and population density influence SARS-CoV-2 transmission in the absence of nonpharmaceutical interventions.

As COVID-19 continues to spread across the world, it is increasingly important to understand the factors that influence its transmission. Seasonal variation driven by responses to changing environment has been shown to affect the transmission intensity of several coronaviruses. However, the impact of the environment on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remains largely unknown, and thus seasonal variation remains a source of uncertainty in forecasts of SARS-CoV-2 transmission. Here we address this issue by assessing the association of temperature, humidity, ultraviolet radiation, and population density with estimates of transmission rate (R). Using data from the United States, we explore correlates of transmission across US states using comparative regression and integrative epidemiological modeling. We find that policy intervention ("lockdown") and reductions in individuals' mobility are the major predictors of SARS-CoV-2 transmission rates, but, in their absence, lower temperatures and higher population densities are correlated with increased SARS-CoV-2 transmission. Our results show that summer weather cannot be considered a substitute for mitigation policies, but that lower autumn and winter temperatures may lead to an increase in transmission intensity in the absence of policy interventions or behavioral changes. We outline how this information may improve the forecasting of COVID-19, reveal its future seasonal dynamics, and inform intervention policies.

Interrelationship between daily COVID-19 cases and average temperature as well as relative humidity in Germany.

COVID-19 pandemic continues to obstruct social lives and the world economy other than questioning the healthcare capacity of many countries. Weather components recently came to notice as the northern hemisphere was hit by escalated incidence in winter. This study investigated the association between COVID-19 cases and two components, average temperature and relative humidity, in the 16 states of Germany. Three main approaches were carried out in this study, namely temporal correlation, spatial auto-correlation, and clustering-integrated panel regression. It is claimed that the daily COVID-19 cases correlate negatively with the average temperature and positively with the average relative humidity. To extract the spatial auto-correlation, both global Moran's [Formula: see text] and global Geary's [Formula: see text] were used whereby no significant difference in the results was observed. It is evident that randomness overwhelms the spatial pattern in all the states for most of the observations, except in recent observations where either local clusters or dispersion occurred. This is further supported by Moran's scatter plot, where states' dynamics to and fro cold and hot spots are identified, rendering a traveling-related early warning system. A random-effects model was used in the sense of case-weather regression including incidence clustering. Our task is to perceive which ranges of the incidence that are well predicted by the existing weather components rather than seeing which ranges of the weather components predicting the incidence. The proposed clustering-integrated model associated with optimal barriers articulates the data well whereby weather components outperform lag incidence cases in the prediction. Practical implications based on marginal effects follow posterior to model diagnostics.

Stability of SARS-CoV-2 Spike G614 Variant Surpasses That of the D614 Variant after Cold Storage.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) carrying the D614G mutation on the spike protein is the predominant circulating variant and is associated with enhanced infectivity. However, whether this dominant variant can potentially spread through the cold chain and whether the spike protein affects virus stability after cold storage remain unclear. To compare the infectivity of two SARS-CoV-2 variants, namely, SARS-CoV-2 variants with spike protein with the D614 mutation (S-D614) and G614 mutation (S-G614), after different periods of refrigeration (4°C) and freezing (-20°C). We also determined the integrity of the viral RNA and the ability of the spike protein to bind angiotensin-converting enzyme 2 (ACE2) after storage at these conditions. The results showed that SARS-CoV-2 was more stable and infectious after storage at -20°C than at 4°C. Particularly, the S-G614 variant was found to be more stable than the S-D614 variant. The spike protein of the S-G614 variant had better binding ability with the ACE2 receptor than that of the S-D614 variant after storage at -20°C for up to 30 days. Our findings revealed that SARS-CoV-2 remains stable and infectious after refrigeration or freezing, and their stability and infectivity up to 30 days depends on the spike variant. Stability and infectivity are related to each other, and the higher stability of S-G614 compared to that of S-D614 may contribute to rapid viral spread of the S-G614 variant.IMPORTANCE It has been observed that variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are more stable and infectious after storage at -20°C than at 4°C. A SARS-CoV-2 S-D614G variant is currently the most dominant variant in circulation and is associated with enhanced infectivity. We compared the stability of two SARS-CoV-2 variants: the early S-D614 variant carrying the D614 spike protein and the new S-G614 variant carrying the G614 spike protein, stored at both 4°C and -20°C for different periods. We observed that SARS-CoV-2 remains stable and infectious after refrigeration or freezing, which further depends on the spike variant, that is, the ability of the spike protein to bind with the ACE2 receptor with higher efficiency. The high stability of the S-G614 variant also explains its rapid spread and infectivity. Therefore, precautions should be taken during and after handling food preserved under cold conditions.

Impaired performance of SARS-CoV-2 antigen-detecting rapid diagnostic tests at elevated and low temperatures.

Antigen-detecting rapid diagnostic tests (Ag-RDTs) can complement molecular diagnostics for COVID-19. The recommended temperature for storage of SARS-CoV-2 Ag-RDTs ranges between 2-30 °C. In the global South, mean temperatures can exceed 30 °C. In the global North, Ag-RDTs are often used in external testing facilities at low ambient temperatures. We assessed analytical sensitivity and specificity of eleven commercially-available SARS-CoV-2 Ag-RDTs using different storage and operational temperatures, including short- or long-term storage and operation at recommended temperatures or at either 2-4 °C or at 37 °C. The limits of detection of SARS-CoV-2 Ag-RDTs under recommended conditions ranged from 1.0×106- 5.5×107 genome copies/mL of infectious SARS-CoV-2 cell culture supernatant. Despite long-term storage at recommended conditions, 10 min pre-incubation of Ag-RDTs and testing at 37 °C resulted in about ten-fold reduced sensitivity for five out of 11 SARS-CoV-2 Ag-RDTs, including both Ag-RDTs currently listed for emergency use by the World Health Organization. After 3 weeks of storage at 37 °C, eight of the 11 SARS-CoV-2 Ag-RDTs exhibited about ten-fold reduced sensitivity. Specificity of SARS-CoV-2 Ag-RDTs using cell culture supernatant from common respiratory viruses was not affected by storage and testing at 37 °C, whereas false-positive results occurred at outside temperatures of 2-4 °C for two out of six tested Ag-RDTs, again including an Ag-RDT recommended by the WHO. In summary, elevated temperatures impair sensitivity, whereas low temperatures impair specificity of SARS-CoV-2 Ag-RDTs. Consequences may include false-negative test results at clinically relevant virus concentrations compatible with transmission and false-positive results entailing unwarranted quarantine assignments. Storage and operation of SARS-CoV-2 Ag-RDTs at recommended conditions is essential for successful usage during the pandemic.

A Numerical Analysis of the Cooling Performance of a Hybrid Personal Cooling System (HPCS): Effects of Ambient Temperature and Relative Humidity.

Hybrid personal cooling systems (HPCS) incorporated with ventilation fans and phase change materials (PCMs) have shown its superior capability for mitigating workers' heat strain while performing heavy labor work in hot environments. In a previous study, the effects of thermal resistance of insulation pads, and latent heat and melting temperature of PCMs on the HPCS's thermal performance have been investigated. In addition to the aforementioned factors, environmental conditions, i.e., ambient temperature and relative humidity, also significantly affect the thermal performance of the HPCS. In this paper, a numerical parametric study was performed to investigate the effects of the environmental temperature and relative humidity (RH) on the thermal management of the HPCS. Five levels of air temperature under RH = 50% (i.e., 32, 34, 36, 38 and 40 °C) and four levels of environmental RH at two ambient temperatures of 36 and 40 °C were selected (i.e., RH = 30, 50, 70 and 90%) for the numerical analysis. Results show that high environmental temperatures could accelerate the PCM melting process and thereby weaken the cooling performance of HPCS. In the moderately hot environment (36 °C), HPCS presented good cooling performance with the maximum core temperature at around 37.5 °C during excise when the ambient RH ≤ 70%, whereas good cooling performance could be only seen under RH ≤ 50% in the extremely hot environment (40 °C). Thus, it may be concluded that the maximum environmental RH under which the HPCS exhibiting good cooling performance decreases with an increase in the environmental temperature.

Production of high-quality SARS-CoV-2 antigens: Impact of bioprocess and storage on glycosylation, biophysical attributes, and ELISA serologic tests performance.

Serological assays are valuable tools to study SARS-CoV-2 spread and, importantly, to identify individuals that were already infected and would be potentially immune to a virus reinfection. SARS-CoV-2 Spike protein and its receptor binding domain (RBD) are the antigens with higher potential to develop SARS-CoV-2 serological assays. Moreover, structural studies of these antigens are key to understand the molecular basis for Spike interaction with angiotensin converting enzyme 2 receptor, hopefully enabling the development of COVID-19 therapeutics. Thus, it is urgent that significant amounts of this protein became available at the highest quality. In this study, we produced Spike and RBD in two human derived cell hosts: HEK293-E6 and Expi293F™. We evaluated the impact of different and scalable bioprocessing approaches on Spike and RBD production yields and, more importantly, on these antigens' quality attributes. Using negative and positive sera collected from human donors, we show an excellent performance of the produced antigens, assessed in serologic enzyme-linked immunosorbent assay (ELISA) tests, as denoted by the high specificity and sensitivity of the test. We show robust Spike productions with final yields of approx. 2 mg/L of culture that were maintained independently of the production scale or cell culture strategy. To the best of our knowledge, the final yield of 90 mg/L of culture obtained for RBD production, was the highest reported to date. An in-depth characterization of SARS-CoV-2 Spike and RBD proteins was performed, namely the antigen's oligomeric state, glycosylation profiles, and thermal stability during storage. The correlation of these quality attributes with ELISA performance show equivalent reactivity to SARS-CoV-2 positive serum, for all Spike and RBD produced, and for all storage conditions tested. Overall, we provide straightforward protocols to produce high-quality SARS-CoV-2 Spike and RBD antigens, that can be easily adapted to both academic and industrial settings; and integrate, for the first time, studies on the impact of bioprocess with an in-depth characterization of these proteins, correlating antigen's glycosylation and biophysical attributes to performance of COVID-19 serologic tests.