SARS-CoV-2 seroprevalence among blood donors in Uganda: 2019-2022.

BACKGROUND: The true burden of COVID-19 in low- and middle-income countries remains poorly characterized, especially in Africa. Even prior to the availability of SARS-CoV-2 vaccines, countries in Africa had lower numbers of reported COVID-19 related hospitalizations and deaths than other regions globally. METHODS: Ugandan blood donors were evaluated between October 2019 and April 2022 for IgG antibodies to SARS-CoV-2 nucleocapsid (N), spike (S), and five variants of the S protein using multiplexed electrochemiluminescence immunoassays (MesoScale Diagnostics, Rockville, MD). Seropositivity for N and S was assigned using manufacturer-provided cutoffs and trends in seroprevalence were estimated by quarter. Statistically significant associations between N and S antibody seropositivity and donor characteristics in November-December 2021 were assessed by chi-square tests. RESULTS: A total of 5393 blood unit samples from donors were evaluated. N and S seropositivity increased throughout the pandemic to 82.6% in January-April 2022. Among seropositive individuals, N and S antibody levels increased ≥9-fold over the study period. In November-December 2021, seropositivity to N and S antibody was higher among repeat donors (61.3%) compared with new donors (55.1%; p = .043) and among donors from Kampala (capital city of Uganda) compared with rural regions (p = .007). Seropositivity to S antibody was significantly lower among HIV-seropositive individuals (58.8% vs. 84.9%; p = .009). CONCLUSIONS: Despite previously reported low numbers of COVID-19 cases and related deaths in Uganda, high SARS-CoV-2 seroprevalence and increasing antibody levels among blood donors indicated that the country experienced high levels of infection over the course of the pandemic.

Anti-PF4 antibodies associated with disease severity in COVID-19.

Severe COVID-19 is characterized by a prothrombotic state associated with thrombocytopenia, with microvascular thrombosis being almost invariably present in the lung and other organs at postmortem examination. We evaluated the presence of antibodies to platelet factor 4 (PF4)-polyanion complexes using a clinically validated immunoassay in 100 hospitalized patients with COVID-19 with moderate or severe disease (World Health Organization score, 4 to 10), 25 patients with acute COVID-19 visiting the emergency department, and 65 convalescent individuals. Anti-PF4 antibodies were detected in 95 of 100 hospitalized patients with COVID-19 (95.0%) irrespective of prior heparin treatment, with a mean optical density value of 0.871 ± 0.405 SD (range, 0.177 to 2.706). In contrast, patients hospitalized for severe acute respiratory disease unrelated to COVID-19 had markedly lower levels of the antibodies. In a high proportion of patients with COVID-19, levels of all three immunoglobulin (Ig) isotypes tested (IgG, IgM, and IgA) were simultaneously elevated. Antibody levels were higher in male than in female patients and higher in African Americans and Hispanics than in White patients. Anti-PF4 antibody levels were correlated with the maximum disease severity score and with significant reductions in circulating platelet counts during hospitalization. In individuals convalescent from COVID-19, the antibody levels returned to near-normal values. Sera from patients with COVID-19 induced higher levels of platelet activation than did sera from healthy blood donors, but the results were not correlated with the levels of anti-PF4 antibodies. These results demonstrate that the vast majority of patients with severe COVID-19 develop anti-PF4 antibodies, which may play a role in the clinical complications of COVID-19.

Absence of pathogenic viruses in COVID-19 convalescent plasma.

BACKGROUND: It is important to maintain the safety of blood products by avoiding the transfusion of units with known and novel viral pathogens. It is unknown whether COVID-19 convalescent plasma (CCP) may contain pathogenic viruses (either newly acquired or reactivated) that are not routinely screened for by blood centers. METHODS: The DNA virome was characterized in potential CCP donors (n = 30) using viral genome specific PCR primers to identify DNA plasma virome members of the Herpesviridae [Epstein Barr Virus (EBV), cytomegalovirus (CMV), human herpesvirus 6A/B, human herpesvirus 7] and Anelloviridae [Torque teno viruses (TTV), Torque teno mini viruses (TTMV), and Torque teno midi viruses (TTMDV)] families. In addition, the RNA plasma virome was characterized using unbiased metagenomic sequencing. Sequencing was done on a HiSeq2500 using high output mode with a read length of 2X100 bp. The sequencing reads were taxonomically classified using Kraken2. CMV and EBV seroprevalence were evaluated using a chemiluminescent immunoassay. RESULTS: TTV and TTMDV were detected in 12 (40%) and 4 (13%) of the 30 study participants, respectively; TTMDV was always associated with infection with TTV. We did not observe TTMV DNAemia. Despite CMV and EBV seroprevalences of 33.3% and 93.3%, respectively, we did not detect Herpesviridae DNA among the study participants. Metagenomic sequencing did not reveal any human RNA viruses in CCP, including no evidence of circulating SARS-CoV-2. DISCUSSION: There was no evidence of pathogenic viruses, whether newly acquired or reactivated, in CCP despite the presence of non-pathogenic Anelloviridae. These results confirm the growing safety data supporting CCP.

Infectious Diseases Institute at Makerere University College of Health Sciences: a case study of a sustainable capacity building model for health care, research and training.

The Infectious Diseases Institute (IDI), established in 2001, was the first autonomous institution of Makerere University set up as an example of what self-governing institutes can do in transforming the academic environment to become a rapidly progressive University addressing the needs of society This paper describes the success factors and lessons learned in development of sustainable centers of excellence to prepare academic institutions to respond appropriately to current and future challenges to global health. Key success factors included a) strong collaboration by local and international experts to combat the HIV pandemic, along with b) seed funding from Pfizer Inc., c) longstanding collaboration with Accordia Global Health Foundation to create and sustain institutional strengthening programs, d) development of a critical mass of multi-disciplinary research leaders and managers of the center, and e) a series of strong directors who built strong governance structures to execute the vision of the institute, with subsequent transition to local leadership. Conclusion: Twenty years of sustained investment in infrastructure, human capital, leadership, and collaborations present Makerere University and the sub-Saharan Africa region with an agile center of excellence with preparedness to meet the current and future challenges to global health.

Differential antibody production by symptomatology in SARS-CoV-2 convalescent individuals.

The association between COVID-19 symptoms and antibody responses against SARS-CoV-2 is poorly characterized. We analyzed antibody levels in individuals with known SARS-CoV-2 infection to identify potential antibody-symptom associations. Convalescent plasma from 216 SARS-CoV-2 RNA+ individuals with symptomatology information were tested for the presence of IgG to the spike S1 subunit (Euroimmun ELISA), IgG to receptor binding domain (RBD, CoronaCHEK rapid test), and for IgG, IgA, and IgM to nucleocapsid (N, Bio-Rad ELISA). Logistic regression was used to estimate the odds of having a COVID-19 symptom from the antibody response, adjusting for sex and age. Cough strongly associated with antibodies against S1 (adjusted odds ratio [aOR] = 5.33; 95% CI from 1.51 to 18.86) and RBD (aOR = 4.36; CI 1.49, 12.78). In contrast, sore throat significantly associated with the absence of antibodies to S1 and N (aOR = 0.25; CI 0.08, 0.80 and aOR = 0.31; 0.11, 0.91). Similarly, lack of symptoms associated with the absence of antibodies to N and RBD (aOR = 0.16; CI 0.03, 0.97 and aOR = 0.16; CI 0.03, 1.01). Cough appeared to be correlated with a seropositive result, suggesting that SARS-CoV-2 infected individuals exhibiting lower respiratory symptoms generate a robust antibody response. Conversely, those without symptoms or limited to a sore throat while infected with SARS-CoV-2 were likely to lack a detectable antibody response. These findings strongly support the notion that severity of infection correlates with robust antibody response.

Severe Acute Respiratory Syndrome Coronavirus 2 Antibody Seroprevalence in Decedents Undergoing Forensic Postmortem Examination: Feasibility for Real-Time Pandemic Surveillance.

Background: Population-based seroprevalence studies offer comprehensive characterization of coronavirus disease 2019 (COVID-19) spread, but barriers exist and marginalized populations may not be captured. We assessed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibody seroprevalence among decedents in Maryland over 6 months in 2020. Methods: Data were collected on decedents undergoing forensic postmortem examination in Maryland from 24 May through 30 November 2020 from whom a blood specimen could be collected. Those with available blood specimens were tested with the CoronaCHEK lateral flow antibody assay. We assessed monthly seroprevalence compared to the statewide estimated number of cases and proportion of positive test results (testing positivity). We used Poisson regression with robust variance to estimate adjusted prevalence ratios (aPRs) with 95% confidence intervals (CIs) for associations of demographic characteristics, homelessness, and manner of death with SARS-CoV-2 antibodies. Results: Among 1906 decedents, 305 (16%) were positive for SARS-CoV-2 antibodies. Monthly seroprevalence increased from 11% to 22% over time and was consistently higher than state-level estimates of testing positivity. Hispanic ethnicity was associated with 2- to 3.2-fold higher seropositivity (P < .05) irrespective of sex. Deaths due to motor vehicle crash were associated with 62% increased seropositivity (aPR, 1.62 [95% CI, 1.15-2.28]) vs natural manner of death. Though seroprevalence was lower in decedents of illicit drug overdose vs nonoverdose in early months, this shifted, and seroprevalence was comparable by November 2020. Conclusions: Decedents undergoing forensic postmortem examination, especially those dying due to motor vehicle trauma, may be a sentinel population for COVID-19 spread in the general population and merits exploration in other states/regions.

A Tale of 3 Pandemics: Severe Acute Respiratory Syndrome Coronavirus 2, Hepatitis C Virus, and Human Immunodeficiency Virus in an Urban Emergency Department in Baltimore, Maryland.

Background: We sought to determine the prevalence and sociodemographic and clinical correlates of acute and convalescent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), hepatitis C virus (HCV), and human immunodeficiency virus (HIV) infections among emergency department (ED) patients in Baltimore. Methods: Remnant blood samples from 7450 unique patients were collected over 4 months in 2020 for SARS-CoV-2 antibody (Ab), HCV Ab, and HIV-1/2 antigen and Ab. Among them, 5012 patients were tested by polymerase chain reaction for SARS-CoV-2 based on clinical suspicion. Sociodemographics, ED clinical presentations, and outcomes associated with coinfections were assessed. Results: Overall, 729 (9.8%) patients had SARS-CoV-2 (acute or convalescent), 934 (12.5%) HCV, 372 (5.0%) HIV infection, and 211 patients (2.8%) had evidence of any coinfection (HCV/HIV, 1.5%; SARS-CoV-2/HCV, 0.7%; SARS-CoV-2/HIV, 0.3%; SARS-CoV-2/HCV/HIV, 0.3%). The prevalence of SARS-CoV-2 (acute or convalescent) was significantly higher in those with HCV or HIV vs those without (13.6% vs 9.1%, P < .001). Key sociodemographic disparities (race, ethnicity, and poverty) and specific ED clinical characteristics were significantly correlated with having any coinfections vs no infection or individual monoinfection. Among those with HCV or HIV, aged 18-34 years, Black race, Hispanic ethnicity, and a cardiovascular-related chief complaint had a significantly higher odds of having SARS-CoV-2 (prevalence ratios: 2.02, 2.37, 5.81, and 2.07, respectively). Conclusions: The burden of SARS-CoV-2, HCV, and HIV co-pandemics and their associations with specific sociodemographic disparities, clinical presentations, and outcomes suggest that urban EDs should consider implementing integrated screening and linkage-to-care programs for these 3 infections.

Differentiation of Individuals Previously Infected with and Vaccinated for SARS-CoV-2 in an Inner-City Emergency Department.

Emergency departments (EDs) can serve as surveillance sites for infectious diseases. The objective of this study was to determine the burden of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and to monitor the prevalence of vaccination against coronavirus disease 2019 (COVID-19) among patients attending an urban ED in Baltimore City. Using 1,914 samples of known exposure status, we developed an algorithm to differentiate previously infected, vaccinated, and unexposed individuals using a combination of antibody assays. We applied this testing algorithm to 4,360 samples from ED patients obtained in the spring of 2020 and 2021. Using multinomial logistic regression, we determined factors associated with infection and vaccination. For the algorithm, sensitivity and specificity for identifying vaccinated individuals were 100% and 99%, respectively, and 84% and 100% for previously infected individuals. Among the ED subjects, seroprevalence to SARS-CoV-2 increased from 2% to 24% between April 2020 and March 2021. Vaccination prevalence rose to 11% by mid-March 2021. Marked differences in burden of disease and vaccination coverage were seen by sex, race, and ethnicity. Hispanic patients, though accounting for 7% of the study population, had the highest relative burden of disease (17% of total infections) but with similar vaccination rates. Women and white individuals were more likely to be vaccinated than men or Black individuals. Individuals previously infected with SARS-CoV-2 can often be differentiated from vaccinated individuals using a serologic testing algorithm. The utility of this algorithm can aid in monitoring SARS-CoV-2 exposure and vaccination uptake frequencies and can potentially reflect gender, race, and ethnic health disparities.

Antibody attributes that predict the neutralization and effector function of polyclonal responses to SARS-CoV-2.

BACKGROUND: While antibodies can provide significant protection from SARS-CoV-2 infection and disease sequelae, the specific attributes of the humoral response that contribute to immunity are incompletely defined. METHODS: We employ machine learning to relate characteristics of the polyclonal antibody response raised by natural infection to diverse antibody effector functions and neutralization potency with the goal of generating both accurate predictions of each activity based on antibody response profiles as well as insights into antibody mechanisms of action. RESULTS: To this end, antibody-mediated phagocytosis, cytotoxicity, complement deposition, and neutralization were accurately predicted from biophysical antibody profiles in both discovery and validation cohorts. These models identified SARS-CoV-2-specific IgM as a key predictor of neutralization activity whose mechanistic relevance was supported experimentally by depletion. CONCLUSIONS: Validated models of how different aspects of the humoral response relate to antiviral antibody activities suggest desirable attributes to recapitulate by vaccination or other antibody-based interventions.

Minimal Crossover between Mutations Associated with Omicron Variant of SARS-CoV-2 and CD8+ T-Cell Epitopes Identified in COVID-19 Convalescent Individuals.

There is a growing concern that ongoing evolution of SARS-CoV-2 could lead to variants of concern (VOC) that are capable of avoiding some or all of the multifaceted immune response generated by both prior infection or vaccination, with the recently described B.1.1.529 (Omicron) VOC being of particular interest. Peripheral blood mononuclear cell samples from PCR-confirmed, recovered COVID-19 convalescent individuals (n = 30) infected with SARS-CoV-2 in the United States collected in April and May 2020 who possessed at least one or more of six different HLA haplotypes were selected for examination of their anti-SARS-CoV-2 CD8+ T-cell responses using a multiplexed peptide-major histocompatibility complex tetramer staining approach. This analysis examined if the previously identified viral epitopes targeted by CD8+ T cells in these individuals (n = 52 distinct epitopes) are mutated in the newly described Omicron VOC (n = 50 mutations). Within this population, only one low-prevalence epitope from the Spike protein, restricted to two HLA alleles and found in 2/30 (7%) individuals, contained a single amino acid change associated with the Omicron VOC. These data suggest that virtually all individuals with existing anti-SARS-CoV-2 CD8+ T-cell responses should recognize the Omicron VOC and that SARS-CoV-2 has not evolved extensive T-cell escape mutations at this time. IMPORTANCE The newly identified Omicron variant of concern contains more mutations than any of the previous variants described to date. In addition, many of the mutations associated with the Omicron variant are found in areas that are likely bound by neutralizing antibodies, suggesting that the first line of immunological defense against COVID-19 is compromised. However, both natural infection and vaccination develop T-cell-based responses in addition to antibodies. This study examined if the parts of the virus, or epitopes, targeted by the CD8+ T-cell response in 30 individuals who recovered from COVID-19 in 2020 were mutated in the Omicron variant. Only one of 52 epitopes identified in this population contained an amino acid that was mutated in Omicron. These data suggest that the T-cell immune response in previously infected, and most likely vaccinated, individuals should still be effective against Omicron.

Severe Acute Respiratory Syndrome Coronavirus-2 seroprevalence in South-Central Uganda, during 2019-2021.

BACKGROUND: Globally, key subpopulations such as healthcare workers (HCW) may have a higher risk of contracting SARS-CoV-2. In Uganda, limited access to Personal Protective Equipment and lack of clarity on the extent/pattern of community spread may exacerbate this situation. The country established infection prevention/control measures such as lockdowns and proper hand hygiene. However, due to resource limitations and fatigue, compliance is low, posing continued onward transmission risk. This study aimed to describe extent of SARS-CoV-2 seroprevalence in selected populations within the Rakai region of Uganda. METHODS: From 30th November 2020 to 8th January 2021, we collected venous blood from 753 HCW at twenty-six health facilities in South-Central Uganda and from 227 population-cohort participants who reported specific COVID-19 like symptoms (fever, cough, loss of taste and appetite) in a prior phone-based survey conducted (between May and August 2020) during the first national lockdown. 636 plasma specimens collected from individuals considered high risk for SARS-CoV-2 infection, prior to the first confirmed COVID-19 case in Uganda were also retrieved. Specimens were tested for antibodies to SARS-CoV-2 using the CoronaChek™ rapid COVID-19 IgM/IgG lateral flow test assay. IgM only positive samples were confirmed using a chemiluminescent microparticle immunoassay (CMIA) (Architect AdviseDx SARS-CoV-2 IgM) which targets the spike protein. SARS-CoV-2 exposure was defined as either confirmed IgM, both IgM and IgG or sole IgG positivity. Overall seroprevalence in each participant group was estimated, adjusting for test performance. RESULTS: The seroprevalence of antibodies to SARS-CoV-2 in HCW was 26.7% [95%CI: 23.5, 29.8] with no difference by sex, age, or cadre. We observed no association between PPE use and seropositivity among exposed healthcare workers. Of the phone-based survey participants, 15.6% [95%CI: 10.9, 20.3] had antibodies to SARS-CoV-2, with no difference by HIV status, sex, age, or occupation. Among 636 plasma specimens collected prior to the first confirmed COVID-19 case, 2.3% [95%CI: 1.2, 3.5] were reactive. CONCLUSIONS: Findings suggest high seroprevalence of antibodies to SARS-CoV-2 among HCW and substantial exposure in persons presenting with specific COVID-19 like symptoms in the general population of South-Central Uganda. Based on current limitations in serological test confirmation, it remains unclear whether seroprevalence among plasma specimens collected prior to confirmation of the first COVID-19 case implies prior SARS-CoV-2 exposure in Uganda.

ABO blood group and SARS-CoV-2 antibody response in a convalescent donor population.

BACKGROUND AND OBJECTIVES: ABO blood group may affect risk of SARS-CoV-2 infection and/or severity of COVID-19. We sought to determine whether IgG, IgA and neutralizing antibody (nAb) to SARS-CoV-2 vary by ABO blood group. MATERIALS AND METHODS: Among eligible convalescent plasma donors, ABO blood group was determined via agglutination of reagent A1 and B cells, IgA and IgG were quantified using the Euroimmun anti-SARS-CoV-2 ELISA, and nAb titres were quantified using a microneutralization assay. Differences in titre distribution were examined by ABO blood group using non-parametric Kruskal-Wallis tests. Adjusted prevalence ratios (aPR) of high nAb titre (≥1:160) were estimated by blood group using multivariable modified Poisson regression models that adjusted for age, sex, hospitalization status and time since SARS-CoV-2 diagnosis. RESULTS: Of the 202 potential donors, 65 (32%) were blood group A, 39 (19%) were group B, 13 (6%) were group AB, and 85 (42%) were group O. Distribution of nAb titres significantly differed by ABO blood group, whereas there were no significant differences in anti-spike IgA or anti-spike IgG titres by ABO blood group. There were significantly more individuals with high nAb titre (≥1:160) among those with blood group B, compared with group O (aPR = 1·9 [95%CI = 1·1-3·3], P = 0·029). Fewer individuals had a high nAb titre among those with blood group A, compared with group B (aPR = 0·6 [95%CI = 0·4-1·0], P = 0·053). CONCLUSION: Eligible CCP donors with blood group B may have relatively higher neutralizing antibody titres. Additional studies evaluating ABO blood groups and antibody titres that incorporate COVID-19 severity are needed.

Differential Performance of CoronaCHEK SARS-CoV-2 Lateral Flow Antibody Assay by Geographic Origin of Samples.

We assessed the performance of the CoronaCHEK lateral flow assay on samples from Uganda and Baltimore to determine the impact of geographic origin on assay performance. Plasma samples from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) PCR-positive individuals (Uganda, 78 samples from 78 individuals, and Baltimore, 266 samples from 38 individuals) and from prepandemic individuals (Uganda, 1,077, and Baltimore, 532) were evaluated. Prevalence ratios (PR) were calculated to identify factors associated with a false-positive test. After the first positive PCR in Ugandan samples, the sensitivity was 45% (95% confidence interval [CI], 24,68) at 0 to 7 days, 79% (95% CI, 64 to 91) at 8 to 14 days, and 76% (95% CI, 50 to 93) at >15 days. In samples from Baltimore, sensitivity was 39% (95% CI, 30 to 49) at 0 to 7 days, 86% (95% CI, 79 to 92) at 8 to 14 days, and 100% (95% CI, 89 to 100) at 15 days after positive PCR. The specificity of 96.5% (95% CI, 97.5 to 95.2) in Ugandan samples was significantly lower than that in samples from Baltimore, 99.3% (95% CI, 98.1 to 99.8; P < 0.01). In Ugandan samples, individuals with a false-positive result were more likely to be male (PR, 2.04; 95% CI, 1.03,3.69) or individuals who had had a fever more than a month prior to sample acquisition (PR, 2.87; 95% CI, 1.12 to 7.35). Sensitivity of the CoronaCHEK was similar in samples from Uganda and Baltimore. The specificity was significantly lower in Ugandan samples than in Baltimore samples. False-positive results in Ugandan samples appear to correlate with a recent history of a febrile illness, potentially indicative of a cross-reactive immune response in individuals from East Africa.

Comparative performance of multiplex salivary and commercially available serologic assays to detect SARS-CoV-2 IgG and neutralization titers.

Oral fluid (hereafter saliva) offers a non-invasive sampling method for detection of SARS-CoV-2 antibodies. However, data comparing performance of salivary tests against commercially-available serologic and neutralizing antibody (nAb) assays are lacking. This study compared the performance of a laboratory-developed multiplex salivary SARS-CoV-2 IgG assay targeting antibodies to nucleocapsid (N), receptor binding domain (RBD) and spike (S) antigens to three commercially-available SARS-CoV-2 serologic enzyme immunoassays (EIAs) (Ortho Vitros, Euroimmun, and BioRad) and nAb. Paired saliva and plasma samples were collected from 101 eligible COVID-19 convalescent plasma (CCP) donors >14 days since PCR+ confirmed diagnosis. Concordance was evaluated using positive (PPA) and negative (NPA) percent agreement, and Cohen's kappa coefficient. The range between salivary and plasma EIAs for SARS-CoV-2-specific N was PPA: 54.4-92.1% and NPA: 69.2-91.7%, for RBD was PPA: 89.9-100% and NPA: 50.0-84.6%, and for S was PPA: 50.6-96.6% and NPA: 50.0-100%. Compared to a plasma nAb assay, the multiplex salivary assay PPA ranged from 62.3% (N) and 98.6% (RBD) and NPA ranged from 18.8% (RBD) to 96.9% (S). Combinations of N, RBD, and S and a summary algorithmic index of all three (N/RBD/S) in saliva produced ranges of PPA: 87.6-98.9% and NPA: 50-91.7% with the three EIAs and ranges of PPA: 88.4-98.6% and NPA: 21.9-34.4% with the nAb assay. A multiplex salivary SARS-CoV-2 IgG assay demonstrated variable, but comparable performance to three commercially-available plasma EIAs and a nAb assay, and may be a viable alternative to assist in monitoring population-based seroprevalence and vaccine antibody response.

Feasibility of Known RNA Polymerase Inhibitors as Anti-SARS-CoV-2 Drugs.

Coronaviruses (CoVs) are positive-stranded RNA viruses that infect humans and animals. Infection by CoVs such as HCoV-229E, -NL63, -OC43 and -HKU1 leads to the common cold, short lasting rhinitis, cough, sore throat and fever. However, CoVs such as Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV), Middle East Respiratory Syndrome Coronavirus (MERS-CoV), and the newest SARS-CoV-2 (the causative agent of COVID-19) lead to severe and deadly diseases with mortality rates ranging between ~1 to 35% depending on factors such as age and pre-existing conditions. Despite continuous global health threats to humans, there are no approved vaccines or drugs targeting human CoVs, and the recent outbreak of COVID-19 emphasizes an urgent need for therapeutic interventions. Using computational and bioinformatics tools, here we present the feasibility of reported broad-spectrum RNA polymerase inhibitors as anti- SARS-CoV-2 drugs targeting its main RNA polymerase, suggesting that investigational and approved nucleoside RNA polymerase inhibitors have potential as anti-SARS-CoV-2 drugs. However, we note that it is also possible for SARS-CoV-2 to evolve and acquire drug resistance mutations against these nucleoside inhibitors.

Evolutionary analysis of the Delta and Delta Plus variants of the SARS-CoV-2 viruses.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has been rapidly evolving in the form of new variants. At least eleven known variants have been reported. The objective of this study was to delineate the differences in the mutational profile of Delta and Delta Plus variants. High-quality sequences (n = 1756) of Delta (B.1.617.2) and Delta Plus (AY.1 or B.1.617.2.1) variants were used to determine the prevalence of mutations (≥20 %) in the entire SARS-CoV-2 genome, their co-existence, and change in prevalence over a period of time. Structural analysis was conducted to get insights into the impact of mutations on antibody binding. A Sankey diagram was generated using phylogenetic analysis coupled with sequence-acquisition dates to infer the migration of the Delta Plus variant and its presence in the United States. The Delta Plus variant had a significant number of high-prevalence mutations (≥20 %) than in the Delta variant. Signature mutations in Spike (G142D, A222V, and T95I) existed at a more significant percentage in the Delta Plus variant than the Delta variant. Three mutations in Spike (K417N, V70F, and W258L) were exclusively present in the Delta Plus variant. A new mutation was identified in ORF1a (A1146T), which was only present in the Delta Plus variant with ~58 % prevalence. Furthermore, five key mutations (T95I, A222V, G142D, R158G, and K417N) were significantly more prevalent in the Delta Plus than in the Delta variant. Structural analyses revealed that mutations alter the sidechain conformation to weaken the interactions with antibodies. Delta Plus, which first emerged in India, reached the United States through England and Japan, followed by its spread to more than 20 the United States. Based on the results presented here, it is clear that the Delta and Delta Plus variants have unique mutation profiles, and the Delta Plus variant is not just a simple addition of K417N to the Delta variant. Highly correlated mutations may have emerged to keep the structural integrity of the virus.

Challenges faced by families of critically ill patients during the first wave of the COVID-19 pandemic.

OBJECTIVE: To understand how surrogates of critically ill patients adjusted to challenges that resulted from the COVID-19 pandemic. METHODS: Participants (N = 62) were surrogates of critically ill adults with COVID-19 in the ICU at an urban, academic medical center from March to June 2020. Participants were recruited using convenience sampling and took part in one-time qualitative individual interviews via telephone. Qualitative data were analyzed using thematic content analysis. RESULTS: Qualitative analyses yielded four types of challenges: Communication with the medical team, communication among family members, understanding and tracking medical information, and distress related to visitor restrictions. To adjust to challenges related to communication, participants developed routines for receiving updates from the medical team and providing updates to other family members. To adjust to the challenge related to comprehension, participants sought information from external sources such as family members in healthcare fields. To adjust to the challenge related to visitation, participants found some comfort in video calls with the patient. CONCLUSIONS: Surrogates of critically ill patients with COVID-19 faced multiple types of challenges yet adjusted to those challenges. PRACTICAL IMPLICATIONS: Future research should focus on ways to support the wellbeing of surrogates during times of restricted hospital visitation. Clinical trial registered at ClinicalTrials.gov (NCT03969810).

CD8+ T-Cell Responses in COVID-19 Convalescent Individuals Target Conserved Epitopes From Multiple Prominent SARS-CoV-2 Circulating Variants.

This study examined whether CD8+ T-cell responses from coronavirus disease 2019 convalescent individuals (n = 30) potentially maintain recognition of the major severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants (alpha, beta, gamma; n = 45 mutations assessed). Only 1 mutation found in Beta variant-spike overlapped with a previously identified epitope (1/52), suggesting that virtually all anti-SARS-CoV-2 CD8+ T-cell responses should recognize these newly described variants.

Evolution, correlation, structural impact and dynamics of emerging SARS-CoV-2 variants.

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) infections remain unmanageable in some parts of the world. As with other RNA viruses, mutations in the SARS-CoV-2 gene have been continuously evolving. Recently, four variants have been identified, B.1.1.7, B.1.351, P.1 and CAL.20C. These variants appear to be more infectious and transmissible than the original Wuhan-Hu-1 virus. Using a combination of bioinformatics and structural analyses, we show that the new SARS-CoV-2 variants emerged in the background of an already known Spike protein mutation D614G together with another mutation P323L in the RNA polymerase of SARS-CoV-2. The phylogenetic analysis showed that the CAL.20C and B.1.351 shared one common ancestor, whereas the B.1.1.7 and P.1 shared a different ancestor. Structural comparisons did not show any significant difference between the wild-type and mutant ACE2/Spike complexes. Structural analysis indicated that the N501Y mutation may increase hydrophobic interactions at the ACE2/Spike interface. However, reported greater binding affinity of N501Y Spike with ACE2 does not seem to be entirely due to increased hydrophobic interactions, given that Spike mutation R417T in P.1 or K417N in B.1.351 results in the loss of a salt-bridge interaction between ACE2 and S-RBD. The calculated change in free energy did not provide a clear trend of S protein stability of mutations in the variants. As expected, we show that the CAL.20C generally migrated from the west coast to the east coast of the USA. Taken together, the analyses suggest that the evolution of variants and their infectivity is complex and may depend upon many factors.