Investigation of Blood Plasma Viral Nucleocapsid Antigen as a Marker of Active Severe Acute Respiratory Syndrome Coronavirus 2 Omicron Variant Infection.

Background: Nasopharyngeal qualitative reverse-transcription polymerase chain reaction (RT-PCR) is the gold standard for diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, but it is not practical or sufficient in every clinical scenario due to its inability to distinguish active from resolved infection. Alternative or adjunct testing may be needed to guide isolation precautions and treatment in patients admitted to the hospital. Methods: We performed a single-center, retrospective analysis of residual clinical specimens and medical record data to examine blood plasma nucleocapsid antigen as a candidate biomarker of active SARS-CoV-2. Adult patients admitted to the hospital or presenting to the emergency department with SARS-CoV-2 ribonucleic acid (RNA) detected by RT-PCR from a nasopharyngeal swab specimen were included. Both nasopharyngeal swab and a paired whole blood sample were required to be available for analysis. Results: Fifty-four patients were included. Eight patients had positive nasopharyngeal swab virus cultures, 7 of whom (87.5%) had concurrent antigenemia. Nineteen (79.2%) of 24 patients with detectable subgenomic RNA and 20 (80.0%) of 25 patients with N2 RT-PCR cycle threshold ≤ 33 had antigenemia. Conclusions: Most individuals with active SARS-CoV-2 infection are likely to have concurrent antigenemia, but there may be some individuals with active infection in whom antigenemia is not detectable. The potential for high sensitivity and convenience of a blood test prompts interest in further investigation as a screening tool to reduce reliance on nasopharyngeal swab sampling and as an adjunct diagnostic test to aid in clinical decision making during the period after acute coronavirus disease 2019.

Outbreak of severe acute respiratory coronavirus virus 2 (SARS-CoV-2) in hospitalized hemodialysis patients: An epidemiologic and genomic investigation.

We performed an epidemiological investigation and genome sequencing of severe acute respiratory coronavirus virus 2 (SARS-CoV-2) to define the source and scope of an outbreak in a cluster of hospitalized patients. Lack of appropriate respiratory hygiene led to SARS-CoV-2 transmission to patients and healthcare workers during a single hemodialysis session, highlighting the importance of infection prevention precautions.

Nucleocapsid Antigenemia in Patients Receiving Anti-CD20 Therapy With Protracted COVID-19.

Immunocompromised patients with prolonged coronavirus disease 2019 symptoms present diagnostic and therapeutic challenges. We measured viral nucleocapsid antigenemia in 3 patients treated with anti-CD20 immunotherapy who acquired severe acute respiratory syndrome coronavirus 2 infection and experienced protracted symptoms. Our results support nucleocapsid antigenemia as a marker of persistent infection and therapeutic response.

Unrecognized introductions of SARS-CoV-2 into the US state of Georgia shaped the early epidemic.

In early 2020, as diagnostic and surveillance responses for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ramped up, attention focused primarily on returning international travelers. Here, we build on existing studies characterizing early patterns of SARS-CoV-2 spread within the USA by analyzing detailed clinical, molecular, and viral genomic data from the state of Georgia through March 2020. We find evidence for multiple early introductions into Georgia, despite relatively sparse sampling. Most sampled sequences likely stemmed from a single or small number of introductions from Asia three weeks prior to the state's first detected infection. Our analysis of sequences from domestic travelers demonstrates widespread circulation of closely related viruses in multiple US states by the end of March 2020. Our findings indicate that the exclusive focus on identifying SARS-CoV-2 in returning international travelers early in the pandemic may have led to a failure to recognize locally circulating infections for several weeks and point toward a critical need for implementing rapid, broadly targeted surveillance efforts for future pandemics.

Association Between Caseload Surge and COVID-19 Survival in 558 U.S. Hospitals, March to August 2020.

BACKGROUND: Several U.S. hospitals had surges in COVID-19 caseload, but their effect on COVID-19 survival rates remains unclear, especially independent of temporal changes in survival. OBJECTIVE: To determine the association between hospitals' severity-weighted COVID-19 caseload and COVID-19 mortality risk and identify effect modifiers of this relationship. DESIGN: Retrospective cohort study. (ClinicalTrials.gov: NCT04688372). SETTING: 558 U.S. hospitals in the Premier Healthcare Database. PARTICIPANTS: Adult COVID-19-coded inpatients admitted from March to August 2020 with discharge dispositions by October 2020. MEASUREMENTS: Each hospital-month was stratified by percentile rank on a surge index (a severity-weighted measure of COVID-19 caseload relative to pre-COVID-19 bed capacity). The effect of surge index on risk-adjusted odds ratio (aOR) of in-hospital mortality or discharge to hospice was calculated using hierarchical modeling; interaction by surge attributes was assessed. RESULTS: Of 144 116 inpatients with COVID-19 at 558 U.S. hospitals, 78 144 (54.2%) were admitted to hospitals in the top surge index decile. Overall, 25 344 (17.6%) died; crude COVID-19 mortality decreased over time across all surge index strata. However, compared with nonsurging (<50th surge index percentile) hospital-months, aORs in the 50th to 75th, 75th to 90th, 90th to 95th, 95th to 99th, and greater than 99th percentiles were 1.11 (95% CI, 1.01 to 1.23), 1.24 (CI, 1.12 to 1.38), 1.42 (CI, 1.27 to 1.60), 1.59 (CI, 1.41 to 1.80), and 2.00 (CI, 1.69 to 2.38), respectively. The surge index was associated with mortality across ward, intensive care unit, and intubated patients. The surge-mortality relationship was stronger in June to August than in March to May (slope difference, 0.10 [CI, 0.033 to 0.16]) despite greater corticosteroid use and more judicious intubation during later and higher-surging months. Nearly 1 in 4 COVID-19 deaths (5868 [CI, 3584 to 8171]; 23.2%) was potentially attributable to hospitals strained by surging caseload. LIMITATION: Residual confounding. CONCLUSION: Despite improvements in COVID-19 survival between March and August 2020, surges in hospital COVID-19 caseload remained detrimental to survival and potentially eroded benefits gained from emerging treatments. Bolstering preventive measures and supporting surging hospitals will save many lives. PRIMARY FUNDING SOURCE: Intramural Research Program of the National Institutes of Health Clinical Center, the National Institute of Allergy and Infectious Diseases, and the National Cancer Institute.

The Distribution and Determinants of COVID-19 in Sudan, 2020/2021: Analysis of Surveillance Data (preprint)

Background: Since the first case of COVID-19 in Sudan was reported in March 2020, the Federal Ministry of Health adopted an active surveillance system to collect and analyze information from the isolation centers and public and private laboratories about all suspected and confirmed COVID-19 cases. This study used the surveillance data to better understand the distribution and determinants of COVID-19 in Sudan and to construct a threshold level beyond which the dramatic surge may occur. Methods: Data of suspected and confirmed COVID-19 cases were extracted from the line list prepared by the Surveillance and Information Department at the Federal Ministry of Health after obtaining ethical approval from the National Ethics Committee. Data were cleaned, coded, and analyzed using SPSS version 21. Frequencies and proportions were used to describe data. A univariate logistic regression analysis was used to determine the association of variables with the positivity of COVID-19. Variables with p-values < 0.05 in the univariate logistic analyses were included in multivariable logistic regression to determine the adjusted odds ratios (ORs) and their 95% confidence interval (CI). A two-sided α of less than 0.05 (p < 0.05) was considered statistically significant.Results: Out of 48,545 suspected cases, 27,453 were positive. Four waves were seen, with a distinct explosion point of around 200 cases observed nationwide. Khartoum reported the highest number of cases. Of those tested positive, 16,444 (59.9%) were male and 11,009 (40.1%) were female. The mean (SD) age of cases was 41.1 (19.0) years with 21.6% of cases above 60 years. 14,780 (53.8%) of cases were asymptomatic. Fever, cough, shortness of breath, and loss of smell and taste were reported in 32.7%, 26.4%, 19.1 and 4.5% of confirmed cases, respectively. A total of 1,793 confirmed cases died; the case fatality rate was 6.5%. A considerable proportion of infection was reported among health workers. A univariate logistic regression analysis revealed that being symptomatic is significantly associated with testing negative for COVID-19 (odds ratio < 1). Conclusions: COVID-19 was widely spread in Sudan with more cases in Khartoum, the capital of Sudan. The country experienced four waves with an observable epidemic explosion point of around 200 positive cases per week nationwide. Around half of the patients were asymptomatic; however, fever, cough, and shortness of breath were the commonest symptoms. The CFR all through was 6.5%, with death having a strong association with age. Further studies are recommended to clarify the image, especially among health workers. The study also highlighted the need to improve the quality of surveillance data.

Immunocompromised Patients with Prolonged Viral Shedding of SARS-COV-2

Background Most individuals diagnosed with mild to moderate COVID-19 are no longer infectious after day 10 of symptom onset and those with severe or critical illness from COVID are typically not infection after day 20 day of symptom onset. Recovered persons can continue to test positive for SARS-CoV-2 by PCR via detection of non-viable RNA in nasopharyngeal specimens for up to three months (or longer) after illness onset. It is also know known that severely immunocompromised patients may produce replication-competent virus greater than 20 days from symptom onset and may require, per CDC recommendations, “additional testing and consultation with infectious diseases specialists and infection control experts”. We aim to discuss four case studies of severely immunocompromised patients who exhibited signs of persistent COVID-19 infection of COVID and how we managed transmission-based precautions in our hospital through sequencing and evaluation of cycle thresholds (CT) values and subgenomic RNA detection. Methods Residual nasopharyngeal (NP) samples were collected on patients exhibiting persistent COVID like symptoms. These samples underwent N gene and N gene subgenomic RNA (sgRNA) real-time reverse transcription polymerase chain reaction (rRT-PCR) testing. Results Analysis of longitudinal SARS-CoV-2 sequence data demonstrated within-patient virus evolution, including mutations in the receptor binding domain and deletions in the N-terminal domain of the spike protein, which have been implicated in antibody escape. See Figures 1 and 2. Figure 1. Timelines of Identified Patients 1 and 2 Patient 1: 46-year-old woman with recently diagnosed stage IV diffuse large B-cell lymphoma for which she was treated with 2 cycles of R-CHOP. Patient 2: 38-year-old woman with history of myelodysplastic syndrome, peripheral blood stem cell transplant with chronic graft versus host disease of the GI tract, skin, and eyes as well as CMV enteritis, and she was maintained on rituximab, mycophenolate mofetil, prednisone, and monthly IVIG without recent changes to her immunosuppression. Figure 2. Timeline of Identified Patients 3 and 4 Patient 3: 44 year-old man with prior history of thymoma s/p thymectomy Patient 4: 46 year-old man who was initially diagnosed with marginal zone lymphoma approximately 2.5 years ago. He was initially treated with bendamustine and rituximab and achieved remission. He was then continued on maintenance rituximab without significant complications for a planned two years. Conclusion Differentiating between prolonged viral shedding of non-infectious RNA and persistent replicating viable virus can be difficult to determine without full evaluation of a patient’s clinical picture and timeline. Consultation between laboratory, infectious diseases, and infection prevention experts to provide appropriate level of guidance for precautions and treatment may be warranted. Testing by PCR and analysis of CT values may provide key findings of viral replication in immunocompromised hosts, indicating the need for evaluation of additional treatment and maintaining isolation status in healthcare settings. Disclosures All Authors: No reported disclosures

Single-Amplicon Multiplex Real-Time Reverse Transcription-PCR with Tiled Probes To Detect SARS-CoV-2 spike Mutations Associated with Variants of Concern.

To provide an accessible and inexpensive method to surveil for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mutations, we developed a multiplex real-time reverse transcription-PCR (rRT-PCR) assay, the Spike single-nucleotide polymorphism (SNP) assay, to detect specific mutations in the spike receptor binding domain. A single primer pair was designed to amplify a 348-bp region of spike, and probes were initially designed to detect K417, E484K, and N501Y. The assay was evaluated using characterized variant sample pools and residual nasopharyngeal samples. Variant calls were confirmed by SARS-CoV-2 genome sequencing in a subset of samples. Subsequently, a fourth probe was designed to detect L452R. The lower limit of 95% detection was 2.46 to 2.48 log10 genome equivalents (GE)/ml for the three initial targets (∼1 to 2 GE/reaction). Among 253 residual nasopharyngeal swabs with detectable SARS-CoV-2 RNA, the Spike SNP assay was positive in 238 (94.1%) samples. All 220 samples with threshold cycle (CT) values of <30 for the SARS-CoV-2 N2 target were detected, whereas 18/33 samples with N2 CT values of ≥30 were detected. Spike SNP results were confirmed by sequencing in 50/50 samples (100%). Addition of the 452R probe did not affect performance for the original targets. The Spike SNP assay accurately identifies SARS-CoV-2 mutations in the receptor binding domain, and it can be quickly modified to detect new mutations that emerge.

Bioaerosol Sampling for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) in a Referral Center with Critically Ill Coronavirus Disease 2019 (COVID-19) Patients March-May 2020.

BACKGROUND: Previous research has shown that rooms of patients with coronavirus disease 2019 (COVID-19) present the potential for healthcare-associated transmission through aerosols containing severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). However, data on the presence of these aerosols outside of patient rooms are limited. We investigated whether virus-containing aerosols were present in nursing stations and patient room hallways in a referral center with critically ill COVID-19 patients. METHODS: Eight National Institute for Occupational Safety and Health BC 251 2-stage cyclone samplers were set up throughout 6 units, including nursing stations and visitor corridors in intensive care units and general medical units, for 6 h each sampling period. Samplers were placed on tripods which held 2 samplers positioned 102 cm and 152 cm above the floor. Units were sampled for 3 days. Extracted samples underwent reverse transcription polymerase chain reaction for selected gene regions of the SARS-CoV-2 virus nucleocapsid and the housekeeping gene human RNase P as an internal control. RESULTS: The units sampled varied in the number of laboratory-confirmed COVID-19 patients present on the days of sampling. Some of the units included patient rooms under negative pressure, while most were maintained at a neutral pressure. Of 528 aerosol samples collected, none were positive for SARS-CoV-2 RNA by the estimated limit of detection of 8 viral copies/m3 of air. CONCLUSIONS: Aerosolized SARS-CoV-2 outside of patient rooms was undetectable. While healthcare personnel should avoid unmasked close contact with each other, these findings may provide reassurance for the use of alternatives to tight-fitting respirators in areas outside of patient rooms during the current pandemic.

The Effect of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Mitigation Strategies on Seasonal Respiratory Viruses: A Tale of 2 Large Metropolitan Centers in the United States.

To assess the impact of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic on seasonal respiratory viruses, absolute case counts and viral reproductive rates from 2019-2020 were compared against previous seasons. Our findings suggest that the public health measures implemented to reduce SARS-CoV-2 transmission significantly reduced the transmission of other respiratory viruses.

Correlation of SARS-CoV-2 Subgenomic RNA with Antigen Detection in Nasal Midturbinate Swab Specimens.

Among symptomatic outpatients, subgenomic RNA of severe acute respiratory syndrome coronavirus 2 in nasal midturbinate swab specimens was concordant with antigen detection but remained detectable in 13 (82.1%) of 16 nasopharyngeal swab specimens from antigen-negative persons. Subgenomic RNA in midturbinate swab specimens might be useful for routine diagnostics to identify active virus replication.

Validation of High-Sensitivity Severe Acute Respiratory Syndrome Coronavirus 2 Testing for Stool-Toward the New Normal for Fecal Microbiota Transplantation.

INTRODUCTION: Mounting evidence demonstrates potential for fecal-oral transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The US Food and Drug Administration now requires SARS-CoV-2 testing of potential feces donors before the use of stool manufactured for fecal microbiota transplantation. We sought to develop and validate a high-sensitivity SARS-CoV-2 reverse transcriptase polymerase chain reaction (RT-PCR) procedure for testing stool specimens. METHODS: A modified extraction method was used with an RT-PCR assay adapted from the Centers for Disease Control and Prevention PCR protocol for respiratory specimens. Contrived specimens were created using pre-COVID-19 banked stool specimens and spiking in known concentrations of SARS-CoV-2-specific nucleic acid. The highest transcript concentration at which 2/2 or 1/2 SARS-CoV-2 targets were detected in 9/10 replicates was defined as the dual-target limit and single-target limit of detection, respectively. The clinical performance of the assay was evaluated with stool samples collected from 17 nasopharyngeal swab RT-PCR-positive patients and 14 nasopharyngeal RT-PCR-negative patients. RESULTS: The dual-target and single-target limit of detection were 56 copies/µL and 3 copies/µL, respectively. SARS-CoV-2 was detected at concentrations as low as 0.6 copies/µL. Clinical stool samples from known COVID-19-positive patients demonstrated the detection of SARS-CoV-2 in stool up to 29 days from symptom onset with a high agreement with nasopharyngeal swab tests (kappa statistic of 0.95, P value < 0.001). DISCUSSION: The described RT-PCR test is a sensitive and flexible approach for the detection of SARS-CoV-2 in stool specimens. We propose an integrated screening approach that incorporates this stool test to support continuation of fecal microbiota transplantation programs.

The Importance and Challenges of Identifying SARS-CoV-2 Reinfections.

Reports of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) reinfection have raised important questions about the strength and durability of the immune response to primary infection, which are key factors in predicting the course of the pandemic. Identifying reinfection requires detecting the virus at two different time points and using viral genomic data to distinguish reinfection from persistent viral carriage. This process is hindered by challenges of logistics and capacity, such as banking samples from primary infection and performing viral genome sequencing. These challenges may help to explain why very few cases have been described to date. In addition, reinfection may be a rare phenomenon, but detailed prospective studies are needed to rigorously assess its frequency. To provide context for future investigations of SARS-CoV-2 reinfection, we review 16 cases that have been published to date or are available in preprint. Reinfection occurred across demographic spectra and in patients whose initial infections were both asymptomatic/mild and moderate/severe. For cases in which severity could be compared between episodes, half of reinfections were less severe, raising the possibility of partial immune protection. Although many patients had a positive total immunoglobulin or IgG result at the time of reinfection, very little examination of their immune response was performed. Further work is needed to elucidate the frequency, determinants, and consequences of SARS-CoV-2 reinfection. Establishing the necessary frameworks for surveillance and investigation will rely heavily on clinical laboratories and clinical investigators, and we propose several considerations to guide the medical community in identifying and characterizing SARS-CoV-2 reinfections.

Infection- and vaccine-induced antibody binding and neutralization of the B.1.351 SARS-CoV-2 variant.

The emergence of SARS-CoV-2 variants with mutations in the spike protein is raising concerns about the efficacy of infection- or vaccine-induced antibodies. We compared antibody binding and live virus neutralization of sera from naturally infected and Moderna-vaccinated individuals against two SARS-CoV-2 variants: B.1 containing the spike mutation D614G and the emerging B.1.351 variant containing additional spike mutations and deletions. Sera from acutely infected and convalescent COVID-19 patients exhibited a 3-fold reduction in binding antibody titers to the B.1.351 variant receptor-binding domain of the spike protein and a 3.5-fold reduction in neutralizing antibody titers against SARS-CoV-2 B.1.351 variant compared to the B.1 variant. Similar results were seen with sera from Moderna-vaccinated individuals. Despite reduced antibody titers against the B.1.351 variant, sera from infected and vaccinated individuals containing polyclonal antibodies to the spike protein could still neutralize SARS-CoV-2 B.1.351, suggesting that protective humoral immunity may be retained against this variant.

Pharmacological Therapy for COVID-19: Where Are We Now and Where Are We Going?

Abstract: Background: Up to the point of writing this review, there is no scientific evidence of any effective medical therapy for coronavirus disease 2019  (COVID-19). In this review, we attempted to discuss the current summary of evidence of some medication, currently in trial for the treatment of COVID-19. Material and Methods: We have done an electronic literature search using the following database: PubMed, Medline, Scopus and Google scholar. These databases were searched using the keywords COVID-19 and pharmacological therapy. Results: At present, there are no well randomized controlled studies which can give evidence for most of the therapy used for COVID-19. Several  medications are in trials for COVID-19, among them: 1/ chloroquine and hydroxychloroquine;2/anti-virals oseltamivir, remdesivir, lopinavir/ritonavir and other protease inhibitors;3/antibiotics macrolide (Azithromycin);4/cytokine therapy interferon;5/ humanized monoclonal antibody tocilizumab;6/adjunct therapies vitamins C, D, and herbal medicine;7/ COVID-19 convalescent plasma;8/systemic steroids;9/expected COVID-19 vaccine. We have also included some of the herbal medicines that are commonly and widely used in the Middle East, Asia as well in Sudan, (black seeds, honey and Acacia Nilotica). It is worth mentioning that these herbal medicines have shown benefits in treating other diseases, but the evidence of their benefit in COVID-19 still needs to be established. Conclusion: Currently there is no pharmacological therapy for the COVID-19. More research and randomized clinical trials are needed to find  effective therapy or vaccine against COVID-19