Nucleocapsid Antigen Research Articles

First-in-Human Phase I Trial to Assess the Safety and Immunogenicity of an Orf Virus-Based COVID-19 Vaccine Booster

The emergence of SARS-CoV-2 has necessitated the development of versatile vaccines capable of addressing evolving variants. Prime-2-CoV_Beta, a novel Orf virus-based COVID-19 vaccine, was developed to express the SARS-CoV-2 spike and nucleocapsid antigens. This first-in-human, phase I, dose-finding clinical trial was conducted to assess the safety, reactogenicity, and immunogenicity of Prime-2-CoV_Beta as a booster in healthy adults. From June 2022 to June 2023, 60 participants in Germany received varying doses of Prime-2-CoV_Beta. The study demonstrated a favorable safety profile, with no serious adverse events (AEs) reported. All AEs were mild (107) or moderate (10), with the most common symptoms being pain at the injection site, fatigue, and headache. Immunogenicity assessments revealed robust vaccine-induced antigen-specific immune responses. High doses notably elicited significant increases in antibodies against the spike and nucleocapsid proteins as well as neutralizing antibodies against SARS-CoV-2 and its variants. Additionally, the vaccine did not induce ORFV-neutralizing antibodies, indicating the potential for repeated administration. In conclusion, Prime-2-CoV_Beta was safe, well tolerated, and immunogenic, demonstrating potential as a broadly protective vaccine against SARS-CoV-2 and its variants. These promising results support further evaluation of higher doses and additional studies to confirm efficacy and long-term protection. This trial was registered at ClinicalTrials, NCT05389319.

A multiantigenic Orf virus-based vaccine efficiently protects hamsters and nonhuman primates against SARS-CoV-2

Among the common strategies to design next-generation COVID-19 vaccines is broadening the antigenic repertoire thereby aiming to increase efficacy against emerging variants of concern (VoC). This study describes a new Orf virus-based vector (ORFV) platform to design a multiantigenic vaccine targeting SARS-CoV-2 spike and nucleocapsid antigens. Vaccine candidates were engineered, either expressing spike protein (ORFV-S) alone or co-expressing nucleocapsid protein (ORFV-S/N). Mono- and multiantigenic vaccines elicited comparable levels of spike-specific antibodies and virus neutralization in mice. Results from a SARS-CoV-2 challenge model in hamsters suggest cross-protective properties of the multiantigenic vaccine against VoC, indicating improved viral clearance with ORFV-S/N, as compared to equal doses of ORFV-S. In a nonhuman primate challenge model, vaccination with the ORFV-S/N vaccine resulted in long-term protection against SARS-CoV-2 infection. These results demonstrate the potential of the ORFV platform for prophylactic vaccination and represent a preclinical development program supporting first-in-man studies with the multiantigenic ORFV vaccine.

Immunomodulatory effect of IFN-γ licensed adipose-mesenchymal stromal cells in an in vitro model of inflammation generated by SARS-CoV-2 antigens

In recent years, clinical studies have shown positive results of the application of Mesenchymal Stromal Cells (MSCs) in severe cases of COVID-19. However, the mechanisms of immunomodulation of IFN-γ licensed MSCs in SARS-CoV-2 infection are only partially understood. In this study, we first tested the effect of IFN-γ licensing in the MSC immunomodulatory profile. Then, we established an in vitro model of inflammation by exposing Calu-3 lung cells to SARS-CoV-2 nucleocapsid and spike (NS) antigens, and determined the toxicity of SARS-CoV-2 NS antigen and/or IFN-γ stimulation to Calu-3. The conditioned medium (iCM) generated by Calu-3 cells exposed to IFN-γ and SARS-CoV-2 NS antigens was used to stimulate T-cells, which were then co-cultured with IFN-γ-licensed MSCs. The exposure to IFN-γ and SARS-CoV-2 NS antigens compromised the viability of Calu-3 cells and induced the expression of the inflammatory mediators ICAM-1, CXCL-10, and IFN-β by these cells. Importantly, despite initially stimulating T-cell activation, IFN-γ-licensed MSCs dramatically reduced IL-6 and IL-10 levels secreted by T-cells exposed to NS antigens and iCM. Moreover, IFN-γ-licensed MSCs were able to significantly inhibit T-cell apoptosis induced by SARS-CoV-2 NS antigens. Taken together, our data show that, in addition to reducing the level of critical cytokines in COVID-19, IFN-γ-licensed MSCs protect T-cells from SARS-CoV-2 antigen-induced apoptosis. Such observations suggest that MSCs may contribute to COVID-19 management by preventing the lymphopenia and immunodeficiency observed in critical cases of the disease.

A novel orf virus vector-based COVID-19 booster vaccine shows cross-neutralizing activity in the absence of anti-vector neutralizing immunity

ABSTRACT Next-generation COVID-19 vaccines are being developed to expand the breadth of coverage against existing and future variants and to extend the duration of protection. Prime-2-CoV_Beta is an orf virus (ORFV) based multi-antigen COVID-19 vaccine that co-expresses Spike (S) and Nucleocapsid (N) antigens. The safety and immunogenicity of Prime-2-CoV_Beta is investigated in a phase 1 first-in-human (FIH) dose-finding trial (ORFEUS study, ClinicalTrials.gov: NCT05367843). Participants of two age groups (18–55 and 65–85 years) who previously completed at least two doses of mRNA vaccines were enrolled and sequentially assigned to different dose groups to receive one intramuscular dose of 3 × 105, 3 × 106, 1.5 × 107, or 3 × 107 plaque-forming units (PFU) of Prime-2-CoV_Beta on day 1 and a second dose on day 29. Here, we report safety and immunogenicity data collected up to 6 months after the first study vaccination. Prime-2-CoV_Beta is safe and well tolerated and elicits immune responses at higher dose levels in participants aged 18–55. A single dose of 3 × 107 PFU boosted binding and cross-neutralizing antibody responses that are maintained through 6 months after the first booster vaccination. Polyfunctional S-specific CD4+ and CD8+ T cell responses are observed after vaccination. No pre-existing or vaccine-induced neutralizing anti-vector antibodies are detected. Our findings highlight the potential of the ORFV vector as a safe platform for future vaccine design, which provides the ability to deliver multiple antigens and allows for repeat immunization.

Symptomatology and IgG Levels before and after SARS-CoV-2 Omicron Breakthrough Infections in Vaccinated Individuals.

(1) Background: After the COVID-19 pandemic, there is concern regarding the immunity of the population to SARS-CoV-2 variants, particularly the Omicron variant and its sub-lineages. (2) Methods: The study involved analyzing the immune response and symptomatology of 27 vaccinated individuals who were subsequently infected by Omicron sub-lineages. Blood samples were collected for serological analysis, including the detection of IgG antibodies reactive to the Nucleocapsid (N) and Spike (S) antigens of SARS-CoV-2. Additionally, participants were interviewed to assess the intensity of symptoms during the infection. (3) Results: Despite the high levels of anti-Spike IgG observed after vaccination, all participants were infected by Omicron sub-lineages. The most common symptoms reported by participants were fever or chills, sore throat, and cough. The levels of anti-Spike IgG found prior to infection did not correlate with symptom intensity post-infection. However, it was observed that high post-infection anti-Nucleocapsid IgG levels correlated with mild symptoms during the course of the disease, suggesting a potential role for anti-N antibodies in symptom intensity. (4) Conclusions: In line with previous studies, the high levels of IgG anti-Spike resulting from vaccination did not provide complete protection against infection by the Omicron variant. Additionally, our data suggest that anti-Nucleocapsid IgG titers are negatively correlated with the intensity of the symptoms during mild infections.

Comparison of Nucleocapsid Antigen, ORIF 1ab and RdRp (RNA-Dependent RNA Polymerase) in the Diagnosis of COVID-19

Comparison of Nucleocapsid Antigen, ORIF 1ab and RdRp (RNA-Dependent RNA Polymerase) in the Diagnosis of COVID-19

Diagnostic Utility of SARS-CoV-2 Nucleocapsid Antigenemia: A Meta-analysis.

Studies of the diagnostic performance of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid antigen in blood (antigenemia) have reached variable conclusions. The potential utility of antigenemia measurements as a clinical diagnostic test needs clarification. We performed a systematic review of Pubmed, Embase, and Scopus through July 15, 2023, and requested source data from corresponding authors. Summary sensitivity from 16 studies (4543 cases) sampled at ≤14 days of symptoms was 0.83 (0.75-0.89), and specificity was 0.98 (0.87-1.00) from 6 studies (792 reverse transcription polymerase chain reaction-negative controls). Summary sensitivity and specificity for paired respiratory specimens with cycle threshold values ≤33 were 0.91 (0.85-0.95) and 0.56 (0.39-0.73) from 10 studies (612 individuals). Source data from 1779 cases reveal that >70% have antigenemia 2 weeks following symptom onset, which persists in <10% at 28 days. The available studies suffer from heterogeneity, and Omicron-era data are scarce. Nucleocapsid antigenemia currently has limited utility due to limitations of existing studies and lack of Omicron-era data. Improved study designs targeting potential clinical uses in screening, surveillance, and complex clinical decision-making-especially in immunocompromised patients-are needed.

Impact of baseline SARS-CoV-2 load in plasma and upper airways on the incidence of acute extrapulmonary complications of COVID-19: a multicentric, prospective, cohort study.

Extrapulmonary complications (EPCs) are common in patients hospitalized for COVID-19, but data on their clinical consequences and association with viral replication and systemic viral dissemination is lacking. Patients hospitalized for COVID-19 and enrolled in the TICO (Therapeutics for Inpatients with COVID-19) platform trial at 114 international sites between August 2020 and November 2021 were included in a prospective cohort study. We categorized EPCs into 39 event types within 9 categories and estimated their frequency through day 28 and their association with clinical outcomes through day 90. We analyzed the association between baseline viral burden (plasma nucleocapsid antigen [N-Ag] and upper airway viral load [VL]) and EPCs, adjusting for other baseline factors. 2,625 trial participants were included in the study. The median age was 57 years (IQR 46-68), 57.7% were male, and 537 (20.5%) had at least one EPC. EPCs were associated with higher day-90 all-cause mortality (HR 9.6, 95% CI 7.3, 12.7) after adjustment for other risk factors. The risk of EPCs increased with increasing baseline plasma N-Ag (HR 1.21 per log10 ng/L increase, 95% CI 1.09, 1.34), and upper airway VL (HR 1.12 per log10 copies/mL increase, 95% CI 1.04, 1.19), after adjusting for comorbidities, disease severity, inflammatory markers, and other baseline factors. Trial treatment allocation had no effect on EPC risk. Systemic viral dissemination as evidenced by high plasma N-Ag and high respiratory viral burden are associated with development of EPCs in COVID-19, which in turn are associated with higher 90-day mortality.

SARS-CoV-2 viral load is linked to remdesivir efficacy in severe Covid-19 admitted to intensive care

Remdesivir therapy has been declared as efficient in the early stages of Covid-19. Of the 339 patients (males 55.8%, age 71(59;77) years) with a detectable viral load, 140 were treated with remdesivir (of those 103 in the ICU and 57 immunosuppressed) and retrospectively compared with 199 patients (of those 82 in the ICU and 28 immunosuppressed) who were denied therapy due to advanced Covid-19. The viral load was estimated by detecting nucleocapsid antigen in serum (n = 155, median 217(28;1524)pg/ml), antigen in sputum (n = 18, COI 18(4.6;32)), nasopharyngeal antigen (n = 44, COI 17(8;35)) and the real-time PCR (n = 122, Ct 21(18;27)). After adjustment for confounders, patients on remdesivir had better 12-month survival (HR 0.66 (0.44;0.98), p = 0.039), particularly when admitted to the ICU (HR 0.49 (0.29;0.81), p = 0.006). For the immunocompromised patients, the difference did not reach statistical significance (HR 0.55 (0.18;1.69), p = 0.3). The other most significant confounders were age, ICU admission, mechanical ventilation, leukocyte/lymphocyte ratio, admission creatinine and immunosuppression. The impact of monoclonal antibodies or previous vaccinations was not significant. Despite frequent immune suppression including haemato-oncology diseases, lymphopenia, and higher inflammatory markers in the remdesivir group, the results support remdesivir administration with respect to widely available estimates of viral load in patients with high illness severity.

Nanobody-based strategy for rapid and accurate pathogen detection: A case of COVID-19 testing

Antibody pairs-based immunoassay platforms served as essential and effective tools in the field of pathogen detection. However, the cumbersome preparation and limited detection sensitivity of antibody pairs challenge in establishment of a highly sensitive detection platform. In this study, using COVID-19 testing as a case, we utilized readily accessible nanobodies as detection antibodies and further proposed an accurate design concept with a more scientific and efficient screening strategy to obtain ultrasensitive antibody pairs. We employed nanobodies capable of binding different antigenic epitopes of the nucleocapsid (NP) or receptor-binding domain (RBD) antigens sandwich as substitutes for monoclonal antibodies (mAbs) sandwich in fast detection formats and utilized time-resolved fluorescence (TRF) microspheres as the signal probe. Consequently, we developed a multi-epitope nanobody sandwich-based fluorescence lateral flow immunoassay (FLFA) strip. Our results suggest that the NP antigen had a detection limit of 12.01pg/mL, while the RBD antigen had a limit of 6.51 pg/mL using our FLFA strip. Based on double mAb sandwiches, the values presented herein demonstrated 4 to 32-fold enhancements in sensitivity, and 32 to 256-fold enhancements compared to commercially available antigen lateral flow assay kits. Furthermore, we demonstrated the excellent characteristics of the proposed test strip, including its specificity, stability, accuracy, and repeatability, which underscores its the prospective utility. Indeed, these findings indicate that our established screening strategy along with the multi-epitope nanobody sandwich mode provides an optimized strategy in the field of pathogen detection.

SARS-CoV-2 Plasma Antibody and Nucleocapsid Antigen Status Predict Outcomes in Outpatients With COVID-19.

Reliable biomarkers of coronavirus disease 2019 (COVID-19) outcomes are critically needed. We evaluated associations of spike antibody (Ab) and plasma nucleocapsid antigen (N Ag) with clinical outcomes in nonhospitalized persons with mild-to-moderate COVID-19. Participants were nonhospitalized adults with mild-to-moderate COVID-19 enrolled in ACTIV-2 between January and July 2021 and randomized to placebo. We used quantitative assays for severe acute respiratory syndrome coronavirus 2 spike Ab and N Ag in blood and determined numbers of hospitalization/death events within 28 days and time to symptom improvement. Of 209 participants, 77 (37%) had quantifiable spike Ab and 139 (67%) quantifiable N Ag. Median age was 50 years; 111 (53%) were female, 182 (87%) White, and 105 (50%) Hispanic/Latino. Higher risk of hospitalization/death was seen with unquantifiable (22/132 [16.7%]) versus quantifiable (1/77 [1.3%]) spike Ab (risk ratio [RR], 12.83 [95% confidence interval {CI}, 1.76-93.34]) and quantifiable (22/139 [15.8%]) vs unquantifiable (1/70 [1.4%]) N Ag (RR, 11.08 [95% CI, 1.52-80.51]). Increasing risk of hospitalizations/deaths was seen with increasing N Ag levels. Time to symptom improvement was longer with unquantifiable versus quantifiable spike Ab (median, 14 [interquartile range {IQR}, 8 to >27] vs 8 [IQR, 4-22] days; adjusted hazard ratio [aHR], 0.66 [95% CI, .45-.96]) and with quantifiable versus unquantifiable N Ag (median, 12 [7 to >27] vs 10 [5-22] days; aHR, 0.79 [95% CI, .52-1.21]). Absence of spike Ab and presence of plasma N Ag predicted hospitalization/death and delayed symptom improvement in COVID-19 outpatients. NCT04518410.

Potential therapeutic biomolecules of hymenopteran venom against SARS-CoV-2 from Egyptian patients

The therapeutic potential of insect-derived bioactive molecules as anti-SARS-CoV-2 agents has shown promising results. Hymenopteran venoms, notably from Apis mellifera (honeybee) and Vespa orientalis (oriental wasp), were examined for the first time in an in vitro setting for their potential anti-COVID-19 activity. This assessment utilized an immunodiagnostic system to detect the SARS-CoV-2 nucleocapsid antigen titer reduction. Further analyses, including cytotoxicity assays, plaque reduction assays, and in silico docking-based screening, were performed to evaluate the efficacy of the most potent venom. Results indicated that bee and wasp venoms contain bioactive molecules with potential therapeutic effects against SARS-CoV-2.Nevertheless, the wasp venom exhibited superior efficacy compared to bee venom, achieving a 90% maximal (EC90) concentration effect of antigen depletion at 0.184 mg/mL, in contrast to 2.23 mg/mL for bee venom. The cytotoxicity of the wasp venom was assessed on Vero E6 cells 48 h post-treatment using the MTT assay. The CC 50 of the cell growth was 0.16617 mg/mL for Vero E6 cells. The plaque reduction assay of wasp venom revealed 50% inhibition (IC50) at a 0.208 mg/mL concentration. The viral count at 50% inhibition was 2.5 × 104 PFU/mL compared to the initial viral count of 5 × 104 PFU/mL. In silico data for the wasp venom revealed a strong attraction to binding sites on the ACE2 protein, indicating ideal interactions. This substantiates the potential of wasp venom as a promising viral inhibitor against SARS-CoV-2, suggesting its consideration as a prospective natural preventive and curative antiviral drug. In conclusion, hymenopteran venoms, particularly wasp venom, hold promise as a source of potential therapeutic biomolecules against SARS-CoV-2. More research and clinical trials are needed to evaluate these results and investigate their potential for translation into innovative antiviral therapies.

Heterotypic immunity from prior SARS-CoV-2 infection but not COVID-19 vaccination associates with lower endemic coronavirus incidence.

Immune responses from prior severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and COVID-19 vaccination mitigate disease severity, but they do not fully prevent subsequent infections, especially from genetically divergent strains. We examined the incidence of and immune differences against human endemic coronaviruses (eCoVs) as a proxy for response against future genetically heterologous coronaviruses (CoVs). We assessed differences in symptomatic eCoV and non-CoV respiratory disease incidence among those with known prior SARS-CoV-2 infection or previous COVID-19 vaccination but no documented SARS-CoV-2 infection or neither exposure. Retrospective cohort analyses suggest that prior SARS-CoV-2 infection, but not previous COVID-19 vaccination alone, associates with a lower incidence of subsequent symptomatic eCoV infection. There was no difference in non-CoV incidence, implying that the observed difference was eCoV specific. In a second cohort where both cellular and humoral immunity were measured, those with prior SARS-CoV-2 spike protein exposure had lower eCoV-directed neutralizing antibodies, suggesting that neutralization is not responsible for the observed decreased eCoV disease. The three groups had similar cellular responses against the eCoV spike protein and nucleocapsid antigens. However, CD8+ T cell responses to the nonstructural eCoV proteins nsp12 and nsp13 were higher in individuals with previous SARS-CoV-2 infection as compared with the other groups. This association between prior SARS-CoV-2 infection and decreased incidence of eCoV disease may therefore be due to a boost in CD8+ T cell responses against eCoV nsp12 and nsp13, suggesting that incorporation of nonstructural viral antigens in a future pan-CoV vaccine may improve vaccine efficacy.

An Evaluation of the Sensitivity and Specificity of Three COVID-19 Rapid Immunochromatographic Test Kits Compared to Real-Time Reverse Transcriptase-Polymerase Chain Reaction (rRT-PCR) Among Clinical Samples.

Background and objective The coronavirus disease 2019 (COVID-19) pandemic has imposed a significant burden on healthcare systems worldwide. This highlights the need for simple, rapid, and affordable diagnostic tests that can serve as alternatives tothe existing costly and demanding polymerase chain reaction (PCR) assay, especially in resource-limited countries like Ghana. In light of this, we aimed to assess thediagnostic efficacy of threeCOVID-19 rapid immunochromatographic antigen test kits vs. real-time reverse transcriptase-PCR (rRT-PCR). Methods This study evaluated the sensitivity and specificity of threeCOVID-19 rapid immunochromatographic antigen test kits: DG Rapid, SD Rapid, and SS Rapid. They were compared with the gold standard RT-PCRfor the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid antigen in 75 randomly selected archived nasopharyngeal samples. Results Of the 75 samples tested, 38 (50.7%) were positive and 37 (49.3%) were negative for SARS-CoV-2 RNA by rRT-PCR assay. No false positives were recorded. On the other hand, the DG Rapid kit detected 30 (78.9%) true positives and eight (21.1%) false negatives. SD Rapid kit detected 28 (73.7%) true positives and 10 (26.3%) false negatives, while the SS Rapid kit detected 19 (50.0%) true positives and 19 (50.0%) false negatives. While the specificity of each test kit was 100% (95% CI),the sensitivity of the DG Rapid, SD Rapid, and SS Rapid kits was 79%, 74%, and 50% (95% CI), respectively. Higher sensitivitieswere recorded among samples with cycle threshold (Ct) values <29.99 for each kit. Also, the DG Rapid kit demonstrated 79% excellent agreement with rRT-PCR, while the SD Rapid and SS Rapid kits demonstrated good agreement with rRT-PCR with 73% and 50% Cohen's kappa values, respectively. Conclusions Based on our findings,DG Rapid and SD Rapid kits are reliable alternatives to rRT-PCR for the detection of SARS-CoV-2 infection, especially in resource-limited settings like Ghana.

Potential Usefulness of IgA for the Early Detection of SARS-CoV-2 Infection: Comparison With IgM.

Serological testing can be a powerful complementary approach to achieve timely diagnosis of severe acute respiratory coronavirus 2 (SARS-CoV-2) infection, along with nucleic acid detection. Immunoglobulin (Ig) A antibodies are less frequently utilized to detect SARS-CoV-2 infection than IgM and IgG antibodies, even though IgA antibodies play an important role in protective immunity against SARS-CoV-2. This review discusses the differences in kinetics and assay performance between IgA and IgM antibodies and the factors influencing antibody responses. It highlights the potential usefulness of analyzing IgA antibodies for the early detection of SARS-CoV-2 infection. The early appearance of IgA and the high sensitivity of IgA-based immunoassays can aid in diagnosing coronavirus disease 2019. However, because of cross-reactivity, it is important to recognize the only moderate specificity of the early detection of SARS-CoV-2 IgA antibodies against spike antigens. Either the analysis of antibodies targeting the nucleocapsid antigen or a combination of antibodies against the nucleocapsid and spike antigens may strengthen the accuracy of serological evaluation.

Early trajectories of virological and immunological biomarkers and clinical outcomes in patients admitted to hospital for COVID-19: an international, prospective cohort study

Serial measurement of virological and immunological biomarkers in patients admitted to hospital with COVID-19 can give valuable insight into the pathogenic roles of viral replication and immune dysregulation. We aimed to characterise biomarker trajectories and their associations with clinical outcomes. In this international, prospective cohort study, patients admitted to hospital with COVID-19 and enrolled in the Therapeutics for Inpatients with COVID-19 platform trial within the Accelerating COVID-19 Therapeutic Interventions and Vaccines programme between Aug 5, 2020 and Sept 30, 2021 were included. Participants were included from 108sites in Denmark, Greece, Poland, Singapore, Spain, Switzerland, Uganda, the UK, and the USA, and randomised to placebo or one of four neutralising monoclonal antibodies: bamlanivimab (Aug 5 to Oct 13, 2020), sotrovimab (Dec16, 2020, to March 1, 2021), amubarvimab-romlusevimab (Dec 16, 2020, to March 1, 2021), and tixagevimab-cilgavimab (Feb 10 to Sept 30, 2021). This trial included an analysis of 2149 participants with plasma nucleocapsid antigen, anti-nucleocapsid antibody, C-reactive protein (CRP), IL-6, and D-dimer measured at baseline and day 1, day3, and day 5 of enrolment. Day-90 follow-up status was available for 1790 participants. Biomarker trajectories were evaluated for associations with baseline characteristics, a 7-day pulmonary ordinal outcome, 90-day mortality, and 90-day rate of sustained recovery. The study included 2149 participants. Participant median age was 57 years (IQR 46-68), 1246 (58·0%) of 2149 participants were male and 903 (42·0%) were female; 1792 (83·4%) had at least one comorbidity, and 1764 (82·1%) were unvaccinated. Mortality to day 90 was 172 (8·0%) of 2149 and 189 (8·8%) participants had sustained recovery. Apattern of less favourable trajectories of low anti-nucleocapsid antibody, high plasma nucleocapsid antigen, and high inflammatory markers over the first 5 days was observed for high-risk baseline clinical characteristics or factors related to SARS-CoV-2 infection. For example, participants with chronic kidney disease demonstrated plasma nucleocapsid antigen 424% higher (95% CI 319-559), CRP 174% higher (150-202), IL-6 173% higher (144-208), D-dimer 149% higher (134-165), and anti-nucleocapsid antibody 39% lower (60-18) to day 5 than those without chronic kidney disease. Participants in the highest quartile for plasma nucleocapsid antigen, CRP, and IL-6 at baseline and day 5 had worse clinical outcomes, including 90-day all-cause mortality (plasma nucleocapsid antigen hazard ratio (HR) 4·50 (95% CI 3·29-6·15), CRP HR 3·37 (2·30-4·94), and IL-6 HR 5·67 (4·12-7·80). This risk persisted for plasma nucleocapsid antigen and CRP after adjustment for baseline biomarker values and other baseline factors. Patients admitted to hospital with less favourable 5-day biomarker trajectories had worse prognosis, suggesting that persistent viral burden might drive inflammation in the pathogenesis of COVID-19, identifying patients that might benefit from escalation of antiviral or anti-inflammatory treatment. US National Institutes of Health.

#1257 Complement system is overactivated in patients with IgA nephropathy after COVID-19

Abstract Background and Aims Since the coronavirus disease of 2019 (COVID-19) pandemic swept across the world, the flare of immune-mediated disease following SARS-CoV-2 infection has emerged rapidly. IgA nephropathy (IgAN), which has been confirmed as a complement mediated autoimmune disease, is also one of the common glomerulonephritis associated with COVID-19. Here, we aim to investigate the clinical and immunological characteristics of patients with IgAN after COVID-19. Method From December 2022 to April 2023, 33 patients with renal biopsy-proven IgAN (Group CoV) at Beijing Anzhen hospital who had experienced COVID-19 before renal biopsy were enrolled. Meanwhile, 44 patients with IgAN (Group non-CoV) diagnosed from December 2018 to April 2019 (the same period in the previous year, with no COVID-19) were enrolled as control. Plasma FHR-5, MBL, C5a, soluble C5b-9, serum Gd-IgA1 and IgA immune complex were detected by enzyme-linked immunosorbent assay. Glomerular staining for FHR-5, C3c, MAC and Gd-IgA1 were detected by immunofluorescence. SARS-CoV-2 nucleocapsid was detected by immunohistochemistry. Clinicopathological and immunological features between the two groups were analyzed. Results The median time between onset of COVID-19 symptoms and the time of renal biopsy was 12 (6, 18) weeks, ranging from 1 to 26 weeks. SARS-CoV-2 nucleocapsid antigen expression in the renal tubular epithelial cells were observed in 11/33 (33.3%) patients. Compared with Group non-CoV, the level of eGFR was significantly lower in Group CoV[67.30 (52.94, 96.43) ml/min per 1.73 m2 vs. 101.04 (74.35, 120.98) ml/min per 1.73 m2, p = 0.010]. Histologically, compared with Group non-CoV, Group CoV presented with more percentages of segmental glomerulosclerosis/adhesion (p = 0.003) and less percentages of mesangial hypercellularity (p = 0.017). The level of plasma level of C5a [16.45 (10.48, 22.07) ng/ml vs. 3.68 (2.70, 4.47) ng/ml], soluble C5b-9 [614.43 (481.66, 770.02) ng/ml vs. 358.08 (304.83, 471.77) ng/ml], FHR5 [4.01 (3.51, 4.48) ug/ml vs. 2.73 (2.25, 3.21) ug/ml]were all significantly higher in Group CoV compared with Group non-CoV, respectively (p &amp;lt; 0.001). Although the plasma level of MBL showed no difference in Group CoV and Group non-CoV, the proportion of high levels of MBL in Group CoV was higher than in Group non-CoV (p = 0.025). Compared with Group non-CoV, the intensity of glomerular MAC deposition was much stronger in Group CoV. There were no significant differences in serum levels of IgA, Gd-IgA1, IgA immune complex between two Groups. However, a greater intensity of Gd-IgA1 deposition in glomerular mesangial and capillary areas has been exhibited in Group CoV than those in Group non-CoV (p = 0.005). Conclusion For IgAN after COVID-19, mucosal immune responses to SARS-CoV-2 infection may result in the overactivation of systemic and renal local complement system, which is the first hit, and increased glomerular deposition of Gd-IgA1 is the second hit. We think that these two hits may lead to renal dysfunction and promote renal progression in IgAN patients. It is crucial to emphasize the necessity for long term follow-up in these patients of IgAN after COVID-19. Additionally, we propose that inhibitors targeting the mediators of complement activation might be a promising therapy for these IgAN patients after SARS-CoV-2 infection.

Does coronavirus disease 2019 history alone increase the risk of postoperative pulmonary complications after surgery? Prospective observational study using serology assessment.

Concern exists about the increasing risk of postoperative pulmonary complications in patients with a history of coronavirus disease 2019 (COVID-19). We conducted a prospective observational study that compared the incidence of postoperative pulmonary complications in patients with and without a history of COVID-19. From August 2022 to November 2022, 244 adult patients undergoing major non-cardiac surgery were enrolled and allocated either to history or no history of COVID-19 groups. For patients without a history of confirming COVID-19 diagnosis, we tested immunoglobulin G to nucleocapsid antigen of SARS-CoV-2 for serology assessment to identify undetected infection. We compared the incidence of postoperative pulmonary complications, defined as a composite of atelectasis, pleural effusion, pulmonary edema, pneumonia, aspiration pneumonitis, and the need for additional oxygen therapy according to a COVID-19 history. After excluding 44 patients without a COVID-19 history who were detected as seropositive, 200 patients were finally enrolled in this study, 100 in each group. All subjects with a COVID-19 history experienced no or mild symptoms during infection. The risk of postoperative pulmonary complications was not significantly different between the groups according to the history of COVID-19 (24.0% vs. 26.0%; odds ratio, 0.99; 95% confidence interval, 0.71-1.37; P-value, 0.92). The incidence of postoperative pulmonary complications was also similar (27.3%) in excluded patients owing to being seropositive. Our study showed patients with a history of no or mild symptomatic COVID-19 did not show an increased risk of PPCs compared to those without a COVID-19 history. Additional precautions may not be needed to prevent PPCs in those patients.

An indirect ELISA for detecting anti-SARS-CoV-2 antibodies in human sera using a baculovirus-expressed recombinant nucleocapsid antigen

This study focuses on the development and initial assessment of an indirect IgG enzyme-linked immunosorbent assay (ELISA) specifically designed to detect of anti-SARS-CoV-2 antibodies. The unique aspect of this ELISA method lies in its utilization of a recombinant nucleocapsid (N) antigen, produced through baculovirus expression in insect cells. Our analysis involved 292 RT-qPCR confirmed positive serum samples and 54 pre-pandemic healthy controls. The process encompassed cloning, expression, and purification of the SARS-CoV-2 N gene in insect cells, with the resulted purified protein employed in our ELISA tests. Statistical analysis yielded an Area Under the Curve of 0.979, and the optimized cut-off exhibited 92 % sensitivity and 94 % specificity. These results highlight the ELISA's potential for robust and reliable serological detection of SARS-CoV-2 antibodies. Further assessments, including a larger panel size, reproducibility tests, and application in diverse populations, could enhance its utility as a valuable biotechnological solution for diseases surveillance.

SARS-CoV-2 RNA and Nucleocapsid Antigen Are Blood Biomarkers Associated With Severe Disease Outcomes That Improve in Response to Remdesivir.

Although antivirals remain important for the treatment COVID-19, methods to assess treatment efficacy are lacking. Here, we investigated the impact of remdesivir on viral dynamics and their contribution to understanding antiviral efficacy in the multicenter Adaptive COVID-19 Treatment Trial 1, which randomized patients to remdesivir or placebo. Longitudinal specimens collected during hospitalization from a substudy of 642 patients with COVID-19 were measured for viral RNA (upper respiratory tract and plasma), viral nucleocapsid antigen (serum), and host immunologic markers. Associations with clinical outcomes and response to therapy were assessed. Higher baseline plasma viral loads were associated with poorer clinical outcomes, and decreases in viral RNA and antigen in blood but not the upper respiratory tract correlated with enhanced benefit from remdesivir. The treatment effect of remdesivir was most pronounced in patients with elevated baseline nucleocapsid antigen levels: the recovery rate ratio was 1.95 (95% CI, 1.40-2.71) for levels >245 pg/mL vs 1.04 (95% CI, .76-1.42) for levels <245 pg/mL. Remdesivir also accelerated the rate of viral RNA and antigen clearance in blood, and patients whose blood levels decreased were more likely to recover and survive. Reductions in SARS-CoV-2 RNA and antigen levels in blood correlated with clinical benefit from antiviral therapy. NCT04280705 (ClinicalTrials.gov).