Proteomic and Cellular Characterization of Omicron Breakthrough Infections and a Third Homologous or Heterologous Boosting Vaccination in a Longitudinal Cohort.

The understanding of dynamic plasma proteome features in hybrid immunity and breakthrough infection is limited. A deeper understanding of the immune differences between heterologous and homologous immunization could assist in the future establishment of vaccination strategies. In this study, 40 participants who received a third dose of either a homologous BBIBP-CorV or a heterologous ZF2001 protein subunit vaccine following two doses of inactivated coronavirus disease 2019 vaccines and 12 patients with BA2.2 breakthrough infections were enrolled. Serum samples were collected at days 0, 28, and 180 following the boosting vaccination and breakthrough and then analyzed using neutralizing antibody tests and mass spectrometer-based proteomics. Mass cytometry of peripheral blood mononuclear cell samples was also performed in this cohort. The chemokine signaling pathway and humoral response markers (IgG2 and IgG3) associated with infection were found to be upregulated in breakthrough infections compared to vaccination-induced immunity. Elevated expression of IGKV, IGHV, IL-17 signaling, and the phagocytosis pathway, along with lower expression of FGL2, were correlated with higher antibody levels in the boosting vaccination groups. The MAPK signaling pathway and Fc gamma R-mediated phagocytosis were more enriched in the heterologous immunization groups than in the homologous immunization groups. Breakthrough infections can trigger more intensive inflammatory chemokine responses than vaccination. T-cell and innate immune activation have been shown to be closely related to enhanced antibody levels after vaccination and therefore might be potential targets for vaccine adjuvant design.

Clinical Characteristics, Treatment, and Prognosis of Osteoarticular Brucellosis: A Retrospective Real-World Study in Shenyang, China, 2014-2019.

This study aimed to assess the clinical characteristics, treatment, and prognosis of osteoarticular brucellosis. We conducted a retrospective study enrolling brucellosis patients from the Sixth People's Hospital of Shenyang between September 2014 and June 2019. A total of 1917 participants were admitted during this period. After applying propensity score matching, we retrospectively analyzed 429 patients with osteoarthritis and 429 patients without osteoarthritis. The primary outcome was treatment completion. The secondary outcome was symptom disappearance and seroconversion. Brucellosis patients with osteoarthritis had longer treatment course (160 [134.3-185.7] vs. 120 [102.3-137.7] d, p = 0.008) than those without osteoarthritis. The most common involved site was lumbar vertebrae (290 [67.6%]) in brucellosis patients with osteoarthritis. Longer symptom duration (90 [83.0-97.0] vs. 42 [40.2-43.8], p < 0.001) along with no significant difference in seroconversion (180 [178.8-181.2] vs. 180 [135.1-224.9], p = 0.212) was observed in osteoarthritis patients with treatment course >90 d. Peripheral joint involvement (adjusted hazard ratio [95% confidence interval] 1.485 [1.103-1.999]; p = 0.009) had a shorter symptom duration compared with shaft joint involvement. No significant differences were observed in treatment therapy between doxycycline plus rifampin (DR) or plus cephalosporins (DRC) in treatment course (p = 0.190), symptom persistence (p = 0.294), and seroconversion (p = 0.086). Lumbar vertebra was the most commonly involved site. Even if all symptoms disappeared, Serum agglutination test potentially remained positive in some patients. Compared with peripheral arthritis, shaft arthritis was the high-risk factor for longer symptom duration. The therapeutic effects were similar between DR and DRC. In summary, our study provided important insights into the clinical characteristics, treatment, and outcomes of osteoarticular brucellosis. Clinical Trial Registration number: NCT04020536.

Early identification and severity prediction of acute respiratory infection (ESAR): a study protocol for a randomized controlled trial.

BACKGROUND: The outbreak of SARS-CoV-2 at the end of 2019 sounded the alarm for early inspection on acute respiratory infection (ARI). However, diagnosis pathway of ARI has still not reached a consensus and its impact on prognosis needs to be further explored. METHODS: ESAR is a multicenter, open-label, randomized controlled, non-inferiority clinical trial on evaluating the diagnosis performance and its impact on prognosis of ARI between mNGS and multiplex PCR. Enrolled patients will be divided into two groups with a ratio of 1:1. Group I will be directly tested by mNGS. Group II will firstly receive multiplex PCR, then mNGS in patients with severe infection if multiplex PCR is negative or inconsistent with clinical manifestations. All patients will be followed up every 7 days for 28 days. The primary endpoint is time to initiate targeted treatment. Secondary endpoints include incidence of significant events (oxygen inhalation, mechanical ventilation, etc.), clinical remission rate, and hospitalization length. A total of 440 participants will be enrolled in both groups. DISCUSSION: ESAR compares the efficacy of different diagnostic strategies and their impact on treatment outcomes in ARI, which is of great significance to make precise diagnosis, balance clinical resources and demands, and ultimately optimize clinical diagnosis pathways and treatment strategies. Trial registration Clinicaltrial.gov, NCT04955756, Registered on July 9th 2021.

Evaluation of droplet digital PCR rapid detection method and precise diagnosis and treatment for suspected sepsis (PROGRESS): a study protocol for a multi-center pragmatic randomized controlled trial.

BACKGROUND: Sepsis is still a major public health concern and a medical emergency due to its high morbidity and mortality. Accurate and timely etiology diagnosis is crucial for sepsis management. As an emerging rapid and sensitive pathogen detection tool, digital droplet PCR (ddPCR) has shown promising potential in rapid identification of pathogens and antimicrobial resistance genes. However, the diagnostic value and clinical impact of ddPCR tests remains to be studied in patients with suspected sepsis. PROGRESS trial is aimed to evaluate the clinical effectiveness of a novel ddPCR assay compared with standard practice. METHODS: PROGRESS is a multicenter, open-label, pragmatic randomized controlled trial (pRCT) set in ten hospitals, including departments of infectious disease and intensive care units. In this study, a total of 2292 patients with suspected sepsis will be randomly assigned to two arms: the ddPCR group and the control group with a ratio of 3:1. The primary outcome is the diagnostic efficacy, that is, the sensitivity and specificity of the ddPCR assay compared with the synchronous blood culture. Secondary outcomes include the mortality rates and the mean Sequential Organ Failure Assessment (SOFA) score at follow-up time points, the length of stay in the hospital, the time to directed antimicrobial therapy, duration of broad-spectrum antibiotic use, and the EQ-5D-5L score on day 90. DISCUSSION: It is the first multicenter pragmatic RCT to explore the diagnostic efficacy and clinical impact of the ddPCR assay in patients with suspected sepsis, taking advantage of both RCT's ability to establish causality and the feasibility of pragmatic approaches in real-world studies (RWS). This trial will help us to get a comprehensive view of the assay's capacity for precise diagnosis and treatment of sepsis. It has the potential to monitor the pathogen load change and to guide the antimicrobial therapy, making a beneficial impact on the prognosis of sepsis patients. TRIAL REGISTRATION: ClinicalTrial.gov, NCT05190861. Registered January 13, 2022-'Retrospectively registered', https://clinicaltrials.gov/ct2/show/NCT05190861 .

Risk of reinfection and severity with the predominant BA.5 Omicron subvariant China, from December 2022 to January 2023.

Data on reinfection in large Asian populations are limited. In this study, we aimed to evaluate the reinfection rate, disease severity, and time interval between the infections in the symptomatic and asymptomatic populations which are firstl infected with BA.2 Omicron Variant. We retrospectively included adult patients with COVID-19 discharged from four designated hospitals between 27 April 2021 and 30 November 2022, who were interviewed via telephone from 29 January to 1 March 2023. Univariable and multivariable analyses were used to explore risk factors associated with reinfection. A total of 16,558 patients were followed up, during the telephone survey of an average of 310.0 days, 1610 (9.72%) participants self-reported reinfection. The mean time range of reinfection was 257.9 days. The risks for reinfection were analysed using multivariable logistic regression. Patients with severe first infection were at higher risk for reinfection (aORs, 2.50; P < 0.001). The male (aORs,0.82; P < 0.001), the elderly (aORs, 0.44; P < 0.001), and patients with full vaccination (aORs, 0.67; P < 0.001) or booster (aORs, 0.63; P < 0.001) had the lower risk of reinfection. Patients over 60 years of age (aORs,9.02; P = 0.006) and those with ≥2 comorbidities (aORs,11.51; P = 0.016). were at higher risk for severe reinfection. The number of clinical manifestations of reinfection increases in people with severe first infection (aORs, 2.82; P = 0.023). The overall reinfection rate was 9.72%, and the reinfection rate of Omicron-to-Omicron subvariants was 9.50% at one year. The severity of Omicron-Omicron reinfection decreased. Data from our clinical study may provide clinical evidence and bolster response preparedness for future COVID-19 reinfection waves.

Alterations of pathogen transmission patterns and attenuated immune stimulation might be the cause of increased adult respiratory infections cases in 2023, results from a multi-center study in mainland China.

Background: Several respiratory infections outbreaks have been observed in mainland China after reduction of non-pharmaceutical interventions. Other countries have seen increases in respiratory infections outside typical seasons post-COVID-19, warranting investigation into underlying causes. Methods: We established monitoring networks for suspected respiratory infection in 14 tertiary hospitals nationwide. PCR for SARS-CoV-2, influenza A and B were performed on 3708 respiratory specimens and deep sequencing were conducted to identify co-infections or newly emerging microbes in 2023. Viral evolutionary analysis was completed. We retrospectively detected serum antibody level for various respiratory pathogens from 4324 adults without respiratory infections over 7 years to observe its dynamic curves. Findings: SARS-CoV-2 and influenza A were the main pathogens during outbreaks in 2023, bacterial-virus and bacterial-bacterial co-infections were most detected, but community co-infections didn't significantly increase pneumonia incidence. Different SARS-CoV-2 and influenza variants were present in different outbreaks, and no novel pathogens were found. The epidemiological patterns of influenza A, COVID-19 and etc. were altered, exhibiting characteristics of being "staggered" compared to most global regions, and potentially led to "overlapping prevalence". Binding antibody testing showed regular fluctuation, without significant decrease against common respiratory pathogens in adults. Influenza A antibody stimulation was attenuated during the 2023 outbreak. Conclusions: "Misaligned" alteration in seasonal respiratory disease patterns possibly caused combined epidemics, leading to cases spike in China, 2023. In adults, antibody levels didn't show significant decline, but reduced immune response to influenza during 2020-2023 emphasizes the need for consistent vaccination during pandemics.

A one-year follow-up study of systematic impact of long COVID symptoms among patients post SARS-CoV-2 omicron variants infection in Shanghai, China.

Long COVID hinders people from normal life and work, posing significant medical and economic challenges. Nevertheless, comprehensive studies assessing its impact on large populations in Asia are still lacking. We tracked over 20,000 patients infected with COVID-19 for the first time during the Omicron BA.2 outbreak in Shanghai from March-June 2022 for one year. Of the 21,799 COVID-19 patients who participated in the 6-month telephone follow-up, 1939 (8.89%) had self-reported long COVID symptoms. 450 long COVID patients participated in the 6-month outpatient follow-up. Participants underwent healthy physical examinations and questionnaires focused on long-COVID-related symptoms and mental health. Mobility problem (P < 0.001), personal care problem (P = 0.003), usual activity problem (P < 0.001), pain/discomfort (P < 0.001), anxiety/depression (P = 0.001) and PTSD (P = 0.001) were more prevalent in long COVID patients than in healthy individuals, but no significant differences were found between the two groups on chest CT and laboratory examinations. Of the 856 long COVID patients who participated in the 12-month follow-up, 587 (68.5%) had their symptoms resolved. In the multivariable logistic analysis, females (P < 0.001), youth (age <40 years) (P < 0.001), ≥ 2 comorbidities (P = 0.009), and severe infection in the acute phase (P = 0.006) were risk factors for developing long COVID. Middle age (40-60 years) was a risk factor for persistent long COVID one year after hospital discharge (P = 0.013). The study found that long COVID mainly manifested as subjective symptoms and impacts partial patients' quality of life and mental status. After one year, most (68.5%) of the patients recovered from long COVID with no impairment of organ function observed.

Utility of clinical metagenomics in diagnosing malignancies in a cohort of patients with Epstein-Barr virus positivity.

Backgrounds: Differentiation between benign and malignant diseases in EBV-positive patients poses a significant challenge due to the lack of efficient diagnostic tools. Metagenomic Next-Generation Sequencing (mNGS) is commonly used to identify pathogens of patients with fevers of unknown-origin (FUO). Recent studies have extended the application of Next-Generation Sequencing (NGS) in identifying tumors in body fluids and cerebrospinal fluids. In light of these, we conducted this study to develop and apply metagenomic methods to validate their role in identifying EBV-associated malignant disease. Methods: We enrolled 29 patients with positive EBV results in the cohort of FUO in the Department of Infectious Diseases of Huashan Hospital affiliated with Fudan University from 2018 to 2019. Upon enrollment, these patients were grouped for benign diseases, CAEBV, and malignant diseases according to their final diagnosis, and CNV analysis was retrospectively performed in 2022 using samples from 2018 to 2019. Results: Among the 29 patients. 16 of them were diagnosed with benign diseases, 3 patients were diagnosed with CAEBV and 10 patients were with malignant diseases. 29 blood samples from 29 patients were tested for mNGS. Among all 10 patients with malignant diagnosis, CNV analysis suggested neoplasms in 9 patients. Of all 19 patients with benign or CAEBV diagnosis, 2 patients showed abnormal CNV results. The sensitivity and specificity of CNV analysis for the identification for tumors were 90% and 89.5%, separately. Conclusions: The application of mNGS could assist in the identification of microbial infection and malignancies in EBV-related diseases. Our results demonstrate that CNV detection through mNGS is faster compared to conventional oncology tests. Moreover, the convenient collection of peripheral blood samples adds to the advantages of this approach.

Neutralization against Omicron subvariants after BA.5/BF.7 breakthrough infection weakened as virus evolution and aging despite repeated prototype-based vaccination1.

BACKGROUND: Omicron had swept the mainland China between December 2022 and January 2023, while SARS-CoV-2 still continued to evolve. To fully prepare for the next wave, it's urgent to evaluate the humoral immune response post BA.5/BF.7 breakthrough infection against predominant sub-lineages among existing vaccination strategies and the elders. METHOD: This study enrolled a longitudinal young-adult cohort from 2/3-dose vaccination to 1 month after breakthrough infection, and an elder cohort at 1 month after breakthrough infection. Seral samples were collected and tested for humoral immune response to SARS-CoV-2 subvariants including WT, BA.2, BA.5, BF.7, BQ.1.1, CH.1.1, XBB.1.5. RESULTS: BA.5/BF.7 breakthrough infection induced higher neutralization activity than solely vaccination in all SARS-CoV-2 strains, while the latest Omicron subvariants, BQ.1.1, CH.1.1, XBB.1.5, exhibited the strongest neutralization evasion ability. There was a negative correlation between age and humoral immune response in WT, BA.5, BQ.1.1, and XBB.1.5. Compared to non-vaccination groups, breakthrough infection in two-dose vaccination groups had significantly higher neutralizing antibody against WT, BA.2, BA.5, BF.7 but not to BQ.1.1, CH.1.1, XBB.1.5 while booster dose against the prototype prior-breakthrough would not further significantly enhance individual's humoral responses against the latest Omicron subvariants. CONCLUSIONS: Newer variants manifest increasing immune evasion from neutralization and repeated prototype-based booster vaccines may not further enhance neutralizing antibody against emerging new variants. Older adults have lower levels of neutralizing antibody. Future vaccination strategies should aim to enhance effective neutralization to contemporary variants.

A diagnostic accuracy study comparing RNA LAMP, direct LAMP, and rapid antigen testing from nasopharyngeal swabs.

Introduction: During the coronavirus disease 2019 (COVID-19) pandemic, the early detection and isolation of individuals infected with severe acute respiratory syndrome coronavirus disease 2 (SARS-CoV-2) through mass testing can effectively prevent disease transmission. SARS-CoV-2 nucleic acid rapid detection based on loop-mediated isothermal amplification (LAMP) may be appropriate to include in testing procedures. Methods: We used 860 nasopharyngeal specimens from healthcare workers of Huashan Hospital and COVID-19 patients collected from April 7th to 21st, 2022, to assess the clinical diagnostic performance of the LAMP assay marketed by Shanghai GeneSc Biotech and compared it to the result of a rapid antigen test (RAT) head-to-head. Results: Overall, the diagnostic performance of LAMP assay and RAT were as follows. The LAMP assay represented higher sensitivity and specificity than RAT, especially in the extracted RNA samples. The sensitivity was 70.92% and 92.91% for direct LAMP and RNA-LAMP assay, respectively, while the specificity was 99.86% and 98.33%. The LAMP assay had overall better diagnostic performance on the specimens with relatively lower C t values or collected in the early phase (≤7 days) of COVID-19. The combination of LAMP assay and RAT improved diagnostic efficiency, providing new strategies for rapidly detecting SARS-CoV-2. Conclusion: The LAMP assay are suitable for mass screenings of SARS-CoV-2 infections in the general population.

Identification of CKD, bedridden history and cancer as higher-risk comorbidities and their impact on prognosis of hospitalized Omicron patients: a multi-centre cohort study.

To further describe the effect of the "fragile population" and their "higher-risk" comorbidities on prognosis among hospitalized Omicron patients, this observational cohort study enrolled hospitalized patients confirmed with SARS-CoV-2 during the 2022 Omicron wave in Shanghai, China. The primary outcome was progression to severe or critical cases. The secondary outcome was viral shedding time from the first positive SARS-CoV-2 detection. A total of 847 participants were enrolled, most of whom featured as advanced age (>70 years old: 30.34%), not fully vaccinated (55.84%), combined with at least 1 comorbidity (65.41%). Multivariate cox regression suggested age >70 years old (aHR[95%CI] 0.78[0.61-0.99]), chronic kidney disease (CKD) stage 4-5 (aHR[95%CI] 0.61[0.46-0.80]), heart conditions (aHR[95%CI] 0.76[0.60-0.97]) would elongate viral shedding time and fully/booster vaccination (aHR[95%CI] 1.4 [1.14-1.72]) would shorten this duration. Multivariate logistic regression suggested CKD stage 4-5 (aHR[95%CI] 3.21[1.45-7.27]), cancer (aHR[95%CI] 9.52[4.19-22.61]), and long-term bedridden status (aHR[95%CI] 4.94[2.36-10.44]) were the "higher" risk factor compared with the elderly, heart conditions, metabolic disorders, isolated hypertension, etc. for severity while female (aHR[95%CI] 0.34[0.16-0.68]) and fully/booster Vaccination (aHR[95%CI] 0.35[0.12-0.87]) could provide protection from illness progression. CKD stage 4-5, cancer and long-term bedridden history were "higher-risk" factors among hospitalized Omicron patients for severity progression while full vaccination could provide protection from illness progression.

Cellular basis of enhanced humoral immunity to SARS-CoV-2 upon homologous or heterologous booster vaccination analyzed by single-cell immune profiling.

SARS-CoV-2 vaccine booster dose can induce a robust humoral immune response, however, its cellular mechanisms remain elusive. Here, we investigated the durability of antibody responses and single-cell immune profiles following booster dose immunization, longitudinally over 6 months, in recipients of a homologous BBIBP-CorV/BBIBP-CorV or a heterologous BBIBP-CorV/ZF2001 regimen. The production of neutralizing antibodies was dramatically enhanced by both booster regimens, and the antibodies could last at least six months. The heterologous booster induced a faster and more robust plasmablast response, characterized by activation of plasma cells than the homologous booster. The response was attributed to recall of memory B cells and the de novo activation of B cells. Expanded B cell clones upon booster dose vaccination could persist for months, and their B cell receptors displayed accumulated mutations. The production of antibody was positively correlated with antigen presentation by conventional dendritic cells (cDCs), which provides support for B cell maturation through activation and development of follicular helper T (Tfh) cells. The proper activation of cDC/Tfh/B cells was likely fueled by active energy metabolism, and glutaminolysis might also play a general role in promoting humoral immunity. Our study unveils the cellular mechanisms of booster-induced memory/adaptive humoral immunity and suggests potential strategies to optimize vaccine efficacy and durability in future iterations.