More rapid blood interferon α2 decline in fatal versus surviving COVID-19 patients.

Background: The clinical outcome of COVID-19 pneumonia is highly variable. Few biological predictive factors have been identified. Genetic and immunological studies suggest that type 1 interferons (IFN) are essential to control SARS-CoV-2 infection. Objective: To study the link between change in blood IFN-α2 level and plasma SARS-Cov2 viral load over time and subsequent death in patients with severe and critical COVID-19. Methods: One hundred and forty patients from the CORIMUNO-19 cohort hospitalized with severe or critical COVID-19 pneumonia, all requiring oxygen or ventilation, were prospectively studied. Blood IFN-α2 was evaluated using the Single Molecule Array technology. Anti-IFN-α2 auto-Abs were determined with a reporter luciferase activity. Plasma SARS-Cov2 viral load was measured using droplet digital PCR targeting the Nucleocapsid gene of the SARS-CoV-2 positive-strand RNA genome. Results: Although the percentage of plasmacytoid dendritic cells was low, the blood IFN-α2 level was higher in patients than in healthy controls and was correlated to SARS-CoV-2 plasma viral load at entry. Neutralizing anti-IFN-α2 auto-antibodies were detected in 5% of patients, associated with a lower baseline level of blood IFN-α2. A longitudinal analysis found that a more rapid decline of blood IFN-α2 was observed in fatal versus surviving patients: mortality HR=3.15 (95% CI 1.14-8.66) in rapid versus slow decliners. Likewise, a high level of plasma SARS-CoV-2 RNA was associated with death risk in patients with severe COVID-19. Conclusion: These findings could suggest an interest in evaluating type 1 IFN treatment in patients with severe COVID-19 and type 1 IFN decline, eventually combined with anti-inflammatory drugs. Clinical trial registration: https://clinicaltrials.gov, identifiers NCT04324073, NCT04331808, NCT04341584.

Hepatitis E Virus Infection in Patients with Chronic Inflammatory Bowel Disease Treated with Immunosuppressive Therapy.

BACKGROUND: Medical treatment of inflammatory bowel disease (IBD) has evolved significantly, and treatment with immunomodulators is recommended. These medications may alter the patient's immune response and increase the risk of opportunistic infections. Our aim was to evaluate the prevalence and the incidence of acute or chronic HEV infection in IBD patients under immunomodulatory treatment. PATIENTS AND METHODS: We conducted a retrospective, multicenter, observational study between 2017 and 2018. IBD outpatients hospitalized for the infusion of immunomodulators were included in 16 French centers. During their daily hospitalization, blood samples were drawn for HEV serology (IgM and IgG) and HEV RNA detection. RESULTS: A total of 488 patients were included, of which 327 (67%) patients had Crohn's disease and 161 (33%) ulcerative colitis. HEV IgM was detected in 3 patients, but HEV RNA was undetectable in all patients. The HEV IgG seroprevalence rate was 14.2%. IgG-positive patients were older at sampling (p = 0.01) and IBD diagnosis (p = 0.03), had higher seafood consumption (p = 0.01) and higher doses of azathioprine (p = 0.03). Ileal and upper digestive tract involvement was more frequent in IgG-positive patients (p = 0.009), and ileocolic involvement was more frequent in IgG-negative patients (p = 0.01). Under multivariate analysis, age > 50 years [OR: 2.21 (1.26, to 3.85), p = 0.004] was associated with previous HEV infection. CONCLUSION: Systematic screening for HEV infection is not needed among IBD patients on immunomodulatory medications. However, in the event of abnormal liver test findings, HEV should be part of the classic diagnostic assessment.

Validation of a SARS-CoV-2 Surrogate Virus Neutralization Test in Recovered and Vaccinated Healthcare Workers.

Vaccination against COVID-19 is the main public health approach to fight against the pandemic. The Spike (S) glycoprotein of SARS-CoV-2 is the principal target of the neutralizing humoral response. We evaluated the analytical and clinical performances of a surrogate virus neutralization test (sVNT) compared to conventional neutralization tests (cVNTs) and anti-S eCLIA assays in recovered and/or vaccinated healthcare workers. Our results indicate that sVNTs displayed high specificity and no cross-reactivity. Both eCLIA and sVNT immunoassays were good at identifying cVNT serum dilutions ≥1:16. The optimal thresholds when identifying cVNT titers ≥1:16, were 74.5 U/mL and 49.4 IU/mL for anti-S eCLIA and sVNT, respectively. Our data show that neutralizing antibody titers (Nab) differ from one individual to another and may diminish over time. Specific assays such as sVNTs could offer a reliable complementary tool to routine anti-S serological assays.

Persistence of Neutralizing Antibodies and Clinical Protection up to 12†Months After Severe Acute Respiratory Syndrome Coronavirus 2 Infection in the Elderly.

Background: Coronavirus disease 2019 (COVID-19) has severely affected the elderly, who are expected to display decreased immune responses due to immunosenescence. Methods: This study retrospectively assesses neutralizing antibody (NAb) production up to 12 months after infection in long-term care patients. We used Roche Diagnostics immunoassay to quantify anti-spike (S) antibodies and a competitive immunoassay from YHLO as a surrogate test for NAb. Results: We included 91 patients (mean age, 86 years). There was no significant variation in anti-S titers over time. There was a significant decrease of NAb titers between month 3 and month 6 but no further significant change up to month 12. Overall, 75 of 91 (82%) and 52 of 91 (57%) patients had, at least once, anti-S titers >75 U/mL and NAb titers >50 AU/mL, respectively, corresponding to a significant neutralizing activity in vitro. All 68 patients studied at M12 had detectable anti-S antibodies and 60 (88%) had detectable NAb; 60 of 68 (88%) and 29 of 68 (42.6%) still had anti-S titers >75 U/mL and NAb titers >50 AU/mL. Higher NAb titers were correlated with severe infection, higher levels of C-reactive protein, and lower lymphocyte counts. No patient developed reinfection. Conclusions: Elderly people can display robust and persistent humoral response after severe acute respiratory syndrome coronavirus 2 infection, with NAb lasting up to 12 months.

Clinical Added Value of SARS-CoV-2 Antigen Detection in Blood Samples.

This study evaluated the performances of immunoassays (LFIA and ELISA) designed for SARS-CoV-2 Antigen (Ag)-detection in nasopharyngeal (NP) and serum samples in comparison to RT-PCR. NP samples from patients with respiratory symptoms (183 RT-PCR-positive and 74 RT-PCR-negative samples) were collected from March to April and November to December 2020. Seroconversion and antigen dynamics were assessed by symptom onset and day of RT-PCR diagnosis. Serum samples from 87 COVID-19 patients were used to investigate the added value of Ag quantification, at diagnosis and during follow-up. The sensitivity of COVID-VIRO-LFIA on samples with Ct ≤ 33, considered as the contagious threshold, was 86% on NPs (CI 95%: 79-90.5) and 76% on serum samples (CI 95%: 59.4-88), with a specificity of 100%. Serum N-Ag was detected during active infection as early as day two from symptom onset, with a diagnostic sensitivity of 81.5%. Within one week of symptom onset, diagnostic sensitivity and specificity reached 90.9% (95% CI, 85.1%-94.6%) and 98.3% (95% CI, 91.1%-99.9%), respectively. Serum N-Ag concentration closely correlated with disease severity. Longitudinal analysis revealed the simultaneous increase of antibodies and decrease of N-Ag. Sensitivities of COVID-VIRO-LFIA and COV-QUANTO-ELISA tests on NP and serum samples were close to 80%. They are suitable COVID-19-laboratory diagnostic tests, particularly when blood samples are available, thus reducing the requirement for NP sampling, and subsequent PCR analysis. ELISA titers may help to identify patients at risk of poor outcomes.

Contribution of Serum Cytomegalovirus PCR to Diagnosis of Early CMV Primary Infection in Pregnant Women.

(1) Background: What is the role of serum CMV PCR in the diagnosis of recent primary infection (PI) in pregnant women when IgG avidity is uninformative? (2) Methods: Retrospective cohort study to compare serum versus whole blood CMV PCR. (a) Qualitative assessment: CMV PCR was performed on 123 serum samples and 74 whole blood samples collected from 132 pregnant women with recent CMV PI. PCR positivity rate was used to calculate sensitivity in serum and whole blood. (b) Quantitative assessment: CMV PCR was performed on 72 paired samples of serum and whole blood collected on the same day from 57 patients. (3) Results: In pregnant women, PCR positivity rate was 89% for serum samples versus 100% in whole blood in the case of very recent PI (<15 days), but only 27% in serum versus 68% in whole blood for PI occurring from 6 weeks to 3 months before. Comparing CMV viral loads between serum and whole blood, we determined the limit of CMV DNA detection in serum as 3 log copies/mL (whole blood equivalent). (4) Conclusions: Serum CMV PCR is reliable in confirming PI in cases when only IgM is detected. It is therefore a valuable tool in introducing valaciclovir treatment as early as possible to prevent mother-to-child CMV transmission.

When EBV serology needs a blot to conclude: a case report / Quand un immunoblot EBV est nécessaire pour conclure une sérologie anti-EBV : cas clinique

A 14-year-old patient underwent liver transplant. Three months after transplantation, EBV associated lymphoma was suspected. EBV serologies before and after transplantation were particularly discrepant. Eighteen months before transplantation, the serological profile suggested a recent EBV primary-infection, four months before transplantation, we had isolated anti-EBNA IgG and five days after transplantation, the serological profile suggested a past infection. All EBV PCR performed on whole blood were negative. Retrospectively, immunoblots anti-EBV IgG were performed. Blots showed no anti-EBV IgG before and until the day of liver transplantation. Five days after transplantation, slight bands of both IgG anti-p18 (VCA) and anti-EBNA-1 were found, without any other serological marker. These were probably due to passive transfers of IgG during surgery. There was therefore no argument for an immunization against EBV before or after liver transplant for this patient.

Un patient de 14 ans a subi une transplantation hépatique. Trois mois après la transplantation, un lymphome associé à l'EBV a été suspecté. Les sérologies anti-EBV (IgG anti-VCA et anti-EBNA, IgM anti-VCA) montraient des résultats particulièrement discordants et ininterprétables : profil de primo-infection EBV récente 18 mois avant la greffe, IgG anti-EBNA isolées 4 mois avant la greffe et profil d'infection ancienne 5 jours après la greffe. Les PCR EBV réalisées sur sang total étaient toujours indétectables. Rétrospectivement, des immunoblots IgG anti-EBV ont été réalisés. Les blots ne montraient pas d'IgG anti-EBV en pré-greffe jusqu'au jour de la transplantation. Cinq jours après la greffe, le blot montrait la présence d'anti-p18 (VCA) et anti-EBNA-1 présents sous forme de traces, sans autre marqueur sérologique d'infection EBV, très probablement dues à des transferts passifs d'anticorps pendant la transplantation. Aucun argument sérologique ni moléculaire n'a été retenu pour une immunisation anti-EBV avant ou après la transplantation hépatique pour ce patient.

SARS-CoV-2 Genomic Characteristics and Clinical Impact of SARS-CoV-2 Viral Diversity in Critically Ill COVID-19 Patients: A Prospective Multicenter Cohort Study.

The SARS-CoV-2 variant of concern, α, spread worldwide at the beginning of 2021. It was suggested that this variant was associated with a higher risk of mortality than other variants. We aimed to characterize the genetic diversity of SARS-CoV-2 variants isolated from patients with severe COVID-19 and unravel the relationships between specific viral mutations/mutational patterns and clinical outcomes. This is a prospective multicenter observational cohort study. Patients aged ≥18 years admitted to 11 intensive care units (ICUs) in hospitals in the Greater Paris area for SARS-CoV-2 infection and acute respiratory failure between 1 October 2020 and 30 May 2021 were included. The primary clinical endpoint was day-28 mortality. Full-length SARS-CoV-2 genomes were sequenced by means of next-generation sequencing (Illumina COVIDSeq). In total, 413 patients were included, 183 (44.3%) were infected with pre-existing variants, 197 (47.7%) were infected with variant α, and 33 (8.0%) were infected with other variants. The patients infected with pre-existing variants were significantly older (64.9 ± 11.9 vs. 60.5 ± 11.8 years; p = 0.0005) and had more frequent COPD (11.5% vs. 4.1%; p = 0.009) and higher SOFA scores (4 [3-8] vs. 3 [2-4]; 0.0002). The day-28 mortality was no different between the patients infected with pre-existing, α, or other variants (31.1% vs. 26.2% vs. 30.3%; p = 0.550). There was no association between day-28 mortality and specific variants or the presence of specific mutations. At ICU admission, the patients infected with pre-existing variants had a different clinical presentation from those infected with variant α, but mortality did not differ between these groups. There was no association between specific variants or SARS-CoV-2 genome mutational pattern and day-28 mortality.

Performance of the QIAprep& Viral RNA UM Kit assay (Qiagen), an automatable method for RT-qPCR detection of SARS-CoV-2 without RNA extraction.

We evaluated the performance of the QIAprep& Viral RNA UM Kit (Qiagen) for SARS-CoV-2 detection. It displayed specificity and sensitivity required for SARS-CoV-2 RNA detection from swab transport media without RNA extraction. This method identifies accurately patients at risk of transmission while saving time and cost of extraction.

Transfusion-Transmitted Hepatitis A Virus, France, 2018.

We report a transfusion-transmitted hepatitis A virus infection in an immunocompromised patient in France, detected shortly after a transfusion of pathogen-reduced pooled platelets. This case raises questions about the efficacy of donor screening methods. Additional safety measures, such as routine donation screening, should be considered.

Rituximab Impairs B Cell Response But Not T Cell Response to COVID-19 Vaccine in Autoimmune Diseases.

OBJECTIVE: Antibody response to the messenger RNA (mRNA) COVID-19 vaccine has been shown to be diminished in rituximab (RTX)-treated patients. We undertook this study to compare humoral and T cell responses between healthy controls, patients with autoimmune diseases treated with RTX, and those treated with other immunosuppressants, all of whom had been vaccinated with 2 doses of the mRNA COVID-19 vaccine. METHODS: We performed anti-spike IgG and neutralization assays just before and 28 days after the second BNT162b2 (Pfizer-BioNTech) vaccine dose. The specific T cell response was assessed in activated CD4 and CD8 T cells using intracellular flow cytometry staining of cytokines (interferon-γ, tumor necrosis factor, and interleukin-2) after stimulation with SARS-CoV-2 spike peptide pools. RESULTS: A lower proportion of responders with neutralizing antibodies to the vaccine was observed in the RTX group (29%; n = 24) compared to the other immunosuppressants group (80%; n = 35) (P = 0.0001) and the healthy control group (92%; n = 26) (P < 0.0001). No patients treated with RTX in the last 6 months showed a response. Time since last infusion was the main factor influencing humoral response in RTX-treated patients. The functional CD4 and CD8 cellular responses to SARS-CoV-2 peptides for each single cytokine or polyfunctionality were not different in the RTX group compared to the other immunosuppressants group or the control group. In RTX-treated patients, the T cell response was not different between patients with and those without a humoral response. CONCLUSION: RTX induced a diminished antibody response to the mRNA COVID-19 vaccine, but the functional T cell response was not altered compared to healthy controls and autoimmune disease patients treated with other immunosuppressants. Further work is needed to assess the clinical protection granted by a functionally active T cell response in the absence of an anti-spike antibody response.

Accuracy assessment of total or IgG Immunoglobulin to hepatitis A virus tests around immunity threshold.

BACKGROUND: Anti-hepatitis A virus (HAV) antibody titers at 20 IU/L are assumed to correlate with protection against HAV challenge. METHODS: We examined the accuracy and precision of currently in use immunoassays for total or anti-HAV IgG determination, by repeated testing of dilutions of the international anti-HAV standard, within a 10-50 IU/mL concentration range. RESULTS AND CONCLUSION: Eight immunoassays were evaluated. All could confidently identify people who need to be vaccinated, or who might benefit from a booster vaccine: no positive interpretation for the 10 and 15 IU/mL concentrations. However, qualitative interpretation may differ from test to test in the 15-30 IU/mL range. This variation has to be taken into account when comparing seroprevalence data.

Emergence of SARS-CoV-2 resistance mutations in a patient who received anti-SARS-COV2 spike protein monoclonal antibodies: a case report.

BACKGROUND: To manage severe or potentially severe cases of CoronaVirus Disease 2019 (COVID-19), therapeutic monoclonal antibodies targeting Spike protein of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) have been designed. It has been noted in vitro that upon exposure to these treatments, mutations could be selected. CASE PRESENTATION: We here report the case of an immunosuppressed patient infected with a B.1.1.7 variant, who received a combination of monoclonal antibodies, and subsequently selected mutations K417N, E484K and Q493R on Spike protein of SARS-CoV-2. CONCLUSIONS: Our case raises the importance of monitoring SARS-CoV-2 mutations in patients receiving monoclonal antibodies and having persistent excretion of the virus, in order to offer optimal management of their infection, and strengthen prevention measures to avoid subsequent transmission of these selected variants.

Response to COVID-19 mRNA vaccination in multiple myeloma is conserved but impaired compared to controls.

Patients with multiple myeloma are at high risk of severe forms of COVID-19. Despite data showing diminished response to vaccine, the era of highly efficient mRNA vaccine might be a gamechanger. We sought to examine response to mRNA vaccine between healthy controls (n = 28) and multiple myeloma (MM) patients (n = 27). Response was analyzed 1 month after the second dose of anti-SARS-CoV-2 BNT162b2 vaccine. Multiple myeloma patients showed diminished levels of Anti-Spike IgG levels compared to controls, but with a high proportion of patients achieving a humoral response (89% vs. 97% in controls). Neutralizing antibodies were present in 74% of patients versus 96% of controls. Patients under current daratumumab treatment had neutralizing activity of anti-SARS-CoV-2 antibodies. Multiple myeloma patients show diminished response to SARS-COV-2 vaccine but with still high response rate. The main potential risk factor of non-response to COVID-19 vaccine was uncontrolled disease under treatment.

Quantitative Assessment of SARS-CoV-2 Virus in Nasopharyngeal Swabs Stored in Transport Medium by a Straightforward LC-MS/MS Assay Targeting Nucleocapsid, Membrane, and Spike Proteins.

Alternative methods to RT-PCR for SARS-CoV-2 detection are investigated to provide complementary data on viral proteins, increase the number of tests performed, or identify false positive/negative results. Here, we have developed a simple mass spectrometry assay for SARS-CoV-2 in nasopharyngeal swab samples using common laboratory reagents. The method employs high sensitivity and selectivity targeted mass spectrometry detection, monitoring nine constitutive peptides representative of the three main viral proteins and a straightforward pellet digestion protocol for convenient routine applications. Absolute quantification of N, M, and S proteins was achieved by addition of isotope-labeled versions of best peptides. Limit of detection, recovery, precision, and linearity were thoroughly evaluated in four representative viral transport media (VTM) containing distinct total protein content. The protocol was sensitive in all swab media with limit of detection determined at 2 × 103 pfu/mL, corresponding to as low as 30 pfu injected into the LC-MS/MS system. When tested on VTM-stored nasopharyngeal swab samples from positive and control patients, sensitivity was similar to or better than rapid immunoassay dipsticks, revealing a corresponding RT-PCR detection threshold at Ct ∼ 24. The study represents the first thorough evaluation of sensitivity and robustness of targeted mass spectrometry in nasal swabs, constituting a promising SARS-CoV-2 antigen assay for the first-line diagnosis of COVID-19 and compatible with the constraints of clinical settings. The raw files generated in this study can be found on PASSEL (Peptide Atlas) under data set identifier PASS01646.

Investigation of Inherited Chromosomally Integrated Human Herpesvirus-6A+ and -6B+ in a Patient with Ulipristal Acetate-Induced Fulminant Hepatic Failure.

Inherited chromosomally integrated (ici) human herpes virus 6 (HHV-6) is estimated to occur in 0.6-2.7% of people worldwide. HHV-6 comprises two distinct species: HHV-6A and HHV-6B. Both HHV-6A and HHV-6B integration have been reported. Several drugs are capable of activating iciHHV-6 in tissues, the consequences of which are poorly understood. We report herein a case of a woman with iciHHV-6A+ and iciHHV-6B+, who developed ulipristal acetate (a selective progesterone receptor modulator)-induced fulminant hepatic failure that required liver transplantation. We confirmed the presence of ~one copy per cell of both HHV-6A and HHV-6B DNA in her hair follicles using multiplex HHV-6A/B real-time PCR and demonstrated the Mendelian inheritance of both iciHHV-6A and iciHHV-6B in her family members over three generations. Because of the rarity of this presentation, we discuss herein the possible links between reactivated HHV-6 from iciHHV-6A and/or iciHHV-6B and adverse drug reactions, suggesting that iciHHV-6 could be screened before the introduction of any hepatotoxic drugs to exclude HHV-6 active disease or combined idiosyncratic drug-induced liver injury in these patients.

Evaluating 10 Commercially Available SARS-CoV-2 Rapid Serological Tests by Use of the STARD (Standards for Reporting of Diagnostic Accuracy Studies) Method.

Numerous severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) rapid serological tests have been developed, but their accuracy has usually been assessed using very few samples, and rigorous comparisons between these tests are scarce. In this study, we evaluated and compared 10 commercially available SARS-CoV-2 rapid serological tests using the STARD (Standards for Reporting of Diagnostic Accuracy Studies) methodology. Two hundred fifty serum samples from 159 PCR-confirmed SARS-CoV-2 patients (collected 0 to 32 days after the onset of symptoms) were tested with rapid serological tests. Control serum samples (n = 254) were retrieved from pre-coronavirus disease (COVID) periods from patients with other coronavirus infections (n = 11), positivity for rheumatoid factors (n = 3), IgG/IgM hyperglobulinemia (n = 9), malaria (n = 5), or no documented viral infection (n = 226). All samples were tested using rapid lateral flow immunoassays (LFIAs) from 10 manufacturers. Only four tests achieved ≥98% specificity, with the specificities ranging from 75.7% to 99.2%. The sensitivities varied by the day of sample collection after the onset of symptoms, from 31.7% to 55.4% (days 0 to 9), 65.9% to 92.9% (days 10 to 14), and 81.0% to 95.2% (>14 days). Only three of the tests evaluated met French health authorities' thresholds for SARS-CoV-2 serological tests (≥90% sensitivity and ≥98% specificity). Overall, the performances varied greatly between tests, with only one-third meeting acceptable specificity and sensitivity thresholds. Knowledge of the analytical performances of these tests will allow clinicians and, most importantly, laboratorians to use them with more confidence; could help determine the general population's immunological status; and may help diagnose some patients with false-negative real-time reverse transcription-PCR (RT-PCR) results.