The healthy donor effect and survey participation, becoming a donor and donor career.

BACKGROUND: The healthy donor effect (HDE) is a selection bias caused by the health criteria blood donors must meet. It obscures investigations of beneficial/adverse health effects of blood donation and complicates the generalizability of findings from blood donor cohorts. To further characterize the HDE we investigated how self-reported health and lifestyle are associated with becoming a blood donor, lapsing, and donation intensity. Furthermore, we examined differences in mortality based on donor status. STUDY DESIGN AND METHODS: The Danish National Health Survey was linked to the Scandinavian Donations and Transfusions (SCANDAT) database and Danish register data. Logistic- and normal regression was used to compare baseline characteristics and participation. Poisson regression was used to investigate future donation choices. Donation intensity was explored by the Anderson-Gill model and Poisson regression. Mortality was investigated using Poisson regression. RESULTS: Blood donors were more likely to participate in the surveys, OR = 2.45 95% confidence interval (2.40-2.49) than non-donors. Among survey participants, better self-reported health and healthier lifestyle were associated with being or becoming a blood donor, donor retention, and to some extent donation intensity, for example, current smoking conveyed lower likelihood of becoming a donor, OR = 0.70 (0.66-0.75). We observed lower mortality for donors and survey participants, respectively, compared with non-participating non-donors. CONCLUSION: We provide evidence that blood donation is associated with increased likelihood to participate in health surveys, possibly a manifestation of the HDE. Furthermore, becoming a blood donor, donor retention, and donation intensity was associated with better self-reported health and healthier lifestyles.

A large cohort study of the effects of Lewis, ABO, 13 other blood groups, and secretor status on COVID-19 susceptibility, severity, and long COVID-19.

BACKGROUND: Previous studies have reported Blood type O to confer a lower risk of SARS-CoV-2 infection, while secretor status and other blood groups have been suspected to have a similar effect as well. STUDY DESIGN AND METHODS: To determine whether any other blood groups influence testing positive for SARS-CoV-2, COVID-19 severity, or prolonged COVID-19, we used a large cohort of 650,156 Danish blood donors with varying available data for secretor status and blood groups ABO, Rh, Colton, Duffy, Diego, Dombrock, Kell, Kidd, Knops, Lewis, Lutheran, MNS, P1PK, Vel, and Yt. Of these, 36,068 tested positive for SARS-CoV-2 whereas 614,088 tested negative between 2020-02-17 and 2021-08-04. Associations between infection and blood groups were assessed using logistic regression models with sex and age as covariates. RESULTS: The Lewis blood group antigen Lea displayed strongly reduced SARS-CoV-2 susceptibility OR 0.85 CI[0.79-0.93] p < .001. Compared to blood type O, the blood types B, A, and AB were found more susceptible toward infection with ORs 1.1 CI[1.06-1.14] p < .001, 1.17 CI[1.14-1.2] p < .001, and 1.2 CI[1.14-1.26] p < .001, respectively. No susceptibility associations were found for the other 13 blood groups investigated. There was no association between any blood groups and COVID-19 hospitalization or long COVID-19. No secretor status associations were found. DISCUSSION: This study uncovers a new association to reduced SARS-CoV-2 susceptibility for Lewis type Lea and confirms the previous link to blood group O. The new association to Lea could be explained by a link between mucosal microbiome and SARS-CoV-2.

Genetic prediction of 33 blood group phenotypes using an existing genotype dataset.

BACKGROUND: Accurate blood type data are essential for blood bank management, but due to costs, few of 43 blood group systems are routinely determined in Danish blood banks. However, a more comprehensive dataset of blood types is useful in scenarios such as rare blood type allocation. We aimed to investigate the viability and accuracy of predicting blood types by leveraging an existing dataset of imputed genotypes for two cohorts of approximately 90,000 each (Danish Blood Donor Study and Copenhagen Biobank) and present a more comprehensive overview of blood types for our Danish donor cohort. STUDY DESIGN AND METHODS: Blood types were predicted from genome array data using known variant determinants. Prediction accuracy was confirmed by comparing with preexisting serological blood types. The Vel blood group was used to test the viability of using genetic prediction to narrow down the list of candidate donors with rare blood types. RESULTS: Predicted phenotypes showed a high balanced accuracy >99.5% in most cases: A, B, C/c, Coa /Cob , Doa /Dob , E/e, Jka /Jkb , Kna /Knb , Kpa /Kpb , M/N, S/s, Sda , Se, and Yta /Ytb , while some performed slightly worse: Fya /Fyb , K/k, Lua /Lub , and Vel ~99%-98% and CW and P1 ~96%. Genetic prediction identified 70 potential Vel negatives in our cohort, 64 of whom were confirmed correct using polymerase chain reaction (negative predictive value: 91.5%). DISCUSSION: High genetic prediction accuracy in most blood groups demonstrated the viability of generating blood types using preexisting genotype data at no cost and successfully narrowed the pool of potential individuals with the rare Vel-negative phenotype from 180,000 to 70.

suPARnostic: an advanced predictive tool for detecting recurrence in renal cell carcinoma.

BACKGROUND: Plasma soluble urokinase-type Plasminogen Activator Receptor (suPAR) predicts disease aggressiveness in renal cell carcinoma (ccRCC), but its prognostic accuracy has not been investigated. To investigate the prognostic accuracy of preoperative plasma suPAR in patients who received curative treatment for initially localized ccRCC. METHODS: We retrospectively analyzed plasma samples stored in the Danish National Biobank between 2010 and 2015 from 235 patients with ccRCC at any stage. Relationships with outcome analyzed using univariate and multiple logistic Cox regression analysis. RESULTS: There were 235 patients with ccRCC. The median follow-up period was 7.7 years. In univariate analysis suPAR ≥ 6 ng/mL was significantly associated with overall survival (OS) and recurrence-free survival (RFS). Patients with elevated suPAR were more likely to recur, with a Hazard Ratio (HR) of 2.3 for RFS. In multiple logistic regression, suPAR ≥ 6 ng/mL remained a negative predictor of OS and RFS. Limitations include retrospective study design, wide confidence intervals, and tumor subtype heterogeneity bias. CONCLUSIONS: ccRCC patients with high plasma suPAR concentrations are at an elevated risk of disease recurrence and see lower OS. suPAR is a promising surveillance tool to more precisely follow up with ccRCC patients and detect future recurrences. In this study, we showed that new type of liquid marker in blood plasma, called suPAR, is associated to a higher risk of kidney cancer recurrence when elevated above 6ng/mL. We also showed suPAR to independently be able to predict patients overall and recurrence free survival in patient with any stage of kidney cancer.

Estimation of SARS-CoV-2 Infection Fatality Rate by Age and Comorbidity Status Using Antibody Screening of Blood Donors During the COVID-19 Epidemic in Denmark.

BACKGROUND: Studies presenting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection fatality rate (IFR) for healthy individuals are warranted. We estimate IFR by age and comorbidity status using data from a large serosurvey among Danish blood donors and nationwide data on coronavirus disease 2019 (COVID-19) mortality. METHODS: Danish blood donors aged 17-69 years donating blood October 2020-February 2021 were tested with a commercial SARS-CoV-2 total antibody assay. IFR was estimated for weeks 11 to 42, 2020 and week 43, 2020 to week 6, 2021, representing the first 2 waves of COVID-19 epidemic in Denmark. RESULTS: In total, 84944 blood donors were tested for antibodies. The seroprevalence was 2% in October 2020 and 7% in February 2021. Among 3898039 Danish residents aged 17-69 years, 249 deaths were recorded. The IFR was low for people <51 years without comorbidity during the 2 waves (combined IFR=3.36 per 100000 infections). The IFR was below 3‰ for people aged 61-69 years without comorbidity. IFR increased with age and comorbidity but declined from the first to second wave. CONCLUSIONS: In this nationwide study, the IFR was very low among people <51 years without comorbidity.

Hyperhidrosis and human leucocyte antigens in the Danish Blood Donor Study.

Familial clustering of the skin disease primary hyperhidrosis suggests a genetic component to the disease. The human leucocyte antigen (HLA) is implicated in a range of diseases, including many comorbidities to hyperhidrosis. No study has investigated whether the HLA genes are involved in the pathogenesis of hyperhidrosis. We, therefore, compared HLA alleles in individuals with and without hyperhidrosis in this study of 65 000 blood donors. In this retrospective cohort study, we retrieved information on individuals with and without hyperhidrosis using self-reported questionnaires, the Danish National Patient Registry and the Danish National Prescription Registry on participants recruited to the Danish Blood Donor Study between 2010 and 2019. Association tests using logistic regression were conducted for each HLA allele corrected for sex, age, body mass index, smoking and principal components. Overall, 145 of 65 795 (0.2%) participants had hospital diagnosed hyperhidrosis. Similarly, 1379 of 15 530 (8.9%) participants had moderate-severe self-reported hyperhidrosis, of whom 447 (2.9%) had severe self-reported hyperhidrosis. Altogether, 28 participants had both hospital diagnosed and moderate-severe self-reported hyperhidrosis. Severe self-reported hyperhidrosis was associated with HLA-A*80:01 (adjusted odds ratio 26.97; 95% confidence interval 5.32-136.70; n = 7; P < .001). Moderate-severe self-reported hyperhidrosis and hospital diagnosed hyperhidrosis were not associated with any HLA. The association between hyperhidrosis and HLA-A*80:01 was based on a very small number of cases and not replicated in other patient subsets, and therefore likely a chance finding. Thus, this study suggests that genes other than the HLA are involved in the pathogenesis of hyperhidrosis.

Seroprevalence of SARS-CoV-2 antibodies in social housing areas in Denmark.

BACKGROUND: COVID-19 is thought to be more prevalent among ethnic minorities and individuals with low socioeconomic status. We aimed to investigate the prevalence of SARS-CoV-2 antibodies during the COVID-19 pandemic among citizens 15 years or older in Denmark living in social housing (SH) areas. METHODS: We conducted a study between January 8th and January 31st, 2021 with recruitment in 13 selected SH areas. Participants were offered a point-of-care rapid SARS-CoV-2 IgM and IgG antibody test and a questionnaire concerning risk factors associated with COVID-19. As a proxy for the general Danish population we accessed data on seroprevalence from Danish blood donors (total Ig ELISA assay) in same time period. RESULTS: Of the 13,279 included participants, 2296 (17.3%) were seropositive (mean age 46.6 (SD 16.4) years, 54.2% female), which was 3 times higher than in the general Danish population (mean age 41.7 (SD 14.1) years, 48.5% female) in the same period (5.8%, risk ratios (RR) 2.96, 95% CI 2.78-3.16, p > 0.001). Seropositivity was higher among males (RR 1.1, 95% CI 1.05-1.22%, p = 0.001) and increased with age, with an OR seropositivity of 1.03 for each 10-year increase in age (95% CI 1.00-1.06, p = 0.031). Close contact with COVID-19-infected individuals was associated with a higher risk of infection, especially among household members (OR 5.0, 95% CI 4.1-6.2 p < 0,001). Living at least four people in a household significantly increased the OR of seropositivity (OR 1.3, 95% CI 1.0-1.6, p = 0.02) as did living in a multi-generational household (OR 1.3 per generation, 95% CI 1.1-1.6, p = 0.003). Only 1.6% of participants reported not following any of the national COVID-19 recommendations. CONCLUSIONS: Danish citizens living in SH areas of low socioeconomic status had a three times higher SARS-CoV-2 seroprevalence compared to the general Danish population. The seroprevalence was significantly higher in males and increased slightly with age. Living in multiple generations households or in households of more than four persons was a strong risk factor for being seropositive. Results of this study can be used for future consideration of the need for preventive measures in the populations living in SH areas.

SARS-CoV-2 antibody prevalence among homeless people and shelter workers in Denmark: a nationwide cross-sectional study.

BACKGROUND: People experiencing homelessness (PEH) and associated shelter workers may be at higher risk of infection with "Severe acute respiratory syndrome coronavirus 2" (SARS-CoV-2). The aim of this study was to determine the prevalence of SARS-CoV-2 among PEH and shelter workers in Denmark. DESIGN AND METHODS: In November 2020, we conducted a nationwide cross-sectional seroprevalence study among PEH and shelter workers at 21 recruitment sites in Denmark. The assessment included a point-of-care test for antibodies against SARS-CoV-2, followed by a questionnaire. The seroprevalence was compared to that of geographically matched blood donors considered as a proxy for the background population, tested using a total Ig ELISA assay. RESULTS: We included 827 participants in the study, of whom 819 provided their SARS-CoV-2 antibody results. Of those, 628 were PEH (median age 50.8 (IQR 40.9-59.1) years, 35.5% female) and 191 were shelter workers (median age 46.6 (IQR 36.1-55.0) years and 74.5% female). The overall seroprevalence was 6.7% and was similar among PEH and shelter workers (6.8% vs 6.3%, p = 0.87); and 12.2% among all participants who engaged in sex work. The overall participant seroprevalence was significantly higher than that of the background population (2.9%, p < 0.001). When combining all participants who reported sex work or were recruited at designated safe havens, we found a significantly increased risk of seropositivity compared to other participants (OR 2.23, 95%CI 1.06-4.43, p = 0.02). Seropositive and seronegative participants reported a similar presence of at least one SARS-CoV-2 associated symptom (49% and 54%, respectively). INTERPRETATIONS: The prevalence of SARS-CoV-2 antibodies was more than twice as high among PEH and associated shelter workers, compared to the background population. These results could be taken into consideration when deciding in which phase PEH are eligible for a vaccine, as part of the Danish national SARS-CoV-2 vaccination program rollout. FUNDING: TrygFonden and HelseFonden.

Estimation of SARS-CoV-2 Infection Fatality Rate by Real-time Antibody Screening of Blood Donors.

BACKGROUND: The pandemic due to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has tremendous consequences for our societies. Knowledge of the seroprevalence of SARS-CoV-2 is needed to accurately monitor the spread of the epidemic and to calculate the infection fatality rate (IFR). These measures may help the authorities make informed decisions and adjust the current societal interventions. The objective was to perform nationwide real-time seroprevalence surveying among blood donors as a tool to estimate previous SARS-CoV-2 infections and the population-based IFR. METHODS: Danish blood donors aged 17-69 years giving blood 6 April to 3 May were tested for SARS-CoV-2 immunoglobulin M and G antibodies using a commercial lateral flow test. Antibody status was compared between geographical areas, and an estimate of the IFR was calculated. Seroprevalence was adjusted for assay sensitivity and specificity taking the uncertainties of the test validation into account when reporting the 95% confidence intervals (CIs). RESULTS: The first 20 640 blood donors were tested, and a combined adjusted seroprevalence of 1.9% (95% CI, .8-2.3) was calculated. The seroprevalence differed across areas. Using available data on fatalities and population numbers, a combined IFR in patients <70 years is estimated at 89 per 100 000 (95% CI, 72-211) infections. CONCLUSIONS: The IFR was estimated to be slightly lower than previously reported from other countries not using seroprevalence data. The IFR is likely severalfold lower than the current estimate. We have initiated real-time nationwide anti-SARS-CoV-2 seroprevalence surveying of blood donations as a tool in monitoring the epidemic.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Infection Fatality Rate Among Elderly Danes: A Cross-sectional Study on Retired Blood Donors.

BACKGROUND: Although the vast majority of individuals succumbing to infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are elderly, infection fatality rate (IFR) estimates for the age group ≥70 years are still scarce. To this end, we assessed SARS-CoV-2 seroprevalence among retired blood donors and combined it with national coronavirus disease 2019 (COVID-19) survey data to provide reliable population-based IFR estimates for this age group. METHODS: We identified 60 926 retired blood donors aged ≥70 years in the rosters of 3 regionwide Danish blood banks and invited them to fill in a questionnaire on COVID-19-related symptoms and behaviors. Among 24 861 (40.8%) responders, we invited a random sample of 3200 individuals for blood testing. Overall, 1201 (37.5%) individuals were tested for SARS-CoV-2 antibodies (Wantai) and compared with 1110 active blood donors aged 17-69 years. Seroprevalence 95% confidence intervals (CIs) were adjusted for assay sensitivity and specificity. RESULTS: Among retired (aged ≥70 years) and active (aged 17-69 years) blood donors, adjusted seroprevalences were 1.4% (95% CI, .3-2.5%) and 2.5% (95% CI, 1.3-3.8%), respectively. Using available population data on COVID-19-related fatalities, IFRs for patients aged ≥70 years and for 17-69 years were estimated at 5.4% (95% CI, 2.7-6.4%) and .083% (95% CI, .054-.18%), respectively. Only 52.4% of SARS-CoV-2-seropositive retired blood donors reported having been sick since the start of the pandemic. CONCLUSIONS: COVID-19 IFR in the age group >69 years is estimated to be 65 times the IFR for people aged 18-69 years.

Peripheral blood immune markers in breast cancer: Differences in regulatory T cell abundance are related to clinical parameters.

BACKGROUND: Cancer development is among other factors driven by tumor immune escape and tumor-mediated changes in the immune response. Investigating systemic immune changes may provide important knowledge for the improvement of patient prognosis and treatment opportunities. METHODS: The systemic immune profile of patients with ER-positive breast cancer (n = 22) and healthy controls (n = 30) was investigated based on complete blood counts, flow cytometric analysis of T cell subsets including regulatory T cells (Tregs), and immune assays investigating soluble (s)HLA-G and the cytokine profile in plasma. We further examined the correlation between the immune markers and clinical parameters including tumor size, tumor grade and lymph node involvement. RESULTS: Results indicated that breast cancer patients possessed a higher amount of neutrophils and monocytes and fewer lymphocytes and eosinophils compared with healthy controls. Breast cancer patients had significantly more CD25+CD127low Tregs than controls, and both lymphocyte and Treg numbers were negatively correlated with tumor size. Furthermore, Treg numbers were elevated in grade I tumors compared with grade II tumors and with healthy controls. No difference in sHLA-G levels was observed between patients and controls. Higher levels of IL-6 and TNF-α were observed in breast cancer patients. Cytokine and sHLA-G levels were not associated with clinical parameters. CONCLUSION: The results of this exploratory study contribute to the elucidation of the systemic immune response in breast cancer indicating a potential use of peripheral immune cell counts and Tregs to distinguish patients from healthy controls and as potential diagnostic and prognostic biomarkers to be investigated in future studies.

Comparison of 16 Serological SARS-CoV-2 Immunoassays in 16 Clinical Laboratories.

Serological assays for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are needed to support clinical diagnosis and epidemiological investigations. Recently, assays for large-scale detection of total antibodies (Ab), immunoglobulin G (IgG), and IgM against SARS-CoV-2 antigens have been developed, but there are limited data on the diagnostic accuracy of these assays. This study was a Danish national collaboration and evaluated 15 commercial and one in-house anti-SARS-CoV-2 assays in 16 laboratories. Sensitivity was evaluated using 150 samples from individuals with asymptomatic, mild, or moderate COVID-19, nonhospitalized or hospitalized, confirmed by nucleic acid amplification tests (NAAT); samples were collected 13 to 73 days either from symptom onset or from positive NAAT (patients without symptoms). Specificity and cross-reactivity were evaluated in samples collected prior to the SARS-CoV-2 epidemic from >586 blood donors and patients with autoimmune diseases, cytomegalovirus or Epstein-Barr virus infections, and acute viral infections. A specificity of ≥99% was achieved by all total-Ab and IgG assays except one, DiaSorin Liaison XL IgG (97.2%). Sensitivities in descending order were Wantai ELISA total Ab (96.7%), CUH-NOVO in-house ELISA total Ab (96.0%), Ortho Vitros total Ab (95.3%), YHLO iFlash IgG (94.0%), Ortho Vitros IgG (93.3%), Siemens Atellica total Ab (93.2%), Roche Elecsys total Ab (92.7%), Abbott Architect IgG (90.0%), Abbott Alinity IgG (median 88.0%), DiaSorin Liaison XL IgG (median 84.6%), Siemens Vista total Ab (81.0%), Euroimmun/ELISA IgG (78.0%), and Snibe Maglumi IgG (median 78.0%). However, confidence intervals overlapped for several assays. The IgM results were variable, with the Wantai IgM ELISA showing the highest sensitivity (82.7%) and specificity (99%). The rate of seropositivity increased with time from symptom onset and symptom severity.

Low transfusion transmission of hepatitis E among 25,637 single-donation, nucleic acid-tested blood donors.

BACKGROUND: Hepatitis E virus genotype-3 (HEV-gt-3) causes autochthonous infections in western countries, with a primary reservoir in animals, especially pigs. HEV transfusion transmission has been reported, and HEV-gt-3 prevalence is high in some European countries. The prevalence of HEV RNA was investigated among Danish blood donors, and the prevalence of HEV transfusion-transmitted infection (TTI) was investigated among recipients. STUDY DESIGN AND METHODS: Samples from 25,637 consenting donors collected during 1 month in 2015 were screened retrospectively using an individual-donation HEV RNA nucleic acid test with a 95% detection probability of 7.9 IU/mL. HEV-positive samples were quantified by real-time polymerase chain reaction and genotyped. Transmission was evaluated among recipients of HEV RNA-positive blood components. Phylogenetic analyses compared HEV sequences from blood donors, symptomatic patients, and swine. RESULTS: Eleven donations (0.04%) were confirmed as positive for HEV RNA (median HEV RNA level, 13 IU/mL). Two donations were successfully genotyped as HEV-gt-3. Only one donor had a travel history outside Europe. Nine of 11 donors were male, but the gender ratio was nonsignificant compared with the total donor population. Seven available recipients tested negative for HEV RNA and anti-HEV immunoglobulin M in follow-up samples. One recipient was HEV RNA-negative but anti-HEV immunoglobulin G-positive. HEV TTI was considered unlikely, but a transfusion-induced secondary immune response could not be excluded. Phylogenetic analysis showed relatively large sequence differences between HEV from donors, symptomatic patients, and swine. CONCLUSIONS: Despite an HEV RNA prevalence of 0.04% in Danish blood donations, all HEV-positive donations carried low viral loads, and no evidence of TTI was found.

Two Novel Human Leukocyte Antigen Alleles Are Associated with Decreased Risk of Onychomycosis in a Large Cohort of Danish Blood Donors.

Introduction: Antigen presentation and antimicrobial immune responses involve the human leukocyte antigen (HLA) system. Onychomycosis is primarily caused by dermatophytes and affects around 5.5% of the population worldwide. Yet, only limited data exist on the associations between the HLA system and onychomycosis. Thus, the objective of the study was to investigate if there is an association between HLA alleles and onychomycosis. Methods: Participants in the Danish Blood Donor Study were defined as cases of onychomycosis and controls based on antifungal prescriptions in the national prescription registry. Associations were investigated using logistic regressions adjusted for confounders and were Bonferroni corrected for multiple tests. Results: A total of 3,665 participants were considered onychomycosis cases, and 24,144 participants were considered controls. We found two protective HLA alleles of onychomycosis: DQB1*06:04, odds ratios (OR) 0.80 (95% confidence interval (CI) 0.71-0.90), and DRB1*13:02, OR 0.79 (95% CI: 0.71-0.89). Conclusion: The finding of two novel protective alleles of onychomycosis indicates that certain HLA alleles have certain antigen presentation properties affecting the risk of fungal infection. These findings may provide the basis for future research identifying immunologically relevant antigens of fungi causing onychomycosis, which could ultimately lead to targets of new drugs with antifungal effects.

Experience of loneliness during the COVID-19 pandemic: a cross-sectional study of 50 968 adult Danes.

OBJECTIVES: To examine the level of loneliness experienced during the COVID-19 pandemic in Denmark and to identify associated behavioural patterns and demographic factors. DESIGN: Cross-sectional cohort study. SETTING: Includes Danish active and former blood donors. PARTICIPANTS: A questionnaire was sent to 124 307 active and former blood donors, of these a total of 50 968 participants completed the study questionnaire (response rate=41%). PRIMARY AND SECONDARY OUTCOME MEASURES: Subjective experience of loneliness was measured using the 3-item University of California, Los Angeles Loneliness Scale (UCLA-3). Besides the UCLA-3, the respondents answered items on sociodemographic and economic characteristics, items on precautionary measures taken to avoid COVID-19 infection as well as on COVID-19 anxiety. RESULTS: The participants indicated their experienced level of loneliness both before and during the pandemic. Comparing the two reports yielded a mean increase in loneliness scores of 14.1% (p<0.001). Exploratory factor analysis identified the factor well-being, which comprised three questionnaire items related to emotional heath, physical health and happiness. A high score on the factor well-being was associated with reduced levels of loneliness (coefficient=-0.47, 95% CI -0.49 to -0.46)). Furthermore, women were more likely than men to have experienced increased levels of loneliness during the pandemic (coefficient=0.27, 95% CI 0.25 to 0.29). Furthermore, a negative correlation between higher age and change in loneliness score was observed. CONCLUSIONS: The findings document an increase in the level of experienced loneliness during the COVID-19 pandemic, particularly affecting individuals with low well-being, women and younger individuals.

Effect of anterior quadratus lumborum block with ropivacaine on the immune response after laparoscopic surgery in colon cancer: a substudy of a randomized clinical trial.

BACKGROUND: Surgery induces a temporal change in the immune system, which might be modified by regional anesthesia. Applying a bilateral preoperative anterior quadratus lumborum block has proven to be a safe and effective technique in pain management after abdominal and retroperitoneal surgery, but the effect on the immune response is not thoroughly investigated. METHODS: This study is a substudy of a randomized, controlled, double-blinded trial of patients undergoing laparoscopic hemicolectomy due to colon cancer. Twenty-two patients were randomized to undergo either a bilateral anterior quadratus lumborum nerve block with a total of 60 mL ropivacaine 0.375% or placebo with corresponding isotonic saline injections. The main objective of this exploratory substudy was to investigate the systemic immune response in the first postoperative day by examining changes in blood transcript levels (n=750) and stimulated secretion of cytokines (n=17) on ex vivo activation with microbial ligands and anti-CD3/CD28. RESULTS: Using unsupervised data analysis tools, we observed no effect of the bilateral anterior quadratus lumborum nerve block on gene expression in immune cells (permutational multivariate analysis of variance using distance matrices: F=0.52, p=0.96), abundances of major immune cell populations (Wilcoxon rank-sum test: p>0.05), and stimulated cytokine secretion (Wilcoxon rank-sum test: p>0.05). CONCLUSIONS: Our study provides evidence that administration of bilateral anterior quadratus lumborum nerve block as a part of a multimodal analgesic regimen in an enhanced recovery after surgery for laparoscopic hemicolectomy in this cohort does not alter the systemic immune response.Trial registration number NCT03570541.

Neoadjuvant intratumoral influenza vaccine treatment in patients with proficient mismatch repair colorectal cancer leads to increased tumor infiltration of CD8+ T cells and upregulation of PD-L1: a phase 1/2 clinical trial.

BACKGROUND: In colorectal cancer, the effects of immune checkpoint inhibitors are mostly limited to patients with deficient mismatch repair tumors, characterized by a high grade infiltration of CD8+T cells. Interventions aimed at increasing intratumoral CD8+T-cell infiltration in proficient mismatch repair tumors are lacking. METHODS: We conducted a proof of concept phase 1/2 clinical trial, where patients with non-metastasizing sigmoid or rectal cancer, scheduled for curative intended surgery, were treated with an endoscopic intratumorally administered neoadjuvant influenza vaccine. Blood and tumor samples were collected before the injection and at the time of surgery. The primary outcome was safety of the intervention. Evaluation of pathological tumor regression grade, immunohistochemistry, flow cytometry of blood, tissue bulk transcriptional analyses, and spatial protein profiling of tumor regions were all secondary outcomes. RESULTS: A total of 10 patients were included in the trial. Median patient age was 70 years (range 54-78), with 30% women. All patients had proficient mismatch repair Union of International Cancer Control stage I-III tumors. No endoscopic safety events occurred, with all patients undergoing curative surgery as scheduled (median 9 days after intervention). Increased CD8+T-cell tumor infiltration was evident after vaccination (median 73 vs 315 cells/mm2, p<0.05), along with significant downregulation of messenger RNA gene expression related to neutrophils and upregulation of transcripts encoding cytotoxic functions. Spatial protein analysis showed significant local upregulation of programmed death-ligand 1 (PD-L1) (adjusted p value<0.05) and downregulation of FOXP3 (adjusted p value<0.05). CONCLUSIONS: Neoadjuvant intratumoral influenza vaccine treatment in this cohort was demonstrated to be safe and feasible, and to induce CD8+T-cell infiltration and upregulation of PD-L1 proficient mismatch repair sigmoid and rectal tumors. Definitive conclusions regarding safety and efficacy can only be made in larger cohorts. TRIAL REGISTRATION NUMBER: NCT04591379.

A Cross-Sectional Study of SARS-CoV-2 Antibodies and Risk Factors for Seropositivity in Staff in Day Care Facilities and Preschools in Denmark.

The aim of this study was to provide information about immunity against COVID-19 along with risk factors and behavior among employees in day care facilities and preschools (DCS) in Denmark. In collaboration with the Danish Union of Pedagogues, during February and March 2021, 47,810 members were offered a point-of-care rapid SARS-CoV-2 antibody test (POCT) at work and were invited to fill in an electronic questionnaire covering COVID-19 exposure. Seroprevalence data from Danish blood donors (total Ig enzyme-linked immunosorbent assay [ELISA]) were used as a proxy for the Danish population. A total of 21,018 (45%) DCS employees completed the questionnaire and reported their POCT result {median age, 44.3 years (interquartile range [IQR], [32.7 to 53.6]); females, 84.1%}, of which 20,267 (96.4%) were unvaccinated and included in analysis. A total of 1,857 (9.2%) participants tested seropositive, significantly higher than a seroprevalence at 7.6% (risk ratio [RR], 1.2; 95% confidence interval [CI], 1.14 to 1.27) among 40,541 healthy blood donors (median age, 42 years [IQR, 28 to 53]; males, 51.3%). Exposure at work (RR, 2.9; 95% CI, 2.3 to 3.6) was less of a risk factor than exposure within the household (RR, 12.7; 95% CI, 10.2 to 15.8). Less than 25% of participants reported wearing face protection at work. Most of the participants expressed some degree of fear of contracting COVID-19 both at work and outside work. SARS-CoV-2 seroprevalence was slightly higher in DCS staff than in blood donors, but possible exposure at home was associated with a higher risk than at work. DCS staff expressed fear of contracting COVID-19, though there was limited use of face protection at work. IMPORTANCE Identifying at-risk groups and evaluating preventive interventions in at-risk groups is imperative for the ongoing pandemic as well as for the control of future epidemics. Although DCS staff have a much higher risk of being infected within their own household than at their workplace, most are fearful of being infected with COVID-19 or bringing COVID-19 to work. This represents an interesting dilemma and an important issue which should be addressed by public health authorities for risk communication and pandemic planning. This study design can be used in a strategy for ongoing surveillance of COVID-19 immunity or other infections in the population. The findings of this study can be used to assess the need for future preventive interventions in DCS, such as the use of personal protective equipment.

Optimization and evaluation of a live virus SARS-CoV-2 neutralization assay.

Virus neutralization assays provide a means to quantitate functional antibody responses that block virus infection. These assays are instrumental in defining vaccine and therapeutic antibody potency, immune evasion by viral variants, and post-infection immunity. Here we describe the development, optimization and evaluation of a live virus microneutralization assay specific for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In this assay, SARS-CoV-2 clinical isolates are pre-incubated with serial diluted antibody and added to Vero E6 cells. Replicating virus is quantitated by enzyme-linked immunosorbent assay (ELISA) targeting the SARS-CoV-2 nucleocapsid protein and the standardized 50% virus inhibition titer calculated. We evaluated critical test parameters that include virus titration, assay linearity, number of cells, viral dose, incubation period post-inoculation, and normalization methods. Virus titration at 96 hours was determined optimal to account for different growth kinetics of clinical isolates. Nucleocapsid protein levels directly correlated with virus inoculum, with the strongest correlation at 24 hours post-inoculation. Variance was minimized by infecting a cell monolayer, rather than a cell suspension. Neutralization titers modestly decreased with increasing numbers of Vero E6 cells and virus amount. Application of two different normalization models effectively reduced the intermediate precision coefficient of variance to <16.5%. The SARS-CoV-2 microneutralization assay described and evaluated here is based on the influenza virus microneutralization assay described by WHO, and are proposed as a standard assay for comparing neutralization investigations.