Design and validation of Dolosigranulum pigrum specific PCR primers using the bacterial core genome.

Dolosigranulum pigrum-a lactic acid bacterium that is increasingly recognized as an important member of the nasal microbiome. Currently, there are limited rapid and low-cost options for confirming D. pigrum isolates and detecting D. pigrum in clinical specimens. Here we describe the design and validation of a novel PCR assay targeting D. pigrum that is both sensitive and specific. We designed a PCR assay targeting murJ, a single-copy core species gene identified through the analysis of 21 D. pigrum whole genome sequences. The assay achieved 100% sensitivity and 100% specificity against D. pigrum and diverse bacterial isolates and an overall 91.1% sensitivity and 100% specificity using nasal swabs, detecting D. pigrum at a threshold of 1.0 × 104 D. pigrum 16S rRNA gene copies per swab. This assay adds a reliable and rapid D. pigrum detection tool to the microbiome researcher toolkit investigating the role of generalist and specialist bacteria in the nasal environment.

Experience with sotrovimab treatment of SARS-CoV-2-infected patients in Denmark.

AIMS: To evaluate the experience with use of sotrovimab following severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection in high-risk groups. METHODS: In a nationwide, population-based cohort study, we identified all individuals treated with sotrovimab (N = 2933) and stratified them by 4 high-risk groups: (A) malignant haematological disease, (B) solid organ transplantation, (C) anti-CD20 therapy ≤1 year and (D) other risks. Cox regression analysis was used to calculate hazard ratios for hospitalization, death and associated prognostic factors. RESULTS: Of 2933 sotrovimab-treated individuals, 83% belonged to high-risk groups (37.6% haematological malignancy, 27.4% solid organ transplantation and 17.5% treatment with anti-CD20 ≤1 year). Only 17.8% had other risks (11.8% were pregnant, 10.7% primary immunodeficiency, 21.2% other malignancy, 4.3% received anti-CD20 >1 year and 52.0% other/unknown causes). Within 90 days of infusion, 30.2% were hospitalized and 5.3% died. The main prognostic factors were the predefined high-risk groups, mainly malignant haematological disease and age ≥65 years. Number of COVID-19 vaccines (≥3) was associated with a decreased risk of hospitalization. The Delta but not the Omicron BA.2 variant was associated with a higher risk of death compared to the BA.1 variant. CONCLUSION: More than 90% of the patients treated with sotrovimab belonged to the very high-risk groups as described in the Danish guidelines. Sotrovimab-treated individuals remained at a high risk of hospitalization and death which was strongly associated with the underlying immunocompromised state and age. Having received >3 COVID-19 vaccines was association with decreased risk of death and hospitalization.

Risk of reinfection, vaccine protection, and severity of infection with the BA.5 omicron subvariant: a nation-wide population-based study in Denmark.

BACKGROUND: Estimates of immunity and severity for the SARS-CoV-2 omicron subvariant BA.5 are important to assess the public health impact associated with its rapid global spread despite vaccination. We estimated natural and vaccine immunity and severity of BA.5 relative to BA.2 in Denmark, a country with high mRNA-vaccination coverage and free-of-charge RT-PCR testing. METHODS: This nation-wide population-based study in Denmark included residents aged 18 years or older who had taken an RT-PCR test between 10 April and 30 June, 2022 (ie, the outcome period), and who the national COVID-19 surveillance system identified as having information since February 2020 on RT-PCR tests, whole-genome sequencing, vaccinations, and hospitalisation with a positive RT-PCR test and COVID-19 as the main diagnosis. First, we used a case-control design, in which cases were people infected with BA.5 or BA.2 during the outcome period and controls were people who tested negative for SARS-CoV-2 infection during the outcome period. We calculated the protection provided by a previous PCR-confirmed omicron infection against BA.5 and BA.2 infection and hospitalisation among triple-vaccinated individuals. Second, we compared vaccination status in people infected with BA.5 versus BA.2 and estimated relative vaccine protection against each subvariant. Third, we compared rates of hospitalisation for COVID-19 among people infected with BA.5 versus BA.2. We estimated effects using logistic regression with adjustment for sex, age, region, PCR-test date, comorbidity and, as appropriate, vaccination and previous infection status. FINDINGS: A total of 210 (2·4%) of 8678 of BA.5 cases, 192 (0·7%) of 29 292 of BA.2 cases, and 33 972 (19·0%) of 178 669 PCR-negative controls previously had an omicron infection, which was estimated in the adjusted analyses to offer 92·7% (95% CI 91·6-93·7) protection against BA.5 infection and 97·1% (96·6-97·5) protection against BA.2 infection. We found similarly high amounts of protection against hospitalisation owing to infection with BA.5 (96·4% [95% CI 74·2-99·5]) and BA.2 (91·2% [76·3-96·7]). Vaccine coverage (three mRNA doses vs none) was 9307 (94·2%) of 9878 among BA.5 cases and 30 581 (94·8%) of 32 272 among BA.2 cases, although in the adjusted analysis, there was a trend towards slightly higher vaccination coverage among BA.5 cases than BA.2 cases (OR 1·18 [95% CI 0·99-1·42]; p=0·064), possibly suggesting marginally poorer vaccine protection against BA.5. The rate of hospitalisation due to COVID-19 was higher among the BA.5 cases (210 [1·9%] of 11 314) than among the BA.2 cases (514 [1·4%] of 36 805), with an OR of 1·34 (95% CI 1·14-1·57) and an adjusted OR of 1·69 (95% CI 1·22-2·33), despite low and stable COVID-19 hospitalisation numbers during the study period. INTERPRETATION: The study provides evidence that a previous omicron infection in triple-vaccinated individuals provides high amounts of protection against BA.5 and BA.2 infections. However, protection estimates greater than 90% might be too high if individuals with a previous infection were more likely than those without one to come forward for a test for reasons other than suspicion of COVID-19. Our analysis also showed that vaccine protection against BA.5 infection was similar to, or slightly weaker than, protection against BA.2 infection. Finally, there was evidence that BA.5 infections were associated with an increased risk of hospitalisation compared with BA.2 infections. FUNDING: There was no funding source for this study.

Risk of reinfection, vaccine protection, and severity of infection with the BA.5 omicron subvariant: a nation-wide population-based study in Denmark

Background Estimates of immunity and severity for the SARS-CoV-2 omicron subvariant BA.5 are important to assess the public health impact associated with its rapid global spread despite vaccination. We estimated natural and vaccine immunity and severity of BA.5 relative to BA.2 in Denmark, a country with high mRNA-vaccination coverage and free-of-charge RT-PCR testing. Methods This nation-wide population-based study in Denmark included residents aged 18 years or older who had taken an RT-PCR test between 10 April and 30 June, 2022 (ie, the outcome period), and who the national COVID-19 surveillance system identified as having information since February 2020 on RT-PCR tests, whole-genome sequencing, vaccinations, and hospitalisation with a positive RT-PCR test and COVID-19 as the main diagnosis. First, we used a case–control design, in which cases were people infected with BA.5 or BA.2 during the outcome period and controls were people who tested negative for SARS-CoV-2 infection during the outcome period. We calculated the protection provided by a previous PCR-confirmed omicron infection against BA.5 and BA.2 infection and hospitalisation among triple-vaccinated individuals. Second, we compared vaccination status in people infected with BA.5 versus BA.2 and estimated relative vaccine protection against each subvariant. Third, we compared rates of hospitalisation for COVID-19 among people infected with BA.5 versus BA.2. We estimated effects using logistic regression with adjustment for sex, age, region, PCR-test date, comorbidity and, as appropriate, vaccination and previous infection status. Findings A total of 210 (2·4%) of 8678 of BA.5 cases, 192 (0·7%) of 29 292 of BA.2 cases, and 33 972 (19·0%) of 178 669 PCR-negative controls previously had an omicron infection, which was estimated in the adjusted analyses to offer 92·7% (95% CI 91·6–93·7) protection against BA.5 infection and 97·1% (96·6–97·5) protection against BA.2 infection. We found similarly high amounts of protection against hospitalisation owing to infection with BA.5 (96·4% [95% CI 74·2–99·5]) and BA.2 (91·2% [76·3–96·7]). Vaccine coverage (three mRNA doses vs none) was 9307 (94·2%) of 9878 among BA.5 cases and 30 581 (94·8%) of 32 272 among BA.2 cases, although in the adjusted analysis, there was a trend towards slightly higher vaccination coverage among BA.5 cases than BA.2 cases (OR 1·18 [95% CI 0·99–1·42];p=0·064), possibly suggesting marginally poorer vaccine protection against BA.5. The rate of hospitalisation due to COVID-19 was higher among the BA.5 cases (210 [1·9%] of 11 314) than among the BA.2 cases (514 [1·4%] of 36 805), with an OR of 1·34 (95% CI 1·14–1·57) and an adjusted OR of 1·69 (95% CI 1·22–2·33), despite low and stable COVID-19 hospitalisation numbers during the study period. Interpretation The study provides evidence that a previous omicron infection in triple-vaccinated individuals provides high amounts of protection against BA.5 and BA.2 infections. However, protection estimates greater than 90% might be too high if individuals with a previous infection were more likely than those without one to come forward for a test for reasons other than suspicion of COVID-19. Our analysis also showed that vaccine protection against BA.5 infection was similar to, or slightly weaker than, protection against BA.2 infection. Finally, there was evidence that BA.5 infections were associated with an increased risk of hospitalisation compared with BA.2 infections. Funding There was no funding source for this study.

Household transmission of SARS-CoV-2 Omicron variant of concern subvariants BA.1 and BA.2 in Denmark.

SARS coronavirus 2 (SARS-CoV-2) continues to evolve and new variants emerge. Using nationwide Danish data, we estimate the transmission dynamics of SARS-CoV-2 Omicron subvariants BA.1 and BA.2 within households. Among 22,678 primary cases, we identified 17,319 secondary infections among 50,588 household contacts during a 1-7 day follow-up. The secondary attack rate (SAR) was 29% and 39% in households infected with Omicron BA.1 and BA.2, respectively. BA.2 was associated with increased susceptibility of infection for unvaccinated household contacts (Odds Ratio (OR) 1.99; 95%-CI 1.72-2.31), fully vaccinated contacts (OR 2.26; 95%-CI 1.95-2.62) and booster-vaccinated contacts (OR 2.65; 95%-CI 2.29-3.08), compared to BA.1. We also found increased infectiousness from unvaccinated primary cases infected with BA.2 compared to BA.1 (OR 2.47; 95%-CI 2.15-2.84), but not for fully vaccinated (OR 0.66; 95%-CI 0.57-0.78) or booster-vaccinated primary cases (OR 0.69; 95%-CI 0.59-0.82). Omicron BA.2 is inherently more transmissible than BA.1. Its immune-evasive properties also reduce the protective effect of vaccination against infection, but do not increase infectiousness of breakthrough infections from vaccinated individuals.

Household transmission of the SARS-CoV-2 Omicron variant in Denmark.

In late 2021, the Omicron SARS-CoV-2 variant overtook the previously dominant Delta variant, but the extent to which this transition was driven by immune evasion or a change in the inherent transmissibility is currently unclear. We estimate SARS-CoV-2 transmission within Danish households during December 2021. Among 26,675 households (8,568 with the Omicron VOC), we identified 14,140 secondary infections within a 1-7-day follow-up period. The secondary attack rate was 29% and 21% in households infected with Omicron and Delta, respectively. For Omicron, the odds of infection were 1.10 (95%-CI: 1.00-1.21) times higher for unvaccinated, 2.38 (95%-CI: 2.23-2.54) times higher for fully vaccinated and 3.20 (95%-CI: 2.67-3.83) times higher for booster-vaccinated contacts compared to Delta. We conclude that the transition from Delta to Omicron VOC was primarily driven by immune evasiveness and to a lesser extent an inherent increase in the basic transmissibility of the Omicron variant.

Vaccine effectiveness against SARS-CoV-2 infection or COVID-19 hospitalization with the Alpha, Delta, or Omicron SARS-CoV-2 variant: A nationwide Danish cohort study.

BACKGROUND: The continued occurrence of more contagious Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) variants and waning immunity over time require ongoing reevaluation of the vaccine effectiveness (VE). This study aimed to estimate the effectiveness in 2 age groups (12 to 59 and 60 years or above) of 2 or 3 vaccine doses (BNT162b2 mRNA or mRNA-1273) by time since vaccination against SARS-CoV-2 infection and Coronavirus Disease 2019 (COVID-19) hospitalization in an Alpha-, Delta-, or Omicron-dominated period. METHODS AND FINDINGS: A Danish nationwide cohort study design was used to estimate VE against SARS-CoV-2 infection and COVID-19 hospitalization with the Alpha, Delta, or Omicron variant. Information was obtained from nationwide registries and linked using a unique personal identification number. The study included all previously uninfected residents in Denmark aged 12 years or above (18 years or above for the analysis of 3 doses) in the Alpha (February 20 to June 15, 2021), Delta (July 4 to November 20, 2021), and Omicron (December 21, 2021 to January 31, 2022) dominated periods. VE estimates including 95% confidence intervals (CIs) were calculated (1-hazard ratio∙100) using Cox proportional hazard regression models with underlying calendar time and adjustments for age, sex, comorbidity, and geographical region. Vaccination status was included as a time-varying exposure. In the oldest age group, VE against infection after 2 doses was 90.7% (95% CI: 88.2; 92.7) for the Alpha variant, 82.3% (95% CI: 75.5; 87.2) for the Delta variant, and 39.9% (95% CI: 26.3; 50.9) for the Omicron variant 14 to 30 days since vaccination. The VE waned over time and was 73.2% (Alpha, 95% CI: 57.1; 83.3), 50.0% (Delta, 95% CI: 46.7; 53.0), and 4.4% (Omicron, 95% CI: -0.1; 8.7) >120 days since vaccination. Higher estimates were observed after the third dose with VE estimates against infection of 86.1% (Delta, 95% CI: 83.3; 88.4) and 57.7% (Omicron, 95% CI: 55.9; 59.5) 14 to 30 days since vaccination. Among both age groups, VE against COVID-19 hospitalization 14 to 30 days since vaccination with 2 or 3 doses was 98.1% or above for the Alpha and Delta variants. Among both age groups, VE against COVID-19 hospitalization 14 to 30 days since vaccination with 2 or 3 doses was 95.5% or above for the Omicron variant. The main limitation of this study is the nonrandomized study design including potential differences between the unvaccinated (reference group) and vaccinated individuals. CONCLUSIONS: Two vaccine doses provided high protection against SARS-CoV-2 infection and COVID-19 hospitalization with the Alpha and Delta variants with protection, notably against infection, waning over time. Two vaccine doses provided only limited and short-lived protection against SARS-CoV-2 infection with Omicron. However, the protection against COVID-19 hospitalization following Omicron SARS-CoV-2 infection was higher. The third vaccine dose substantially increased the level and duration of protection against infection with the Omicron variant and provided a high level of sustained protection against COVID-19 hospitalization among the +60-year-olds.

Introduction and transmission of SARS-CoV-2 lineage B.1.1.7, Alpha variant, in Denmark.

BACKGROUND: In early 2021, the SARS-CoV-2 lineage B.1.1.7 (Alpha variant) became dominant across large parts of the world. In Denmark, comprehensive and real-time test, contact-tracing, and sequencing efforts were applied to sustain epidemic control. Here, we use these data to investigate the transmissibility, introduction, and onward transmission of B.1.1.7 in Denmark. METHODS: We analyzed a comprehensive set of 60,178 SARS-CoV-2 genomes generated from high-throughput sequencing by the Danish COVID-19 Genome Consortium, representing 34% of all positive cases in the period 14 November 2020 to 7 February 2021. We calculated the transmissibility of B.1.1.7 relative to other lineages using Poisson regression. Including all 1976 high-quality B.1.1.7 genomes collected in the study period, we constructed a time-scaled phylogeny, which was coupled with detailed travel history and register data to outline the introduction and onward transmission of B.1.1.7 in Denmark. RESULTS: In a period with unchanged restrictions, we estimated an increased B.1.1.7 transmissibility of 58% (95% CI: [56%, 60%]) relative to other lineages. Epidemiological and phylogenetic analyses revealed that 37% of B.1.1.7 cases were related to the initial introduction in November 2020. The relative number of cases directly linked to introductions varied between 10 and 50% throughout the study period. CONCLUSIONS: Our findings corroborate early estimates of increased transmissibility of B.1.1.7. Both substantial early expansion when B.1.1.7 was still unmonitored and continuous foreign introductions contributed considerably to case numbers. Finally, our study highlights the benefit of balanced travel restrictions and self-isolation procedures coupled with comprehensive surveillance efforts, to sustain epidemic control in the face of emerging variants.

Effect of vaccination on household transmission of SARS-CoV-2 Delta variant of concern.

Effective vaccines protect individuals by not only reducing the susceptibility to infection, but also reducing the infectiousness of breakthrough infections in vaccinated cases. To disentangle the vaccine effectiveness against susceptibility to infection (VES) and vaccine effectiveness against infectiousness (VEI), we took advantage of Danish national data comprising 24,693 households with a primary case of SARS-CoV-2 infection (Delta Variant of Concern, 2021) including 53,584 household contacts. In this setting, we estimated VES as 61% (95%-CI: 59-63), when the primary case was unvaccinated, and VEI as 31% (95%-CI: 26-36), when the household contact was unvaccinated. Furthermore, unvaccinated secondary cases with an infection exhibited a three-fold higher viral load compared to fully vaccinated secondary cases with a breakthrough infection. Our results demonstrate that vaccinations reduce susceptibility to infection as well as infectiousness, which should be considered by policy makers when seeking to understand the public health impact of vaccination against transmission of SARS-CoV-2.

Risk of hospitalisation associated with infection with SARS-CoV-2 omicron variant versus delta variant in Denmark: an observational cohort study.

BACKGROUND: Estimates of the severity of the SARS-CoV-2 omicron variant (B.1.1.529) are crucial to assess the public health impact associated with its rapid global dissemination. We estimated the risk of SARS-CoV-2-related hospitalisations after infection with omicron compared with the delta variant (B.1.617.2) in Denmark, a country with high mRNA vaccination coverage and extensive free-of-charge PCR testing capacity. METHODS: In this observational cohort study, we included all RT-PCR-confirmed cases of SARS-CoV-2 infection in Denmark, with samples taken between Nov 21 (date of first omicron-positive sample) and Dec 19, 2021. Individuals were identified in the national COVID-19 surveillance system database, which included results of a variant-specific RT-PCR that detected omicron cases, and data on SARS-CoV-2-related hospitalisations (primary outcome of the study). We calculated the risk ratio (RR) of hospitalisation after infection with omicron compared with delta, overall and stratified by vaccination status, in a Poisson regression model with robust SEs, adjusted a priori for reinfection status, sex, age, region, comorbidities, and time period. FINDINGS: Between Nov 21 and Dec 19, 2021, among the 188 980 individuals with SARS-CoV-2 infection, 38 669 (20·5%) had the omicron variant. SARS-CoV-2-related hospitalisations and omicron cases increased during the study period. Overall, 124 313 (65·8%) of 188 980 individuals were vaccinated, and vaccination was associated with a lower risk of hospitalisation (adjusted RR 0·24, 95% CI 0·22-0·26) compared with cases with no doses or only one dose of vaccine. Compared with delta infection, omicron infection was associated with an adjusted RR of hospitalisation of 0·64 (95% CI 0·56-0·75; 222 [0·6%] of 38 669 omicron cases admitted to hospital vs 2213 [1·5%] of 150 311 delta cases). For a similar comparison by vaccination status, the RR of hospitalisation was 0·57 (0·44-0·75) among cases with no or only one dose of vaccine, 0·71 (0·60-0·86) among those who received two doses, and 0·50 (0·32-0·76) among those who received three doses. INTERPRETATION: We found a significantly lower risk of hospitalisation with omicron infection compared with delta infection among both vaccinated and unvaccinated individuals, suggesting an inherent reduced severity of omicron. Our results could guide modelling of the effect of the ongoing global omicron wave and thus health-care system preparedness. FUNDING: None.

Increased transmission of SARS-CoV-2 in Denmark during UEFA European championships.

Denmark hosted four games during the 2020 UEFA European championships (EC2020). After declining positive SARS-CoV-2 test rates in Denmark, a rise occurred during and after the tournament, concomitant with the replacement of the dominant Alpha lineage (B.1.1.7) by the Delta lineage (B.1.617.2), increasing vaccination rates and cessation of several restrictions. A cohort study including 33 227 cases was conducted from 30 May to 25 July 2021, 14 days before and after the EC2020. Included was a nested cohort with event information from big-screen events and matches at the Danish national stadium, Parken (DNSP) in Copenhagen, held from 12 June to 28 June 2021. Information from whole-genome sequencing, contact tracing and Danish registries was collected. Case-case connections were used to establish transmission trees. Cases infected on match days were compared to cases not infected on match days as a reference. The crude incidence rate ratio (IRR) of transmissions was 1.55, corresponding to 584 (1.76%) cases attributable to EC2020 celebrations. The IRR adjusted for covariates was lower (IRR 1.41) but still significant, and also pointed to a reduced number of transmissions from fully vaccinated cases (IRR 0.59). These data support the hypothesis that the EC2020 celebrations contributed to the rise of cases in Denmark in the early summer of 2021.

Molecular epidemiology of the SARS-CoV-2 variant Omicron BA.2 sub-lineage in Denmark, 29 November 2021 to 2 January 2022.

Following emergence of the SARS-CoV-2 variant Omicron in November 2021, the dominant BA.1 sub-lineage was replaced by the BA.2 sub-lineage in Denmark. We analysed the first 2,623 BA.2 cases from 29 November 2021 to 2 January 2022. No epidemiological or clinical differences were found between individuals infected with BA.1 versus BA.2. Phylogenetic analyses showed a geographic east-to-west transmission of BA.2 from the Capital Region with clusters expanding after the Christmas holidays. Mutational analysis shows distinct differences between BA.1 and BA.2.

Epidemiological characterisation of the first 785 SARS-CoV-2 Omicron variant cases in Denmark, December 2021.

By 9 December 2021, 785 SARS-CoV-2 Omicron variant cases have been identified in Denmark. Most cases were fully (76%) or booster-vaccinated (7.1%); 34 (4.3%) had a previous SARS-CoV-2 infection. The majority of cases with available information reported symptoms (509/666; 76%) and most were infected in Denmark (588/644; 91%). One in five cases cannot be linked to previous cases, indicating widespread community transmission. Nine cases have been hospitalised, one required intensive care and no deaths have been registered.