Combining epidemiological data and whole genome sequencing to understand SARS-CoV-2 transmission dynamics in a large tertiary care hospital during the first COVID-19 wave in The Netherlands focusing on healthcare workers.

BACKGROUND: Healthcare facilities have been challenged by the risk of SARS-CoV-2 transmission between healthcare workers (HCW) and patients. During the first wave of the COVID-19 pandemic, infections among HCW were observed, questioning infection prevention and control (IPC) measures implemented at that time. AIM: This study aimed to identify nosocomial transmission routes of SARS-CoV-2 between HCW and patients in a tertiary care hospital. METHODS: All SARS-CoV-2 PCR positive HCW and patients identified between 1 March and 19 May 2020, were included in the analysis. Epidemiological data were collected from patient files and HCW contact tracing interviews. Whole genome sequences of SARS-CoV-2 were generated using Nanopore sequencing (WGS). Epidemiological clusters were identified, whereafter WGS and epidemiological data were combined for re-evaluation of epidemiological clusters and identification of potential transmission clusters. HCW infections were further classified into categories based on the likelihood that the infection was acquired via nosocomial transmission. Secondary cases were defined as COVID-19 cases in our hospital, part of a transmission cluster, of which the index case was either a patient or HCW from our hospital. FINDINGS: The study population consisted of 293 HCW and 245 patients. Epidemiological data revealed 36 potential epidemiological clusters, with an estimated 222 (75.7%) HCW as secondary cases. WGS results were available for 195 HCW (88.2%) and 20 patients (12.8%) who belonged to an epidemiological cluster. Re-evaluation of the epidemiological clusters, with the available WGS data identified 31 transmission clusters with 65 (29.4%) HCW as secondary cases. Transmission clusters were all part of 18 (50.0%) previously determined epidemiological clusters, demonstrating that several larger outbreaks actually consisted, of several smaller transmission clusters. A total of 21 (7.2%) HCW infections were classified as from confirmed nosocomial, of which 18 were acquired from another HCW and 3 from a patient. CONCLUSION: The majority of SARS-CoV-2 infections among HCW could be attributed to community-acquired infection. Infections among HCW that could be classified as due to nosocomial transmission, were mainly caused by HCW-to-HCW transmission rather than patient-to-HCW transmission. It is important to recognize the uncertainties of cluster analyses based solely on epidemiological data.

Immunogenicity of bivalent omicron (BA.1) booster vaccination after different priming regimens in health-care workers in the Netherlands (SWITCH ON): results from the direct boost group of an open-label, multicentre, randomised controlled trial.

BACKGROUND: Bivalent mRNA-based COVID-19 vaccines encoding the ancestral and omicron spike (S) protein were developed as a countermeasure against antigenically distinct SARS-CoV-2 variants. We aimed to assess the (variant-specific) immunogenicity and reactogenicity of mRNA-based bivalent omicron (BA.1) vaccines in individuals who were primed with adenovirus-based or mRNA-based vaccines encoding the ancestral spike protein. METHODS: We analysed results of the direct boost group of the SWITCH ON study, an open-label, multicentre, randomised controlled trial. Health-care workers from four academic hospitals in the Netherlands aged 18-65 years who had completed a primary COVID-19 vaccination regimen and received one booster of an mRNA-based vaccine, given no later than 3 months previously, were eligible. Participants were randomly assigned (1:1) using computer software in block sizes of 16 and 24 to receive an omicron BA.1 bivalent booster straight away (direct boost group) or a bivalent omicron BA.5 booster, postponed for 90 days (postponed boost group), stratified by priming regimen. The BNT162b2 OMI BA.1 boost was given to participants younger than 45 years, and the mRNA-1273.214 boost was given to participants 45 years or older, as per Dutch guidelines. The direct boost group, whose results are presented here, were divided into four subgroups for analysis: (1) Ad26.COV2.S (Johnson & Johnson) prime and BNT162b2 OMI BA.1 (BioNTech-Pfizer) boost (Ad/P), (2) mRNA-based prime and BNT162b2 OMI BA.1 boost (mRNA/P), (3) Ad26.COV2.S prime and mRNA-1273.214 (Moderna) boost (Ad/M), and (4) mRNA-based prime and mRNA-1273.214 boost (mRNA/M). The primary outcome was fold change in S protein S1 subunit-specific IgG antibodies before and 28 days after booster vaccination. The primary outcome and safety were assessed in all participants except those who withdrew, had a SARS-CoV-2 breakthrough infection, or had a missing blood sample at day 0 or day 28. This trial is registered with ClinicalTrials.gov, NCT05471440. FINDINGS: Between Sept 2 and Oct 4, 2022, 219 (50%) of 434 eligible participants were randomly assigned to the direct boost group; 187 participants were included in the primary analyses; exclusions were mainly due to SARS-CoV-2 infection between days 0 and 28. From the 187 included participants, 138 (74%) were female and 49 (26%) were male. 42 (22%) of 187 participants received Ad/P and 44 (24%) mRNA/P (those aged <45 years), and 45 (24%) had received Ad/M and 56 (30%) mRNA/M (those aged ≥45 years). S1-specific binding antibody concentrations increased 7 days after bivalent booster vaccination and remained stable over 28 days in all four subgroups (geometric mean ratio [GMR] between day 0 and day 28 was 1·15 [95% CI 1·12-1·19] for the Ad/P group, 1·17 [1·14-1·20] for the mRNA/P group, 1·20 [1·17-1·23] for the Ad/M group, and 1·16 [1·13-1·19] for the mRNA/M group). We observed no significant difference in the GMR between the Ad/P and mRNA/P groups (p=0·51). The GMR appeared to be higher in the Ad/M group than in the mRNA/M group, but was not significant (p=0·073). Most side-effects were mild to moderate in severity and resolved within 48 h in most individuals. INTERPRETATION: Booster vaccination with mRNA-1273.214 or BNT162b2 OMI BA.1 in adult healthcare workers resulted in a rapid recall of humoral and cellular immune responses independent of the priming regimen. Monitoring of SARS-CoV-2 immunity at the population level, and simultaneously antigenic drift at the virus level, remains crucial to assess the necessity and timing of COVID-19 variant-specific booster vaccinations. FUNDING: The Netherlands Organization for Health Research and Development (ZonMw).

Looking back on the COVID-19 pandemic in an elite sports team using whole genome sequencing.

OBJECTIVES: The aim of this study was to investigate the effectiveness of infection control measures to prevent transmission of SARS-CoV-2 within a professional sports team using whole genome sequencing. DESIGN: Prospective cohort study. METHODS: 74 players and staff members of a Dutch professional male football team were followed from August 2020 until May 2021. A set of health and safety measures were introduced and all participants underwent regular SARS-CoV-2 RNA testing. All positive samples were subsequently sequenced (Nanopore sequencing) to assess whether infections were acquired within the training center or in the community. RESULTS: Throughout the study period, 13 participants tested positive for SARS-CoV-2. The phylogenetic analysis revealed 2 clusters (of 2 and 3 cases respectively), indicating that 3/13 cases (23%) acquired infection from another player or staff member. The first cluster was diagnosed upon enrolment, thus transmission had occurred prior to the implementation of health and safety protocols. Finally, 4 cases were diagnosed prior to symptom onset, emphasizing that frequent testing leads to early detection and isolation. CONCLUSIONS: These data show that a combination of regular testing and basic control measures can prevent outbreaks of COVID-19 in a professional sports team. Whole genome sequencing is an important tool to distinguish between infections introduced from the community and infections transmitted between athletes.

Capturing the SARS-CoV-2 infection pyramid within the municipality of Rotterdam using longitudinal sewage surveillance.

Despite high vaccination rates in the Netherlands, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to circulate. Longitudinal sewage surveillance was implemented along with the notification of cases as two parts of the surveillance pyramid to validate the use of sewage for surveillance, as an early warning tool, and to measure the effect of interventions. Sewage samples were collected from nine neighborhoods between September 2020 and November 2021. Comparative analysis and modeling were performed to understand the correlation between wastewater and case trends. Using high resolution sampling, normalization of wastewater SARS-CoV-2 concentrations, and 'normalization' of reported positive tests for testing delay and intensity, the incidence of reported positive tests could be modeled based on sewage data, and trends in both surveillance systems coincided. The high collinearity implied that high levels of viral shedding around the onset of disease largely determined SARS-CoV-2 levels in wastewater, and that the observed relationship was independent of variants of concern and vaccination levels. Sewage surveillance alongside a large-scale testing effort where 58 % of a municipality was tested, indicated a five-fold difference in the number of SARS-CoV-2-positive individuals and reported cases through standard testing. Where trends in reported positive cases were biased due to testing delay and testing behavior, wastewater surveillance can objectively display SARS-CoV-2 dynamics for both small and large locations and is sensitive enough to measure small variations in the number of infected individuals within or between neighborhoods. With the transition to a post-acute phase of the pandemic, sewage surveillance can help to keep track of re-emergence, but continued validation studies are needed to assess the predictive value of sewage surveillance with new variants. Our findings and model aid in interpreting SARS-CoV-2 surveillance data for public health decision-making and show its potential as one of the pillars of future surveillance of (re)emerging viruses.

Looking back on the COVID-19 pandemic in an elite sports team using whole genome sequencing

Objectives The aim of this study was to investigate the effectiveness of infection control measures to prevent transmission of SARS-CoV-2 within a professional sports team using whole genome sequencing (WGS). Design Prospective cohort study. Methods 74 players and staff members of a Dutch professional male football team were followed from August 2020 until May 2021. A set of health and safety measures were introduced and all participants underwent regular SARS-CoV-2 RNA testing. All positive samples were subsequently sequenced (Nanopore sequencing) to assess whether infections were acquired within the training center or in the community. Results Throughout the study period, 13 participants tested positive for SARS-CoV-2. The phylogenetic analysis revealed 3 clusters (of 2 and 3 cases respectively), indicating that 3/13 cases (23%) acquired infection from another player or staff member. The first cluster was diagnosed upon enrolment, thus transmission had occurred prior to the implementation of health and safety protocols. Finally, 4 cases were diagnosed prior to symptom onset, emphasizing that frequent testing leads to early detection and isolation. Conclusion These data show that a combination of regular testing and basic control measures can prevent outbreaks of COVID-19 in a professional sports team. WGS is an important tool to distinguish between infections introduced from the community and infections transmitted between athletes.

Risks of SARS-CoV-2 transmission between free-ranging animals and captive mink in the Netherlands.

In the Netherlands, 69 of the 126 (55%) mink farms in total became infected with SARS-CoV-2 in 2020. Despite strict biosecurity measures and extensive epidemiological investigations, the main transmission route remained unclear. A better understanding of SARS-CoV-2 transmission between mink farms is of relevance for countries where mink farming is still common practice and can be used as a case study to improve future emerging disease preparedness. We assessed whether SARS-CoV-2 spilled over from mink to free-ranging animals, and whether free-ranging animals may have played a role in farm-to-farm transmission in the Netherlands. The study encompassed farm visits, farm questionnaires, expert workshops and SARS-CoV-2 RNA and antibody testing of samples from target animal species (bats, birds and free-ranging carnivores). In this study, we show that the open housing system of mink allowed access to birds, bats and most free-ranging carnivores, and that direct and indirect contact with mink was likely after entry, especially for free-ranging carnivores and birds. This allowed SARS-CoV-2 exposure to animals entering the mink farm, and subsequent infection or mechanical carriage by the target animal species. Moreover, mink can escape farms in some cases, and two SARS-CoV-2-positive mink were found outside farm premises. No other SARS-CoV-2-RNA-positive free-ranging animals were detected, suggesting there was no abundant circulation in the species tested during the study period. To investigate previous SARS-CoV-2 infections, SARS-CoV-2 antibody detection using lung extracts of carcasses was set up and validated. One tested beech marten did have SARS-CoV-2 antibodies, but the closest SARS-CoV-2-infected mink farm was outside of its home range, making infection at a mink farm unlikely. Knowing that virus exchange between different species and the formation of animal reservoirs affects SARS-CoV-2 evolution, continued vigilance and monitoring of mink farms and surrounding wildlife remains vital.

Standardization of SARS-CoV-2 Nucleic Acid Amplification Techniques by Calibration and Quantification to the First WHO International Standard for SARS-CoV-2 RNA.

Clinical decision-making regarding isolation of SARS-CoV-2 patients is usually based on semiquantitative cycle-threshold (Ct) values without standardization. However, not all molecular assays produce Ct values, and there is ongoing discussion about whether Ct values can be safely used for decision-making. In this study, we standardized two molecular assays which use different nucleic acid amplification techniques (NAAT): the Hologic Aptima SARS-CoV-2/Flu (TMA) and Roche Cobas 6800 SARS-CoV-2 assays. We calibrated these assays against the first WHO international standard for SARS-CoV-2 RNA by using linear regression of log10 dilution series. These calibration curves were used to calculate viral loads for clinical samples. Clinical performance was assessed retrospectively using samples collected between January 2020 and November 2021, including known positives of the wild-type SARS-CoV-2 virus, the VOCs (alpha, beta, gamma, delta, and omicron) and quality control panels. Linear regression and Bland-Altman analysis showed good correlations for SARS-CoV-2 between Panther TMA and Cobas 6800 when standardized viral loads were used. These standardized quantitative results can benefit clinical decision-making and standardization of infection control guidelines.

Detection of SARS-CoV-2 in Air and on Surfaces in Rooms of Infected Nursing Home Residents.

There is an ongoing debate on airborne transmission of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) as a risk factor for infection. In this study, the level of SARS-CoV-2 in air and on surfaces of SARS-CoV-2 infected nursing home residents was assessed to gain insight in potential transmission routes. During outbreaks, air samples were collected using three different active and one passive air sampling technique in rooms of infected patients. Oropharyngeal swabs (OPS) of the residents and dry surface swabs were collected. Additionally, longitudinal passive air samples were collected during a period of 4 months in common areas of the wards. Presence of SARS-CoV-2 RNA was determined using RT-qPCR, targeting the RdRp- and E-genes. OPS, samples of two active air samplers and surface swabs with Ct-value ≤35 were tested for the presence of infectious virus by cell culture. In total, 360 air and 319 surface samples from patient rooms and common areas were collected. In rooms of 10 residents with detected SARS-CoV-2 RNA in OPS, SARS-CoV-2 RNA was detected in 93 of 184 collected environmental samples (50.5%) (lowest Ct 29.5), substantially more than in the rooms of residents with negative OPS on the day of environmental sampling (n = 2) (3.6%). SARS-CoV-2 RNA was most frequently present in the larger particle size fractions [>4 µm 60% (6/10); 1-4 µm 50% (5/10); <1 µm 20% (2/10)] (Fischer exact test P = 0.076). The highest proportion of RNA-positive air samples on room level was found with a filtration-based sampler 80% (8/10) and the cyclone-based sampler 70% (7/10), and impingement-based sampler 50% (5/10). SARS-CoV-2 RNA was detected in 10 out of 12 (83%) passive air samples in patient rooms. Both high-touch and low-touch surfaces contained SARS-CoV-2 genome in rooms of residents with positive OPS [high 38% (21/55); low 50% (22/44)]. In one active air sample, infectious virus in vitro was detected. In conclusion, SARS-CoV-2 is frequently detected in air and on surfaces in the immediate surroundings of room-isolated COVID-19 patients, providing evidence of environmental contamination. The environmental contamination of SARS-CoV-2 and infectious aerosols confirm the potential for transmission via air up to several meters.

Rise and fall of SARS-CoV-2 variants in Rotterdam: Comparison of wastewater and clinical surveillance.

Monitoring of SARS-CoV-2 in wastewater (WW) is a promising tool for epidemiological surveillance, correlating not only viral RNA levels with the infection dynamics within the population, but also to viral diversity. However, the complex mixture of viral lineages in WW samples makes tracking of specific variants or lineages circulating in the population a challenging task. We sequenced sewage samples of 9 WW-catchment areas within the city of Rotterdam, used specific signature mutations from individual SARS-CoV-2 lineages to estimate their relative abundances in WW and compared them against those observed in clinical genomic surveillance of infected individuals between September 2020 and December 2021. We showed that especially for dominant lineages, the median of the frequencies of signature mutations coincides with the occurrence of those lineages in Rotterdam's clinical genomic surveillance. This, along with digital droplet RT-PCR targeting signature mutations of specific variants of concern (VOCs), showed that several VOCs emerged, became dominant and were replaced by the next VOC in Rotterdam at different time points during the study. In addition, single nucleotide variant (SNV) analysis provided evidence that spatio-temporal clusters can also be discerned from WW samples. We were able to detect specific SNVs in sewage, including one resulting in the Q183H amino acid change in the Spike gene, that was not captured by clinical genomic surveillance. Our results highlight the potential use of WW samples for genomic surveillance, increasing the set of epidemiological tools to monitor SARS-CoV-2 diversity.

Durability of immune responses after boosting in Ad26.COV2.S-primed healthcare workers.

The emergence of SARS-CoV-2 variants raised questions regarding the durability of immune responses after homologous or heterologous booster vaccination after Ad26.COV2.S priming. We found that SARS-CoV-2-specific binding antibodies, neutralizing antibodies and T-cells are detectable 5 months after boosting, although waning of antibodies and limited cross-reactivity with Omicron BA.1 was observed.

SARS-CoV-2 outbreaks in secondary school settings in the Netherlands during fall 2020; silent circulation.

BACKGROUND: In fall 2020 when schools in the Netherlands operated under a limited set of COVID-19 measures, we conducted outbreaks studies in four secondary schools to gain insight in the level of school transmission and the role of SARS-CoV-2 transmission via air and surfaces. METHODS: Outbreak studies were performed between 11 November and 15 December 2020 when the wild-type variant of SARS-CoV-2 was dominant. Clusters of SARS-CoV-2 infections within schools were identified through a prospective school surveillance study. All school contacts of cluster cases, irrespective of symptoms, were invited for PCR testing twice within 48 h and 4-7 days later. Combined NTS and saliva samples were collected at each time point along with data on recent exposure and symptoms. Surface and active air samples were collected in the school environment. All samples were PCR-tested and sequenced when possible. RESULTS: Out of 263 sampled school contacts, 24 tested SARS-CoV-2 positive (secondary attack rate 9.1%), of which 62% remained asymptomatic and 42% had a weakly positive test result. Phylogenetic analysis on 12 subjects from 2 schools indicated a cluster of 8 and 2 secondary cases, respectively, but also other distinct strains within outbreaks. Of 51 collected air and 53 surface samples, none were SARS-CoV-2 positive. CONCLUSION: Our study confirmed within school SARS-CoV-2 transmission and substantial silent circulation, but also multiple introductions in some cases. Absence of air or surface contamination suggests environmental contamination is not widespread during school outbreaks.

A Comprehensive Sampling Study on SARS-CoV-2 Contamination of Air and Surfaces in a Large Meat Processing Plant Experiencing COVID-19 Clusters in June 2020.

OBJECTIVE: We aimed to assess SARS-CoV-2 contamination of air and surfaces to gain insight into potential occupational exposure in a large meat processing plant experiencing COVID-19 clusters. Methods: Oro-nasopharyngeal SARS-CoV-2 screening was performed in 76 workers. Environmental samples ( n = 275) including air, ventilation systems, sewage, and swabs of high-touch surfaces and workers' hands were tested for SARS-CoV-2 RNA by real-time quantitative polymerase chain reaction. Results: Twenty-seven (35.5%) of the (predominantly asymptomatic) workers tested positive with modest to low viral loads (cycle threshold ≥ 29.7). Six of 203 surface swabs, 1 of 12 personal air samples, and one of four sewage samples tested positive; other samples tested negative. Conclusions: Although one third of workers tested positive, environmental contamination was limited. Widespread SARS-CoV-2 transmission via air and surfaces was considered unlikely within this plant at the time of investigation while strict COVID-19 control measures were already implemented.

Reduced Seasonal Coronavirus Antibody Responses in Children Following COVID-19 Mitigation Measures, The Netherlands.

SARS-CoV-2 prevention and control measures did not only impact SARS-CoV-2 circulation, but also the timing and prevalence of other seasonal respiratory viruses. Especially in children, information on exposure and infections to seasonal coronaviruses as well as SARS-CoV-2 in the first year of the pandemic is largely lacking. Therefore, we set up a one-year serological survey in a large tertiary hospital in the Netherlands. We show that seasonal coronavirus seroprevalence significantly decreased in 2021 in children less than one year, most likely due to COVID-19 control measures. The SARS-CoV-2 seroprevalence in children and adolescents increased from 0.4% to 11.3%, the highest in adolescents. This implies higher exposure rates in adolescents as compared to the general population (>18 years old). It is clear that there have been significant changes in the circulation and subsequent immunity against most respiratory pathogens as a result of the mitigation measures. The implications on shorter as well as longer term are still largely unknown, but the impact of the SARS-CoV-2 pandemic and subsequent control measures will continue to affect the dynamics of other pathogens.

Normalisation of SARS-CoV-2 concentrations in wastewater: The use of flow, electrical conductivity and crAssphage.

Over the course of the Corona Virus Disease-19 (COVID-19) pandemic in 2020-2022, monitoring of the severe acute respiratory syndrome coronavirus 2 ribonucleic acid (SARS-CoV-2 RNA) in wastewater has rapidly evolved into a supplementary surveillance instrument for public health. Short term trends (2 weeks) are used as a basis for policy and decision making on measures for dealing with the pandemic. Normalisation is required to account for the dilution rate of the domestic wastewater that can strongly vary due to time- and location-dependent sewer inflow of runoff, industrial discharges and extraneous waters. The standard approach in sewage surveillance is normalisation using flow measurements, although flow based normalisation is not effective in case the wastewater volume sampled does not match the wastewater volume produced. In this paper, two alternative normalisation methods, using electrical conductivity and crAssphage have been studied and compared with the standard approach using flow measurements. For this, a total of 1116 24-h flow-proportional samples have been collected between September 2020 and August 2021 at nine monitoring locations. In addition, 221 stool samples have been analysed to determine the daily crAssphage load per person. Results show that, although crAssphage shedding rates per person vary greatly, on a population-level crAssphage loads per person per day were constant over time and similar for all catchments. Consequently, crAssphage can be used as a quantitative biomarker for populations above 5595 persons. Electrical conductivity is particularly suitable to determine dilution rates relative to dry weather flow concentrations. The overall conclusion is that flow normalisation is necessary to reliably determine short-term trends in virus circulation, and can be enhanced using crAssphage and/or electrical conductivity measurement as a quality check.

Analyzing the immunogenicity of bivalent booster vaccinations in healthcare workers: The SWITCH ON trial protocol.

Vaccination against coronavirus disease 2019 (COVID-19) has contributed greatly to providing protection against severe disease, thereby reducing hospital admissions and deaths. Several studies have reported reduction in vaccine effectiveness over time against the Omicron sub-lineages. However, the willingness to receive regular booster doses in the general population is declining. To determine the need for repeated booster vaccinations in healthy individuals and to aid policymakers in future public health interventions for COVID-19, we aim to gain insight into the immunogenicity of the additional bivalent booster vaccination in a representative sample of the healthy Dutch population. The SWITCH ON study was initiated to investigate three main topics: i) immunogenicity of bivalent vaccines after priming with adenovirus- or mRNA-based vaccines, ii) immunological recall responses and reactivity with relevant variants after booster vaccination, and iii) the necessity of booster vaccinations for the healthy population in the future. Clinical trial registration: https://clinicaltrials.gov/, identifier NCT05471440.

Ad26.COV2.S priming provided a solid immunological base for mRNA-based COVID-19 booster vaccination.

The emergence of novel SARS-CoV-2 variants led to the recommendation of booster vaccinations after Ad26.COV2.S priming. It was previously shown that heterologous booster vaccination induces high antibody levels, but how heterologous boosters affect other functional aspects of the immune response remained unknown. Here, we performed immunological profiling of Ad26.COV2.S-primed individuals before and after homologous or heterologous (mRNA-1273 or BNT162b2) booster. Booster vaccinations increased functional antibodies targeting ancestral SARS-CoV-2 and emerging variants. Especially heterologous booster vaccinations induced high levels of functional antibodies. In contrast, T-cell responses were similar in magnitude following homologous or heterologous booster vaccination and retained cross-reactivity towards variants. Booster vaccination led to a minimal expansion of SARS-CoV-2-specific T-cell clones and no increase in the breadth of the T-cell repertoire. In conclusion, we show that Ad26.COV2.S priming vaccination provided a solid immunological base for heterologous boosting, increasing humoral and cellular responses targeting emerging variants of concern.

In vitro and in vivo differences in neurovirulence between D614G, Delta And Omicron BA.1 SARS-CoV-2 variants.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is associated with various neurological complications. Although the mechanism is not fully understood, several studies have shown that neuroinflammation occurs in the acute and post-acute phase. As these studies have predominantly been performed with isolates from 2020, it is unknown if there are differences among SARS-CoV-2 variants in their ability to cause neuroinflammation. Here, we compared the neuroinvasiveness, neurotropism and neurovirulence of the SARS-CoV-2 ancestral strain D614G, the Delta (B.1.617.2) and Omicron BA.1 (B.1.1.529) variants using in vitro and in vivo models. The Omicron BA.1 variant showed reduced neurotropism and neurovirulence compared to Delta and D614G in human induced pluripotent stem cell (hiPSC)-derived cortical neurons co-cultured with astrocytes. Similar differences were obtained in Syrian hamsters inoculated with D614G, Delta and the Omicron BA.1 variant 5 days post infection. Replication in the olfactory mucosa was observed in all hamsters, but most prominently in D614G inoculated hamsters. Furthermore, neuroinvasion into the CNS via the olfactory nerve was observed in D614G, but not Delta or Omicron BA.1 inoculated hamsters. Furthermore, neuroinvasion was associated with neuroinflammation in the olfactory bulb of hamsters inoculated with D614G. Altogether, our findings suggest differences in the neuroinvasive, neurotropic and neurovirulent potential between SARS-CoV-2 variants using in vitro hiPSC-derived neural cultures and in vivo in hamsters during the acute phase of the infection.

Manifestation of SARS-CoV-2 Infections in Mink Related to Host-, Virus- and Farm-Associated Factors, The Netherlands 2020.

SARS-CoV-2 outbreaks on 69 Dutch mink farms in 2020 were studied to identify risk factors for virus introduction and transmission and to improve surveillance and containment measures. Clinical signs, laboratory test results, and epidemiological aspects were investigated, such as the date and reason of suspicion, housing, farm size and distances, human contact structure, biosecurity measures, and presence of wildlife, pets, pests, and manure management. On seven farms, extensive random sampling was performed, and age, coat color, sex, and clinical signs were recorded. Mild to severe respiratory signs and general diseases such as apathy, reduced feed intake, and increased mortality were detected on 62/69 farms. Throat swabs were more likely to result in virus detection than rectal swabs. Clinical signs differed between virus clusters and were more severe for dark-colored mink, males, and animals infected later during the year. Geographical clustering was found for one virus cluster. Shared personnel could explain some cases, but other transmission routes explaining farm-to-farm spread were not elucidated. An early warning surveillance system, strict biosecurity measures, and a (temporary) ban on mink farming and vaccinating animals and humans can contribute to reducing the risks of the virus spreading and acquisition of potential mutations relevant to human and animal health.