An Observational Cohort Study on the Incidence of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Infection and B.1.1.7 Variant Infection in Healthcare Workers by Antibody and Vaccination Status

Abstract Background Natural and vaccine-induced immunity will play a key role in controlling the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic. SARS-CoV-2 variants have the potential to evade natural and vaccine-induced immunity. Methods In a longitudinal cohort study of healthcare workers (HCWs) in Oxfordshire, United Kingdom, we investigated the protection from symptomatic and asymptomatic polymerase chain reaction (PCR)-confirmed SARS-CoV-2 infection conferred by vaccination (Pfizer-BioNTech BNT162b2, Oxford-AstraZeneca ChAdOx1 nCOV-19) and prior infection (determined using anti-spike antibody status), using Poisson regression adjusted for age, sex, temporal changes in incidence and role. We estimated protection conferred after 1 versus 2 vaccinations and from infections with the B.1.1.7 variant identified using whole genome sequencing. Results In total, 13 109 HCWs participated; 8285 received the Pfizer-BioNTech vaccine (1407 two doses), and 2738 the Oxford-AstraZeneca vaccine (49 two doses). Compared to unvaccinated seronegative HCWs, natural immunity and 2 vaccination doses provided similar protection against symptomatic infection: no HCW vaccinated twice had symptomatic infection, and incidence was 98% lower in seropositive HCWs (adjusted incidence rate ratio 0.02 [95% confidence interval {CI} < .01–.18]). Two vaccine doses or seropositivity reduced the incidence of any PCR-positive result with or without symptoms by 90% (0.10 [95% CI .02–.38]) and 85% (0.15 [95% CI .08–.26]), respectively. Single-dose vaccination reduced the incidence of symptomatic infection by 67% (0.33 [95% CI .21–.52]) and any PCR-positive result by 64% (0.36 [95% CI .26–.50]). There was no evidence of differences in immunity induced by natural infection and vaccination for infections with S-gene target failure and B.1.1.7. Conclusions Natural infection resulting in detectable anti-spike antibodies and 2 vaccine doses both provide robust protection against SARS-CoV-2 infection, including against the B.1.1.7 variant.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has had a global impact on morbidity and mortality [1]. Natural and vaccine-induced immunity will play a key role in controlling the pandemic, by reducing transmission, hospitalization and mortality. However, the ability of new SARS-CoV-2 variants to evade natural and vaccine-induced immunity mounted against ancestral viruses is of major public health concern.
Prior SARS-CoV-2 infection protects against polymerase chain reaction (PCR)-confirmed symptomatic/asymptomatic SARS-CoV-2 infection by 83-88% up to 5-6 months postinfection, with greater reductions in symptomatic reinfections [2][3][4]. Ongoing longitudinal studies are required to determine the duration of protection conferred by natural immunity; however evaluating this will be more difficult with widespread vaccination. Understanding the interaction between prior infection/serostatus and vaccination on protection from infection is also important.
Three vaccines have been approved for use in the United Kingdom to date [5], with Pfizer-BioNTech BNT162b2 and Oxford-AstraZeneca ChAdOx1 nCoV-19 (AZD1222) currently the most widely deployed, with many individuals receiving only one dose to date following a government decision to extend the dosing interval to 12 weeks to maximize initial coverage. For BNT162b2, trials demonstrated 95% efficacy in preventing symptomatic PCR-confirmed infection >7 days post-second dose; these findings have been replicated in several real-world studies including in Israel (92% effectiveness) [6] and the United Kingdom (88% effectiveness in individuals >80 years [7]; 85% reduction in all-PCR positives in a cohort of healthcare workers [HCWs]) [8]. Vaccine efficacy of 50-90% is seen following a single dose, dependent on population demographics, exposures and time-frame studied [6,7,[9][10][11][12][13]. Fewer real-world data are available for ChAdOx1 nCoV-19, due to its later regulatory approval. Trials demonstrated vaccine efficacy of 62% against PCR-positive infection >14 days post-second dose using a standard dose/standard dose regimen, with subsequent analysis showing a higher efficacy of 81% in those with a longer dosing interval (>12 weeks). Single dose vaccine efficacy >22 days postfirst dose has been reported as 69-76% [14,15]. No real-world data on vaccine effectiveness against PCR-positive infections has been published, but preliminary analyses show a reduction in hospital admissions in the United Kingdom [16].
A novel SARS-CoV-2 variant, B.1.1.7, identified in September 2020 in the United Kingdom, has spread rapidly. Estimates suggest increased transmissibility and disease severity [17][18][19][20]. The lineage carries several mutations of immunologic significance, including N501Y located in the receptor-binding domain (RBD), a key neutralizing antibody target; deletions in the N-terminal domain at residues 69/70, associated with viral escape in the immunocompromised and S-gene target failure (SGTF) in PCR assays; and a deletion at residue 144 resulting in decreased monoclonal antibody binding [21].
We use an observational longitudinal cohort study of hospital HCWs to investigate and compare the protection from SARS-CoV-2 infection conferred by vaccination and prior infection (determined using anti-spike antibody status). Additionally, we estimate the protection provided by different vaccines, after 1 versus 2 doses and from infections with the B.1.1.7 variant confirmed by whole-genome sequencing.

Setting
Oxford University Hospitals (OUH) offers symptomatic and asymptomatic SARS-CoV-2 testing to all staff at 4 hospitals and associated facilities in Oxfordshire, United Kingdom. SARS-CoV-2 PCR testing of nasal and oropharyngeal swabs for symptomatic (new persistent cough, fever ≥37.8°C, anosmia/ageusia) staff was offered from 27 March 2020. Asymptomatic HCWs were offered voluntary nasal and oropharyngeal swab PCR testing every 2 weeks and serological testing every 2 months from 23 April 2020, as previously described [2,37,38]. We report data to 28 February 2021. To minimize underascertainment of outcomes arising from staff leaving OUH's employment, only those who participated in asymptomatic screening, symptomatic testing or vaccination from 1 September 2020 onward were included. We also performed a sensitivity analysis restricted to staff participating in asymptomatic screening or symptomatic testing from 1 September 2020. All staff working for the hospitals were eligible to participate.

Laboratory Assays
Antibody status was determined using an anti-trimeric spike immunoglobulin G (IgG) enzyme linked immunosorbent assay (ELISA) [39] using an 8 million units threshold to determine antibody-positivity. PCR tests were performed by OUH using a range of PCR assays (see Supplementary materials). PCRpositive results from symptomatic community testing were also recorded. From 16 November 2020, OUH used the Thermo-Fisher TaqPath PCR assay as their first-line diagnostic assay, which includes orf1ab, S and N gene targets. As such SGTF indicative of the B.1.1.7 variant [20] could be identified, that is, orf1ab-positive/N-positive only. Oxford Nanopore sequencing was undertaken of all stored PCR-positive primary samples from 1 December 2020 onward to identify the infecting lineage (see Supplementary materials).

Study Groups
Staff members were classified into 5 groups: (a) unvaccinated and consistently seronegative during follow-up; (b) unvaccinated and ever seropositive; (c) vaccinated once, always seronegative prior to vaccination; (d) vaccinated twice, always seronegative prior to first vaccination; (e) vaccinated (once or twice) and ever seropositive prior to first vaccination. The latter group were combined as relatively few staff were previously seropositive and vaccinated twice. Vaccinated groups were considered at-risk of infection >14 days after each vaccine dose (see Table 1 for further details of at-risk periods).
Staff remained at risk of infection in each follow-up group until the earliest of the study end, first vaccination, second vaccination in previously seronegative HCWs, a positive PCR test, or for unvaccinated HCWs, a positive antibody test. Staff could transition from one group to another following seroconversion or vaccination after 60 or 14 days, respectively, disregarding any PCR-positive result during this crossover period, including the 14 days following a second vaccination for previously seronegative HCWs vaccinated twice.
The staff vaccination program began on 8 December 2020, starting with the Pfizer-BioNTech BNT162b2 vaccine, with the addition of the Oxford-AstraZeneca ChAdOx1 nCoV-19 vaccine from 4 January 2021. Some staff members received the ChAdOx1 nCoV-19 vaccine in clinical trials beginning 23 April 2020 and were included following unblinding.

Outcomes
The main outcome was PCR-confirmed symptomatic SARS-CoV-2 infection. We also considered any PCR-positive result (ie, either symptomatic or asymptomatic). To assess the impact of the B.1.1.7 variant on (re)infection risk, we also analyzed PCR-positive results with and without SGTF, and those confirmed as B.1.1.7 on sequencing.

Statistical Analysis
We used Poisson regression to model incidence of each outcome per day-at-risk by study group. We adjusted for calendar month, age, sex, self-reported ethnicity and staff occupational role, patient contact and working on a non-intensive care unit (ICU) ward caring for coronavirus disease 2019 (COVID- 19) patients (previously shown to increase risk [37]) (details in Supplementary materials). We compared incidence in each follow-up group to unvaccinated seronegative HCWs, using incidence rate ratios (IRRs), such that 100*(1-IRR) is the percentage protection arising from being seropositive or vaccinated. We tested for heterogeneity by vaccine type. To assess timing of onset of protection we also fitted models in vaccinated individuals from day 1 postvaccination.
We used stacked Poisson regression to test for variation in the incidence of SGTF versus non-SGTF PCR-positive results, and B.1.1.7 versus non-B.1.1.7, considering only results from 1 December 2020 where S-gene PCR and sequencing were most complete.
We compared PCR cycle threshold (Ct) values between symptomatic and asymptomatic infections and by follow-up group using quantile regression.

Ethics Statement
Deidentified data were obtained from the Infections in Oxfordshire Research Database which has generic Research Ethics Committee, Health Research Authority and Confidentiality Advisory Group approvals (19/SC/0403, 19/ CAG/0144).
Thirty-eight unvaccinated seronegative HCWs attended hospital within −2 to + 28 days of a SARS-CoV-2 PCR-positive result (14.2/million person-days); of these, 27 had a COVID-19 primary diagnostic code, and 16 required admission for COVID-19. Two previously seronegative vaccinated HCWs required hospital review (6.9/million person-days); however, neither required admission. No HCW vaccinated twice or unvaccinated seropositive HCW required hospital review or admission.

PCR-Positive Results Following Vaccination
The incidence of PCR-positive results fell from >14 days after the first vaccination for both the Pfizer-BioNTech and Oxford-AstraZeneca vaccines, with similar levels of protection seen up to 42 days postvaccine ( Figure 3). There was an unexpected rise in incidence above baseline levels in the first two weeks following vaccination, which remained to some extent after adjustment ( Figure 3B). Considering efficacy against any PCRpositive result >14 days post first dose, there was no evidence of a difference by vaccine type following the first (heterogeneity P = .33) or second (P = .16) dose. Similarly, there was no evidence of difference in PCR-confirmed symptomatic SARS-CoV-2 infection (P = .21 and P > .99, respectively).  Table 4).   in mid-November 2020, rising to 90% in the second half of January 2021, before declining again ( Figure 5A). There was no evidence that SGTF changed the extent of protection against any PCR-positive infection in unvaccinated seropositive We used viral whole-genome sequencing to determine infecting lineages from 1 December 2020 onward (Supplementary Table 5

DISCUSSION
In this longitudinal cohort study of HCWs receiving Pfizer-BioNTech and Oxford-AstraZeneca vaccines, vaccination reduced the incidence of PCR-positive symptomatic SARS-CoV-2 infection, with 2 doses providing similar levels of protection to natural immunity. No symptomatic infections were seen following two vaccine doses and there was a 98% reduction in symptomatic infections in unvaccinated seropositive HCWs. Protection was still afforded >14 days after a single vaccine dose, albeit at lower levels (67% reduction). No vaccinated HCW required hospital admission. Furthermore, vaccination Abbreviations: CI, confidence interval; ICU, intensive care unit; PCR, polymerase chain reaction; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
reduced the incidence of any PCR-positivity by 64% and 90% >14 days post-first and second vaccine dose, respectively, compared to an 85% reduction post-natural infection. This suggests that both vaccination and previous infection are also likely to reduce transmission. Additionally, there was a trend toward reduced viral loads in reinfected individuals compared to infected seronegative HCWs, with a smaller observed reduction postvaccination. The comparable protection offered by seropositivity to 2 doses of vaccine suggests that the immunoassay used provides an accurate correlate of immunity, which could potentially be used to support individualized relaxation of societal restrictions. Furthermore, where vaccine supplies are limited prioritizing seronegative infection-naïve individuals may be appropriate.
Protection following 2 vaccine doses was comparable to other real-world studies [6,8]. Protection following a single dose was toward the lower range of previous reports, potentially reflecting occupational exposure in HCWs. Although an unexpected rise in incidence was seen in the first 2 weeks postvaccination, this time period was excluded from effectiveness calculations. Possible explanations include increased ascertainment of asymptomatic infection due to vaccinerelated symptoms leading to testing, behavior change, acquisition at vaccination facilities, or staff attending for vaccination prompted by high levels of exposure to infected colleagues or patients. A similar rise in incidence was noted in the Israeli mass vaccination program, attributed to behavior change postvaccination [11,12].
Immunity induced by natural infection and vaccination was robust to lineage, including cases confirmed to be B.1.1.7 by whole-genome sequencing, at least within the power of the study. Sequencing was important to confirm the lineage of SGTF cases: although >99% del69-70 sequences from Southeast England were due to B.1.1.7 over this period [20], locally 17% of SGTF was due to other non-B.1.1.7 lineages. Assuming all SGTF is B.1.1.7 risks misestimating the impact of this lineage on natural and vaccine-induced immunity. This reinforces the need to understand local genomic epidemiology and the reliability of SGTF as a proxy for B.1.1.7 over time. Our results are comparable with the Oxford-AstraZeneca analysis of vaccine efficacy against B.1.1.7 based on a relatively low proportion of successfully sequenced cases (179/499, 36%) and no documentation of SGTF status [35], compared to this study, where PCR and WGS confirmed SGTF/lineage status in 78% cases.
One important finding is that despite universal use of personal protective equipment (gloves, plastic aprons, surgical marks for all patient care and FFP3 masks, gowns and eye protection for aerosol generating procedures), social distancing, and use of surgical masks throughout all areas of the hospital, staff working in COVID-19 wards remained at higher risk of SARS-CoV-2 infection independent of vaccine and antibody status. Possible explanations include acquisition from patients   with or without subsequent amplification by staff-to-staff spread. Nurses, healthcare assistants, and Asian staff were also at higher risk of infection, possibly reflecting both hospital and community-based exposures as we have discussed previously [37].
One study limitation is that staff working in roles more likely to be exposed to SARS-CoV-2 were initially prioritized for vaccination; these staff were also at the greatest risk of occupationally-acquired SARS-CoV-2 infection. We adjusted for this by including working in a COVID-19 ward and staff roles, but incomplete adjustment could lead to underestimation of vaccine efficacy. Similarly, vaccinated staff were potentially more likely to be current employees than unvaccinated staff; if unvaccinated seronegative staff left employment this would potentially lead to underascertainment of infection in this group. We address this by only including staff using testing and/or vaccination services in the last 6 months of the study. Testing rates were lower in seropositive HCWs and to a lesser extent following vaccination, leading to underascertainment of PCRpositive results in these groups; however, we have previously demonstrated the impact of this is relatively small [2]. Other limitations include limited power to detect differences in efficacy between vaccines. We were also unable to sequence all PCR-positives, in particular because those with higher Ct values are less likely to generate high-quality sequences, and some samples were not stored, including those processed by community testing facilities. Similar studies will be needed to assess the vaccine effectiveness against other, novel emerging SARS-CoV-2 lineages. Finally, this is a study of HCWs of working age, so findings may not generalize to other settings.
In summary, by pooling data from unvaccinated and Pfizer-BioNTech and AstraZeneca vaccinated HCWs, we show that natural infection resulting in detectable anti-spike antibodies and 2 doses of vaccine both provide robust protection against SARS-CoV-2 infection, including against the B.1.1.7 variant of concern.  Week begining  to an application and research proposal meeting the ethical and governance requirements of the Database. Sequence data generated during the study are available from the European Nucleotide Archive under study accession number PRJEB43319.