Teacher Trainees' Perspectives on Remote Instruction for Multilingual Learners of English

The COVID-19 pandemic prompted a shift to virtual learning across many countries and school systems. It is worthwhile to examine the specific ways in which this shift is significant to teacher trainees preparing to work with multilingual learners (MLs). Considering the perspectives of teacher trainees preparing to teach MLs offers an opportunity to identify the questions and concerns that they are likely to have upon graduation. Examining these perspectives can also help to identify ways that teacher trainees can use virtual and remote teaching approaches more constructively. This paper presents findings from a qualitative study of an educator preparation program focused on preparing trainees in content areas along with English to Speakers of Other Languages (ESOL), with a focus on the perspectives of teacher trainees who worked with MLs through virtual and remote modalities during the COVID-19 pandemic. The paper draws on data from an analysis of nine teacher trainees' response journals and course assignments, and includes themes identified from the teacher trainees' perceptions of virtual learning for MLs. The findings from the analysis revealed that teacher trainees emphasized the importance of establishing meaningful professional relationships in the virtual setting with their MLs, especially as a way to facilitate effective instruction and online classroom management. Participants also spoke about the importance of developing culturally responsive and sensitive instruction, and stressed the importance of engaging students and families in appropriate, linguistically accessible ways. Implications for future virtual instruction as well as teacher preparation are also discussed. © 2023 by the authors.

Lower B-Cell Responses Rather Than Circulating Antibody Titres Are Associated With SARS-CoV-2 Infection Post Third Dose COVID-19 Vaccination

Background. Which components of the immune response to SARS-CoV-2 vaccination best protect against subsequent infection remains unclear. We explored SARS-CoV-2 specific antibody and B-cell responses post 3rd dose vaccine and their relationship to subsequent SARS-CoV-2 infection. Methods. In a multicentre prospective cohort, adult subjects provided samples before and 14 days (d14) post 3rd dose vaccine with Pfizer-BioNTech 162b2. At 18-22 weeks post vaccine, subjects self-reported SARS-CoV-2 infection (confirmed by PCR or antigen test). We used electrochemiluminescence assays to quantify antibodies to SARS-CoV-2 spike subunit 1 (S1), subunit 2 (S2) and receptor-binding domain (RBD) in plasma (reported inWHOIU/mL). In a subset of subjects, we assessed SARS-CoV-2 specific differentiated B-cell (plasma cell) and memory B-cell responses from peripheral blood mononuclear cells. Unstimulated plasma cells, and memory B cells stimulated with R848 and IL2, were seeded on plates coated with RBD or full Spike antigen and antigen-specific responses measured by ELISpot (Mabtech ELISpot, Sweden). We compared between group differences by Wilcoxon signed rank or Mann-Whitney tests. Data are median [IQR] unless specified. Results. Of 133 subjects (age 43 [32-50], 81.2% female (table 1), 77 subjects in the B-cell subgroup (table 2)), 47 (35.3%) reported SARS-CoV-2 infection post 3rd vaccine. Antibody titres, plasma cell and memory B-cell responses all increased significantly at d14 post 3rd vaccine (Table 1 & 2, all P< 0.001). Although d14 antibody titres did not differ in those with and without subsequent infection (table 1), those reporting subsequent infection had significantly lower d14 RBD-specific plasma cells and a lower proportion of RBD-specific memory B-cells (Figure 1a-b, both P< 0.05). Similar results were observed with full-spike-specific memory B-cell responses (Figure 1d). The differences persisted when the non-infected group was restricted only to those reporting a confirmed close contact (n=26). Conclusion. Infection following 3rd dose vaccine is associated with lower d14 circulating and memory B cell responses, but not antibody titres, suggesting B-cell responses better predict protection against subsequent SARS-CoV-2 infection.

Performance and validation of an adaptable multiplex assay for detection of serologic response to SARS-CoV-2 infection or vaccination

Background. A wide array of assays to detect the serologic response to SARS-CoV-2 have been developed since the emergence of the pandemic. The majority of these are either qualitative or semi-quantitative, detect antibodies against one antigenic target, and are not adaptable to new antigens. Methods. We developed a new, multiplex immunoassay to detect antibodies against the receptor binding domain, S1 and S2 spike subunits and nucleocapsid (N) antigens of SARS-CoV-2 (the CEPHR SARS-CoV-2 Serology Assay). This assay uses electrochemiluminescence technology which allows for a broad dynamic range, and a linker format which allows for the addition of new antigenic targets. We tested this assay on a series of biobanked samples and validated its performance against the Abbott SARS-CoV-2 IgG and Abbott SARS-CoV-2 IgG II assays, and the MesoScale Diagnostics V-PLEX SARS-CoV-2 Panel 2 Kit. Results. Participant demographics are shown in Table 1. The mean (standard deviation (SD)) intra-assay (within plate) coefficient of variation (CV) of 80 plasma samples run on the same plate was 3.9% (2.9) for N, 3.8% (6.2) for RBD, 3.8% (5.9) for S1 and 3.9% (5.3) for S2. The mean (SD) inter-assay CV derived from 5 samples run across 3 days by two different operators was 11% (6.5) for N, 13% (5.7) for RBD, 14% (8.9) for S1 and 13% (5.1) for S2. In the convalescent group (n=193), overall sensitivity for each assay was;RBD 82% (95% CI 76-87), S1 86% (81-91%), S2 88% (83 - 92%) and N 72% (64 - 78%). Sensitivity improved when analysis included only individuals who were sampled more than 14 days from onset of symptoms (n=166), RBD 87% (81 - 95%), S1 91% (85 - 95%), S2 91% (85 - 95%) but not for the N-target (73% (66-80%)). In vaccinated individuals (n = 58), 100% (94-100%) had both detectable RBD and S1 antibodies. Overall specificity was 96% (87-99%) for RBD, 90% (78-97%) for S1, 94% (84-99%) for S2 and 90% (78-97%) for N. There was excellent correlation between the Abbott IgG II and both CEPHR anti-RBD IgG (rho 0.91) and CEPHR anti-S1 IgG (rho 0.9, both p < 0.001, Figure 1.) and the V-PLEX full spike and both CEPHR RBD IgG (rho 0.83) and S1 IgG (rho 0.82, both p < 0.001, Figure 4). Conclusion. The CEPHR SARS-CoV-2 Serology Assay is a robust, customisable, multiplex serologic assay for the detection of several different IgG specific to SARS-CoV-2.

Demonstration of stable clusters of symptoms in long COVID

Background. Long COVID is a heterogenous condition. We previously demonstrated distinct phenotypes of long COVID, but the impact of later waves caused by SARS-CoV-2 variants on long COVID presentations has not been described. Methods. We selected individuals with ongoing symptoms > 4 weeks from PCR-confirmed COVID-19 in a multicentre, prospective cohort study. We used multiple correspondence analysis and hierarchical clustering on self-reported symptoms to identify symptom clusters, in individuals recruited during two periods;cohort 1 from March 2020 to April 2021, and cohort 2 from April 2021 to March 2022. We explored differences in symptoms by mapping acute infection to one of four COVID-19 waves in Ireland (table 1) as well as vaccination status, and used Chi2 test to compare symptoms frequencies. Results. Demographics are shown in Table 2. Cluster analysis of each cohort demonstrated 3 distinct clusters in both cohorts, which shared similar clinical characteristics;a musculoskeletal/pain symptom cluster, a cardiorespiratory cluster and a third less symptomatic cluster (Figure 1). While symptoms within clusters were similar across both periods, in the cardiorespiratory cluster, the frequency of palpitations decreased (56% vs 16%) and cough increased (14% vs 45%) between reporting periods (both P< 0.01). Furthermore, a greater proportion of palpitations were reported in those with COVID-19 from waves 1 and 2 (35% and 28%) compared to 3 and 4 (both 12%, P< 0.001), and a greater proportion of chest pain in waves 1, 2 and 4 compared to wave 3. There were no differences in other symptoms (Table 3). Additionally there were significantly less palpitations reported in those vaccinated at the time of review (17% vs 31% P=0.002), but not chest pain (30% vs 39% P=0.13). In multivariate analysis, infection in wave 3 and 4 but not vaccination status remained significantly associated with lower reported palpitations (OR (95% CI) 0.28 (0.13-0.97) and 0.5 (0.06-0.87) for waves 3 and 4, both P< 0.05), and wave 3 infection remained independently associated with lower reported chest pain (OR 0.3 (0.25-0.7)). Conclusion. Three symptom clusters define long COVID across the two cohorts, but characteristics of the cardiorespiratory phenotype have evolved over time with evolution of SARS-CoV-2 variants. (Table Presented).

Reduced Serological Response to Covid-19 Booster Vaccines in Patients with Ibd Is Further Diminished by Tnf Inhibitor Therapy, Variation Study Part 2 (Variability Inresponsein Ibdagainstsars-Cov-2immunisation)

Introduction: Evidence suggests patients with inflammatory bowel disease (IBD) receiving TNF-antagonists have attenuated response to vaccination against COVID-19 {1}. Aims & Methods: We sought to determine the impact of IBD and various medications for treatment of IBD on antibody responses after 3rd vaccine dose against COVID-19. Patients with IBD (n=202) and healthy controls (HC, n= 92) were recruited prospectively. Quantitative antibody responses were assessed following COVID-19 vaccination and ACE2 binding inhibition assay to assess viral inhibition (or neutralization) were also assessed. The impact of IBD and medications for treatment of IBD on vaccine response rates was investigated. Result(s): Median age of IBD patients was 37.3 compared to 41.4 in HCs (p = 0.3). 60% of IBD patients were male compared to 23% of HCS (p =<0.001). Median time from 3rd vaccine dose to serum collection was 9.9 weeks in HCs versus 9.0 weeks in our IBD cohort (p = 0.75). 100% of HC seroconverted post 3rd vaccination. 1% (n=2) of patients with IBD failed to seroconvert. Median anti-spike protein (SP) immunoglobulin (Ig)G levels post-third vaccination in our IBD cohort was significantly lower than HC (7,862 AU/mL versus 19,622 AU/mL, p=<0.001). Median ACE2 binding inhibition (IQR) in our IBD cohort was significantly lower than HCs (97.1% (72.9 - 99.1) versus 99.9% (99.1 - 99.9), p = <0.001) with 28 (13.9%) IBD patients having ACE2 binding inhibition < 50% compared to 0 (0%) HCs (p = < 0.001). All IBD patients with ACE 2 inhibition levels < 50% were receiving biologic therapy. Breakdown of biologics received is as follows: 21 (75%) infliximab, 4 (14.2%) adalimumab, 1 (3.6%) golimumab and 1 (3.6%) vedolizumab therapy. than IBD patients not receiving TNF-inhibitors (n = 72) (10731AU/mL) (p = 0.001). Patients with IBD not receiving TNF-inhibitors still showed attenuated responses compared to HC (10730AU/mL versus 19622AU/mL p = 0.02). Conclusion(s): Patients with IBD have attenuated serological responses to SARS-CoV-2 vaccination after post booster vaccine. Use of anti-TNF therapy negatively impacts anti-SP IgG levels further. Patients who do not seroconvert post-vaccination are a particularly vulnerable cohort and causes for attenuated vaccine response need to be further investigated.

REDUCED SEROLOGICAL RESPONSE TO COVID-19 VACCINES IN PATIENTS WITH IBD IS FURTHER DIMINISHED BY TNF INHIBITOR THERAPY;EARLY RESULTS OF THE VARIATION STUDY (VARIABILITY IN RESPONSE IN IBD AGAINST SARS-COV2-IMMUNISATION)

Objective: Patients with inflammatory bowel disease (IBD) have attenuated responses to current vaccinations. There is a limited body of evidence suggesting patients with IBD receiving TNF antagonists have an attenuated response to vaccination against COVID-19. We sought to determine the impact of IBD and various medications for the treatment of IBD on antibody responses to vaccination against COVID-19. Design: Patients with IBD (n=270) and healthy controls (HC, n=116) were recruited prospectively and quantitative antibody responses assessed following COVID-19 vaccination. The impact of IBD and medications for treatment of IBD on vaccine response rates was investigated. Results: All HC seroconvert post complete vaccination with two vaccine doses [100%]. A small proportion of patients with IBD failed to seroconvert [2%]. Median anti-spike protein (SP) immunoglobulin (Ig)G levels post one vaccination and complete vaccination in our IBD cohort was significantly lower than HC [2,613 AU/mL versus 6,871 AU/mL, p=<0.001] [Figure 1]. A diagnosis of IBD was independently associated with lower anti-SP IgG levels [β coefficient -0.2, p = 0.001] whereas use of mRNA vaccines was independently associated with higher anti-SP IgG levels [β coefficient 0.25, p = < 0.001]. Patients with IBD receiving anti-TNF therapy had significantly lower anti-SP IgG levels [2444.6 AU/mL] than IBD patients not receiving these agents [3867.6 AU/mL] [p = < 0.001]. Patients with IBD not receiving TNF inhibitors still showed attenuated responses compared to HC receiving a similar vaccine [p = 0.001] [Figure 2]. 58 patients had an additional follow-up serology sample at a median of 12 weeks to complete vaccination to allow assessment of the durability of the response after their initial post-vaccination IgG level. There was a significant drop in IgG levels from 3952.85 AU/mL at the first timepoint checked post-complete vaccination to 921.1 AU/mL (343.1 – 2102.7) on follow-up sampling (p = <0.001). Median anti-SP IgG levels were numerically lower in our cohort receiving anti-TNF therapy (794.8 AU/mL) compared to those not receiving anti-TNF therapy (3136.9 AU/mL) on final follow-up samples (p =0.28). HC participants with previous COVID-19 infection (n= 5) had significantly higher anti-SP IgG levels post complete vaccination (20,719.6 AU/mL) compared to IBD patients (n=4) with prior infection (3,938.2 AU/mL) (p = < 0.001). Conclusions: Patients with IBD have attenuated serological responses to SARS-CoV-2 vaccination. Patients with IBD who do not seroconvert post-vaccination against COVID-19 are a particularly vulnerable cohort. Use of anti-TNF therapy negatively impacts anti-SP IgG levels. Impaired responses to vaccination in our study highlights the importance of booster vaccination programmes for patients with IBD. (Figure Presented) Differences in median IgG levels across three time points (Figure Presented) Differences in median anti-SP Levels dependent on medication for treatment of IBD.

ESTABLISHMENT of AN ANTI-RBD THRESHOLD THAT PREDICTS ROBUST NEUTRALIZING ACTIVITY

Background: Although presence of SARS-CoV-2 neutralising antibodies can provide protection against development of COVID-19, how reflective circulating anti-SARS-CoV-2 antibody levels are of underlying neutralising capacity, and whether a threshold exists to predict sufficient neutralising capacity remains unclear. Methods: In plasma from individuals with PCR-confirmed COVID-19 recruited to the All Ireland Infectious Diseases Cohort Study, we measured IgG concentrations against RBD, Spike protein sub-unit 1 and 2 (S1, S2) and Nucleocapsid (NC) using multiplex electrochemiluminescence (normalised to World Health Organisation reference serum as IU/mL). Neutralising capacity was measured against live SARS-CoV-2 virus (clinical isolate 2019-nCoV/Italy-INMI1) by determining the maximum plasma dilution required to maintain 50% inhibition of infection of Vero E6 cells (50% Neutralisation Titre (NT50)), by flow cytometry-based micro-neutralisation assay. Given that the Beta SARS-CoV-2 variant of concern (VOC) reduces neutralising activity up to six fold, we estimated a NT50 of 1:1000 against wild type SARS-CoV-2 would maintain neutralising activity against VOC. We used Spearman correlation and linear regression to model relationships between NT50 and IgG concentrations. Data are presented as median (IQR) unless specified. Results: In 190 individuals (age 50 (40-64) years, 55% female, time from symptom onset 98 (35-179) days), NT50 most highly correlated with anti-RBD IgG (Rho 0.81 p<0.001, Fig 1a) compared with other IgG classes (S1;Rho 0.8, S2;0.73, NC;0.72, all p<0.001). Median RBD titre was 246 (71-662) but trended lower over time, with a median of 319 (61-1012) IU/ml at 0-90 days, 244 (86-523) IU/ml at 90-180 days and 157 (80-364) IU/ml at >180 days post symptom onset respectively (p=0.08, Fig 1b). RBD IgG titres of 476 IU/ml corresponded to a NT50 of 1:1000. Overall, RBD ≥476 IU/ml predicted NT50 ≥1:1000 with a sensitivity of 77 (95% CI 65-87)% and specificity 89 (95% CI 82-93)%. This improved in an analysis restricted to convalescent samples (>30 days post symptom onset, n=148), with a sensitivity 88% (95% CI 74-96%) and specificity 90% (95%CI 82-95%) respectively. Conclusion: In convalescent plasma, RBD IgG titres ≥476IU/mL is sensitive and specific for predicting robust underlying neutralising capacity. Further research is required to validate these findings in other cohorts and confirm these thresholds in post-vaccinated individuals.

DIFFERENTIALLY EXPRESSED IMMUNOLOGICAL GENES in MILD and SEVERE CASES of COVID-19

Background: Coronavirus disease 2019 (COVID19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has varied clinical presentations from mild subclinical to severe disease with high mortality. Our aim was to determine whether examining immune-related gene expression early in infection could predict progression to severe disease. Methods: In subjects of the All Ireland Infectious Diseases Cohort study, we analysed expression of 579 genes with the NanoString nCounter Immunology panel in peripheral blood mononuclear cells in those with confirmed SARS-CoV-2 infection collected within 5 days of symptom onset and matched SARS-CoV-2 negative controls with respiratory infection. Subsequent maximum COVID19 disease severity was classified as mild or severe. Read counts were normalized using panel housekeeping genes. Expression changes in severity groups were estimated against control baseline. Results: Between April and July of 2020, we recruited 120 subjects, 62 with COVID19 and 58 controls, with average age 59 y.o. (IQR 34-88), 66% males and 69% Caucasian ethnicity. Maximal disease severity was used to separate COVID19 cases into mild (n=31) and severe (n=31). We identified 20 significantly deregulated genes between those with COVID19 and controls (;log2 fold;>0.5, p<0.05, Benjamin-Yekutieli p-adjustment). Function of 12 of these genes related to cytokine signaling, 9 upregulated genes to type I interferon signaling (MX1, IRF7, IFITM1, IFI35, STAT2, IRF4, PML, BST2, STAT1), while 7 downregulated genes mapped to innate immune function (IRF7, ICAM2, SERPING1, IFI16, BST2, FCER1A, PTK2). Expression in the severe group showed downregulation of FCER1A (innate immunity regulation), IL1B and TNF (inflammatory cytokines), and PTGS2 (inflammatory mediator) and greater upregulation of TNFSF4 (cytokine signaling) and PTK2 (innate immunity). Mild cases presented higher upregulation of IFIT2 (type I interferon signaling). Conclusion: Observed early downregulation of regulators and mediators of inflammation in those who developed severe COVID19, suggested dysregulation of inflammation. Specifically, IFIT2 upregulation in mild cases and FCER1A downregulation in severe cases, points to early differences in host responses centered on deregulation of the interferon and inflammation responses. Whether these patterns reflect delayed interferon involvement in pathways to control the infection and contribute to pathological inflammation and cytokine storms observed in severe COVID19 requires further research.

Performance and dynamic change in SARS-CoV-2 antibody responses

Background: Although reports suggest that most individuals with COVID-19 infection develop detectable antibodies post infection, the kinetics, durability, and relative differences between IgM and IgG responses remain poorly understood beyond the first few weeks after symptom onset. Methods: Within a large, well-phenotyped, diverse, prospective cohort of subjects with and without SARS-CoV-2 PCR-confirmed infection and historical controls derived from cohorts with high prevalence of viral coinfections and samples taken during prior flu seasons, we measured SARS-CoV-2 serological responses (both IgG and IgM) using three commercially available assays. We calculated sensitivity and specificity, relationship with disease severity and mapped the kinetics of antibody seropositivity and antibody levels over time using generalised additive models. Results: We analysed 1,001 samples (327 confirmed SARS-CoV-2, of whom 30% developed severe disease) from 752 subjects spanning a period of 90 days from symptom onset. Overall sensitivity was lower (44.1-47.1%) early (<10 days) after symptom onset but increased to >80% after 10 days. IgM positivity increased earlier than IgG-targeted assay but positivity peaked between day 32 and 38 post onset of symptoms and declined thereafter, a dynamic that was confirmed when antibody levels were analysed and was more rapid with IgM. Early (<10 days) IgM but not IgG levels were significantly higher in those who subsequently developed severe disease (signal / cut-off 4.20 (0.75-17.93) versus 1.07 (0.21-5.46), P=0.048). Conclusion: This study suggests that post-infectious antibody responses in those with confirmed COVID-19 infection begin to decline relatively early post infection and suggests a potential role for higher IgM levels early in infection predicting subsequent disease severity.