The effect of dose-interval on antibody response to mRNA COVID-19 vaccines: a prospective cohort study.

Background: Vaccination against COVID-19 is highly effective in preventing severe disease and hospitalization, but primary COVID mRNA vaccination schedules often differed from those recommended by the manufacturers due to supply chain issues. We investigated the impact of delaying the second dose on antibody responses to COVID mRNA-vaccines in a prospective cohort of health-care workers in Quebec. Methods: We recruited participants from the McGill University Health Centre who provided serum or participant-collected dried blood samples (DBS) at 28-days, 3 months, and 6 months post-second dose and at 28-days after a third dose. IgG antibodies to SARS-CoV2 spike (S), the receptor-binding domain (RBD), nucleocapsid (N) and neutralizing antibodies to the ancestral strain were assessed by enzyme-linked immunosorbent assay (ELISA). We examined associations between long (≤89 days) versus short (<89 days) between-dose intervals and antibody response through multivariable mixed-effects models adjusted for age, sex, prior covid infection status, time since vaccine dose, and assay batch. Findings: The cohort included 328 participants who received up to three vaccine doses (>80% Pfizer-BioNTech). Weighted averages of the serum (n=744) and DBS (n=216) cohort results from the multivariable models showed that IgG anti-S was 31% higher (95% CI: 12% to 53%) and IgG anti-RBD was 37% higher (95% CI: 14% to 65%) in the long vs. short interval participants, across all time points. Interpretation: Our study indicates that extending the covid primary series between-dose interval beyond 89 days (approximately 3 months) provides stronger antibody responses than intervals less than 89 days. Our demonstration of a more robust antibody response with a longer between dose interval is reassuring as logistical and supply challenges are navigated in low-resource settings.

Skin prick test in milk allergic patients undergoing oral immunotherapy: Does the milk form used for skin tests matter?

SPT is the most commonly used confirmatory test for an IgE-mediated milk allergy. However, food SPTs are not standardized. We aimed to assess the accuracy of SPTs with extract, diluted, and undiluted milk to detect desensitization in children with milk allergy undergoing OIT. Children with milk allergy undergoing OIT and controls were recruited from Montreal Children's Hospital (MCH), British Columbia Children's Hospital (BCCH) and The Hospital for Sick Children (SickKids). Participants in the active arm received a weekly increase in milk until 200 ml of pure milk was tolerated. SPT using milk extract (Omega), diluted 2% milk (1:10), and undiluted milk was done at the study entry and when 200 ml of pure milk was reached. Participants in the control arm had SPT at study entry and 12 months later before they entered the active arm. Among 53 children who reached 200 ml, the median age was 12 years and 54.7% were males. The mean decrease in wheal size at 200 ml from the baseline was 3.78 mm (95%CI, 2.55-5.01), 5.05 mm (95% CI, 3.68-6.41), and 5.05 mm (95% CI, 3.29-6.80) for milk extract, diluted and undiluted milk respectively. Among 32 controls, the median age was 10 years and 62.5% were males. There was no significant change in wheal diameter over a one-year period regardless of the skin test method. Response to extract behaved similarly to whole food (Diluted and undiluted) and thus can be used to follow sensitization in the context of a desensitization program.

Real-world serological responses to extended-interval and heterologous COVID-19 mRNA vaccination in frail, older people (UNCoVER): an interim report from a prospective observational cohort study.

BACKGROUND: The use of COVID-19 vaccines has been prioritised to protect the most vulnerable-notably, older people. Because of fluctuations in vaccine availability, strategies such as delayed second dose and heterologous prime-boost have been used. However, the effectiveness of these strategies in frail, older people are unknown. We aimed to assess the antigenicity of mRNA-based COVID-19 vaccines in frail, older people in a real-world setting, with a rationed interval dosing of 16 weeks between the prime and boost doses. METHODS: This prospective observational cohort study was done across 12 long-term care facilities of the Montréal Centre-Sud - Integrated University Health and Social Services Centre in Montréal, Québec, Canada. Under a rationing strategy mandated by the provincial government, adults aged 65 years and older residing in long-term care facilities in Québec, Canada, with or without previously documented SARS-CoV-2 infection, were administered homologous or heterologous mRNA vaccines, with an extended 16-week interval between doses. All older residents in participating long-term care facilities who received two vaccine doses were eligible for inclusion in this study. Participants were enrolled from Dec 31, 2020, to Feb 16, 2021, and data were collected up to June 9, 2021. Clinical data and blood samples were serially collected from participants at the following timepoints: at baseline, before the first dose; 4 weeks after the first dose; 6-10 weeks after the first dose; 16 weeks after the first dose, up to 2 days before administration of the second dose; and 4 weeks after the second dose. Sera were tested for SARS-CoV-2-specific IgG antibodies (to the trimeric spike protein, the receptor-binding domain [RBD] of the spike protein, and the nucleocapsid protein) by automated chemiluminescent ELISA. Two cohorts were used in this study: a discovery cohort, for which blood samples were collected before administration of the first vaccine dose and longitudinally thereafter; and a confirmatory cohort, for which blood samples were only collected from 4 weeks after the prime dose. Analyses were done in the discovery cohort, with validation in the confirmatory cohort, when applicable. FINDINGS: The total study sample consisted of 185 participants. 65 participants received two doses of mRNA-1273 (Spikevax; Moderna), 36 received two doses of BNT162b2 (Comirnaty; Pfizer-BioNTech), and 84 received mRNA-1273 followed by BNT162b2. In the discovery cohort, after a significant increase in anti-RBD and anti-spike IgG concentrations 4 weeks after the prime dose (from 4·86 log binding antibody units [BAU]/mL to 8·53 log BAU/mL for anti-RBD IgG and from 5·21 log BAU/mL to 8·05 log BAU/mL for anti-spike IgG), there was a significant decline in anti-RBD and anti-spike IgG concentrations until the boost dose (7·10 log BAU/mL for anti-RBD IgG and 7·60 log BAU/mL for anti-spike IgG), followed by an increase 4 weeks later for both vaccines (9·58 log BAU/mL for anti-RBD IgG and 9·23 log BAU/mL for anti-spike IgG). SARS-CoV-2-naive individuals showed lower antibody responses than previously infected individuals at all timepoints tested up to 16 weeks after the prime dose, but achieved similar antibody responses to previously infected participants by 4 weeks after the second dose. Individuals primed with the BNT162b2 vaccine showed a larger decrease in mean anti-RBD and anti-spike IgG concentrations with a 16-week interval between doses (from 8·12 log BAU/mL to 4·25 log BAU/mL for anti-RBD IgG responses and from 8·18 log BAU/mL to 6·66 log BAU/mL for anti-spike IgG responses) than did those who received the mRNA-1273 vaccine (two doses of mRNA-1273: from 8·06 log BAU/mL to 7·49 log BAU/mL for anti-RBD IgG responses and from 6·82 log BAU/mL to 7·56 log BAU/mL for anti-spike IgG responses; mRNA-1273 followed by BNT162b2: from 8·83 log BAU/mL to 7·95 log BAU/mL for anti-RBD IgG responses and from 8·50 log BAU/mL to 7·97 log BAU/mL for anti-spike IgG responses). No differences in antibody responses 4 weeks after the second dose were noted between the two vaccines, in either homologous or heterologous combinations. INTERPRETATION: Interim results of this ongoing longitudinal study show that among frail, older people, previous SARS-CoV-2 infection and the type of mRNA vaccine influenced antibody responses when used with a 16-week interval between doses. In these cohorts of frail, older individuals with a similar age and comorbidity distribution, we found that serological responses were similar and clinically equivalent between the discovery and confirmatory cohorts. Homologous and heterologous use of mRNA vaccines was not associated with significant differences in antibody responses 4 weeks following the second dose, supporting their interchangeability. FUNDING: Public Health Agency of Canada, Vaccine Surveillance Reference Group; and the COVID-19 Immunity Task Force. TRANSLATION: For the French translation of the abstract see Supplementary Materials section.

JOTROL, a Novel Formulation of Resveratrol, Shows Beneficial Effects in the 3xTg-AD Mouse Model.

BACKGROUND: Alzheimer's disease (AD) has minimally effective treatments currently. High concentrations of resveratrol, a polyphenol antioxidant found in plants, have been reported to affect several AD-related and neuroprotective genes. To address the low bioavailability of resveratrol, we investigated a novel oral formulation of resveratrol, JOTROL™, that has shown increased pharmacokinetic properties compared to non-formulated resveratrol in animals and in humans. OBJECTIVE: We hypothesized that equivalent doses of JOTROL, compared to non-formulated resveratrol, would result in greater brain exposure to resveratrol, and more efficacious responses on AD biomarkers. METHODS: For sub-chronic reversal studies, 15-month-old male triple transgenic (APPSW/PS1M146V/TauP301L; 3xTg-AD) AD mice were treated orally with vehicle or 50 mg/kg JOTROL for 36 days. For prophylactic studies, male and female 3xTg-AD mice were similarly administered vehicle, 50 mg/kg JOTROL, or 50 mg/kg resveratrol for 9 months starting at 4 months of age. A behavioral battery was run, and mRNA and protein from brain and blood were analyzed for changes in AD-related gene and protein expression. RESULTS: JOTROL displays significantly increased bioavailability over non-formulated resveratrol. Treatment with JOTROL resulted in AD-related gene expression changes (Adam10, Bace1, Bdnf, Psen1) some of which were brain region-dependent and sex-specific, as well as changes in inflammatory gene and cytokine levels. CONCLUSION: JOTROL may be effective as a prophylaxis and/or treatment for AD through increased expression and/or activation of neuroprotective genes, suppression of pro-inflammatory genes, and regulation of central and peripheral cytokine levels.

Elevated Cow's Milk-Specific IgE Levels Prior to Oral Immunotherapy Decrease the Likelihood of Reaching the Maintenance Dose.

BACKGROUND: Food desensitization via oral immunotherapy (OIT) is gaining acceptance in clinical practice. Owing to adverse reactions, the duration of the buildup phase until a maintenance dose is achieved may be prolonged, and in a minority of cases, OIT is stopped. OBJECTIVE: We aimed to assess factors associated with the probability of reaching the maintenance dose in cow's milk (CM) OIT. METHODS: We collected data from patients undergoing CM OIT at the Montreal Children's Hospital, BC Children's Hospital, and Hospital for Sick Children. We compared univariable and multivariable Cox regressions to evaluate sociodemographic factors, comorbidities, clinical characteristics, and biomarkers at study entry associated with the likelihood of reaching a maintenance dose of 200 mL of CM. RESULTS: Among 69 children who reached 4 mL of milk, the median age was 12 years (interquartile range, 9-15 years); 59% were male. Median duration of buildup phase from 4 to 200 mL was 24.0 weeks (interquartile range, 17.7-33.4 weeks). After adjusting for age and sex, higher baseline levels of specific IgE antibodies for α-lactalbumin (hazard ratio [HR] = 0.80; 95% confidence interval [CI], 0.67-0.95), ß-lactoglobulin (HR = 0.86; 95% CI, 0.76-0.98), casein (HR = 0.82; 95% CI, 0.72-0.94), and total CM (HR = 0.79; 95% CI, 0.65-0.97) were associated with a decreased probability of reaching maintenance. In addition, for every 10-mL increase in CM tolerated at entry challenge, the probability of reaching maintenance increased by 10%. CONCLUSIONS: The data suggest that higher levels of CM-specific IgE decreased the likelihood of reaching maintenance, whereas an increased cumulative CM dose at entry challenge increased the likelihood. Assessing these factors before therapy may assist in predicting the success of CM OIT.

Age- and sex-mediated differences in T lymphocyte populations of kidney transplant recipients.

BACKGROUND: Graft failure rates increase through childhood and adolescence, decline in adulthood, and are higher in female than male kidney transplant recipients (KTR) until middle age. We aimed to describe age- and sex-related differences in T-cell subsets among KTR to determine which differences may help to explain the differences in kidney graft failure rates. METHODS: Effector T (Teff)-cell and regulatory T (Treg)-cell phenotypes in PBMCs from healthy controls and KTR, who were at least 1 year post-transplant with stable graft function under immunosuppression, were analyzed by flow cytometry. The effects of age, sex, and status (KTR or control) were analyzed using linear regressions. RESULTS: We enrolled 20 male and 21 female KTR and 20 male and 20 female controls between 3 and 29 years of age. CD3+ T-cell frequencies were not associated with age or sex but were higher in KTR than controls. There were no differences in CD4+ and CD8+ frequencies. Th1 (IFNγ+ IL-4- IL-17A-) and Th17 (IL-17A+) frequencies within the CD4+ T-cell population were higher at older ages. The frequencies of FOXP3 + Helios + Treg cells in CD4+ CD25+ CD127- T cells were lower in females than males and in KTR than controls. CONCLUSIONS: Increasing frequencies of Th1 and Th17 cells with increasing age mirrors the increasing graft failure rates from childhood to young adulthood. Importantly, sex differences in frequencies of circulating Treg cells may suggest a role in the sex differences in graft failure rates.

Enhanced interleukin-8 production in mononuclear cells in severe pediatric obstructive sleep apnea.

BACKGROUND: Obstructive sleep apnea (OSA) is a risk factor for cardiovascular disease, metabolic disorders, and cognitive dysfunction. Current thinking links chronic intermittent hypoxia (CIH) with oxidative stress and systemic inflammation. However, the sequence of events leading to the morbidities associated with OSA is poorly understood in children. Monocytes are known to be altered by chronic hypoxia. Thus in this prospective study, we investigated inflammatory cytokine profiles from cultures of peripheral blood mononuclear cells (PBMC) obtained from children with severe OSA and sleep-related CIH. METHODS: Ten children with OSA (cases) and 5 age-matched children without OSA (controls) were recruited for study. Samples of plasma and PBMC were obtained before and after adenotonsillectomy. The levels of the inflammatory cytokines, interleukin (IL)-1ß, IL-6, IL-8, IL-10, IL-12p70, and tumor necrosis factor-α (TNFα), were measured in both plasma and ex vivo culture supernatants of PBMC incubated with lipopolysaccharide (LPS) using the cytometric bead assay. RESULTS: Upon activation of PBMC by LPS, the levels of IL-8 in the culture supernatants from cases were threefold higher than in controls. The levels of the other cytokines including IL-1ß, IL-6, and TNFα, in culture supernatant of PBMC from cases showed no difference from controls; nor were there significant differences in plasma cytokine levels. CONCLUSION: We speculate that in young children with sleep-related CIH, an enhanced production capacity of IL-8 precedes the development of systemic inflammatory markers. Future work should evaluate IL-8 production capacity as a potential biomarker for OSA severity.

Biophysical characterization of bladder cancer cells with different metastatic potential.

Specific membrane capacitance (SMC) and Young's modulus are two important parameters characterizing the biophysical properties of a cell. In this work, the SMC and Young's modulus of two cell lines, RT4 and T24, corresponding to well differentiated (low grade) and poorly differentiated (high grade) urothelial cell carcinoma (UCC), respectively, were quantified using microfluidic and AFM measurements. Quantitative differences in SMC and Young's modulus values of the high-grade and low-grade UCC cells are, for the first time, reported.

Staphylococcus aureus strains that express serotype 5 or serotype 8 capsular polysaccharides differ in virulence.

Most isolates of Staphylococcus aureus produce a serotype 5 (CP5) or 8 (CP8) capsular polysaccharide. To investigate whether CP5 and CP8 differ in their biological properties, we created isogenic mutants of S. aureus Reynolds that expressed CP5, CP8, or no capsule. Biochemical analyses of CP5 and CP8 purified from the isogenic S. aureus strains were consistent with published structures. The degree of O acetylation of each polysaccharide was similar, but CP5 showed a greater degree of N acetylation. Mice challenged with the CP5(+) strain showed a significantly higher bacteremia level than mice challenged with the CP8(+) strain. Similarly, the CP5(+) strain survived preferentially in the bloodstream and kidneys of infected mice challenged with a mixed inoculum containing both strains. The enhanced virulence of the CP5(+) strain in vivo correlated with its greater resistance to in vitro killing in whole mouse blood. Likewise, in vitro opsonophagocytic killing assays with human neutrophils and sera revealed greater survival of the Reynolds (CP5) strain, even though the kinetics of opsonization by C3b and iC3b was similar for both the CP5(+) and CP8(+) strains. Electron micrographs demonstrated C3 molecules on the cell wall beneath the capsule layer for both serotype 5 and 8 strains. Purified CP5 and CP8 stimulated a modest oxidative burst in human neutrophils but failed to activate the alternative complement pathway. These results indicate that CP5 and CP8 differ in a number of biological properties, and these differences likely contribute to the relative virulence of serotype 5 and 8 S. aureus in vivo.

Use of tuf sequences for genus-specific PCR detection and phylogenetic analysis of 28 streptococcal species.

A 761-bp portion of the tuf gene (encoding the elongation factor Tu) from 28 clinically relevant streptococcal species was obtained by sequencing amplicons generated using broad-range PCR primers. These tuf sequences were used to select Streptococcus-specific PCR primers and to perform phylogenetic analysis. The specificity of the PCR assay was verified using 102 different bacterial species, including the 28 streptococcal species. Genomic DNA purified from all streptococcal species was efficiently detected, whereas there was no amplification with DNA from 72 of the 74 nonstreptococcal bacterial species tested. There was cross-amplification with DNAs from Enterococcus durans and Lactococcus lactis. However, the 15 to 31% nucleotide sequence divergence in the 761-bp tuf portion of these two species compared to any streptococcal tuf sequence provides ample sequence divergence to allow the development of internal probes specific to streptococci. The Streptococcus-specific assay was highly sensitive for all 28 streptococcal species tested (i.e., detection limit of 1 to 10 genome copies per PCR). The tuf sequence data was also used to perform extensive phylogenetic analysis, which was generally in agreement with phylogeny determined on the basis of 16S rRNA gene data. However, the tuf gene provided a better discrimination at the streptococcal species level that should be particularly useful for the identification of very closely related species. In conclusion, tuf appears more suitable than the 16S ribosomal RNA gene for the development of diagnostic assays for the detection and identification of streptococcal species because of its higher level of species-specific genetic divergence.