Salivary IgG Research Articles

PSV-10 Vaccination of beef cattle to reduce enteric methane emissions

Abstract Livestock methane (CH4) emissions total over 3 billion tonnes per year of carbon dioxide equivalents (CO2e) and are responsible for approximately 6% of total annual greenhouse gas emissions. The contribution of livestock CH4 emissions are only to be exacerbated as the global demand for meat and dairy products increases. Greater than 75% of livestock CH4 emissions are generated in dispersed production environments (e.g., cow-calf and stocker segments), which are not addressable by methane mitigation strategies requiring constant inputs. Thus, strategies that fit into standard agronomic practices, in particular dispersed production environments, are urgently needed to address the increasing carbon footprint associated with livestock production. Methane-reducing vaccines are a promising solution for addressing this need, due to their longevity of action, low cost, and ease of integration into standard agronomic practices, which could lower the barrier for adoption. Thus, this study aimed to assess the effects of a prototype vaccine on total and antigen-specific immune response, CH4 yield, and average daily gain (ADG) in cattle. Angus crossbred steers [n = 20; initial body weight (BW) = 537 ± 15 kg) fed on a high-forage diet were randomly assigned to either placebo- or vaccine-treated groups blocked by BW, breed, and feed intake. Steers were subcutaneously inoculated (2 mL dose) in the anterior region of the neck. Blood and saliva samples were collected, and total sera and salivary immunoglobulin (Ig) G and IgA were quantified using ELISA (Bethyl laboratories, Montgomery, TX). Antigen-specific sera IgG was measured using ELISA. Daily CH4 emissions and dry matter intake (DMI) were measured using GreenFeed systems (C-Lock Inc. Rapid City, SD) and GrowSafe feed bunks (Vytelle, Lenexa, KS), respectively. Body weights were recorded bi-weekly. Total sera IgA and IgG did not differ between the placebo-treated and vaccinated steers (P = 0.584 and P = 0.425, respectively), nor did total salivary IgA and IgG (P = 0.577 and P = 0.548, respectively). However, antigen-specific sera IgG did significantly increase following booster vaccination in the vaccinated steers, as compared with placebo-treated steers (d 0 vs. d 28; P < 0.001). CH4 yield was significantly reduced in vaccinated steers following booster vaccination despite significantly increasing in placebo-treated steers over the same period (P = 0.002). This reduction in CH4 yield was not associated with a reduction in DMI nor ADG (P = 0.181 and P = 0.314, respectively), suggesting that vaccination did not negatively affect treated animals. These findings underscore the potential of vaccine-based solutions to mitigate the environmental impact of livestock CH4 emissions, especially from dispersed production environments where current options for mitigation strategies are limited.

Salivary immune responses after COVID-19 vaccination.

mRNA-based COVID-19 vaccines have played a critical role in reducing severe outcomes of COVID-19. Humoral immune responses against SARS-CoV-2 after vaccination have been extensively studied in blood; however, limited information is available on the presence and duration of SARS-CoV-2 specific antibodies in saliva and other mucosal fluids. Saliva offers a non-invasive sampling method that may also provide a better understanding of mucosal immunity at sites where the virus enters the body. Our objective was to evaluate the salivary immune response after vaccination with the COVID-19 Moderna mRNA-1273 vaccine. Two hundred three staff members of the U.S. Centers for Disease Control and Prevention were enrolled prior to receiving their first dose of the mRNA-1273 vaccine. Participants were asked to self-collect 6 saliva specimens at days 0 (prior to first dose), 14, 28 (prior to second dose), 42, and 56 using a SalivaBio saliva collection device. Saliva specimens were tested for anti-spike protein SARS-CoV-2 specific IgA and IgG enzyme immunoassays. Overall, SARS-CoV-2-specific salivary IgA titers peaked 2 weeks after each vaccine dose, followed by a sharp decrease during the following weeks. In contrast to IgA titers, IgG antibody titers increased substantially 2 weeks after the first vaccine dose, peaked 2 weeks after the second dose and persisted at an elevated level until at least 8 weeks after the first vaccine dose. Additionally, no significant differences in IgA/IgG titers were observed based on age, sex, or race/ethnicity. All participants mounted salivary IgA and IgG immune responses against SARS-CoV-2 after receiving the mRNA-1273 COVID-19 vaccine. Because of the limited follow-up time for this study, more data are needed to assess the antibody levels beyond 2 months after the first dose. Our results confirm the potential utility of saliva in assessing immune responses elicited by immunization and possibly by infection.

The impact of vaccine type and booster dose on the magnitude and breadth of SARS-CoV-2-specific systemic and mucosal antibodies among COVID-19 vaccine recipients

The COVID-19 pandemic has had a major impact on global health and economy, which was significantly mitigated by the availability of COVID-19 vaccines. The levels of systemic and mucosal antibodies against SARS-CoV-2 correlated with protection. However, there is limited data on how vaccine type and booster doses affect mucosal antibody response, and how the breadth of mucosal and systemic antibodies compares. In this cross-sectional study, we compared the magnitude and breadth of mucosal and systemic antibodies in 108 individuals who received either the BNT162b2 (Pfizer) or CoronaVac (SinoVac) vaccine. We found that BNT162b2 (vs CoronaVac) or booster doses (vs two doses) were significantly associated with higher serum IgG levels, but were not significantly associated with salivary IgA levels, regardless of prior infection status. Among non-infected individuals, serum IgG, serum IgA and salivary IgG levels were significantly higher against the ancestral strain than the Omicron BA.2 sublineage, but salivary IgA levels did not differ between the strains. Salivary IgA had the weakest correlation with serum IgG (r = 0.34) compared with salivary IgG (r = 0.63) and serum IgA (r = 0.60). Our findings suggest that intramuscular COVID-19 vaccines elicit a distinct mucosal IgA response that differs from the systemic IgG response. As mucosal IgA independently correlates with protection, vaccine trials should include mucosal IgA as an outcome measure.

In silico study to explore the mechanism of Toxoplasma-induced inflammation and target therapy based on sero and salivary Toxoplasma

We aimed to assess salivary and seroprevalence of Toxoplasma immunoglobulins in risky populations and evaluate drug docking targeting TgERP. A cross-sectional study was conducted in Alexandria University hospitals’ outpatient clinics. 192 participants were enrolled from September 2022 to November 2023. Anti-Toxoplasma IgG and IgM were determined in serum and saliva by ELISA. An in-Silico study examined TgERP’s protein–protein interactions (PPIs) with pro-inflammatory cytokine receptors, anti-inflammatory cytokine, cell cycle progression regulatory proteins, a proliferation marker, and nuclear envelope integrity-related protein Lamin B1. Our findings revealed that anti-T. gondii IgG were detected in serum (66.1%) and saliva (54.7%), with 2.1% of both samples were positive for IgM. Salivary IgG had 75.59% sensitivity, 86.15% specificity, 91.40% PPV, 64.40% NPP, 79.17% accuracy and fair agreement with serum IgG. On the other hand, the sensitivity, specificity, PPV, NPV, and accuracy in detecting salivary IgM were 75.0%, 99.47%, 75.0%, 99.47%, and 98.96%. AUC 0.859 indicates good discriminatory power. Examined synthetic drugs and natural products can target specific amino acids residues of TgERP that lie at the same binding interface with LB1 and Ki67, subsequently, hindering their interaction. Hence, salivary samples can be a promising diagnostic approach. The studied drugs can counteract the pro-inflammatory action of TgERP.

Serum and Salivary IgG and IgA Response After COVID-19 Messenger RNA Vaccination

There is still considerable controversy in the literature regarding the capacity of intramuscular messenger RNA (mRNA) vaccination to induce a mucosal immune response. To compare serum and salivary IgG and IgA levels among mRNA-vaccinated individuals with or without previous SARS-CoV-2 infection. In this cohort study, SARS-CoV-2-naive participants and those with previous infection were consecutively included in the CoviCompare P and CoviCompare M mRNA vaccination trials and followed up to day 180 after vaccination with either the BNT162b2 (Pfizer-BioNTech) vaccine or the mRNA-1273 (Moderna) vaccine at the beginning of the COVID-19 vaccination campaign (from February 19 to June 8, 2021) in France. Data were analyzed from October 25, 2022, to July 13, 2023. An ultrasensitive digital enzyme-linked immunosorbent assay was used for the comparison of SARS-CoV-2 spike-specific serum and salivary IgG and IgA levels. Spike-specific secretory IgA level was also quantified at selected times. A total of 427 individuals were included in 3 groups: participants with SARS-CoV-2 prior to vaccination who received 1 single dose of BNT162b2 (Pfizer-BioNTech) (n = 120) and SARS-CoV-2-naive individuals who received 2 doses of mRNA-1273 (Moderna) (n = 172) or 2 doses of BNT162b2 (Pfizer-BioNTech) (n = 135). The median age was 68 (IQR, 39-75) years, and 228 (53.4%) were men. SARS-CoV-2 spike-specific IgG saliva levels increased after 1 or 2 vaccine injections in individuals with previous infection and SARS-CoV-2-naive individuals. After vaccination, SARS-CoV-2-specific saliva IgA levels, normalized with respect to total IgA levels, were significantly higher in participants with previous infection, as compared with the most responsive mRNA-1273 (Moderna) recipients (median normalized levels, 155 × 10-5 vs 37 × 10-5 at day 29; 107 × 10-5 vs 54 × 10-5 at day 57; and 104 × 10-5 vs 70 × 10-5 at day 180 [P < .001]). In contrast, compared with day 1, spike-specific IgA levels in the BNT162b2-vaccinated SARS-CoV-2-naive group increased only at day 57 (36 × 10-5 vs 49 × 10-5 [P = .01]). Bona fide multimeric secretory IgA levels were significantly higher in individuals with previous infection compared with SARS-CoV-2-naive individuals after 2 antigenic stimulations (median optical density, 0.36 [IQR, 0.16-0.63] vs 0.16 [IQR, 0.10-0.22]; P < .001). The findings of this cohort study suggest that mRNA vaccination was associated with mucosal immunity in individuals without prior SARS-CoV-2 infection, but at much lower levels than in previously infected individuals. Further studies are needed to determine the association between specific saliva IgA levels and prevention of infection or transmission.

Detection of SARS-CoV-2-specific mucosal antibodies in saliva following concomitant COVID-19 and influenza vaccination in the ComFluCOV trial

The ComFluCOV trial randomized 679 participants to receive an age-appropriate influenza vaccine, or placebo, alongside their second COVID-19 vaccine. Concomitant administration was shown to be safe, and to preserve systemic immune responses to both vaccines. Here we report on a secondary outcome of the trial investigating SARS-CoV-2-specific mucosal antibody responses. Anti-spike IgG and IgA levels in saliva were measured with in-house ELISAs. Concomitant administration of an influenza vaccine did not affect salivary anti-spike IgG positivity rates to Pfizer/BioNTech BNT162b2 (99.1 cf. 95.6%), or AstraZeneca ChAdOx1 (67.8% cf. 64.9%), at 3-weeks post-vaccination relative to placebo. Furthermore, saliva IgG positively correlated with serum titres highlighting the potential utility of saliva for assessing differences in immunogenicity in future vaccine studies. Mucosal IgA was not detected in response to either COVID-19 vaccine, reinforcing the need for novel vaccines capable of inducing sterilising immunity or otherwise reducing transmission. The trial is registered as ISRCTN 14391248.

Comparison of Omicron breakthrough infection versus monovalent SARS-CoV-2 intramuscular booster reveals differences in mucosal and systemic humoral immunity

Our understanding of the quality of cellular and humoral immunity conferred by coronavirus disease 2019 (COVID-19) vaccination alone versus vaccination plus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) breakthrough (BT) infection remains incomplete. While the current (2023) SARS-CoV-2 immune landscape of Canadians is complex, in late 2021 most Canadians had either just received a third dose of COVID-19 vaccine, or had received their two dose primary series and then experienced an Omicron BT. Herein we took advantage of this coincident timing to contrast cellular and humoral immunity conferred by three doses of vaccine versus two doses plus BT. Our results show that mild BT infection induces cell-mediated immune responses to variants comparable to an intramuscular vaccine booster dose. In contrast, BT subjects had higher salivary IgG and IgA levels against the Omicron spike and enhanced reactivity to the ancestral spike for the IgA isotype, which also reacted with SARS-CoV-1. Serum neutralizing antibody levels against the ancestral strain and the variants were also higher after BT infection. Our results support the need for vaccines that emulate the enhanced mucosal and humoral immunity induced by Omicron BT without exposing individuals to the risks associated with SARS-CoV-2 infection.One Sentence SummaryOmicron breakthrough elicits cross-reactive systemic and mucosal immune responses in vaccinated adults.

Detection of Specific Immunoglobulins in the Saliva of Patients With Mild COVID-19.

Saliva has many advantages over blood as a biofluid, so using it for measuring and monitoring antibody responses in COVID-19 would be highly valuable. To assess the value of saliva-based IgG and IgM/IgA antibody testing in COVID-19, this cross-sectional pilot study evaluated the accuracy of salivary and serum IgG and IgM/IgA for detecting mild COVID-19 and their correlation. Fifty-one patients with mild COVID-19 (14-28 days post-symptom onset) were included in the study. Enzyme-linked immunosorbent assays (ELISA) were used to measure IgG and IgM/IgA responses to SARS-CoV-2 spike protein in both serum and saliva samples using a slightly modified protocol for saliva samples. Saliva-based IgG testing had 30% sensitivity and 100% specificity, with a positive predictive value (PPV) of 100% and a negative predictive value (NPV) of 50%. Saliva-based IgM/IgA testing had 13.2% sensitivity and 100% specificity, with a PPV of 100% and an NPV of 28.3%. Blood and saliva IgG values were positively correlated. Saliva currently has limited diagnostic value for COVID-19 testing, at least for mild disease. Nevertheless, the significant positive correlation between blood and saliva IgG titers indicates that saliva might be a complementary biofluid for assessing systemic antibody responses to the virus, especially if the assay is further optimized across the full disease spectrum.

Clinical and Analytical Performance of ELISA Salivary Serologic Assay to Detect SARS-CoV-2 IgG in Children and Adults.

Saliva is a promising matrix with several purposes. Our aim is to verify if salivary anti-SARS-CoV-2 antibody determination is suitable for monitoring immune responses. One hundred eighty-seven subjects were enrolled at University-Hospital Padova: 105 females (56.1%) and 82 males (43.9%), 95 (50.8%) children and 92 (49.2%) adults. Subjects self-collected saliva using Salivette; nineteen subjects collected three different samples within the day. A serum sample was obtained for all individuals. The N/S anti-SARS-CoV-2 salivary IgG (sal-IgG) and serum anti-SARS-CoV-2 S-RBD IgG (ser-IgG) were used for determining anti-SARS-CoV-2 antibodies. The mean (min-max) age was 9.0 (1-18) for children and 42.5 (20-61) for adults. Of 187 samples, 63 were negative for sal-IgG (33.7%), while 7 were negative for ser-IgG (3.7%). Spearman's correlation was 0.56 (p < 0.001). Sal-IgG and ser-IgG levels were correlated with age but not with gender, comorbidities, prolonged therapy, previous SARS-CoV-2 infection, or time from last COVID-19 infection/vaccination. The repeatability ranged from 23.8% (7.4 kAU/L) to 4.0% (3.77 kAU/L). The linearity of the assay was missed in 4/6 samples. No significant intrasubject differences were observed in sal-IgG across samples collected at different time points. Sal-IgG has good agreement with ser-IgG. Noninvasive saliva collection represents an alternative method for antibody measurement, especially in children.

Tixagevimab/cilgavimab for the prevention of COVID-19 in vaccine-refractory patients with autoimmune diseases: a prospective cohort study.

To investigate the effects of passive immunization with the anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) monoclonal antibodies tixagevimab/cilgavimab on humoral responses and on coronavirus disease 2019 (COVID-19) outcomes in vaccine-refractory patients with immune-mediated inflammatory diseases (IMIDs) at high risk of severe COVID-19. A prospective cohort study was performed on a cohort of high-risk vaccine-refractory IMID patients treated with a single dose of tixagevimab/cilgavimab (150 mg/150 mg). COVID-19 outcomes as well as serum and salivary anti-SARS-CoV-2 IgG were assessed at baseline and for at least 6 months. Results were compared with an untreated high-risk vaccine-refractory IMID population. Standardized incidence ratios (SIRs) of COVID-19 compared with the general population were calculated for both groups. A total of 38 high-risk IMID patients received tixagevimab/cilgavimab and were compared with 114 untreated high-risk IMID controls. Serum anti-spike IgG increased to 6.6 OD (s.d. 0.8) at day 1 and remained positive up to month 6 [6.3 OD (s.d. 1.4)]. Salivary anti-spike IgG peaked at month 2 [1.6 OD (s.d. 1.1)] and decreased from month 3 [0.8 OD (s.d. 0.3)]. No severe or extended infection was observed in the tixagevimab/cilgavimab group. Compared with the general population, the SIR of COVID-19 in treated patients was 0.76 (95% CI 0.24, 1.58) despite the increased risk profile. The SIR of the control group was 1.51 (95% CI 1.07, 2.02), corresponding to a significantly increased incidence. Passive immunization with tixagevimab/cilgavimab is safe and effective in inducing anti-SARS-CoV-2 immunity and potentially in preventing COVID-19 in high-risk vaccine-refractory IMID patients. These data provide a proof of concept for the use of monoclonal antibodies as a preventative strategy against SARS-CoV-2 in vulnerable populations.

Sensitive SARS-CoV-2 salivary antibody assays for clinical saline gargle samples using smartphone-based competitive particle immunoassay platforms

Antibody assay for SARS-CoV-2 has become increasingly important to track latent and asymptomatic infections, check the individual's immune status, and confirm vaccine efficacy and durability. However, current SARS-CoV-2 antibody assays require invasive blood collection, requiring a remote laboratory and a trained phlebotomist. Direct detection of SARS-CoV-2 antibodies from clinical saline gargle samples has been considered challenging due to the smaller number of antibodies in such specimens and the high limit of detection of currently available rapid tests. This work demonstrates simple and non-invasive methods for detecting SARS-CoV-2 salivary antibodies. Competitive particle immunoassays were developed on a paper microfluidic chip using the receptor-binding domain (RBD) antigens on spike proteins. Using a smartphone, they were monitored by counting the captured fluorescent particles or evaluating the capillary flow velocities. The limit of detection (LOD), cross-binding between alpha- and omicron-strains, and the effect of angiotensin-converting enzyme 2 (ACE2) presence were investigated. LODs were 1-5ng/mL in both 10% and 1% saliva. Clinical saline gargle samples were assayed using both methods, showing a statistical difference between virus-negative and virus-positive samples, although the assays targeted antibodies. Only a small number of virus-positive samples were antibody-negative. The high assay sensitivity detected a small number of antibodies developed even during the early phase of infections. Overall, this work demonstrates the ability to detect SARS-CoV-2 salivary IgG antibodies on simple, cost-effective, portable platforms towards mitigating SARS-CoV-2 and potentially other respiratory viruses.

Omicron Breakthrough Infection Elicits Superior Humoral and Mucosal Immunity to SARS-CoV-2 Variants than Booster Doses

Abstract In light of ongoing booster vaccination campaigns, one recurring question is whether individuals infected with SARS-CoV-2 should still receive a booster dose if they were infected after completing their primary series. We aimed to assess the intensity of functional humoral and cellular immune responses induced by breakthrough infections during the B.1.1.529 wave in Canada compared to individuals who received three doses of vaccine. Because hybrid immunity has been shown to induce particularly potent immunity, we hypothesized that breakthrough infections could induce superior immune response to variants than booster doses. To address this important public health question, we analyzed samples from two cohorts of individuals naïve to SARS-CoV-2 or who were primary infected before vaccination. We compared responses 45 days after a third vaccine dose or an Omicron breakthrough infection. Our results show that mild breakthrough infection induces cell-mediated immune response to variants comparable to a vaccine booster dose. In contrast, breakthrough subjects had higher salivary IgG and IgA levels against the Omicron Spike and enhanced reactivity to the ancestral Spike for the IgA isotype, which also reacted with SARS-CoV-1. Serum neutralizing antibody levels against the ancestral strain and the variants were also higher after breakthrough infection, especially in individuals naïve to the infection prior to vaccination. Altogether our results demonstrate that an Omicron breakthrough infection can substitute a booster dose of vaccine, while also supporting the need for mucosal vaccine boosts to potentially replicate the improvement on mucosal and humoral immunity without the risks associated to being infected with SARS-CoV-2. Research supported by grants from the Canadian Institutes of Health Research, Public Health Agency of Canada and the Ontario COVID-19 Rapid Research Fund through Ontario Together.

Comparative ELISA Detection of SARS-COV-2 Monoclonal Antibodies in Patients` Serum, Saliva, and Nasal Fluid in Iraq

Precise management of COVID-19 infection and global pandemic requires a precise assay of SARS-COV-2 antibodies. Spike protein and RBD specific serum monoclonal antibodies has been known to be elicited with the currently approved mRNA vaccine for COVID-19. Since combating SARS-CoV-2 infection begins as early at the viral entry port, mediated by mucosal immunity; whether the currently approved vaccines have the capacity to elicit mucosal immunity is still to be studied. Method; We conducted our research to detect SARS-Cov-2 IgA-RBD and IgG-NCP, in a set of a randomized three patients’ groups (total 55 subjects, randomized into 20 (Non previously infected control, 20 fully vaccinated people from AlSalam teaching hospital vaccination ward upon receiving the vaccine doses, and 15 patients newly recovered from SARS-CoV-2 infection from AlShifaa quarantine hospital after remission from COVID-19 infection and preparing to discharge) in three fluid samples serum, saliva and nasal fluid, for a total of three samples per participant, to make a total of 165 samples to be tested, using a commercial immunoassay kit (Anti-SARS-CoV-2 ELISA RBD IgA, and Anti-SARS- CoV-2 NCP ELISA IgG) the study was designed to assess the immunoglobulins levels in each sample from different health status and anatomical locations to give the statistical difference and the correct conclusion regarding this study. Statistical analysis was performed using SPSS version 24, as well as Microsoft excel version (16.0.15028), test equations were performed via Mann-Whitney U Test, as well as The Kruskal-Wallis H test, to evaluate the non-gaussian distribution among the study group, for each of the outcomes, p &lt; 0.05 was undertaken as statistically significant. Results; Our work showed that two doses of mRNA BNT162b2 vaccine Comirnaty (Pfizer/BioNTech, New York, NY, USA), elicited a robust anti SARS-CoV-2 nasal and salivary IgA and IgG monoclonal antibody protection (as compared to control 0.625 COI, the vaccinated subjects resulted in a higher median salivary concentration of IgG NCP values as 1.69 COI, with a p-value of 0.0001. same goes for median salivary concentration of IgA RBD with value of 2.875 ± 1.276, as compared to control median values of 0.667 ± 0.208), (as compared to control 0.462 ± 0.2369, the vaccinated subjects resulted in a higher median nasal concentration of IgG NCP values as 1.944 ± 0.694, with a p-value of 0.0001. Same goes for median nasal concentration of IgA RBD with value of 1.941 ± 0.53, as compared to control median values of 0.681 ± 0.226). Two injections 25 days a part shown to trigger a stronger titer of protective immunity as compared to single early shot, still it`s less robust compared to the titers measured for the recovered subjects from COVID-19 infection. Taking into consideration that there was a lack of trustable ELISA based assays with specially designed kit for this purpose, focusing on nasal and salivary secretions, the current study that our pre investigational and investigational data are consistent. The results of this study indicates that a protective antigen specific nasal and salivary monoclonal antibody, can be triggered following proper vaccination regimen with mRNA COVID-19 vaccine, so this paves the way for using mucosal protective antibodies detection kits, as a suitable alternative option as a less invasive method compared to serum investigations, for detecting the protection against SARS-Cov-2 infection, induced by vaccine. The vaccinated subjects are significantly better tested via IgA in Saliva as compared to IgG, because of the higher median values for the IgA vs IgG, 2.875 ± 1.276 and 2.083 ± 1.610 respectively with a p-value of 0.008. in a similar state the IgA is the best test for the convalescent subjects in term of nasal fluid samples investigations, as the median value are comparatively higher than median value of IgG, 2.024 ± 0.520 and 1.786 ± 1.115 respectively.

Asociaciones de hallazgos diagnósticos con actividad de la enfermedad en el síndrome de Sjögren primario: un análisis de conglomerados

ObjectiveThe diagnosis of primary Sjögren's syndrome still relies upon a constellation of clinical, laboratory, imaging, and pathological findings. We aimed to evaluate the relation of the disease activity with the results of diagnostic tests for primary Sjögren's syndrome. MethodsA principal component with cluster analysis was performed to classify 69 patients with primary Sjögren's syndrome based on the results of diagnostic evaluations. ResultsAnti-SSA autoantibody was the most represented feature on the principal components. The anti-SSA and ultrasound score were positively correlated (p=0.001). We identified two distinct clusters of low or high disease activity (p<0.001). Except for disease duration and serum beta2-microglobulin, the clusters were significantly different in salivary flow (p=0.004), ultrasound findings (p<0.001), IgG (p=0.001), and salivary beta2-microglobulin (p=0.048). Also, positive findings were significantly different between the clusters in rheumatoid factor, antinuclear antibody, anti-SSA, and anti-SSB (all p≤0.013). ConclusionPatients with higher syndrome activity were best recognized with serological and ultrasound assessments. However, patients with lower syndrome activity had a longer disease duration, higher stimulated salivary flow rate, and a positive biopsy of minor salivary glands (56%).

Evaluation and deployment of isotype-specific salivary antibody assays for detecting previous SARS-CoV-2 infection in children and adults

BackgroundSaliva is easily obtainable non-invasively and potentially suitable for detecting both current and previous SARS-CoV-2 infection, but there is limited evidence on the utility of salivary antibody testing for community surveillance.MethodsWe established 6 ELISAs detecting IgA and IgG antibodies to whole SARS-CoV-2 spike protein, to its receptor binding domain region and to nucleocapsid protein in saliva. We evaluated diagnostic performance, and using paired saliva and serum samples, correlated mucosal and systemic antibody responses. The best-performing assays were field-tested in 20 household outbreaks.ResultsWe demonstrate in test accuracy (N = 320), spike IgG (ROC AUC: 95.0%, 92.8–97.3%) and spike IgA (ROC AUC: 89.9%, 86.5–93.2%) assays to discriminate best between pre-pandemic and post COVID-19 saliva samples. Specificity was 100% in younger age groups (0–19 years) for spike IgA and IgG. However, sensitivity was low for the best-performing assay (spike IgG: 50.6%, 39.8–61.4%). Using machine learning, diagnostic performance was improved when a combination of tests was used. As expected, salivary IgA was poorly correlated with serum, indicating an oral mucosal response whereas salivary IgG responses were predictive of those in serum. When deployed to household outbreaks, antibody responses were heterogeneous but remained a reliable indicator of recent infection. Intriguingly, unvaccinated children without confirmed infection showed evidence of exposure almost exclusively through specific IgA responses.ConclusionsThrough robust standardisation, evaluation and field-testing, this work provides a platform for further studies investigating SARS-CoV-2 transmission and mucosal immunity with the potential for expanding salivo-surveillance to other respiratory infections in hard-to-reach settings.

Short-Lived Antibody-Mediated Saliva Immunity against SARS-CoV-2 after Vaccination.

Knowledge about the effect of vaccination against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on immunity reflected in the saliva is sparse. We examined the antibody response in saliva compared to that in serum 2 and 6 months after the first vaccination with the BNT162b2 vaccine. Four hundred fifty-nine health care professionals were included in a prospective observational study measuring antibody levels in saliva and corresponding serum samples at 2 and 6 months after BNT162b2 vaccination. Vaccinated, previously SARS-CoV-2-infected individuals (hybrid immunity) had higher IgG levels in saliva at 2 months than vaccinated, infection-naive individuals (P < 0.001). After 6 months, saliva IgG levels declined in both groups (P < 0.001), with no difference between groups (P = 0.37). Furthermore, serum IgG levels declined from 2 to 6 months in both groups (P < 0.001). IgG antibodies in saliva and serum correlated in individuals with hybrid immunity at 2 and 6 months (ρ = 0.58, P = 0.001, and ρ = 0.53, P = 0.052, respectively). In vaccinated, infection-naive individuals, a correlation was observed at 2 months (ρ = 0.42, P < 0.001) but not after 6 months (ρ = 0.14, P = 0.055). IgA and IgM antibodies were hardly detectable in saliva at any time point, regardless of previous infection. In serum, IgA was detected at 2 months in previously infected individuals. BNT162b2 vaccination induced a detectable IgG anti-SARS-CoV-2 RBD response in saliva at both 2 and 6 months after vaccination, being more prominent in previously infected than infection-naive individuals. However, a significant decrease in salivary IgG was observed after 6 months, suggesting a rapid decline in antibody-mediated saliva immunity against SARS-CoV-2, after both infection and systemic vaccination. IMPORTANCE Knowledge about the persistence of salivary immunity after SARS-CoV-2 vaccination is limited, and information on this topic could prove important for vaccine strategy and development. We hypothesized that salivary immunity would wane rapidly after vaccination. We measured anti-SARS-CoV-2 IgG, IgA, and IgM concentrations in saliva and serum in both previously infected and infection-naive individuals, 2 and 6 months after first vaccination with BNT162b2, in 459 hospital employees from Copenhagen University Hospital. We observed that IgG was the primary salivary antibody 2 months after vaccination in both previously infected and infection-naive individuals, but dropped significantly after 6 months. Neither IgA nor IgM was detectable in saliva at either time point. Findings indicate that salivary immunity against SARS-CoV-2 rapidly declines following vaccination in both previously infected and infection-naive individuals. We believe this study shines a light on the workings of salivary immunity after SARS-CoV-2 infection, which could prove relevant for vaccine development.

Associations of diagnostic findings with disease activity in primary Sjӧgren's syndrome: a cluster analysis

ObjectiveThe diagnosis of primary Sjӧgren's syndrome still relies upon a constellation of clinical, laboratory, imaging, and pathological findings. We aimed to evaluate the relation of the disease activity with the results of diagnostic tests for primary Sjӧgren's syndrome. MethodsA principal component with cluster analysis was performed to classify 69 patients with primary Sjӧgren's syndrome based on the results of diagnostic evaluations. ResultsAnti-SSA autoantibody was the most represented feature on the principal components. The anti-SSA and ultrasound score were positively correlated (p=0.001). We identified two distinct clusters of low or high disease activity (p<0.001). Except for disease duration and serum beta2-microglobulin, the clusters were significantly different in salivary flow (p= 0.004), ultrasound findings (p<0.001), IgG (p= 0.001), and salivary beta2-microglobulin (p= 0.048). Also, positive findings were significantly different between the clusters in rheumatoid factor, antinuclear antibody, anti-SSA, and anti-SSB (all p≤0.013). ConclusionPatients with higher syndrome activity were best recognized with serological and ultrasound assessments. However, patients with lower syndrome activity had a longer disease duration, higher stimulated salivary flow rate, and a positive biopsy of minor salivary glands (56%).

Methodological approaches to optimize multiplex oral fluid SARS-CoV-2 IgG assay performance and correlation with serologic and neutralizing antibody responses

BackgroundOral fluid (hereafter, saliva) is a non-invasive and attractive alternative to blood for SARS-CoV-2 IgG testing; however, the heterogeneity of saliva as a matrix poses challenges for immunoassay performance. ObjectivesTo optimize performance of a magnetic microparticle-based multiplex immunoassay (MIA) for SARS-CoV-2 IgG measurement in saliva, with consideration of: i) threshold setting and validation across different MIA bead batches; ii) sample qualification based on salivary total IgG concentration; iii) calibration to U.S. SARS-CoV-2 serological standard binding antibody units (BAU); and iv) correlations with blood-based SARS-CoV-2 serological and neutralizing antibody (nAb) assays. MethodsThe salivary SARS-CoV-2 IgG MIA included 2 nucleocapsid (N), 3 receptor-binding domain (RBD), and 2 spike protein (S) antigens. Gingival crevicular fluid (GCF) swab saliva samples were collected before December 2019 (n = 555) and after molecular test-confirmed SARS-CoV-2 infection from 113 individuals (providing up to 5 repeated-measures; n = 398) and used to optimize and validate MIA performance (total n = 953). Combinations of IgG responses to N, RBD and S and total salivary IgG concentration (μg/mL) as a qualifier of nonreactive samples were optimized and validated, calibrated to the U.S. SARS-CoV-2 serological standard, and correlated with blood-based SARS-CoV-2 IgG ELISA and nAb assays. ResultsThe sum of signal to cutoff (S/Co) to all seven MIA SARS-CoV-2 antigens and disqualification of nonreactive saliva samples with ≤15 μg/mL total IgG led to correct classification of 62/62 positives (sensitivity [Se] = 100.0%; 95% confidence interval [CI] = 94.8%, 100.0%) and 108/109 negatives (specificity [Sp] = 99.1%; 95% CI = 97.3%, 100.0%) at 8-million beads coupling scale and 80/81 positives (Se = 98.8%; 95% CI = 93.3%, 100.0%] and 127/127 negatives (Sp = 100%; 95% CI = 97.1%, 100.0%) at 20-million beads coupling scale. Salivary SARS-CoV-2 IgG crossed the MIA cutoff of 0.1 BAU/mL on average 9 days post-COVID-19 symptom onset and peaked around day 30. Among n = 30 matched saliva and plasma samples, salivary SARS-CoV-2 MIA IgG levels correlated with corresponding-antigen plasma ELISA IgG (N: ρ = 0.76, RBD: ρ = 0.83, S: ρ = 0.82; all p < 0.001). Correlations of plasma SARS-CoV-2 nAb assay area under the curve (AUC) with salivary MIA IgG (N: ρ = 0.68, RBD: ρ = 0.78, S: ρ = 0.79; all p < 0.001) and with plasma ELISA IgG (N: ρ = 0.76, RBD: ρ = 0.79, S: ρ = 0.76; p < 0.001) were similar. ConclusionsA salivary SARS-CoV-2 IgG MIA produced consistently high Se (> 98.8%) and Sp (> 99.1%) across two bead coupling scales and correlations with nAb responses that were similar to blood-based SARS-CoV-2 IgG ELISA data. This non-invasive salivary SARS-CoV-2 IgG MIA could increase engagement of vulnerable populations and improve broad understanding of humoral immunity (kinetics and gaps) within the evolving context of booster vaccination, viral variants and waning immunity.

Salivary IgA and IgG Antibody Responses against Periodontitis-Associated Bacteria in Crohn's Disease.

Elevated serum immunoglobulin (Ig) antibody levels are observed in Crohn's disease patients. The aim of this study was to evaluate the salivary IgA and IgG antibody levels against Porphyromonas gingivalis, Tannerella forsythia, Aggregatibacter actinomycetemcomitans, and Prevotella intermedia in Crohn's disease patients. Eighty-eight participants (47 Crohn's disease patients and 41 systemically healthy age- and gender-matched controls) were included in the study. Oral and medical health statuses were recorded and salivary samples were collected. Salivary P. gingivalis, T. forsythia, A. actinomycetemcomitans, and P. intermedia carriage were analyzed with DNA sequencing technique, salivary levels of IgG1, IgG2, IgG3, IgG4, and IgM were measured with the Luminex® xMAP™ technique, and salivary IgA and IgG antibody levels against P. gingivalis, T. forsythia, A. actinomycetemcomitans, and P. intermedia were detected by ELISA. As result, higher salivary IgG2 (p = 0.011) and IgG3 (p = 0.006), P. gingivalis IgA (p < 0.001), A. actinomycetemcomitans IgG (p = 0.001), and P. intermedia IgG (p < 0.001) antibody levels were detected in the Crohn's disease group compared to the controls. Salivary P. gingivalis carriage was lower in the Crohn's disease group in comparison to the controls (p = 0.024). In conclusion, salivary IgA antibody responses against P. gingivalis and IgG antibody responses against P. intermedia have independent associations with Crohn's disease.

Point-of-Care Diagnostic Biosensors to Monitor Anti-SARS-CoV-2 Neutralizing IgG/sIgA Antibodies and Antioxidant Activity in Saliva.

Monitoring biomarkers is a great way to assess daily physical condition, and using saliva instead of blood samples is more advantageous as the process is simple and allows individuals to test themselves. In the present study, we analyzed the titers of neutralizing antibodies, IgG and secretory IgA (sIgA), in response to the SARS-CoV-2 vaccine, in saliva. A total of 19 saliva and serum samples were collected over a 10-month period 3 weeks after the first vaccine, 8 months after the second vaccine, and 1 month after the third vaccine. The ranges of antibody concentrations post-vaccination were: serum IgG: 81-15,000 U/mL, salivary IgG: 3.4-330 U/mL, and salivary IgA: 58-870 ng/mL. A sharp increase in salivary IgG levels was observed after the second vaccination. sIgA levels also showed an increasing trend. A correlation with trends in serum IgG levels was observed, indicating the possibility of using saliva to routinely assess vaccine efficacy. The electrochemical immunosensor assay developed in this study based on the gold-linked electrochemical immunoassay, and the antioxidant activity measurement based on luminol electrochemiluminescence (ECL), can be performed using portable devices, which would prove useful for individual-based diagnosis using saliva samples.