Comparison of SARS-CoV-2 spike-specific IgA and IgG in nasal secretions, saliva and serum.

Introduction: Several novel vaccine platforms aim at mucosal immunity in the respiratory tract to block SARS-CoV-2 transmission. Standardized methods for mucosal sample collection and quantification of mucosal antibodies are therefore urgently needed for harmonized comparisons and interpretations across mucosal vaccine trials and real-world data. Methods: Using commercial electrochemiluminescence antibody panels, we compared SARS-CoV-2 spike-specific IgA and IgG in paired saliva, nasal secretions, and serum from 1048 healthcare workers with and without prior infection. Results: Spike-specific IgA correlated well in nasal secretions and saliva (r>0.65, p<0.0001), but the levels were more than three-fold higher in nasal secretions as compared to in saliva (p<0.01). Correlations between the total population of spike-specific IgA and spike-specific secretory IgA (SIgA) were significantly stronger (p<0.0001) in nasal secretions (r=0.96, p<0.0001) as opposed to in saliva (r=0.77, p<0.0001), and spike-specific IgA correlated stronger (p<0.0001) between serum and saliva (r=0.73, p<0.001) as opposed to between serum and nasal secretions (r=0.54, p<0.001), suggesting transudation of monomeric spike specific IgA from the circulation to saliva. Notably, spike-specific SIgA had a markedly higher SARS-CoV-2 variant cross-binding capacity as compared to the total population of spike specific IgA and IgG in both nasal secretions, saliva and serum, (all p<0.0001), which emphasizes the importance of taking potential serum derived monomeric IgA into consideration when investigating mucosal immune responses. Discussion: Taken together, although spike-specific IgA can be reliably measured in both nasal secretions and saliva, our findings imply an advantage of higher levels and likely also a larger proportion of SIgA in nasal secretions as compared to in saliva. We further corroborate the superior variant cross-binding capacity of SIgA in mucosal secretions, highlighting the potential protective benefits of a vaccine targeting the upper respiratory tract.

Memory T cells effectively recognize the SARS-CoV-2 hypermutated BA.2.86 variant.

T cells are critical in mediating the early control of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) breakthrough infection. However, it remains unknown whether memory T cells can effectively cross-recognize new SARS-CoV-2 variants with a broad array of mutations, such as the emergent hypermutated BA.2.86 variant. Here, we report in two separate cohorts, including healthy controls and individuals with chronic lymphocytic leukemia, that SARS-CoV-2 spike-specific CD4+ and CD8+ T cells induced by prior infection or vaccination demonstrate resilient immune recognition of BA.2.86. In both cohorts, we found largely preserved SARS-CoV-2 spike-specific CD4+ and CD8+ T cell magnitudes against mutated spike epitopes of BA.2.86. Functional analysis confirmed that both cytokine expression and proliferative capacity of SARS-CoV-2 spike-specific T cells to BA.2.86-mutated spike epitopes are similarly sustained. In summary, our findings indicate that memory CD4+ and CD8+ T cells continue to provide cell-mediated immune recognition to highly mutated emerging variants such as BA.2.86.

Impact of the gut microbiome on immunological responses to COVID-19 vaccination in healthy controls and people living with HIV.

Although mRNA SARS-CoV-2 vaccines are generally safe and effective, in certain immunocompromised individuals they can elicit poor immunogenic responses. Among these individuals, people living with HIV (PLWH) have poor immunogenicity to several oral and parenteral vaccines. As the gut microbiome is known to affect vaccine immunogenicity, we investigated whether baseline gut microbiota predicts immune responses to the BNT162b2 mRNA SARS-CoV-2 vaccine in healthy controls and PLWH after two doses of BNT162b2. Individuals with high spike IgG titers and high spike-specific CD4+ T-cell responses against SARS-CoV-2 showed low α-diversity in the gut. Here, we investigated and presented initial evidence that the gut microbial composition influences the response to BNT162b2 in PLWH. From our predictive models, Bifidobacterium and Faecalibacterium appeared to be microbial markers of individuals with higher spike IgG titers, while Cloacibacillus was associated with low spike IgG titers. We therefore propose that microbiome modulation could optimize immunogenicity of SARS-CoV-2 mRNA vaccines.

Local and Systemic Immunity During Five Vaccinations Against SARS-CoV-2 in Zanubrutinib-Treated Patients With Chronic Lymphocytic Leukemia.

Background: Patients with chronic lymphocytic leukemia (CLL) are vulnerable to coronavirus disease 2019 (COVID-19) and are at risk of inferior response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccination, especially if treated with the first-generation Bruton's tyrosine kinase inhibitor (BTKi) ibrutinib. We aimed to evaluate the impact of the third-generation BTKi, zanubrutinib, on systemic and mucosal response to SARS-CoV-2 vaccination. Methods: Nine patients with CLL with ongoing zanubrutinib therapy were included and donated blood and saliva during SARS-CoV-2 vaccination, before vaccine doses 3 and 5 and 2 - 3 weeks after doses 3, 4, and 5. Ibrutinib-treated control patients (n = 7) and healthy aged-matched controls (n = 7) gave blood 2 - 3 weeks after vaccine dose 5. We quantified reactivity and neutralization capacity of SARS-CoV-2-specific IgG and IgA antibodies (Abs) in both serum and saliva, and reactivity of T cells activated with viral peptides. Results: Both zanubrutinib- and ibrutinib-treated patients had significantly, up to 1,000-fold, lower total spike-specific Ab levels after dose 5 compared to healthy controls (P < 0.01). Spike-IgG levels in serum from zanubrutinib-treated patients correlated well to neutralization capacity (r = 0.68; P < 0.0001) and were thus functional. Mucosal immunity (specific IgA in serum and saliva) was practically absent in zanubrutinib-treated patients even after five vaccine doses, whereas healthy controls had significantly higher levels (tested in serum after vaccine dose 5) (P < 0.05). In contrast, T-cell reactivity against SARS-CoV-2 peptides was equally high in zanubrutinib- and ibrutinib-treated patients as in healthy control donors. Conclusions: In our small cohort of zanubrutinib-treated CLL patients, we conclude that up to five doses of SARS-CoV-2 vaccination induced no detectable IgA mucosal immunity, which likely will impair the primary barrier defence against the infection. Systemic IgG responses were also impaired, whereas T-cell responses were normal. Further and larger studies are needed to evaluate the impact of these findings on disease protection.

Analyzing the Effect of the 2022 Formula Shortage on Postpartum Parents' Feeding Decisions in an Urban Hospital Setting.

Introduction: In early 2022, more than 40% of the U.S. formula supply was out of stock due to product recalls and manufacturing plant shutdowns. While previous studies have explored parents' opinions on formula feeding in response to formula safety concerns and offered advice on adapting to the 2022 formula shortage, there is currently a lack of data assessing the relationship between the formula shortage and parents' feeding decisions. Our study aims to understand how new parents' feeding decisions were impacted in the aftermath of the 2022 formula shortage, in comparison to demographic factors and feeding methods with previous children. We hypothesized that parents may either exclusively breastfeed their new baby when they did not do so previously or breastfeed their new baby more frequently than they did previously due to the formula shortage. Methods: Ninety-nine postpartum parents were administered a Qualtrics survey during their admission at a private urban hospital in St. Louis, Missouri, and answered questions about demographics, feeding decisions with previous children, current feeding decisions, and how influential the formula shortage was on these decisions. Data was analyzed using chi-square and Fisher's exact tests. Results: There is a significant relationship between parents' feeding decisions for previous children and feeding decisions for their new baby (p < 0.0001) but no significant influence of the formula shortage on feeding decisions for their new baby (p = 0.80). Conclusion: When making feeding decisions for their newborn, parents were more influenced by decisions with previous children than by the formula shortage, which highlights the importance of supporting parents with breastfeeding, especially for their first child.

Mycobiota profile of oral fungal infections in head and neck cancer patients receiving radiotherapy: A 6-year retrospective MALDI-TOF mass spectrometry study.

OBJECTIVES: Head and neck cancer (HNC) impairs patient immunity and increases susceptibility to oral fungal infections (OFIs). Effectively treating such infections requires accurate identification of the causative pathogens. This study aimed to characterize the mycobiota profile of OFIs in HNC patients undergoing radiation treatment (RT). MATERIALS AND METHODS: A 6-year retrospective analysis of oral mucosal samples from HNC patients with a history of RT and OFIs between 2014 and 2019 was conducted using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) profiling. Samples from the Clinical Microbiology Laboratory at Karolinska University Hospital were evaluated for mycobiota diversity and species co-occurrence patterns in the ongoing-RT and post-RT groups. RESULTS: A total of 190 oral fungi (88% Candida, 5% Pichia) were isolated from 162 HNC patients receiving RT. In the ongoing-RT group, the emergent non-albicans Candida (NAC) species; F. solani and C. jadinii, were detected for the first time. The dominant pathogens in both ongoing and post-RT groups were C. albicans, C. glabrata, P. kudriavzevii, C. parapsilosis, and C. tropicalis, as shown by Venn analysis. Network analysis revealed greater fungi diversity and multi-species co-occurrence in the ongoing-RT group. C. albicans commonly co-occurred with C. glabrata in both ongoing-RT (21%) and post-RT groups (30%). CONCLUSION: MALDI-TOF MS identified a wide range of oral fungal species in HNC patients receiving RT. While C. albicans remains the most prevalent OFIs pathogen, multi-species co-occurrence and novel NACs were noted. Understanding the ecological interactions among these causative pathogens could significantly advance the development of effective therapeutics for treating OFIs in HNC patients.

Additive effects of booster mRNA vaccination and SARS-CoV-2 Omicron infection on T cell immunity across immunocompromised states.

Suboptimal immunity to SARS-CoV-2 mRNA vaccination has frequently been observed in individuals with various immunodeficiencies. Given the increased antibody evasion properties of emerging SARS-CoV-2 subvariants, it is necessary to assess whether other components of adaptive immunity generate resilient and protective responses against infection. We assessed T cell responses in 279 individuals, covering five different immunodeficiencies and healthy controls, before and after booster mRNA vaccination, as well as after Omicron infection in a subset of patients. We observed robust and persistent Omicron-reactive T cell responses that increased markedly upon booster vaccination and correlated directly with antibody titers across all patient groups. Poor vaccination responsiveness in immunocompromised or elderly individuals was effectively counteracted by the administration of additional vaccine doses. Functionally, Omicron-reactive T cell responses exhibited a pronounced cytotoxic profile and signs of longevity, characterized by CD45RA+ effector memory subpopulations with stem cell-like properties and increased proliferative capacity. Regardless of underlying immunodeficiency, booster-vaccinated and Omicron-infected individuals appeared protected against severe disease and exhibited enhanced and diversified T cell responses against conserved and Omicron-specific epitopes. Our findings indicate that T cells retain the ability to generate highly functional responses against newly emerging variants, even after repeated antigen exposure and a robust immunological imprint from ancestral SARS-CoV-2 mRNA vaccination.

Real-world assessment of immunogenicity in immunocompromised individuals following SARS-CoV-2 mRNA vaccination: a one-year follow-up of the prospective clinical trial COVAXID.

BACKGROUND: Immunocompromised patients have varying responses to SARS-CoV-2 mRNA vaccination. However, there is limited information available from prospective clinical trial cohorts with respect to long-term immunogenicity-related responses in these patient groups following three or four vaccine doses, and in applicable cases infection. METHODS: In a real-world setting, we assessed the long-term immunogenicity-related responses in patients with primary and secondary immunodeficiencies from the prospective open-label clinical trial COVAXID. The original clinical trial protocol included two vaccine doses given on days 0 and 21, with antibody titres measured at six different timepoints over six months. The study cohort has subsequently been followed for one year with antibody responses evaluated in relation to the third and fourth vaccine dose, and in applicable cases SARS-CoV-2 infection. In total 356/539 patients were included in the extended cohort. Blood samples were analysed for binding antibody titres and neutralisation against the Spike protein for all SARS-CoV-2 variants prevailing during the study period, including Omicron subvariants. SARS-CoV-2 infections that did not require hospital care were recorded through quarterly in-person, or phone-, interviews and assessment of IgG antibody titres against SARS-CoV-2 Nucleocapsid. The original clinical trial was registered in EudraCT (2021-000175-37) and clinicaltrials.gov (NCT04780659). FINDINGS: The third vaccine dose significantly increased Spike IgG titres against all the SARS-CoV-2 variants analysed in all immunocompromised patient groups. Similarly, neutralisation also increased against all variants studied, except for Omicron. Omicron-specific neutralisation, however, increased after a fourth dose as well as after three doses and infection in many of the patient subgroups. Noteworthy, however, while many patient groups mounted strong serological responses after three and four vaccine doses, comparably weak responders were found among patient subgroups with specific primary immunodeficiencies and subgroups with immunosuppressive medication. INTERPRETATION: The study identifies particularly affected patient groups in terms of development of long-term immunity among a larger group of immunocompromised patients. In particular, the results highlight poor vaccine-elicited neutralising responses towards Omicron subvariants in specific subgroups. The results provide additional knowledge of relevance for future vaccination strategies. FUNDING: The present studies were supported by grants from the Swedish Research Council, the Knut and Alice Wallenberg Foundation, Nordstjernan AB, Region Stockholm, and Karolinska Institutet.

Clinical Microbial Identification of Severe Oral Infections by MALDI-TOF Mass Spectrometry in Stockholm County: an 11-Year (2010 to 2020) Epidemiological Investigation.

Growing evidence suggests that oral infections can modify the course of systemic diseases. To date, epidemiological data on microbial oral infections are scarce. Here, we performed a comprehensive analysis of the trend and microbial diversity in oral infection specimens referred for clinical microbiology analysis from 2010 to 2020. The microbes were isolated by culture and were identified via matrix-assisted laser desorption ionization-time of flight mass spectrometry technology (MALDI-TOF MS) throughout the study period. A total of 1,014 referred samples from dental clinics in Stockholm County with dentoalveolar abscesses and jaw osteomyelitis being the main reason were identified. Overall, the microbial composition was dominated by Firmicutes (51%), followed by Bacteroidetes (19%), Proteobacteria (12%), and Actinobacteria (5%). At the genus level, Streptococcus spp. (36%), Prevotella spp. (18%), and Staphylococcus spp. (11%) were among the most frequently reported. Interestingly, a strong increase in trend was noted for Streptococcus anginosus, Streptococcus mitis, Streptococcus sanguinis, Eikenella corrodens, Actinomyces spp., Aggregatibacter aphrophilus, Staphylococcus epidermidis, and Granulicatella adiacens during the study time (R = 0.66 to 0.89, P < 0.05), and a minor increase was noted for Enterococcus faecalis and Klebsiella spp., whereas steady levels were noted for most of the others. The present study shows the diversity of bacteria that have been involved in dental infections during the last decade in the capital of Sweden, as well as the emerging oral microbiota trend, with clear clinical implications on the oral-systemic link. IMPORTANCE Oral diseases and associated microbes are a risk factor for systemic diseases and can change the courses of these diseases. To date, epidemiological data on microbial oral infections are scarce, and longitudinal reports are lacking. We present for the first time the microbial composition of severe oral bacterial infections determined via the MALDI-TOF mass spectrometry technique in a comprehensive study between 2010 and 2020 (11 years) in Stockholm County. The trend and microbial diversity of oral infections were analyzed on referred clinical microbiological samples and were processed by standardized protocols. Trend increase was noted for Streptococcus anginosus, Streptococcus mitis, Streptococcus sanguinis, Eikenella corrodens, Actinomyces spp., Aggregatibacter aphrophilus, Staphylococcus epidermidis, Granulicatella adiacens, Enterococcus faecalis, and Klebsiella spp. Our results provide new insights into the diversity and trend of oral microbiota that were involved in serious oral infections over the past decade in the capital of Sweden and may influence the oral-systemic link.

Poor dental health and risk of pancreatic cancer: a nationwide registry-based cohort study in Sweden, 2009-2016.

BACKGROUND: Previous studies have reported inconsistent results regarding the association between poor dental health and pancreatic cancer risk. This study aimed to assess this association using a well-functioning nationwide dental health registry in Sweden. METHODS: Information of exposures (dental caries, root canal infection, mild inflammation, and periodontitis; the number of teeth) was ascertained from the Swedish Dental Health Register, and occurrence of pancreatic cancer was identified from both cancer and cause of death registries. Hazard ratios (HRs) were estimated using Cox models. RESULTS: During a median of 7.2 years of follow-up, 10,081 pancreatic cancers were identified among 5,889,441 individuals. Compared with the healthy status, a higher risk of pancreatic cancer was observed in individuals with root canal infection, mild inflammation, and periodontitis in the <50 age group (P for trend <0.001). In the 50-70 age group, only the subgroup with periodontitis had an excess risk (multivariable-adjusted HR = 1.20, 95% confidence interval [CI] 1.11-1.29). No positive association with statistical significance was observed in the 70+ age group. Individuals with fewer teeth tended to have a higher risk in all age groups. CONCLUSIONS: Our results confirmed the association between poor dental health and pancreatic cancer risk, which warrants further studies on underlying mechanisms.

Immunodeficiency syndromes differentially impact the functional profile of SARS-CoV-2-specific T cells elicited by mRNA vaccination.

Many immunocompromised patients mount suboptimal humoral immunity after SARS-CoV-2 mRNA vaccination. Here, we assessed the single-cell profile of SARS-CoV-2-specific T cells post-mRNA vaccination in healthy individuals and patients with various forms of immunodeficiencies. Impaired vaccine-induced cell-mediated immunity was observed in many immunocompromised patients, particularly in solid-organ transplant and chronic lymphocytic leukemia patients. Notably, individuals with an inherited lack of mature B cells, i.e., X-linked agammaglobulinemia (XLA) displayed highly functional spike-specific T cell responses. Single-cell RNA-sequencing further revealed that mRNA vaccination induced a broad functional spectrum of spike-specific CD4+ and CD8+ T cells in healthy individuals and patients with XLA. These responses were founded on polyclonal repertoires of CD4+ T cells and robust expansions of oligoclonal effector-memory CD45RA+ CD8+ T cells with stem-like characteristics. Collectively, our data provide the functional continuum of SARS-CoV-2-specific T cell responses post-mRNA vaccination, highlighting that cell-mediated immunity is of variable functional quality across immunodeficiency syndromes.

Process Development for Adoptive Cell Therapy in Academia: A Pipeline for Clinical-Scale Manufacturing of Multiple TCR-T Cell Products.

Cellular immunotherapies based on T cell receptor (TCR) transfer are promising approaches for the treatment of cancer and chronic viral infections. The discovery of novel receptors is expanding considerably; however, the clinical development of TCR-T cell therapies still lags. Here we provide a pipeline for process development and clinical-scale manufacturing of TCR-T cells in academia. We utilized two TCRs specific for hepatitis C virus (HCV) as models because of their marked differences in avidity and functional profile in TCR-redirected cells. With our clinical-scale pipeline, we reproduced the functional profile associated with each TCR. Moreover, the two TCR-T cell products demonstrated similar yield, purity, transduction efficiency as well as phenotype. The TCR-T cell products had a highly reproducible yield of over 1.4 × 109 cells, with an average viability of 93%; 97.8-99% of cells were CD3+, of which 47.66 ± 2.02% were CD8+ T cells; the phenotype was markedly associated with central memory (CD62L+CD45RO+) for CD4+ (93.70 ± 5.23%) and CD8+ (94.26 ± 4.04%). The functional assessments in 2D and 3D cell culture assays showed that TCR-T cells mounted a polyfunctional response to the cognate HCV peptide target in tumor cell lines, including killing. Collectively, we report a solid strategy for the efficient large-scale manufacturing of TCR-T cells.

Evaluating the prevalence of Hepatitis E virus infection in a large cohort of European blood donors, 2015-2018.

Hepatitis E virus (HEV) is endemic in Europe. However, standardized methods for the surveillance of HEV viremia in the general population are lacking. This study aimed to compare the incidence of HEV among blood donors in two European countries, Germany and Portugal, during the period 2015-2018. The seasonal distribution of HEV infection, as well as host risk factors including age, sex, and blood group phenotype were explored. A total of 191,236 donations from Germany and Portugal were tested for HEV RNA in plasma mini-pools of up to 96 donations using an internally controlled reverse transcription real-time PCR (RT-PCR) assay. The 95% cut-off of the assay was 15 International Units (IU)/mL (CI 10-35 IU/mL) as determined by dilution of the WHO International Standard for HEV RNA. Blood type was determined by agglutination and pattern recognition using the Beckmann Coulter PK 7300 AB0- and Rhesus-Assay. The overall positivity rate was 0.09% with significantly more infections observed in the German cohort (p < 0.0001). Infections peaked in the summer months, and investigation of risk factors revealed that incidence was significantly higher amongst males (p = 0.0002), but was not associated with ABO or Rh(D) blood group phenotypes. No significant relationships between risk factors and viral load were observed. Our findings confirm that HEV infections are highly prevalent in Europe, even amongst otherwise healthy blood donors. Increasing awareness of the seasonal spread and risk factors for HEV transmission is of great importance for individuals susceptible to more severe forms of the disease, such as immunocompromised patients.

Sleep enhances reconsolidation-based strengthening of visuospatial memories.

Consolidated memories can be returned to a labile state upon reactivation. The re-stabilization of reactivated memories, or reconsolidation, can allow for change in previously established memories. Given the role of sleep in the initial consolidation of memories, sleep may be important for reconsolidation as well. However, effects of sleep on reconsolidation and specific aspects of sleep that may contribute are unclear. Here, participants learned 30 picture-location pairs. After overnight sleep, initial consolidation was tested. Following either one day (Experiment 1) or one week (Experiment 2), participants were tested again to reactivate their memory and then learned 30 novel picture-location pairs. Control groups (Experiment 1) received no reactivation prior to new learning. Twelve hours later, after daytime wakefulness or overnight sleep, participants completed a final memory test. Sleep participants underwent polysomnography between reactivation and final tests. In Experiment 1, reactivation led to preservation of memory compared to no reactivation. Sleep was associated with less post-reactivation memory decline than waking, with memory preservation positively related to time spent in non-rapid-eye movement sleep. In Experiment 2, sleep was associated with greater post-reactivation memory improvement than waking, with improvement positively related to sigma activity. These results suggest sleep enhances reconsolidation-based strengthening of episodic memories.

Preserved Mucosal-Associated Invariant T Cells in the Cervical Mucosa of HIV-Infected Women with Dominant Expression of the TRAV1-2-TRAJ20 T Cell Receptor α-Chain.

BACKGROUND: Mucosa-associated invariant T (MAIT) cells are innate-like T cells with specialized antimicrobial functions. Circulating MAIT cells are depleted in chronic human immunodeficiency virus (HIV) infection, but studies examining this effect in peripheral tissues, such as the female genital tract, are lacking. METHODS: Flow cytometry was used to investigate circulating MAIT cells in a cohort of HIV-seropositive (HIV+) and HIV-seronegative (HIV-) female sex workers (FSWs), and HIV- lower-risk women (LRW). In situ staining and quantitative polymerase chain reaction were performed to explore the phenotype of MAIT cells residing in paired cervicovaginal tissue. The cervicovaginal microbiome was assessed by means of 16S ribosomal RNA gene sequencing. RESULTS: MAIT cells in the HIV+ FSW group were low in frequency in the circulation but preserved in the ectocervix. MAIT cell T-cell receptor gene segment usage differed between the HIV+ and HIV- FSW groups. The TRAV1-2-TRAJ20 transcript was the most highly expressed MAIT TRAJ gene detected in the ectocervix in the HIV+ FSW group. MAIT TRAVJ usage was not associated with specific genera in the vaginal microbiome. CONCLUSIONS: MAIT cells residing in the ectocervix are numerically preserved irrespective of HIV infection status and displayed dominant expression of TRAV1-2-TRAJ20. These findings have implications for understanding the role of cervical MAIT cells in health and disease.