Specific IgG Antibodies Research Articles

Lactobacillus paracasei-derived extracellular vesicles alleviate neutrophilic asthma by inhibiting the JNK pathway in airway epithelium

BackgroundLactobacillus paracasei has been known to reduce airway resistance and inflammation in asthma. However, the therapeutic effect of its extracellular vesicles (EVs) in patients with asthma remains unclear. MethodsTo validate the clinical relevance of L. paracasei-derived EVs (LpEV) in asthma, the composition of gut microbial EVs was verified by metagenomics in LPS-induced C57BL/6 mice. The components of proteins and metabolites in LpEV were identified by peptide mass fingerprinting and metabolomic analysis. The serum levels of specific IgG1 or IgG4 antibodies to LpEV were compared by ELISA between patients with eosinophilic asthma (EA, n = 10) and those with neutrophilic asthma (NA, n = 10) as well as with healthy controls (HCs, n = 10). Finally, therapeutic effects of LpEV and their metabolites in asthma were validated in vivo/in vitro. ResultsSignificantly lower proportions of EVs derived from Lactobacillus at the genus level were noted in mice with NA than in control mice. Moreover, the serum levels of LpEV-specific IgG4, but not IgG1, were lower in patients with NA than in those with EA or in HCs and positively correlated with FEV1 (%) values. In addition, oral administration of LpEV reduced airway resistance and inflammation in mice with NA. Finally, LpEV and their 3 metabolites (dodecanoic acid, palmitoleic acid, and D-(−)-tagatose) significantly inhibited JNK phosphorylation/IL-8 production in airway epithelium in vitro. ConclusionsThese findings suggest that LpEV may have a therapeutic potential targeting NA by suppressing the JNK pathway and proinflammatory cytokine production in airway epithelium.

ApoB-specific CD4+ T cells circulate in human blood and formation of a memory population is driven by coronary artery disease and an elevated cardiovascular risk profile

Abstract Aim Atherosclerosis is a chronic inflammatory disease involving an autoimmune response against Apolioprotein B (ApoB), the core protein of low-density lipoprotein (LDL) cholesterol. Using a translational bench-to-bedside approach, we linked the presence and immuno-phenotype of ApoB-specific CD4+ T cells to clinical risk profiles for cardiovascular disease in humans. Methods Peripheral blood mononuclear cells (PBMCs) from 230 patients that underwent coronary angiography were co-incubated in vitro with a pool of immunodominant peptides from human ApoB for 6h. Reactive ApoB-specific T cells (ApoB+) were defined in flow cytometry by expression of the activation marker CD40L. Partition of ApoBpos into naïve and memory T cells was achieved by extracellular fluorochrome-based staining of CD45RO, CCR7 and CD45RA. Cytokine expression of ApoBpos was determined by intracellular staining. Results We found that a median of 0.69±0.1% of circulating CD4+ T cells were reactive to Apolipoprotein B. We found detectable concentrations of auto reactive ApoB+ T cells in 86,44% of all healthy individuals. However, overall frequencies of ApoB reactive T cells did not correlate with diagnosed coronary artery disease (CAD). Compared to ApoBneg. T cells, we detected a higher fraction of memory T cells in ApoBpos (p<0,0001), suggesting a previously occurred ApoB directed immune response. Accordingly, elevated serum levels of ApoB specific IgG Antibodies associated with high proportions of ApoB specific memory cells. Patients diagnosed with coronary artery disease presented higher percentages of central memory T cells among ApoBpos compared to healthy individuals. Correspondingly, these observations applied to patients with an elevated 10-year risk for cardiovascular events estimated by the ESC-SCORE2. These effects largely grounded on strong associations with the clinical diagnosis of arterial hypertension, elevated levels of Lp(a) and obesity. Interestingly, systolic blood pressure and LDL-serum levels correlated with elevated proportions of memory-status ApoBpos T cells in patients without statin intake, although this was not the case for patients with LDL-lowering medication. Analysis of cytokine expression in ApoBpos CD4+ T cells showed significantly increased expression of anti-inflammatory Il-2 and IL-10 in addition to pro-inflammatory IFNγ. Cytokine secretion was mainly driven by memory-status ApoBpos when compared to naïve ApoBpos T cells. Interestingly, a significant number of patients showed a mixed TH1/Treg phenotype among ApoBpos with simultaneous expression of IL-10 and IFN-γ. Conclusions Our data show for the first time that ApoB-reactive T cells exist even in healthy individuals. Cardiovascular risk factors and clinically relevant atherosclerotic disease drive memory formation and activation of ApoB+ T cells. ApoB+ Memory T cells are prone to secrete pro- and anti-inflammatory cytokines and may modulate systemic inflammation in atherosclerosis.

Antibody response and soluble mediator profile in the first six months following acute SARS-CoV-2 infection

The COVID-19 pandemic has caused a severe global health and economic crisis, with significant consequences for human mortality and morbidity. Therefore, there is an urgent need for more studies on the immune response to SARS-CoV-2 infection, both to enhance its effectiveness and prevent its deleterious effects. This study presents the chronology of antibodies during six months after infection in hospitalized patients and the kinetics of serum soluble mediators of the cellular response triggered by SARS-CoV-2. Samples and clinical data from 330 patients hospitalized at the Hospital da Baleia in Belo Horizonte, Brazil, who were suspected of having COVID-19, were collected at the time of hospitalization and during 6 months after infection. The immune response was analyzed by enzyme-linked immunosorbent assay (ELISA) and flow cytometry. There was a significant difference in IgM specific antibody titers from the 7th to 60th days after infection between COVID-19 negative and positive patients. Soon after 60 days after infection, antibody levels started to reduce, becoming similar to the antibody levels of the COVID-19 negative patients. IgG specific antibodies started to be detectable after 9 days of infection and antibody levels were comparatively higher in positive patients as soon as after 7 days. Furthermore, IgG levels remained higher in these patients during the complete period of 180 days after infection. The study observed similar antibody profiles between different patient groups. The soluble systemic biomarkers evaluated showed a decrease during the six months after hospitalization, except for CCL11, CXCL8, CCL3, CCL4, CCL5, IL-6, IFN-g, IL-17, IL-5, FGF-basic, PDGF, VEGF, G-CSF, and GM-CSF. The results indicate that IgM antibodies are more prominent in the early stages of infection, while IgG antibodies persist for a longer period. Additionally, the study identified that patients with COVID-19 have elevated levels of biomarkers after symptom onset, which decrease over time.

Safety and immunogenicity of an inactivated SARS-CoV-2 vaccine (FAKHRAVAC®) in healthy adults aged 18–55 years: Randomized, double-blind, placebo-controlled, phase I clinical trial

BackgroundThe FAKHRAVAC®, an inactivated SARS-CoV-2 vaccine, was assessed for safety and immunogenicity. Methods and findingsIn this double-blind, placebo-controlled, phase I trial, we randomly assigned 135 healthy adults between 18 and 55 to receive vaccine strengths of 5 or 10 μg/dose or placebo (adjuvant only) in 0–14 or 0–21 schedules. This trial was conducted in a single center in a community setting. The safety outcomes in this study were reactogenicity, local and systemic adverse reactions, abnormal laboratory findings, and Medically Attended Adverse Events (MAAE). Immunogenicity outcomes include serum neutralizing antibody activity and specific IgG antibody levels.The most frequent local adverse reaction was tenderness (28.9%), and the most frequent systemic adverse reaction was headache (9.6%). All adverse reactions were mild, occurred at a similar incidence in all six groups, and were resolved within a few days. In the 10-µg/dose vaccine group, the geometric mean ratio for neutralizing antibody titers at two weeks after the second injection compared to the placebo group was 9.03 (95% CI: 3.89–20.95) in the 0–14 schedule and 11.77 (95% CI: 2.77–49.94) in the 0–21 schedule. The corresponding figures for the 5-µg/dose group were 2.74 (1.2–6.28) and 5.2 (1.63–16.55). The highest seroconversion rate (four-fold increase) was related to the 10-µg/dose group (71% and 67% in the 0–14 and 0–21 schedules, respectively). ConclusionsFAKHRAVAC® is safe and induces a strong humoral immune response to the SARS-CoV-2 virus at 10-µg/dose vaccine strength in adults aged 18–55. This vaccine strength was used for further assessment in the phase II trial.Trial registrationThis study is registered with https://www.irct.ir; IRCT20210206050259N1.

Features of the epidemic process in the spread of a new coronavirus infection COVID-19 on the territory of the metropolitan area in the period 2020–2022

Introduction Conducting a comprehensive epidemiological etiological analysis of the development of the СОVID-19 pandemic in a metropolitan area is of interest to specialists of didderent profiles (epidemiologists, infectious disease specialists, pediatricians, therapists, immunologists, virologists). An important element of epidemiological surveillance is the assessment of collective immunity to infectious agents, including the SARS-CoV-2 virus. The natural ability of viruses to variability leads to the emergence of new types and strains. Currently, a number of mutations and polymorphisms have been identified that affect the structure and stability of proteins, factors of susceptibility to SARS-CoV-2 infection, pathogenicity and virulence of the virus, and the ability to “elude” the immune response of the host organism.The aim of the work was to assess evaluate the character of the epidemic process development of a new coronavirus infection on the territory of the metropolitan area in the period 2020–2022.Materials and methods A total of 331,013 people were examined by PCR for the presence of SARS-CoV-2 RNA in nasopharyngeal and oropharyngeal swabs. Specific IgM and IgG antibodies to SARS-CoV-2 in serum were detected by ELISA in 42,955 people. Genetic variants of the virus were typed (by PCR and sequencing).Results During the entire follow-up period, the highest percentage of “positive for Covid-19” was observed in February 2022 (58.9 %) and September 2022 (46.8 %). Among the total number of patients examined (n = 331,013), adult patients accounted for the largest proportion, – 83.9 %, children – 16.1 %. The proportion of individuals with specific IgG to SARSCoV-2 in 2020–2021 ranged from 53.3–82.6 %.Discussion In 2020–2022, there was an increase in the proportion of individuals with detectable RNA virus. The greatest infection was observed in the age groups 19–27 years and 28–65 years. It was found that the linear dependence of the increase in the specific weight of seropositive persons with respect to IgG among the residents of Ekaterinburg has a progressive trend, the approximation reliability coefficient R2 corresponds to 0.8172. In samples of patients examined in April–August 2022, markers of the genetic variant Omicron were detected in 96.9 % of cases. The increase in the structure of infected pediatric population (almost twofold over 2020–2022 years) is probably caused by the adaptation of the pathogen to a new susceptible cohort of people.Conclusion The dynamics of the number of people examined and the proprtion of detectability of SARS-CoV-2 RNA in Ekaterinburg during the entire period of the pandemic (March 2020–December 2022) had a wave-like character. By the end of 2022, the level of seropositive individuals in all age groups increased significantly, which confirms the presence of collective immunity in the territory.

Immunogenicity and Tolerability of a SARS-CoV-2 TNX-1800, a Live Recombinant Poxvirus Vaccine Candidate, in Syrian Hamsters and New Zealand White Rabbits.

TNX-1800 is a preclinical stage synthetic-derived live attenuated chimeric horsepox virus vaccine engineered to express the SARS-CoV-2 spike (S) gene. The objectives of this study were to assess the safety, tolerability, and immunogenicity of TNX-1800 administration in Syrian golden hamsters and New Zealand white rabbits. Animals were vaccinated at three doses via percutaneous inoculation. The data showed that the single percutaneous administration of three TNX-1800 vaccine dose levels was well tolerated in both hamsters and rabbits. At all dose levels, rabbits were more decerning regarding vaccine site reaction than hamsters. Lastly, no TNX-1800 genomes could be detected at the site of vaccination. Post-vaccination, all animals had anti-SARS-CoV-2 spike protein IgG specific antibody responses. These data demonstrate that TNX-1800 infection was limited, asymptomatic, and cleared by the end of this study, and a single dose was able to generate immune responses.

Immune-mediated cholangiopathies in children: the need to better understand the pathophysiology for finding the future possible treatment targets.

Cholangiopathies are defined as focal or extensive damage of the bile ducts. According to the pathogenetic mechanism, it may be immune-mediated or due to genetic, infectious, toxic, vascular, and obstructive causes. Their chronic evolution is characterized by inflammation, obstruction of bile flow, cholangiocyte proliferation, and progression toward fibrosis and cirrhosis. Immune-mediated cholangiopathies comprise primary sclerosing cholangitis (PSC), autoimmune cholangitis and IgG4-associated cholangitis in adults and biliary atresia (BA), neonatal sclerosing cholangitis (NSC) in children. The main purpose of this narrative review was to highlight the similarities and differences among immune-mediated cholangiopathies, especially those frequent in children in which cholangiocyte senescence plays a key role (BA, NSC, and PSC). These three entities have many similarities in terms of clinical and histopathological manifestations, and the distinction between them can be hard to achieve. In BA, bile duct destruction occurs due to aggression of the biliary cells due to viral infections or toxins during the intrauterine period or immediately after birth. The consequence is the activation of the immune system leading to severe inflammation and fibrosis of the extrahepatic biliary tract, lumen stenosis, and impairment of the biliary flow. PSC is characterized by inflammation and fibrosis of intra- and extrahepatic bile ducts, leading to secondary biliary cirrhosis. It is a multifactorial disease that occurs because of genetic predisposition [human leukocyte antigen (HLA) and non-HLA haplotypes], autoimmunity (cellular immune response, autoantibodies, association with inflammatory bowel disease), environmental factors (infections or toxic bile), and host factors (intestinal microbiota). NSC seems to be a distinct subgroup of childhood PSC that appears due to the interaction between genetic predisposition (HLA B8 and DR3) and the disruption of the immune system, validated by elevated IgG levels or specific antibodies [antinuclear antibody (ANA), anti-smooth muscle antibody (ASMA)]. Currently, the exact mechanism of immune cholangiopathy is not fully understood, and further data are required to identify individuals at high risk of developing these conditions. A better understanding of the immune mechanisms and pathophysiology of BA, NSC, and PSC will open new perspectives for future treatments and better methods of preventing severe evolution.

Cross-sectional assessment of a history of SARS-CoV-2 infections using IgG

IntroductionSevere acute respiratory syndrome, coronavirus type 2 (SARS-CoV-2) has become a global threat for every healthcare system, and the coronavirus disease 2019 (COVID-19) pandemic has resulted in over 3 million deaths worldwide. SARS-CoV-2 brings symptoms such as fever, cough, shortness of breath, headache, sore throat and loss of taste or smell. Diagnostic of COVID-19 may include specific RT-PCR for viral ribonucleic acid detection, and ELISA testing for virus-derived spike protein or nucleocapsid.AimThe aim of this study was to measure the antinucleocapsid level of SARS-CoV-2 IgG to identify the number of asymptomatic cases of COVID-19 after infection in a population of workers from a production company.Material and methodsHuman anti-SARS-CoV-2(N) IgG ELISA kit was used to determine serum IgG level. Study includes 107 individuals (48% female, 52% male) in different ages (18–60 years).Results and discussionOf 107 tested individuals in 80 (74.7%) cases SARS-CoV-2(N)-specific IgG antibodies were detected, with an average antibody concentration of the whole study group 4.08 µg/mL (<i>n</i> = 107 with the range 0.59–7.91 µg/mL; <i>n</i> = 80 were included in the study with the sensitivity of the method above 2.344 µg/mL). In only 9 cases, SARS-CoV-2 infection was confirmed before using the PCR test. Our data underscore the need for a population study in Poland to test the proportion of asymptomatic IgG positive for SARS-CoV-2 individuals.ConclusionsThis study indicates that within studied sample large proportion of asymptomatic people have undergone SARS-CoV-2 infection and suggests that isolation of only symptomatic patients would not stop the transmission of the virus.

Prevalence of hypersensitive pneumonitis specific IgG antibodies in the Indian population: A retrospective study

Identifying the underlying antigen responsible for Hypersensitivity Pneumonitis (HP) in patients poses a significant challenge within the confines of a typical clinical environment. Our primary objective in this study was to investigate the distribution and prevalence of G antibodies among individuals diagnosed with HP, taking into account factors such as age, gender, and geographical location.A retrospective study spanning 5 years (from January 2018 to June 2023) was conducted, involving patients over the age of 18 who had undergone screening for HP. Data collected from 1087 patients was analysed, and the patients were categorized based on age, gender, location, and their antibody status.Out of the total cohort of 1087 patients, 62.47% were female, while 37.53% were male. The overall positivity rate for HP panel testing was determined to be 49.22%. Among the patients subjected to testing, 174 individuals (16.01%) tested positive for at least one specific antibody. Notably, among patients aged 18 to 30 years, Penicillium emerged as the most prevalent (48.28%), closely followed by fumigatus Fumigatus (44.83%). In contrast, for patients aged 31 to 45 years, Pigeon serum feathers exhibited the highest prevalence (39.69%). Moreover, the positivity rates varied across different regions in India.HP can be attributed to an array of agents commonly encountered in both workplace and home settings, encompassing microorganisms, animal and plant proteins, as well as organic and inorganic chemicals. Discerning the causative antigen not only holds immense value for physicians in optimizing patient treatment but also plays a pivotal role in pinpointing the source of exposure. Armed with this knowledge, physicians can provide patients with tailored advice to minimize their exposure risk, potentially leading to disease stabilization or even reversal. In summary, the successful identification of the etiologic antigen emerges as a formidable tool for enhancing the overall quality of patient care.

Phase II, Safety and Immunogenicity of RAZI Cov Pars (RCP) SARS Cov-2 Vaccine in Adults Aged 18–70 Years; A Randomized, Double-Blind Clinical Trial

BackgroundThis study explores the safety and immunogenicity of the Razi-Cov-Pars (RCP) SARS Cov-2 recombinant spike protein vaccine. MethodIn a randomized, double-blind, placebo-controlled trial, adults aged 18–70 were randomly allocated to receive selected 10 µg/200 µl vaccine strengths or placebo (adjuvant). It included two intramuscular injections at days 0 and 21, followed by an intranasal dose at day 51. Immediate and delayed solicited local and systemic adverse reactions after each dose up to a week, and specific IgG antibodies against SARS Cov-2 spike antigens two weeks after the 2nd dose were assessed as primary outcomes. Secondary safety outcomes were abnormal laboratory findings and medically attended adverse events (MAAE) over six months follow up. Secondary immunogenicity outcomes were neutralizing antibody activity and cell-mediated immune response. ResultBetween May 27th and July 15th, 2021, 500 participants were enrolled. Participants' mean (SD) age was 37.8 (9.0), and 67.0 % were male. No immediate adverse reaction was observed following the intervention. All solicited local and systemic adverse events were moderate (Grade I-II). Specific IgG antibody response against S antigen in the vaccine group was 5.28 times (95 %CI: 4.02–6.94) the placebo group with a 75 % seroconversion rate. During six months of follow-up, 8 SAEs were reported, unrelated to the study intervention. The participants sustained their acquired humoral responses at the end of the sixth month. The vaccine predominantly resulted in T-helper 1 cell-mediated immunity, CD8+ cytotoxic T-cell increase, and no increase in inflammatory IL-6 cytokine. ConclusionRCP vaccine is safe and creates strong and durable humoral and cellular immunity. Trial registration(IRCT20201214049709N2).

Influence of antigen density and TLR ligands on preclinical efficacy of a VLP-based vaccine against peanut allergy.

Virus-like particle (VLP) Peanut is a novel immunotherapeutic vaccine candidate for the treatment of peanut allergy. The active pharmaceutical ingredient represents cucumber mosaic VLPs (CuMVTT -VLPs) that are genetically fused with one of the major peanut allergens, Ara h 2 (CuMVTT -Ara h 2). We previously demonstrated the immunogenicity and the protective capacity of VLP Peanut-based immunization in a murine model for peanut allergy. Moreover, a Phase I clinical trial has been initiated using VLP Peanut material manufactured following a GMP-compliant manufacturing process. Key product characterization studies were undertaken here to understand the role and contribution of critical quality attributes that translate as predictive markers of immunogenicity and protective efficacy for clinical vaccine development. The role of prokaryotic RNA encapsulated within VLP Peanut on vaccine immunogenicity was assessed by producing a VLP Peanut batch with a reduced RNA content (VLP Peanut low RNA). Immunogenicity and peanut allergen challenge studies were conducted with VLP Peanut low RNA, as well as with VLP Peanut in WT and TLR 7 KO mice. Furthermore, mass spectrometry and SDS-PAGE based methods were used to determine Ara h 2 antigen density on the surface of VLP Peanut particles. This methodology was subsequently applied to investigate the relationship between Ara h 2 antigen density and immunogenicity of VLP Peanut. A TLR 7 dependent formation of Ara h 2 specific high-avidity IgG antibodies, as well as a TLR 7 dependent change in the dominant IgG subclass, was observed following VLP Peanut vaccination, while total allergen-specific IgG remained relatively unaffected. Consistently, a missing TLR 7 signal caused only a weak decrease in allergen tolerability after vaccination. In contrast, a reduced RNA content for VLP Peanut resulted in diminished total Ara h 2 specific IgG responses, followed by a significant impairment in peanut allergen tolerability. The discrepant effect on allergen tolerance caused by an absent TLR 7 signal versus a reduced RNA content is explained by the observation that VLP Peanut-derived RNA not only stimulates TLR 7 but also TLR 3. Additionally, a strong correlation was observed between the number of Ara h 2 antigens displayed on the surface of VLP Peanut particles and the vaccine's immunogenicity and protective capacity. Our findings demonstrate that prokaryotic RNA encapsulated within VLP Peanut, including antigen density of Ara h 2 on viral particles, are key contributors to the immunogenicity and protective capacity of the vaccine. Thus, antigenicity and RNA content are two critical quality attributes that need to be determined at the stage of manufacturing, providing robust information regarding the immunogenicity and protective capacity of VLP Peanut in the mouse which has translational relevance to the human setting.

Raman spectroscopic analysis of human serum samples of convalescing COVID-19 positive patients

Rapid screening, detection and monitoring of viral infection is of critical importance, as exemplified by the rapid spread of SARS-CoV-2, leading to the worldwide pandemic of COVID-19. This is equally the case for the stages of patient convalescence as for the initial stages of infection, to understand the medium and long terms effects, as well as the efficacy of therapeutic interventions. Optical spectroscopic techniques potentially offer an alternative to currently employed techniques of screening for the presence, or the response to infection. In this study, the ability of Raman spectroscopy to distinguish between samples of the serum of convalescent COVID-19 positive patients and COVID-19 negative serum samples, and to further analyse and quantify systemic responses, was explored. The study included serum samples of patients who had been tested for SARS-CoV-2 specific IgG and IgM responses between 25 and 134 days after the infection was identified. Both COVID-19 positive and negative groups included males and females who ranged in age from 21 to 81 years old. No correlation was apparent between the specified SARS-CoV-2 specific IgG and IgM immunoglobulin levels of the positive group, their sex, or age. Raman spectroscopic measurements were performed at 785 nm, in liquid serum, thawed from frozen, and spectra were pre-processed to remove the contribution of water, normalising to the water content. Principal components analysis of the spectral dataset over the range 400–1800 cm-1 provided no clear indication of a difference between normal serum and SARS-CoV-2 positive serum. A selection of 5 of the samples, which were available in sufficient volume, were fractionated by centrifugal filtration, and the 100 kDa, 50 kDa, 30 kDa, and 10 kDa concentrates similarly analysed by Raman spectroscopy. Partial least squares regression analysis revealed a negative correlation between the spectral profile of the 30 kDa fractions and SARS-CoV-2 specific IgG antibody levels, potentially indicating an association with depleted glutathione levels. The study supports a potential role of Raman screening of blood serum for monitoring of SARS-CoV-2 infection, but also in longitudinal studies of disease progression, long term effects, and therapeutic interventions.

An enterovirus A71 virus-like particle with replaced loops confers partial cross-protection in mice

Enterovirus A71 (EV-A71), coxsackievirus A16 (CV-A16), and CV-A10 belong to the main prevailing types causing hand-foot-and-mouth disease. Since EV-A71 monovalent vaccine does not confer cross-protection, developing a multivalent vaccine is essential. In this study, a trivalent chimeric virus-like particle of EV-A71 (EV-A71-VLPCHI3) was constructed based on EV-A71-VLP backbone by replacing the corresponding surface loops with CV-A16 VP1 G-H, CV-A10 VP1 B-C and E-F loops, which are critical for immunogenic neutralization. The baculovirus-insect cell expression system was employed for EV-A71-VLPCHI3 production. EV-A71-VLPCHI3 was purified by sucrose density gradient and observed by transmission electron microscopy. The immunogenicity and protective efficacy of EV-A71-VLPCHI3 were evaluated in mice. Our results revealed that EV-A71-VLPCHI3 had a similar morphology to inactivated EV-A71 particles and could induce specific IgG antibodies against EV-A71, CV-A16 and CV-A10 in mice. More importantly, EV-A71-VLPCHI3 enhanced cross-reactive protection against CV-A16 and CV-A10, by 20 % and 40 %, compared to inactivated EV-A71 counterparts, respectively. In conclusion, the successful construction of EV-A71-VLPCHI3 suggested that loop-dependent heterologous protection could be transferred by loops replacement on the surface of viral capsid. This strategy may also supplement the development of multivalent vaccines against other infectious viral diseases.

Measuring the clustering effect of the SARS-CoV-2 transmission in a school population: a cross-sectional study in a high incidence region

Since the beginning of the pandemic, there has been a great deal of controversy regarding the role of schools in the spread of SARS-CoV-2 infection, and the relative contribution of students, teachers, and others. To quantify the clustering effect of SARS-CoV-2 infection within classes and schools considering the seroprevalence of specific antibodies among students and school staff (teachers and non-teachers) evaluated in schools located in the Northern region of Portugal. 1517 individuals (1307 students and 210 school staff) from 4 public and 2 private schools, comprising daycare to secondary levels, were evaluated. A rapid point-of-care test for SARS-CoV-2 specific IgM and IgG antibodies was performed and a questionnaire was completed providing sociodemographic and clinical information. We calculated the seroprevalence of IgM and IgG antibodies and estimated the Median Odds Ratio (OR) and 95% confidence interval (CI) to assess the clustering effect, using a multilevel (school and class) logistic regression. SARS-CoV-2 seroprevalence (IgM or IgG) was 21.8% and 23.8% (p = 0.575) in students and school staff, respectively. A total of 84 (8.6%) students and 35 (16.7%) school staff reported a previous molecular diagnosis. Among students, those who reported high-risk contacts only at school (OR = 1.13; 95% CI 0.72–1.78) had a seroprevalence similar to those without high-risk contacts; however, seroprevalence was significantly higher among those who only reported a high-risk contact outside the school (OR = 6.56; 95% CI 3.68–11.72), or in both places (OR = 7.83; 95% CI 5.14–11.93). Similar associations were found for school staff. The median OR was 1.00 (95% CI 1.00, 1.38) at the school-level and 1.78 (95% CI 1.40, 2.06) at the class-level. SARS-CoV-2 seroprevalence was similar between students and staff, without a clustering effect observed at the school level, and only a moderate clustering effect documented within classes. These results indicate that the mitigation measures in the school environment can prevent the spread of class outbreaks to the remaining school community.

Trivalent mRNA Vaccine against SARS-CoV-2 and Variants with Effective Immunization.

mRNA vaccines encoding a single spike protein effectively prevent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. However, the emergence of SARS-CoV-2 variants leads to a wide range of immune evasion. Herein, a unique trivalent mRNA vaccine based on ancestral SARS-CoV-2, Delta, and Omicron variant spike receptor-binding domain (RBD) mRNAs was developed to tackle the immune evasion of the variants. First, three RBD mRNAs of SARS-CoV-2, Delta, and Omicron were coencapsulated into lipid nanoparticles (LNPs) by using microfluidic technology. After that, the physicochemical properties and time-dependent storage stability of the trivalent mRNA vaccine nanoformulation were tested by using dynamic light scattering (DLS). In vitro, the trivalent mRNA vaccine exhibited better lysosomal escape ability, transfection efficiency, and biocompatibility than did the commercial transfection reagent Lipo3000. In addition, Western blot analyses confirmed that the three RBD proteins can be detected in cells transfected with the trivalent mRNA vaccine. Furthermore, ex vivo imaging analysis indicated that the livers of BALB/c mice had the strongest protein expression levels after intramuscular (IM) injection. Using a prime-boost strategy, this trivalent vaccine elicited robust humoral and T-cell immune responses in both the high-dose and low-dose groups and showed no toxicity in BALB/c mice. Three specific IgG antibodies in the high-dose group against SARS-CoV-2, Delta, and Omicron variants approached ∼1/1,833,333, ∼1/1,866,667, and ∼1/925,000, respectively. Taken together, two doses of inoculation with the trivalent mRNA vaccine may provide broad and effective immunization responses against SARS-CoV-2 and variants.

Obtaining a high titer of polyclonal antibodies from rats to the SARS-CoV-2 nucleocapsid protein and its N- and C-terminal domains for diagnostic test development

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is an enveloped, plus-stranded RNA virus responsible for the Coronavirus Disease 2019 (COVID-19). Patients infected with COVID-19 may be asymptomatic or have symptoms ranging from mild manifestations to severe cases of the disease that could lead to death. The SARS-CoV-2 genome encodes 4 structural proteins, including the Spike protein (S), the Nucleocapsid protein (N), Membrane protein (M) and, the Envelope protein (E). The N protein forms a major component of the ribonucleoprotein complex within the virus particle and play a vital role in its transcription and replication. Nevertheless, the S protein was the most important protein in the development of vaccines against COVID-19. However, the decrease in number of registered immunizations against the disease and the rapid drop in neutralizing antibody titers together with looser preventive measures for virus transmission, favored the rapid appearance of new variants of concerns (VOCs) that primarily show mutations in the S protein. This fact makes the N protein a good candidate for the development of diagnostic tests, due to its stability, amino acid conservation, high immunogenicity, and the smaller likelihood of mutation. With the aim of developing a new diagnostic kit based on the N protein, we evaluated the humoral response in female Wistar rats against this target. Three constructions of the N protein were used to inoculate the animals: the full-length protein (Cfull), the N- (NTD), and the C-terminal (CTD) portion of the protein. The immunizations induced the animal's immune response, with specific polyclonal IgG antibodies against the Cfull protein and its fragments. There were not non-specific bind to the protein used as negative control. Anti-Cfull antibodies demonstrated high efficiency in binding to the NTD protein and the antibodies present in the anti-CTD and anti-NTD sera have recognized the Cfull protein, but they were not able to recognize the NTD and CTD proteins, respectively. Our results indicate an efficient protocol for obtaining high antibody titers against the N recombinant protein of SARS-CoV-2 and its fragments highlighting the Cfull protein, which can be used in the development of new diagnostic kits.

A seromolecular study to determine the prevalence of cytomegalovirus in pregnant women referred to health centers in the north of Iran.

Because of the controversial aspects of the CMV virus during pregnancy, it should be considered a serious health threat, especially in developing countries. The present seromolecular study aimed to determine cytomegalovirus prevalence in pregnant women referred to health centers in the north of Iran. One hundred and twenty-five pregnant women who were referred to health centers in Mazandaran province for regular health checks were randomly selected from Jan 2022 to Oct 2022. To detect the presence of the CMV genome and specific IgM and IgG antibodies against cytomegalovirus, the conventional PCR and ELISA tests were applied respectively. All 125 pregnant women that attended the study were from Mazandaran province with a mean age of 30 years ranging from 20 to 42 years. The result showed that 2 (1.6%), 92 (73.6%), and 2 (1.6%) of the cases were positive for IgM, IgG, and IgM/IgG, respectively. The PCR test results indicated that the CMV DNA was present in 10 (8%) pregnant women. Our study shows that all PCR-positive cases were negative for the IgM test. Of the 10 PCR-positive samples 3 were positive and 1 was suspicious for the IgG test. Our study revealed that there is an urgent need for vaccination or other strategies to prevent and treat congenital CMV infection. Reducing the burden of congenital CMV infection requires global awareness. Further studies are recommended to obtain accurate estimates of the risk of congenital CMV infection.

Evaluation of SARS-CoV-2 Specific Antibodies in Recovered Patients by Different ELISA Kits.

The coronavirus disease 2019 (COVID-19) was first reported in December 2019 in Wuhan, Hubei Province of China. As long as the 27th of December 2021, approximately 280 million people have been infected with coronavirus, resulting in more than 5,418,421 deaths worldwide. Since the beginning of the COVID-19 pandemic, different methods were introduced for diagnosing coronavirus-infected patients and evaluating the immune response, following the vaccination. The current study aimed to compare the level of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) specific IgG in a group of patients who recovered from COVID-19, measured by three different enzyme-linked immunosorbent assay (ELISA) kits. This cross-sectional study was conducted on sera from patients who recovered from a real-time reverse transcriptase-polymerase chain reaction (RT-PCR)-confirmed COVID-19 in Birjand, South Khorasan, Iran. SARS-CoV-2 anti-nucleocapsid (N) and spike (S) protein IgG levels were measured using commercial ELISA kits. Comparison between groups was made using one-way ANOVA and Tukey post hoc tests. The mean titer of anti-N IgG was significantly higher for the PishtazTeb Diagnostics kit than the Ideal Tashkhis Atieh kit (p<0.05). There was no correlation between the titer of anti-N IgG (PishtazTeb Diagnostics and Ideal Tashkhis Atieh) and anti-S IgG (Chemobind Company) antibodies. This study indicates that the domestic ELISA kits have variable but acceptable sensitivity for detecting SARS-CoV-2 specific IgG antibodies.

Is there a reactive IgG antibody for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in maternal breast milk?

Restricted data on the transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) reactive IgG antibodies by breast milk to infants, born to mothers who did not know if they had coronavirus disease 2019 (COVID-19), are presented. The objective of the present work was to demonstrate the existence of specific IgG antibodies against the SARS‐CoV‐2 virus in the milk of breastfeeding mothers in the suburban area of Şanlıurfa, Türkiye. This retrospective analysis examined the medical records of maternal and neonatal data for all 51 newborns to 50 mothers from September to October 2020. The present work was approved by the local medical ethics committee. Written informed consent was obtained from the mothers. Testing SARS-CoV-2 reactive IgG was performed using ELISA assay. SARS-CoV-2 reactive IgG were detected in 58% of breast milk. Over the past year, SARS-CoV-2 reactive IgG identified in breast milk of mothers without symptoms of viral respiratory infection (self-reported diagnosis) was 12.5 times lower than mothers with symptoms. The IgG responses were different between symptomatic and asymptomatic mothers’ and infants’ samples. These results propose that along with the COVID-19 pandemic, breastfeeding could be safe, and the existence of SARS-CoV-2-reactive IgG antibodies in breast milk could provide passive immunity to breastfed infants, and keep them safe against COVID-19 infection.

Immunogenicity and safety assessment of a SARS-CoV-2 recombinant spike RBD protein vaccine (Abdala) in paediatric ages 3–18 years old: a double-blinded, multicentre, randomised, phase 1/2 clinical trial (ISMAELILLO study)

COVID-19 in paediatric ages could result in hospitalizations and death. In addition, excluding children from vaccination could turn them into reservoirs of the SARS-COV-2. Safe and effective COVID-19 vaccines are urgently needed for large-scale paediatric vaccination. ISMAELILLO study aimed to evaluate safety and immunogenicity of two strengths of a new recombinant receptor-binding domain (RBD) protein vaccine (Abdala) in paediatric population. A double-blinded, multicentre, randomised, phase 1/2 clinical trial was conducted in nine polyclinics in the province of Camagüey, Cuba. Healthy children and adolescents were stratified according to age (3-11 years old, or 12-18 years old) and they were randomly assigned (1:1; block size four) in two dosage level groups of vaccine to receive three intramuscular doses of 25μg or 50μg of RBD, 14 days apart. Main safety endpoint was analyzed as the percentage of serious adverse reactions during vaccination up to 28 days after the third dose (Day 56) in participants who received at least one dose vaccination. The primary immunogenicity endpoint assessed was seroconversion rate of anti-RBD IgG antibody at day 56. The immunogenicity outcomes were assessed in the per-protocol population. This trial is registered with Cuban Public Registry of Clinical Trials, RPCEC00000381. Between July 15, 2021, and August 16, 2021, 644 paediatric subjects were screened, of whom 592 were enrolled after verifying that they met the selection criteria: firstly 88 were included in Phase 1 of the study and 504 who completed Phase 2. The vaccine was well tolerated. Injection site pain was the most frequently reported local event (143 [8·4%] of 1707 total doses applied), taking place in 66/851 (7·8%) in the 25μg group and in 77/856 (9·0%) in the 50μg. The most common systemic adverse event (AE) was headache: 23/851 (2·7%) in the 25μg group and 19/856 (2·2%) in the 50μg. Reactogenicity was mild or moderate in severity, represented in 75% of cases by local symptoms, completely resolved in the first 24-48h. Twenty-eight days after the third dose, seroconversion anti-RBD IgG were observed in 98·2% of the children and adolescents (231/234) for the 50μg group and 98·7% (224/228) for the 25μg group without differences between both strength. The specific IgG antibody geometric mean titres (GMT) showed higher titres between participants who received Abdala 50μg (231·3; 95% CI 222·6-240·4) compared to those who received 25μg (126·7; 95% CI 121·9-131·7). The mean ACE2 inhibition %, were 59·4% for 25μg, and for 50μg, 72·9% (p<0·01). Both strength elicited neutralising activity against the SARS-CoV-2, specifically (18·3; 95% CI 14·7-22·78) for Abdala 25μg and (36·4; 95% CI 30·26-43·8) for 50μg to the selected sample analyzed. Abdala vaccine was safe and well tolerated at both antigenic strength levels tested in participants aged between 3 and 18 years. Regarding immunogenicity, Abdala Vaccine stimulated the production of specific IgG antibodies against the RBD of SARS-CoV-2 as well as the production of ACE2 inhibition titres and neutralising antibodies (Nab) in children and adolescents. Centre for Genetic Engineering and Biotechnology (CIGB), Havana, Cuba.