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The progression of monoclonal gammopathies is affected by a range of factors, including the microenvironment surrounding plasma cells. It is recognized that TGF‐β1 plays a distinct role in stimulating IgA production. Hence, this study aims to investigate whether individuals with serum IgA monoclonal immunoglobulins (paraproteins) exhibit elevated total TGF‐β1 levels compared to those with IgG or IgM paraproteins. To achieve this goal, individuals with a positive laboratory finding of monoclonal gammopathy were segregated according to the paraprotein class as well as according to the type of the light chain. Total TGF‐β1 levels were assessed in blood serum samples containing IgG (n = 50), IgA (n = 46), and IgM (n = 31) paraproteins. Elevated level of TGF‐β1 was confirmed in sera with IgA paraproteins (median 25.8 ng/mL; interquartile range IQR: 19.0–33.7) compared to those having IgG (median: 18.2 ng/mL; IQR: 14.3–22.1; p < 0.001) or IgM paraproteins (21.5 ng/mL; IQR: 15.0–27.4; p = 0.043). Also, a higher TGF‐β1 level was detected in sera with IgMλ than those with IgMκ paraproteins (p = 0.043). This research affirms the role of TGF‐β1 in the pathophysiology of IgA monoclonal gammopathies and the potential switch towards the IgA isotype, known for a less favourable prognosis. [ABSTRACT FROM AUTHOR]

Unlike Fc receptors for switched immunoglobulin (Ig) isotypes, Fc receptor for IgM (FcµR) is selectively expressed by lymphocytes. The ablation of the FcµR gene in mice impairs B cell tolerance as evidenced by concomitant production of autoantibodies of IgM and IgG isotypes. In this essay, we reiterate the autoimmune phenotypes observed in mutant mice, ie IgM homeostasis, dysregulated humoral immune responses including autoantibodies, and Mott cell formation. We also propose the potential phenotypes in individuals with FCMR deficiency and the model for FcµR-mediated regulation of self-reactive B cells. [ABSTRACT FROM AUTHOR]

In teleosts, the immunoglobulin classes produced by B cells are IgM, IgD, and IgT/IgZ. IgT was initially described as an immunoglobulin specialized in mucosal responses; accumulating evidence, however, shows that it is also involved in systemic immune responses. Two types of IgT/IgZ (IgT1 and IgT2) were previously described in common carp, but their further characterization was hampered by the lack of specific tool. In the current study, we developed and validated polyclonal antibodies against carp IgT1 and IgT2 and used them in combination with well validated monoclonal antibody against carp IgM (WCI12), to study the distribution of IgM+, IgT1+ and IgT2+ B cells or their secreted immunoglobulins in various mucosal and systemic organs of carp. Finally, we also preliminary assessed the B cell response to infection with the blood-borne parasite Trypanoplasma borreli. Using these tools, we report on the distinct expression of soluble immunoglobulins in systemic and mucosal compartments. IgT1 and IgM were expressed in mucosal as well as systemic organs and responded to systemic parasitic infection, whereas IgT2 was preferentially expressed at mucosal sites and did not respond to systemic infections. By studying the distribution of B cells in different organs, compartmentalization of the three B cell subtypes was observed in gills and gut, whereas splenic B cells appeared as organized clusters around ellipsoids. Our results provide insights into the distribution and to some extent the function of B cells in carp, indicating that our newly developed tools are valuable for future studies aiming at the further characterization of immune responses of carp to infections and vaccination. [ABSTRACT FROM AUTHOR]

Introduction: Seroprevalence of anti-severe acute respiratory syndrome coronavirus 2 (anti-SARS-CoV-2) antibodies in the postvaccination period in Kenya remains to be elucidated. This study aimed to determine the seroprevalence of anti-SARS-CoV-2 IgM and IgG and evaluate Covid-19 vaccination uptake in a university setting in Nairobi. Methods: This cross-sectional study assayed serum anti-SARS-CoV-2 IgM and IgG levels using enzyme-linked immunosorbent assays. A structured questionnaire was used to determine vaccine uptake, vaccine hesitancy and reasons for hesitancy. Results: A total of 189 participants were enrolled (median age, 21 years; female, 50.8%). The seroprevalence of anti-SARS-CoV-2 was 12.7% for IgM and 87.8% for IgG. Anti-SARS-CoV-2 IgG titers were higher among the vaccinated vs. non-vaccinated individuals (p < 0.001, U = 2817.5), females vs. males (p = 0.024, U = 3616), and those vaccinated ≤ 6 months before the study vs. those vaccinated >1 year earlier (p = 0.002, H = 12.359). The vaccination hesitancy rate was 43.4% and the underlying reasons included mistrust (22.4%), health concerns (19.7%), and lack of information (18.4%). Discussion: The high seroprevalence of anti-SARS-CoV-2 IgG is an indication of high exposure to SARS-CoV-2 either through natural infection or through vaccination. The high vaccine hesitancy noted necessitates community engagement, and public education to dispel myths and misinformation prior to roll out of new vaccines and other health interventions. [ABSTRACT FROM AUTHOR]

Background and purpose Methods Results Conclusions Complement factor C2 is a potential therapeutic target in immune‐mediated neuropathies. However, literature suggests that classical complement pathway activation may proceed to C3 in the absence of C2, a so‐called “C2 bypass.” Here, we evaluated a C2 bypass mechanism during complement activation by pathogenic human IgM from patients with immune‐mediated neuropathies.IgM autoantibodies from 51 patients with multifocal motor neuropathy (MMN) or anti‐myelin‐associated glycoprotein (MAG) neuropathy (AMN) were used to activate complement in ex vivo disease models. C2 bypass was evaluated using C2‐depleted (C2D) serum and a therapeutic anti‐C2 antibody.In two different disease models of MMN, IgM anti‐GM1 and IgM anti‐GM2 autoantibodies from MMN patients were bound to induced pluripotent stem cell‐derived motor neurons and Schwann cells, respectively, and fixed C3 upon incubation with fresh serum. C3 fixation was inhibited by anti‐C2 and did not occur with C2D serum. Similarly, in an AMN model, IgM anti‐MAG antibodies were incubated with fresh serum fixed C3, which in all cases was abrogated in the absence of C2 or in the presence of anti‐C2.In ex vivo disease models of MMN and AMN, complement activation by IgM autoantibodies from 51 patients was in all cases dependent on C2 and was inhibited by an antihuman C2 antibody. No evidence of a C2 bypass mechanism was found. [ABSTRACT FROM AUTHOR]

Avian influenza viruses (AIVs) are a major economic burden to the poultry industry and pose serious zoonotic risks, with human infections being reported every year. To date, the vaccination of birds remains the most important method for the prevention and control of AIV outbreaks. Most national vaccination strategies against AIV infection use whole virus-inactivated vaccines, which predominantly trigger a systemic antibody-mediated immune response. There are currently no studies that have examined the antibody repertoire of birds that were infected with and/or vaccinated against AIV. To this end, we evaluate the changes in the H9N2-specific IgM and IgY repertoires in chickens subjected to vaccination(s) and/or infectious challenge. We show that a large proportion of the IgM and IgY clones were shared across multiple individuals, and these public clonal responses are dependent on both the immunisation status of the birds and the specific tissue that was examined. Furthermore, the analysis revealed specific clonal expansions that are restricted to particular H9N2 immunisation regimes. These results indicate that both the nature and number of immunisations are important drivers of the antibody responses and repertoire profiles in chickens following H9N2 antigenic stimulation. We discuss how the repertoire biology of avian B-cell responses may affect the success of AIV vaccination in chickens, in particular the implications of public versus private clonal selection. [ABSTRACT FROM AUTHOR]

Introduction: Antibody Fc regions harbour the binding sites for receptors that mediate effector functions following antigen engagement by the Fab regions. An extended "hinge" region in IgG allows flexibility between Fab and Fc, but in both the most primitive antibody, IgM, and in the evolutionarily more recent IgE, the hinge is replaced by an additional domain pair in the homodimeric six-domain Fc region. This permits additional flexibility within the Fc region, which has been exploited by nature to modulate antibody effector functions. Thus, in pentameric or hexameric IgM, the Fc regions appear to adopt a planar conformation in solution until antigen binding causes a conformational change and exposes the complement binding sites. In contrast, IgE-Fc principally adopts an acutely bent conformation in solution, but the binding of different receptors is controlled by the degree of bending, and there is allosteric communication between receptor binding sites. Methods: We sought to trace the evolution of Fc conformational diversity from IgM to IgE via the intermediate avian IgY by studying the solution conformations of their Fc regions by small-angle X-ray scattering. We compared four extant proteins: human IgM-Fc homodimer, chicken IgY-Fc, platypus IgE-Fc, and human IgE-Fc. These are examples of proteins that first appeared in the jawed fish [425 million years ago (mya)], tetrapod (310 mya), monotreme (166 mya), and hominid (2.5 mya) clades, respectively. Results and discussion: We analysed the scattering curves in terms of contributions from a pool of variously bent models chosen by a non-negative linear least-squares algorithm and found that the four proteins form a series in which the proportion of acutely bent material increases: IgM-Fc < IgY-Fc < plIgEFc < huIgE-Fc. This follows their order of appearance in evolution. For the huIgMFc homodimer, although none are acutely bent, and a significant fraction of the protein is sufficiently bent to expose the C1q-binding site, it predominantly adopts a fully extended conformation. In contrast, huIgE-Fc is found principally to be acutely bent, as expected from earlier studies. IgY-Fc, in this first structural analysis of the complete Fc region, exhibits an ensemble of conformations from acutely bent to fully extended, reflecting IgY's position as an evolutionary intermediate between IgM and IgE. [ABSTRACT FROM AUTHOR]

A 42-year-old male was referred to the internal medicine department because of renal failure and persistent malaise after a recent SARS-CoV-2 infection. Blood results showed anemia and severe renal insufficiency (hemoglobin of 10.3 g/dL and a creatinine of 2.19 mg/dL). Additional tests revealed a type I cryoglobulinemia with a cryoprecipitate composed of dual IgM (kappa and lambda). Further investigations on the cryoprecipitate revealed that the immunoglobulins were directed against SARS-CoV-2 antigens. In the meanwhile, our patient noticed improvement of his symptoms accompanied by resolution of laboratory abnormalities. Three months later, the cryoglobulin could no longer be detected. Type 1 cryoglobulinemia is usually associated with lymphoproliferative disorders and is characterized by various symptoms caused by cryoprecipitates occluding small blood vessels. This is, to our knowledge, the first case of type I cryoglobulinemia with proven precipitation of SARS-CoV-19 antibodies. COVID-19 induced cryoglobulinemia appears to have a mild disease course and to be self-limiting upon viral clearance. [ABSTRACT FROM AUTHOR]

Significance: Fidelity of intercellular communication depends on unambiguous interactions between protein ligands and membrane receptors. Most proteins destined to the extracellular space adopt the required three-dimensional shape as they travel through the endoplasmic reticulum (ER), Golgi complex, and other organelles of the exocytic pathway. However, some proteins, many of which are involved in inflammation, avoid this classical secretory route and follow unconventional pathways to leave the cell. Recent Advances: Stringent quality control systems operate in the ER and cis-Golgi, restricting transport to native conformers, devoid of non-native disulfides and/or reactive thiols. However, some proteins released by living cells require reduced cysteines to exert their extracellular function(s). Remarkably, these proteins lack the secretory signal sequence normally required by secretory proteins for translocation into the ER lumen. Critical Issues: Why do interleukin-1β, high mobility group box 1, and other proinflammatory proteins avoid the ER–Golgi route to reach the intercellular space? These proteins require reactive cysteines for exerting their function. Therefore, eluding thiol-mediated quality control along the exocytic pathway is likely one of the main reasons why extracellular proteins that need to be reduced utilize unconventional pathways of secretion, where a quality control aimed at oxidating native cysteines is not present. Future Directions: Particularly under stress conditions, cells release redox-active enzymes and nonprotein thiol compounds that exert an extracellular control of redox-sensitive protein activity, shaping inflammatory responses. This post-secretion, redox-dependent editing of protein messages is still largely undefined. Understanding the underlying mechanistic events will hopefully provide new tools to control inflammation. Antioxid. Redox Signal. 41, 693–705. [ABSTRACT FROM AUTHOR]

Background: The coronavirus, SARS-CoV-2, is the causative agent for COVID-19, first registered in Wuhan, China and responsible for more than 6 million deaths worldwide. Currently, RT-PCR is the gold-standard method for diagnosing COVID-19. However, serological tests are needed for screening acute disease diagnosis and screening large populations during the COVID-19 outbreak. Objectives: Herein, we described the development and validation of an in-house enzyme-linked immunosorbent assay (ELISA) for detecting the levels of anti-spike-1-RBD IgM antibody (CovIgM-ELISA) in well-defined serum/plasma panel for screening and identifying subjects infected with SARS-CoV-2 in a Latin population. Method: In-house CovIgM-ELISA has the format of an indirect ELISA. It was optimized by checkerboard titration using recombinant SARS-CoV-2 spike-S1-RBD protein as an antigen. Results: We found that, compared to the RT-PCR as the standard method, the in-house CovIgM-ELISA displayed sensitivities of 96.15% and 93.22% for samples collected up to 30 or 60 days after infection, respectively, as well as 95.59% specificity with 97.3% accuracy. The agreement kappa value (κ) of our CovIgM-ELISA was substantial when compared to RT-PCR ( κ = 0.873) and the anti-SARS-CoV-2 IgM ELISA (InBios Int) ( κ = 0.684). The IgM levels detected in the population positively correlated with the neutralizing activity against the wild-type, Alpha and Delta variants of concern, but failed to neutralize Omicron. Conclusions: These data indicate that our in-house CovIgM-ELISA is a compatible performing assay for the detection of SARS-CoV-2 infection. [ABSTRACT FROM AUTHOR]

Aim: This study investigated hepatitis E virus (HEV) prevalence among pregnant women in Siem Reap, Cambodia, by developing a cost‐effective, user‐friendly in‐house enzyme‐linked immunosorbent assay (ELISA) for detecting total anti‐HEV immunoglobulins (Ig). Methods: The in‐house ELISA was designed for large‐scale screening in resource‐limited settings. Its performance was benchmarked against two commercial tests: the Anti‐HEV IgG EIA (Institute of Immunology, Co. Ltd) and the Anti‐HEV IgG RecomLine LIA (Mikrogen). The in‐house ELISA demonstrated a sensitivity of 76% and 71.4%, and a specificity of 94.1% and 98.6%, against the two commercial tests, respectively, with overall agreement rates of 92.4% and 94.3%. Results: Among 1565 tested pregnant women, 11.6% were anti‐HEV positive. Prevalence increased with age, particularly in women aged 35–40 years and over 40 years. No significant associations were found with education, number of children, family size, or history of blood transfusion and surgery, except for the occupation of the family head as a public officer. Of the total anti‐HEV positive women, 22.7% had anti‐HEV IgM, indicating recent or ongoing infection. Conclusion: The study concluded that the in‐house ELISA is a viable option for HEV screening in regions with limited resources due to its high accuracy and cost‐effectiveness. It is particularly suitable for large‐scale studies and public health interventions in areas where HEV is endemic and poses a significant risk to pregnant women. [ABSTRACT FROM AUTHOR]

The aim of the study was to find out the pattern of IgM and IgG changes in patients with autoimmune thyroiditis depending on the pathological process duration. Materials and methods. A single-center cross-sectional study with randomization elements enrolled 170 patients with autoimmune thyroiditis, and 65 patients without thyroid pathology or other autoimmune diseases were assigned to sex- and age-matched comparison group (p = 0.6155 and p = 0.3093, respectively). The patients were classified according to thyroid status parameters into subclinical and manifest groups. All study participants were examined on IgM and IgG levels based not only on the clinical form of the disease, but also on the disease duration (up to 5 years and more than 5 years). The control group comprised 65 healthy individuals, including 26 men and 39 women (mean age 38.7 ± 10.8 years). Results. A slight decrease in IgM levels was observed in patients with subclinical form and longer disease duration, which was 1.5 (1.5; 1.7) g/l with the disease duration of up to 5 years and 1.4 (1.2; 1.4) g/l with the disease duration of more than 5 years, while there were no differences in IgM levels in patients with manifest form with longer disease duration. IgG concentrations were statistically significantly higher in both clinical groups of patients with the disease duration of up to 5 years compared to those in patients with the disease duration of more than 5 years (13 (11; 14) g/l up to 5 years and 11 (10; 12) g/l more than 5 years in subclinical group, p < 0.05); 13 (12; 14) g/l up to 5 years and 12 (10; 15) g/l more than 5 years in manifest group, p < 0.05). Conclusions. A downward trend in IgG concentrations is noted with the disease progression and longer duration, while IgM levels are uninformative.

Since May 2022, mpox has been identified in 108 countries without endemic disease; most cases have been in gay, bisexual, or other men who have sex with men. To determine number of missed cases, we conducted 2 studies during June-September 2022: a prospective serologic survey detecting orthopoxvirus antibodies among men who have sex with men in San Francisco, California, and a retrospective monkeypox virus PCR testing of swab specimens submitted for other infectious disease testing among all patients across the United States. The serosurvey of 225 participants (median age 34 years) detected 18 (8.0%) who were orthopoxvirus IgG positive and 3 (1.3%) who were also orthopoxvirus IgM positive. The retrospective PCR study of 1,196 patients (median age 30 years; 54.8% male) detected 67 (5.6%) specimens positive for monkeypox virus. There are likely few undiagnosed cases of mpox in regions where sexual healthcare is accessible and patient and clinician awareness about mpox is increased.

Abstract Using a modified proximity extension assay, total and immunoglobulin (Ig) class-specific anti-SARS-CoV-2 antibodies were sensitively and conveniently detected directly from ø1.2 mm discs cut from dried blood and saliva spots (DBS and DSS) without the need for elution. For total Ig detection, antigen probes were prepared by conjugating recombinant spike protein subunit 1 (S1-RBD) to a pair of oligonucleotides. To detect isotype-specific antibody reactivity, one antigen probe was replaced with oligonucleotide-conjugated antibodies specific for antibody isotypes. Binding of pairs of oligonucleotide-conjugated probes to antibodies in patient samples brings oligonucleotides in proximity. An added DNA polymerase uses a transient hybridization between the oligonucleotides to prime synthesis of a DNA strand, which serves as a DNA amplicon that is quantified by real-time PCR. The S1-RBD-specific IgG, IgM, and IgA antibodies in DBS samples collected over the course of a first and second vaccination exhibited kinetics consistent with previous reports. Both DBS and DSS collected from 42 individuals in the autumn of 2023 showed significant level of total S1-RBD antibodies with a correlation of R = 0.70. However, levels in DSS were generally 10 to 100-fold lower than in DBS. Anti-S1-RBD IgG and IgA in DSS demonstrated a correlation of R = 0.6.

Abstract Streptococcus suis (S. suis) is one of the most important porcine pathogens, causing severe pathologies such as meningitis or polyarthritis. It is also a very successful colonizer of mucosal surfaces. The IgM-degrading enzyme of S. suis (Ide Ssuis ) specifically cleaves porcine IgM, which results in complement evasion. On the basis of our previous finding that Ide Ssuis also cleaves the IgM B cell receptor in vitro, we verified IgM B cell receptor cleavage ex vivo in whole regional lymph nodes and investigated the working hypothesis that this IgM B cell receptor cleavage results in a long-lasting impaired B cell function. The number of IgM-secreting cells was determined via ELISpot analysis after porcine peripheral blood mononuclear cells had initially been treated with different recombinant S. suis proteins and subsequently stimulated with interleukin-2 and the toll-like receptor 7/8 ligand R848. Compared with treatment with medium or recombinant muramidase-released protein, treatment with rIde Ssuis but also with a cleavage-deficient variant led to a reduction in the number of IgM-secreting cells as well as the level of secreted IgM. Flow cytometry analysis confirmed that the IgM B cell receptor was cleaved only by rIde Ssuis, and the receptor recovered to pretreatment levels on day 2 after treatment. Flow cytometry analysis of B and T cells incubated with fluorescein-labelled recombinant proteins revealed that different rIde Ssuis variants bind specifically to B cells, most prominently the cleavage-deficient variant. Our results indicate that in vitro interference of rIde Ssuis with the IgM B cell receptor results in long-lasting impaired IgM secretion by B cells after toll-like receptor activation. Further studies are warranted to prove that the modulation of B cell function by Ide Ssuis could play a role in vivo.

Relevance. The production of brucellosis diagnosticums is currently being established in the medical and veterinary practice of the Republic of Uzbekistan. However, due to the lack of a national standard serum in the country and the high cost of analog standard serums on the market, problems may arise in the production of diagnostic tools. In addition, the lack of national standard serums causes problems in quality control of commercial and local antigens, hinders the development of local drugs – brucellosis diagnostics, which reduces the effectiveness of serological diagnosis of brucellosis in humans and animals. Given the above, it becomes necessary to develop and produce a standard serum with a high level of safety, which will improve the methods of laboratory diagnosis of various forms of brucellosis in patients and animals to identify hidden foci of infection in Uzbekistan. The aim of the study is to obtain and characterize experimental standard diagnostic brucellosis rabbit sera for the development of national and industry standard serum with appropriate titers. Materials and Methods. The experiments were carried out in full compliance with the European Council Directive 2010/63/EU on compliance with ethical principles in working with laboratory animals and the experiments were carried out on the basis of the methodological manual «Methods and rules of work» approved by the Ministry of Health of the Republic of Uzbekistan in 2016 with laboratory animals in experimental microbiological and immunological studies. To obtain positive standard sera against brucellosis pathogens, 12 rabbits weighing from 2.2 kg to 4.6 kg, aged 6 to 12 months, were used. The animals were kept under identical conditions on a standard vivarium diet. For the experiment, animals adapted to the conditions of the study were selected and kept in quarantine for 21 days. The maintenance and conduct of the studies followed international rules for the humane treatment of animals. Results and Discussion. During the evaluation of the heterologic specificity of the obtained sera for 23 collection and hospital strains, a weakly positive response (+) was observed in 16.7% of cases. Rabbit sera positive for heterologic specificity were adsorbed on suspensions of inactivated microorganisms (Citrobacter spp. and Enterobacteriaceae spp.) - no significant drop in titer was observed, and cross-agglutination was eliminated. A significant increase in the total protein level was noted after the first and fourth immunizations. Before the first immunization, the total protein level was 59.57 g/l, and after the fourth immunization, it increased threefold to 157.17 g/l. There was also an increase in albumin and globulin (33.2 g/L and 26.37 g/L, respectively) to 79.37 g/L and 77.43 g/L, respectively. IgM levels remained almost unchanged, increasing 3.5-fold after the second immunization and decreasing to 0.24 mg/ml after the fourth immunization. An increase in the level of IgG after the second and third immunizations was revealed, followed by a decrease after the fourth immunization. Conclusion. The obtained national and industry standard diagnostic polyvalent brucellosis sera, as well as the developed methods of laboratory control of its quality and efficiency testing will make it possible to produce candidate standard serum samples for agglutination reaction test and detection of Brucella strains. The obtained standard serum samples will be further used for control of the produced immunobiological diagnostic preparations and provision of unified requirements to their quality. The goal was achieved – improving the serological diagnosis of brucellosis, laying the foundation for further study and standardization of serum.

Streptococcus suis (S. suis) is one of the most important porcine pathogens, causing severe pathologies such as meningitis or polyarthritis. It is also a very successful colonizer of mucosal surfaces. The IgM-degrading enzyme of S. suis (IdeSsuis) specifically cleaves porcine IgM, which results in complement evasion. On the basis of our previous finding that IdeSsuis also cleaves the IgM B cell receptor in vitro, we verified IgM B cell receptor cleavage ex vivo in whole regional lymph nodes and investigated the working hypothesis that this IgM B cell receptor cleavage results in a long-lasting impaired B cell function. The number of IgM-secreting cells was determined via ELISpot analysis after porcine peripheral blood mononuclear cells had initially been treated with different recombinant S. suis proteins and subsequently stimulated with interleukin-2 and the toll-like receptor 7/8 ligand R848. Compared with treatment with medium or recombinant muramidase-released protein, treatment with rIdeSsuis but also with a cleavage-deficient variant led to a reduction in the number of IgM-secreting cells as well as the level of secreted IgM. Flow cytometry analysis confirmed that the IgM B cell receptor was cleaved only by rIdeSsuis, and the receptor recovered to pretreatment levels on day 2 after treatment. Flow cytometry analysis of B and T cells incubated with fluorescein-labelled recombinant proteins revealed that different rIdeSsuis variants bind specifically to B cells, most prominently the cleavage-deficient variant. Our results indicate that in vitro interference of rIdeSsuis with the IgM B cell receptor results in long-lasting impaired IgM secretion by B cells after toll-like receptor activation. Further studies are warranted to prove that the modulation of B cell function by IdeSsuis could play a role in vivo. [ABSTRACT FROM AUTHOR]

Natural antibodies (NAbs) occurring in individuals without prior exposure to specific antigens, provide direct first barrier protection against pathogens, and exert immunoregulation thus actively contributing to the maintenance of immune homeostasis, controlling inflammatory processes and preventing autoimmunity. Common variable immunodeficiency (CVID) is a heterogeneous group of disorders characterized by a compromised immune function that brings into focus the role of NAbs. Our aim was to explore whether NAb levels could serve as potential key indicators in CVID for monitoring disease progression and predicting outcomes. In this study, we analyzed a Hellenic cohort of 56 patients with CVID (31 newly diagnosed and 25 under immunoglobulin replacement therapy-IgRT) and 33 healthy controls, for total Ig levels and serum IgM and IgG NAb levels against five informative target-antigens of NAbs, namely, actin, DNA, carbonic anhydrase, F(ab΄)2 fragments of human IgG and TriNitroPhenyl. In addition, follow-up pre- and post- IgRT samples were analyzed in ten (10) patients of our cohort. Results showed that Ig-treated patients exhibited significantly lower IgM NAb levels than untreated patients and healthy controls against all panel antigens. In the follow-up samples, pre-treatment IgM NAb levels negatively correlated with total serum IgM. This imbalance was only partially restored after IgRT, with a significant decrease in IgM NAb levels observed in nine out of ten patients. Moreover, post-treatment patients with recurrent infections presented significantly lower IgM NAb levels, a reduction also observed in patients with bronchiectasis independently of treatment status. On the contrary, post-treatment patients with enteropathy had significantly higher IgM NAb levels against all panel antigens, an increase also noted in patients with autoimmune diseases. Regarding IgG NAbs, replacement therapy restored levels to those of healthy controls. In conclusion, impaired NAb levels are found in CVID patients, particularly related to certain phenotypes. Moreover, the significant decrease in IgM NAb levels after IgRT suggests a potential association with disease course and complications. The results suggest that administration of human IgM NAbs may be an effective combinatorial treatment in selected patients. Further research is needed to understand the functional roles of NAbs in CVID and its complex clinical phenotypes. [ABSTRACT FROM AUTHOR]

Using a modified proximity extension assay, total and immunoglobulin (Ig) class-specific anti-SARS-CoV-2 antibodies were sensitively and conveniently detected directly from ø1.2 mm discs cut from dried blood and saliva spots (DBS and DSS) without the need for elution. For total Ig detection, antigen probes were prepared by conjugating recombinant spike protein subunit 1 (S1-RBD) to a pair of oligonucleotides. To detect isotype-specific antibody reactivity, one antigen probe was replaced with oligonucleotide-conjugated antibodies specific for antibody isotypes. Binding of pairs of oligonucleotide-conjugated probes to antibodies in patient samples brings oligonucleotides in proximity. An added DNA polymerase uses a transient hybridization between the oligonucleotides to prime synthesis of a DNA strand, which serves as a DNA amplicon that is quantified by real-time PCR. The S1-RBD-specific IgG, IgM, and IgA antibodies in DBS samples collected over the course of a first and second vaccination exhibited kinetics consistent with previous reports. Both DBS and DSS collected from 42 individuals in the autumn of 2023 showed significant level of total S1-RBD antibodies with a correlation of R = 0.70. However, levels in DSS were generally 10 to 100-fold lower than in DBS. Anti-S1-RBD IgG and IgA in DSS demonstrated a correlation of R = 0.6. [ABSTRACT FROM AUTHOR]

Background: Pregnancy is a critical time for women, making them more susceptible to infectious diseases like COVID-19. This study aims to determine the immunogenicity of COVID-19 in pregnant women who have been infected compared to those who have received the inactive COVID-19 vaccine. Materials and methods: In this retrospective cohort study, pregnant women who received the inactivated COVID-19 vaccine (Sinopharm) and those with a history of COVID-19 infection during pregnancy were studied. Participants who had experienced stillbirth, received different COVID-19 vaccines, or had intrauterine fetal death were excluded from the study. Overall, the study included 140 participants. The participants were divided into two groups of 70 participants - pregnant women who received the Sinopharm vaccine and pregnant women who had COVID-19 infection during pregnancy. Before delivery, blood samples were collected from all mothers to evaluate the maternal immunoglobulin G (IgG) level. Blood samples were also taken from the baby's umbilical cord during delivery to measure the newborn's IgG level. Additionally, blood samples were collected from babies whose mothers showed signs of acute infection to measure their IgM levels and evaluate vertical transmission. Findings: The study found a significant relationship between the mean level of maternal IgG and umbilical cord IgG within the groups (P < 0.001). The highest levels of maternal IgG (2.50 ± 2.17) and umbilical cord IgG (2.43 ± 2.09) were observed in pregnant women with a previous COVID-19 infection and no history of vaccination (P < 0.001). Only one baby was born with a positive IgM, and this baby was born to a mother who showed signs of COVID-19 infection in the last five days of pregnancy. The mother was 28 years old, with a BMI of 33; it was her first pregnancy, and she gave birth to a male newborn at term. Conclusion: Administering an inactivated vaccine during pregnancy can generate immunity in both the mother and the child. However, the vaccine's immunity level may not be as potent as that conferred by COVID-19 infection during pregnancy. Nonetheless, the risk of vertical transmission of COVID-19 is considered minimal and can be classified as negligible. [ABSTRACT FROM AUTHOR]

The aim of the study was to find out the pattern of IgM and IgG changes in patients with autoimmune thyroiditis depending on the pathological process duration. Materials and methods. A single-center cross-sectional study with randomization elements enrolled 170 patients with autoimmune thyroiditis, and 65 patients without thyroid pathology or other autoimmune diseases were assigned to sex- and age-matched comparison group (p = 0.6155 and p = 0.3093, respectively). The patients were classified according to thyroid status parameters into subclinical and manifest groups. All study participants were examined on IgM and IgG levels based not only on the clinical form of the disease, but also on the disease duration (up to 5 years and more than 5 years). The control group comprised 65 healthy individuals, including 26 men and 39 women (mean age 38.7 ± 10.8 years). Results. A slight decrease in IgM levels was observed in patients with subclinical form and longer disease duration, which was 1.5 (1.5; 1.7) g/l with the disease duration of up to 5 years and 1.4 (1.2; 1.4) g/l with the disease duration of more than 5 years, while there were no differences in IgM levels in patients with manifest form with longer disease duration. IgG concentrations were statistically significantly higher in both clinical groups of patients with the disease duration of up to 5 years compared to those in patients with the disease duration of more than 5 years (13 (11; 14) g/l up to 5 years and 11 (10; 12) g/l more than 5 years in subclinical group, p < 0.05); 13 (12; 14) g/l up to 5 years and 12 (10; 15) g/l more than 5 years in manifest group, p < 0.05). Conclusions. A downward trend in IgG concentrations is noted with the disease progression and longer duration, while IgM levels are uninformative. [ABSTRACT FROM AUTHOR]

(1) Background: Horses infected by a tick-borne encephalitis virus (TBEV) can develop clinically apparent infections. In humans, vaccination is the most effective preventive measure, while a vaccine is not available for horses. The objective of this study was to describe the immune response in horses after a TBEV vaccination with a human vaccine. (2) Materials and Methods: Seven healthy horses were randomised to a treatment or a control group in a stratified fashion based on TBEV–IgG concentrations on day −4. The treatment group (n = 4) was intramuscularly vaccinated using an inactivated human TBEV vaccine on days 0 and 28; the control group (n = 3) did not receive an injection. A clinical examination and blood sampling were performed on day –4, 0, 2, 4, 6, 8, 10, 14, 28, 30, 32, 34, 36, 38, 43, 56, 84, and 373. A linear mixed model analysis was used to compare IgG and IgM concentrations, neutralising antibody (nAb) titres, leucocyte count, serum amyloid A (SAA), and fibrinogen and globulin concentrations between the groups and time points. (3) Results: The clinical examination was normal in all horses at all time points. There were no significant changes in SAA, globulin, and fibrinogen concentrations and leucocyte count between the groups or time points (all p > 0.05). There was no significant increase in IgG, IgM, or nAb titres in the control group over time (all p > 0.05). In the vaccination group, there was a significant increase in IgG concentration and nAb titres after the second vaccination (p < 0.0001). There was no significant increase in IgM antibodies after the TBEV vaccination (all p > 0.05). One horse in the vaccination group had an IgM concentration above the laboratory reference on day 10. (4) Conclusions: The human TBEV vaccine did not have side effects when used in healthy horses in this study. A significant rise in TBEV-specific IgG antibodies and nAbs after the second vaccination was observed. However, IgG and nAb titres have been shown to decrease within 1 year after vaccination. The results of this study indicate that a vaccination with a human vaccine only induces a mild rise in IgM antibodies and only in previously naive horses. With no significant changes to inflammatory parameters in the vaccinated horses, it remains unclear whether vaccination with the human vaccine leads to protective immunity. [ABSTRACT FROM AUTHOR]

B-1a cells, a regulatory subset of B lymphocytes, produce natural IgM and interleukin-10. Neutrophil extracellular traps (NETs) play a crucial role in pathogen defense, but their excessive formation during sepsis can cause further inflammation and tissue damage. In sepsis, extracellular cold-inducible RNA-binding protein (eCIRP), a damage-associated molecular pattern, is released to induce NET formation. We hypothesize that B-1a cells clear NETs to prevent sepsis-induced injury. Sepsis in mice was induced by injecting 1 × 107 and 5 × 107 colony-forming units of Escherichia coli intraperitoneally. After 4 and 20 h, we assessed the number of B-1a cells in the peritoneal cavity using flow cytometry. Our results showed that the number of peritoneal B-1a cells was significantly decreased in E. coli sepsis mice. Importantly, replenishing B-1a cells via intraperitoneal injection in sepsis mice significantly decreased NETs in peritoneal neutrophils. We also observed a decrease in serum inflammation and injury markers and a significant increase in the overall survival rate in B-1a cell–treated septic mice. To understand the mechanism, we cocultured bone marrow–derived neutrophils with peritoneal B-1a cells in a contact or noncontact condition using an insert and stimulated them with eCIRP. After 4 h, we found that eCIRP significantly increased NET formation in bone marrow–derived neutrophils. Interestingly, we observed that B-1a cells inhibited NETs by 67% in a contact-dependent manner. Surprisingly, when B-1a cells were cultured in inserts, there was no significant decrease in NET formation, suggesting that direct cell-to-cell contact is crucial for this inhibitory effect. We further determined that B-1a cells promoted NET phagocytosis, and this was mediated through natural IgM, as blocking the IgM receptor attenuated the engulfment of NETs by B-1a cells. Finally, we identified that following their engulfment, NETs were localized into the lysosomal compartment for lysis. Thus, our study suggests that B-1a cells decrease NET content in eCIRP-treated neutrophils and E. coli sepsis mice. [ABSTRACT FROM AUTHOR]

HEV antibody detection constitutes the main screening test for HEV infection. The aim of this study is to compare the sensitivity and specificity of four techniques: LIAISON® MUREX DiaSorin anti-HEV IgG and anti-HEV IgM assays, Hepatitis E VIRCLIA® IgM and IgG monotests, WANTAI HEV-IgM and IgG ELISA and VIDAS® anti-HEV IgM and IgG tests in five panels of samples configurated according to the immunoblot (RecomLine, Mikrogen, Neuss, Germany). Anti-HEV IgM sensitivity in the acute phase was 100% in all techniques, while sensitivity, including the immediate convalescence phase, was 96.74% for LIAISON®, 83.14% for VIRCLIA®, 84.78% for WANTAI and 88.04% for VIDAS®. Anti-HEV IgM specificity was 100% for both LIAISON® and VIRCLIA®. Anti-HEV IgM WANTAI agreed with VIRCLIA® with a good Kappa coefficient (κ = 0.71). Anti-HEV IgG post-infection sensitivity was 100% for LIAISON®, VIDAS® and VIRCLIA® and 99% for WANTAI. Anti-HEV IgG specificity reached 97.17% for LIAISON and 88.68% for VIRCLIA®. Our results demonstrated a better capacity of LIAISON® MUREX anti-HEV IgM than that of competitors for detecting acute infections as well as accurate anti-HEV IgG results and in how to resolve them. [ABSTRACT FROM AUTHOR]

Background: Infections frequently occur after orthotopic liver transplantation (OLT) and are associated with increased mortality. In 2018, we introduced perioperative administration of intravenous immunoglobulin enriched in IgM as an optional therapy in recipients at a high risk of infection. This preliminary study evaluated whether this preparation reduced infections in the early post-transplantation period. Methods: Adult patients with a high risk of postoperative infections who underwent OLT between January 2014 and December 2021 in our center were included in the study. The primary outcome was the occurrence of new postoperative bacterial and fungal infections within the first 30 days after OLT. Results: Ninety recipients at a high risk of postoperative infections who underwent OLT were included, of whom 51 (57%) received IgM preparation. Patients treated and not treated with IgM were similar in terms of demographics, model of end-stage liver disease score, and risk factors for postoperative infections. The occurrence of new infections was lower (absolute risk reduction (ARR) 21.2%; p = 0.038) in patients who received IgM than in those who did not. Multivariate analysis adjusted for confounders (OR 0.348; p = 0.033) and propensity score-based matching analysis (ARR 21.2%, p = 0.067) confirmed an association between IgM preparation and lower occurrence of postoperative infections. The 90-day mortality rate was lower (ARR 13.4%, p = 0.018) in patients who received IgM preparation. Conclusions: In OLT recipients at high risk for infections, perioperative administration of an IgM-enriched preparation seems to reduce the development of new infections within the first 30 days after OLT. [ABSTRACT FROM AUTHOR]

Background: Production of anti-phosphatidylserine (anti-PS) antibodies has been associated with malaria and can aggravate pathology. How these autoantibodies develop during early childhood in a malaria context is not known. We examined levels of anti-PS IgG and IgM antibodies in a longitudinal cohort of mother-baby pairs during birth, in the infants at 2.5, 6 months, and in mothers and their babies at 9 months postpartum. Results: There was no difference between levels of anti-PS IgG in cord blood and the mothers' peripheral blood at birth. However, anti-PS IgM levels were significantly higher in the mothers compared to the infants' cord blood, and IgM levels were steadily increasing during the first 9 months of the infants' life. In infants that had the highest anti-PS IgM levels at birth, there was a decline until 6 months with a rise at 9 months. Infants that possessed high anti-PS IgG at birth also exhibited a progressive decline in levels. When anti-PS were correlated to different fractions of B-cells, there were several correlations with P. falciparum specific atypical B cells both at birth and at 2.5 months for the infants, especially for anti-PS IgM. Anti-PS also correlated strongly to C1q-fixing antibodies at birth. Conclusion: These results show that anti-PS IgG acquired by mothers could be transferred transplacentally and that IgM antibodies targeting PS are acquired during the first year of life. These results have increased the knowledge about autoimmune responses associated with infections in early life and is critical for a comprehensive understanding of malaria vaccine functionality in endemic areas. [ABSTRACT FROM AUTHOR]

Introduction: Dengue, also known as "break bone fever," is a viral infection transmitted by Aedes aegypti followed by Aedes albopictus. It is more prevalent in tropical and subtropical climates. While many individuals infected with dengue may not experience any symptoms, those who do may commonly exhibit high fever, headache, body aches, nausea, and rash. In severe cases, dengue can be fatal, with some individuals developing dengue haemorrhagic fever characterized by bleeding, low blood platelet levels, and blood plasma leakage, or dengue shock syndrome, which involves dangerously low blood pressure. Aim: To detect Dengue IgM with IgM capture ELISA in NS1 positive samples of patients attending a tertiary care hospital. Material and Methods: This study was conducted as a prospective cohort study, conducting within the study period June 2022 to December 2022 involving patients admitted to GBCM and Subharti hospital, Dehradun. A Comparative study to detect Dengue IgM with IgM capture ELISA in NS1 positive samples of patients attending a tertiary care hospital among 498 patients. Results: Among 258 female patients, 24 were NS1-positive, 36 were IgM-positive, and 46 were IgG-positive and in male patients (240), 59 were NS1 positive, 40 were IgM positive, and 39 were IgG positive. In July 2022, a higher correlation coefficient of 0.422029 (p-value: 0.000570) was observed, indicating a stronger association between NS1 antigen and IgM seropositivity. Thrombocytopenia with most cases showing platelet counts ranging from 50,000 to 100,000 platelets/cu mm. Conclusion: This study enhances our understanding of the correlation between IgM seropositivity and NS1 antigen positivity in dengue cases within rural populations. The findings have significant implications for early detection, diagnosis, and management of dengue infection, emphasizing the value of IgM as a diagnostic marker. [ABSTRACT FROM AUTHOR]

Background: SARS-Cov-2 caused a viral pandemic that started in late December 2019. This study aimed to investigate the humoral immunity response against SARS-Cov-2 by evaluating the B cell counts and serum levels of IgG and IgM in different groups of patients. Methods: A total of 74 samples were collected from different groups of confirmed COVID-19 patients to evaluate the concentrations of IgG and IgM in groups including control, mild, severe, and asymptomatic. In addition, 84 samples were collected for CD19+ and CD27+ B cells analysis in the same group. Results: Levels of IgG and IgM were found to be significantly elevated in COVID-19 patients; the severe groups showed the highest level among the groups of patients. Furthermore, a significant decrease in B cell counts in patients was observed. Conclusion: Peripheral B cells, IgG and IgM were shown to be related to the severity of the disease. While CD19+ and CD27+ B cells tended to decrease in groups of patients, antibodies’ concentrations increased along with disease severity [ABSTRACT FROM AUTHOR]

Purpose: Tick-borne encephalitis virus (TBEV) infections result in severe central nervous system diseases in humans across Asia and Europe. In China, cases of tick-borne encephalitis are primarily caused by the Far East subtype of TBEV, which exhibits a distinct disease course compared to other extensively studied subtypes. However, there is limited knowledge regarding the nucleic acid and serological diagnostic characteristics of patients infected with the TBEV in China, which is the focus of investigation in the present study. Methods: This study established a TaqMan qPCR approach to detect TBEV RNA in the serum with optimal specificity, sensitivity, and precision. Using TaqMan qPCR and ELISA assay for TBEV IgM detection, serum samples from 63 hospitalized patients bitten by ticks in Northeast China were investigated for diagnostic characteristics. Results: Twenty-five patients were positive for viral RNA; nineteen patients were positive for IgM, and nine were positive for both viral RNA and IgM. Through comparative analysis, TBEV RNA copies were negatively correlated with the virus incubation period. IgM levels were positively correlated with the clinical symptom scores of patients. The severity of clinical symptoms and the length after the tick bite could be used to predict the IgM occurrence. Furthermore, IgM levels and viral RNA copies were not correlated in double-positive patients. Conclusion: Both nucleic acid and serological detection methods exhibited distinct windows for detecting TBEV infection, with some overlap, and were associated with specific correlated factors. This study provided novel insights into the diagnosis and course of TBEV-induced tick-borne encephalitis in China. [ABSTRACT FROM AUTHOR]

Simple Summary: Toxoplasmosis is a disease caused by the parasite Toxoplasma gondii, which can infect humans and animals, including dogs. Dogs become infected by accidentally ingesting the parasite's oocysts from the environment and eating infected rodents or other small animals. Following infection, dogs develop antibodies against T. gondii. The current study investigated the percentage of dogs living in Greece that have antibodies against the parasite as an indirect way to assess the level of environmental contamination with T. gondii oocysts. Blood samples were collected from 1282 dogs living in both urban and rural regions of Greece, including Attica and Thessaloniki. Overall, 47.6% of the dogs had antibodies against the parasite, indicating they had been exposed to T. gondii. Seropositive dogs were more commonly found in rural areas (53.8%) than in urban areas (43.9%) (p < 0.001). The findings of this study suggest that T. gondii infection is widespread in dogs across Greece, especially in rural regions, and the parasite is highly present in the environment, posing a potential risk for human exposure in these areas. Control measures are necessary to prevent dogs from being infected and to reduce this risk for dog owners. Dog owners should also follow basic hygiene practices, like washing their hands after petting their dogs, to protect themselves. Toxoplasmosis, caused by the protozoan Toxoplasma gondii, is a zoonotic disease that affects various animal species, including dogs, that can serve as sentinels for indirectly estimating the environmental contamination. The current study aimed to determine the seroprevalence of T. gondii IgG and IgM antibodies in dogs across different regions of Greece and assess their living area as a potential risk factor. In total, 1282 blood samples were collected from dogs in urban and rural areas of Greece, including Attica and Thessaloniki. Serum samples were tested for T. gondii-specific IgG and IgM antibodies using an indirect immunofluorescent antibody test. A chi-square test was performed to assess the association between seropositivity for T. gondii and geographical location (urban/rural). The overall T. gondii seroprevalence was 47.6%, while the seroprevalence of IgG and IgM antibodies was 34.3% and 22.2%, respectively. Dogs from rural areas exhibited a significantly higher seroprevalence (53.8%) than those from urban areas (43.9%) (p < 0.001), with the estimated odds ratio being equal to 1.49 (95% CI, 1.18 to 1.65) and the relative risk increased by 22.4%. Dogs in Greece are highly exposed to T. gondii, particularly in rural areas. Measures to prevent canine infections are necessary, and basic hygiene practices, such as hand washing after petting dogs, are required to reduce human infection risk and safeguard public health. [ABSTRACT FROM AUTHOR]

Introduction: Coronavirus Disease 2019 (COVID-19) is a severe respiratory illness caused by the RNA virus SARS-CoV-2. Globally, there have been over 759.4 million cases and 6.74 million deaths, while Ecuador has reported more than 1.06 million cases and 35.9 thousand deaths. To describe the COVID-19 pandemic impact and the vaccinations effectiveness in a low-income country like Ecuador, we aim to assess the seroprevalence of IgG and IgM antibodies against SARS-CoV-2 in a sample from healthy blood donors at the Cruz Roja Ecuatoriana. Methods: The present seroprevalence study used a lateral flow immunoassay (LFIA) to detect anti-SARS-CoV-2 IgG and IgM antibodies in months with the highest confirmed case rates (May 2020; January, April 2021; January, February, June, July 2022) and months with the highest vaccination rates (May, June, July, August, December 2021) in Quito, Ecuador. The IgG and IgM seroprevalence were also assessed based on sex, age range, blood type and RhD antigen type. The sample size was 8,159, and sampling was performed based on the availability of each blood type. Results: The results showed an overall IgG and IgM seroprevalence of 47.76% and 3.44%, respectively. There were no differences in IgG and IgM seroprevalences between blood groups and sex, whereas statistical differences were found based on months, age range groups, and RhD antigen type. For instance, the highest IgG seroprevalence was observed in February 2022 and within the 17-26 years age range group, while the highest IgM seroprevalence was in April 2021 and within the 47-56 years age range group. Lastly, only IgG seroprevalence was higher in RhD+ individuals while IgM seroprevalence was similar across RhD types. Discussion: This project contributes to limited data on IgG and IgM antibodies against SARS-CoV-2 in Ecuador. It suggests that herd immunity may have been achieved in the last evaluated months, and highlights a potential link between the RhD antigen type and COVID-19 susceptibility. These findings have implications for public health strategies and vaccine distribution not only in Ecuador but also in regions with similar characteristics. [ABSTRACT FROM AUTHOR]

In response to the 2015 Zika virus (ZIKV) epidemic that occurred in Brazil, numerous commercial serological assays have been developed for clinical and research applications. Diagnosis of recent infection in pregnant women remains challenging. Having standardized, comparative studies of ZIKV tests is important for implementing optimal diagnostic testing and disease surveillance. This is especially important for serology tests used to detect ZIKV infection given that antibodies against ZIKV can cross-react with other arboviruses in the same virus family, such as dengue virus (DENV), yellow fever virus (YFV) and West Nile virus (WNV). We looked at the sensitivity and specificity of tests detecting ZIKV antibodies (IgM, IgG) from multiple manufacturers using panels of samples previously collected with known exposure to ZIKV and other arboviruses. We found that performance of the IgM tests was highly variable, with only one test (Inbios 2.0 IgM capture ELISA) having both high sensitivity and specificity. All IgG tests showed good sensitivity; however, specificity was highly variable, with some assays giving false-positive results on samples infected by another flavivirus. Overall, the results confirmed that accurate ZIKV antibody testing is challenging, especially in specimens from regions endemic for multiple other flaviviruses, and highlight the importance of available and suitable reference samples to evaluate ZIKV diagnostics. [ABSTRACT FROM AUTHOR]

SARS-CoV-2 is a highly pathogenic respiratory virus that successfully initiates and establishes its infection at the respiratory mucosa. However, little is known about how SARS-CoV-2 antagonizes the host's mucosal immunity. Recent findings have shown a marked reduction in the expression of the polymeric Ig receptor (pIgR) in COVID-19 patients. This receptor maintains mucosal homeostasis by transporting the dimeric IgA (dIgA) and pentameric IgM (pIgM) across mucosal epithelial cells to neutralize the invading respiratory pathogens. By studying the interaction between pIgR and SARS-CoV-2 proteins, we discovered that the viral accessory protein Open Reading Frame 8 (ORF8) potently downregulates pIgR expression and that this downregulation activity of ORF8 correlates with its ability to interact with pIgR. Importantly, the ORF8-mediated downregulation of pIgR diminishes the binding of dIgA or pIgM, and the ORF8 proteins of the variants of concern of SARS-CoV-2 preserve the function of downregulating pIgR, indicating the importance of this conserved activity of ORF8 in SARS-CoV-2 pathogenesis. We further observed that the secreted ORF8 binds to cell surface pIgR, but that this interaction does not trigger the cellular internalization of ORF8, which requires the binding of dIgA to pIgR. These findings suggest the role of ORF8 in SARS-CoV-2 mucosal immune evasion. [ABSTRACT FROM AUTHOR]

Cell adhesion molecule 1 (CADM1), a single-pass transmembrane protein, is involved in oncogenesis. We previously demonstrated the therapeutic efficacy of anti-CADM1 ectodomain monoclonal antibodies against mesothelioma; however, the underlying mechanism is unclear. In the present study, we explored the molecular behavior of anti-CADM1 antibodies in CADM1-expressing tumor cells. Sequencing analyses revealed that the anti-CADM1 chicken monoclonal antibodies 3E1 and 9D2 are IgY and IgM isotype antibodies, respectively. Co-administration of 3E1 and 9D2 altered the subcellular distribution of CADM1 from the detergent-soluble fraction to the detergent-resistant fraction in tumor cells. Using recombinant chicken–mouse chimeric antibodies that had been isotype-switched from IgG to IgM, we demonstrated that the combination of the variable region of 3E1 and the constant region of IgM was required for CADM1 relocation. Cytochemical studies showed that 3E1 colocalized with late endosomes/lysosomes after co-administration with 9D2, suggesting that the CADM1–antibody complex is internalized from the cell surface to intracellular compartments by lipid-raft mediated endocytosis. Finally, 3E1 was conjugated with the antimitotic agent monomethyl auristatin E (MMAE) via a cathepsin-cleavable linker. Co-administration of 3E1–monomethyl auristatin E and 9D2 suppressed the growth of multiple types of tumor cells, and this anti-tumor activity was confirmed in a syngeneic mouse model of melanoma. 3E1 and 9D2 are promising drug delivery vehicles for CADM1-expressing tumor cells. Proposed mechanism of the effects of 3E1 and 9D2 on CADM1 relocation. (1) 9D2-IgM interferes with CADM1 homophilic trans -binding in cell–cell interactions. (2) 3E1-IgG (along with 9D2-IgM) strongly binds to cell-surface CADM1, inducing CADM1 clustering by the combined effect of 3E1 epitope binding and the IgM structure, leading to CADM1 relocation to detergent-resistant microdomains (DRM). (3) As a result of clustering in DRMs, the CADM1–antibody complex is internalized by lipid raft-mediated endocytosis. [Display omitted] [ABSTRACT FROM AUTHOR]

The mechanisms driving metabolic reprogramming during B cell activation are unclear, particularly roles for enzymatic pathways involved in lipid remodeling. We found that murine B cell activation with lipopolysaccharide (LPS) led to a 1.6-fold increase in total lipids that included higher levels of phosphatidylethanolamine (PE) and plasmenyl PE. Selenoprotein I (SELENOI) is an ethanolamine phospholipid transferase involved in the synthesis of both PE and plasmenyl PE, and SELENOI expression was also upregulated during activation. Selenoi knockout (KO) B cells exhibited decreased levels of plasmenyl PE, which plays an important antioxidant role. Lipid peroxidation was measured and found to increase ∼2-fold in KO vs. wild-type (WT) B cells. Cell death was not impacted by KO in LPS-treated B cells and proliferation was only slightly reduced, but differentiation into CD138 + Blimp-1+ plasma B cells was decreased ∼2-fold. This led to examination of B cell receptors important for differentiation that recognize the ligand B cell activating factor, and levels of TACI (transmembrane activator, calcium-modulator, and cytophilin ligand interactor) (CD267) were significantly decreased on KO B cells compared with WT control cells. Vaccination with ovalbumin/adjuvant led to decreased ovalbumin-specific immunoglobulin M (IgM) levels in sera of KO mice compared with WT mice. Real-time polymerase chain reaction analyses revealed a decreased switch from surface to secreted IgM in spleens of KO mice induced by vaccination or LP-BM5 retrovirus infection. Overall, these findings detail the lipidomic response of B cells to LPS activation and reveal the importance of upregulated SELENOI for promoting differentiation into IgM-secreting plasma B cells. [ABSTRACT FROM AUTHOR]

Rift Valley fever (RVF) is a viral zoonosis, which is considered as a threat to food security in the Horn of Africa. In Kenya, RVF is the 5th ranked priority zoonotic disease due to its high morbidity and mortality, frequent outbreak events, and associated socioeconomic impacts during outbreak events. In 2018, an RVF outbreak was confirmed in Kenya's Siaya, Wajir, and Marsabit counties. During this outbreak, 30 people were confirmed infected with RVF through laboratory tests; 21 in Wajir, 8 in Marsabit, and 1 in Siaya Counties.Seventy-five (75) households (15 cases and 60 controls) were selected and interviewed using a case-control study design in 2021 (?). A case was a household with a member who was diagnosed with RVF in 2018. In addition, a total of 1029 animals were purposively selected within these households and serologically tested for RVF. The study aimed to estimate the contribution of various risk factors to RVF human occurrence in Kenya with a special focus on Wajir County. Wajir County was chosen due to high number of confirmed human cases reported in the 2018 outbreak. A univariable regression model revealed that owner-reported RVF virus exposure in livestock significantly increased the odds of an RVF human case in the household by 32.7 times (95 % CI 4.0–267.4). The respondent being linked to a goat flock that was IgG-positive increased the odds of an RVF human case by 3.8 times (95 % CI 1.17–12.3). In the final multivariable analysis, the respondent being linked to their own animals affected by RVF increased odds of having an RVF human case in the household by 56.9 times (95 % CI 4.6–700.4), while the respondent being linked to a neighbor household member affected decreased odds of having a RVF human case by 0.1 times (95 % CI 0.08–0.75).In summary, these results have revealed a potential link for the spread of RVF infection from animals to humans in pastoralist households, hence it is critical to carry out targeted, community education, One Health surveillance, prevention, and control measures against the disease. This will be critical to protecting humans against potential spillovers of infections during outbreak events in livestock.

Aim: To correlate hematological, inflammatory indicators and serological responses among COVID-19 patients to point out the significant biomarkers for disease management and prognosis.Materials & methods: Standard analytical and molecular methods were used to assess various inflammatory and serological Responses among COVID-19 patients (ICU- (n = 99) and non-ICU patients (n = 64) as compared with health control (n = 40).Results: Significant differences in the Hb, WBC, Lymphocyte count, CRP and serum ferritin (p

ABSTRACTDengue is often misclassified and underreported in Africa due to inaccurate differential diagnoses of nonspecific febrile illnesses such as malaria, sparsity of diagnostic testing and poor clinical and genomic surveillance. There are limited reports on the seroprevalence and genetic diversity of dengue virus (DENV) in humans and vectors in Nigeria. In this study, we investigated the epidemiology and genetic diversity of dengue in the rainforest region of Nigeria. We screened 515 febrile patients who tested negative for malaria and typhoid fever in three hospitals in Oyo and Ekiti States in southern Nigeria with a combination of anti-dengue IgG/IgM/NS1 rapid test kits and metagenomic sequencing. We found that approximately 28% of screened patients had previous DENV exposure, with the highest prevalence in persons over sixty. Approximately 8% of the patients showed evidence of recent or current infection, and 2.7% had acute infection. Following sequencing of sixty samples, we assembled twenty DENV-1 genomes (3 complete and 17 partial). We found that all assembled genomes belonged to DENV-1 genotype III. Our phylogenetic analyses showed evidence of prolonged cryptic circulation of divergent DENV lineages in Oyo state. We were unable to resolve the source of DENV in Nigeria owing to limited sequencing data from the region. However, our sequences clustered closely with sequences in Tanzania and sequences reported in Chinese with travel history to Tanzania in 2019. This may reflect the wider unsampled bidirectional transmission of DENV-1 in Africa, which strongly emphasizes the importance of genomic surveillance in monitoring ongoing DENV transmission in Africa.

ABSTRACT Serology testing is commonly used to evaluate the immunogenicity of COVID-19 vaccines and measure antibodies as a marker of previous infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In this study, four laboratory-developed serology enzyme-linked immunosorbent assays (SARS-CoV-2 anti-Spike and anti-Nucleocapsid immunoglobin G [IgG] and immunoglobin M [IgM]) calibrated to the WHO International Standard 20/136 were validated via analytical measuring interval (limit of blank [LOB], limit of detection [LOD], and limit of quantification [LOQ]), linearity, and precision according to the Clinical and Laboratory Standards Institute (CLSI) guidelines EP17-A2, EP06 2nd Edition, and EP05-A3. For Spike IgG, LOB was 3.0 binding antibody units per milliliter (BAU/mL), LOD was 4.1 BAU/mL, and LOQ was 27.1 BAU/mL. For Nucleocapsid IgG, LOB was 1.9 BAU/mL, LOD was 3.2 BAU/mL, and LOQ was 24.6 BAU/mL. For Spike IgM, LOB was 57.1 BAU/mL, LOD was 69.0 BAU/mL, and LOQ was 113.5 BAU/mL. For Nucleocapsid IgM, LOD was 242.2 BAU/mL, LOD was 289.9 BAU/mL, and LOQ was 572.4 BAU/mL. Each assay displayed good linearity (max % deviation from linearity (≥LOQ) = 10.7%). The result of within-run repeatability evaluation for medium positive samples was 7.7% for Spike IgG, 4.6% for Nucleocapsid IgG, 7.5% for Spike IgM, and 10.1% for Nucleocapsid IgM. The total precision, including medium positive sample variability across 20 days, three reagent kits, and two operators, was 13.5% for Spike IgG, 14.5% for Nucleocapsid IgG, 17.6% for Spike IgM, and 16.2% for Nucleocapsid IgM. The assays were successfully validated following the applicable CLSI guidelines. All assays met the ±20% deviation from linearity and the ±20% coefficient of variation specification for precision and repeatability.IMPORTANCEReliable and validated serology assays are of increasing importance as the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus continues to evolve and cause outbreaks. Validation of serology assays along with calibration to the International and National Standards (such as anti-SARS-CoV-2 Immunoglobulin WHO International Standard 20/136 or Frederick National Laboratory for Cancer Research’s National Serology Standard COVID-NS01097) is critical to ensuring that results from clinical studies are reliable and comparable among various assays and laboratories. We describe the design and execution of a comprehensive study that established the analytical measuring intervals, linearity, precision, and repeatability of four in-house developed serology enzyme-linked immunosorbent assays (SARS-CoV-2 anti-Spike immunoglobin G [IgG] and immunoglobin M [IgM] and anti-Nucleocapsid IgG and IgM) following applicable Clinical and Laboratory Standards Institute (CLSI) guidelines. Overall, this study provides practical guidance on experimental design strategies and data analysis techniques, pertaining to the validation of COVID-19 serology assays according to CLSI guidelines, for use in clinical research studies.

IntroductionStreptococcus suis serotype (cps) 2 is an important bacterial pathogen in piglet nurseries and an emerging zoonotic agent without effective vaccines available. Immunoglobulin (Ig)M plays an essential role in host defense against S. suis. In mice, non-conventional B-1 cells are a major source of protective IgM against encapsulated bacterial pathogens, such as S. pneumoniae. Two IgM+CD21− B-1-like cell subpopulations, distinguishable by CD11R1 expression, were described in pigs, but their properties and functions are poorly understood. This study aimed at a first characterization of the porcine early IgM B cell response against S. suis cps 2.MethodsWe analyzed the same healthy pigs, naturally colonized by different S. suis serotypes, including cps 2, at four and eight weeks of age serologically and determined the frequency of different peripheral B cell subpopulations by flow cytometry. Furthermore, we isolated conventional IgM+CD21+ B-2 cells as well as non-conventional B-1-like cell subpopulations from peripheral blood of eight-weeks-old pigs to evaluate their potential of IgM secretion in response to innate and adaptive stimuli in vitro.ResultsBetween the fourth and eighth week of life, a characteristic increase of S. suis cps 2-binding serum IgM antibodies, restricting bacterial growth, was observed. Moreover, we show for the first time that the significant increase of anti-S. suis serum IgM is associated with a relative increase of peripheral non-conventional IgM+CD21− B-1-like cells in vivo, particularly of the IgM+CD21− CD11R1− subpopulation. Noteworthy, sorted IgM+CD21− CD11R1− B-1-like cells from eight-weeks-old pigs spontaneously secreted IgM in vitro. In addition, both non-conventional IgM+CD21− B cell subpopulations, in contrast to conventional IgM+CD21+ B-2 cells, produced anti-S. suis IgM upon toll-like receptor (TLR) stimulation underlining their innate-like characteristics. We furthermore observed that both B-1-like subpopulations secrete S. suis cps 2-binding IgM upon stimulation with T cell-associated factors with highest amounts in IgM+CD21−CD11R1− B-1-like cells even exceeding anti-S. suis IgM levels produced by B-2 cells.ConclusionPorcine non-conventional B-1-like cells are a potent source of S. suis-binding IgM indicating a role in immunity during a critical phase of piglet rearing.

In teleosts, the immunoglobulin classes produced by B cells are IgM, IgD, and IgT/IgZ. IgT was initially described as an immunoglobulin specialized in mucosal responses; accumulating evidence, however, shows that it is also involved in systemic immune responses. Two types of IgT/IgZ (IgT1 and IgT2) were previously described in common carp, but their further characterization was hampered by the lack of specific tool. In the current study, we developed and validated polyclonal antibodies against carp IgT1 and IgT2 and used them in combination with well validated monoclonal antibody against carp IgM (WCI12), to study the distribution of IgM+, IgT1+ and IgT2+ B cells or their secreted immunoglobulins in various mucosal and systemic organs of carp. Finally, we also preliminary assessed the B cell response to infection with the blood-borne parasite Trypanoplasma borreli. Using these tools, we report on the distinct expression of soluble immunoglobulins in systemic and mucosal compartments. IgT1 and IgM were expressed in mucosal as well as systemic organs and responded to systemic parasitic infection, whereas IgT2 was preferentially expressed at mucosal sites and did not respond to systemic infections. By studying the distribution of B cells in different organs, compartmentalization of the three B cell subtypes was observed in gills and gut, whereas splenic B cells appeared as organized clusters around ellipsoids. Our results provide insights into the distribution and to some extent the function of B cells in carp, indicating that our newly developed tools are valuable for future studies aiming at the further characterization of immune responses of carp to infections and vaccination.

IntroductionSeroprevalence of anti-severe acute respiratory syndrome coronavirus 2 (anti-SARS-CoV-2) antibodies in the postvaccination period in Kenya remains to be elucidated. This study aimed to determine the seroprevalence of anti-SARS-CoV-2 IgM and IgG and evaluate Covid-19 vaccination uptake in a university setting in Nairobi.MethodsThis cross-sectional study assayed serum anti-SARS-CoV-2 IgM and IgG levels using enzyme-linked immunosorbent assays. A structured questionnaire was used to determine vaccine uptake, vaccine hesitancy and reasons for hesitancy.ResultsA total of 189 participants were enrolled (median age, 21 years; female, 50.8%). The seroprevalence of anti-SARS-CoV-2 was 12.7% for IgM and 87.8% for IgG. Anti-SARS-CoV-2 IgG titers were higher among the vaccinated vs. non-vaccinated individuals (p < 0.001, U = 2817.5), females vs. males (p = 0.024, U = 3616), and those vaccinated ≤ 6 months before the study vs. those vaccinated >1 year earlier (p = 0.002, H = 12.359). The vaccination hesitancy rate was 43.4% and the underlying reasons included mistrust (22.4%), health concerns (19.7%), and lack of information (18.4%).DiscussionThe high seroprevalence of anti-SARS-CoV-2 IgG is an indication of high exposure to SARS-CoV-2 either through natural infection or through vaccination. The high vaccine hesitancy noted necessitates community engagement, and public education to dispel myths and misinformation prior to roll out of new vaccines and other health interventions.

Ehrlichia canis, a widely distributed tick-borne pathogen, requires prompt and accurate diagnosis for effective management. Real-time PCR serves as the reference standard for diagnosing E. canis infection, whereas serological tests are crucial for detecting antibody responses indicative of infection or post-exposure status. Early detection during the acute phase of the disease is essential for optimal treatment and recovery. E. canis Tandem Repeat Protein 36 (TRP36) elicits early acute phase responses. We developed an enzyme-linked immunosorbent assay (ELISA) using recombinant TRP36 to detect IgM and IgG antibodies against E. canis. We assessed the diagnostic agreement, sensitivity, and specificity of the rTRP36-ELISA and a commercial test kit (SNAP 4Dx Plus Test) with TaqMan Real-time PCR (qPCR) using the NCSS 2023 program version 23.0.1. Leftover serum samples from 32 dogs at the Veterinary Teaching Hospital were randomly selected and examined for E. canis infection using qPCR, rTRP36-ELISA, and SNAP 4Dx Plus Test. qPCR detected positive results in 12 (37.5 %), whereas ELISA and SNAP 4Dx Plus detected positive results in 10 (31.25 %) and 13 (40.62 %), respectively. Moderate agreement (κ = 0.545) was observed between rTRP36-ELISA and qPCR, and fair agreement (κ = 0.379) between SNAP 4Dx Plus and qPCR. Substantial agreement (κ = 0.79) was found between rTRP36-ELISA and SNAP 4Dx Plus. Compared to qPCR, the sensitivity and specificity of rTRP36-ELISA were 70 % and 77.27 %, respectively, compared to 53.85 % and 73.86 %, respectively, for SNAP 4Dx Plus. Our findings suggest that TRP36 is a promising antigen for E. canis serodiagnosis, potentially improving sensitivity and specificity.

Avian influenza viruses (AIVs) are a major economic burden to the poultry industry and pose serious zoonotic risks, with human infections being reported every year. To date, the vaccination of birds remains the most important method for the prevention and control of AIV outbreaks. Most national vaccination strategies against AIV infection use whole virus-inactivated vaccines, which predominantly trigger a systemic antibody-mediated immune response. There are currently no studies that have examined the antibody repertoire of birds that were infected with and/or vaccinated against AIV. To this end, we evaluate the changes in the H9N2-specific IgM and IgY repertoires in chickens subjected to vaccination(s) and/or infectious challenge. We show that a large proportion of the IgM and IgY clones were shared across multiple individuals, and these public clonal responses are dependent on both the immunisation status of the birds and the specific tissue that was examined. Furthermore, the analysis revealed specific clonal expansions that are restricted to particular H9N2 immunisation regimes. These results indicate that both the nature and number of immunisations are important drivers of the antibody responses and repertoire profiles in chickens following H9N2 antigenic stimulation. We discuss how the repertoire biology of avian B-cell responses may affect the success of AIV vaccination in chickens, in particular the implications of public versus private clonal selection.

IntroductionAntibody Fc regions harbour the binding sites for receptors that mediate effector functions following antigen engagement by the Fab regions. An extended “hinge” region in IgG allows flexibility between Fab and Fc, but in both the most primitive antibody, IgM, and in the evolutionarily more recent IgE, the hinge is replaced by an additional domain pair in the homodimeric six-domain Fc region. This permits additional flexibility within the Fc region, which has been exploited by nature to modulate antibody effector functions. Thus, in pentameric or hexameric IgM, the Fc regions appear to adopt a planar conformation in solution until antigen binding causes a conformational change and exposes the complement binding sites. In contrast, IgE-Fc principally adopts an acutely bent conformation in solution, but the binding of different receptors is controlled by the degree of bending, and there is allosteric communication between receptor binding sites.MethodsWe sought to trace the evolution of Fc conformational diversity from IgM to IgE via the intermediate avian IgY by studying the solution conformations of their Fc regions by small-angle X-ray scattering. We compared four extant proteins: human IgM-Fc homodimer, chicken IgY-Fc, platypus IgE-Fc, and human IgE-Fc. These are examples of proteins that first appeared in the jawed fish [425 million years ago (mya)], tetrapod (310 mya), monotreme (166 mya), and hominid (2.5 mya) clades, respectively.Results and discussionWe analysed the scattering curves in terms of contributions from a pool of variously bent models chosen by a non-negative linear least-squares algorithm and found that the four proteins form a series in which the proportion of acutely bent material increases: IgM-Fc < IgY-Fc < plIgE-Fc < huIgE-Fc. This follows their order of appearance in evolution. For the huIgM-Fc homodimer, although none are acutely bent, and a significant fraction of the protein is sufficiently bent to expose the C1q-binding site, it predominantly adopts a fully extended conformation. In contrast, huIgE-Fc is found principally to be acutely bent, as expected from earlier studies. IgY-Fc, in this first structural analysis of the complete Fc region, exhibits an ensemble of conformations from acutely bent to fully extended, reflecting IgY’s position as an evolutionary intermediate between IgM and IgE.

Background The Chikungunya (CHIK) is a viral infection caused by CHIK virus which is an arbovirus that belongs to the genus alphavirus under the Togaviridae family. CHIK infection is transmitted to humans by the bite of mosquitoes namely Aedes albopictus and Aedes aegypti. Since Chikungunya fever leads to long term residual arthropathy – like complications which causes hampering in routine work of person after recovery, it is still believed that chikungunya fever is a major burden on public health even in post COVID-19 era. Aims and Objectives We need to understand and evaluate the seroprevalence of chikungunya with reference to its study period, seasonal tendency, need for hospitalization and age-sex distribution. Materials and Methods A retrospective study was done in 305 suspected samples of chikungunya infection with history of Fever along with joint pain, myalgia between January 2020 to December 2023. Separated serum samples suspected to have Chikungunya infection were subjected to IgM capture ELISA for detection of IgM anti-chikungunya antibodies using IgM antibody capture ELISA produced by National Institute of Virology, Pune, India (NIV, Pune, India). Various statistical tests were done to find out statistical significance with p-value < 0.05 kept as statistically significant. Results Total 305 samples were tested for Chikungunya IgM antibody detection. 55 out of 305 samples were tested positive for CHIK-IgM antibody (18.03%). Out of total 55 positive samples, 32 (58.18%) were belonged to Males and remaining 23 (41.81%) to females (p-value for gender distribution=0.224). The cases varied significantly by age group (p=0.0328), with the highest number of positives from 41-60 years age-group (38.18%). All positive cases having fever and joint pain as chief complaints (p