Synergistic polyreactivity of antibodies—where distinct oligo- or polyreactive clones combine to enable broad-spectrum antigen binding—was recently demonstrated for human anti-galactose-α-1,3-galactose (αGal) antibodies. However, it remains unclear whether this phenomenon also occurs in other antigen-defined antibody populations. Here, we investigated synergistic polyreactivity in isohemagglutinins (naturally occurring antibodies against ABO blood group antigens) and its potential role in antimicrobial defense. Isohemagglutinins were affinity-purified from pooled plasma from groups of healthy donors sharing the same ABO blood group, then characterized by solid-phase immunoassays, SDS-PAGE, and Western blotting. Binding to formaldehyde-fixed Streptococcus pneumoniae representing 30 serotypes was assessed by flow cytometry. Specificity was further examined using inhibition assays with soluble pneumococcal polysaccharides. To evaluate clinical relevance, ABO blood group types (used as proxies for isohemagglutinin profiles) were linked to serotype-specific invasive pneumococcal disease (IPD) events using Danish nationwide registry data. The purified isohemagglutinins retained specificity for their cognate ABO antigens. Notably, they bound all tested pneumococcal strains, primarily through reactivity with the common cell wall polysaccharide. Adsorption studies further revealed serotype-specific capsule binding mediated by distinct isohemagglutinin clones, demonstrating synergistic polyreactivity. Despite this, population-level analyses showed only modest protection: most notably, individuals with blood types B or O (producers of anti-A and anti-A,B isohemagglutinins, respectively) had a 19% lower prevalence of serotype 9V IPD than blood type A or AB individuals. Although the overall protective effect of isohemagglutinins against IPD seems limited, the observed synergistic polyreactivity supports the concept that this is a general feature of antigen-defined antibodies, extending beyond anti-αGal antibodies.
Bernth Jensen JM, Dinh KM, Hindhede L, Erikstrup LT, Hansen AG, et al.
Medical microbiology and immunology •
IgG antibodies against terminal galactose-α-1,3-galactose (anti-αGal antibodies) are naturally occurring in humans, but their origins remain poorly understood. These antibodies target various microorganisms including Staphylococcus aureus, a common nasal commensal and the major cause of skin abscesses. This study investigates the impact of S. aureus colonization and abscess events on plasma anti-αGal antibody levels. We measured plasma anti-αGal antibody levels using a quantitative immunoassay in: (i) 101 pairs of healthy individuals with and without nasal S. aureus colonization, (ii) 106 healthy individuals before and after abscess formation, and (iii) 43 patients with recurrent skin abscesses compared with 75 patient controls and 60 healthy controls. We observed a 35% reduction (95%CI: 7-54%) in anti-αGal antibody levels in nasal S. aureus carriers. Conversely, we found a 30% increase (95%CI: 4-66%) in individuals within 187 days post-skin abscess, and patients with recurrent skin abscesses exhibited 81% higher (95%CI: 14-190%) levels than patient controls, and 110% higher (95%CI: 39-230%) than healthy controls. This study suggests that skin abscesses lead to elevated plasma anti-αGal antibody levels and that these antibodies might convey or correlate with mucosal immunity to S. aureus.
Leth-Møller KB, van Hage M, Apostolovic D, Sørensen JA, Vestergaard C, et al.
Allergy •
Alpha-gal syndrome is a novel food allergy to the oligosaccharide galactose-α-1,3-galactose (alpha-gal) present in mammalian meat. Tick bites are considered an important route of sensitization to alpha-gal. Data on alpha-gal sensitization in the general population is scant. We utilized a unique data source of repeated population-based health examination studies to assess prevalence, time trends, risk factors, and characteristics of alpha-gal sensitization. Alpha-gal sensitization was assessed in >11.000 adults from four health examination studies of randomly invited residents in the Copenhagen region conducted in 1990-1991, 2011-2012, 2012-2015, and 2016-2017. Alpha-gal sensitization was defined as serum specific IgE (sIgE) to alpha-gal ≥0.1 kU/L; ≥0.35 kU/L; ≥0.7 kU/L; ≥3.5 kU/L. The population was characterized according to genetically determined ABO blood group, aeroallergen sensitization, and pets at home. The prevalence of sIgE to alpha-gal ≥0.1 kUA/L was 1.3% in 1990-1991, 3.7% in 2012-2015 and 3.2% in 2016-2017. Of those sensitized to alpha-gal >97% reported to consume red meat at least once a week, even for sIgE to alpha-gal ≥3.5 kU/L. Male sex, older age, aeroallergen sensitization, cat at home, and blood group A were associated with increased odds of alpha-gal sensitization. The known protective effect of blood group B was confirmed. In this general adult population, the prevalence of alpha-gal sensitization had doubled from 1990-1991 to 2016-2017. This could potentially be due to increased tick exposure and an increased atopic predisposition.
Pedersen HST, Sørensen JA, Madsen F, Linneberg A, Leth-Møller KB, et al.
Clinical and translational allergy •
Little is known about α-gal (galactose-α-1,3-galactose) sensitization in patients with chronic urticaria (CU). The aim of this study was to examine the prevalence, predictors and clinical relevance of α-gal sensitization in patients with CU. Two consecutive cohorts of newly referred patients with CU from a primary care allergology practice and a tertiary hospital dermatology department, plus a control group with allergic disease, but not CU, from the allergology practice, were interviewed and screened for α-gal sensitization (serum specific-IgE ≥0.35 KU/L). Of 733 patients included, 21 (5.6%) and 11 (3.9%) of CU patients from private practice and hospital, respectively, were α-gal sensitized. In total, 8 patients (38.1% of sensitized patients, and 2.1% of all CU patients) from private practice, and 2 patients (18.2% of sensitized patients, and 0.7% of all CU patients) from hospital, had clinically relevant α-gal allergy. In private practice, male sex (47.6 vs. 24.7%), p = 0.020, obesity (33.3 vs. 23.6%), p = 0.302, and frequency of angioedema (61.9 vs. 51.4%), p = 0.350; and in hospital, male sex (72.7 vs. 27.9%), p = 0.003, and high total immunoglobulin E (median 168 vs. 70.5 KU/L), p = 0.022 were associated with α-gal sensitization. α-gal sensitization is observed in a small fraction of CU patients with only few patients experiencing clinically relevant sensitization. Certain patients, particularly from primary care, may constitute a relevant population for aimed testing.
Bernth Jensen JM, Hansen AT, Söderström A, Jørgensen CS, Larsen CS, et al.
Clinical immunology (Orlando, Fla.) •
Functional antibody deficiency is clinically assessed from antibody responses to vaccination. However, diagnostic vaccination is complex and may fail in practice. We hypothesized that the levels of naturally occurring antibodies against galactose-α-1,3-galactose (αGal) may represent alternative markers of functional antibody capacity. We included data from 229 patients with suspected primary immunodeficiency in a retrospective study. Antibody levels against αGal and twelve pneumococcal serotypes were determined with solid-phase immunoassays. Pneumococcal vaccinations and treatment with normal human immunoglobulin were assessed from medical records. Anti-αGal antibody levels correlated positively with anti-pneumococcal antibody levels measured before and after pneumococcal vaccination. Contrary to the anti-pneumococcal antibody levels, the anti-αGal antibody level showed potential for predicting subsequent immunoglobulin treatment - a marker of disease severity. Naturally occurring antibodies may reflect the functional capacity tested by diagnostic vaccination but add more useful clinical data. The clinical utility of this easy test should be evaluated in prospective studies.
Naturally occurring antibodies are abundant in human plasma, but their importance in the defence against bacterial pathogens is unclear. We studied the role of the most abundant of such antibodies, the antibody against terminal galactose-α-1,3-galactose (anti-αGal), in the protection against pneumococcal infections (Streptococcus pneumonia). All known pneumococcal capsular polysaccharides lack terminal galactose-α-1,3-galactose, yet highly purified human anti-αGal antibody of the IgG class reacted with 48 of 91 pneumococcal serotypes. Anti-αGal was found to contain multiple antibody subsets that possess distinct specificities beyond their general reactivity with terminal galactose-α-1,3-galactose. These subsets in concert targeted a wide range of microbial polysaccharides. We found that anti-αGal constituted up to 40% of the total antibody reactivity to pneumococci in normal human plasma, that anti-αGal drives phagocytosis of pneumococci by human neutrophils and that the anti-αGal level was twofold lower in patients prone to pneumococcal infections compared with controls. Moreover, during a 48-year period in Denmark, the 48 anti-αGal-reactive serotypes caused fewer invasive pneumococcal infections (n = 10 927) than the 43 non-reactive serotypes (n = 18 107), supporting protection on the population level. Our findings explain the broad-spectrum pathogen reactivity of anti-αGal and support that these naturally occurring polyreactive antibodies contribute significantly to human protective immunity.
Some human antibodies may paradoxically inhibit complement activation on bacteria and enhance pathogen survival in humans. This property was also claimed for IgG antibodies reacting with terminal galactose-α-1,3-galactose (Galα3Gal; IgG anti-αGal), a naturally occurring and abundant antibody in human plasma that targets numerous different pathogens. To reinvestigate these effects, we used IgG anti-αGal affinity isolated from a pool of normal human IgG and human hypogammaglobulinaemia serum as a complement source. Flow cytometry was performed to examine antibody binding and complement deposition on pig erythrocytes, Escherichia coli O86 and Streptococcus pneumoniae serotype 9V. Specific nanobodies were used to block the effect of single complement factors and to delineate the complement pathways involved. IgG anti-αGal was capable of activating the classical complement pathway on all the tested target cells. The degree of activation was exponentially related to the density of bound antibody on E. coli O86 and pig erythrocytes, but more linearly on S. pneumoniae 9V. The alternative pathway of complement amplified complement deposition. Deposited C3 fragments covered the activating IgG anti-αGal, obstructing its detection and highlighting this as a likely general caveat in studies of antibody density and complement deposition. The inherent capacity for complement activation by the purified carbohydrate reactive IgG anti-αGal was similar to that of normal human IgG. We propose that the previously reported complement inhibition by IgG anti-αGal relates to suboptimal assay configurations, in contrast to the complement activating property of the antibodies demonstrated in this paper.
