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Antigenically sequence variable M proteins of the major bacterial pathogen Streptococcus pyogenes (Strep A) are responsible for recruiting human C4b-binding protein (C4BP) to the bacterial surface, which enables Strep A to evade destruction by the immune system. The most sequence divergent portion of M proteins, the hypervariable region (HVR), is responsible for binding C4BP. Structural evidence points to the conservation of two C4BP-binding sequence patterns (M2 and M22) in the HVR of numerous M proteins, with this conservation applicable to vaccine immunogen design. These two patterns, however, only partially explain C4BP binding by Strep A. Here, we identified several M proteins that lack these patterns but still bind C4BP and determined the structures of two, M68 and M87 HVRs, in complex with a C4BP fragment. Mutagenesis of these M proteins led to the identification of amino acids that are crucial for C4BP binding, enabling formulation of new C4BP-binding patterns. Mutagenesis was also carried out on M2 and M22 proteins to refine or generate experimentally grounded C4BP-binding patterns. The M22 pattern was the most prevalent among M proteins, followed by the M87 and M2 patterns, while the M68 pattern was rare. These patterns, except for M68, were also evident in numerous M-like Enn proteins. Binding of C4BP via these patterns to Enn proteins was verified. We conclude that C4BP-binding patterns occur frequently in Strep A strains of differing M types, being present in their M or Enn proteins, or frequently both, providing further impetus for their use as vaccine immunogens.

BACKGROUND: High levels of neutrophil extracellular trap (NET) formation or NETosis and autoantibodies are related to poor prognosis and disease severity of COVID-19 patients. Human angiotensin-converting enzyme 2 (ACE2) cross-reactive anti-severe acute respiratory syndrome coronavirus 2 spike protein receptor-binding domain (SARS-CoV-2 RBD) antibodies (CR Abs) have been reported as one of the sources of anti-ACE2 autoantibodies. However, the pathological implications of CR Abs in NET formation remain unknown. METHODS: In this study, we first assessed the presence of CR Abs in the sera of COVID-19 patients with different severity by serological analysis. Sera and purified IgG from CR Abs positive COVID-19 patients as well as a mouse monoclonal Ab (mAb 127) that can recognize both ACE2 and the RBD were tested for their influence on NETosis and the possible mechanisms involved were studied. RESULTS: An association between CR Abs levels and the severity of COVID-19 in 120 patients was found. The CR Abs-positive sera and IgG from severe COVID-19 patients and mAb 127 significantly activated human leukocytes and triggered NETosis, in the presence of RBD. This NETosis, triggered by the coexistence of CR Abs and RBD, activated thrombus-related cells but was abolished when the interaction between CR Abs and ACE2 or Fc receptors was disrupted. We also revealed that CR Abs-induced NETosis was suppressed in the presence of recombinant ACE2 or the Src family kinase inhibitor, dasatinib. Furthermore, we found that COVID-19 vaccination not only reduced COVID-19 severity but also prevented the production of CR Abs after SARS-CoV-2 infection. CONCLUSIONS: Our findings provide possible pathogenic effects of CR Abs in exacerbating COVID-19 by enhancing NETosis, highlighting ACE2 and dasatinib as potential treatments, and supporting the benefit of vaccination in reducing disease severity and CR Abs production in COVID-19 patients.

Background: Immunoglobulin (Ig)E cross‐reactivity has been shown between Dermatophagoides farinae (Df; house dust mite) and the nematode Toxocara canis (Tc), yet its allergen basis is unknown. Objective s : To identify the Df allergens IgE‐cross‐reactive with those of Tc. Animals: Archived sera from 73 dogs with suspected allergy sensitised to Df. Materials and Methods: We performed a combination of Pet Allergy Xplorer (PAX) and enzyme‐linked immunosorbent assay (ELISA) inhibitions with excretory–secretory and somatic (i.e. nematode body) extracts of Tc or recombinant Tc tropomyosin on coats of Df, Der f 15 and Zen‐1 (ELISA) or PAX allergens. Results: The ELISA and PAX inhibitions established that there is mutual yet variable cross‐reactivity between the Tc excretory–secretory extract, purified Der f 15 and purified Zen‐1. This cross‐reactivity is likely to involve cross‐reactive glycans, as there is no inhibition between the Tc excretory–secretory extract and recombinant Der f 15 without its predicted natural O‐glycans. We also confirmed a heterogeneous cross‐reactivity between the somatic Tc extract and Der p 11 (paramyosin), as well as between the recombinant Toxo c 3 and Der p 10 tropomyosins. The cross‐reactivity among tropomyosins and paramyosins is likely to involve peptidic epitopes, as these recombinant allergens are not glycosylated. Conclusions and Clinical Relevance: In dogs with suspected allergies, the cross‐reactivity between Tc and Df for dogs is complex and heterogeneous. Some of the cross‐reactive IgE recognises shared glycans on Der f 15 and Zen‐1, while some targets peptidic epitopes on shared paramyosins and tropomyosins. We do not exclude that additional cross‐reactive allergens between Df and Tc also might exist. [ABSTRACT FROM AUTHOR]

Accurate detection is a prerequisite for effective prevention and control of Mycoplasma synoviae infection. ELISA is the most popular method for the clinical detection of M. synoviae because of its convenience, low cost, and high detection rate. However, the cross-reactivity of commercially available ELISA kits with other avian pathogen-positive sera needs to be addressed. The aim of this study was to establish an ELISA method with high specificity for the detection of anti- M. synoviae antibodies in chicken serum to evaluate the M. synoviae infection status on poultry farms. The recombinant MS087 (rMS087) protein was expressed in Escherichia coli BL21 (DE3) and purified by Ni2+ affinity chromatography. An antibody against rMS087 was generated by immunizing BALB/c mice. Bioinformatic analysis revealed that MS087 was conserved among M. synoviae strains. Western blotting and indirect immunofluorescence results indicated that MS087 was not only localized in the cytoplasm and on the membrane but also secreted by the organism. For the established ELISA method based on rMS087, the optimal antigen concentration, blocking buffer, blocking duration, serum dilution, serum incubation duration, secondary antibody dilution, secondary antibody incubation duration and colorimetric reaction duration were 2 μg/mL, 1% BSA, 3 h, 1:500, 1.5 h, 1:20,000, 2 h and 5 min, respectively. Validation of the rMS087-based ELISA revealed a cut-off value of 0.5. The coefficients of variation of both the intra-batch and inter-batch methods were less than 9%. The assay was able to differentiate positive serum against M. synoviae from antisera against nine other avian pathogens and was able to recognize M. synoviae -positive sera at a dilution of 1:1,000. Compared with the commercial ELISA method, the rMS087-based ELISA has the potential to recognize more positive sera against M. synoviae. Collectively, the rMS087-based ELISA is a reproducible, specific, and sensitive serological method for detecting antibodies against M. synoviae in chicken serum and has robust potential for large-scale serological epidemiology of M. synoviae infection on poultry farms. [ABSTRACT FROM AUTHOR]

Anti‐immunocomplex (Anti‐IC) antibodies have been used in developing noncompetitive immunoassays for detecting small molecule analytics (haptens). These antibodies bind specifically to the primary antibody in complex with hapten. Although several anti‐IC antibody–based immunoassays have been developed, structural studies of these systems are very limited. In this study, we determined the crystal structures of anti‐testosterone Fab220 in complex with testosterone and the corresponding anti‐IC antibody FabB12. The structure of the ternary complex of testosterone, Fab220, and FabB12 was predicted using LightDock and AlphaFold. The ternary complex has a large (~ 1100 Å2) interface between antibodies. The A‐ring of the testosterone bound by Fab220 also participates in the binding of the anti‐IC antibody. The structural analysis was complemented by native mass spectrometry. The affinities for testosterone (TES) and three cross‐reactive steroids [dihydrotestosterone (DHT), androstenedione (A4), and dehydroepiandrosterone sulfate (DHEA‐S)] were measured, and ternary complex formation was studied. The results clearly show the ternary complex formation in the solution. Although DHT showed significant cross‐reactivity, A4 and DHEA‐S exhibited minor cross‐reactivity. [ABSTRACT FROM AUTHOR]

Patch testing for multiple cross-reactive allergens for allergic contact dermatitis (ACD) may not be necessary because of copositivity. We evaluated the formaldehyde group allergens to determine the optimal, most cost-effective allergens to test. A retrospective analysis of Mayo Clinic (1997-2022) examined the well-established copositive formaldehyde group: formaldehyde, quaternium 15, hexahydro-1,3,5-tris(2-hydroxyethyl)triazine, diazolidinyl urea, imidazolidinyl urea, toluenesulphonamide formaldehyde resin, DMDM hydantoin, and ethyleneurea melamine formaldehyde mix. Patch Optimization Platform identified which single formaldehyde-related allergen optimally captures patients with clinically relevant ACD. Next, Patch Optimization Platform determined the optimal additional 1, 2, 3, etc. allergens. Cost per patch test was $5.19 (Medicare 2022). A total of 9832 patients were tested for all listed allergens, with 830 having positive patch test results. Patch Optimization Platform determined that quaternium 15 alone captures 53% of patients with ACD to the formaldehyde group; adding the optimal second allergen (formaldehyde 1%) captures 78%; the optimal 5 top allergens capture >94% of patients. The incremental cost per additional diagnosis increased up to 44-fold as the number of allergens tested increased. Data are from a single institution, and the cost per test was fixed according to Medicare Part B in 2022. For diagnosing ACD, we recommend considering an optimized allergen selection algorithm. [ABSTRACT FROM AUTHOR]

Following successful oral immunotherapy (OIT) for peanut allergy using boiled peanuts (BOPI trial), this study investigated the potential of wet-thermal-processing-induced allergen modification, specifically soaking and boiling (1–4 h) to reduce the allergenicity of cashew and pistachio allergens. In addition, this study provides a foundation of understanding for developing safer forms of cashew/pistachio administration for application in OIT by gradual exposure to increasing doses of modified allergens with reduced potency as an "allergen ladder". An SDS-PAGE analysis and an intrinsic-fluorescence spectroscopy revealed altered tertiary structures of the allergens, leading to their denaturation and even degradation. Notably, the reduction in both allergen-specific polyclonal IgG and human-specific IgE (sIgE) binding correlated with the treatment time, with the most significant decrease observed after 4 h of boiling. In contrast, shorter soaking treatments showed negligible effects on the IgE-binding capacity of these nuts, therefore indicating a further need for optimization. These findings indicate that extended boiling effectively reduced the amounts of the highly potent allergenic component Ana o 3 in cashew and Pis v 1 in pistachio, as confirmed by ELISA using polyclonal anti-Ana o 3 antibodies, and an immunoblot showed decreased IgE epitope binding in cashew and pistachio allergens, which further modified their allergenic profiles. This approach shows promise as a viable method for offering a safer therapeutic option for cashew/pistachio allergy. [ABSTRACT FROM AUTHOR]

Lyme disease, a zoonotic infection caused by the bacterium Borrelia burgdorferi, is transmitted to humans through the bites of infected ticks. Its diagnosis primarily relies on serological methods; however, the existing borreliosis techniques have shown a variable sensitivity and specificity. Our study aimed to map IgG epitopes from five outer membrane proteins (Omp) from B. burgdorferi [Filament flagellar 41kD (PI1089), flagellar hook-associated protein (Q44767), Flagellar hook k2 protein (O51173), Putative Omp BURGA03 (Q44849), and 31 kDa OspA (P0CL66)] lipoprotein to find specific epitopes for the development of accurate diagnosis methods. Using the spot synthesis technique, a library of 380 peptides was constructed to identify linear B cell epitopes recognized by human IgG in response to specific B. burgdorferi-associated proteins. The reactivity of this epitope when chemically synthesized was then evaluated using ELISA with a panel of the patient's sera. Cross-reactivity was assessed through data bank access and in vitro analysis. Among the 19 epitopes identified, four were selected for further investigation based on their signal intensity, secondary structure, and peptide matching. Validation was performed using ELISA, and ROC curve analysis demonstrated a sensitivity of ≥85.71%, specificity of ≥92.31, accuracy of ≥90.7, and AUC value of ≥0.91 for all peptides. Our cross-reactivity analysis demonstrated that the Burg/02/huG, Burg/03/huG, and Burg/12/huG peptides were not reactive to antibodies from patients with Leptospirosis and syphilis compared to those from the B. burgdorferi group. These peptides indicated an excellent performance in distinguishing between B. burgdorferi-infected and non-infected individuals and exhibited a neglected reactivity to antibodies in sera from patients with Leptospirosis and syphilis. These peptides are promising targets for recombinant development, potentially leading to more accurate serological tests and vaccines. [ABSTRACT FROM AUTHOR]

Background: Common hops (Humulus lupulus L.) play a key role in brewing, providing the bitterness, flavor, and aroma of beer, and are widely used in supplements for their antibacterial, anti-inflammatory, and antioxidant properties. However, despite their broad applications, the allergenic potential of common hops remains underexplored, particularly when compared to the closely related Humulus japonicus. This preliminary study aimed to investigate the allergenic potential of common hops and their potential cross-reactivity with common pollen allergens. Methods: The immunoreactivity of hop stalks, leaves, and cones was assessed using antibodies against major allergens from birch (Bet v1a), mugwort (Art v1), and timothy grass (Phl p5b), as well as three sera from pollen-allergic patients. Slot Blot analysis was performed using phosphate-buffered saline extracts from the stalks, leaves, and cones of three hop cultivars, while Western Blotting followed SDS-PAGE protein separation. Results: The results revealed significant immunoreactivity in native hop proteins, with diminished reactivity observed in denatured proteins. Cross-reactivity between hop proteins and major pollen allergens was confirmed, indicating that hop proteins may contribute to allergic sensitization in pollen-sensitive individuals. Conclusions: These findings underscore the potential allergenic risks associated with the consumption or exposure to hop-containing products. [ABSTRACT FROM AUTHOR]

The prevalence of allergic diseases such as Allergic Rhinitis and Asthma is steadily increasing globally, with pollen allergy being one of the most significant sensitizing factors. However, the cross-reactivity of different pollen allergies remains unclear, posing challenges in the diagnosis and treatment of individuals with multiple sensitivities. In this study, the Component Resolved Diagnosis technique was performed to simultaneously measure the specific IgE concentrations of 52 patients against Art v and its components (Art v 1), Phl p and its components (Phl p 1, Phl p 4, Phl p 5, Phl p 6, Phl p 7, Phl p 12), Bet v and its components (Bet v 1, Bet v 2), Amb a and its component (Amb a 1), and Amb p. Additionally, sIgE inhibition tests were conducted by Art v, Phl p, and Bet v extracts. Among Art v-positive patients, 64.6% showed positivity for Art v 1. In Phl p-positive patients, Phl p 12 had the highest positivity rate (75.0%). Among Bet v-positive patients, 75.6% exhibited positivity for Bet v 2, whereas for Amb a and Amb p-positive patients, 23.7% and 29.0% respectively showed positivity for Amb a 1. The sIgE inhibition assays results revealed that Art v extract had inhibition rates greater than 73.2% against Phl p and its component Phl p 12, as well as Bet v and its component Bet v 2. Simultaneously, Phl p extract showed inhibition rates of 80.70–89.87% against Phl p 12, Bet v and Bet v 2. Bet v extract showed inhibition rates ranging from 21.9 to 59.8% against Phl p and Bet v 2, with a better inhibition rate (76.80%) against Phl p 12. In conclusion, Art v 1 is identified as the principal component of Art v. The profilin proteins of Phl p and Bet v (Phl p 12 and Bet v 2), are implicated as potential cross-reactive elements contributing to polysensitization in patients with respiratory allergies in the Northwest region of China. This cross-reactivity leads to a shared sensitization mechanism among pollen allergens such as Art v, Phl p, and Bet v. [ABSTRACT FROM AUTHOR]

Background Rapidly evolving RNA viruses, such as human norovirus, generate extraordinary sequence diversity, posing a significant challenge to vaccine design. This diversity, coupled with short-lasting natural immunity, leads to reinfection throughout one's lifetime. How reexposure shapes humoral immunity to future norovirus strains remains incompletely understood. Methods We profiled the antibody responses following 2 community gastroenteritis outbreaks with GII.2 and GII.6 noroviruses in 1971. Using diverse virus-like particles (VLPs), enzyme-linked immunosorbent assay (ELISA), and carbohydrate-blocking assays (surrogate for neutralization), we examined the antibody response at acute and convalescent timepoints following GII.6 infection. Results Convalescent sera displayed strong homologous blocking, demonstrating a 5-fold increase in GII.6 carbohydrate blockade over acute samples, and broad blocking of diverse archival and modern GII.6 noroviruses. Convalescent sera displayed limited carbohydrate blocking of heterotypic VLPs, despite high ELISA binding titers. Select individuals developed broad cross-genotype blockade, but this response was established before the second outbreak. Finally, we applied a novel competitive carbohydrate-blocking assay to demonstrate the epitope specificity and discrete compartments of the neutralizing response. Conclusions Our data show that infection generates narrow, focused immunity directed toward the infecting genotype. We did detect broad cross-blocking in specific individuals, but these responses could be attributed to diverse, genotype-specific antibodies predating GII.6 infection. [ABSTRACT FROM AUTHOR]

Background: Fenugreek, or Trigonella foenum greacum, is an ancient medicinal plant native to the eastern Mediterranean that later spread to Asia; it is used as a medicinal herb, spice, or food. It is a component of spice mixes, such as curry, and it is also used as a supplement in wheat and corn flour for bread making. Fenugreek appears to have many health benefits and potential medicinal properties; for this reason, it is increasingly being used in nutraceutical formulations. This study aimed to describe patients who visited our operative unit for a suspected adverse reaction after ingestion of fenugreek and to highlight possible cross-reactivity between fenugreek and other legumes or other foods. Methods: A total of 13 patients (5 males and 8 females; mean age: 30.6; range: 20–42 years) visited our outpatient clinic with a clinical history suggestive of an allergic reaction to spices or legumes. They underwent a skin prick test for fenugreek, fresh foods, and food extracts. Results: A total of 6 out of 13 patients positive for fenugreek had experienced allergic symptoms after ingesting foods containing spices, and 10 patients presented polysensitization with other legumes, of which 8 also presented sensitization to pistachios. Conclusions: Given the increasing use of novel spices in Mediterranean cuisine and the possible spread of fenugreek-based nutraceuticals, it is relevant to draw attention to possible allergic reactions. Further studies are also needed to be able to establish its cross-reactivity pattern. [ABSTRACT FROM AUTHOR]

Background: Hypersensitivity reactions (HSR) are reported for the macrolides, lincosamides, and streptogramins (MLS) antibiotic family. Data about cross‐reactivity among and between MLS remain scarce or controversial. Objectives: The aim of this study was to provide an overview of hypersensitivity cross‐reactions among MLSs based on data extracted from the French National Pharmacovigilance Database (FPVD). Methods: Cases of HSR to MLSs reported between January 1985 and December 2019 were extracted from the FPVD using standardized MedDRA queries (SMQ). Cases including an allergological test involving multiple MLSs and giving at least one positive result were included. Results: Of the 8394 cases reviewed, 149 were included. HSR mainly involved pristinamycin (n = 83; 53.2%) and spiramycin (n = 31; 19.9%). HSR to MLS was immediate in 54 cases and delayed in 94 cases. Skin tests represented the majority of the allergological tests performed (n = 728; 84.7%), followed by reintroduction tests (n = 79; 9.2%). Eighty‐six cross‐reactivities among MLS were identified in 62 cases (41.6%). All the 25 explorations performed for streptogramins showed cross‐reactivities, but only 30/253 among macrolides (11.9%). Cross‐reactivities between the three MLS were observed in 31/322 (9.6%) of the allergological explorations. Conclusion: This study highlights the possibility of cross‐reactivity among and between MLSs. Dermatologists and allergologists managing patients with HSR to MLSs should be aware of a risk of cross‐reactivity among the macrolides and between the different classes of MLS and to perform MLSs allergological testing before recommending an alternative antibiotic, especially in severe drug hypersensitivity from the MLS family. [ABSTRACT FROM AUTHOR]

Public health systems reported low mortality from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in East Asia, in low-income countries, and for children during the first year of the SARS-CoV-2 pandemic. These reports led commentators to suggest that cross-reactive immunity from prior exposure to other pathogens reduced fatality risk. Resolution of initial infection waves also contributed to speculation that herd immunity prevented further waves prior to vaccination. Serology instead implied that immunity was too limited to achieve herd immunity and that there was little impact from cross-reactive protection. Paediatric deaths exceeded those from influenza, with higher age-specific fatality risk in lower-income nations and similar fatality risk in East Asia compared with demographically similar regions. Neither pre-outbreak exposure to related pathogens nor immunity induced by initial infection waves are necessarily a reliable response to future pathogen outbreaks. Preparedness for future pathogen outbreaks should instead focus on strategies such as voluntary behavioural changes, nonpharmaceutical interventions, and vaccination. [ABSTRACT FROM AUTHOR]

In accordance with numerous national and international criteria, screening donated blood for abnormal antibodies against red cell antigens is crucial for patient safety. Antibodies can be formed by three mechanisms: by immune responses, naturally occurring, or passively acquired. In accordance with departmental policy, ICT was performed on a 30-yearold male blood donor's sample in addition to ABO Rh grouping. He tested A Rh Dnegative and had a positive ICT (+ 2). Anti-D was identified by using the antibody screening and identification panel. Retesting revealed similar results, so we called the donor. We were unable to find a source for the Rh isoimmunization during the course of his history. On further work-up, we found that antibodies reacted only in the antiglobulin phase with no reaction in the saline phase, suggesting an IgG type of antibody. Also, the anti-D titers (IgG) of the donor were up to 1:32. All potential sources of Rh isoimmunization have been ruled out for a male donor. Therefore, we assume that this donor has naturally occurring anti-D, perhaps created by certain environmental factors, bacteria, or components of the COVID vaccine that have cross-reactivity with the D antigen. This instance emphasizes how crucial the ICT test on the donor blood is. In our case, all potential causes of alloimmunization were ruled out, leaving only exposure to antigens with antigenic similarity to D antigens as a viable explanation. [ABSTRACT FROM AUTHOR]

Purpose of Review: There is growing evidence that enolase is involved in allergy. This manuscript reviews the impact of enolase in allergic disease and describes several sources of this allergen including molds, plants, animals, and pollens, among others. IgE epitopes are carefully analyzed as they may account for cross-reactivity. Recent Findings: Enolase has been previously associated to food allergy and contact dermatitis. However, other groups and we have identified recently novel enolases derived from diverse pollens in patients suffering asthma and allergic rhinitis. Exposure to outdoor enolases may cause respiratory disease. Summary: Enolase has been identified across various species and its amino acid sequence is highly conserved among different sources of this allergen. The demonstration that enolase is involved in many allergic diseases including respiratory allergies, is of clinic relevance. Thus, the development of novel molecular-based diagnostic and therapeutic strategies may pave the way for improved diagnosis and therapeutics. [ABSTRACT FROM AUTHOR]

Abstract The prevalence of allergic diseases such as Allergic Rhinitis and Asthma is steadily increasing globally, with pollen allergy being one of the most significant sensitizing factors. However, the cross-reactivity of different pollen allergies remains unclear, posing challenges in the diagnosis and treatment of individuals with multiple sensitivities. In this study, the Component Resolved Diagnosis technique was performed to simultaneously measure the specific IgE concentrations of 52 patients against Art v and its components (Art v 1), Phl p and its components (Phl p 1, Phl p 4, Phl p 5, Phl p 6, Phl p 7, Phl p 12), Bet v and its components (Bet v 1, Bet v 2), Amb a and its component (Amb a 1), and Amb p. Additionally, sIgE inhibition tests were conducted by Art v, Phl p, and Bet v extracts. Among Art v-positive patients, 64.6% showed positivity for Art v 1. In Phl p-positive patients, Phl p 12 had the highest positivity rate (75.0%). Among Bet v-positive patients, 75.6% exhibited positivity for Bet v 2, whereas for Amb a and Amb p-positive patients, 23.7% and 29.0% respectively showed positivity for Amb a 1. The sIgE inhibition assays results revealed that Art v extract had inhibition rates greater than 73.2% against Phl p and its component Phl p 12, as well as Bet v and its component Bet v 2. Simultaneously, Phl p extract showed inhibition rates of 80.70–89.87% against Phl p 12, Bet v and Bet v 2. Bet v extract showed inhibition rates ranging from 21.9 to 59.8% against Phl p and Bet v 2, with a better inhibition rate (76.80%) against Phl p 12. In conclusion, Art v 1 is identified as the principal component of Art v. The profilin proteins of Phl p and Bet v (Phl p 12 and Bet v 2), are implicated as potential cross-reactive elements contributing to polysensitization in patients with respiratory allergies in the Northwest region of China. This cross-reactivity leads to a shared sensitization mechanism among pollen allergens such as Art v, Phl p, and Bet v.

Abstract Background Enterococcus faecium and Staphylococcus aureus are the Gram-positive pathogens of the ESKAPE group, known to represent a great threat to human health due to their high virulence and multiple resistances to antibiotics. Combined, enterococci and S. aureus account for 26% of healthcare-associated infections and are the most common organisms responsible for blood stream infections. We previously showed that the peptidyl-prolyl cis/trans isomerase (PPIase) PpiC of E. faecium elicits the production of specific, opsonic, and protective antibodies that are effective against several strains of E. faecium and E. faecalis. Due to the ubiquitous characteristics of PPIases and their essential function within Gram-positive cells, we hypothesized a potential cross-reactive effect of anti-PpiC antibodies. Results Opsonophagocytic assays combined with bioinformatics led to the identification of the foldase protein PrsA as a new potential vaccine antigen in S. aureus. We show that PrsA is a stable dimeric protein able to elicit opsonic antibodies against the S. aureus strain MW2, as well as cross-binding and cross-opsonic in several S. aureus, E. faecium and E. faecalis strains. Conclusions Given the multiple antibiotic resistances S. aureus and enterococci present, finding preventive strategies is essential to fight those two nosocomial pathogens. The study shows the potential of PrsA as an antigen to use in vaccine formulation against the two dangerous Gram-positive ESKAPE bacteria. Our findings support the idea that PPIases should be further investigated as vaccine targets in the frame of pan-vaccinomics strategy.

Background: COVID-19 began in December 2019, rapidly spreading worldwide. China implemented a dynamic zero-COVID strategy and strict control measures after the outbreak. However, Guangzhou city ended closed-off management by the end of November 2022, leading to exposure to SARS-CoV-2. Despite most hospitalized patients being infected or co-infected with SARS-CoV-2, some remained uninfected. We report two cases of bacterial pneumonia with elevated globulin levels not infected with SARS-CoV-2, aiming to identify protection factors of SARS-CoV-2 infection and provide a scientific basis for SARS-CoV-2 prevention. Case presentation: Case 1, a 92-year-old male, admitted on October 21, 2022, developed worsening cough and sputum after aspiration, diagnosed with bacterial pneumonia with Pseudomonas aeruginosa, Escherichia coli (CRE) and carbapenem-resistant Acinetobacter baumannii (CRAB) infections. He was treated with imipenem anti-infective therapy and mechanical ventilation, then switched to a combination of meropenem, voriconazole and amikacin antiinfective therapy due to recurrent infections and septic shock, and died of sepsis on 8 January 2023. Case 2 is an 82-year-old male admitted on 30 September 2022, with recurrent cough, sputum, and shortness of breath, diagnosed with bacterial pneumonia with carbapenem-resistant Klebsiella pneumoniae (CRKP) and Mycobacterium pneumoniae infections. He was treated with ventilator-assisted ventilation, meropenem, amikacin, tigecycline and mucomycin nebulization and discharged with improvement on 26 October. He was readmitted on 21 November 2022 and diagnosed with bacterial pneumonia. He was treated with cefoperazone sulbactam, amikacin, meropenem and fluconazole and discharged on 31 December. Neither patient was infected with SARS-CoV-2 during hospitalization. Notably, their globulin levels were elevated before SARS-CoV-2 exposure, gradually decreasing afterward. Conclusions: Patients with bacterial pneumonia with high globulin levels likely have large amounts of immunoglobulin, and that immunoglobulin crossreactivity causes this protein to be involved in clearing SARS-CoV-2 and preventing infection. Therefore, bacterial pneumonia patients with high globulin levels included in this study were not infected with SARS-CoV-2. After exposure to SARS-CoV-2, the amount of globulin in the patient's body was reduced because it was used to clear SARS-CoV-2. The results of this study are expected to provide a theoretical basis for the study of the mechanism of prevention and treatment of SARS-CoV-2 infection. [ABSTRACT FROM AUTHOR]

The COVID-19 pandemic caused by the rapid spread of the novel coronavirus SARS-CoV-2, has promoted an interest in studying the T-cell immune response. It was found that the polyclonal and cross-reactive T-cell response against seasonal coronaviruses and other SARS-CoV-2 strains reduced disease severity. We investigated the immunodominant T-cell epitope SPRWYFYYYL from the nucleocapsid protein of SARS-CoV-2. The immune response to this epitope is characterized by the formation of highly homologous (convergent) receptors that have been found in the T-cell receptor (TCR) repertoires of different individuals. This epitope belongs to a group of highly conserved peptides that are rarely mutated in novel SARS-CoV-2 strains and are homologous to the epitopes of seasonal coronaviruses. It has been suggested that the cross-reactive response to homologous peptides contributes to the reduction of COVID-19 severity. However, some investigators have questioned this hypothesis, suggesting that the low affinity of the cross-reactive receptors reduces the strength of the immune response. The aim of this study was to evaluate the effect of amino acid substitutions in the SPR epitope on its binding affinity to specific TCRs. For this, we performed antigen-dependent cellular expansions were performed using samples from four COVID-19-transfected donors and sequenced their TCR repertoires. The resulting SPR-specific repertoire of β-chains in TCRs had a greater sequence diversity than the repertoire of α-chains. However, the TCR repertoires of all four donors contained public receptors, three of which were cloned and used to generate the Jurkat E6-1 TPR cell line. Only one of these receptors was activated by the SPR peptide and recognized with the same affinity by its mutant homologue LPRWYFYYY from seasonal coronaviruses. This indicates that the presence of the mutation did not affect the strength of the immune response, which may explain why the cross-reactive response to the SPR epitope is so frequent and contributes positively to COVID-19 infection. [ABSTRACT FROM AUTHOR]

Legumes are an inexpensive and essential protein source worldwide. The most consumed legumes include peanuts, soybeans, lentils, lupines, peas, common bean and chickpeas. In addition, the food industry is growing interested in expanding the use of legumes to partially replace or substitute cereals. Legumes were described to cause IgE-mediated allergies, and their growing use may also increase the incidence of allergy. The epidemiology of legume allergy varies by region; peanuts and soybeans are the legumes most involved in food allergies in Western countries, whereas lentils, peas, and chickpeas are reported as culprit allergens mainly in the Mediterranean area and India. This review, edited by the Italian Society of Pediatric Allergology and Immunology, summarizes the scientific literature on legume allergy in children and proposes a diagnostic workup and therapeutic approach. [ABSTRACT FROM AUTHOR]

Recent mumps outbreaks have been observed in vaccinated young adults due to the mumps virus (MuV) of genotype G, whereas the current vaccine is a mixture of two genotype A strains. These outbreaks could be attributed to waning vaccine immunity or the antigenic differences between the HN and F glycoproteins in the vaccine and circulating MuV. These glycoproteins are essential targets for the immune system, and antigenic variations may reduce the recognition of mumps antibodies, rendering the population susceptible to the MuV. We established stable cell lines expressing the MuV glycoproteins to study cross-reactivity between genotype A and genotype G. Cross-reactivity between the genotypes was evaluated via immunofluorescence using patient sera from vaccinated individuals, infected individuals, and vaccinated individuals infected with genotype G. Titer ratios showed that the vaccinated individuals exhibited a titer 3.68 times higher for the HN protein and 2.3 times higher for the F protein when comparing genotype A with genotype G. In contrast, the infected individuals showed a lower titer for genotype A compared with genotype G, at 0.43 and 0.33 for the HN and F proteins, respectively. No difference in titer ratio was observed for individuals vaccinated and subsequently infected with mumps. These findings suggest that antigenic variations between the two genotypes may potentially result in immune escape of the circulating strain, resulting in individuals susceptible to the MuV. [ABSTRACT FROM AUTHOR]

Lipid Transfer Protein (LTP) syndrome refers to a complex clinical pattern in which affected individuals have allergies to multiple plant foods due to cross-reactions between botanically unrelated pollens and plant foods that contain LTPs. Pollens and plants containing LTPs include mugwort, plane and olive pollens, and peach, apricot and walnut respectively. Published reports of LTP syndrome are few in the paediatric population in non-Mediterranean regions. [ABSTRACT FROM AUTHOR]

Background: Cross-reactivity between nonspecific lipid transfer proteins could cause anaphylaxis, further influencing food avoidance and nutrient deficiencies. The one affecting olive pollen (Ole e 7) and peach (Pru p 3) may underlie a variety of pollen-food syndromes, though a deep molecular analysis is necessary. Methods: Three Ole e 7-monosensitised patients (MON_OLE), three Pru p 3-monosensitised patients (MON_PRU) and three bisensitised patients (BI) were selected. For epitope mapping, both digested proteins were incubated with patient sera, and the captured IgE-bound peptides were characterised by LC-MS. Results: The analysis revealed two Ole e 7 epitopes and the three Pru p 3 epitopes previously described. Interestingly, the "KSALALVGNKV" Ole e 7 peptide was recognised by MON_OLE, BI and MON_PRU patients. Conversely, all patients recognised the "ISASTNCATVK" Pru p 3 peptide. Although complete sequence alignment between both proteins revealed 32.6% identity, local alignment considering seven residue fragments showed 50 and 57% identity when comparing "ISASTNCATVK" with Ole e 7 and "KSALALVGNKV" with Pru p 3. Conclusions: This study mapped sIgE-Ole e 7-binding epitopes, paving the way for more precise diagnostic tools. Assuming non-significant sequence similarity, structural homology and shared key residues may underlie the potential cross-reactivity between Ole e 7 and Pru p 3 nsLTPs. [ABSTRACT FROM AUTHOR]

Pneumococcal vaccines are a cornerstone for the prevention of pneumococcal diseases, reducing morbidity and mortality in children and adults worldwide. Pneumococcal vaccine composition is based on the polysaccharide capsule of Streptococcus pneumoniae, which is one of the most important identified contributors to the pathogen's virulence. Similarities in the structural composition of polysaccharides included in licensed pneumococcal vaccines may result in cross-reactivity of immune response against closely related serotypes, including serotypes not included in the vaccine. Therefore, it is important to understand whether cross-reactive antibodies offer clinical protection against pneumococcal disease. This review explores available evidence of cross-reactivity and cross-protection associated with pneumococcal vaccines, the challenges associated with the assessment of cross-reactivity and cross-protection, and implications for vaccine design and development. [ABSTRACT FROM AUTHOR]

Purpose of Review: Allergy diagnostics and immunotherapeutics in Asia heavily rely on imported products from Western countries, raising concerns about the accuracy and efficacy of these products for the management of Asian allergy patients. Recent Findings : Recent advancements in allergen research have led to the identification and characterization of novel allergens from indigenous Korean species. While some allergens share homology with well-known allergens, others lack counterparts in imported allergen extracts. Summary: Classifying regional allergens in Asia into three categories based on their cross-reactivity with imported allergens offers valuable insights. Highly cross-reactive allergens, such as oak allergens Que m 1 from Quercus mongolica and Que ac 1 from Q. acutissima, can be effectively substituted with the imported allergens. Allergens with partial cross-reactivity, like the Asian needle ant allergen Pac c 3 (Antigen 5), permit limited diagnostic value by the currently available products. Unique allergens, including the Japanese hop allergen Hum j 6 (pectin methylesterase inhibitor) and the silkworm pupa allergen Bomb m 4 (30 kDa hemolymph lipoprotein) lack alternatives in the available product list. Greater attention is needed, particularly for species listed as ecologically invasive in Western regions. Additionally, allergens from domestic fruits and vegetables causing pollen food allergy syndrome require characterization for the development of improved diagnostics. [ABSTRACT FROM AUTHOR]

Objective: This study aims to assess the outcome of challenging documented moderate, severe, or unknown beta‐lactam allergies with full dose administration of a beta‐lactam antibiotic in emergency department (ED) patients admitted for acute bacterial infection. Methods: A single‐center, retrospective, descriptive study of adult patients challenged with a full dose of beta‐lactam in the ED from January 2021 to December 2022 was conducted. Included patients had at least one documented moderate, severe, or unknown beta‐lactam allergy in the electronic medical record (EMR) without documentation of prior tolerance. Patient demographics, prior beta‐lactam antibiotic reaction, beta‐lactam administered in the ED, inpatient beta‐lactam continuation, adverse drug reactions, and updates to allergy profiles were collected. Descriptive statistics for data analysis were performed using SPSS Version 22. Results: Of the 184 ED encounters with full‐dose beta‐lactam challenges, five (2.7%) patients with documented moderate, severe, or unknown beta‐lactam allergies experienced an allergic reaction after the challenge; one (0.5%) patient had an allergic reaction in the ED, and the remaining four (2.2%) occurred after admission. No anaphylactic reactions occurred. All allergic reactions were limited to mild rash or itching. Most patients (98.9%) were challenged with a cephalosporin. A beta‐lactam was continued in 86.4% of cases, and the allergy profile was updated for future utilization in 73.4% of patients. Conclusions: This study suggests that full‐dose challenge of moderate, severe, or unknown beta‐lactam allergies can be safely accomplished in the ED. This approach avoids unnecessary penicillin allergy skin testing and reduces utilization of suboptimal alternative antibiotic regimens. [ABSTRACT FROM AUTHOR]

Tick‐borne encephalitis (TBE) virus is the most prevalent tick‐transmitted orthoflavivirus in Europe. Due to the nonspecific nature of its symptoms, TBE is primarily diagnosed by ELISA‐based detection of specific antibodies in the patient serum. However, cross‐reactivity between orthoflaviviruses complicates the diagnosis. Specificity issues may be mitigated by serum neutralization assays (SNT), although the handling of clinically relevant orthoflaviviruses requires biosafety level (BSL) 3 conditions and they have highly divergent viral kinetics and cell tropisms. In the present study, we established a reporter virus particle (RVP)‐based SNT in which the infectivity is measured by luminescence and that can be performed under BSL‐2 conditions. The RVP‐based SNT for TBEV exhibited a highly significant correlation with the traditional virus‐based SNT (R2 = 0.8637, p < 0.0001). The RVP‐based assay demonstrated a sensitivity of 92.3% (95% CI: 79.7%–97.4%) and specificity of 100% (95% CI: 81.6%–100%). We also tested the cross‐reactivity of serum samples in RVP‐based assays against other orthoflaviviruses (yellow fever virus, dengue virus type 2, Zika virus, West Nile virus and Japanese encephalitis virus). Interestingly, all serum samples which had tested TBEV‐positive by ELISA but negative by RVP‐based SNT were reactive for antibodies against other orthoflaviviruses. Thus, the RVP‐based seroneutralization assay provides an added value in clinical diagnostics as well as in epidemiological studies. [ABSTRACT FROM AUTHOR]

Background: Enterococcus faecium and Staphylococcus aureus are the Gram-positive pathogens of the ESKAPE group, known to represent a great threat to human health due to their high virulence and multiple resistances to antibiotics. Combined, enterococci and S. aureus account for 26% of healthcare-associated infections and are the most common organisms responsible for blood stream infections. We previously showed that the peptidyl-prolyl cis/trans isomerase (PPIase) PpiC of E. faecium elicits the production of specific, opsonic, and protective antibodies that are effective against several strains of E. faecium and E. faecalis. Due to the ubiquitous characteristics of PPIases and their essential function within Gram-positive cells, we hypothesized a potential cross-reactive effect of anti-PpiC antibodies. Results: Opsonophagocytic assays combined with bioinformatics led to the identification of the foldase protein PrsA as a new potential vaccine antigen in S. aureus. We show that PrsA is a stable dimeric protein able to elicit opsonic antibodies against the S. aureus strain MW2, as well as cross-binding and cross-opsonic in several S. aureus, E. faecium and E. faecalis strains. Conclusions: Given the multiple antibiotic resistances S. aureus and enterococci present, finding preventive strategies is essential to fight those two nosocomial pathogens. The study shows the potential of PrsA as an antigen to use in vaccine formulation against the two dangerous Gram-positive ESKAPE bacteria. Our findings support the idea that PPIases should be further investigated as vaccine targets in the frame of pan-vaccinomics strategy. [ABSTRACT FROM AUTHOR]

Thirty years have passed since the concept of latex-fruit syndrome (LFS) was first introduced. Since then, this phenomenon, characterized by cross-reactivity between natural latex rubber allergens and certain fruit allergens, has been extensively studied. This literature review sought to determine the prevalence of LFS in latex-allergic patients, identify the most common cross-reactions with fruit allergens in individuals with LFS, and understand the clinical manifestations of this syndrome. An extensive literature search was carried out using PubMed and Scopus databases, while applying the Preferred Reporting Items for Systematic Reviews and Meta-Analysis methodology. The analysis of original studies revealed a wide variation in LFS prevalence (4–88%) influenced by diverse diagnostic tools, different geographical regions, and the size of study populations. Our findings indicate that the most prevalent allergenic fruits in patients with LFS are banana, avocado, kiwifruit, and papaya. After evaluating the symptoms of the fruit hypersensitivity of patients with LFS, the clinical manifestation of hypersensitivity constituted 73% of systemic allergy symptoms and only 27% of reported symptoms described the localized allergy. Furthermore, the clinical picture of latex-fruit syndrome is illustrated through two cases, one typical and one with an unusual presentation. Their clinical features were assessed and contrasted utilizing different anaphylaxis severity grading criteria. To properly manage LFS, it is essential to establish standardized diagnostic criteria and severity grading systems, as these are crucial for accurate diagnosis and effective treatment. [ABSTRACT FROM AUTHOR]

Dengue (DENV) and Chikungunya (CHIKV) viruses can be transmitted simultaneously by Aedes mosquitoes, and there may be co-infections in humans. However, how the adaptive immune response is modified in the host has yet to be known entirely. In this study, we analyzed the cross-reactivity and neutralizing activity of IgG antibodies against DENV and CHIKV in sera of patients from the Mexican Institute of Social Security in Veracruz, Mexico, collected in 2013 and 2015 and using IgG antibodies of BALB/c mice inoculated with DENV and/or CHIKV. Mice first inoculated with DENV and then with CHIKV produced IgG antibodies that neutralized both viruses. Mice were inoculated with CHIKV, and then with DENV; they had IgG antibodies with more significant anti-CHIKV IgG antibody neutralizing activity. However, the inoculation only with CHIKV resulted in better neutralization of DENV2. In sera obtained from patients in 2013, significant cross-reactivity and low anti-CHIKV IgG antibody neutralizing activity were observed. In CHIKV-positive 2015 sera, the anti-DENV IgG antibody neutralizing activity was high. These results suggest that CHIKV stimulates DENV2-induced memory responses and vice versa. Furthermore, cross-reactivity between the two viruses generated neutralizing antibodies, but exchanging CHIKV for DENV2 generated a better anti-CHIKV neutralizing response. [ABSTRACT FROM AUTHOR]

Infectious diseases are commonly implicated as potential initiators of autoimmune diseases (ADs) and represent the most commonly known factor in the development of autoimmunity in susceptible individuals. Epidemiological data and animal studies on multiple ADs suggest that molecular mimicry is one of the likely mechanisms for the loss of peripheral tolerance and the development of clinical disease. Besides molecular mimicry, other mechanisms such as defects in central tolerance, nonspecific bystander activation, epitope-determinant spreading, and/or constant antigenic stimuli, may also contribute for breach of tolerance and to the development of ADs. Linear peptide homology is not the only mechanism by which molecular mimicry is established. Peptide modeling (i.e., 3D structure), molecular docking analyses, and affinity estimation for HLAs are emerging as critical strategies when studying the links of molecular mimicry in the development of autoimmunity. In the current pandemic, several reports have confirmed an influence of SARS-CoV-2 on subsequent autoimmunity. Bioinformatic and experimental evidence support the potential role of molecular mimicry. Peptide dimensional analysis requires more research and will be increasingly important for designing and distributing vaccines and better understanding the role of environmental factors related to autoimmunity.

ABSTRACT The FilmArray Pneumonia Panel (FilmArrayPN; bioMérieux) was tagged for potential false-positive seasonal coronavirus results, possibly caused by non-specific amplification or cross-reactivity with human genomic DNA. This study was conducted to determine the real-world false-positive rate of seasonal coronavirus infections using FilmArrayPN at a tertiary care hospital. We retrospectively reviewed FilmArrayPN results to detect lower respiratory tract pathogens from July 2023 to April 2024. Allplex Respiratory Panel (AllplexRP; Seegene) and/or FilmArray Respiratory Panel (FilmArrayRP) results for the same patient were considered as reference results. In the last month of the study period, all positive samples in FilmArrayPN were confirmed by AllplexRP and pan-coronavirus targeted PCR (pan-CoV PCR). A total of 2,120 specimens, including 1,847 expectorated sputum and 273 bronchial aspirate (BA) samples, were tested by FilmArrayPN. Of these, 161 (8.7%) sputum and 7 (2.6%) BA specimens from 152 patients were positive for coronavirus targets in FilmArrayPN. Of the 122 cases also tested by AllplexRP and/or FilmArrayRP, 106 specimens (86.9%) were coronavirus-negative, except for 9 OC43-positive, 6 229E-positive, and 1 NL63-positive. Notably, 83.1% of the 106 cases that tested negative in the other tests had melting temperatures above 83℃. A total of 61 specimens that tested positive for coronavirus in FilmArrayPN but negative in AllplexRP were confirmed to be negative in pan-CoV PCR. The coronavirus positivity of 7.8% in the FilmArrayPN resulted in 5% of samples being potentially misreported as false-positives. This report thus highlights the need for continuous monitoring of melting temperatures to avoid potential false-positives.IMPORTANCEThe FilmArray Pneumonia Panel (FilmArrayPN; bioMérieux) was tagged for potential false-positive seasonal coronavirus results, possibly caused by non-specific amplification or cross-reactivity with human genomic DNA. FilmArrayPN results were retrospectively reviewed from July 2023 to May 2024. Of 2,120 tested specimens, 168 specimens from 152 patients were positive for coronavirus targets in FilmArrayPN. Of the 122 cases also tested by Allplex Respiratory Panel and/or FilmArray Respiratory Panel, 106 specimens (86.9%) were coronavirus-negative. Notably, 83.1% of the 106 cases that tested negative in the other tests had melting temperatures above 83℃. A total of 61 specimens that tested positive for coronavirus in FilmArrayPN but negative in Allplex were confirmed to be negative in pan-coronavirus targeted PCR. The coronavirus positivity of 7.8% in the FilmArrayPN resulted in 5% of samples being potentially misreported as false-positives. This report highlights the need for continuous monitoring of melting temperatures to avoid potential false-positives.

Background: The diagnosis of the culprit allergen depends on exposure, symptoms at exposure, and the presence of specific IgE (sIgE). Pet allergens are sticky and can sensitize individuals without adoption history. Exclusive dog owners frequently exhibit both dog (e5) and cat dander sIgE (e1). We assessed whether the measurement of Fel d 1 sIgE (e94) can discriminate true cat sensitization from false positivity by cross-reactivity in the exclusive dog owners. Methods: Thirty-one patients with respiratory allergies who exclusively owned dogs were enrolled for this study. e5, e1, and e94 were measured with ImmunoCAP. ELISA inhibition was performed to assess cross-reactivity. Results: About 81% of patients (25/31) were both e5 and e1 positive, and 8 were also positive for e94. In the e94 positive, cat dander exhibited higher maximum inhibition of cat sIgE (94% vs 88%) and demonstrated lower IC50 (6.5 vs 737.9 BAU/mL) compared to dog dander. Conversely, in the e94 negative, dog dander demonstrated higher maximum inhibition of cat dander sIgE (71.9% vs 56.2%) and lower IC50 (172 vs 1947 BAU/mL) compared to cat dander. In the e94 positive, dog dander exhibited higher maximal inhibition for dog sIgE (91.5 vs 76.1%) and lower IC50 (10.6 vs 1679 BAU/mL) compared to cat dander, whereas in the e94 negative, the IC50 for cat dander could not be determined. Conclusions: Genuine co-sensitization to cats is notable even in individuals who exclusively own dogs. Positive e94 results could discriminate authentic cat sensitization from false positivity by cross-reactivity in these patients, underscoring the importance of comprehensive allergy assessment.

Specificity profiling is a requirement for monoclonal antibodies (mAbs) and antibody-directed biotherapeutics such as CAR-T cells prior to initiating human trials. However, traditional approaches to assess the specificity of mAbs, primarily tissue cross-reactivity studies, have been unreliable, leading to off-target binding going undetected. Here, we review the emergence of cell-based protein arrays as an alternative and improved assessment of mAb specificity. Cell-based protein arrays assess binding across the full human membrane proteome, ~6,000 membrane proteins each individually expressed in their native structural configuration within live or unfixed cells. Our own profiling indicates a surprisingly high off-target rate across the industry, with 33% of lead candidates displaying off-target binding. Moreover, about 20% of therapeutic mAbs in clinical development and currently on the market display off-target binding. Case studies and off-target rates at different phases of biotherapeutic drug approval suggest that off-target binding is likely a major cause of adverse events and drug attrition.

Monoclonal neutralizing antibodies (mAbs) are considered an important prophylactic against SARS-CoV-2 infection in at-risk populations and a strategy to counteract future sarbecovirus-induced disease. However, most mAbs isolated so far neutralize only a few sarbecovirus strains. Therefore, there is a growing interest in bispecific antibodies (bsAbs) which can simultaneously target different spike epitopes and thereby increase neutralizing breadth and prevent viral escape. Here, we generate and characterize a panel of 30 novel broadly reactive bsAbs using an efficient controlled Fab-arm exchange protocol. We specifically combine some of the broadest mAbs described so far, which target conserved epitopes on the receptor binding domain (RBD). Several bsAbs show superior cross-binding and neutralization compared to the parental mAbs and cocktails against sarbecoviruses from diverse clades, including recent SARS-CoV-2 variants. BsAbs which include mAb COVA2–02 are among the most potent and broad combinations. As a result, we study the unknown epitope of COVA2–02 and show that this mAb targets a distinct conserved region at the base of the RBD, which could be of interest when designing next-generation bsAb constructs to contribute to a better pandemic preparedness.

The immune system is endowed with the capacity to distinguish between self and non-self, so-called immune tolerance or “consciousness of the immune system.” This type of awareness is designed to achieve host protection by eliminating cells expressing a wide range of non-self antigens including microbial-derived peptides. Such a successful immune response is associated with the secretion of a whole spectrum of soluble mediators, e.g., cytokines and chemokines, which not only contribute to the clearance of infected host cells but also activate T cells that are not specific to the original cognate antigen. This kind of non-specific T-cell activation is called “bystander activation.” Although it is well-established that this phenomenon is cytokine-dependent, there is evidence in the literature showing the involvement of peptide/MHC recognition depending on the type of T-cell subset (naive vs. memory). Here, we will summarize our current understanding of the mechanism(s) of bystander T-cell activation as well as its biological significance in a wide range of diseases including microbial infections, cancer, auto- and alloimmunity, and chronic inflammatory diseases such as atherosclerosis.

Background: The simultaneous occurrence of the COVID-19 pandemic and a dengue outbreak has posed significant challenges for governments and medical personnel in dengue-endemic countries like Indonesia. Several studies in dengue-endemic countries have reported cases of misdiagnosis between COVID-19 and dengue. Therefore, it is crucial to evaluate the potential cross-reactivity between SARS-CoV-2 antibodies and dengue. Methods: This study aimed to confirm the serological cross-reaction between dengue virus and SARS-CoV-2 in Surabaya, East Java, which is a highly dengue-endemic city in Indonesia. In total, 238 serum samples with confirmed dengue that were collected before the emergence of COVID-19 were tested to detect the presence of reacting IgG and IgM antibodies (Abs) against SARS-CoV-2 via a rapid detection test (RDT) and enzyme-linked immunosorbent assay (ELISA). Samples from patients with dengue infection collected during the pandemic, from healthy volunteers predating the pandemic, and from patients with COVID-19 were used for comparison. Results and conclusion: Few (6.7 %) of the pre-COVID-19 dengue Ab-positive serum samples showed reactive on SARS-CoV-2 in the RDT, with significantly lower IgG and IgM levels detected in ELISA compared with the dengue samples collected during the pandemic and the COVID-19 samples (P

Accurate detection is a prerequisite for effective prevention and control of Mycoplasma synoviae infection. ELISA is the most popular method for the clinical detection of M. synoviae because of its convenience, low cost, and high detection rate. However, the cross-reactivity of commercially available ELISA kits with other avian pathogen-positive sera needs to be addressed. The aim of this study was to establish an ELISA method with high specificity for the detection of anti-M. synoviae antibodies in chicken serum to evaluate the M. synoviae infection status on poultry farms. The recombinant MS087 (rMS087) protein was expressed in Escherichia coli BL21 (DE3) and purified by Ni2+ affinity chromatography. An antibody against rMS087 was generated by immunizing BALB/c mice. Bioinformatic analysis revealed that MS087 was conserved among M. synoviae strains. Western blotting and indirect immunofluorescence results indicated that MS087 was not only localized in the cytoplasm and on the membrane but also secreted by the organism. For the established ELISA method based on rMS087, the optimal antigen concentration, blocking buffer, blocking duration, serum dilution, serum incubation duration, secondary antibody dilution, secondary antibody incubation duration and colorimetric reaction duration were 2 μg/mL, 1% BSA, 3 h, 1:500, 1.5 h, 1:20,000, 2 h and 5 min, respectively. Validation of the rMS087-based ELISA revealed a cut-off value of 0.5. The coefficients of variation of both the intra-batch and inter-batch methods were less than 9%. The assay was able to differentiate positive serum against M. synoviae from antisera against nine other avian pathogens and was able to recognize M. synoviae-positive sera at a dilution of 1:1,000. Compared with the commercial ELISA method, the rMS087-based ELISA has the potential to recognize more positive sera against M. synoviae. Collectively, the rMS087-based ELISA is a reproducible, specific, and sensitive serological method for detecting antibodies against M. synoviae in chicken serum and has robust potential for large-scale serological epidemiology of M. synoviae infection on poultry farms.

Abstract Background Remimazolam is a recently developed, ultrashort-acting benzodiazepine that is used as a general anesthetic. Some cases of remimazolam anaphylaxis have been reported, but its characteristics are not fully understood. We present an interesting case report and review of the literature to better understand remimazolam anaphylaxis. Case presentation A 75-year-old man scheduled for robot-assisted gastrectomy was administered remimazolam for the induction of general anesthesia. After intubation, low end-expiratory CO2, high airway pressure and concurrent circulatory collapse were observed. Bronchoscopy revealed marked tracheal and bronchial edema, which we diagnosed as anaphylaxis. The patient suffered cardiac arrest after bronchoscopy but recovered immediately with intravenous adrenaline administration and chest compressions. We performed skin prick tests for the drugs used during induction except for remimazolam, considering the high risk of systemic adverse reactions to remimazolam. We diagnosed remimazolam anaphylaxis because the skin prick test results for the other drugs used during anesthesia were negative, and these drugs could have been used without allergic reactions during the subsequent surgery. Furthermore, this patient had experienced severe anaphylactic-like reactions when he underwent cardiac surgery a year earlier, in which midazolam had been used, but it was not thought to be the allergen at that time. Based on these findings, cross-reactivity to remimazolam and midazolam was suspected. However, the patient had previously received another benzodiazepine, brotizolam, to which he was not allergic, suggesting that cross-reactivity of remimazolam may vary among benzodiazepines. In this article, we reviewed the 11 cases of remimazolam anaphylaxis that have been described in the literature. Conclusions Remimazolam is an ultrashort-acting sedative; however, it can cause life-threatening anaphylaxis. In addition, its cross-reactivity with other benzodiazepines is not fully understood. To increase the safety of this drug, further research and more experience in its use are needed.

Background: After the eradication of smallpox, there have been no specific public health measures for any Orthopoxviruses (OPXVs). Therefore, it is necessary to countermeasure OPXV infections after Mpox (formerly monkeypox) occurrences, such as the latest global outbreak in 2022–2023. This study aimed to provide crucial insights for the development of effective public health policy making against mpox in populations residing in regions where the virus is not prevalent. Methods: This study used enzyme-linked immunosorbent assays (ELISA) to examine smallpox and mpox antibodies in Koreans with three different age groups. We analyzed 56 sera obtained from a tertiary care hospital in South Korea between September 2022 and April 2023. Plasma levels of antibodies against the viral proteins of smallpox (variola cytokine response-modifying protein B) and MPXV (A29) were measured using enzyme-linked immunosorbent assays. Results: Plasma samples from participants in their early 40 s and older exhibited higher reactivity to viral antigens than those from younger participants. Furthermore, there was a strong positive correlation in antibody positivity for the two different viruses across the sera. Conclusions: The presence of low antibody levels in participants ˂40 years may hinder their ability to defend against OPXV. Therefore, it is imperative to implement effective public health measures to mitigate the transmission of OPXV within the community. These findings serve as fundamental information for devising strategies to combat mpox efficiently, particularly in regions where the virus is not prevalent.

Abstract Background High levels of neutrophil extracellular trap (NET) formation or NETosis and autoantibodies are related to poor prognosis and disease severity of COVID-19 patients. Human angiotensin-converting enzyme 2 (ACE2) cross-reactive anti-severe acute respiratory syndrome coronavirus 2 spike protein receptor-binding domain (SARS-CoV-2 RBD) antibodies (CR Abs) have been reported as one of the sources of anti-ACE2 autoantibodies. However, the pathological implications of CR Abs in NET formation remain unknown. Methods In this study, we first assessed the presence of CR Abs in the sera of COVID-19 patients with different severity by serological analysis. Sera and purified IgG from CR Abs positive COVID-19 patients as well as a mouse monoclonal Ab (mAb 127) that can recognize both ACE2 and the RBD were tested for their influence on NETosis and the possible mechanisms involved were studied. Results An association between CR Abs levels and the severity of COVID-19 in 120 patients was found. The CR Abs-positive sera and IgG from severe COVID-19 patients and mAb 127 significantly activated human leukocytes and triggered NETosis, in the presence of RBD. This NETosis, triggered by the coexistence of CR Abs and RBD, activated thrombus-related cells but was abolished when the interaction between CR Abs and ACE2 or Fc receptors was disrupted. We also revealed that CR Abs-induced NETosis was suppressed in the presence of recombinant ACE2 or the Src family kinase inhibitor, dasatinib. Furthermore, we found that COVID-19 vaccination not only reduced COVID-19 severity but also prevented the production of CR Abs after SARS-CoV-2 infection. Conclusions Our findings provide possible pathogenic effects of CR Abs in exacerbating COVID-19 by enhancing NETosis, highlighting ACE2 and dasatinib as potential treatments, and supporting the benefit of vaccination in reducing disease severity and CR Abs production in COVID-19 patients.

The Nipah virus (NiV), a highly deadly bat-borne paramyxovirus, poses a substantial threat due to recurrent outbreaks in specific regions, causing severe respiratory and neurological diseases with high morbidity. Two distinct strains, NiV-Malaysia (NiV-M) and NiV-Bangladesh (NiV-B), contribute to outbreaks in different geographical areas. Currently, there are no commercially licensed vaccines or drugs available for prevention or treatment. In response to this urgent need for protection against NiV and related henipaviruses infections, we developed a novel homotypic virus-like nanoparticle (VLP) vaccine co-displaying NiV attachment glycoproteins (G) from both strains, utilizing the self-assembling properties of ferritin protein. In comparison to the NiV G subunit vaccine, our nanoparticle vaccine elicited significantly higher levels of neutralizing antibodies and provided complete protection against a lethal challenge with NiV infection in Syrian hamsters. Remarkably, the nanoparticle vaccine stimulated the production of antibodies that exhibited superior cross-reactivity to homologous or heterologous henipavirus. These findings underscore the potential utility of ferritin-based nanoparticle vaccines in providing both broad-spectrum and long-term protection against NiV and emerging zoonotic henipaviruses challenges. [ABSTRACT FROM AUTHOR]

Background: Remimazolam is a recently developed, ultrashort-acting benzodiazepine that is used as a general anesthetic. Some cases of remimazolam anaphylaxis have been reported, but its characteristics are not fully understood. We present an interesting case report and review of the literature to better understand remimazolam anaphylaxis. Case presentation: A 75-year-old man scheduled for robot-assisted gastrectomy was administered remimazolam for the induction of general anesthesia. After intubation, low end-expiratory CO2, high airway pressure and concurrent circulatory collapse were observed. Bronchoscopy revealed marked tracheal and bronchial edema, which we diagnosed as anaphylaxis. The patient suffered cardiac arrest after bronchoscopy but recovered immediately with intravenous adrenaline administration and chest compressions. We performed skin prick tests for the drugs used during induction except for remimazolam, considering the high risk of systemic adverse reactions to remimazolam. We diagnosed remimazolam anaphylaxis because the skin prick test results for the other drugs used during anesthesia were negative, and these drugs could have been used without allergic reactions during the subsequent surgery. Furthermore, this patient had experienced severe anaphylactic-like reactions when he underwent cardiac surgery a year earlier, in which midazolam had been used, but it was not thought to be the allergen at that time. Based on these findings, cross-reactivity to remimazolam and midazolam was suspected. However, the patient had previously received another benzodiazepine, brotizolam, to which he was not allergic, suggesting that cross-reactivity of remimazolam may vary among benzodiazepines. In this article, we reviewed the 11 cases of remimazolam anaphylaxis that have been described in the literature. Conclusions: Remimazolam is an ultrashort-acting sedative; however, it can cause life-threatening anaphylaxis. In addition, its cross-reactivity with other benzodiazepines is not fully understood. To increase the safety of this drug, further research and more experience in its use are needed. [ABSTRACT FROM AUTHOR]

Background: The Coronaviridae family comprises seven viruses known to infect humans, classified into alphacoronaviruses (HCoV-229E and HCoV-NL63) and betacoronaviruses (HCoV-OC43 and HCoV-HKU1), which are considered endemic. Additionally, it includes SARS-CoV (severe acute respiratory syndrome), MERS-CoV (Middle East respiratory syndrome), and the novel coronavirus SARSCoV- 2, responsible for COVID-19. SARS-CoV-2 induces severe respiratory complications, particularly in the elderly, immunocompromised individuals and those with underlying diseases. An essential question since the onset of the COVID-19 pandemic has been to determine whether prior exposure to seasonal coronaviruses influences immunity or protection against SARS-CoV-2. Methods: In this study, we investigated a cohort of 47 couples (N=94), where one partner tested positive for SARS-CoV-2 infection via real-time PCR while the other remained negative. Plasma samples, collected at least 30 days post-PCR reaction, were assessed using indirect ELISA and competition assays to measure specific antibodies against the receptor-binding domain (RBD) portion of the Spike (S) protein from SARS-CoV-2, HCoV-229E, HCoV-NL63, HCoV-OC43, and HCoV-HKU1. Results: IgG antibody levels against the four endemic coronavirus RBD proteins were similar between the PCR-positive and PCR-negative individuals, suggesting that IgG against endemic coronavirus RBD regions was not associated with protection from infection. Moreover, we found no significant IgG antibody crossreactivity between endemic coronaviruses and SARS-CoV-2 RBDs. Conclusions: Taken together, results suggest that anti-RBD antibodies induced by a previous infection with endemic HCoVs do not protect against acquisition of COVID-19 among exposed uninfected individuals. [ABSTRACT FROM AUTHOR]

Primary antibodies are one of the main tools used in molecular biology research. However, the often-occurring cross-reactivity of primary antibodies complicates accurate data analysis. Our results show that three commercial polyclonal antibodies raised against N-6 adenine-specific DNA methyltransferase 1 (N6AMT1) strongly cross-react with endogenous and recombinant mitosis-associated protein Aurora kinase A (AURKA). The cross-reactivity was verified through immunofluorescence, immunoblot, and immunoprecipitation assays combined with mass spectrometry. N6AMT1 and AURKA are evolutionarily conserved proteins that are vital for cellular processes. Both proteins share the motif ENNPEE, which is unique to only these two proteins. We suggest that N6AMT1 antibodies recognise this motif in N6AMT1 and AURKA proteins and exhibit an example of "specific" non-specificity. This serves as an example of the importance of controls and critical data interpretation in molecular biology research. [ABSTRACT FROM AUTHOR]

Food allergies mediated by specific IgE (sIgE) have a significant socioeconomic impact on society. Evaluating the IgE cross-reactivity between allergens from different allergen sources can enable the better management of these potentially life-threatening adverse reactions to food proteins and enhance food safety. A novel banana fruit allergen, S-adenosyl-L-homocysteine hydrolase (SAHH), has been recently identified and its recombinant homolog was heterologously overproduced in E. coli. In this study, we performed a search in the NCBI (National Center for Biotechnology Information) for SAHH homologs in ryegrass, latex, and kiwifruit, all of which are commonly associated with pollen-latex-fruit syndrome. In addition, Western immunoblot analysis was utilized to identify the cross-reactive IgE to banana SAHH in the sera of patients with a latex allergy, kiwifruit allergy, and ryegrass allergy. ClustalOmega analysis showed more than 92% amino acid sequence identity among the banana SAHH homologs in ryegrass, latex, and kiwifruit. In addition to five B-cell epitopes, in silico analysis predicted eleven T-cell epitopes in banana SAHH, seventeen in kiwifruit SAHH, twelve in ryegrass SAHH, and eight in latex SAHH, which were related to the seven-allele HLA reference set (HLA-DRB1*03:01, HLA-DRB1*07:01, HLA-DRB1*15:01, HLA-DRB3*01:01, HLA-DRB3*02:02, HLA-DRB4*01:01, HLA-DRB5*01:01). Four T-cell epitopes were identical in banana and kiwifruit SAHH (positions 328, 278, 142, 341), as well as banana and ryegrass SAHH (positions 278, 142, 96, and 341). All four SAHHs shared two T-cell epitopes (positions 278 and 341). In line with the high amino acid sequence identity and B-cell epitope homology among the analyzed proteins, the cross-reactive IgE to banana SAHH was detected in three of three latex-allergic patients, five of six ryegrass-allergic patients, and two of three kiwifruit-allergic patients. Although banana SAHH has only been studied in a small group of allergic individuals, it is a novel cross-reactive food allergen that should be considered when testing for pollen-latex-fruit syndrome. [ABSTRACT FROM AUTHOR]

After the eradication of smallpox, there have been no specific public health measures for any Orthopoxviruses (OPXVs). Therefore, it is necessary to countermeasure OPXV infections after Mpox (formerly monkeypox) occurrences, such as the latest global outbreak in 2022–2023. This study aimed to provide crucial insights for the development of effective public health policy making against mpox in populations residing in regions where the virus is not prevalent. This study used enzyme-linked immunosorbent assays (ELISA) to examine smallpox and mpox antibodies in Koreans with three different age groups. We analyzed 56 sera obtained from a tertiary care hospital in South Korea between September 2022 and April 2023. Plasma levels of antibodies against the viral proteins of smallpox (variola cytokine response-modifying protein B) and MPXV (A29) were measured using enzyme-linked immunosorbent assays. Plasma samples from participants in their early 40 s and older exhibited higher reactivity to viral antigens than those from younger participants. Furthermore, there was a strong positive correlation in antibody positivity for the two different viruses across the sera. The presence of low antibody levels in participants ˂40 years may hinder their ability to defend against OPXV. Therefore, it is imperative to implement effective public health measures to mitigate the transmission of OPXV within the community. These findings serve as fundamental information for devising strategies to combat mpox efficiently, particularly in regions where the virus is not prevalent. [ABSTRACT FROM AUTHOR]

Cannabis allergy is a relatively new phenomenon described in the 1970s. Its increased frequency has been observed over the last years due to the increasing therapeutic and recreational use of cannabis-based products. Sensitization possibly leading to allergy symptoms can occur not only through the smoking of cannabis, but also through ingestion, the inhalation of pollen, or direct contact. The severity of symptoms varies from benign pruritus to anaphylaxis. There is scant information available to support clinicians throughout the entire therapeutic process, starting from diagnosis and ending in treatment. In this review, we present six cases of patients in whom molecular in vitro testing revealed sensitization to cannabis extract and/or cannabis-derived nsLTP molecules (Can s 3). Based on these cases, we raise important questions regarding this topic. The article discusses current proposals and highlights the importance of further research not only on cannabis allergy but also on asymptomatic sensitization to cannabis allergens, which may be ascertained in some percentage of the population. [ABSTRACT FROM AUTHOR]