Assessment of In Vitro Digestive Behavior of Lactic-Acid-Bacteria Fermented Soy Proteins: A Study Comparing Colloidal Solutions and Curds.

This study investigated the effect of lactic-acid-bacteria fermentation on the microstructure and gastrointestinal digestibility of soy proteins using a digestomics approach. Fermented soy protein isolates (FSPIs) under varied fermentation-terminal pH demonstrated a colloidal solution (FSPI-7.0/6.0) or yogurt-like curd (FSPI-5.0/4.0) state. Cryo-electron microscopy figures demonstrated the loosely stacked layer of FSPI-7.0/6.0 samples, whereas a denser gel network was observed for FSPI-5.0/4.0 samples. Molecular interactions shifted from dominant ionic bonds to hydrophobic forces and disulfide bonds. The gastric/intestinal digestion demonstrated that the curd samples afforded a significantly low particle size and high-soluble protein and peptide contents in the medium and late digestive phases. A peptidomics study showed that the FSPI-6.0 digestate at early intestinal digestion had a high peptidome abundance, whereas FSPI curd digestates (FSPI-5.0/4.0) elicited a postponed but more extensive promotion during medium and late digestion. Glycinin G2/G4 and ß-conglycinin α/α' subunits were the major subunits promoted by FSPI-curds. The spatial structures of glycinin G2 and ß-conglycinin α subunits demonstrated variations located in seven regions. Glycinin G2 region 6 (A349-K356) and ß-conglycinin α subunit region 7 (E556-E575), which were located at the interior of the 3D structure, were the key regions contributing to discrepancies at the late stage.

The potential of plant-made vaccines to fight picornavirus.

INTRODUCTION: Several Picornaviruses are pathogens that generate serious problems for human and animal health worldwide. Vaccination is an attractive approach to fight against picornaviruses. In this regard, the development of low-cost vaccines is a priority to ensure coverage; especially in developing and low-income countries. In this context, plant-made vaccines are a convenient technology since plant cells are low-cost bioreactors capable of producing complex antigens that preserve their antigenic determinants; moreover, they can serve as biocapsules to achieve oral delivery. AREAS COVERED: In the present review the advances in the development of plant-made vaccines against picornaviruses are summarized and placed in perspective. The main diseases that have been targeted using this approach include Poliovirus, Food and mouth disease virus, Hepatitis A virus, and Enterovirus 71. EXPERT OPINION: Several vaccine candidates against picornavirus have been characterized at the preclinical level; with many of them capable of inducing humoral and cellular responses that led to neutralization of pathogens when evaluated in vitro and test animal challenge assays. Plant-made vaccines are a promise to fight picornaviruses; especially in the developing world where limited resources hamper vaccination coverage. A critical analysis of the road ahead for this technology is provided.

Clinical assessment of tolerability, immunological and cutaneous reactivity effects of an abbreviated schedule with Olea europaeanative extract of subcutaneous immunotherapy.

Summary: Objectives. To evaluate the tolerability and efficacy of Olea europaea subcutaneous immunotherapy (SCIT) on patients with rhinoconjunctivitis. Methods. In this open clinical trial patients were assigned to an abbreviated build-up scheme. The outcomes were: number, percentage, and severity of adverse reactions. Secondary outcomes included: changes in immunoglobulin titers and changes in dose-response skin prick tests. Results. Only 8 systemic reactions were registered, which represented 7/47 (14.9%) of patients and 8/429 (1.9%) of administered doses. Regarding immunological parameters the significant increases of sIgG and sIgG4 evidenced the changes in the patient immune system. Cutaneous reactivity decreased significantly. Conclusions. Olea europaea SCIT (Allergovac® depot ROXALL Medicina España S.A.) showed a good safety and tolerability profile. Immunological changes with induction of blocking IgG and decreases in cutaneous reactivity were detected in the patients.

Optimization of induction parameters, structure quality assessment by ATR-FTIR and in silico characterization of expressed recombinant polcalcin in three different strains of Escherichia coli.

The inhalation of Chenopodium album (C. album) pollen, especially polcalcin (Che a 3), has been reported as a significant source of allergic respiratory symptoms. This study was conducted to characterize biophysical differences of recombinant polcalcin come from three different Escherichia coli strains using attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR), optimize recombinant polcalcin expression, and linear B-cell epitopes identification using in silico methods. ATR-FTIR results of purified proteins showed spectra intensity variance in the amide I region, while there were no changes in pick position and shape of the bands. SHuffle® T7 Express was selected for subsequent optimization due to ability in the correction of the mis-oxidized bonds and promotes proper folding which was validated by ATR-FTIR analysis results. Then, Response Surface Methodology was performed to optimize critical factors including induction temperature, duration of induction, and concentration of inducer. The best partitioning conditions were achieved in 1.5 mM IPTG for 10.97 h at 29.9 °C. Finally, prediction of polcalcin B-cell epitopes was carried out which indicated the presence of 4 different epitopes. Together, the results may help to the development of diagnostic approaches and also vaccine manufacture for desensitization and modulation of the allergic response in patients.

An economic evaluation of immediate vs non-immediate activation of emergency medical services after epinephrine use for peanut-induced anaphylaxis.

BACKGROUND: Layperson food allergy management plans commonly stipulate that if epinephrine is used to immediately call 911 and seek care in the nearest medical facility for observation. OBJECTIVE: To evaluate the cost-effectiveness of this strategy, vs a watchful waiting approach before activating emergency medical services (EMS). METHODS: We performed a cost-effectiveness analysis using Markov modeling simulated over a 20-year horizon comparing activating EMS immediately after epinephrine use for allergic reactions to peanut vs a "wait and see" approach in which EMS was only activated if symptoms of the reaction did not promptly resolve after treatment. The base-case model assumed a 10-fold increased fatality risk with delayed EMS activation. RESULTS: The fatality risk associated with early EMS use was minimal, with a per-patient fatality rate over a 20-year horizon of 1.2 × 10-6, vs 1.9 × 10-6 for a wait and see approach. The incremental cost per life-year saved was $142,943,447 for early EMS vs wait and see, with the cost per death prevented reaching $1,349,335,651 as the simulation concluded. Cost of early EMS activation rose to $321,625,534 per life-year saved ($3,035,454,848 per death prevented) if a 5-fold increase in fatality risk was assumed, and was $12,997,173 per life-year saved ($122,689,936 per death prevented) if a 100-fold increase in fatality risk was assumed. CONCLUSION: Medical observation of a treated and promptly resolved peanut allergic reaction has minimal benefit and excessive costs. Immediately activating EMS after using epinephrine for a peanut allergic reaction in this context is not cost-effective.

Ragweed Pollen Allergy: Burden, Characteristics, and Management of an Imported Allergen Source in Europe.

Ambrosia artemisiifolia, also known as common or short ragweed, is an invasive annual flowering herbaceous plant that has its origin in North America. Nowadays, ragweed can be found in many areas worldwide. Ragweed pollen is known for its high potential to cause type I allergic reactions in late summer and autumn and represents a major health problem in America and several countries in Europe. Climate change and urbanization, as well as long distance transport capacity, enhance the spread of ragweed pollen. Therefore ragweed is becoming domestic in non-invaded areas which in turn will increase the sensitization rate. So far 11 ragweed allergens have been described and, according to IgE reactivity, Amb a 1 and Amb a 11 seem to be major allergens. Sensitization rates of the other allergens vary between 10 and 50%. Most of the allergens have already been recombinantly produced, but most of them have not been characterized regarding their allergenic activity, therefore no conclusion on the clinical relevance of all the allergens can be made, which is important and necessary for an accurate diagnosis. Pharmacotherapy is the most common treatment for ragweed pollen allergy but fails to impact on the course of allergy. Allergen-specific immunotherapy (AIT) is the only causative and disease-modifying treatment of allergy with long-lasting effects, but currently it is based on the administration of ragweed pollen extract or Amb a 1 only. In order to improve ragweed pollen AIT, new strategies are required with higher efficacy and safety.

Assessment of peanut allergen Ara h1 in processed foods using a SWCNTs-based nanobiosensor.

The goals of this research were to develop a rapid single-walled carbon nanotube (SWCNT)-based biosensor and to employ it to commercial food products for Ara h1 detection. The SWCNT-based biosensor was fabricated with SWCNTs immobilized with antibody (pAb) through hybridization of 1-pyrenebutanoic acid succinimidyl ester (1-PBASE) as a linker. The resistance difference (ΔR) was calculated by measuring linear sweep voltammetry (LSV) using a potentiostat. Resistance values increased as the concentration of Ara h1 increased over the range of 1 to 105 ng/L. The specific binding of anti-Ara h1 pAb to antigen including Ara h1 was confirmed by both indirect ELISA kit and biosensor assay. The biosensor was exposed to extracts prepared from commercial processed food containing peanuts, or no peanuts, and could successfully distinguish the peanut containing foods. In addition, the application of present biosensor approach documented the precise detection of Ara h1 concentrations in commercially available peanut containing foods.

Clinical Characteristics and Allergen Sensitization Patterns of Patients with Local Allergic Rhinitis in Southern China.

BACKGROUND: Local allergic rhinitis (LAR) is characterized by the production of specific IgE (sIgE) in the nasal mucosa without evidence of systemic atopy. The characteristics of LAR in Caucasians have been well documented. LAR is understudied in China, with prevalence, patient demographics, symptomatology, and the allergen sensitization profile being poorly understood. The purpose of this study is to investigate the demographics, characteristics, and allergen sensitization profile of patients with LAR in Southern China. METHODS: A total of 194 patients with rhinitis and 13 healthy subjects were enrolled in the current study. The patients' demographic data, clinical history, and symptoms were recorded. Local and systemic sIgE to a wide panel of specific allergens were measured in the nasal secretion and serum samples. RESULTS: Among the rhinitis patients, 115 were classified as allergic rhinitis (AR; 59.3%), 15 as LAR (7.7%), and 64 as non-AR (33.0%). The demographic characteristics, duration, frequency, and severity of symptoms were similar, although LAR exhibited higher symptom scores for nasal itch. Monosensitization was the predominant pattern of sensitization in both AR (109 out of 115, 95%) and LAR (14 out of 15, 93%). House dust mite was the dominant allergen in AR patients (109 out of 115, 95%), while pollen was the dominant allergen in LAR patients (11 out of 15, 73%). CONCLUSION: The prevalence of LAR patients in Southern China was 7.7%. Pollen was the most common sensitizing allergen for the local LAR patients, which differs from Caucasian studies, in which house dust mite was the dominant sensitizing allergen. Monosensitization was the predominant pattern in both AR and LAR.

Cost-effectiveness of grass pollen subcutaneous immunotherapy (SCIT) compared to sublingual immunotherapy (SLIT) and symptomatic treatment in Austria, Spain, and Switzerland.

BACKGROUND: While specific immunotherapy (SIT) has been proven to be cost-effective for the treatment of allergic rhinitis compared to symptomatic treatment, there is a lack of European studies in which sublingual (SLIT) and subcutaneous (SCIT) immunotherapy were compared. The present analysis is focused on the cost-effectiveness of SCIT compared to SLIT and symptomatic treatment of grass pollen allergy in Austria, Spain, and Switzerland. It will address specific properties of the underlying healthcare systems. METHODS: The investigation is based on a previously published health economic model calculation. This was designed as a Markov model with pre-defined health stages and a duration of 9 years covering specific preparations for SCIT (Allergovit) and SLIT (Oralair). The effectiveness was assessed as symptom-score based quality-adjusted life years (QALYs). Additionally, total cost has been determined as well as the cost-effectiveness of SCIT. The robustness of model results was proved in further sensitivity analyses. RESULTS: With regard to the effectiveness of both SCIT and SLIT, preparations were dominant compared to pharmacological symptomatic therapy. Both strategies were associated with additional cost, but, combined with the results on effectiveness, both have to be regarded as cost-effective. A direct comparison of the SCIT (Allergovit) and SLIT (Oralair) showed lower total costs of SCIT vs SLIT for Austria, Spain, and Switzerland (€1,368 vs €2,012, €2,229 vs €2,547, and €1,901 vs €2,220) and superior effectiveness (SCIT =8.02 QALYs; SLIT =7.98 QALYs; and symptomatic therapy =7.90 QALYs). CONCLUSION: In patients with allergic rhinitis, SIT offers cost-effective treatment options compared to symptomatic treatment. When comparing SCIT (Allergovit) and SLIT (Oralair), SCIT was dominant in terms of QALYs as well as costs, in particular due to a slightly higher patient compliance and lower drug costs.

Allergenicity Assessment of Transgenic Wheat Lines In Silico.

Agriculture biotechnology is a promising tool for developing varieties with enhanced quality and quantity. Transgenic proteins expressed by genetically modified (GM) food crops improve crop characteristics like nutritional value, taste, and texture, and endow plants with resistance against fungus, pests, and insects. Despite such potential benefits, there are concerns regarding possible adverse effects of GM crops on human health, animals and the environment. Among the proposed guidelines for GM food safety testing-the weight-of-evidence approach proposed by the Codex Alimentarius Commission (ALINORM 03/34A) is the most recent. Till date, several transgenic wheat lines have been developed and research is underway for further improvement. However, GM wheat is not being grown or consumed in any part of the world. In the present study, in silico tools were employed for safety testing of eight transgenes used for the development of transgenic wheat lines. Among the genes studied, none of them shared sequence homology with the reported allergens and may be safe for use in genetic engineering. In conclusion, gene selection for developing transgenic wheat lines should be done with utmost care to ensure its safety for feed and fodder.

Static and elevated pollen traps do not provide an accurate assessment of personal pollen exposure.

SUMMARY: Background. Volumetric pollen traps are commonly used to assess pollen exposure. These traps are well suited for estimating the regional mean airborne pollen concentration but are likely not to provide an accurate index of personal exposure. In this study, we tested the hypothesis that hair sampling may provide different pollen counts from those from pollen traps, especially when the pollen exposure is diverse. Methods. We compared pollen counts in hair washes to counts provided by stationary volumetric and gravimetric pollen traps in 2 different settings: urban with volunteers living in short distance from one another and from the static trap and suburban in which volunteers live in a scattered environment, quite far from the static trap. Results. Pollen counts in hair washes are in full agreement with trap counts for uniform pollen exposure. In contrast, for diverse pollen exposure, .individual pollen counts in hair washes vary strongly in quantity and taxa composition between individuals and dates. These results demonstrate that the pollen counts method (hair washes vs. stationary pollen traps) may lead to different absolute and relative contributions of taxa to the total pollen count. Conclusions. In a geographic area with a high diversity of environmental exposure to pollen, static pollen traps, in contrast to hair washes, do not provide a reliable estimate of this higher diversity.

Molecular diagnosis and immunotherapy.

PURPOSE OF REVIEW: To describe recent insights into how molecular diagnosis can improve indication and selection of suitable allergens for specific immunotherapy and increase the safety of this therapy. RECENT FINDINGS: As specific allergen immunotherapy targets specific allergens, identification of the disease-eliciting allergen is a prerequisite for accurate prescription of treatment. In areas of complex sensitization to aeroallergens or in cases of hymenoptera venom allergy, the use of molecular diagnosis has demonstrated that it may lead to a change in indication and selection of allergens for immunotherapy in a large proportion of patients when compared with diagnosis based on skin prick testing and/or specific IgE determination with commercial extracts. These changes in immunotherapy prescription aided by molecular diagnosis have been demonstrated to be cost-effective in some scenarios. Certain patterns of sensitization to grass or olive pollen and bee allergens may identify patients with higher risk of adverse reaction during immunotherapy. SUMMARY: Molecular diagnosis, when used with other tools and patients' clinical records, can help clinicians better to select the most appropriate patients and allergens for specific immunotherapy and, in some cases, predict the risk of adverse reactions. The pattern of sensitization to allergens could potentially predict the efficacy of allergen immunotherapy provided that these immunotherapy products contain a sufficient amount of these allergens. Nevertheless, multiplex assay remains a third-level approach, not to be used as screening method in current practice.

Crosslinking of peanut allergen Ara h 2 by polyphenol oxidase: digestibility and potential allergenicity assessment.

BACKGROUND: Peanut is one of the eight major food allergens. Its allergen, Ara h 2, can be recognized by over 90% of serum IgE samples from peanut-allergic patients. Therefore, reducing the allergenicity of Ara h 2 is especially important. RESULTS: In the present study, polyphenol oxidase (PPO), a protein cross-linking reaction catalyst that acts on tyrosine residue, was used to modify Ara h 2. After crosslinking, the microstructure, digestibility, IgG binding capability and IgE binding capability of Ara h 2 were analyzed. Cross-linking decreased the potential allergenicity of Ara h 2 by masking the allergen epitope, while the antigenicity of Ara h 2 changed slightly. After crosslinking, the apparent diameter of Ara h 2 was altered from 300 to 1700 nm or 220 nm, indicating that polymerization could either be inter- or intramolecular. Regarding digestibility, crosslinked Ara h 2 was relatively more easily digested by gastric fluid compared with the untreated Ara h 2, but much more difficult in the intestinal fluid. CONCLUSION: The crosslinking reaction catalyzed by PPO, as a non-thermal process, may be beneficial for avoiding food allergy. The reaction could mask allergen epitopes, decreasing the allergenicity of Ara h 2. © 2015 Society of Chemical Industry.

Assessment of the Allergenic Potential of Transgenic Wheat (Triticum aestivum) with Reduced Levels of ω5-Gliadins, the Major Sensitizing Allergen in Wheat-Dependent Exercise-Induced Anaphylaxis.

The ω5-gliadins are the major sensitizing allergens in wheat-dependent exercise-induced anaphylaxis (WDEIA). In this study, two-dimensional immunoblot analysis was used to assess the allergenic potential of two transgenic wheat lines in which ω5-gliadin genes were silenced by RNA interference. Sera from 7 of 11 WDEIA patients showed greatly reduced levels of immunoglobulin E (IgE) reactivity to ω5-gliadins in both transgenic lines. However, these sera also showed low levels of reactivity to other gluten proteins. Sera from three patients showed the greatest reactivity to proteins other than ω5-gliadins, either high-molecular-weight glutenin subunits (HMW-GSs), α-gliadins, or non-gluten proteins. The complexity of immunological responses among these patients suggests that flour from the transgenic lines would not be suitable for individuals already diagnosed with WDEIA. However, the introduction of wheat lacking ω5-gliadins could reduce the number of people sensitized to these proteins and thereby decrease the overall incidence of this serious food allergy.

Food safety assessment of an antifungal protein from Moringa oleifera seeds in an agricultural biotechnology perspective.

Mo-CBP3 is an antifungal protein produced by Moringa oleifera which has been investigated as potential candidate for developing transgenic crops. Before the use of novel proteins, food safety tests must be conducted. This work represents an early food safety assessment of Mo-CBP3, using the two-tiered approach proposed by ILSI. The history of safe use, mode of action and results for amino acid sequence homology using the full-length and short contiguous amino acids sequences indicate low risk associated to this protein. Mo-CBP3 isoforms presented a reasonable number of alignments (>35% identity) with allergens in a window of 80 amino acids. This protein was resistant to pepsin degradation up to 2 h, but it was susceptible to digestion using pancreatin. Many positive attributes were presented for Mo-CBP3. However, this protein showed high sequence homology with allergens and resistance to pepsin digestion that indicates that further hypothesis-based testing on its potential allergenicity must be done. Additionally, animal toxicity evaluations (e.g. acute and repeated dose oral exposure assays) must be performed to meet the mandatory requirements of several regulatory agencies. Finally, the approach adopted here exemplified the importance of performing an early risk assessment of candidate proteins for use in plant transformation programs.

Objective eliciting doses of peanut-allergic adults and children can be combined for risk assessment purposes.

BACKGROUND: To improve food labelling strategies, information regarding eliciting doses (EDs) and the effect of patient characteristics on these EDs is necessary. OBJECTIVE: To establish EDs for objective and subjective symptoms and analyse the effect of sensitization levels and other patient characteristics on threshold distribution curves (TDCs). METHODS: Threshold data from 100 adults and 262 children with a positive food challenge were analysed with interval-censoring survival analysis (ICSA) and fitted to a TDC from which EDs could be extracted. Possible influencing factors were analysed as covariates by ICSA. A hazard ratio (HR) was calculated in case of a significant effect. RESULTS: TDCs for both objective and subjective symptoms were significantly different between adults and children (P < 0.001). Objective ED05 values, however, were comparable (2.86 mg peanut protein in adults and 6.38 mg in children). Higher levels of sIgE to Ara h 2 and peanut extract were associated with a larger proportion of patient groups reacting to a dose increase with objective symptoms (adults and children) or subjective symptoms (adults, in children a trend). Age had a similar effect in children (HR 1.05 for objective symptoms and 1.09 for subjective symptoms). Gender had no effect on TDCs. CONCLUSION AND CLINICAL RELEVANCE: Subjective and objective TDCs were different between adults and children, but objective ED05 values were comparable, meaning that threshold data from children and adults can be combined for elaboration of reference doses for risk assessment. Higher sIgE levels to Ara h 2 and peanut extract were associated with a larger proportion of both patient groups to react to a certain dose increase.

Sublingual or subcutaneous immunotherapy for seasonal allergic rhinitis: an indirect analysis of efficacy, safety and cost.

RATIONALE, AIMS AND OBJECTIVES: The standard of preventive care for poorly controlled seasonal allergic rhinitis (AR) is subcutaneous immunotherapy (SCIT) with allergen extracts, administered in a physician's office. As an alternative to SCIT, sublingual immunotherapy (SLIT) is now an option for patients with seasonal AR. Oralair, a SLIT tablet containing freeze-dried allergen extracts of five grasses [cocksfoot (Dactylis glomerata), meadow grass (Poa pratensis), rye grass (Lolium perenne), sweet vernal grass (Anthoxanthum odoratum) and timothy grass (Phleum pratense)], and Grazax, a SLIT tablet containing a standardized extract of grass pollen allergen from timothy grass (P pratenase), are two such agents currently available in many countries. However, head-to-head comparative data are not available. In this study, an indirect comparison on efficacy, safety and cost was undertaken between Oralair, Grazax and SCIT. METHODS: A systematic review was conducted for double-blind placebo-controlled randomized trials evaluating Oralair, Grazax or SCIT in patients with grass-induced seasonal AR. Using placebo as the common control, an indirect statistical comparison between treatments was performed using meta regression analysis with active drug as the primary independent variable. An economic analysis, which included both direct and indirect costs for the Canadian setting, was also undertaken. RESULTS: Overall, 20 placebo-controlled trials met the study inclusion criteria. The indirect analysis suggested improved efficacy with Oralair over SCIT [standardized mean difference (SMD) in AR symptom control = -0.21; P = 0.007] and Grazax (SMD = -0.18; P = 0.018). In addition, there were no significant differences in the risk of discontinuation due to adverse events between therapies. Oralair was associated with cost savings against year-round SCIT ($2471), seasonal SCIT ($948) and Grazax ($1168) during the first year of therapy. CONCLUSIONS: Oralair has at least non-inferior efficacy and comparable safety against SCIT and Grazax at a lower annual cost.

Comparative assessment of multiple criteria for the in silico prediction of cross-reactivity of proteins to known allergens.

Genetically modified crops are becoming important components of a sustainable food supply and must be brought to market efficiently while also safeguarding the public from cross-reactivity of novel proteins to known allergens. Bioinformatic assessments can help to identify proteins warranting further experimental checks for cross-reactivity. This study is a large-scale in silico evaluation of assessment criteria, including searches for: alignments between a query and an allergen having ≥ 35% identity over a length ≥ 80; any sequence (of some minimum length) found in both a query and an allergen; any alignment between a query and an allergen with an E-value below some threshold. The criteria and an allergen database (AllergenOnline) are used to assess 27,243 Viridiplantae proteins for potential allergenicity. (A protein is classed as a "real allergen" if it exceeds a test-specific level of identity to an AllergenOnline entry; assessment of real allergens in the query set is against a reduced database from which the identifying allergen has been removed.) Each criterion's ability to minimize false positives without increasing false negative levels of current methods is determined. At best, the data show a reduction in false positives to ∼6% (from ∼10% under current methods) without any increase in false negatives.