Helios-Negative Regulatory T Cells as a Key Factor of Immune Tolerance in Nonallergic Beekeepers.

BACKGROUND AND OBJECTIVES: Although exposure to stings has been identified as the leading risk factor for anaphylaxis due to Hymenoptera venom allergy, professional beekeepers receive hundreds of stings yearly without developing systemic reactions. This study aims to analyze the mechanisms underlying bee venom tolerance in beekeepers. METHODS: A cross-sectional study was conducted. Participants were recruited and classified into 3 groups: allergic patients (APs), who experienced systemic reactions after bee stings, with a positive intradermal test and specific IgE (sIgE) to Apis mellifera venom (AmV); tolerant beekeepers (TBKs), who received ≥50 stings/year; and healthy nonexposed controls (HCs). We measured serum levels of sIgE and specific IgG4 (sIgG4) to AmV, rApi m 1, rApi m 2, rApi m 3, Api m 4, rApi m 5, and rApi m10, as well as AmV-induced basophil degranulation, percentage of T-cell subsets, regulatory T cells (Treg), and IL-10 production. RESULTS: Compared with TBKs, APs had high levels of sIgE to AmV and all its allergic components (P<.001), together with a high basophil activation rate (P<.001). Conversely, compared with APs, TBKs had higher levels of sIgG4 (P<.001) and IL-10 (P<.0001), as well as an enhanced CTLA-4+ Treg population (P=.001), expanded Helios- Treg (P<.003), and reduced type 1 helper T cells (TH1) (P=.008), TH2 (P=.004), and TH17 (P=.007) subsets. CONCLUSIONS: The profile of TBKs, which was strongly marked by Treg activity, differed from that of TBKs. This natural tolerance would be led by the expansion of inducible Helios- Treg cells at the peripheral level. The Helios- Treg population could be a novel candidate biomarker for monitoring tolerance.

The Lights and the Shadows of Controlled Sting Challenge With Hymenoptera.

Hymenoptera venom immunotherapy (VIT) is effective for protecting individuals with systemic allergic reactions caused by Hymenoptera stings. The need for a tool that shows the degree of protection afforded by VIT and the lack of useful biomarkers have made the sting challenge test (SCT) the gold standard for this disorder, although its use has both lights and shadows. SCT with Hymenoptera involves causing a real sting in a patient diagnosed with allergy to the venom of the stinging insect and who is undergoing treatment with specific immunotherapy. In Spain, SCT is included in the list of services offered by some hospitals and forms part of their daily clinical practice. This review aims to analyze the strengths and weaknesses of this test and to describe the standardized procedure and necessary resources, based on the experience of a group of Spanish experts and a review of the literature.

Seroepidemiological evidence for Taenia solium taeniasis/cysticercosis in three Venezuelan rural communities.

Taenia solium is the most common parasite infection of the brain, causing neurocysticercosis and typically found in rural communities with free-ranging pigs. Identification of transmission in rural areas is essential for its control. Risk factors and transmission of the parasite were evaluated in three rural Venezuelan communities (Valle del Rio and Potrero Largo, Cojedes state; and Palmarito, Portuguesa state) by a questionnaire (112 households) and coprological (492 samples) and serological (433 human and 230 porcine sera) analysis, respectively. Typical risk factors were found in all three communities: free-foraging pig husbandry, deficient sanitary conditions, high open defecation and ignorance of the parasite life cycle. Coprological examinations revealed a high level of soil-transmitted parasites. Importantly, two T. solium adult worm carriers were identified in each of the three communities. Anti-metacestode antibodies and the HP10 secreted metacestode glycoprotein were detected at significant levels in human and porcine sera in Valle del Rio, Potrero Largo and Palmarito. In conclusion, these communities may be considered to be endemic for taeniasis/cysticercosis, and the instigation of an appropriate control programme is recommended.

Quality Standards for Allergen Immunotherapy Clinics in Spain: Consensus Document.

BACKGROUND AND OBJECTIVE: Allergen immunotherapy clinics (AITCs) in Spain differ widely in terms of structure, organization, resources, and portfolio of services. Therefore, it is essential to unify treatment criteria and define quality standards for the most complex AITCs. Objective: To establish a series of recommendations that make it possible to guarantee quality and safety in the administration of immunotherapy and define quality standards for the most complex AITCs. METHODS: This project began with an online survey of 65 allergy departments/units throughout Spain in 2013. Next, a 2-phase consensus process was carried out. In the first phase, 10 experts defined and agreed on the standards using the RAND/UCLA Appropriateness method; in the second, the agreements were validated by means of a 2-round Delphi consultation with 84 experts. RESULTS: Consensus was reached on minimum safety and quality criteria in the administration of allergen immunotherapy, and 2 levels of highly complex AITCs were defined: accredited AITCs and accredited AITCs with excellence. Consensus was also reached on quality standards and accreditation criteria for both levels. CONCLUSIONS: This project is pioneering in terms of its purpose (the definition of quality standards for AITCs) and of the use of structured participation techniques (combination of the RAND/UCLA and Delphi methods). It enabled the design of minimum standards for quality and safety in administering AIT, as well as quality criteria for accreditation of AITCs supported by a broad panel of experts from the Spanish Society of Allergology and Clinical Immunology.

Nuclear medicine: workplace monitoring and internal occupational exposure during a ventilation/perfusion single-photon emission tomography.

The administration of 99mTc-HDP to diagnose pulmonary thromboembolisms leads to the presence of 99mTc in the environment of a nuclear medicine department, which could pose a potential risk of internal contamination to medical staff. Therefore, air samples from the administration room, gamma camera room and corridor of such a department were taken for the purpose of performing a workplace monitoring program of the medical centre under study, with maximum activity values of 640 ± 30 kBq/m3, 1.5 ± 0.1 kBq/m3 and 54 ± 3 kBq/m3, respectively, being obtained. These results correspond to committed effective doses received by exposed employees, via inhalation, when one ventilation/perfusion single-photon emission tomography study was performed, of 0.7 µSv, 0.004 µSv and 0.2 µSv, respectively. As inhalation is the employees' main exposure pathway to radio-aerosols, the internal dose of the nuclear medicine department's medical staff was also evaluated via urine bioassay measurements. Nuclear medicine nurses showed the highest 99mTc activity in 24-h urine samples (2100 ± 130 Bq/day), resulting in a committed effective dose of 21 µSv for each diagnostic study performed. Even so, the performance of ventilation/perfusion diagnostic studies did not constitute a substantial radiological risk since the annual dose limit for exposed employees was not exceeded.

Molecular characterization of Trypanosoma cruzi and Leishmania spp. coinfection in mammals of Venezuelan coendemic areas.

BACKGROUND & OBJECTIVES: Trypanosoma cruzi and Leishmania spp. are protozoans that cause American trypanosomiasis and leishmaniasis, respectively. In endemic foci where both diseases coincide, coinfection can occur. The objective of this work was the characterization of the parasites involved in coinfection in several endemic areas of Venezuela. METHODS: Molecular characterization was done in 30 samples of several species of mammals (Didelphis marsupialis, Equus mulus, Rattus rattus, Canis familiaris, Felis catus, and Sciurus granatensis) from the states of Anzoategui, Cojedes and Capital District diagnosed with T. cruzi and Leishmania spp. coinfections. For the typing of T. cruzi DTUs, the markers of miniexon, 24Sa rDNA, 18Sa rDNA, and hsp60-PCR-RFLP (EcoRV) were used. Infection by Leishmania spp. was characterized by miniexon multiplex PCR for complexes of Leishmania and ITS1-PCR-RFLP (HaeIII, HhaI, and RsaI) for the identification of the species. RESULTS: The T. cruzi TcI was present in 100% of the coinfected mammals, which included 76.7% of triple infection by T. cruzi TcI-complex-L. (L) mexicana-L. infantum/chagasi, 13.3% of double infection by T. cruzi TcI-L. mexicana and 10% of double infection by T. cruzi Tcl-L. infantum/chagasi. INTERPRETATION & CONCLUSION: These results suggest that the double or triple infection is a phenomenon existing in almost all the coendemics areas and mammals studied, which might influence the mechanisms of adaptation and pathogenicity of these parasites.

Successful desensitization to cloxacillin in a patient with sepsis, with infective endocarditis and clinical suspicion of hypersensitivity to penicillins, a case report.

WHAT IS KNOWN AND OBJECTIVE: Beta-lactam antibiotic (BLA) therapy is frequently needed to treat infective endocarditis (IE). Hypersensitive reactions to BLA restrict BLA therapy in allergic patients. In the current case, we aim to describe the utility of desensitization (DS) in this context. Although the evidence is limited, DS is recommended. CASE DESCRIPTION: This case report deals with a 79-year-old woman with a clinical suspicion of allergy to BLA and a methicillin-sensitive Staphylococcus aureus (MSSA) IE. A cloxacillin DS protocol was developed to enable treatment with cloxacillin. WHAT IS NEW AND CONCLUSION: Alternative antibiotic treatments may be less effective or not available in MSSA IE. In this case report, DS allowed optimal cloxacillin treatment.

Component-resolved diagnosis in hymenoptera allergy.

Component-resolved diagnosis based on the use of well-defined, properly characterised and purified natural and recombinant allergens constitutes a new approach in the diagnosis of venom allergy. Prospective readers may benefit from an up-to-date review on the allergens. The best characterised venom is that of Apis mellifera, whose main allergens are phospholipase A2 (Api m1), hyaluronidase (Api m2) and melittin (Api m4). Additionally, in recent years, new allergens of Vespula vulgaris have been identified and include phospholipase A1 (Ves v1), hyaluronidase (Ves v2) and antigen 5 (Ves v5). Polistes species are becoming an increasing cause of allergy in Europe, although only few allergens have been identified in this venom. In this review, we evaluate the current knowledge about molecular diagnosis in hymenoptera venom allergy.

Control of Bull's-Eye Rot of Apple Caused by Neofabraea perennans and Neofabraea kienholzii Using Pre- and Postharvest Fungicides.

Bull's-eye rot is a major postharvest disease of apple caused by several fungi belonging to the Neofabraea and Phlyctema genera. Chemical control of these fungi is a crucial component of disease management for apples that are conventionally grown. The efficacy of several preharvest and postharvest applied fungicides were evaluated to identify effective chemistries that can control bull's-eye rot incited by Neofabraea perennans and N. kienholzii on apples. In general, the preharvest fungicide thiophanate-methyl was found to be effective at reducing disease caused by N. perennans and N. kienholzii. Two postharvest fungicides, thiabendazole and pyrimethanil, also provided disease control that was far superior to other chemical compounds evaluated in this study. The efficacy of thiabendazole and pyrimethanil was unaffected by application method (fungicide dip compared with thermofog). Despite providing satisfactory control of bull's-eye rot, integration of these three chemicals into disease management programs should proceed judiciously with consideration of their impact on the development of fungicide resistance and influence on diversity in populations of apple postharvest pathogens.

Application of recombinant antigen 5 allergens from seven allergy-relevant Hymenoptera species in diagnostics.

BACKGROUND: Hymenoptera stings can cause severe anaphylaxis in untreated venom-allergic patients. A correct diagnosis regarding the relevant species for immunotherapy is often hampered by clinically irrelevant cross-reactivity. In vespid venom allergy, cross-reactivity between venoms of different species can be a diagnostic challenge. To address immunological IgE cross-reactivity on molecular level, seven recombinant antigens 5 of the most important Vespoidea groups were assessed by different diagnostic setups. METHODS: The antigens 5 of yellow jackets, hornets, European and American paper wasps, fire ants, white-faced hornets, and Polybia wasps were recombinantly produced in insect cells, immunologically and structurally characterized, and their sIgE reactivity assessed by ImmunoCAP, ELISA, cross-inhibition, and basophil activation test (BAT) in patients with yellow jacket or Polistes venom allergy of two European geographical areas. RESULTS: All recombinant allergens were correctly folded and structural models and patient reactivity profiles suggested the presence of conserved and unique B-cell epitopes. All antigens 5 showed extensive cross-reactivity in sIgE analyses, inhibition assays, and BAT. This cross-reactivity was more pronounced in ImmunoCAP measurements with venom extracts than in sIgE analyses with recombinant antigens 5. Dose-response curves with the allergens in BAT allowed a differentiated individual dissection of relevant sensitization. CONCLUSIONS: Due to extensive cross-reactivity in various diagnostic settings, antigens 5 are inappropriate markers for differential sIgE diagnostics in vespid venom allergy. However, the newly available antigens 5 from further vespid species and the combination of recombinant allergen-based sIgE measurements with BAT represents a practicable way to diagnose clinically relevant sensitization in vespid venom allergy.

Variability among strains of Aspergillus section Nigri with capacity to degrade tannic acid isolated from extreme environments.

Tannins are polyphenolic compounds that cause astringent flavor and turbidity in food. Tannase is an enzyme that catalyzes the hydrolysis of tannins and is used in food industry. This study was conducted to determine the genetic variability and the tannase alleles variation in fungal strains isolated from soil and plants at five extreme areas of Coahuila, México. Two screening assays under 1 and 20 % of tannic acid were performed, with the isolations. In these assays, it was possible to identify 756 and 128 fungal strains, respectively. The major fungal variability was observed in "Cuatro Ciénegas" with 26 strains. The microorganisms were distributed in 11 groups, which correspond to Aspergillus section Nigri. AN7 and AN1 groups showed the major number of isolates from "Paila" and "Cuatro Ciénegas" locations, respectively. In the last location, the major diversity and specific richness were found. But in "Ojo Caliente," tannase allele conservations were observed.

Daytime restricted feeding modifies the daily regulation of fatty acid ß-oxidation and the lipoprotein profile in rats.

Daytime restricted feeding (2 h of food access from 12.00 to 14.00 hours for 3 weeks) is an experimental protocol that modifies the relationship between metabolic networks and the circadian molecular clock. The precise anatomical locus that controls the biochemical and physiological adaptations to optimise nutrient use is unknown. We explored the changes in liver oxidative lipid handling, such as ß-oxidation and its regulation, as well as adaptations in the lipoprotein profile. It was found that daytime restricted feeding promoted an elevation of circulating ketone bodies before mealtime, an altered hepatic daily rhythmicity of 14CO2 production from radioactive palmitic acid, and an up-regulation of the fatty acid oxidation activators, the α-subunit of AMP-activated protein kinase (AMPK), the deacetylase silent mating type information regulation homolog 1, and the transcriptional factor PPARγ-1α coactivator. An increased localisation of phosphorylated α-subunit of AMPK in the periportal hepatocytes was also observed. Liver hepatic lipase C, important for lipoprotein transformation, showed a change of daily phase with a peak at the time of food access. In serum, there was an increase of LDL, which was responsible for a net elevation of circulating cholesterol. We conclude that our results indicate an enhanced fasting response in the liver during daily synchronisation to food access, which involves altered metabolic and cellular control of fatty acid oxidation as well a significant elevation of serum LDL. These adaptations could be part of the metabolic input that underlies the expression of the food-entrained oscillator.

Posttraumatic growth and well-being among adolescents and young adults (AYAs) with cancer: a longitudinal study.

PURPOSE: This study examines posttraumatic growth (PTG) among adolescents and young adults (AYAs) with cancer, as well as its correlates and trajectories over time. The study also explores the buffering role of PTG on the associations between posttraumatic stress (PTS), health-related quality of life (HRQoL), and psychological distress. METHODS: A multicenter, longitudinal, prospective study was conducted among AYA cancer patients aged 14-39 years. One hundred sixty-nine patients completed a self-report measure of PTG (PTGI) and PTS (PDS) 6, 12, and 24 months after baseline (within the first 4 months of diagnosis). At 24-month follow-up, HRQoL (SF-36) and psychological distress (BSI-18) were also assessed. RESULTS: Among participants, 14% showed increasing PTG, 45% remained at a stable high PTG level, 14% showed decreasing PTG, and 27% remained at a stable low PTG level. AYAs who remained high on PTG were more often younger, female, and received chemotherapy. PTG level at 6-month follow-up was predictive of mental HRQoL (ß = 0.19; p = 0.026) and psychological distress (ß = -0.14; p = 0.043) at 24-month follow-up when corrected for PTS and sociodemographic and clinical covariates. No relationship between PTG and physical HRQoL was found. The interactive effects of PTS and PTG on outcomes were not significant, indicating that buffering did not take place. CONCLUSION: This study indicates that PTG is dynamic and predicts mental well-being outcomes but does not buffer the effects of PTS. Psychosocial interventions should focus on promoting PTG and reducing PTS in order to promote the adjustment of AYAs diagnosed with cancer.

Fermentation of Dietetic Fiber from Green Bean and Prickly Pear Shell by Pure and Mixture Culture of Lactobacillus acidophilus LA-5 and Bifidobacterium bifidum 450B.

The aim of this study was to evaluate the fermentation of dietary fiber from green bean (Phaseolus vulgaris) and prickly pear shell (Opuntia ficus-indica) by Lactobacillus acidophilus LA-5 and Bifidobacterium bifidum 450B growing as mono-culture and co-culture, the fermentation products, and proteins expressed during this process. The analysis of the fermentation profile showed a major growth of bacteria in the culture media of each dietary fiber supplemented with glucose, and particularly B. bifidum 450B at 48 h showed the highest growth. In the case of the co-culture, the growth was lower indicating the possible negative interaction between L. acidophilus LA-5 and B. bifidum 450B and may be due to the less amount of carbohydrates and the high content of non-soluble fiber that affected the nutrients availability for the bacterial strains. The pH changes indicated the presence of short-chain fatty acids (SCFAs), being acetate (46-100%) the main SCFA. Changes in the proteome concerned proteins that are involved in carbohydrate and other carbohydrate pathways. The characterization of the bacteria according to the growth, metabolites, and proteins expressed allows understanding the response to the change of environmental conditions and could be useful to understand L. acidophilus LA-5 and B. bifidum 450B strains' adaptation to specific applications.

Timing of Apple Fruit Infection by Neofabraea perennans and Neofabraea kienholzii in Relation to Bull's-eye Rot Development in Stored Apple Fruit.

Bull's-eye rot is a postharvest disease of pome fruit in the U.S. Pacific Northwest. The disease is caused by the fungi Neofabraea kienholzii, N. malicorticis, N. perennans, and N. vagabunda. Fruit infection by these pathogens is initiated in the orchard during the fruit-growing season but remains latent at harvest. For fruit held in postharvest cold storage, bull's-eye rot symptom development is slow to progress, requiring at least 3 months before symptoms are first visible. In order to determine the timing of preharvest fruit infection in relation to bull's-eye rot development in cold storage, 'Red Delicious' and 'Fuji' apple fruit were inoculated with a conidial suspension of N. perennans or N. kienholzii at different inoculation timings throughout the fruit growing seasons of 2012 to 2014. Fruit were harvested and stored at 0°C for up to 10 months, during which time disease incidence was recorded periodically. Results from this study demonstrate that apple fruit infection by either pathogen may occur at any point during the growing season. However, infections occurring over the 8-week period immediately prior to harvest yield a higher incidence of bull's-eye rot in stored fruit compared with infections initiated earlier in the growing season.

Design and application of a 23-gene panel by next-generation sequencing for inherited coagulation bleeding disorders.

INTRODUCTION: Molecular testing of Inherited bleeding coagulation disorders (IBCDs) not only offers confirmation of diagnosis but also aids in genetic counselling, prenatal diagnosis and in certain cases genotype-phenotype correlations are important for predicting the clinical course of the disease and to allow tailor-made follow-up of individuals. Until recently, genotyping has been mainly performed by Sanger sequencing, a technique known to be time consuming and expensive. Currently, next-generation sequencing (NGS) offers a new potential approach that enables the simultaneous investigation of multiple genes at manageable cost. AIM: The aim of this study was to design and to analyse the applicability of a 23-gene NGS panel in the molecular diagnosis of patients with IBCDs. METHODS: A custom target enrichment library was designed to capture 31 genes known to be associated with IBCDs. Probes were generated for 296 targets to cover 86.3 kb regions (all exons and flanking regions) of these genes. Twenty patients with an IBCDs phenotype were studied using NGS technology. RESULTS: In all patients, our NGS approach detected causative mutations. Twenty-one pathogenic variants were found; while most of them were missense (18), three deletions were also identified. Six novel mutations affecting F8, FGA, F11, F10 and VWF genes, and 15 previously reported variants were detected. NGS and Sanger sequencing were 100% concordant. CONCLUSION: Our results demonstrate that this approach could be an accurate, reproducible and reliable tool in the rapid genetic diagnosis of IBCDs.

Respiratory tract infection caused by Mycobacterium bovis in a black swan (Cygnus atratus).

A 3-year-old male black swan (Cygnus atratus), belonging to a private collection, died suddenly and was subjected to post mortem examination. At necropsy, caseous exudate was observed in the lungs and air sacs; granulomatous lesions characterized by epithelioid macrophages and abundant mycobacteria were observed microscopically. Avian tuberculosis associated with Mycobacterium bovis was confirmed by bacteriologic isolation, biochemical tests and molecular methods. The organism was identified as spoligotype SB0140, which is frequently found in cattle and people in North America. In this case, interspecies transmission could have been the source of infection because the swan cohabited with cattle.