Bird fancier's lung serodiagnosis by automated r-IgLL1 ELISA.

CONTEXT: Bird fancier's lung (BFL) is the most prevalent form of hypersensitivity pneumonitis (HP) worldwide. The current techniques used for the serological diagnosis of BFL all use crude extracts from feathers, droppings, and blooms as test antigens, which is associated with a lack of standardization and variability of the results. An antigenic protein, immunoglobulin lambda-like polypeptide-1 (IgLL1), isolated from pigeon droppings, was recently identified to be associated with BFL. We used genetic engineering to produce IgLL1 as a recombinant antigen. AIM: We aimed to prospectively validate the use of an automated ELISA based on recombinant IgLL1 protein (r-IgLL1) as the test antigen for the serological diagnosis of BFL. METHODS: Immunoprecipitation (IP) techniques (immunodiffusion (ID), immunoelectrophoresis (IEP)) and ELISA using r-IgLL1 were performed concomitantly over 10 months on 634 sera from patients with a BFL serodiagnosis request. Questionnaires were sent to obtain details on the avian exposure, clinical data, and final diagnosis. Concordance, sensitivity (Se), and specificity (Sp) of the two techniques were compared. RESULTS: In total, 72 completed questionnaires were returned with 18 cases of BFL diagnosed and 54 of non-BFL. The concordance between the ELISA and ID+IEP precipitation techniques was 71%. The combination of immunoprecipitation techniques showed a Se of 78% and a Sp of 67%. The ELISA using r-IgLL1 showed a Se of 89% and a Sp of 91%. The automated r-IgLL1 ELISA test is sufficiently efficient to be used alone for the diagnosis of patients exposed solely to Columbidae. In cases of other avian exposure, the Se and Sp of the r-IgLL1 ELISA used for screening combined with the immunodiffusion test for confirmation were 89% and 93%, respectively. CONCLUSIONS: The automated ELISA using r-IgLL1 is a promising tool for BFL serodiagnosis. Replacing immunodiffusion by the automated ELISA using r-IgLL1 as a screening technique will be the basis of our future strategy for BFL serodiagnosis.

Pilot Study Using Recombinant Antigens r-PROE and r-IGLL1 for the Serodiagnosis of Feather Duvet Lung.

BACKGROUND: Feather duvet lung (FDL) is an underestimated form of acute and chronic hypersensitivity pneumonitis. Serological tests for FDL need to be validated. We investigated the ability of recombinant pigeon Proproteinase E (r-PROE) and Immunoglobulin-lambda-like-polypeptide-1 (r-IGLL1) proteins to support the serological diagnosis of FDL, and propose them as a serological tool for clinicians to differentiate cases from FDL and Bird fancier's lung (BFL). METHODS: Specific IgG antibodies against r-PROE and r-IGLL1, analyzed with ELISA, were measured in patients diagnosed with FDL (n=31), BFL (n=15) controls exposed (n=15) and unexposed to feathers (n=15). RESULTS: The sensitivity and specificity of the r-PROE ELISA for the serological diagnosis of FDL cases versus exposed and unexposed controls were 74.2% and 86.7% respectively, with an index threshold of 0.5 (AUC: 0.89). In addition, this serological test was effective to support the serological diagnosis of FDL and BFL cases with significantly different thresholds. The r-IGLL1 ELISA was only effective for the serological diagnosis of BFL. Also, these two serological tests were useful for the diagnosis of both chronic and acute forms. CONCLUSIONS: The new diagnostic test for FDL using r-PROE protein should help to detect overt and hidden cases of FDL. The combination of both test will help the clinician in distinguish between the etiology of birds or feathers duvet.

Pilot Study Using Recombinant Antigens r-PROE and r-IGLL1 for the Serodiagnosis of Feather Duvet Lung.

BACKGROUND: Feather duvet lung (FDL) is an underestimated form of acute and chronic hypersensitivity pneumonitis. Serological tests for FDL need to be validated. We investigated the ability of recombinant pigeon Proproteinase E (r-PROE) and Immunoglobulin-lambda-like-polypeptide-1 (r-IGLL1) proteins to support the serological diagnosis of FDL, and propose them as a serological tool for clinicians to differentiate cases from FDL and Bird fancier's lung (BFL). METHODS: Specific IgG antibodies against r-PROE and r-IGLL1, analyzed with ELISA, were measured in patients diagnosed with FDL (n=31), BFL (n=15) controls exposed (n=15) and unexposed to feathers (n=15). RESULTS: The sensitivity and specificity of the r-PROE ELISA for the serological diagnosis of FDL cases versus exposed and unexposed controls were 74.2% and 86.7% respectively, with an index threshold of 0.5 (AUC: 0.89). In addition, this serological test was effective to support the serological diagnosis of FDL and BFL cases with significantly different thresholds. The r-IGLL1 ELISA was only effective for the serological diagnosis of BFL. Also, these two serological tests were useful for the diagnosis of both chronic and acute forms. CONCLUSIONS: The new diagnostic test for FDL using r-PROE protein should help to detect overt and hidden cases of FDL. The combination of both test will help the clinician in distinguish between the etiology of birds or feathers duvet.

Indoor Microbiome: Quantification of Exposure and Association with Geographical Location, Meteorological Factors, and Land Use in France.

The indoor microbial community is a mixture of microorganisms resulting from outdoor ecosystems that seed the built environment. However, the biogeography of the indoor microbial community is still inadequately studied. Dust from more than 3000 dwellings across France was analyzed by qPCR using 17 targets: 10 molds, 3 bacteria groups, and 4 mites. Thus, the first spatial description of the main indoor microbial allergens on the French territory, in relation with biogeographical factors influencing the distribution of microorganisms, was realized in this study. Ten microorganisms out of 17 exhibited increasing abundance profiles across the country: Five microorganisms (Dermatophagoïdes pteronyssinus, Dermatophagoïdes spp., Streptomyces spp., Cladosporium sphaerospermum, Epicoccum nigrum) from northeast to southwest, two (Cryptococcus spp., Alternaria alternata) from northwest to southeast, Mycobacteria from east to west, Aspergillus fumigatus from south to north, and Penicillium chrysogenum from south to northeast. These geographical patterns were partly linked to climate and land cover. Multivariate analysis showed that composition of communities seemed to depend on landscapes, with species related to closed and rather cold and humid landscapes (forests, located in the northeast) and others to more open, hot, and dry landscapes (herbaceous and coastal regions, located in the west). This study highlights the importance of geographical location and outdoor factors that shape communities. In order to study the effect of microorganisms on human health (allergic diseases in particular), it is important to identify biogeographic factors that structure microbial communities on large spatial scales and to quantify the exposure with quantitative tools, such as the multi-qPCR approach.

Hypersensitivity pneumonitis: A new strategy for serodiagnosis and environmental surveys.

We propose a strategy for serodiagnosis of hypersensitivity pneumonitis (HP): 1) question patients about their private or occupational activity, or visit him on site; 2) select panels of six somatic specific antigens appropriate for each type of exposure; 3) and use ELISA to test concomitantly two recombinant antigens highly specific to Farmer's lung, Metalworking-fluid HP, and for Bird fancier's lung. The serodiagnosis provides an immunological argument that may complete radiological, functional lung exploration and clinical features; 4) If the serodiagnosis is negative but the suspicion of HP is strong, a microbial analysis of the patient's specific exposure is conducted; 5) "A la carte" antigens are produced from the microorganisms isolated in the patient's environment sample and tested; 6) Finally, the patient may be asked to undergo a specific inhalation challenge with the offending antigens in a safety cabin, or to avoid his usual environment for a few days.

Usefulness of à la carte antigens for bird fancier's lung serodiagnosis: total dropping extract and/or dropping's microflora antigens.

Bird fancier's lung (BFL) is a pulmonary disease caused by inhalation of avian proteins. The involvement of the microorganisms of droppings has been assumed in the past and this idea still persists today. Our study aimed to compare by immunoprecipitation assay the detection of antibodies against both droppings and microorganisms in the sera of patients (n=15) and asymptomatic exposed controls (n=18). We found that 14/15 BFL patients had negative serological results for isolated microorganisms of the droppings, only one positive against Enterobacter sakasakii. Serological arguments were in accordance with diagnosis in 87 % of cases by testing à la carte antigens from each bird dropping versus 20 % using the standard antigenic panel. Otherwise, the microorganisms antigens issued from dropping flora were negative in 93 % of cases. Consequently, it's preferable to use the total extract from the patient's bird droppings to establish the serodiagnosis of the disease.

An immunoproteomic approach revealed antigenic proteins enhancing serodiagnosis performance of bird fancier's lung.

BACKGROUND: Bird fancier's lung (BFL) caused by repeated inhalation of avian proteins is the most common form of hypersensitivity pneumonitis. However, the exact identification of proteins involved is unknown, and serological test use for diagnosis need to be standardized. The objectives of this study were (i) to identify antigenic proteins from pigeon droppings (ii) to provide information about their location in avian matrices and (iii) to produce them in recombinant proteins to evaluate their diagnostic performances. METHOD: Antigenic proteins of pigeon dropping extracts were investigated using 2-dimensional immunoblotting with sera from patients with BFL, asymptomatic exposed controls and healthy volunteers. We investigated the origin of these antigenic proteins by analyzing droppings, blooms and sera using a shotgun proteomic analysis. BFL-associated proteins were produced as recombinant antigens in E. coli and were assessed in ELISA with sera from patients (n=25) and subject exposed controls (n=30). These diagnostic performances were compared with those obtained by precipitin techniques (agar gel double diffusion, immunoelectrophoresis). RESULTS: We identified 14 antigenic proteins mainly located in droppings and blooms. These proteins were involved in either the digestive or immune systems of pigeons. Using the recombinant BFL-associated proteins: Immunoglobulin lambda-like polypeptide-1 (IGLL1: sensitivity: 76%; specificity: 100%; AUC: 0.93) and Proproteinase E (ProE: sensitivity: 84%; specificity: 80%; AUC: 0.85), the ELISA test showed better performance than precipitin assays with pigeon dropping extracts (sensitivity: 60%; specificity: 93.3%; AUC: 0.76). CONCLUSION: IGLL1 and ProE were identified as the biomarkers of the disease. The use of these highly standardized antigens discriminates BFL cases from exposed subjects in serological assays. The results of this study offer new possibilities for the serological diagnosis of the disease. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov: Identifier NCT03056404.

Immunogenic proteins specific to different bird species in bird fancier's lung.

Bird fancier's lung (BFL) is a disease produced by exposure to avian proteins present in droppings, blooms, and serum of a variety of birds. Although serological test results are currently used to confirm clinical diagnosis of the disease, bird species specificity is poorly understood. This study aimed to contribute to a better understanding of the specificity of immunogenic proteins revealed from the droppings of three bird species. Sera from four patients with BFL and two controls without exposure were analyzed by Western blotting with antigens from droppings of two pigeon and budgerigar strains and two hen species. When the antigens from the droppings of the three bird species were compared, the profile of immunogenic proteins was different and there were similarities between strains of the same species. Only one 68-kD protein was common to pigeon and budgerigar droppings, while proteins of 200, 175, 140, 100, and 35 kD were detected as specific in one bird species. These results provide insight to further characterize these proteins, and to design new serological tests specific to different bird species. These tests may help to refine strategies of antigenic exclusion and also to allow a patient compensation in case of BFL of occupational origin.