Detection of Aspergillus antibodies by a new indirect haemagglutination assay

Purpose: Measuring Aspergillus antibodies is an important part of the diagnostic pathway for allergic bronchopulmonary aspergillosis (ABPA) and chronic pulmonary aspergillosis (CPA). It may represent a major public health issue on a global scale as 20-35% of patients develop Aspergillus antibodies following tuberculosis treatment and 63% of these develop pulmonary aspergillosis within 3 years. The worldwide 5 year period prevalence of (CPA) secondary to tuberculosis in, for example, the Congo and Nigeria has been estimated at between 0.8 and 1.37 million cases, with 43 cases per 100,000 population. Detection of specific antibodies provides key diagnostic evidence in chronic aspergillosis. There are numerous EIA formats for quantifying antibodies but these are not suitable for use in developing countries with limited laboratory resources. Haemagglutination tests involve coating erythrocytes with antigens. Erythrocytes clump together when antibodies cross-react with antigens on more than one cell and become visible to the human eye. The method also detects all Aspergillus antibody types. Its simplicity and speed (~2 hours) and commercial production make it highly suited for epidemiological and prevalence studies in low and middle resource countries. The goal was to compare the efficacy of an indirect haemagglutination assay designed to detect Aspergillus agglutinating antibodies with an agar double diffusion system used for the detection of Aspergillus precipitins. Methods: Serum samples from patients with a diagnosis of chronic pulmonary aspergillosis were tested by ELI.H.A. Aspergillus indirect haemagglutination (ELITech MICROBIO, Signes, France), and by an Aspergillus immunodiffusion system (Microgen Bioproducts Ltd, Camberley, UK). For the indirect haemagglutination assay sera were serially diluted to 1:2560. For the immunodiffusion assay patient sera were diluted to 1:16. The antigens used in these assays were a combination of cytoplasmic (somatic) and culture filtration extracts. Results: In the indirect haemagglutination assay sera with a titre <1:320 were considered to be a nonsignificant reaction. Sera with a titre equal to 1:320 were considered to be an indeterminant reaction. Titres 8805;1:640 indicated a significant reaction in favour of aspergillosis. A positive precipitin reaction in the immunodiffusion test signified the presence of Aspergillus precipitating antibodies. The performance of positive control sera in both assays was excellent, and reproducible. All sera positive in one test were positive in the other. The concordance between titres determined by either the indirect haemagglutination assay or the precipitin test were varied. High precipitin titres (1:8) were recorded as >1:2560 in the agglutination assay. Precipitin titres of 1:4 were all in the positive range of the agglutination assay. Sera recorded as weak precipitin positive (titre of 1:2) were all positive in the agglutination test but with a range of agglutinin titres (1:320 – >1:2560). The immunodiffusion test takes 5 days to perform. The total performance time of the indirect haemagglutination test is 3 hours. Conclusion: The EliTech indirect haemagglutination assay for the detection of Aspergillus antibody in patients with chronic manifestations of pulmonary demonstrated has many advantages compared with precipitin tests: it was rapid, very user friendly and easy to read. This is an ideal near point-of-care test for field studies and for community clinics. Furthermore, this test can be used for screening patients and support our efforts to understand the global epidemiology of chronic pulmonary aspergillosis. 2014 abstract No: 97 Full conference title: 6th Advances Against Aspergillosis 2014 http://www.AAA2014.org Conference abstracts