Comparative immunochemical analysis of five Thermoactinomyces strains.

The antigen structures in mycelial extracts of 4 strains of Thermoactinomyces vulgaris and 1 strain of Thermoactinomyces candidus were compared using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining. For immunological characterization, antigens were transferred electrophoretically to nitrocellulose membranes and blotted with T. vulgaris strain-specific antisera and with sera of farmer's lung patients. Cross-reactions between the strains were also studied using immunodiffusion and crossed immunoelectrophoresis. Protein components between 50 and 60 kilodaltons were found to be the most immunogenic. Patient sera showed heterogeneous responses, but all reacted with an antigen at 55 kilodaltons that was also common to all strains studied.

Circulating IgG antibodies against fungal and actinomycete antigens in the sera of farmer's lung patients from different countries.

Sixty-nine farmer's lung patients and 28 normal controls from four countries (Finland, Switzerland, Canada and the United States) were investigated for antibody levels against 13 antigens commonly used for the screening panel for hypersensitivity pneumonitis. Of these antigens, eight were from the Medical College of Wisconsin (United States) and five were from the University of Kuopio (Finland). IgG antibodies against these antigens were studied in 97 sera using a sensitive biotin-avidin-linked enzyme immunoassay. The results indicate that the mean antibody titer against Micropolyspora faeni was highest in the United States (U.S.) followed by Finland. Both Finnish and U.S. antigens reacted almost identically against various groups of patients, although the degree of reactivity varied considerably. Higher antibody levels against Thermoactinomyces vulgaris were detected in Finnish patients than patients from other countries while patients from all four countries showed elevated levels of antibodies against T. candidus. This study demonstrates that antigens from identical species, irrespective of geographic origin, reacted similarly. However, variability between antigens of the same species was still considerably significant. Since the microbiological flora of moldy hay varies widely in different regions, the microbial species associated with the disease at a given geographical area has to be determined before selecting antigens for serological studies. The antigens currently used in various laboratories are crude preparations and need to be purified and standardized for dependable results. Until such antigens are available, all antigenic preparations used in the immunological evaluation of patients should be immunochemically characterized for their reproducibility and reliability although the ultimate goal should be to obtain standardized pure antigens for dependable immunodiagnosis of farmer's lung.

Significance of IgG antibodies against environmental microbial antigens in a farming population.

A group of dairy farmers studied 6 years earlier in a field survey was re-surveyed for respiratory symptoms, occupational capability and the presence of antibodies against environmental micro-organisms. Specific IgG antibodies to Micropolyspora faeni, Thermoactinomyces vulgaris, Aspergillus umbrosus and Aspergillus fumigatus were measured by ELISA from the serum samples obtained in the two surveys. Antibody titres remained constant in 70% of both farmers and controls, but where changes took place, the titres against the actinomycete antigens tended to rise, whereas both increases and decreases were detected equally against the Aspergillus antigens. The titre of specific antibody to any of the four micro-organisms, when measured from a single serum specimen, seemed to be of little diagnostic value. Observed changes however, were more diagnostic, in that a fall in titre, especially against the Aspergillus antigens, was closely associated with a definite decrease in exposure, such as after retirement. Increased titres occurred in farmers with continued exposure, and those against the actinomycetes were associated with the appearance of symptoms in previously symptom-free individuals. In a case of farmer's lung which developed in this population during the follow-up period, significant increases were detectable against T. vulgaris and M. faeni.

Familial aggregation of IgG antibody response to antigens associated with farmer's lung.

The levels of circulating IgG antibodies to Aspergillus umbrosus, Aspergillus fumigatus, Thermoactinomyces vulgaris, and Micropolyspora faeni were determined by enzyme immunoassay in 197 subjects selected for a study of farmer's lung (FL). The material consisted of five study groups: 37 patients with clinically confirmed FL, 31 spouses of the patients, 44 immediate relatives of the patients, 35 immediate relatives of the patients' spouses, and 50 unrelated people who were spouses of the 79 people in both relative groups. The mean titres of IgG antibodies to all four microbes were highest in patients with clinically established FL. In the other groups the mean titre of Aspergillus umbrosus, a mould found much more frequently in Finnish farm environments than other moulds under study, was significantly higher (p less than 0.01) in the relatives of FL patients than in other people. This finding remained irrespective of whether the subjects had suffered from FL symptoms or not or whether they worked or lived on the same farm as the patient or on a different one. The difference in the mean titre was not due to the differences between the study groups in age, sex, smoking habits, atopic background, frequency of handling of plant materials, or time interval from the most recent handling of visibly mouldy hay. The results imply that genetic factors may be important in the IgG antibody response to microbial antigens associated with FL.

Exposure of water consumers to mesophilic actinomycetes.

In autumn 1978 an epidemic of respiratory disease resembling allergic alveolitis occurred in a small Finnish community. The disease was caused by repeated exposures to tap water aerosol. The raw water of the community and the sand filters of the purification system were heavily contaminated with mesophilic actinomycetes. Fourteen different strains of actinomycetes were isolated. Exposed persons with and without symptoms as well as unexposed control persons were tested for antibodies against five of these actinomycetes and against Enterobacter agglomerans. Both the exposed and the control persons had antibodies to actinomycetes but the exposed persons had antibodies against more actinomycete strains than the control persons. Precipitating antibodies against E. agglomerans were also found in control persons as well as in patients. There was a significant difference between the patients and the exposed healthy persons in bacterial agglutination tests with flagellar antigen of one E. agglomerans strain. However, the role of mesophilic actinomycetes and E. agglomerans in the aetiology of the disease could not be firmly established.

Antigen release from Thermoactinomyces vulgaris by lysozyme treatment.

Lysozyme treatment was used to release antigenic material from Thermoactinomyces vulgaris, one of the microbes associated with farmer's lung. Lysozyme caused degradation of the murein layer visualized as changes and disappearance of the bacterial morphology in scanning electron microscopy. Enrichment of different antigenic components in the lysozyme extract and in the cell residue, respectively, was detected by immunoprecipitation. When lysozyme extract and cell residue antigens were used in enzyme-linked immunosorbent assay to test sera of farmer's lung patients and control persons, it became evident that there was no significant difference between the reactions against the two antigens. However, a number of sera reacted preferentially against one or the other of the two antigens.

Comparison of Aspergillus fumigatus and Aspergillus umbrosus antigens in serological tests of farmer's lung.

A comparison was made between Aspergillus fumigatus and A. umbrosus antigens regarding their usefulness in serological tests of farmer's lung. Mycelial and culture fluid antigens of both microbes were used to test sera of 30 Finnish farmer's lung patients for antibodies in the ELISA and in the gel precipitation (GP) test. Antibodies to A. umbrosus were more common than to A. fumigatus; this was especially evident in ELISA, ELISA gave more positive results with all four antigens than GP. Mycelial antigens showed more positive reactions in ELISA than the corresponding culture fluid antigens. The situation was reversed in GP. Selection of antigens in the diagnostic panel of farmer's lung requires careful consideration. Our results suggest that other Aspergilli (in the Finnish population A. umbrosus) might be preferable to A. fumigatus in the antigen panel.

Sensitisation against environmental antigens and respiratory symptoms in swine workers.

Adverse effects caused by airborne material to the respiratory tract are due either to non-specific irritation or to hypersensitivity. In this study 20 people employed in swine barns and 18 controls were tested for sensitisation against dusts present in the barn. Immunoprecipitation and enzyme-linked immunoassay (ELISA) were used to test for IgG antibodies; IgE antibodies against swine epithelium were tested using solid phase radioimmunoassay. Precipitins against swine antigens were found in two swine workers; while ELISA found six to be sensitised. Sensitisation against swine antigens correlated with exposure but not with the presence of symptoms. No IgE antibodies were found. Precipitins against feed antigens were detected in 12 workers; in nine of the 12 with symptoms, and in three of the eight asymptomatic workers. No single antigen was of special importance as an inducer of sensitisation. Sensitisation against feed dusts in barns, as indicated by the presence of circulating antibodies, suggests an immunological background for persistent symptoms. A large antigen penal should be used in testing for sensitisation because of the many immunogenic dusts present in the air in swine barns.

The use of enzyme-linked immunosorbent assay (ELISA) in the diagnosis of farmer's lung.

Enzyme-linked immunosorbent assay (ELISA) was adapted to measure antibodies against Aspergillus fumigatus, Thermoactinomyces vulgaris and Micropolyspora faeni. Mycelial and culture fluid antigens were used separately to analyze antibodies in three study groups: farmer's lung patients, patients with bronchitis and healthy control persons. The mycelial antigen of A. fumigatus gave more positive results than the corresponding culture fluid antigen. The situation was reversed for the actinomycetes. For this reason either a mycelial antigen or a combination of mycelial and culture fluid antigens should be used in ELISA in routine tests. The prevalence and the titers of antibodies against the three microbes detected by ELISA were higher in the farmer's lung group than in the other two groups.