[Immunologic diagnosis of severe Candida infections].
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Biomedical subjects
Publications and source records attributed to K Holmberg.
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46 reference strains and clinical isolated of Actinomyces israelli and 8 strains of Arachnia propionica--the causative microorganisms of actinomycosis in man--were tested for their in vitro susceptibility to penicillin, sulfaisodimidine, erythromycin, tetracycline, lincomycin, clindamycin, metronidazole and tinidazole by the agar dilution method. All strains were susceptible to benzylpenicillin (minimum inhibitory concentrations, MICs of less than or equal to 0.064 mug/ml). Many strains were resistant in vitro to sulfaisodimidine (MICs 4.0-32.0 mug/ml). Erythromycin, tetracycline, clindiamycin and lincomycin possessed in vitro activity at concentrations readily attainable in serum. Due to adverse side-effects associated with clindamycin and lincomycin therapy, erythromycin and tetracycline may be the best alternative drugs to penicillin in the treatment of actinomycosis. In contrast, in vitro resistance (MICs greater than or equal to 4.0 mug/ml) of the pathogenic actinomycetes to metronidazole and tinidazole implies that these drugs may not be valuable in the therapy of human actinomycosis.
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A numerical taxonomic study was made on 49 facultative anaerobic Gram-positive filamentous and/or diphtheroidal organisms isolated from dental plaques, carious dentin and faeces, together with 63 reference strains belonging to the genera Actinomyces, Arachnia, Bifidobacterium, Actinobacterium, Propionibacterium, Eubacterium and Lactobacillus. They were examined for 90 unit characters covering a wide range of tests and properties. The data were subjected to computer analysis in which the simple matching coefficient (SSM) and the similarity index (SJ) were calculated, and the results of single linkage techniques and an unweighted average linkage cluster analysis technique were compared. The strains fell into six major groups (phena). The Actinomyces strains were recovered in two phena; the first contained Actinomyces israelii and the other facultative anaerobic Actinomyces, including subclusters equal to taxospecies of A. odontolyticus and A. viscosus/A. naeslundii, while the other phenon corresponded to the genera Arachnia, Actinobacterium, Bifidobacterium and Propionibacterium. The groups of Arachnia and Actinobacterium each contained one species, representing taxospecies of Arachnia propionica and Actinobacterium meyerii. Taxonomic criteria, both constant and discriminative, were selected to form a diagnostic table useful for laboratory identification of this group of organisms. Immunofluorescence supported the numerical data.
Standard preparations of crude cytoplasmic and whole cell-associated antigen mixtures of Actinomyces israelii were analyzed by crossed immunoelectrophoresis (CIE), with a standard polyvalent antiserum comprising purified and concentrated immunoglobulin G antibodies to formolized whole cells of A. israelii serotypes 1 and 2. The standard antigens provided four antigen-antibody systems for A. israelii. The immunoprecipitation patterns of the system were compared, and the immunochemical characteristics of individual precipitates were analyzed. Each system contained specific precipitates, but also one or two precipitates which were immunochemically identical to precipitates of the other systems. The standard system for A. israelii based on cytoplasmic antigens was best reproducible and revealed the highest number of immunoprecipitates. These precipitates possessed immunochemical properties which made them suitable for CIE studies. The cytoplasmic antigen mixture of A. israelii was, therefore, adopted as the most suitable for further development of a crossed immunoelectrophoretic system for A. israelii. In subsequent assays the cytoplasmic antigen mixture was raised in rabbit against cell lysates of A. israelii, serotypes 1 and 2. A standard antigen-antibody system for A. israelii was obtained which revealed an immunoprecipitation pattern of 10 distinguishable precipitates. The resolving power and separation by CIE of this standard system for A. israelii was compared with that of crossed immunoelectrofocusing. The results suggest that these methods supplement each other. Crossed immunoelectrofocusing appeared to be a useful tool for separation of specific components of the protein-antigen complex of A. israelii for analytic serology. The CIE in conjunction with a standard reference antigen-antibody system for A. israelii based on cytoplasmic antigens offers great potentialities in diagnostic A. israelii serology.
Crossed immunoelectrophoresis (CIE) with intermediate gel was applied to the serological analysis of Actinomyces israelii to develop a test with high efficiency in the laboratory diagnosis of human actinomycosis and classification of A. israelii. Recently developed standard antigen-antibody systems for A. israelii by CIE were used as reference. The reference systems were based on standard preparations of cytoplasmic and whole cell-associated antigens of A. israelii and a standard immunoglobulin G pool purified from rabbit antisera to formalin-treated whole cells and cell lysates of A. israelii. The specificity of the standard antigens for A. israelii was evaluated in CIE studies by screening for antibodies to components of the antigens in rabbit antisera raised against related bacteria. The standard system for A. israelii based on cytoplasmic antigens formed species-specific precipitins whereas antisera raised against A. naeslundii and/or Propionibacterium acnes precipitated components of the other standard antigens. As a result of these analyses, the standard system for A. israelii based on 10 cytoplasmic antigens was used as reference for CIE studies to detect humoral antibodies to A. israelii in sera from nine patients with actinomycosis. All the sera from the patients formed at the time of diagnosis one or more precipitins in terms of the 10 reference precipitins. Up to five precipitins were found in single sera. Follow-up studies covering a period of one-half year after treatment showed a gradually decreased precipitin response in the course of time. In control sera from patients with newly diagnosed tuberculosis, nocardiosis, deep Candida infection, and aspergillosis, and in sera from healthy blood donors, no antibodies were detected with specificity for the reference antigens.
In 30 patients with peritonsillar abscesses, pus was obtained by aspiration and by taking a swab after incision; bacterial recovery was compared. Although processed in the laboratory within 2 h, swab speciments gave results comparable to syringe specimens in only 9 of 13 patients with beta-hemolytic streptococci and 7 of 25 patients with anaerobic bacteria. Both kinds of microorganisms were lost in some cases but appeared as additional flora in others. The poor results from the swab technique was ascribed to overgrowth of respiratory flora contaminating the sample after incision. In aspirated pus kept in the syringe, or transferred to anaerobic transporters, the microbial flora was unchanged for 24 to 48 h. Some anaerobes also survived on agar slants for 24 h, but specially designed anaerobic transporters are recommended.
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Fractionated fluorescein-isothiocyanate (FITC)-conjugated immunoglobulin G (dye-to-protein ratio <10), produced against whole cells of Actinomyces spp., Arachnia, Bacterionena, Rothia, and Propionibacterium spp., give species-specific conjugates with controlled nonspecific staining reactions when appropriately diluted on the basis of their antibody content (10 mg/ml). Using this standardization in immunofluorescence, serotype-specific conjugates are also available after dilution for all serotypes of these organisms except for Actinomyces viscosus type 2, and Propionibacterium acnes type 1. Adequately adsorbed conjugates could be used to differentiate these serotypes from A. viscosus type 1 and P. acnes type 2, respectively. A serological classification in defined immunofluorescence corresponded to species and serotype designation proposed on the basis of other serological analysis and biochemical characteristics. This includes a separation in immunofluorescence of two serotypes of Propionibacterium acnes. The detection of certain actinomycetes of the family Actinomycetaceae and Propionibacterium species by the defined immunofluorescence in direct smears prepared from clinical specimens agreed to 88% with parallel culturing when including a prereduced (PRAS) medium technique for isolation. Qualitative studies revealed that single cells of these organisms could be specifically identified by immunofluorescence when admixed with morphologically similar bacteria and a large number of other contaminants.
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