J. B. Hertz

Taxonomic Application of Crossed Immunoelectrophoresis

We used crossed immunoelectrophoresis, crossed-line immunoelectrophoresis, tandem-crossed immunoelectrophoresis, and crossed immunoelectrophoresis with an intermediate gel (containing antibodies) in studies of bacterial taxonomy. By these quantitative immunoelectrophoretic methods, as many as 90 antigens from a bacterial species can easily and reproducibly be analyzed for immunological cross-reactivity with antigens from other bacterial species without prior purification of antigens. The immunological cross-reactivity between various bacterial species is expressed as the matching coefficient (MC), i.e., the ratio between the number of cross-reactive antigens with another bacterial species and the total number of antigens of the reference bacterial species. The MC has been found to be significantly correlated with phenotypic similarities (biochemical characteristics) and nucleic acid hybridization results obtained with the taxa investigated so far. In general, closely related strains (serotypes within a species) have a high MC (≥0.90), while different species of the same genus have a lower MC (0.90 to 0.70 or lower in some cases); genera within a family have an intermediate MC (0.70 to 0.15), and distantly related taxa have a very low MC (≤0.15).

Studies on Hypersensitivity to Bacterial Antigens in Intrinsic Asthma

Twelve children, aged 4 to 14 years, with moderate to severe intrinsic asthma (IA) were studied. Symptom‐score charts were used to confirm the relationship of acute respiratory tract infections to exacerbations of asthma. Hypersensitivity to eight commonly occurring bacteria from the normal flora of the upper respiratory tract was studied by skin test, by crossed immunoelectrophoresis, and by basophil histamine release in vitro, using ultrasonicates of the bacteria as antigens. Skin tests were all negative. All children contained low titers of precipitating antibodies against most of the bacteria, but in this respect they did not differ from normal children. In contrast. release of histamine was induced in leukocytes from the IA children by all, or most sonicates, while such reactions, were less frequent in control children. The pattern of responses indicated an element of specificity. There was no correlation to precipitating antibodies, or to the microbial flora of the children. Positive responses were characterized by low values of maximal histamine release, and by a tendency to fluctuations with time. Because of these fluctuations, and because the IA children and control children were tested on separate occasions, we cannot be certain as to the real difference between these two groups. Our studies do, however, demonstrate that water‐soluble constituents of all real bacterial strains tested were capable of causing the release of histamine in vitro, but that this phenomenon is not restricted to IA. The clinical significance of these findings awaits further investigations on the mechanism(s) of release in vitro by such agents.

An antigen common to a wide range of bacteria. 2. A biochemical study of a "common antigen" from Pseudomonas aeruginosa.

Common Antigen (CA) of Pseudomonas aeruginosa has been shown to be a protein composed of polypeptide subunits of a molecular weight (MW) of about 62 000. The MW of this protein was estimated to 665 000 by gel filtration on sepharose CL-6B, to 800 000 by electrophoresis on polyacrylamide gradient gels and to about 900 000 by ultracentrifugation, on a sucrose gradient. By analytical ultracentrifugation with Schlieren optics a sedimentation coefficient (S20 degrees, W) of 22.65 was calculated. The isoelectrical point was determined to pH 4.4. The antigen was decomposed on exposure to proteolytic enzymes. Polysaccharide, lipid, deoxyribonucleic acid or ribonucleic acid were not demonstrated in CA. The amino acid content of CA was determined, and no hexosamine or abnormal residues were observed. The amino acid content of CA was determined, and no hexosamine or abnormal residues were observed. The antigen was degraded when heated to 100 degrees C for 4 min or when exposed to pH below 4 or above 11 at 4 degree C. CA has been isolated from the cytoplasmic water-soluble fraction of disintegrated bacteria and only trace-amounts could be obtained from envelope fractions after solubilization with Triton X-100.

PRECIPITATING ANTIBODIES AGAINST ESCHERICHIA COLI, BACTEROIDES FRAGILIS SS. THETAIOTAOMICRON AND PSEUDOMONAS AERUGINOSA IN SERUM FROM NORMAL PERSONS AND CYSTIC FIBROSIS PATIENTS, DETERMINED BY MEANS OF CROSSED IMMUNOELECTROPHORESIS

Abstract. Serum from normal persons and from 133 cystic fibrosis patients was examined for precipitins against faecal bacteria (E. coli and B. fragilis) and against P. aeruginosa, by means of crossed immunoelectrophoresis. 27% of the normal sera contained 1–2 precipitins against E. coli, 16% contained one precipitin against B. fragilis and 6% contained one precipitin against P. aeruginosa. In sera from cystic fibrosis patients, there was a significantly increased prevalence of precipitins against E. coli (76%), B. fragilis (38%) and P. aeruginosa (63%), and the mean number of precipitins against each of these bacteria was 3 (range: 1–12), 1.4 (range: 1–4), and 16 (range: 1–60), respectively. Increased numbers of precipitins against P. aeruginosa were correlated with chronic lung infection caused by this species and with poor prognosis, whereas such associations were not revealed as regards precipitins against the two other bacterial species. Increased numbers of precipitins against E. coli and B. fragilis were significantly associated with increased numbers of precipitins against P. aeruginosa. It is suggested that this association is due to immune reactions in the intestinal mucosa, resulting in increased absorption of antigens from the gut.