David H. Smith

Human Serum Activities against Hemophilus influenzae, Type b

Humoral immunity to Hemophilus influenzae, type b was studied in normal human adults by means of assays for serum bactericidal and opsonizing activities against the organism and for passive hemagglutinating activity using erythrocytes sensitized with polyribophosphate, the type-specific capsular antigen. Hemagglutinating activity was detectable in about 60% of the 114 sera tested. Serum bactericidal and opsonizing activities were found in all sera tested; the levels in some sera, however, were quite low. The antibacterial activities were due not only to antibodies directed against the polyribophosphate capsule but also to antibodies that appear to be directed against somatic antigens. Type b strains differed in their susceptibility to the antisomatic antibodies of particular sera but were uniformly sensitive to anticapsular antibody.

R Factors Mediate Resistance to Mercury, Nickel, and Cobalt

Fifty-five clinical isolates and laboratory stocks of Escherichia coli and Salmonella were studied for resistance to each of ten metals. Eleven clinical isolates carrying R factors were resistant to mercury, and, in each case, the resistance was mediated by a previously undefined R-factor gene. The gene was phenotypically expressed within 2 to 4 minutes after entry into sensitive bacteria, but the basis for the resistance remains undefined. Fourteen strains, 12 infected with R factors, were resistant to cobalt and nickel, but these resistances were mediated by R-factor genes in only two strains; separate R-factor genes mediated the resistances to nickel and cobalt. These and other results indicate that the genetic composition of R factors is greater than that originally defined.

Circulating polyribophosphate in Hemophilus influenzae, type b meningitis. Correlation with clinical course and antibody response.

In systemic infections caused by Hemophilus influenzae, type b, the capsular polysaccharide, polyribophosphate, is released into the circulation. Polyribophosphate was quantitated in serial serum and cerebrospinal fluid samples from 45 children with H. influenzae, type b meningitis by means of a radiolabeled antigen-binding inhibition assay. Polyribophosphate was regularly found in acute serum and cerebrospinal fluid samples and could be detected in unbound form for periods of 1-30 days after initiation of effective therapy. Complexes of polyribophosphate dissociable with acid and pepsin were detected in serum samples from 17 patients, in one case for a period of 145 days after hospitalization. Polyribophosphate levels and patterns of clearance were studied in relation to hospital course and antibody response. Patients with prolonged antigenemia had protracted fevers and severe neurological symptoms during hospitalization, frequently with focal complications.Antipolyribophosphate antibody responses were detected during the first 100 days of convalescence by radioimmunoassay in 79% of the patients studied, including 60% of the children 1 yr or less in age. The intensity of antibody response although clearly related to the age of the patient, was more reliably predicted by the efficiency of antigen clearance. Antibody responses were uniformly of low magnitude in patients with prolonged antigenemia, irrespective of age. Paients who failed to develop antibody to polyribophosphate after meningitis also exhibited impaired antigen clearance. These studies suggest that mechanisms necessary for clearance of polyribophosphate may influence the development and intensity of the humoral immune response and raise the possibility of developmental deficiencies in the clearance system in infants and children.

Impaired Humoral Immunity in Treated Hodgkin's Disease

To define the contribution of aggressive lymphoma treatment to the risk of post-splenectomy septicemia, we investigated the humoral immunity of 44 patients with Hodgkins disease. Specific antibody against Haemophilus influenzae Type b was significantly reduced (mean, 147 ng per milliliter, P less than 0.01) in patients receiving combined treatment (radiotherapy and chemotherapy), whereas single treatment reduced titers marginally (chemotherapy) or not at all (radiotherapy). Untreated patients had normal values (396 ng per milliliter), and splenectomy was without effect. In some patients who received combined treatment, titers were reduced to levels seen in infants. IgM levels were likewise normal in untreated patients. Chemotherapy, however, significantly reduced IgM levels (P less than 0.025), an effect potentiated by prior splenectomy. IgG, IgA, alternate-pathway activity, C3, C4 and CH50 were all normal or elevated. Aggressive treatment with chemotherapy and radiation impairs humoral defense against encapsulated micro-organisms, and thus magnifies the risk of post-splenectomy septicemia in patients with Hodgkins disease.

Immunization of Humans with Polyribophosphate, the Capsular Antigen of Hemophilus influenzae, Type b

In human volunteers, single injections of purified polyribophosphate elicited antibodies detectable by passive hemagglutination and by serum bactericidal and opsonizing activities against viable Hemophilus influenzae, type b. All three activities rose by 2 wk to maximal levels, at which they remained for at least 6 months. Doses of 1 mug elicited antibody responses in nearly all recipients; higher doses of the antigen, however, produced larger increases in titer. Booster doses of 1 mug given at 6 months did not further increase the antibody titers. A tuberculin-like response was often observed at the site of injections given intradermally.

The paradox of Hemophilus infuenzae type B bacteremia in the presence of serum bactericidal activity.

We investigated the role of serum bactericidal activity in Hemophiplus influenzae type b infections in infants with meningitis and in a rat model. In infected infants, 13/22 admission sera had bactericidal activity against the infecting strain, and bacteremia was as frequent in those with bactericidal activity (54%) as those without (56%). The coexistence of bactericidal activity and bacteremia was reproduced and studied in experimentally infected weanling rats. Serum from such rats kills in vitro 95% of conventionally broth-grown bacteria within 10 min, but does not kill organisms obtained from the infected animals. Thus bactericidal activity as conventionally determined for H. influenzae b may have no relevance in vivo, Incubation of broth-grown bacteria in normal rat serum for 30 min at 37 degrees C produces a resistance like that of in vivo organisms. This phenotypic conversion depends on factors that are of molecular weight less than 1,000, stable to 100 degrees C, but destroyed by ashing. When injected intravenously into nonimmune animals, broth-grown bacteria are quickly cleared, while serum-preincubated bacteria are not. The latter, however, are cleared when injected into bacteremic rats (half-life 30 min). Bacteremia in the rats may persist despite this capacity for clearance because bacteria are entering the blood from extravascular fluids, which contain greater than 90% of the total bacterial burden.

Catabolite repression of chloramphenicol acetyl transferase synthesis in E. coli K12

Abstract The specific activity of chloramphenicol acetyl transferase (CAT) in R + E. coli K 12 . strains grown on glucose was 3–5 fold lower than when grown on glycerol. This decrease was eliminated completely when the cells were grown in the presence of cyclic 3′5′ AMP but not 5′ AMP. When an adenyl cyclase-defective mutant and its parent strain, both harboring the R factor, were grown on glucose plus 5′ AMP, the level of CAT was 3–4 fold lower in the mutant than in the parent. Replacing 5′ AMP with cyclic AMP increased the CAT level 20–30 fold.

Acquisition of type-specific antibodies to Hemophilus influenzae type b.

Seven of eight 2- to 3-year-old healthy children associated with a child with H. influenzae type b meningitis in a day care center had or developed type-specific antibody activity in high titer. These antibody responses were similar to those seen following invasive H. influenzae type b diseases in children of the same age but differed in titer and prevalence from those of children in the “general population.” The nature of the antigenic experience in the day care center children producing the antibodies to the capsule of H. influenzae type b appeared to be mild or asymptomatic infection, possibly independent of prolonged nasopharyngeal carriage.

Inhibition of nucleic acid synthesis by novobiocin

Abstract In E. coli novobiocin immediately inhibits DNA synthesis and to a lesser extent RNA synthesis, while inhibition of protein synthesis and growth, and increased membrane permeability, appear considerably later. The drug is not bound detectably to DNA, and therefore may act directly on the polymerase system.

Unclassified mycobacterial infection and disease in children residing in Massachusetts

Twenty-eight children with lymphadenitis of mycobacterial origin were studied and tested with PPD-B (Battey) and PPD-S (Standard). Eleven were seen in 1963, the others from 1956 to 1962. Unclassified mycobacteria were most likely the etiological agents in 22 children, including 6 with proved cultures. Complete excision was curative, but other surgical procedures were not. The effect of antituberculous drugs could not be defined. Twenty of 135 school children in Cambridge, Massachusetts, with a history of sensitivity to PPD-S had significantly greater reactions to PPD-B than to PPD-S. This number included 23 per cent of the children given antituberculous chemoprophylaxis because of recent tuberculin conversion. Most of the “Battey reactors” in both groups were lifelong residents of Massachusetts, only a few having been in areas previously described as endemic for unclassified mycobacteria.