Strains ofH Research Articles

Studies on pathogenic and non-pathogenic small free-living amoebae and the bearing of nuclear division on the classification of the order amoebida.

A systematic study of small free-living amoebae has been made under standardized and reproducible cultural conditions, and their pathogenicity has been tested in mice.Naegleria aerobia, Hartmannella culbertsoniandH. rhysodesare pathogenic;H. castellanii, H. astronyxis, H. palestinensis, H. glebae, H. exundans, H. vermiformis, Schizopyrenus russelli, Didascalus thorntoniandTetramitus rostratusare non-pathogenic. Strains ofH. culbertsoniandH. rhysodesare present in Indian soils. A classification of the order Amoebida Kent into families Schizopyrenidae Singh, Hartmannellidae Volkonsky, 1931 emend. Singh, 1952, and Endamoebidae (Calkins), based on nuclear division, is proposed, and the relation of this classification to previously defined families and genera of amoebae and its bearing on phylogeny are discussed. Metronidazole and other anti-amoebic drugs are ineffective againstN. aerobiaandH. culbertsoni in vitroand in meningo-encephalitis in mice.

Selective culture medium to survey the incidence of Haemophilus species.

A culture medium for the selective isolation of Haemophilus species is described. Bacitracin and nutritional supplements were incorporated in a rich basal agar medium to which rabbit blood was added to distinguish hemolytic species. Colony counts of seven typed strains of H. influenzae on this medium were within practical limits of counts on other media tested for clinical use. The bacitracin medium was as reliable as hemoglobin-agar for detecting H. influenzae and more sensitive for detecting other Haemophilus species in a clinical survey with the advantage of selectivity.

Antibodies in histoplasmosis.

Production of precipitating and complement-fixing antibody in rabbits and other animals was induced by immunization with live yeast-phase cells of Histoplasma capsulatum. Results of studies of polysaccharide antigens from three strains of H. capsulatum, by quantitative complement-fixation with human and rabbit antisera, strongly suggest the presence of type specificity. The variations of titer during 11 weeks in one patient with histoplasmosis and the variations of titer among a group of patients with histoplasmosis were studied by use of quantitative complement-fixation tests.

Occurrence of serotypes among Histoplasma capsulatum strains.

Kaufman, Leo (Communicable Disease Center, Atlanta, Ga.), and Sharon Blumer. Occurrence of serotypes among Histoplasma capsulatum strains. J. Bacteriol. 91:1434-1439. 1966.-Extensive studies with a fluorescent antibody specific for the yeast form of Histoplasma capsulatum indicated that, although most strains of H. capsulatum stained intensely, others stained weakly and some did not stain. These observations suggested that antigenic variants of the fungus existed. To test this hypothesis, reciprocal cross-staining and adsorption procedures were performed with fluorescein-labeled antiglobulins to four strains of H. capsulatum. Fifty-six strains of H. capsulatum and 11 strains of H. duboisii were studied. The data indicate that at least five serotypes of H. capsulatum occur. One is closely related antigenically to H. duboisii and Blastomyces dermatitidis.

COMPARATIVE FLUORESCENT ANTIBODY STAINING OF HISTOPLASMA CAPSULATUM AND HISTOPLASMA DUBOISII WITH A SPECIFIC ANTI-YEAST PHASE H. CAPSULATUM CONJUGATE.

The specific anti-yeast phaseHistoplasma capsulatum conjugate has been tested against 13 yeast phase strains ofH. capsulatum and 9 ofH. duboisii. The conjugate was specific forH. capsulatum, no yeast phase form ofH. duboisii obtained in vitro or in vivo reacted with it. The taxonomic implications of these results are discussed.

ACTION OF HAEMOPHILUS CULTURES ON δ-AMINOLEVULINIC ACID

Biberstein, Ernst L. (University of California, Davis), Patricia D. Mini, and Marjorie G. Gills. Action of Haemophilus cultures on delta-aminolevulinic acid. J. Bacteriol. 86:814-819. 1963.-Utilization of delta-aminolevulinic acid (ALA) by strains of Haemophilus recovered from various sources was investigated. With the 37 cultures studied, there was perfect correlation between absence of hemin requirement and ability to convert ALA to porphyrin. A total of 29 strains, including representatives of H. parainfluenzae, H. parahaemolyticus, H. "parasuis," and H. gallinarum, fell into this group. The remaining eight isolates, which were incapable of porphyrin synthesis from ALA, were strains of H. influenzae, H. suis, and a Haemophilus culture of uncertain classification obtained from a splenic abscess in a deer. In the active preparations, the products of synthesis included a mixture of porphyrins, porphobilinogen (PBG), and a pigment which absorbed light strongly at 470 to 480 mmu. Paper chromatographic studies of fractions of supernatants and cells revealed the presence of porphyrins having R(F) values similar to those of uro- and protoporphyrins, as well as some intermediate rates of migration probably representing coproporphyrins. Porphyrins were found both intra- and extracellularly, and PBG and the pigment absorbing at 470 to 480 mmu were confined to the supernatant.

Oxidative metabolism of Haemophilus species grown at different levels of hemin supplementation.

Biberstein, Ernst L. (University of California, Davis) and Patricia D. Spencer. Oxidative metabolism of Haemophilus species grown at different levels of hemin supplementation. J. Bacteriol. 84:916-920. 1962.-Haemophilus strains were propagated on hemin-deficient and hemin-supplemented media and tested on various substrates for their ability to carry on oxidative metabolism. H. parainfluenzae and H. parahaemolyticus exhibited equally vigorousoxygen uptake under both methods of culture. Strains of H. influenzae showed substantially lower rates of oxygen uptake when cultivated in the absence of hemin than when grown in its presence. Some oxidative capacity remained, however, even when the organisms had been grown for several passages in the absence of hemin supplements.

The histamine-sensitizing property of Haemophilus pertussis.

Summary and ConclusionsMice and rats which are normally resistant to histamine become more susceptible to its lethal action after an injection ofH. pertussis. This so-called sensitization to histamine is not an anaphylactic phenomenon. It is due to the action of a component ofH. pertussis, the histamine-sensitizing factor (HSF), which in some unknown way overcomes the physiological mechanism in rats and mice that makes them more resistant than other species to histamine. Guinea-pigs which appear not to possess this mechanism and are about 200-fold more susceptible, weight for weight, than rats and mice, were not made more sensitive byH. pertussis.After a stated dose of vaccine, sensitization was detectable in 48 hr., reached a maximum in 3–4 days, remained at this level for about 2 weeks and gradually disappeared. The effects of dosage, route of injection, and weight and sex of mice have been examined.The HSF was found in strains ofH. pertussis; it was not found inH. parapertussis, H. bronchisepticusorH. influenzae. It was only slightly affected by heating at 70° C. for 1 hr. but was destroyed at 80° C. in ½ hr. It was destroyed when bacteria were disintegrated by shaking with glass beads, or by grinding after being freeze-dried. It was found in the supernatant fluid of a partially autolysed vaccine.HSF was antigenic. Antisera were prepared in rabbits. Anti-HSF combined with HSF and neutralized its histamine-sensitizing activity. Bacteria treated with antiserumin vitroabsorbed anti-HSF and did not thereafter sensitize mice.Antiserum protected mice passively against the sensitizing action of vaccine, presumably by combining with HSF. After sensitization had developed the sensitive state was not affected by antiserum.Although HSF is an antigen there was no indication that histamine-sensitization was due to its antigenicity.The HSF was differentiated from heat-labile and heat-stable toxin, haemagglutinin, capsular material and agglutinogen.The preparation V 17, which is a small fraction of the disintegrated bacteria, adsorbed on red cell stromata (Pillemeret al.1954) had a high histamine-sensitizing value. Compared with whole bacterial vaccine it caused little production of agglutinin in mice; in rabbits it caused a slower and smaller production of agglutinin and a faster and greater production of anti-HSF. For this reason immune rabbit sera may have a high agglutinin titre and a low anti-HSF value or vice versa. Anti-HSF rabbit serum protected mice against sensitization by V 17.Vaccines could be graded according to their histamine-sensitizing activity. This did not always correspond to their grading by agglutinin production in mice. The relation of HSF to the immunizing antigen ofH. pertussisand the use of histamine-sensitization to indicate the immunizing potency of pertussis vaccines are discussed.We wish to thank Glaxo Laboratories Ltd. for the supply of reference vaccine, and the Whooping Cough Immunization Committee of the Medical Research Council for vaccines and sera.

The type specificity of haemophilus pertussis.

1. One hundred and eighty-seven freshly isolated strains ofHaemophilus pertussis, together with ten laboratory strains and eleven strains ofH. parapertussiswere submitted to tests of their morphological, biochemical and antigenic properties in relation to their virulence to mice.2. The naturally occurringH. pertussisstrains form a single antigenic type differing from that of the attenuated laboratory strains.3.H. parapertussisforms, as previously claimed, a distinctive type with characteristic properties closely related toH. pertussis.Our thanks are due to Col. H. J. Bensted, Prof. J. W. McLeod and Drs A. Beck, D. G. Evans, G. Eldering, J. Marks, M. T. Parker, M. Pittman and H. Proom for supplying strains used in this investigation.