bluuebunny/crossref_metadata_2025 huggingface dataset:
Lysozyme-mediated aggregation and lysis of the periodontal microorganism Capnocytophaga gingivalis 2010
Authors: Iacono V J , Zove S M , Grossbard B L , Pollock J J , Fine D H , Greene L S
Published: January 2020

The ability of lysozyme to aggregate and lyse the gram-negative capnophilic periodontal microorganism Capnocytophaga gingivalis 2010 was monitored optically at 540 nm. Both hen egg white and chromatographically purified human lysozymes had significant but similar aggregation potentials for both logarithmic- and stationary-phase bacteria. In general, an increase in enzyme concentration resulted in a graded increase in both the initial and maximum changes in turbidity which occurred during the reaction period. The greatest change in turbidity occurred within the initial minutes of interaction of lysozyme and the cells, and the extent of aggregation paralleled a rapid depletion of lysozyme by the suspensions during the first minute of its incubation with the bacteria. Interestingly, the muramidase inhibitors N-acetyl-D-glucosamine and histamine did not block aggregation, whereas maleylation of lysozyme completely inhibited its aggregating ability. Demaleylation, however, restored aggregation activity comparable to the native enzyme, indicating that maleylated lysozyme retained its integrity and that aggregation was primarily dependent on charge. The addition of up to physiological concentrations of NaHCO3 and NaCl to cell aggregates resulted in varying degrees of deaggregation and lysis. Surprisingly, ultrastructural analysis of lysozyme-treated cells revealed morphological changes with or without the addition of salt. Damage appeared to occur at the blunted polar end of the cells where there was a large spherical outpouching bordered by a damaged cell envelope. Damaged cells uniformly contained dense granular cytoplasmic debris. In effect, the cationic enzyme lysed C. gingivalis 2010, which was not apparent in the spectrophotometric assay. The paradoxical finding that during bacterial aggregation there was lysis may be of significance to the further elucidation of lysozyme's antibacterial role in the gingival sulcus.
Role of tetracycline in pathogenesis of chronic candidiasis of rat tongues
Authors: Allen C M , Beck F M , Lurie F A , Pinsky H M
Published: January 2020

Forty female Sprague-Dawley rats were randomly divided into two groups. Group 1 animals received tetracycline solution (0.1% during week 1 and 0.01% thereafter) as drinking water, as in previous studies. Group 2 animals received double-distilled demineralized water. Animals in both groups were inoculated orally with an equal number of viable, mucosally pathogenic Candida albicans organisms. After 20 weeks, inspection of the tongues showed gross lesions in 16 of the 20 animals in group 1 and 17 of the 20 in group 2. These lesions were confirmed histologically. No significant difference in the number of lesions was noted between the two groups. However, the lesions in group 1 animals were significantly larger than those in group 2 animals. These results suggest that, given this mucosally pathogenic strain of C. albicans, the establishment of a chronic infection of the rat tongue is apparently not influenced by tetracycline-treated drinking water as administered in this study. This suggests that antibiotic exposure is not an essential factor in the pathogenesis of this lesion. A larger lesion area, however, does appear to be related to the use of tetracycline solution as drinking water.
Cellular and humoral immune responses of mice subclinically infected with Pneumocystis carinii
Authors: Furuta T , Ueda K , Fujiwara K , Yamanouchi K
Published: January 2020

Cellular and humoral immune responses to Pneumocystis carinii were investigated. ICR and DDD mice were intranasally infected with 10(4) mouse lung-derived P. carinii, and the delayed-type hypersensitivity (DTH) reaction and antibody titers to P. carinii were measured along with the number of P. carinii cysts in the lungs after infection. The number of P. carinii cysts in the lungs peaked at 2 weeks after infection and then decreased to barely detectable levels by 4 weeks. Serum antibody (immunoglobulin G) titers measured by indirect immunofluorescence increased up to 4 weeks. The DTH footpad reaction was most prominent at 2 weeks postinfection and declined thereafter. Thus, the decline in the number of P. carinii cysts in the lungs corresponded well with the time of the peak of the DTH reaction but not with the serum antibody response. Spleen T cells from infected mice mediated the DTH reaction when transferred intravenously into normal recipients and reduced the number of P. carinii cysts in the lungs when transferred intravenously into P. carinii-infected mice. The results indicated that cellular immunity is important for protection from subclinical P. carinii infections.
Antigenic variation in clones of Trypanosoma brucei grown in immune-deficient mice
Authors: Myler P J , Allen A L , Agabian N , Stuart K
Published: January 2020

We have produced monoclonal antibodies against six variant surface glycoproteins from early variant antigen types (VATs) of the IsTaR 1 serodeme of Trypanosoma brucei brucei. We have used these in fixed cell immunofluorescence assays to follow the VAT composition of populations of each early VAT when passaged through irradiated mice. The IsTat 1.A and 1.7a populations were stable for more than 30 days (approximately 150 generations), but 1.1a, 1.3a, 1.5a, and 1.11a all changed to 1.A within this time. The time and rate of this antigenic switch were characteristic for each VAT. Growth rates of the VATs were determined when they were both grown separately and grown with 1.A. It appeared that the order of growth rates was 1.7a greater than 1.A = 1.1a greater than 1.11a greater than 1.5a greater than 1.3a. We have generated theoretical curves for the replacement of one VAT by another based on differences in their growth rates and the rate at which one VAT switches to another (switch frequency). These curves closely match those derived experimentally. We postulate that the differences in growth rates between VATs and the different switch frequencies for VATs may be sufficient to generate the loosely defined sequence of VATs seen in chronic infections.
Loss of virulence in a protease-deficient mutant of Aeromonas salmonicida
Authors: Sakai D K
Published: January 2020

The importance of extracellular protease production by Aeromonas salmonicida, the bacterial pathogen of fish furunculosis, was investigated with four virulent strains (which were autoagglutinative, hemagglutinative, resistant to fish serum, adhesive to fish tissue culture, protease positive, hemolysin positive, and leukocytolysin positive) and three avirulent strains (which were nonagglutinative, nonhemagglutinative, sensitive to serum, nonadhesive, protease positive, hemolysin positive, and leukocytolysin positive). A protease-deficient mutant (NTG-1) was induced by mutagenesis with N-methyl-N'-nitro-N-nitrosoguanidine from virulent strain A-7301, showing protease production, which was common to the two strain groups. Strain NTG-1 showed loss of virulence in determinations of 50% lethal doses, although it remained autoagglutinative, hemagglutinative, serum resistant, adhesive, hemolysin positive, and leukocytolysin positive. A protease fraction separated from extracellular products of strain A-7301 by DEAE-cellulose column chromatography had the capacity to produce skin lesions (furuncles) and high mortality in sockeye salmon. A comparable protein fraction from extracellular products of NTG-1 resulted in no protease activity and no pathological effects on the fish. The avirulent strains were eliminated from rainbow trout in a short time, whereas the virulent strains (including A-7301) were highly infective and proliferated in hosts. NTG-1 preserved its infectivity, but fish showed no signs of disease and no mortality. These findings indicate that extracellular protease is a major virulence factor and that protease production in the host is closely implicated in the pathogenesis of fish furunculosis.
Epidemiology of virulence-associated plasmids and outer membrane protein patterns within seven common Salmonella serotypes
Authors: Helmuth R , Stephan R , Bunge C , Hoog B , Steinbeck A , Bulling E
Published: January 2020

Antibiotic-sensitive Salmonella isolates belonging to seven common serotypes and originating from 29 different countries from all continents were investigated for their plasmid DNA content (337 isolates) and their outer membrane protein profiles (216 isolates). Of the S. typhimurium, S. enteritidis, S. dublin, and S. choleraesuis isolates, 90% or more carried a serotype-specific plasmid. The molecular sizes of the plasmids were 60 megadaltons (Md) for S. typhimurium, 37 Md for S. enteritidis, 56 Md for S. dublin, and 30 Md for S. choleraesuis. The outer membrane protein profiles were homogeneous within each of the seven serotypes, except that a minority of S. enteritidis and S. dublin strains were lacking one major outer membrane protein. Virulence studies were performed with 39 representative strains by measuring the 50% lethal doses (LD50S) after oral infection of mice. The LD50 values obtained for plasmid-positive strains of S. typhimurium, S. enteritidis, and S. dublin were up to 10(6)-fold lower than the values obtained for the plasmid-free strains of the same serotype. Only the plasmid-positive strains could invade the livers of orally infected mice, and only they were resistant to the bactericidal activity of 90% guinea pig serum. Strains of S. infantis were generally plasmid free, whereas S. panama and S. heidelberg isolates carried heterogeneous plasmid populations. The virulence properties of the latter three serotypes could not be correlated with the predominant plasmids found in these strains.
Dissipation of the proton motive force in oral streptococci by fluoride
Authors: Kashket S , Kashket E R
Published: January 2020

Strains of oral streptococci maintain an intracellular pH (pHi) that is more alkaline than the external pH (pHo). The delta pH (pHi-pHo) was about 0.6 in neutral media and about 0.9 in media of pH 5.5. Addition of 10 micrograms of F- per ml at pH 7.0 reduced the delta pH in Streptococcus salivarius to 0.17. The reduction of delta pH in S. sanguis H7PR3 was less pronounced, whereas the delta pH in S. mutans IB 1600 was unaffected. The F- -resistant mutant of S. salivarius, strain Flr103, maintained a delta pH of 0.51 with 100 micrograms of F- per ml. Addition of F- to cells in media below pH 6.0 led to a reduction of delta pH in all strains. The anion had no effect on the transmembrane electrical gradient of either mutant or parental cells of S. salivarius at pH 7.0. The principal effect of F- addition at neutral pH, therefore, was on the delta pH component of the proton motive force. At pH 5.5, 100 micrograms of F- per ml reduced the transmembrane electrical gradient from 71 to 40 mV in the parent and from 80 to 42 mV in the mutant. We propose that the greater sensitivity of cells to F- at lower medium pH stems from the rapid dissipation of delta pH by the anion. Thus, pH equilibration in media of low pH would lead to a greater reduction of metabolic activity than when it occurs in media at neutral pH. The data also suggest that the growth of streptococci, with or without added F-, is limited when the intracellular pH falls below about 5.7.
Cloning and genetic analysis of serotype 5 M protein determinant of group A streptococci: evidence for multiple copies of the M5 determinant in the Streptococcus pyogenes genome
Authors: Kehoe M A , Poirier T P , Beachey E H , Timmis K N
Published: January 2020

A gene bank of group A Streptococcus strain Manfredo (M protein serotype 5) was constructed with a bacteriophage lambda vector-Escherichia coli K-12 host system and screened by immunoblotting hybrid phage plaques with antisera raised to purified pep M5 (serotype 5 M protein fragment released from the streptococcal cell surface by pepsin). Hybrid phage expressing M5 antigen (lambda M5) were detected in the gene bank at an unexpectedly high frequency. The cloned streptococcal DNA sequences from one lambda M5 phage were subcloned into an E. coli plasmid vector. The M5 gene (smp5) was mapped, and its transcriptional orientation was determined by isolating and characterizing deletion and transposon insertion mutants of the M5+ hybrid plasmid pMK207. This analysis indicated that the intact smp5 gene had been cloned. Anti-pep M5 sera reacted with five pMK207-encoded polypeptides having relative molecular sizes of 64,000, 56,000, 55,500, 52,500, and 50,000. All of these polypeptides were encoded by the same DNA sequences, and all reacted with antisera raised to a synthetic peptide corresponding to the amino-terminal end of pep M5, suggesting that proteolytic cleavage at the carboxy-terminal end of the smp5 gene product generates at least some of the lower relative molecular size forms. Southern blotting experiments with smp5 gene sequences as probes identified multiple copies of DNA sequences sharing homology with the smp5 gene in the type 5 group A streptococcal genome.
Recognition of serogroup A Neisseria meningitidis serotype antigens by human antisera
Authors: Sugasawara R J
Published: January 2020

The antigens of Neisseria meningitidis serogroup A which were recognized by human antisera were identified by Western blot and enzyme-linked immunosorbent assay techniques. The components of six prototype strains used for serotyping serogroup A meningococci were resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and then transferred to nitrocellulose for immunoperoxidase staining with sera collected from 10 acute-phase and 14 convalescent-phase patients. Six acute-phase sera detected six major antigens having apparent molecular weights between 14,000 and 82,000. In addition to recognizing these antigens, the convalescent-phase sera detected a protease-sensitive antigen with an apparent molecular weight of 20,000 for one strain and 27,000 for five strains, lipopolysaccharide, and the heat-modifiable proteins. The sera recognized lipopolysaccharide in a serotype-specific manner, whereas their reactions with the heat-modifiable protein were not serotype specific. Convalescent-phase sera recognized components from eight meningococcal serogroups. The concentrations of immunoglobulin G directed to capsular polysaccharide were determined by the enzyme-linked immunosorbent assay; seven acute-phase sera had less than 0.39 micrograms of antibody per ml, whereas the average concentration in convalescent-phase sera was 3.22 micrograms/ml and the range was 0.40 to 7.50 micrograms/ml.
In vivo evidence that bacteria in urinary tract infection grow under iron-restricted conditions
Authors: Shand G H , Anwar H , Kadurugamuwa J , Brown M R , Silverman S H , Melling J
Published: January 2020

The outer membrane protein composition of bacteria isolated directly and without subculturing from the urine of two patients with urinary tract infections was investigated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The results indicated that the bacteria grew under iron-restricted conditions, as revealed by the expression of several high-molecular-weight outer membrane proteins which could also be observed when the same isolates were grown under iron-depleted conditions in laboratory media. The antigenicity of outer membrane components of the bacteria isolated was studied by immunoblotting with serum samples from the patients. The results indicated that the sera from the patients contained antibodies against major outer membrane components of the bacteria present in the urine, including the iron-regulated membrane proteins.
Comparison of the capacity of two lipid A precursor molecules to express the local Shwartzman phenomenon
Authors: Galanos C , Hansen-Hagge T , Lehmann V , Lüderitz O
Published: January 2020

It has been shown recently that a Salmonella lipid A precursor molecule (Ia) and its synthetic counterpart are inactive in expressing the local Shwartzman reaction in both homologous and heterologous systems in combination with lipid A. Precursor Ia contains a bisphosphoryl-beta-1,6-glucosamine disaccharide substituted by 4 mol of (D)-3-hydroxytetradecanoyl residues. Escherichia coli lipid A, on the other hand, which contains two additional non-hydroxylated acyl residues in the form of two 3-acyloxyacyl units, is highly active. We have recently isolated a lipid A precursor molecule (Ib) with the same basic structure as precursor Ia, which contains, however, one additional non-hydroxylated (hexadecanoic) fatty acid forming one 3-acyloxyacyl residue. A comparison of precursor Ia and Ib in homologous and cross-reacting Shwartzman systems confirmed that precursor Ia completely lacked the capacity to prepare the skin for, or to elicit, the Shwartzman reaction. In contrast, precursor Ib was strongly active in inducing the local Shwartzman reaction both when administered intradermally as a preparatory agent and when administered intravenously as a provocatory agent. The results indicate that the additional presence of at least one fatty acid either as such or as an acyloxyacyl residue (as in precursor Ib) is a prerequisite for the ability of the molecule to induce the local Shwartzman phenomenon.
Succinic acid, a metabolic by-product of Bacteroides species, inhibits polymorphonuclear leukocyte function
Authors: Rotstein O D , Pruett T L , Fiegel V D , Nelson R D , Simmons R L
Published: January 2020

Anaerobes, in particular Bacteroides spp., are the predominant bacteria present in mixed intra-abdominal infections, yet their critical importance in the pathogenicity of these infections is not clearly defined. Succinic acid, a major fatty acid by-product of Bacteroides metabolism, was tested for its effect on neutrophil function to determine whether it might play a role in enhancing the virulence of Bacteroides-containing infections. At pH 5.5 but not pH 7.0, succinic acid at concentrations commonly found in clinical abscesses profoundly inhibits in vitro neutrophil function. It virtually obliterates phagocytic killing of Escherichia coli and reduces neutrophil random migration and chemotactic response to formyl-methionyl-leucyl-phenylalanine and C5a. These effects occur in conjunction with a reduced chemiluminescent peak and delayed time to the peak. The effect on neutrophils is only partially reversible by multiple washings. These findings suggest that succinic acid may be an important Bacteroides virulence factor when present in the microenvironment of a mixed intra-abdominal infection in which concentrations are high and the pH of the medium is reduced.
Recombination-deficient Streptococcus sanguis
Authors: Daneo-Moore L , Volpe A
Published: January 2020

A UV-sensitive derivative was obtained from Streptococcus sanguis Challis. The organism could be transformed with a number of small streptococcal plasmids at frequencies equal to, or 1 logarithm below, the transformation frequencies for the parent organism. However, transformation with chromosomal DNA was greatly impaired in the UV-sensitive derivative.
Antigenic mimicry of mammalian intermediate filaments by mycoplasmas
Authors: Wise K S , Watson R K
Published: January 2020

A murine monoclonal antibody raised against Mycoplasma hyorhinis specifically reacted by indirect immunofluorescence with mammalian intermediate filaments. The antibody recognized a related epitope on a 74,000-molecular-weight protein of M. hyorhinis and on components of similar size from other pathogenic mycoplasmas. This defines a shared antigenic structure of interest in autoantibody development during mycoplasmal diseases.
Inhibitory effect of estradiol-17 beta and progesterone on bactericidal activity in uteri of rabbits infected with Escherichia coli
Authors: Matsuda H , Okuda K , Fukui K , Kamata Y
Published: January 2020

The influence of ovarian hormones at different estrous stages on the bactericidal activity of the uterus in rabbits was investigated. When Escherichia coli cells were inoculated in ligated uteri, the survival period of the bacteria in the uterus at the luteal phase was clearly longer than that at the follicular phase. At the luteal phase, high levels of plasma estradiol-17 beta and progesterone were detected. A luteolytic treatment with prostaglandin F2 alpha and human chorionic gonadotropin at the luteal phase lowered plasma progesterone levels and prompted bacterial clearance from the uterus. In ovariectomized rabbits, E. coli from the uterine exudates was not detected 6 days after the inoculation in both the nontreated and estradiol-17 beta-treated animals. In the progesterone-treated rabbits, the survival period of E. coli was longer than that in the nontreated and estradiol-17 beta-treated animals. When estradiol-17 beta and progesterone at the ratio of 1:100 were administered concurrently, E. coli survived for the longest period in the rabbits treated with various doses of different hormones. Formalin-killed E. coli cells were inoculated into the uterine lumen, and 4 h later the proportion of heterophils phagocytizing the bacteria dropped in the progesterone-treated rabbits and in the estradiol-17 beta- and progesterone-treated rabbits, but there was no significant difference in heterophil numbers among the rabbits treated with different hormones. The present results suggest that progesterone inhibits the bactericidal activity of the uterus and that estrogen concurrently secreted at the luteal phase promotes the inhibitory action of progesterone, although estrogen alone hardly affects the uterine defense. In addition, the lowering of the bactericidal activity of the uterus at the luteal phase may be attributable to lower activity of phagocytosis by heterophils infiltrated into the uterine lumen.
Immune response to plasmid- and chromosome-encoded Yersinia antigens
Authors: Mazza G , Karu A E , Kingsbury D T
Published: January 2020

The immune response of humans and mice to temperature-specific, plasmid- or chromosome-encoded proteins of yersinia pestis and Yersinia enterocolitica was investigated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting. Extracts from Y. pestis and Y. enterocolitica strains with and without the virulence plasmids pYV019 and pYV8081, respectively, were resolved by denaturing electrophoresis, and the major antigens were visualized with sera from convalescing plague patients, individuals immunized with plague vaccine, and mice and rabbits immunized with avirulent live yersiniae. The Y. pestis grown in vitro in this study did not express detectable amounts of plasmid-encoded antigens. The sera from plague patients recognized Y. pestis and Y. enterocolitica antigens ranging from 15 to 72 kilodaltons (kDa), whereas sera from immunized subjects recognized four antigenic components in Y. pestis ranging from 17 to 64 kDa and five antigens in Y. enterocolitica ranging from 16 to 68 kDa. Sera from mice reacted with 7 antigens in Y. pestis and 12 antigens in Y. enterocolitica ranging from 14 to 68 kDa, and sera from rabbits reacted with 7 and 10 antigens in Y. pestis and Y. enterocolitica, respectively. All of the plague patient sera, as well as the sera from immunized mice and rabbits, reacted with a 22-kDa Y. enterocolitica plasmid-associated polypeptide, and five of the patient sera also recognized a Y. enterocolitica plasmid-associated 31-kDa protein. The results indicate a common immune response to at least these two plasmid-specified Yersinia outer membrane proteins. Y. pestis apparently expresses these components only in vivo, and in vitro, Y. enterocolitica expresses a greater number of plasmid-associated antigens than does Y. pestis.
Effects of siderophores on the growth of Pseudomonas aeruginosa in human serum and transferrin
Authors: Ankenbauer R , Sriyosachati S , Cox C D
Published: January 2020

A combination of the siderophores produced by Pseudomonas aeruginosa, pyochelin and pyoverdin, dramatically stimulates the growth of this bacterium in medium containing human transferrin. The amount of growth stimulation observed when each siderophore was added alone was only slightly less than the amount observed with the combination. Siderophore-defective mutants of strain PAO1 were isolated to test the effects of siderophore production on growth in transferrin and human serum. The pyoverdin-proficient (Pvd+), pyochelin-deficient (Pch-) strain (IA5) grows just as well as the parent (PAO1), which produces both siderophores. On the other hand, the Pvd- Pch+ strain (211-5) has severely retarded growth, similar to that demonstrated by a mutant lacking production of both siderophores (IA1), but has an accelerated log phase compared with strain IA1 at the later stages of the growth curve. However, the Pvd- Pch+ strain (211-5) had no observable advantage over the Pvd- Pch- strain, IA1, during incubation in human serum. The inability of P. aeruginosa strains to produce pyochelin in glucose-minimal medium may explain the poor growth of 211-5 in this medium and in human serum. The 211-5 strain grows much better than the IA1 strain in the medium that allows pyochelin synthesis, but it still does not grow as well as the Pvd+ Pch- strain (IA5). Therefore, pyoverdin appears to be the most important siderophore for growth in human serum.
Enhancing effects of gamma interferon on phagocytic cell association with and killing of Trypanosoma cruzi
Authors: Wirth J J , Kierszenbaum F , Sonnenfeld G , Zlotnik A
Published: January 2020

The effects of gamma interferon (IFN-gamma) on P388D1 cell or mouse resident peritoneal macrophage association (i.e., binding and internalization) with the protozoan Trypanosoma cruzi were studied, as well as the effects of this lymphokine on intracellular parasite killing. Incubation of either type of cell with a conditioned medium containing IFN-gamma and traces of interleukin 2 markedly increased the capacities of the cells to associate with virulent blood forms of T. cruzi, as evidenced by significant increases in both the proportion of parasite-associated cells and the number of parasites associated with the cells. Three lines of evidence pointed to IFN-gamma, and not interleukin 2, as the lymphokine responsible for the noted effect. First, a conditioned medium containing interleukin 2 but not IFN-gamma failed to enhance P338D1 cell-parasite association. Second, treatment of the IFN-gamma preparation at pH 2 to selectively inactivate IFN-gamma reduced its enhancing effect. Third, recombinant IFN-gamma, devoid of other lymphokines, also enhanced parasite association with P388D1 cells. Incubation of P388D1 cells with IFN-gamma for 24, 48, or 72 h increased cell association with T. cruzi, whereas a 12-h incubation period was insufficient, suggesting that IFN-gamma triggered time-dependent cellular events leading to the enhancement. Treatment of mouse resident peritoneal macrophages with the IFN-gamma-containing conditioned medium also increased the capacity of these cells to kill internalized trypanosomes. P388D1 cells, which showed minimal or no cytotoxicity after mock treatment with medium, displayed cytotoxicity after incubation with the IFN-gamma-containing conditioned medium; similar results were obtained with recombinant IFN-gamma. Catalase prevented parasite killing by P388D1 cells, indicating that H2O2 mediated the cytotoxicity. These results, underscoring the regulatory effects of IFN-gamma on macrophage-parasite interactions, suggest a possible role for this lymphokine in the mechanisms of host defense active against T. cruzi infection.
Mechanism of polymyxin B-mediated lysis of lipopolysaccharide-treated erythrocytes
Authors: Carr C , Morrison D C
Published: January 2020

A novel system was used previously to characterize the dynamic interaction of a polysaccharide-deficient, lipid-rich lipopolysaccharide (LPS) with rabbit erythrocytes (RaRBC). Exposure of the RaRBC to the LPS rendered them sensitive to induction of hemolysis by the cationic antibiotic polymyxin B (PB) in a time- and temperature-independent manner. Subsequent decay in the response of LPS-sensitized cells to PB was shown to be critically dependent on both the time and temperature of incubation of RaRBC with LPS and to be independent of a change in LPS binding (Carr and Morrison, Infect. Immun. 43:600-606, 1984). In the present study, we performed experiments designed to define the mechanism by which PB mediates hemolysis of LPS-sensitized RaRBC. Experiments were performed to examine the molecular requirements of the LPS and the PB that were essential for hemolytic activity. The capacity of various cations to mediate hemolysis of LPS-sensitized RaRBC or to block PB-mediated hemolysis and the temperature dependence of the PB lytic reaction were investigated. The results of these experiments suggest that PB-mediated hemolysis of LPS-treated erythrocytes is dependent upon an initial ionic association of PB with erythrocyte membrane-bound LPS, followed by hydrophobic insertion of the PB fatty acid into the erythrocyte membrane lipid bilayer.
Dissociation between interleukin-1 and interleukin-2 production in proliferative response to microbial antigens: restorative effect of exogenous interleukin-2
Authors: Vismara D , Lombardi G , Piccolella E , Colizzi V
Published: January 2020

The relationship between the production of interleukin-1 (IL-1) and interleukin-2 (IL-2) after stimulation of human mononuclear cells within an antigenic extract from Candida albicans was analyzed in both responder and nonresponder donors. Culture supernatants from responders contained both IL-1 and IL-2 activity, whereas the supernatants from nonresponders contained only IL-1 and no appreciable IL-2. However, the addition of exogenous IL-2 to nonresponder cultures restored the normal proliferative response. Similar observations were made when cells from mice infected intravenously with high doses of Mycobacterium bovis BCG were cultured; these cells showed a marked impairment of the proliferative response to purified protein derivative. Spleen cells from BCG-induced unresponsive mice failed to produce IL-2 despite the fact that normal IL-1 activity was present in the culture. Again, the addition of exogenous IL-2 fully reversed the proliferative unresponsiveness. Thus, the presence of IL-1 does not necessarily induce production of IL-2, and the proliferative unresponsiveness is therefore due to a primary lack of IL-2.
Trypsin-sensitive, bovine serum albumin-dependent hemolysis activity in Mycoplasma pulmonis
Authors: Minion F C , Goguen J D
Published: January 2020

Although Mycoplasma pulmonis did not lyse normal erythrocytes, it rapidly lysed erythrocytes that had cytoskeletal deficiencies which allow increased diffusion of membrane glycophorin or that had been treated with trypsin to remove surface proteins. This hemolysis occurred only in the presence of bovine serum albumin and was eliminated by trypsin treatment of the mycoplasma. Hemolytic activity was restored after such trypsin treatment when mycoplasma protein synthesis was allowed. M. pulmonis hemolytic activity was not diffusible and thus differed from the activities reported for other mycoplasmas, which involve small diffusible intermediates such as hydrogen peroxide. With the exception of the requirement for bovine serum albumin, the factors which affected hemolysis were similar to those which we have previously reported to affect M. pulmonis hemagglutination, suggesting that these two activities are functionally related.
Kinetics of growth and toxigenicity of Clostridium botulinum in experimental wound botulism
Authors: Dezfulian M , Bartlett J G
Published: January 2020

An animal model of wound botulism was developed in mice using an inoculum of Clostridium botulinum type A spores. The number of C. botulinum in infected wounds was quantitated by culturing on egg yolk agar, and the level of C. botulinum toxin in infected wound tissue was measured by a bioassay in mice and by an enzyme-linked immunosorbent assay. All infected mice receiving no further treatment developed neuroparalytic symptoms consistent with botulism after an incubation period of ca. 48 h, and all of these animals died. Serotherapy with C. botulinum type A antitoxin initiated 24 h postchallenge reduced the mortality rate to 5%. Treatment with metronidazole 2 to 24 h postchallenge resulted in recovery rates of 40 to 91%.
Correlation between adherence to HeLa cells and serogroups, serotypes, and bioserotypes of Escherichia coli
Authors: Scaletsky I C , Silva M L , Toledo M R , Davis B R , Blake P A , Trabulsi L R
Published: January 2020

Four hundred fifty Escherichia coli strains of 45 O serogroups and subgroups and 112 serotypes were studied to determine their patterns of adherence to HeLa cells. Adherence was exhibited by strains of 17 O serogroups and subgroups, but within these groups more than one adherence pattern was frequently observed. However, within each serotype, the adherence pattern was highly consistent. Localized adherence (LA) was observed much more frequently in serotypes that we considered to be enteropathogenic E. coli serotypes (93%) than in other serotypes (14%), whereas diffuse adherence (DA) occurred predominantly among nonenteropathogenic E. coli strains. Determination of biochemical characteristics showed that within O serogroups, nonmotile strains tended to have the same behavior as motile strains with the LA adherence pattern, suggesting that they were derived from these motile strains. LA and non-LA strains of the same serotype differed biochemically. LA appears to be a property of most E. coli commonly considered to be enteropathogenic and should assist attempts to determine which E. coli are enteropathogenic and to elucidate their pathogenic mechanisms.
Immunoglobulin M and immunoglobulin G responses in BALB/c mice to conjugated outer membrane extracts of four Salmonella serotypes
Authors: Kudrna D A , Teresa G W , Arnzen J M , Beard K S
Published: January 2020

Outer membranes (OMs) of Salmonella enteritidis, S. anatum, S. typhimurium, and S. infantis were extracted and cross-linked with glutaraldehyde to form a large macromolecular antigen. The antigen consisted of OM proteins and lipopolysaccharide and was designated 4-OMP-LPS. Polyacrylamide gel electrophoresis of extracted OMs from each serotype revealed differences in protein profiles. S. enteritidis and S. infantis possessed a greater variety of proteins than did S. anatum and S. typhimurium. Immunizations with 4-OMP-LPS in phosphate-buffered saline (4-OMP-LPS-C) and 4-OMP-LPS emulsified with muramyl dipeptide in the oil phase of a hexadecane-water emulsion (4-OMP-LPS-MDP) revealed that BALB/c mice were capable of eliciting specific primary and secondary immunoglobulin M (IgM) and IgG responses. Both antigen preparations were capable of eliciting IgM and IgG specific for the cell surfaces of each live Salmonella serotype. Also, 4-OMP-LPS-MDP and 4-OMP-LPS-C were capable of evoking a substantial anamnestic response. Adsorption studies revealed that the combined serotypes had the antigenic capacity to adsorb up to 94% of the antibodies, but 4-OMP-LPS-MDP antibodies were more effectively adsorbed than were 4-OMP-LPS-C antibodies. Adsorption of pooled antiserum with heterologous bacteria yielded a variety of adsorption profiles.
Transient Increase in Capillary Permeability Induced by
Clostridium perfringens
Type A Enterotoxin
Authors: Stark Raymond L. , Duncan Charles L.
Published: January 2020

A transient increase in capillary permeability was induced in the skin of guinea pigs by
Clostridium perfringens
type A enterotoxin. Maximum intensity and maximum diameter of the area of increased capillary permeability were obtained with injection of enterotoxin 10 to 20 min before dye injection.
Classification of
Neisseria meningitidis
Group B into Distinct Serotypes I. Serological Typing by a Microbactericidal Method
Authors: Frasch Carl E. , Chapman S. Stephen
Published: January 2020

Neisseria meningitidis
group B was subdivided into a number of different serotypes. A sensitive microbactericidal assay procedure was developed, and, with this method, over 10 different serotypes were provisionally established. The method by which distinct serotype designations were assigned is described. Almost half of the strains examined thus far contained a common serotype antigen which was distinct from the grouping antigen. Differentiation of group B meningococci into distinct types will aid in epidemiological studies and in the selection of appropriate strains for vaccines against group B meningococci.
Development of Bactericidal Capacity and Phagocytosis-Associated Metabolism of Fetal Pig Leukocytes
Authors: Holmes Beulah , Day Noorbibi , Haseman Judy , Good Robert A.
Published: January 2020

Evidence that the bactericidal ability and the stimulated oxidative metabolism of leukocytes appear in parallel during fetal development of the Minnesota Miniature pig has been obtained by application of the techniques applied to studies of human cells. It was demonstrated that leukocytes from 87- to 90-day fetuses were fully capable of ingesting
Staphylococcus aureus
but greatly diminished in bactericidal capacity as compared to leukocytes of older fetuses and adults. Although resting levels of oxygen consumption and hexose monophosphate pathway activity of leukocytes from the younger fetuses compared well with those of leukocytes from older animals, the phagocytosis-stimulated increments of metabolism were much less at 87 to 90 days of gestation than at later developmental stages. Both bactericidal capacity and increased metabolism of leukocytes reach adult levels by 100 days of gestation (normal gestation period of 115 to 120 days). Acrylamide gels stained for reduced nicotinamide adenine dinucleotide (NADH) and NADH phosphate (NADPH) diaphorase activity after disc electrophoresis of leukocyte extracts revealed normal mobility and intensity of NADH diaphorase bands. Three NADPH diaphorase bands were present in adult leukocyte extracts. Only the fast-migrating NADPH diaphorase band of 87- to 90-day cells stained with decreased intensity. This “deficiency” was no longer present at the later fetal period. The fast-migrating NADPH diaphorase band may represent an electron transfer protein which functions in cyanide-insensitive respiration of the leukocytes of the pig.
Latex Agglutination Test for Measurement of Antibodies to Meningococcal Polysaccharides
Authors: Tramont Edmund C. , Artenstein Malcolm S.
Published: January 2020

A latex agglutination test employing serogroup-specific meningococcal polysaccharides as the antigen has been developed. The test has been used to measure serological responses in patients with meningococcal disease, in meningococcal carriers, and in volunteers who received meningococcal vaccines. It has been shown to be a sensitive and highly specific test for the detection of group-specific meningococcal antibody. The advantages of standardized particles and antigens which are simple to prepare and stable make this assay feasible for many nonspecialized or small laboratories.
Suppression of the Intracellular Growth of
Shigella flexneri
in Cell Cultures by Interferon Preparations and Polyinosinic-Polycytidylic Acid
Authors: Gober L. L. , Friedman-Kien A. E. , Havell E. A. , Vilček J.
Published: January 2020

A strain of
Shigella flexneri
type 2a was found to multiply intracellularly in cultures of the diploid human cell strain FS-1 and in secondary rabbit kidney cells. When inoculated cultures were stained and observed under the light microscope, it appeared that the cytoplasm of infected cells became gradually filled with bacteria. Various preparations of human and rabbit interferon were found to suppress the intracellular bacterial growth in homologous cells. Polyinosinic-polycytidylic acid (poly I·poly C) had a similar inhibitory effect. Rabbit interferon preparations did not cause a significant suppression in human cells. Suppression of the bacterial growth could be demonstrated in two ways: (i) by showing that treatment with either homologous interferon or poly I·poly C reduced the proportion of infected cells determined by counting the total number of cells and the number of cells with 10 or more bacteria in several microscopic fields selected at random, or (ii) by showing that a suppression in
3
H-uridine incorporation by bacteria occurs in infected cultures treated with actinomycin D after incubation with interferon or poly I·poly C. (Uridine incorporation by the bacterium is insensitive to actinomycin D.) Treatment of cells with actinomycin D before incubation with interferon prevented the development of cellular resistance to bacterial infection. Interferon preparations did not have an inhibitory effect on the extracellular growth of
S. flexneri
in a broth culture. These findings show that the range of activity of the interferon system apparently extends to intracellularly growing bacteria.
Adherence Inhibition of
Streptococcus mutans:
an Assay Reflecting a Possible Role of Antibody in Dental Caries Prophylaxis
Authors: Olson Gerald A. , Bleiweis Arnold S. , Small Parker A.
Published: January 2020

Adherence of cariogenic streptococci to the smooth surfaces of teeth has been shown to lead to plaque formation and ultimately to caries production. This adherence can be reproduced in vitro and requires sucrose in the growth media. The work reported here describes the development of an assay based on the ability of immune rabbit serum to inhibit adherence of
Streptococcus mutans
6715 to glass surfaces. Rabbits were immunized with Formalin-killed whole bacteria. Sera were titered for adherence inhibition and bacterial agglutination. The former assay was found to be dependent upon immunoglobulin G (IgG) antibody, whereas the latter detected either IgG or IgM antibodies. These two assays appeared to detect different antigens. It was hypothesized that the adherence inhibition assay detects antibody directed against an enzyme, perhaps dextransucrase, responsible for dextran synthesis, since immune serum was found to inhibit incorporation of radiolabel from
14
C-sucrose into cell wall extract. The adherence inhibition assay has potential application for study of other cariogenic bacteria. Studies are being pursued in the hope that this assay may lead to a means of control of dental caries.
Studies of the Rickettsial Plaque Assay Technique
Authors: Wike David A. , Tallent George , Peacock Marius G. , Ormsbee Richard A.
Published: January 2020

A plaque assay system for pathogenic rickettsiae, which utilizes primary chick embryo tissue cultures, is described. It proved to be a highly reproducible measure of infectiousness for
Rickettsia rickettsi
and
R. typhi
, which were employed in most studies; as well as for
R. canada, R. prowazeki, R. sibirica, R. akari, R. conori
, and
Coxiella burneti.
Plaque-forming units (PFU) were compared to direct rickettsial counts and to 50% infectious dose (ID
50
) values for embryonated eggs, mice, and guinea pigs. Plaque size, appearance, and number were influenced by diluent, incubation temperature after nutrient overlay, centrifugation of inoculated tissue cultures, and number of host cells planted initially in each flask. The most critical factors in plaque formation were diluent used in making rickettsial suspensions and incubation temperature (32 C) after nutrient overlay. Brain Heart Infusion was the only diluent capable of preventing significant delay in plaque formation and decreases in PFU and mouse ID
50
. Plaque formation was unaffected by genetic background of host cells, volume of inoculum, temperature and length of incubation period before nutrient overlay, and rapid freezing and thawing of rickettsial seed. Centrifugation of inoculated cultures at 600 ×
g
resulted in 100% irreversible absorption of rickettsiae to host cells within 5 min, whereas without centrifugation at least 4 hr was required to achieve the same effect.
M Protein-Associated Adherence of
Streptococcus pyogenes
to Epithelial Surfaces: Prerequisite for Virulence
Authors: Ellen R. P. , Gibbons R. J.
Published: January 2020

Virulent strains of
Streptococcus pyogenes
containing M protein were found to adhere well to human cheek epithelial cells in vitro, whereas an avirulent M − mutant strain adhered feebly. Pretreatment of M + strains with trypsin to remove their M protein surface coating or reacting them with type-specific antiserum markedly impaired their abilities to attach to epithelial cells. Electron microscopy revealed that the attachment of an M + strain to germfree rat epithelial cells was mediated by a fuzzy surface structure previously shown to contain M protein. When mixtures of streptomycin-resistant M + and M − strains were introduced into the mouths of mice, the proportions of the M + strain increased on tongue and cheek surfaces relative to its M − mutant. These data indicate that the surface fuzz of
S. pyogenes
which contains M protein functions in the attachment of the organism to epithelial surfaces, thereby permitting its colonization.
Interferon-Inducing Polycarboxylates: Mechanism of Protection Against Vaccinia Virus Infection in Mice
Authors: Billiau A. , Muyembe J. J. , De Somer P.
Published: January 2020

The present study was undertaken to examine possible correlations between long-term protection of mice treated with polycarboxylates and infected with vaccinia virus, and the presence of subdetectable amounts of interferon in the tissues, as determined by tissue or organ resistance to replication of viruses. After a single dose of polycarboxylate, splenic resistance to virus replication could be detected. It persisted for 7 to 14 days, but no attempt was made to prove that it was due to subdetectable amounts of interferon. Whole-animal protection lasted longer than splenic resistance. Moreover, when different polycarboxylates and different virus strains were used, patterns of early protection were not correlated with those of splenic resistance. These data, as others presented earlier, suggest that, in addition to interferon, other antiviral mechanisms are stimulated by polycarboxylates.
Adenovirus Susceptibility to Human Interferon During One-Step Replication
Authors: Gallagher James G. , Khoobyarian Newton
Published: January 2020

Susceptibility of adenovirus types 2, 7, and 12 to human interferon was measured in three human diploid cell strains during a single-cycle infection. Although the relative susceptibility of adenovirus to interferon varied in these cell strains, the final yield of each type in each cell strain decreased as the interferon dose increased. On the other hand, wide difference in interferon susceptibility of adenoviruses and vesicular stomatitis virus (VSV) was noted, as interferon doses above 100 units profoundly inhibited VSV but not the adenoviruses.
Respiratory Diseases in Cyclophosphamide-Treated Mice I. Increased Virulence of
Mycoplasma pulmonis
Authors: Singer S. H. , Ford M. , Kirschstein R. L.
Published: January 2020

Mice infected intranasally with
Mycoplasma pulmonis
were treated with cyclophosphamide, a potent immunologic suppressor. In place of a chronic smouldering infection with little mortality (2%), a rapidly lethal infection with high mortality (66%) was produced.
M. pulmonis
was able to be isolated from several organs during the course of the unmodified infection. In the infected animals treated with cyclophosphamide, dissemination occurred earlier, and higher titers of mycoplasma were found. Reconstitution experiments with spleen cells from previously infected animals reversed the effect of cyclophosphamide, indicating that immunity plays an important role in containment of the infection and eventual recovery.
Cryoprotein Produced by
Clostridium botulinum
Type G
Authors: Ciccarelli Alberto S. , Giménez Domingo F.
Published: January 2020

Strain 89 of
Clostridium botulinum
type G produces a protein which behaves as a cryoprotein.