bluuebunny/crossref_metadata_2025 huggingface dataset:
Chemical Composition and Immunological Specificity of the Streptococcal Group O Cell Wall Polysaccharide Antigen
Authors: Mukasa Hidehiko , Slade Hutton D.
Published: January 2020

The group O streptococcal group antigen was shown to be a polysaccharide located in the cell wall of the organism. The antigen could be extracted by one of several methods: (i) 0.5
n
NaOH at 37 C, (ii) phenol-water (50:50) at 68 C, (iii) 0.2
n
HCl at 100 C, or (iv) 10% trichloroacetic acid at 4 C. The last method yielded more polysaccharide with less protein contamination. The polysaccharide was purified on diethylaminoethyl-Sephadex A-25 and Sephadex G-200. It was composed of two-thirds glucosamine and galactosamine, and the remainder glucose plus galactose. Rhamnose, glycerol, ribitol, and muramic acid were absent. Total phosphorus and amino acids were each less than 0.1%.
N
-Acetyl-β-
d
-glucosamine exerted a strong inhibition of the precipitin reaction and is considered the immunodominant sugar. Glucosamine and glucose possessed a partial inhibitory activity. Galactose and galactosamine were essentially negative. No evidence of cross-reactivity was found between the O polysaccharide and group A and L polysaccharides, and group A and
Staphylococcus aureus
teichoic acids, which posesss
N
-acetylglucosamine specificity. The release of limited quantities of
N
-acetyl-glucosamine from its terminal location by enzyme, and glucose by acid hydrolysis, indicates a limited number of side chains in the O antigen. The glucosamine is in acid-stable linkage in the polysaccharide. Glucose was not released by β-glucosidase and probably does not occupy a terminal position. The O antigen is the only known streptococcal polysaccharide antigen which does not contain rhamnose. The effect of these factors on the immunological specificity is discussed. O serum, after adsorption with the purified polysaccharide, was used to demonstrate the presence of protein antigens in acid extracts of cells from each of the nine strains examined. These antigens may represent type antigens. Two of these strains, originally described as group O, did not contain the O polysaccharide.
Experimental Infection of Horses with an Attenuated Venezuelan Equine Encephalomyelitis Vaccine (Strain TC-83)
Authors: Walton Thomas E. , Alvarez Otto , Buckwalter Ross M. , Johnson Karl M.
Published: January 2020

Ten horses (
Equus caballus
) were vaccinated with strain TC-83 Venezuelan equine encephalomyelitis (VEE) virus vaccine. Febrile responses and leukopenia due to a reduction of lymphocytes and neutrophils were observed in all animals. Viremias were demonstrable in eight horses, with a maximum of 10
3.5
median tissue culture infectious dose units per ml of serum in two horses. Clinical illness with depression and anorexia were observed in five horses. Neutralizing (N), hemagglutination-inhibiting, and complement-fixing antibodies to the vaccine virus were demonstrable by 5, 6.5, and 7 days, respectively, after vaccination. Differential titrations of serum to six VEE strains revealed high titers of N antibody to vaccine virus, moderate titers to the epizootic Trinidad donkey no. 1 strain (VEE antigenic subtype I, variant A) from which TC-83 was derived, and low titers to two other epizootic strains (subtype I, variants B and C) in all horses at 1 month after vaccination; some animals responded with low levels of N antibody to the enzootic viruses (subtype I, variants D and E). Fourteen months after vaccination, six animals with detectable N antibody were challenged with MF-8 (subtype I, variant B), an epidemic-epizootic strain isolated in 1969 from a man in Honduras. All horses resisted challenge with the equine pathogenic strain of VEE. Marked increases of N antibody in most horses were demonstrable to some VEE strains when tested 1 month after challenge.
Effect of Antithymocyte Serum on Experimental Toxoplasmosis in Mice
Authors: Strannegård Örjan , Lycke Erik
Published: January 2020

The effect of antithymocyte serum (ATS) on mice infected with high-virulent or low-virulent strains of
Toxoplasma gondii
was studied. Treatment with ATS reduced the survival time of mice infected with high-virulent
Toxoplasma
and aggravated the disease; treatment prolonged parasitemia in animals infected with low-virulent
Toxoplasma.
Although ATS adversely affected the humoral antibody response, its main effect on toxoplasmosis was probably due to depression of cell-mediated immunity. ATS appeared to be able to activate chronic
Toxoplasma
infection, but this effect was much less pronounced than that on acquisition of immunity. The results suggest that
Toxoplasma
infection should be routinely looked for in patients treated with antilymphocyte serum.
Isolation of an Additional Capsular-Type Strain of
Staphylococcus aureus
by the Serum-Soft Agar Technique
Authors: Yoshida Kosaku
Published: January 2020

An additional capsular-type strain of
Staphylococcus aureus
was isolated by the serum-soft agar technique and designated as capsule type D.
Indirect Hemagglutination Test for Varicella-Zoster Infection
Authors: Furukawa Toru , Plotkin Stanley A.
Published: January 2020

An indirect hemagglutination test has been adapted for use with varicella-zoster serology. Antigen was made in infected WI-38 human diploid cell line. The indirect hemagglutination antigen seemed to be a subunit of the virus particle. The antibodies measured in the test are well correlated with neutralizing and complement-fixing antibodies in cases of zoster but not in cases of varicella.
Phagocytosis of Microbial L-Forms
Authors: Harwick Herbert J. , Barajas Luciano , Montgomerie John Z. , Kalmanson George M. , Guze Lucien B.
Published: January 2020

Little data are available regarding L-form phagocyte interactions. Due to difficulty with more conventional methods, a serum-free technique of surface phagocytosis employing cellulose ester micropore filters, fluorochrome staining, and reflecting fluorescence microscopy was developed. Intracellular localization of L-frms was confirmed by electron microscopy. In timed experiments, phagocytosis of a group D
Streptococcus
(ATCC 23241) and its L-form (T53I) were maximal at 30 min, although the bacteria were consistently better phagocyted. Phagocytosis of an
Escherichia coli
(strain Yale) and its L-form was also demonstrated with the same technique. The lesser phagocytosis of L-forms may be related to their lower chemotactic activity.
Effect of Vitamin E on the Immune Response of Hypoxic and Normal Chickens
Authors: Tengerdy Robert P. , Heinzerling Rollin H. , Nockels Cheryl F.
Published: January 2020

Chicks and hens raised on a diet enriched with 60 mg of vitamin E per lb. of feed had a significantly increased immune response as measured by the antibody plaque-forming cell test or by hemagglutination. Since the effect was much greater on hypoxic (simulated altitude exposed) chicks, a synergistic effect between hypoxia and vitamin E may be suggested.
Evolution of inflammatory response and cellular immune responses in a murine model of disseminated blastomycosis
Authors: Deepe G S , Taylor C L , Bullock W E
Published: January 2020

A reproducible model of disseminated blastomycosis was established in C57BL/6 mice by intravenous injection of 10(6) yeast-phase Blastomyces dermatiditis organisms. The infection progressed over 5 weeks to involve lungs, brains, superficial fascia, livers, and spleens of mice. By week 5, there was a greater number of organisms in lungs and brains than in livers and spleens. The tissue response in lungs, brains, and livers progressed from acute neutrophilic invasion before week 1 to pyogranuloma formation by week 5. Lymph nodes and spleens were remarkably spared. By week 5, infected mice became anergic to intradermal challenge with both specific Blastomyces antigen and a nonspecific antigen (sheep erythrocytes). At this time, the response to concanavalin A or phytohemagglutinin by splenocytes was markedly less than that of normal controls. Likewise, the plaque-forming cell response to sheep erythrocytes by splenocytes from infected mice was diminished. In coculture studies, splenocytes from 5-week-infected mice reduced the plaque-forming cell response by normal splenocytes. The development of this murine model should prove useful for elucidating the perturbations of immunoregulation associated with disseminated blastomycosis.
Brucella fractions behave as nonspecific mitogens and polyclonal B-cell activators for human lymphocytes
Authors: Vendrell J P , Rabesandratana H , Huguet M F , Cannat A , Serre A
Published: January 2020

Two lipid-A-free fractions which were extracted from Brucella melitensis and were designated PI and SF stimulated human unsensitized mononuclear cells to proliferate and to secrete immunoglobulins. Both of these effects were observed in cultures of peripheral blood, tonsils, and cord blood lymphocytes. Neither B cells nor T cells alone proliferated in the presence of these fractions, whereas the proliferative response of T cells plus B cells was largely independent of accessory cells. Polyclonal activation was estimated by counting the cells which secreted immunoglobulins of different isotypes into culture supernatants. This phenomenon was strongly T dependent.
Demonstration of cholera toxin-related factor in cultures of Aeromonas species by enzyme-linked immunosorbent assay
Authors: Honda T , Sato M , Nishimura T , Higashitsutsumi M , Fukai K , Miwatani T
Published: January 2020

The production of toxins by Aeromonas species was examined by the suckling mouse test, the hemolysin test, and the enzyme-linked immunosorbent assay with anticholera enterotoxin. A factor that was immunologically related to cholera enterotoxin was produced by 5 of 14 strains of Aeromonas hydrophila and 4 of 15 strains of Aeromonas sobria. Analysis by these assays and by a test for heat stability suggested that the factor differed from hemolysin and from toxin that was active in the suckling mouse test.
Shwartzman reaction in germfree rabbits
Authors: Ito M
Published: January 2020

The local Shwartzman reaction occurred in germfree rabbits which had no natural antibody to endotoxin and none or only a very small amount of immunoglobulin G. From the results it was concluded that the presence of natural antibody to endotoxin is not a prerequisite of the production of the local Shwartzman reaction by bacterial endotoxin.
Effects of carbon dioxide and pH on adhesion of Candida albicans to vaginal epithelial cells
Authors: Persi M A , Burnham J C , Duhring J L
Published: January 2020

A controlled-environment membrane model for use in vitro was developed and employed in an attempt to mimic the environment of the vagina in order to study yeast-vaginal cell adhesion. Adhesion in vitro of four strains of Candida albicans (NIH 3181A, NIH 526B, ATCC 18804, and MCO 2400) to vaginal epithelial cells (VEC) appeared to be affected by the pH and the level of carbon dioxide that have been found to be present in the vagina in vivo. Strain 3181A had a greater adhesion ability than 526B when the concentration of yeast cells was increased and when the yeast cells were incubated with VEC at pH 5 in sodium phosphate buffer in ambient air supplemented with 10% CO2. Of the four strains of C. albicans used, 3181A had the greatest adhesion ability, with strains 2400, 18804, and 526B ranked in order of decreasing adhesion ability. Also, an enhanced, electron-dense, matted outer region of the cell walls of the yeasts was observed frequently when they were incubated in ambient air supplemented with 10% CO2. In addition, of the vaginal cells that had yeast cells attached to them, an average of 94.4% of the total yeast cells were attached to the microridge side of the VEC, whereas an average of only 5.6% of the total were found on the nonmicroridge side of the VEC. The results from this study indicate that adhesion of C. albicans to the VEC surface was affected by the strain of yeast used, by the side of the vaginal cell exposed, and by the pH and CO2 levels present in the adhesion assay.
Interleukin 2 enhances specific and nonspecific immune responses in experimental Chagas' disease
Authors: Choromanski L , Kuhn R E
Published: January 2020

Mice infected with Trypanosoma cruzi develop an early and profound immunosuppression of responses to heterologous antigens. Recently it has been demonstrated that this immunosuppression is linked, in part, to deficiency in the production of interleukin 2 (IL-2), and that the addition of IL-2 to cultures of normally unresponsive spleen cells from infected mice will restore responsiveness to sheep erythrocytes (SRBC) and enhance parasite-specific immune responses. In the present study, the effect of administration of ultrapure or recombinant IL-2 on immune responses to SRBC and parasite-specific responses in vivo was examined. It was found that a single injection of 1,500 U of IL-2 provided at the same time as SRBC more than doubled the number of direct plaque-forming cells to SRBC and that multiple injections of 1,500 U of IL-2 were no more restorative than a single injection. Anti-SRBC responses of normal mice were unaffected by injection of IL-2. Single or multiple injections of recombinant human IL-2, with and without gelatin, into highly susceptible C3H(He) mice induced greater parasite-specific immunity as reflected by significantly reduced levels of parasitemia and increased longevity. Three injections of 1,500 U each of recombinant human IL-2 on days 10, 14, and 18 was found to be the most efficacious in reducing parasitemia and increasing longevity. Injection of IL-2 with gelatin did not enhance the effect of IL-2 alone.
Role of type 1 pili and effects of phase variation on lower urinary tract infections produced by Escherichia coli
Authors: Hultgren S J , Porter T N , Schaeffer A J , Duncan J L
Published: January 2020

Phase variation of type 1 pili (fimbriae) was studied during the in vivo growth of Escherichia coli in two animal models. In the first, a heavily piliated urinary tract isolate (strain 149) was placed in 1-cm polypropylene chambers sealed with 0.22-micron-pore-size filters. The chambers were surgically implanted intraperitoneally in mice and recovered at various times. Piliation, as determined by electron microscopy and by measuring the minimum number of bacteria needed to produce mannose-sensitive hemagglutination, gradually decreased, and by day 5, most of the organisms were nonpiliated. In the second model, piliated and nonpiliated E. coli phase variants were inoculated into the bladders of BALB/c mice via urinary catheters, and their fate in the lower urinary tract was studied. Viable counts of bladder homogenates revealed that piliated phase variants were significantly more effective in colonizing the bladder urothelium than were their nonpiliated counterparts. Specific antibody to type 1 pili prevented colonization by the piliated organisms. After inoculation of piliated variants, the bladder-associated bacteria gave rise to approximately 80% mannose-sensitive hemagglutination-positive colonies, and immunocytochemistry of bladder lavages revealed large numbers of type 1 piliated bacteria adhering to the bladder transitional cells. Electron microscopy confirmed the presence of piliated bacteria in association with the bladder urothelium. The urine of these mice, whose bladders were colonized with piliated bacteria, frequently showed no growth, and when bacteria were present, strain 149 yielded less than 30% hemagglutination-positive colonies. The results suggest that for some E. coli strains, phase variation may be a factor in determining the fate of the E. coli in the urinary tract and that the urine may not necessarily reflect the bacteriologic state of the bladder mucosa.
Structural integrity of host defense factors in dental plaque
Authors: Hsu S D , Cole M F
Published: January 2020

The structural integrity of immunoglobulin A (IgA), IgG, and IgM and lactoferrin in dental plaque fluid samples from two populations of Colombian children with contrasting levels of dental caries was examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by electrophoretic transfer to nitrocellulose. The immune factors or their fragments or both were detected with monospecific antibody conjugated with horseradish peroxidase. All the immune factors examined were extensively degraded, although there appeared to be small amounts of intact IgA and IgG in some samples. Analysis of the samples with antibody to secretory component showed that secretory IgA as well as serum IgA was degraded. IgG appeared to be cleaved into two major fragments, one fragment having a relative mobility similar to the F(ab')2 fragment of IgG and the other a relative mobility slightly greater than Fc. IgM and lactoferrin were virtually completely degraded. There was no apparent relationship between the fragmentation patterns of IgA and IgG in the plaque fluid samples from the two communities and their susceptibility to dental caries.
Defective immunoglobulin M responses to vaccination or infection with Schistosoma mansoni in xid mice
Authors: Correa-Oliveira R , Sher A
Published: January 2020

Mice vaccinated with irradiated Schistosoma mansoni cercariae develop a persistent immunoglobulin M (IgM) antischistosomulum antibody response. To investigate the possible role of antilarval IgM antibodies in the effector mechanism of vaccine-induced immunity, CBA/N mice, which have an X-linked genetic defect resulting in impaired IgM antibody responses to certain antigens, were analyzed for their resistance to a challenge infection. When either infected with unattenuated parasites or vaccinated with irradiated cercariae, mice of this inbred strain failed to produce detectable IgM antibodies to schistosomulum surface membrane and soluble worm antigens. To analyze the effect of this IgM deficiency on immunity, F1 hybrids were constructed between CBA/N females and nondefective C57BL/6J males. As expected, vaccinated (CBA/N X C57BL/6J)F1 females, as well as (CBA/J X C57BL/6J)F1 males and females, produced normal IgM antibodies to both surface antigens and worm antigen extracts. However, such antibodies were not produced by (CBA/N X C57BL/6J)F1 males (hemizygous for xid). Nevertheless, (CBA/N + C57BL/6J)F1 males displayed the same high levels of immunity to challenge infection as (CBA/N X C57BL/6J)F1 females and (CBA/J X C57BL/6J)F1 males and females. These results indicate that vaccine-induced immunity is not dependent on an IgM response to schistosome antigens.
Modification of interactions between neutrophils and staphylococci by lysosomotropic weak bases
Authors: Styrt B , Klempner M S
Published: January 2020

Weak bases that alkalinize the pH within neutrophil lysosomes inhibit in vitro cell functions, including lysosomal enzyme release and superoxide production. To determine the relevance of this inhibition to microbicidal activity, the effect of lysosomotropic weak bases on interactions between human neutrophils and Staphylococcus aureus 502a was studied. After treatment with 1 mM chloroquine, neutrophils showed significantly impaired phagocytosis of 14C-labeled S. aureus. However, 50 mM ammonium chloride had no effect on phagocytosis, although we have previously shown that this concentration raises lysosomal pH and inhibits degranulation and superoxide production. This base was therefore used to study effects on intracellular microbicidal activity. Incubation of neutrophils with 50 mM ammonium chloride diminished killing of S. aureus (22.9 +/- 6.3% of bacteria surviving versus 8.2 +/- 1.3% in suspensions without ammonium chloride). At 1 mM, ammonium chloride had no significant effect. The inhibition of cellular function could be neither explained as a function of neutrophil death, as measured by trypan blue dye exclusion, nor attributed to direct promotion of bacterial growth (in the absence of neutrophils, colony counts were similar in the presence or absence of ammonium chloride) or enhanced resistance to neutrophil microbicidal mechanisms (bacteria treated with ammonium chloride and washed before neutrophil exposure showed no improvement in survival). Ammonium chloride at 50 mM also impaired neutrophil killing of S. aureus in an anaerobic chamber, but microbicidal activity against Escherichia coli S15 was not affected. These findings suggest that optimal neutrophil killing of staphylococci requires a highly acid intralysosomal compartment, but ingestion of bacteria does not. This may reflect primary failure of acidification of the phagocytic vacuole or differential pH requirements for fusion of the plasma membrane with itself and with lysosome membranes. The difference between effects on killing of S. aureus and E. coli is probably a result of the relative importance of the components of neutrophil microbicidal activity against the two organisms.
Three-dimensional structure of fimbriae determines specificity of immune response
Authors: Karch H , Leying H , Goroncy-Bermes P , Kroll H P , Opferkuch W
Published: January 2020

We recently described how a fraction of isolated fimbriae from a multifimbriated strain of Escherichia coli O7:K1:H6 (WF96) could be subdivided by sequential disaggregation in disrupting agents into individual subunits with different molecular weights. In this study, antibodies were raised in rabbits against these isolated fimbrial subunits and against purified intact WF96 fimbriae. These sera were tested by Western blot analysis or by enzyme-linked immunosorbent assays for reactivity against the following antigens: intact WF96 fimbriae, dissociated WF96 fimbriae, dissociated and reaggregated WF96 fimbriae, the WF96 21K fimbrial subunit, reaggregated WF96 21K subunits, the WF96 16K subunits, reaggregated WF96 16K subunits, intact fimbriae from four other E. coli strains, and deaggregated fimbriae from these strains. We found that antibody against intact WF96 fimbriae only reacted strongly with intact WF96 fimbriae, depolymerized and reaggregated WF96 fimbriae, or reaggregated fimbrial subunits; no reactions were evident with intact fimbriae from four other E. coli strains. Conversely, antisera prepared against the WF96 16K subunit and against the WF96 21K subunit did not react with intact WF96 fimbriae or with depolymerized and reaggregated WF96 fimbriae, but did react with homologous isolated subunits. One cross-reaction between fimbrial subunits was apparent: anti-WF96 16K subunit bound to a 21K subunit of deaggregated fimbriae, from another E. coli strain. Taken together, the findings indicate that the three-dimensional structure of the fimbrial preparation used to immunize animals determines the specificity of the immune response.
Production and partial characterization of an elastolytic protease of Vibrio vulnificus
Authors: Kothary M H , Kreger A S
Published: January 2020

Conditions are described for the production of large amounts of an extracellular elastolytic protease by Vibrio vulnificus. The yield of enzyme was maximal during the late exponential growth phase and was stable during the stationary growth phase in a medium composed of 2% Proteose Peptone and 1.5% NaCl. The protease has a molecular weight of ca. 50,500 (estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis), an isoelectric point of ca. 5.8, and a pH optimum range against azocasein and elastin of pH 7 to 8. The caseinolytic and elastase activities in protease preparations partially purified by ammonium sulfate precipitation were inseparable by gel filtration, hydrophobic interaction chromatography, and isoelectric focusing. Both activities were deleteriously affected by heat, low pH, heavy-metal ions, chelating agents, reducing agents, sodium cyanide, N-bromosuccinimide, alpha-2-macroglobulin, and phosphoramidon, but were unaffected by various trypsin inhibitors, chymostatin, aprotinin, leupeptin, pepstatin A, phenylmethylsulfonyl fluoride, and N-ethylmaleimide.
Avirulence and altered physiological properties of cystic fibrosis strains of Pseudomonas aeruginosa
Authors: Luzar M A , Montie T C
Published: January 2020

Twenty Pseudomonas aeruginosa strains isolated from patients with cystic fibrosis in good and poor clinical condition were typed by the American Scientific (Difco Laboratories, Detroit, Mich.) Typing Scheme. Only five strains were agglutinated with a single typing serum. Ten strains were agglutinated with more than one serum, and five were not agglutinated with any serum, suggesting some type of lipopolysaccharide alteration in the majority of these strains. Of the strains from patients in good clinical condition, 72% demonstrated proteolytic activity, while 60% of the strains from patients in poor clinical condition demonstrated no proteolytic activity. Twenty-three cystic fibrosis strains of P. aeruginosa examined demonstrated reduced bacteremic virulence when compared with a virulent burn strain with a 50% lethal dose (LD50) of 1.5 X 10(1) CFU in an invasive burned mouse model. Ninety-two percent of the strains tested were avirulent at doses of 10(3) to 10(5) CFU. The LD50s were determined for 10 selected strains which exhibited specific important morphological and physiological deficiencies. Five of the strains tested gave LD50s greater than 10(6) CFU. Reduced virulence of these strains was associated with loss of two or more physiological characteristics associated with virulence. The cystic fibrosis strains of P. aeruginosa which morphologically and physiologically resembled the virulent burn strain were the most virulent (LD50s of 10(2) to 10(4). Results suggest that some degree of virulence is associated only with classic strains prevalent in early infections. The data suggest that a selection transition occurs in the lungs of patients with cystic fibrosis that favors P. aeruginosa avirulence. The avirulent state may be caused by alterations in the cell envelope, including associated factors such as motility and chemotaxis and protease production.
Structural studies of the rhamnose-glucose polysaccharide antigen from Streptococcus sobrinus B13 and 6715-T2
Authors: Linzer R , Reddy M S , Levine M J
Published: January 2020

The rhamnose-glucose polysaccharide antigens from the cell walls of Streptococcus sobrinus B13 and 6715-T2 (formerly Streptococcus mutans serotypes d and g, respectively) were structurally examined by using gas chromatography-mass spectroscopy. These data confirmed earlier chemical and serological studies suggesting that these polysaccharides had identical structures. The polysaccharides appeared to have a backbone of alternating 1,2- and 1,3-linked rhamnose units. Branching occurred at 1,2,3-linked rhamnose units. Side chains appeared to be composed of 1,2- and 1,6-linked glucose units with glucose as the only terminal carbohydrate.
Neutralization of Chlamydia trachomatis cell culture infection by serovar-specific monoclonal antibodies
Authors: Lucero M E , Kuo C C
Published: January 2020

Seventeen monoclonal antibodies (MAs) were tested. Five of seven serovar-specific MAs from five different serovars and two of five subspecies-specific MAs showed neutralizing activity as well as serovar specificity. No species- or genus-specific MAs showed neutralizing activity. No neutralization occurred without complement. Results indicate neutralization was serovar specific and complement dependent.
Identification and partial characterization of Rickettsia tsutsugamushi major protein immunogens
Authors: Hanson B
Published: January 2020

Strains of Rickettsia tsutsugamushi so far examined have either three or four quantitatively predominant proteins, which apparently are surface proteins and which range in size between 50 and 63 kilodaltons. These polypeptides also were the major immunogens detected by polyacrylamide gel electrophoresis of extracted rickettsial proteins which had been precipitated by hyperimmune rabbit sera. The major proteins from different rickettsial strains share some epitopes, as evidenced by the lack of strain specificity of the rabbit sera in the immunoprecipitation tests. However, similar experiments with a limited number of monoclonal antibodies showed that strain-specific determinants also are associated with at least the 58/60-kilodalton polypeptide. A lack of strain-specific epitopes on the rickettsial surface was indicated by our inability to detect binding of heterologous antisera to the rickettsial surface by immunoferritin labeling. Because the three major proteins of the Karp and Gilliam strains are accessible to antibody in unextracted organisms, it is possible that the exteriorly exposed epitopes of these three polypeptides are strain specific and that their common determinants are normally buried in the membrane or otherwise inaccessible. Attempts to absorb out specific antibody with intact rickettsiae gave equivocal results; however, when immune complexes formed before rickettsial extraction were examined by electrophoresis, antibody appeared to have bound strain specifically with at least the 60-kilodalton protein.
Receptor-mediated entry of diphtheria toxin into monkey kidney (Vero) cells: electron microscopic evaluation
Authors: Morris R E , Gerstein A S , Bonventre P F , Saelinger C B
Published: January 2020

To express toxicity in living cells, diphtheria toxin (DT) must cross a membrane barrier and reach its target in the cytosol. Here we examine the entry of DT into the toxin-sensitive monkey kidney (Vero) cells. Using electron microscopy we directly demonstrated for the first time that DT is internalized by receptor-mediated endocytosis, i.e., via clathrin-coated pits, and enters the endosomal system. Methylamine, which is known to protect cells from DT, stopped the movement of toxin to coated areas of the cell membrane. In the presence of amine, prebound biotinyl-DT was internalized, but toxicity was inhibited. Biochemical evidence revealed that methylamine maintained toxin molecules at a site accessible to neutralization by antitoxin. The data suggest that DT entering Vero cells in the presence of methylamine is sequestered within the cell and does not express toxicity.
Established macrophagelike cell lines synthesize interleukin-1 in response to toxic shock syndrome toxin
Authors: Hirose A , Ikejima T , Gill D M
Published: January 2020

Toxic shock syndrome toxin is already known to induce the production of interleukin-1 (IL-1) by preparations of monocytes and macrophages that are presumably contaminated with other types of cells. The response is enhanced by increasing the density of such monocytes, suggesting that the monocyte's response to toxic shock syndrome toxin may be augmented by its interaction with some other cell. Nevertheless, we now show that several human and murine macrophagelike cell lines (U937, J774, P388D1, and WEHI 3) produce IL-1 when exposed to toxic shock syndrome toxin, and therefore the basic response does not require the presence of cells of other lineages. The cultured cells generally produce less IL-1 than do monocytes, but considerably more IL-1 is induced from cells that have undergone a terminal differentiation as a result of exposure to 1 alpha,25-dihydroxyvitamin D3. High concentrations of cultured cels suppress the production of IL-1; this effect is apparently not due to long-lived inhibitors of IL-1 production or of IL-1 activity, but may be due to a short-lived inhibitor of production.
Importance of a lipopolysaccharide-containing extracellular toxic complex in infections produced by Klebsiella pneumoniae
Authors: Straus D C , Atkisson D L , Garner C W
Published: January 2020

A Klebsiella pneumoniae serotype 2 strain was examined for its ability to produce extracellular toxic material. The organism was grown to the stationary phase in a defined medium, and the toxic material was isolated by ultrafiltration-ion-exchange chromatography on DEAE-Sephacel and gel filtration chromatography on Sepharose 4B or 2B. It was found to be comprised of 63% capsular polysaccharide, 30% lipopolysaccharide, and 7% protein and possessed a 50% lethal dose (when injected intraperitoneally into mice) of 393 +/- 45 micrograms. The toxicity appeared to be associated with the endotoxin portion of the compound, because boiling for 15 min and exposure to proteolytic enzymes had no effect on the toxicity. However, saponification destroyed the toxicity of the compound. Studies employing radial immunodiffusion examining the sera of mice infected with this organism demonstrated in vivo production of the complex at levels sufficiently high to produce death. When sublethal amounts of this complex were placed in the lungs of specific-pathogen-free mice, the lung pathology observed after 24, 48, and 72 h was similar to the damage caused by an active K. pneumoniae lobar pneumonia. These data indicate that this extracellular toxic compound produced by K. pneumoniae may be responsible for the lethality and lung tissue destruction normally associated with an active lobar pneumonia caused by this organism.
In vitro inhibition of adherence of Streptococcus mutans strains by nonadherent mutants of S. mutans 6715
Authors: Murchison H , Larrimore S , Curtiss R
Published: January 2020

Four nonadherent mutants from Streptococcus mutans 6715 mutant UAB66 (serotype g) with similar phenotypes were shown to inhibit the adherence of adherence-proficient S. mutans serotypes c and g strains. One mutant, UAB108, was shown to inhibit adherence by wild-type strains representing serotypes a, d, and e as well. This inhibition of adherence was seen with pairs of strains grown in partially defined (PD) medium supplemented with 1% sucrose in both microtiter plates and glass tubes. The inhibiting factor was present in culture supernatant fluids of inhibiting strains grown in PD medium plus 1% sucrose and was heat stable. Ethanol precipitation of culture supernatant fluids of these strains yielded a water-soluble polymer which effectively inhibited the adherence of UAB66. This polymer, isolated from UAB108, was also shown to inhibit the adherence of UAB66 at lower concentrations than that needed to inhibit adherence with dextran T10. Partially purified glucosyltransferase, isolated from the culture supernatant fluids of glucose-grown UAB108, produced a water-soluble glucan which was shown to inhibit the adherence of UAB66 as well. The methods developed permit rapid screening for strains or mutants of strains or both that inhibit adherence or plaque formation or both by wild-type strains of S. mutans.
Oxidative metabolism in cord blood monocytes and monocyte-derived macrophages
Authors: Speer C P , Ambruso D R , Grimsley J , Johnston R B
Published: January 2020

Little is known about phagocytosis-associated oxidative metabolism in mononuclear phagocytes from the human neonate. We investigated this phenomenon in monocytes from the cord blood of term newborn infants by measuring generation of superoxide anion (O2-) and hydroxyl radical (X OH) after stimulation with opsonized zymosan or phorbol myristate acetate. Production of these microbicidal oxygen metabolites by monocytes from neonates and healthy adult volunteers was equivalent. When cultured in the presence of the macrophage activator lipopolysaccharide or muramyl dipeptide, monocytes from neonates and adults differentiated into cells with the appearance of macrophages and with an enhanced capacity to release O2- compared with cells cultured in the absence of an activator. Monocyte-derived macrophages from neonates produced only slightly less O2- than did adult cells. Thus, unlike the cord blood neutrophil, which exhibits abnormalities in oxidative metabolism, the cord blood mononuclear phagocyte has a respiratory burst that is quantitatively comparable to that of the adult cell.
Reversible expression of flagella in Campylobacter jejuni
Authors: Caldwell M B , Guerry P , Lee E C , Burans J P , Walker R I
Published: January 2020

Campylobacter jejuni 81116 and A3249 undergo a bidirectional transition between flagellated (Fla+) and aflagellated (Fla-) phenotypes. When measured in culture medium, the Fla+----Fla- transition occurred at a rate of 3.1 X 10(-3) to 5.9 X 10(-3) per cell per generation, and the Fla- to Fla+ transition occurred at a rate of 4.0 X 10(-7) to 8.0 X 10(-7) per cell per generation. However, passage through a rabbit intestine markedly favored the Fla+ phenotype.
Molecular specificities of monoclonal antibodies directed against virulent Treponema pallidum
Authors: Marchitto K S , Selland-Grossling C K , Norgard M V
Published: January 2020

Radioimmunoprecipitation (RIP) and Western blot analyses with specific anti-Treponema pallidum subsp. pallidum monoclonal antibodies were used to identify antigens with apparent masses of 102, 84, 54, 53, 52, 47, 32, 29, and 24 kilodaltons (kDa). Cross-reactivity of these antibodies with T. pallidum subsp. pertenue antigens and lack of cross-reactivity with T. phagedenis biotype Reiter, T. vincentii, T. refringens, T. scoliodontum, and T. denticola were also demonstrated by RIP and Western blot analyses. Reactivities in the T. pallidum immobilization test, along with the RIP of lactoperoxidase-catalyzed iodination products, suggested that the identified antigens were surface associated. The abundance and surface association of the 47- and 84-kDa antigens were supported by reactivity in the microhemagglutination test for T. pallidum and by strong reactivity of monoclonal antibodies upon indirect immunofluorescence assays with rabbit-cultivated T. pallidum subsp. pallidum, respectively, but not with T. phagedenis biotype Reiter. Anti-47-kDa and anti-84-kDa monoclonal antibodies were also reactive in indirect immunofluorescence assays using treponemes found in dark-field-positive smears of human genital ulcers.
Immunoresponses to Neisseria meningitidis epitopes: suppression of secondary response to phosphorylcholine is carrier specific
Authors: Faro J , Seoane R , Eiras A , Lareo I , Couceiro J , Regueiro B J
Published: January 2020

Results of our previous work have shown that Neisseria meningitidis serogroup B M986 can induce a phosphorylcholine (PC)-specific plaque-forming cell immunoresponse in mice. Also, a single injection of a relatively low dose of meningococci in NBF1 female mice induced a priming time-dependent suppression on subsequent meningococcus challenge. This suppression was not due to switching to another class of immunoglobulin nor to the presence of a capsule on N. meningitidis. In this study we show that suppression induced by meningococcus is carrier specific. Furthermore, we offer evidence suggesting that the structure(s) on meningococcus that trigger this suppression is heat labile and different from the antigenic structure(s) recognized by the suppressed B cells. In addition, we found that there is a gradual increase in antibody secretion rates of N. meningitidis-induced anti-PC plaque-forming cells that correlates with N. meningitidis priming time. Rather unexpected was the fact that pretreatment of mice with PC-keyhole limpet hemocyanin (thymus-dependent antigen) had a great influence on the subsequent PC-specific immunoresponses induced by N. meningitidis and PC-coupled heat-inactivated meningococcus [PC-(NMB)HI], as shown by (i) a striking decrease in T15 idiotype expression, (ii) concomitant direct anti-PC plaque-forming cells reduction, (iii) switching to immunoglobulin G (N. meningitidis-induced immunoresponse) or immunoglobulin G plus immunoglobulin A [PC-(NMB)HI-induced immunoresponse], and (iv) a significant increase in heterogeneity of plaque-forming cell secretion rates. The possibility that N. meningitidis, PC-(NMB)HI, and PC-KLH stimulate B lymphocytes pertaining to three different subpopulations embedded in distinct regulatory circuits is discussed, with emphasis on the interrelationships between T-dependent and T-independent lymphocyte compartments. We focus on the possibility of the existence of high-level regulatory circuits in which lymphocyte subpopulations or sets of lymphocyte subpopulations with different requirements of activation are connected.
Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and monoclonal antibodies as tools for the subgrouping of Escherichia coli lipopolysaccharide O18 and O23 antigens
Authors: Pluschke G , Moll A , Kusecek B , Achtman M
Published: January 2020

The lipopolysaccharide (LPS) of Escherichia coli O18 isolated from a wide variety of sources was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Four different LPS types, designated O18A, O18A1, O18B, and O18B1, were identified. Most O18 strains possess O18A, O18A1, or O18B LPS types, and these types are clonally associated. A reference test strain with the classical O18ab designation possessed O18B LPS, while two reference O18ac strains possessed O18A and O18A1 LPS, respectively. A panel of 15 anti-O18A B-cell hybridomas was isolated. Enzyme-linked immunosorbent assays revealed that some of the monoclonal antibodies produced by these cells recognize different epitopes. Four of these antibodies suffice to distinguish the four O18 types. Numerous strains whose LPS had been typed by SDS-PAGE were tested by agglutination with seven monoclonal antibodies whose specificities had been determined by enzyme-linked immunosorbent assays. The results indicated a perfect correlation between the two methods. Rabbit antisera raised against O18A bacteria agglutinated boiled bacteria of each of the O18 LPS types efficiently. The antisera were adsorbed with bacteria possessing each of the LPS types. The adsorbed sera only distinguished between two groups: O18A and O18A1 versus O18B and O18B1, as shown by agglutination assays and Western blotting. E. coli O4 and O23 and Serratia marcescens O8 antigens, which are reputed to cross-react with O18, were also analyzed. One O4, one O8, and four O23 strains were tested. All made an LPS which was distinguishable from O18 LPS types by SDS-PAGE. Each O23 strain synthesized a different LPS, and three of them synthesized only few short chains. Some of the monoclonal antibodies reacted with O4, O8, and O23A LPSs. The results are interpreted as indicating that numerous E. coli O serogroups will prove to be chemically heterogeneous and that future analyses of subgroup heterogeneity should be guided by results from SDS-PAGE and rely preferentially on monoclonal antibodies as opposed to rabbit hyperimmune sera.
Characterization of the T-cell proliferative response to a purified glycopeptide antigen (GP-25) present on the Trypanosoma cruzi cell surface
Authors: dos Reis G A , Maldonado M S , Mendonça-Previato L , Barcinski M A
Published: January 2020

A glycoconjugate (GP-25) was previously purified from Trypanosoma cruzi and shown to be a major immunogenic constituent of the parasite cell surface, capable of inducing specific humoral responses in the vast majority of patients with Chagas' disease. In the present study, the T-cell proliferative response to GP-25 was studied in mice immunized with T. cruzi fractions or whole parasites. Recognition of GP-25 by proliferating T cells requires the participation of syngeneic, accessory spleen cells and is specifically blocked by anti-la antibodies. Furthermore, recognition of GP-25 is influenced by the MHC haplotype of accessory antigen-presenting cells. Short-term, GP-25-specific T-cell lines were used to demonstrate the specificity of anti-GP-25 T cells and to show that this glycoconjugate is not involved in T-cell cross-reactivity with heart antigens. T cells primed with nonpathogenic trypanosomatids are able to recognize the purified T. cruzi GP-25 molecule, indicating that T cells recognize a GP-25 epitope which is shared among trypanosomatids.
Characterization of cellular immune response to chemically defined glycoconjugates from Leishmania mexicana subsp. amazonensis
Authors: Rodrigues M M , Xavier M T , Previato L M , Barcinski M A
Published: January 2020

Two defined glycoconjugates (GP-10/20 and FR II Phe) purified from Leishmania mexicana subsp. amazonensis were analyzed with respect to their ability to induce cellular responses in immunized and infected mice. Each glycoconjugate was recognized by specific immune cells, as assessed by the proliferative response of lymph node cells of immunized mice. The response to GP-10/20 depended on helper T cells and antigen-presenting cells and was restricted by a major histocompatibility complex class II gene product. A specific anti-GP-10/20 T-cell line was established, and it was able to transfer a delayed-type hypersensitivity (DTH) response to normal mice. Both antigens were also recognized during an ongoing disease, as assessed by DTH response of infected mice. By this response, it was possible to distinguish susceptible from resistant strains of mice. In the course of the disease in resistant mice a correlation between the size of the primary lesion and the DTH response to GP-10/20 was observed. The presence of the glycoproteins on both promastigote and amastigote forms of the parasite, the antigenic similarities between both fractions, and the distribution of the GP-10/20 antigen in other trypanosomatids were studied. The results showed that both antigens were present on promastigotes and amastigotes. GP-10/20 shared no epitopes with FR II Phe, was included as part of the crude preparation leishmanin, and had some cross-reactive determinants with Leishmania donovani and Crithidia deanei.
Purification of lipoteichoic acid by chromatography in water-organic solvent systems
Authors: Josephson S L , Stinson M W , Millar S J , Cohen R E
Published: January 2020

Lipoteichoic acid (LTA), extracted from Streptococcus mutans 10449 by hot aqueous phenol, was partially purified by Sepharose 6B column chromatography in 0.01 M sodium acetate, pH 6.0, containing 0.25 M sodium chloride and 0.001 M EDTA. Nucleic acid and polysaccharide were precipitated from the LTA-containing column peak by the addition of 2 volumes of chloroform-methanol (1:5). The resulting single-phase chloroform-methanol-water (1:5:3) supernatant contained LTA and small amounts of several contaminating substances as indicated by reverse-phase high-pressure liquid chromatography and chemical analyses. LTA was purified further by DEAE-cellulose chromatography, using a concentration gradient of sodium chloride in chloroform-methanol-water (1:5:3). Two column peaks of LTA were found to contain phosphate, glycerol, glucose, and fatty acids at molar ratios of 1:1:0.11:0.10 and 1:1:0.09:0.04, respectively. The LTA polymers contained 18 and 22 repeating units of unsubstituted glycerophosphate and two glucose residues. The LTA in one column peak had two fatty acids per molecule, whereas that in the second peak contained only one. The yield of LTA was 1.68 mg per g of cell dry weight or 65 mg per g of phenol-water-extracted material. The specific activity of the LTA preparation was increased 128-fold by the purification scheme as determined by a erythrocyte-binding assay. Reverse-phase high-pressure liquid chromatography may be used for rapid separation of LTA molecules containing different numbers of acyl groups.