bluuebunny/crossref_metadata_2025 huggingface dataset:
Flow microfluorometry analysis of alterations in T-lymphocyte subsets during murine listeriosis
Authors: Watson S R , Redington T J , Miller T B , Bullock W E
Published: January 2020

C57BL/6 mice were infected intravenously with 6 X 10(3) Listeria monocytogenes organisms. As early as day 3 of infection, there was a marked reduction in the number of lymphocytes recovered from the peripheral blood, bone marrow, and thymuses of infected animals. Concomitantly, there was an increase in the number of splenic lymphocytes. By day 14, both the total and differential cell counts were similar in both infected and normal animals. Flow microfluorometric studies comparing the Thy-1.2, Lyt-1, Lyt-2, and surface immunoglobulin (SIg) phenotypes of lymphocytes from normal and infected mice were performed. Between days 3 and 5, there was a decrease in the percentage of Thy-1.2+ cells in the spleens of L. monocytogenes-infected animals. Conversely, the percentages of Lyt-1+, Lyt-2+, and SIg+ cells remained constant. At day 7 of infection, the percentage of Thy-1.2+ splenocytes was within normal limits, and at day 10, the percentage of Thy-1.2+ cells was elevated slightly. The absolute numbers of Thy-1.2+ cells were comparable in both infected and normal animals at early stages (days 3 to 5) of L. monocytogenes infection, but there was a marked elevation of Thy-1.2+ splenocytes at days 7 to 14 of infection. Lyt-1+, Lyt-2+, and SIg+ splenocytes increased in absolute numbers as early as day 3 of infection and were still elevated at day 14. Adrenalectomy before infection had no effect on the results obtained, suggesting that these changes were not mediated by endogenous steroids.
Comparison of Chlamydia psittaci isolates by DNA restriction endonuclease analysis
Authors: McClenaghan M , Herring A J , Aitken I D
Published: January 2020

Preparations of DNA from 12 Chlamydia psittaci isolates and one Chlamydia trachomatis strain were compared by restriction endonuclease analysis. Polyacrylamide gel electrophoresis, followed by silver staining, resulted in optimal resolution of fragments generated by digestion. By this technique, four distinct electropherotypes were demonstrated when ovine abortion, ovine arthritis, and avian and Cal10 strains of C. psittaci were examined. Minor profile differences allowed the discrimination of avian isolates derived from psittacine and columbiforme species, and the Cal10 DNA electropherotype was shown to have features in common with these profiles. However, there were no detectable differences in the DNA patterns of eight ovine abortion isolates.
Distinctive patterns of adherence of enteropathogenic Escherichia coli to HeLa cells
Authors: Scaletsky I C , Silva M L , Trabulsi L R
Published: January 2020

We showed that Escherichia coli strains attach to HeLa cells in two different patterns. In one, the bacteria cover the whole surface of the cell (diffuse adherence), and in the other, attachment is limited to one or a few sites of the cell surface (localized adherence). Among the enteropathogenic strains, serogroups O55, O86, O11ab, O119, O125, O128ab, and O142 usually showed localized adherence when tested in the presence of D-mannose. Localized adherence was not shown either by E. coli strains isolated from urine or by enteroinvasive and enterotoxigenic E. coli strains. Some of these strains showed diffuse adherence. Some strains of serogroups O55, O111, and O119 showed both localized and diffuse adherence in the same preparation. Mannose-resistant adherence was not related to colonization factor antigens.
T-cell-independent elimination of Borrelia turicatae
Authors: Newman K , Johnson R C
Published: January 2020

Mice deficient or deprived of thymus-derived lymphocytes eliminated blood-borne Borrelia turicatae with efficiency comparable to that observed in normal littermates. When challenged with 10(5) borreliae, nude mice had mean (+/- standard deviation) primary spirochetemias lasting 3.1 +/- 0.2 days and mean (+/- standard deviation) peak bacterial counts of 3.0 X 10(7) +/- 0.5 X 10(7) cells per ml of blood; in comparison, heterozygous littermates and normal mice had respective primary spirochetemias lasting 3.4 +/- 0.6 and 3.2 +/- 0.2 days and respective peak counts of 8.0 X 10(7) +/- 1.5 X 10(7) and 5.5 X 10(7) +/- 0.9 X 10(7) bacterial cells per ml of blood. No increased responsiveness to concanavalin A was observed in infected nude mice, indicating the sustained lack of maturate T cells in these animals. Thymectomized and steroid-treated mice were also found to eliminate circulating borreliae with efficiency comparable to that observed in control animals. Irradiation of mice abrogated responsiveness to borreliae, but reconstitution with T-cell-depleted splenocytes restored antibody production. It is proposed that elimination of B. turicatae is mediated by a T-cell-independent immune response mechanism.
Vaccination against Legionella pneumophila: serum antibody correlates with protection induced by heat-killed or acetone-killed cells against intraperitoneal but not aerosol infection in guinea pigs
Authors: Eisenstein T K , Tamada R , Meissler J , Flesher A , Oels H C
Published: January 2020

An aerosol model of Legionella infection has been established in guinea pigs. Infected animals showed growth of Legionella in their lungs, dissemination of organisms to the spleen, development of pneumonia and fever, and weight loss. Vaccination studies using heat-killed or acetone-killed cells were carried out, and guinea pigs were challenged intraperitoneally or by using the aerosol model of infection. Both vaccines were shown to give moderately high levels of protection against intraperitoneal challenge (28 to 145 50% lethal doses). Protection was found to be dose dependent and correlated with antibody levels as measured by enzyme-linked immunosorbent assay to an outer membrane antigen and by indirect immunofluorescence to heat-killed cells. In contrast, the same vaccination regimens that protected against intraperitoneal challenge failed to protect guinea pigs against aerosol challenge with comparable doses of Legionella, despite the presence of serum antibody. The results are discussed in terms of the possible requirements for immunity to aerosolized Legionella, including secretory immunoglobulin or cell-mediated immunity.
Common and isolate-restricted antigens of Anaplasma marginale detected with monoclonal antibodies
Authors: McGuire T C , Palmer G H , Goff W L , Johnson M I , Davis W C
Published: January 2020

Anaplasma marginale-infected erythrocytes were examined for the presence of maturation, isolate-restricted, and isolate-common antigens by indirect immunofluorescence with monoclonal antibodies. A panel of 18 monoclonal antibodies was used; none of the antibodies reacted with Anaplasma ovis, Babesia bigemina, babesia bovis, Trypanosoma brucei, Trypanosoma congolense, or uninfected bovine erythrocytes. Antigens common to all six A. marginale isolates were detected by nine antibodies. Single isolates from Florida, Southern Idaho, Northern Texas, and Virginia and two isolates from Washington state had four patterns of reactivity with a second panel of nine antibodies. Antigenically distinct stages were not detected, as sequential smears taken daily during acute infection had the same pattern of reactivity. The results demonstrate antigenic heterogeneity among isolates of A. marginale and the presence of common antigens. This information allows grouping of different isolates and, more importantly, provides a method for the identification and isolation of common antigens for diagnostic tests.
Capsule reduces adherence of enterotoxigenic Escherichia coli to isolated intestinal epithelial cells of pigs
Authors: Runnels P L , Moon H W
Published: January 2020

Previous reports have demonstrated that heat-stable (A-type) capsule on piliated enterotoxigenic Escherichia coli enhances colonization of enterotoxigenic E. coli in the small intestine and enhances virulence of enterotoxigenic E. coli. In this report, four encapsulated enterotoxigenic E. coli strains and one encapsulated nonenterotoxigenic strain of E. coli and their nonencapsulated mutants were tested for adhesion to isolated intestinal epithelial cells or brush borders from neonatal pigs. The enterotoxigenic E. coli also expressed the K99 pilus antigen. The nonencapsulated mutants of the four enterotoxigenic E. coli adhered in higher numbers than did the encapsulated parental strains. Both the encapsulated and nonencapsulated forms of enterotoxigenic E. coli 431 grown at 18 degrees C (K99 production suppressed) adhered poorly to the isolated cells. The nonenterotoxigenic E. coli 1793 which does not express K99 antigen also adhered poorly in both encapsulated and nonencapsulated forms. Fab fragments of anticapsular immunoglobulin G failed to block the effect of capsule on adherence of strain 431. The results indicated that K99 was the principal mediator of in vitro adhesion of the enterotoxigenic E. coli strains and that capsule impedes the in vitro adhesion. They also suggested that the capsular enhancement of colonization by such strains in vivo probably is by some mechanism other than enhanced adhesion to epithelium.
Chlamydia trachomatis species-specific epitope detected on mouse biovar outer membrane protein
Authors: Stephens R S , Kuo C C
Published: January 2020

A common antigenic determinant on the chlamydial major outer membrane protein was detected on each of the three Chlamydia trachomatis biovars (trachoma, lymphogranuloma venereum, and mouse). This determinant was prominently displayed on the surface of chlamydial strains from both the trachoma and lymphogranuloma venereum biovars. However, detection of this determinant on a mouse biovar strain required denaturation by sodium dodecyl sulfate or periodate oxidation. This determinant provides a definable taxonomic link between the three biovars of C. trachomatis.
Stage-specific variations in lectin binding to Leishmania donovani
Authors: Wilson M E , Pearson R D
Published: January 2020

Visceral leishmaniasis is caused by the dimorphic protozoan Leishmania donovani, which exists as an aflagellar amastigote within mammalian mononuclear phagocytes and as a flagellated extracellular promastigote in its sandfly vector. We have identified four plant lectins that bind to the L. donovani surface, and through these we have documented stage-specific differences in exposed surface carbohydrates. Concanavalin A bound to both promastigotes and amastigotes; binding was inhibited by mannose or alpha-methyl-mannoside, implying a mannose-containing residue on the surface of both parasite stages. Ricinus communis agglutinin, which binds to galactose-containing residues, also bound to both stages and was inhibited by lactose, implying a galactose-containing glycoconjugate on the parasite surface. Two other lectins, wheat germ agglutinin (WGA) and peanut agglutinin (PNA), exhibited stage specificity in their binding characteristics. Amastigotes bound WGA but not PNA. During the process of conversion from the amastigote to the promastigote stage, the WGA-binding glycoconjugate was lost, and a PNA-binding residue was newly displayed. WGA binding was inhibited by N-acetyl-D-glucosamine and was not altered by neuraminidase treatment, suggesting the presence of an exposed N-acetyl-D-glucosamine moiety on the amastigote surface. The PNA binding site is known to accommodate the oligosaccharide beta-D-galactose-(1----3)-N-acetyl-D-galactosamine; in our system, PNA may have identified an internal rather than a terminal galactose on the promastigote surface. Localized binding of WGA and PNA to the surface of intermediate phases of the parasite suggested inhomogeneous and changing surface characteristics during conversion from amastigote to promastigote stages. This evolution of L. donovani surface glycoconjugates may be important in the adaptation of the organism to its divergent mammalian host and arthropod vector environments.
Comparison of sodium dodecyl sulfate-polyacrylamide gel electrophoresis profiles and antigenic relatedness among outer membrane proteins of 49 Brucella abortus strains
Authors: Verstreate D R , Winter A J
Published: January 2020

Outer membrane proteins were solubilized from 49 strains of Brucella abortus by sequential extraction of physically disrupted cells with N-lauroylsarcosinate and a dipolar ionic detergent (Verstreate et al., Infect. Immun. 35:979-989, 1982). The strains tested included standard agglutination test strain 1119, virulent strain 2308, and eight reference strains representing each of the biotypes; the remainder were isolates from cattle in North America with natural infections and included biotypes 1, 2, and 4. Three principal protein groups with apparent molecular weights of 88,000 to 94,000 (group 1), 35,000 to 40,000 (group 2, now established as porins [Douglas et al., Infect. Immun. 44:16-21, 1984]), and 25,000 to 30,000 (group 3) were observed in every strain. Some variability in banding patterns occurred among strains, but intrastrain variation was sufficient to preclude the use of sodium dodecyl sulfate-polyacrylamide gel electrophoresis profiles of outer membrane proteins for differentiating among strains of B. abortus. One antigen ([b]) was shared among the porin proteins, and three others ([c], ([d], and ([ e]) were shared among the group 3 proteins of all of the strains tested, indicating that these relationships are probably species wide. These results suggest that it may be possible to use outer membrane proteins from a representative strain of B. abortus in a vaccine for species-wide immunization.
Lactose transport in Streptococcus mutans: isolation and characterization of factor IIIlac, a specific protein component of the phosphoenolpyruvate-lactose phosphotransferase system
Authors: Vadeboncoeur C , Proulx M
Published: January 2020

The transport of lactose in Streptococcus mutans is mediated via an inducible phosphoenolpyruvate-lactose phosphotransferase system. This system requires for catalytic activity a membrane fraction (enzyme II), two general proteins called enzyme I and HPr, and a soluble specific protein termed factor IIIlac. This protein factor was purified from S. mutans ATCC 27352 by chromatographies on DEAE-cellulose, hydroxylapatite, Ultrogel AcA 34, and phosphocellulose. The purified protein migrated as a single band with a molecular weight of 10,000 on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and urea. The molecular weight calculated from the amino acid composition was 10,541. Gel filtration of the native protein gave a molecular weight of 41,500. Its isoelectric point was ca. 4.70. A specific antiserum was prepared against purified factor IIIlac. Immunodiffusion experiments revealed that only cellular extracts from lactose-grown cells contained factor IIIlac. A cross-reaction was observed with all of the S. mutans strains tested as well as with Streptococcus sanguis 10556, Streptococcus lactis 11454, and Staphylococcus aureus 6538. No precipitin band was observed with extracts of Streptococcus salivarius, Streptococcus faecalis, Lactobacillus casei, and Bacillus subtilis.
Ingestion and intracellular survival of Brucella abortus in human and bovine polymorphonuclear leukocytes
Authors: Riley L K , Robertson D C
Published: January 2020

Bovine polymorphonuclear leukocytes (PMNs) were found to be significantly more bactericidal than human PMNs against a smooth-intermediate strain of Brucella abortus (45/0), whereas there was no difference in bactericidal activity of the two kinds of PMNs against a rough strain of B. abortus (45/20). Electron microscopy of thin sections of PMNs revealed that both strains of B. abortus were readily ingested; however, the extent of degranulation was significantly less than in PMNs incubated with an extracellular parasite, Staphylococcus epidermidis. Amounts of myeloperoxidase and lactoferrin released through exocytosis by PMNs incubated with S. epidermidis were 4.7- and 1.2-fold greater, respectively, than those released from PMNs incubated with B. abortus 45/0. When azurophil and specific granules were isolated after incubation of PMNs with either B. abortus 45/0 or S. epidermidis, results showed that the extent of degranulation by both types of granules was greater in PMNs incubated with S. epidermidis than in those incubated with B. abortus 45/0. Amounts of degranulation by azurophil and specific granules were similar in PMNs incubated with either the smooth-intermediate strain 45/0 or the rough strain 45/20. Degranulation was not stimulated when glutaraldehyde-killed strain 45/0 was substituted for viable cells. These data suggest that B. abortus does not stimulate an effective level of degranulation after ingestion, as observed with extracellular parasites, and that the smooth intermediate strain 45/0 is more resistant to intraleukocytic killing system than the rough strain 45/20.
Cytotoxicity of human peripheral blood and colostral leukocytes against Shigella species
Authors: Morgan D R , DuPont H L , Gonik B , Kohl S
Published: January 2020

We examined the ability of human peripheral blood leukocytes to kill strains of Shigella sonnei and Shigella flexneri by using a modified bactericidal assay. Antibody-dependent cellular cytotoxicity (ADCC) was demonstrated in the presence of specific rabbit immune serum directed against S. sonnei. With peripheral blood leukocytes from adults, ADCC was found only in the mononuclear cell and purified lymphocyte populations. Monocyte-macrophages and polymorphonuclear leukocytes were unable to demonstrate ADCC. Lymphocyte ADCC, which was not affected by the addition of phenylbutazone (an inhibitor of phagocytosis), was mediated by a non-T, Fc receptor-positive, HNK-1- cell. ADCC (using antiserum directed against virulent S. sonnei) was demonstrated against virulent S. sonnei but not against virulent S. sonnei or virulent S. flexneri. In contrast to leukocytes from adults, both mononuclear and polymorphonuclear cells from neonatal cord blood and from a patient with chronic granulomatous disease mediated anti-Shigella ADCC. Breast milk leukocytes (BMLs) collected 1 to 3 days postpartum were used as effector cells against virulent S. sonnei. The entire BML population, BMLs which did not adhere to plastic and BMLs which passed through nylon wool columns mediated both natural killer cytotoxicity and ADCC. In paired experiments, natural killer cytotoxicity and ADCC were significantly lower (30 to 45% inhibition) but not ablated, when phenylbutazone was added to BMLs and nylon wool-purified BMLs (P less than 0.05). These experiments suggest that colostral leukocytes mediated both extracellular and intracellular bacteriolysis in the presence and absence of specific antiserum. These mechanisms may be active in vivo in protection against shigellosis.
Blastomyces dermatitidis chemotactic factor: kinetics of production and biological characterization evaluated by a modified neutrophil chemotaxis assay
Authors: Thurmond L M , Mitchell T G
Published: January 2020

Chemotactic activity for human polymorphonuclear neutrophils (PMNs) was detectable in culture filtrates (CFs) of Blastomyces dermatitidis and may have influenced the pathogenesis of blastomycosis. Production of this chemotaxin depended upon culture age and medium; peak levels were achieved after incubation for 17 days or more in minimal essential medium. This factor was also chemotactic for human monocytes. CF was temperature stable even after treatment at 100 degrees C for 60 min. The activity was stable under alkaline conditions but was destroyed below pH 7. Dialyzed, chemotactically active CF contained approximately 60 micrograms of carbohydrate per ml; total protein was estimated to be less than 0.8 micrograms/ml. Preincubation of PMNs with CF or N-formyl-L-methionyl-L-leucyl-L-phenylalanine deactivated their chemotactic response to each agent, whereas the chemotactic response to zymosan-activated serum was not affected. In addition, deactivation with N-formyl-L-methionyl-L-leucyl-L-phenylalanine reduced the response to CF. Pretreatment of CF with PMNs decreased chemotactic activity, which may reflect binding of the chemotaxin molecules to PMN receptors. A modified chemotaxis assay was developed in which commercial, disposable multiwell plates are used. This method was rapid, efficient, and inexpensive and permitted the assay of larger numbers of samples than was previously feasible with conventional chemotaxis methods.
Comparative observations of fever and associated clinical hematological and blood biochemical changes after intravenous administration of staphylococcal enterotoxins B and F (toxic shock syndrome toxin-1) in goats
Authors: Van Miert A S , Van Duin C T , Schotman A J
Published: January 2020

The present investigation was undertaken to examine the characteristics of purified toxic shock syndrome toxin-1 (staphylococcal enterotoxin F) given intravenously to dwarf goats (dose, 0.02 to 20 micrograms kg-1). Rectal temperature, heart rate, rumen motility, plasma zinc and iron concentrations, and certain other blood biochemical and hematological values were studied and compared with the changes seen after intravenous administration of staphylococcal enterotoxin B (dose, 0.02 to 0.5 micrograms kg-1). Similar changes such as fever, tachycardia, inhibition of rumen contractions, drop in plasma zinc and iron concentrations, lymphopenia, and a decrease in serum alkaline phosphatase activity were observed. In contrast to the effects of toxic shock syndrome toxin-1, staphylococcal enterotoxin B induced colic, watery diarrhea with pseudomembranes, hemoconcentration, and a more pronounced increase in blood urea nitrogen. The results obtained demonstrate that (i) in the goat staphylococcal enterotoxin B is much more potent than toxic shock syndrome toxin-1 and (ii) the goat is a useful model to study the gastro-intestinal effects caused by staphylococcal enterotoxin B. The present finding that no clear relationship could be found between the temperature response and the alterations in zinc and iron levels in plasma support the theory that the febrile reactions and the changes in plasma trace metals are mediated by different polypeptides released by activated macrophages.
Mechanisms of invasion and replication of the intracellular stage in Trypanosoma cruzi
Authors: McCabe R E , Remington J S , Araujo F G
Published: January 2020

Amastigotes obtained from spleens of mice infected with different strains of Trypanosoma cruzi were examined for their ability to invade macrophages and L929 cells and to initiate infection in mice. Both types of cells were readily invaded by organisms of the strains Y, MR, and Tulahuen. Organisms of the CL strain were taken up by both types of cells at a rate that was significantly lower than that for organisms of the other strains. However, all strains multiplied intracellularly. Activated macrophages inhibited the replication of intracellular organisms. Treatment of normal macrophages with cytochalasin B, trypsin, chymotrypsin, or pronase significantly inhibited phagocytosis, but the inhibitory effect was reversible. Mice injected with spleen amastigotes developed parasitemia and died of the infection. These results demonstrate that spleen amastigotes are able to infect, survive, and replicate within professional and nonprofessional phagocytes and to initiate infection in vivo. Interiorization of spleen amastigotes is by phagocytosis and is dependent upon a protease-sensitive receptor(s) on the cell surfaces of host macrophages.
Purification of dermonecrotic toxin from a sonic extract of Pasteurella multocida SP-72 serotype D
Authors: Nakai T , Sawata A , Tsuji M , Samejima Y , Kume K
Published: January 2020

A procedure was developed to purify dermonecrotic toxin (DNT) from a sonic extract of a serotype D strain of Pasteurella multocida. Sonic extract containing DNT was applied to a DEAE-Sephacel column and eluted by a linear gradient of NaCl. Upon rechromatographing, fractions with dermonecrotic activity for guinea pigs were applied on a second Sephacel column, and a pooled fraction with the toxic activity was filtered through a Sephadex G-200 column. Pooled fractions with the toxic activity were subjected to polyacrylamide disc gel electrophoresis (PAGE), and the toxic substance was eluted from each sliced gel. Eluted fractions with the toxic activity were rechromatographed on a second Sephadex G-200 column, and a pooled fraction with high dermonecrotic activity was referred to as a purified DNT. The activity of purified DNT was increased by 1,000 times, and the average yield was about 1.8%. The purified DNT was homogeneous as determined by Ouchterlony double immunodiffusion, crossed immunoelectrophoresis, and thin-layer isoelectric focusing in polyacrylamide gels and gave a single band on PAGE and sodium dodecyl sulfate-PAGE. The molecular weight of the toxin was ca. 160,000 as determined by sodium dodecyl sulfate-PAGE. The isoelectric point of the toxin was ca. 4.7 to 4.8. Amino acid analysis of the purified DNT revealed that the toxin was composed of characteristically high proportions of glutamic acid, aspartic acid, glycine, proline, alanine, and leucine. The minimal necrotizing dose of the toxin was about 1 ng of protein, and the 50% lethal dose per mouse was 0.2 micrograms. The purified DNT was heat labile and sensitive to inactivation by trypsin, Formalin, and glutaraldehyde.
Expression of lipopolysaccharide O antigen in Escherichia coli K-12 hybrids containing plasmid and chromosomal genes from Shigella dysenteriae 1
Authors: Hale T L , Guerry P , Seid R C , Kapfer C , Wingfield M E , Reaves C B , Baron L S , Formal S B
Published: January 2020

The requirement for both plasmid and chromosomal genes in the biosynthesis of Shigella dysenteriae 1 lipopolysaccharide O antigen was demonstrated in Escherichia coli-Shigella hybrids. A 6-megadalton S. dysenteriae 1 plasmid, designated pWR23, was phenotypically tagged with the Tn3 ampicillin-resistance transposon. The tagged plasmid, designated pWR24, was transferred by transformation or conjugal mobilization to a rough E. coli K-12 recipient. Although the resultant hybrids were agglutinated in S. dysenteriae 1 antiserum, they did not remove all of the anti-Shiga agglutinins in absorption experiments. Modified lipid A core structure was detected in these hybrids, but Shiga O antigen was not expressed. When the his+ locus of the S. dysenteriae 1 chromosome was transferred by transduction to E. coli K-12 containing pWR24, complete Shiga O antigen was expressed. Lipopolysaccharide extracted from these hybrids was indistinguishable chemically, electrophoretically, and serologically from native S. dysenteriae 1 lipopolysaccharide.
Haemophilus influenzae type b isolates show antigenic variation in a major outer membrane protein
Authors: Erwin A L , Kenny G E
Published: January 2020

Antigenic variation of the outer membrane proteins among isolates of Haemophilus influenzae was examined by immunoblotting. Rabbit antisera were raised against six strains of H. influenzae type b and tested against outer membrane preparations of 50 isolates. The principal outer membrane band was not reactive on immunoblotting, so its antigenic heterogeneity could not be examined. Most of the other outer membrane proteins shared common determinants among all strains tested. Absorption of serum with heterologous bacteria removed antibody to nearly all proteins, confirming the extensive cross-reactivity among isolates. The greatest antigenic variation was seen in one major outer membrane band, a heat-modifiable, Zwittergent-soluble protein with a molecular weight of 49,000 to 51,000. One antiserum reacted with the 49,000-to-51,000-molecular-weight protein of the homologous isolate only; the remaining five antisera showed differing patterns of reactivity with heterologous 49,000-to-51,000-molecular-weight proteins. We were able to divide the 50 H. influenzae isolates into 13 antigenic groups based on their reaction patterns. The antigenic groupings may provide an epidemiological tool for studying the prevalence and transmission of strains of H. influenzae type b.
Binding of pseudomonal leukocidin to rabbit polymorphonuclear leukocytes
Authors: Hirayama T , Noda M , Matsuda F , Nagamori M , Kato I
Published: January 2020

The leukocytotoxic toxin pseudomonal leukocidin, produced by Pseudomonas aeruginosa, was radioiodinated with chloramine-T reagent. Binding of [125I]leukocidin to rabbit polymorphonuclear leukocytes was found to be concentration dependent at 37 degrees C. A Scatchard plot of binding data was linear, indicating that leukocidin binds to a single population of sites. The dissociation constant, KD, was calculated from the Scatchard plot to be 2.5 X 10(-7) M, and the number of binding sites per leukocyte was approximately 4.4 X 10(5). Unlabeled leukocidin or antileukocidin antibody reduced the binding of [125I]leukocidin to the leukocytes. A leukocidin-binding protein was extracted from rabbit polymorphonuclear leukocytes with Triton X-100 and purified by leukocidin-Sepharose 4B affinity column chromatography. Approximately 60 micrograms of binding protein was obtained from 8.1 mg of material extracted from the leukocytes. The binding protein had a molecular weight of about 50,000 as shown by sodium dodecyl sulfate-polyacrylamide slab gel electrophoresis and staining with silver nitrate. Under nondenaturing conditions, its molecular weight was also about 50,000, as shown by gel filtration-Sephadex G-200 chromatography. The 50,000-dalton protein purified in this way from rabbit polymorphonuclear leukocytes competitively inhibited the binding of leukocidin to leukocytes and inactivated leukocidin activity. With equimolar amounts of the 50,000-dalton protein and leukocidin, up to 90% inactivation of leukocidin was observed.
Human opsonins to meningococci after vaccination
Authors: Halstensen A , Haneberg B , Frøholm L O , Lehmann V , Frasch C E , Solberg C O
Published: January 2020

Two groups of volunteers were immunized with either a serogroup A plus C meningococcal polysaccharide vaccine or a combined serogroup B polysaccharide-serotype 2 protein vaccine. Serum opsonin responses were measured by chemiluminescence of polymorphonuclear leukocytes exposed to opsonized live meningococci. Two of the six volunteers immunized with the A plus C vaccine had an increase in serum opsonins to group A meningococci, four responded to group C meningococci, and none to group B meningococci. Five other volunteers who were immunized with the combined group B polysaccharide-serotype 2 protein vaccine responded with an increase in serum opsonins to group B meningococci of two different protein serotypes, as well as to a group C-serotype 2 meningococcal strain. Although no booster effect was observed after a second dose of the combined vaccine, both the polysaccharide and the protein components appear to be able to stimulate an opsonin response.
Effects of oxygen on pyruvate formate-lyase in situ and sugar metabolism of Streptococcus mutans and Streptococcus sanguis
Authors: Yamada T , Takahashi-Abbe S , Abbe K
Published: January 2020

The strictly anaerobic metabolism of sugar in strains of Streptococcus mutans and Streptococcus sanguis was studied because deep layers of dental plaque are strictly anaerobic. Galactose-grown cells of these streptococcal strains had higher pyruvate formate-lyase activity than did glucose-grown cells. Among these strains, two strains of S. mutans had a significantly higher pyruvate formate-lyase activity than did the others. This enzyme is extremely sensitive to oxygen, and even in situ the enzyme was inactivated by exposure of the cells to air. Lactate was less than 50% of the total end product of the strictly anaerobic incubation of the galactose-grown cells of S. mutans with excess glucose, and a significant amount of formate, acetate, and ethanol was produced through the catalysis of pyruvate formate-lyase. But the cells exclusively produced lactate when exposed to air for 2 min before the anaerobic incubation. The metabolism of sorbitol by S. mutans was seriously impaired by the exposure of the cells to oxygen, and the metabolic rate was reduced to less than 1/20 of that found under strictly anaerobic conditions because of the inactivation of pyruvate formate-lyase. S. sanguis produced a smaller amount of the volatile products from glucose than did S. mutans because of the low level of pyruvate formate-lyase. However, the pyruvate formate-lyase in situ in S. sanguis was less sensitive to oxygen than was that in S. mutans. Because of this low sensitivity, S. sanguis metabolized glucose more rapidly under aerobic conditions, whereas the rates of the aerobic and anaerobic metabolism of glucose by S. mutans were similar, which suggests that S. mutans rather than S. sanguis can sustain the rapid sugar metabolism in the deep layers of dental plaque.
Defective neutrophil and monocyte motility in patients with early onset periodontitis
Authors: Page R C , Sims T J , Geissler F , Altman L C , Baab D A
Published: January 2020

Several studies have documented suppressed polymorphonuclear neutrophil (PMN) chemotaxis in most patients with juvenile periodontitis. In contrast, data regarding PMN chemotaxis in patients with rapidly progressive periodontitis are very limited, and monocyte (MN) chemotaxis and random migration of PMNs or MNs from these patients have not been studied previously. Accordingly, we examined cell motility of PMNs and MNs from 27 patients with rapidly progressive periodontitis, 5 patients with juvenile periodontitis, and 37 normal control subjects by using a microchamber technique and the synthetic peptide N-formylmethionyl-leucyl-phenylalanine (FMLP) as the chemoattractant. As a group, PMNs and MNs from patients with rapidly progressive periodontitis manifested significantly enhanced random migration relative to control cells (P less than 0.001), suppressed directed migration (chemotaxis) at FMLP doses of 10(-9) and 10(-8) M (P less than 0.05), and enhanced directed migration at a dose of 10(-6) M FMLP (P less than 0.01). In contrast, PMNs from patients with juvenile periodontitis exhibited normal random migration, and directed migration was significantly suppressed at all doses of FMLP tested (P less than 0.05). An abnormality of either PMN or MN motility was observed in 26 of 27 patients with rapidly progressive periodontitis. Enhanced random migration was seen in PMNs in 63%, MNs in 39%, and both cell types in 26% of the patients. Suppressed chemotaxis was seen in PMNs in 85%, in MNs in 74%, and in both cell types in 69% of the patients. The prevalence and magnitude of abnormalities in motility were somewhat lower in treated than in untreated patients. Thus, most, if not all, of this subgroup of patients with early onset, highly destructive periodontitis have abnormalities in PMN or MN motility, and these defects may differ from those seen in cells from patients with the juvenile form of the disease.
An adhesive protein capsule of Escherichia coli
Authors: Orskov I , Birch-Andersen A , Duguid J P , Stenderup J , Orskov F
Published: January 2020

The nature of the adhesive capacity of three hemagglutinating Escherichia coli strains that had earlier been described as nonfimbriated was studied. The strains that were isolated from human disease adhered to human buccal and urinary tract epithelial cells, an adhesion that was not inhibited by D-mannose. By crossed immunoelectrophoresis it was shown that the three strains produced a common antigen, Z1, developed after growth at 37 degrees C but not 18 degrees C. One of the strains produced an additional antigen, Z2, of almost the same electrophoretic mobility in crossed immunoelectrophoresis. A mutant of this strain deficient of its polysaccharide K antigen had maintained the adhesive capacity, indicating that the K antigen was not responsible for adhesion. A further mutant of the acapsular mutant produced a strongly reduced amount of the Z antigens and had lost the ability to adhere. The Z1 (and Z2?) antigens were therefore deemed to be responsible for adhesion. In sodium dodecyl sulfate-polyacrylamide gel electrophoresis of extracts of cells of the three strains, a heavy Coomassie-blue stained line was seen, indicating the presence of a protein subunit of molecular weight slightly above 14,400. By immunoblotting with absorbed antiserum, it was shown that this protein was the same as that detected by crossed immunoelectrophoresis. Protease from Streptomyces griseus, but not trypsin, digested the protein. Heating to 100 degrees C did not affect it. By immunoelectron microscopy of embedded and sectioned bacteria that had first been treated with specific antisera and ferritin-labeled antirabbit immunoglobulin, the protein adhesin-antibody complex was found to surround the bacteria as a heavy capsule. After negative staining with uranylacetate (pH approximately 4), the capsule appeared as a mesh of very fine filaments. The possible role of this capsule in the pathogenesis of disease is discussed.
Enterotoxin production, presence of colonization factor antigen I, and adherence to HeLa cells by Escherichia coli O128 strains belonging to different O subgroups
Authors: Guth B E , Silva L M , Scaletsky I C , Toledo M R , Trabulsi L R
Published: January 2020

Strains of three subgroups of Escherichia coli O128 were studied. Enterotoxin production was observed in 30 (91%) O128ac strains, whereas strains of subgroups O128ab and O128ad were not toxigenic. CFA/I was only found in two serotypes of subgroup O128ac, all of them producing heat-stable enterotoxin except for one which produced both toxins. None of the strains studied produced CFA/II. In a binding test with HeLa cells, localized adherence was found only in strains of subgroup O128ab; diffuse adherence occurred in strains of subgroup O128ac. As flagellar antigens were specific in subgroups ab and ac and toxin production was observed only in subgroup ac, the present results suggest that subgroup and serotype are useful markers for O128 strains that are enterotoxigenic or enteropathogenic.
Intracellular acid phosphatase content and ability of different macrophage populations to kill Nocardia asteroides
Authors: Black C M , Beaman B L , Donovan R M , Goldstein E
Published: January 2020

It has been reported that the activity of lysosomal acid phosphatase decreases inversely with numbers of ingested virulent Nocardia spp. in normal murine peritoneal and alveolar macrophages. These studies suggested that this relationship correlated with the effectiveness of these macrophage populations in killing Nocardia asteroides. Experiments were designed to determine if acid phosphatase activity is affected by infection with N. asteroides in four different macrophage populations isolated from normal and nocardia-immunized mice. Macrophages were also tested simultaneously for their ability to kill N. asteroides. Peritoneal, alveolar, and splenic macrophages and Kupffer cells were infected in vitro with strains of N. asteroides of differing virulence. Uptake and killing assays were performed. Acid phosphatase levels and numbers of intracellular nocardiae were quantitated in the same macrophages, using a computer-assisted cytophotometry system. Acid phosphatase activity decreased inversely with numbers of intracellular nocardiae in macrophages that could not kill or inhibit this pathogen. Acid phosphatase activity was not significantly changed in macrophages that inhibited growth of, but did not kill, N. asteroides, whereas activity was increased or enhanced in macrophages that killed most of the ingested nocardiae. The order of nocardicidal effectiveness (and resistance to enzyme activity reduction with infection) for normal macrophages was splenic greater than peritoneal greater than alveolar greater than Kupffer. In contrast, the order of these two parameters for macrophages isolated from immunized mice was Kupffer greater than peritoneal greater than alveolar greater than splenic. These results demonstrate that lysosomal acid phosphatase activity is an effective marker of the ability of macrophages to inhibit growth of and kill N. asteroides and that macrophages isolated from different anatomical sites differ functionally from each other with respect to nocardicidal and acid phosphatase activities.
Response of Neisseria gonorrhoeae to iron limitation: alterations in expression of membrane proteins without apparent siderophore production
Authors: West S E , Sparling P F
Published: January 2020

For acquisition of iron, an essential nutrient, most microorganisms produce siderophores (low-molecular-weight iron-chelating compounds) and membrane proteins to serve as receptors for the iron-siderophore complexes. The gonococcus does not appear to produce a siderophore, since the quantity of siderophore detected by bioassays of culture supernatants from strains F62 and FA19 was never greater than the amount present in the uninoculated medium. Iron limitation of the laboratory strains F62 and FA19 and 12 recent clinical isolates resulted in the expression of several iron-repressible membrane proteins. The expression of proteins in the apparent molecular weight range of 70,000 to 100,000 was strain dependent. All strains expressed 36,000-dalton (36K) and 19.5K proteins. FA19 and F62 were also grown in medium containing iron sources commonly encountered in vivo (i.e., transferrin, lactoferrin, hemoglobin, or hemin). Comparison of growth rates indicates that transferrin and lactoferrin were more readily utilized as iron sources than hemin and hemoglobin were. Expression of the iron-repressible proteins varied depending upon the iron source. Fewer iron-repressible proteins were observed when cells were supplied with transferrin or lactoferrin than when the cultures were grown with either hemin or hemoglobin. The 36K protein was expressed with all four iron sources.
Isolation and separation of physicochemically distinct fimbrial types expressed on a single culture of Escherichia coli O7:K1:H6
Authors: Karch H , Leying H , Büscher K H , Kroll H P , Opferkuch W
Published: January 2020

The fimbrial (pili) profile of a single strain of Escherichia coli O7:K1:H6 (WF96) was evaluated. Fimbriae were isolated by sucrose density gradient ultracentrifugation, purified from flagellae by the use of 0.4% sodium dodecyl sulfate (SDS), and separated into distinct fimbrial types. Analysis of the purified WF96 fimbriae by SDS-polyacrylamide gel electrophoresis revealed two polypeptide bands with molecular weights of 16,000 and 21,000. Treatment of the fimbrial mixture with saturated guanidine hydrochloride resulted in the appearance of a third band with a molecular weight of 19,500. The relative susceptibilities of the WF96 fimbrial types to disrupting chemicals (octyl-glucoside, urea, SDS, and guanidine hydrochloride) were assessed by exposure of the fimbrial mixture to each agent, separation of the depolymerized fimbriae from intact fimbriae by gel filtration on Sepharose CL-4B, and identification of the disaggregated fimbrial types by SDS-polyacrylamide gel electrophoresis of column fractions. The physicochemical heterogeneity of the three fimbrial types coexpressed on WF96 was exploited to develop a method for separation of individual fimbriae.
Synergistic rotavirus and Escherichia coli diarrheal infection of mice
Authors: Newsome P M , Coney K A
Published: January 2020

Experimental or naturally acquired subclinical infections with rotavirus caused a significant increase in mortality of infant mice after challenge with enterotoxigenic Escherichia coli B44.
Oxidation of microbial iron-sulfur centers by the myeloperoxidase-H2O2-halide antimicrobial system
Authors: Rosen H , Klebanoff S J
Published: January 2020

Myeloperoxidase, H2O2, and a halide (chloride, bromide, or iodide) form a potent microbicidal system that contributes to the antimicrobial activity of neutrophils. The mechanism of toxicity is not completely understood. Powerful oxidants are formed that presumably attack the microbe at a variety of sites. Among the consequences of this attack is the release of a large proportion of 59Fe of prelabeled organisms. We report here that the myeloperoxidase-H2O2-halide system oxidizes the iron-sulfur centers of model compounds (spinach ferredoxin) and intact microorganisms (Escherichia coli) with the loss of labile sulfide. The oxidation of the iron-sulfur centers of ferredoxin was measured by the fall in absorbance at 420 nm (bleaching) and by the loss of 5,5'-dithiobis-(2-nitrobenzoic acid) reducing activity. The latter compound is a sulfhydryl reagent that is reduced by ferredoxin labile sulfide during denaturation. The oxidation of E. coli iron-sulfur centers by the peroxidase system was determined by the loss of labile sulfide content, as measured by the release of H2S by acid and its reaction with zinc acetate to form ZnS. The halides were effective as components of the peroxidase system in the order I greater than Br greater than Cl. The oxidation of E. coli iron-sulfur centers by the peroxidase system was rapid and preceded the loss of viability. Gentamicin, at a concentration which produced a loss of viability comparable to that of the peroxidase system, did not cause a loss of labile sulfide from E. coli, suggesting that labile sulfide loss is not a nonspecific reflection of the loss of viability, but a direct consequence of the action of the myeloperoxidase system. The oxidation of iron-sulfur centers in microorganisms by the myeloperoxidase-H2O2-halide system may contribute to the death of the organism.
Biochemical and immunobiological properties of lipopolysaccharide (LPS) from Bacteroides gingivalis and comparison with LPS from Escherichia coli
Authors: Koga T , Nishihara T , Fujiwara T , Nisizawa T , Okahashi N , Noguchi T , Hamada S
Published: January 2020

Lipopolysaccharides (LPSs) were isolated from Bacteroides gingivalis and Escherichia coli by the phenol-water and butanol-water procedures. The phenol-water-extracted LPS from B. gingivalis 381 was composed of 46% carbohydrate, 23% hexosamine, 18% fatty acid, and 5% protein. The major component sugars of this preparation were glucose, glucosamine, rhamnose, galactose, galactosamine, and mannose, and their molecular ratio was 1:0.9:0.7:0.6:0.6:0.4, respectively. Neither heptose nor 2-keto-3-deoxyoctonate was detected. The butanol-water-extracted LPS from this strain was composed of 76% glucose, 7% fatty acid, and 13% protein, and it was associated with a number of polypeptides (13 to 56 kilodaltons). The main fatty acid of both LPS preparations was palmitic acid. It was found that biological activities of LPS from B. gingivalis were comparable to those of LPS from E. coli in terms of activation of the clotting enzyme of Limulus amebocyte lysate, mitogenicity, polyclonal B cell activation, and stimulation of interleukin 1 production in BALB/c mice. Furthermore, LPS-nonresponsive C3H/HeJ spleen cells were found to yield good mitogenic responses to both phenol-water-extracted LPS and butanol-water-extracted LPS from B. gingivalis or butanol-water-extracted LPS from E. coli. On the other hand, spleen cells from LPS-responsive C3H/HeN mice responded well to all these LPS preparations.