bluuebunny/crossref_metadata_2025 huggingface dataset:
Enhancement of endotoxin lethality and generation of anaphylactoid reactions by lipopolysaccharides in muramyl-dipeptide-treated mice
Authors: Takada H , Galanos C
Published: January 2020

Intravenous injection of muramyl dipeptide (MDP) and Salmonella lipopolysaccharides (LPS) enhanced lethal toxicity of the LPS in C57BL/6 mice. This was true for S (smooth)- and R (rough)-form LPS and free lipid A. Enhancement of toxicity was maximum when the LPS was administered 4 h after MDP, at which time the lethal doses for 50% of mice of S- and R-form LPS and free lipid A were between 1 and 10 micrograms, compared with more than 100 micrograms in normal animals. This sensitization was absent in endotoxin-resistant C3H/HeJ mice. Lethality usually commenced 15 h after LPS injection and was complete after 72 h. Higher doses of some S-form LPS (100 micrograms or more) administered 4 h after MDP led to a strong anaphylactoid reaction within 10 to 20 min of injection, with lethal outcomes in less than 1 h after LPS administration. This early anaphylactoid reaction was observed for various mouse strains, including LPS-resistant C3H/HeJ mice, but it was very weak or completely absent with R-form LPS or free lipid A even in concentrations of up to 1,000 micrograms. A strong anaphylactoid reaction comparable to that seen with S-form LPS was also obtained, after MDP treatment, with an LPS of low toxicity prepared from Bacteroides gingivalis. It is noteworthy that oral administration of MDP also contributed to the anaphylactoid reaction and enhanced the late-phase lethality of LPS. The present findings strongly suggest that the early- and late-phase reactions induced by MDP and LPS are caused by different mechanisms.
Use of synthetic antigens to determine the epitope specificities of monoclonal antibodies against the 3-deoxy-D-manno-octulosonate region of bacterial lipopolysaccharide
Authors: Brade L , Kosma P , Appelmelk B J , Paulsen H , Brade H
Published: January 2020

Mouse monoclonal antibodies were raised against heat-killed bacteria of the Re mutant R595 of Salmonella minnesota and characterized by the passive hemolysis and passive hemolysis inhibition tests and by double immunodiffusion experiments using lipopolysaccharide (LPS) from different rough mutants of S. minnesota and synthetic antigens. The latter were copolymerization products of acrylamide with the alpha- and beta-allylglycosides of 3-deoxy-D-manno-octulosonic acid (KDO) and the alpha-2,4-linked KDO disaccharide [poly-alpha-KDO, poly-beta-KDO, and poly-(alpha-KDO)2, respectively], and sodium (3-deoxy-D-manno-octulopyranosyl)onate-(2----6)-(2-deoxy-2-[ (R)-3- hydroxytetradecanoylamino]- beta-D-glucopyranosyl)-(1----6)-(2-deoxy-2-[(R)-3-hydroxytetradecanoy lam ino]-D-glucose) [alpha-KDO-(GlcNhm)2], representing a part structure of Re LPS. One antibody (clone 20, immunoglobulin M) was found to recognize a terminal alpha-linked KDO residue, since it reacted in the passive hemolysis assay with alpha-KDO-(GlcNhm)2 and all LPS tested, it was inhibited by all synthetic antigens containing alpha-linked KDO residues, and it gave a reaction of identity with poly-alpha-KDO and poly-(alpha-KDO)2 in double immunodiffusion experiments. A second antibody (clone 25, immunoglobulin G3) was identified as specific for an alpha-2,4-linked KDO disaccharide, since it reacted in immunodiffusion exclusively with synthetic poly-(alpha-KDO)2 and not with the monosaccharide derivatives in either anomeric configuration, and it was inhibited only with poly-(alpha-KDO)2 and with LPS from S. minnesota R595 (Re) and R345 (Rb2). The reaction of this antibody with R345 LPS is attributed to the quantitative substitution with KDO disaccharide present as a side chain, which is not present in stoichiometric amounts in the other LPS.
Conservation of epitopes in the oligosaccharide portion of the lipooligosaccharide of Haemophilus influenzae type b
Authors: Gulig P A , Patrick C C , Hermanstorfer L , McCracken G H , Hansen E J
Published: January 2020

The antigenic characteristics of the lipooligosaccharide (LOS) of Haemophilus influenzae type b (Hib) were examined in strains obtained over an extended period of time. These Hib strains were isolated from patients with systemic Hib disease in Dallas, Tex., over a 20-year period and in New York City between 1941 and 1956. The antigenic characteristics of the LOS of these Hib strains were examined by using a set of four murine monoclonal antibodies directed against epitopes present in the oligosaccharide portion of the LOS molecule. The same basic set of LOS antigenic determinants that is expressed by recent Hib isolates was also found to be present in this collection of Hib strains spanning a 40-year period. Some variation with time was detected in the distribution of the systemic disease isolates among four Hib LOS antigenic groups; however, only 2 of 188 Hib isolates failed to react with a set of two LOS-specific monoclonal antibodies. Therefore, little variation has occurred among Hib strains with regard to the LOS epitopes defined by these monoclonal antibodies over a considerable period of time.
Plasmid-mediated early killing of eucaryotic cells by Shigella flexneri as studied by infection of J774 macrophages
Authors: Clerc P L , Ryter A , Mounier J , Sansonetti P J
Published: January 2020

In Shigella flexneri a 220-kilobase plasmid encodes the ability to invade nonprofessional phagocytes by a mechanism similar to phagocytosis. In this report, the continuous macrophage cell line J774 was used to study the intracellular fate of both invasive and noninvasive strains. pWR100, the virulence plasmid of S. flexneri serotype 5, mediated very efficient and rapid killing of J774 macrophages, as measured by cellular detachment and uptake of trypan blue. For this to occur, the bacteria had to be within the cells, since the macrophages were protected by cytochalasin D. A battery of strains differing in their levels of Shiga toxin production showed that inhibition of protein synthesis by Shiga toxin, as measured by [35S]methionine incorporation into infected macrophages, was not required for early killing of cells. Damage to J774 macrophages rather correlated with the ability of invasive bacteria to rapidly and efficiently lyse the membrane of the phagocytic vacuole. The role of the release of bacteria within the cytosol for subsequent expression of cytotoxic activity is discussed, and mitochondria are proposed as a potential target for this activity.
Dissociation between delayed-type hypersensitivity and resistance to pathogenic mycobacteria demonstrated by T-cell clones
Authors: Hussein S , Curtis J , Akuffo H , Turk J L
Published: January 2020

One Lyt-2+ clone and fourteen T-helper clones (Lyt-1+ L3T4+ Lyt-2-) were isolated from Mycobacterium lepraemurium-infected BALB/c and C57BL/6 mice. All the clones were tested for their ability to transfer delayed-type hypersensitivity adoptively, and six clones were tested for their ability to transfer resistance. It was found that four L3T4+ clones that transferred delayed-type hypersensitivity responses locally and one L3T4+ clone that did not had no effect on resistance. The Lyt-2+ clone transferred increased resistance locally (77% reduction in the numbers of organisms recovered from the infection site) but did not transfer delayed footpad responses.
Antigenic differences between mannoproteins of germ tubes and blastospores of Candida albicans
Authors: Sundstrom P M , Nichols E J , Kenny G E
Published: January 2020

To determine the nature of germ tube-specific antigens of Candida albicans, procedures for intrinsically labeling cell wall antigens metabolically were developed. Blastospores or germ tubes labeled either in their proteins with L-[35S]methionine or in mannose-containing carbohydrates with D[2-3H]mannose contained surface components similar to those found previously with 125I-labeled organisms. Germ tube-specific determinants, were found on a 200-kilodalton protein in digests from germ tubes, whereas a component of similar molecular size in blastospore extracts reacted weakly or not at all with germ tube-specific antibody. In addition, a glycan fraction prepared from germ tubes reacted with the unadsorbed anti-C. albicans polyvalent antibody but not with the germ tube-specific antibody, suggesting that the germ tube-specific determinants are not carbohydrates.
A bacteriocin of Actinobacillus actinomycetemcomitans
Authors: Hammond B F , Lillard S E , Stevens R H
Published: January 2020

An inhibitory factor from Actinobacillus actinomycetemcomitans Y4 was isolated, and its properties indicated that it was a bacteriocin (actinobacillicin). The bacteriocin was active against Streptococcus sanguis strains, Streptococcus uberis (FDC1), and Actinomyces viscosus T14 as well as other strains of A. actinomycetemcomitans, but not against other crevicular bacteria, including other streptococci and actinomycetes. The activity of this bacteriocin was inhibited by pronase, trypsin, and heat (45 min at 56 degrees C) but not by DNase, RNase, phospholipase, exposure to UV light, or low pH (1.0 to 6.5). Although actinobacillicin markedly inhibited glycolysis in S. sanguis, the primary mechanism of its bactericidal action appears to be alterations in cell permeability, with the resultant leakage of RNA, DNA, and other essential intracellular macromolecules. These findings provide an ecologic explanation for the reciprocal growth relationship between A. actinomycetemcomitans and S. sanguis/Actinomyces viscosus observed in localized juvenile periodontitis.
Histoenzymological study of selected dehydrogenase enzymes in Pneumocystis carinii
Authors: Mazer M A , Kovacs J A , Swan J C , Parrillo J E , Masur H
Published: January 2020

The metabolic activity of Pneumocystis carinii cysts was studied histochemically by a tetrazolium dye technique to assess substrate-specific dehydrogenase activity. Lactate dehydrogenase, succinate dehydrogenase, and glutamate dehydrogenase produced moderate-to-strong reactions in the cysts, whereas glucose-6-phosphate dehydrogenase had little if any reactivity. These results suggest that pneumocystis cysts have some of the enzymes necessary for glycolysis, Krebs cycle activity, and intermediary protein metabolism. These studies provide a method of directly assessing metabolic pathways in P. carinii which circumvents the uncertainties of specificity inherent in previous investigations with partially purified suspensions.
Phospholipid profile of Pneumocystis carinii and its interaction with alveolar type II epithelial cells
Authors: Pesanti E L
Published: January 2020

Pneumocystis carinii is an obligate parasite of mammalian lungs, attaching to but not invading the alveolar epithelium. The alveolar air spaces are rich in phospholipids, which are secreted by steroid-responsive alveolar type II epithelial cells. P. carinii isolated from rat lungs was found to contain the expected structural phospholipids as well as a large amount of firmly attached disaturated phosphatidylcholine, the characteristic phospholipid of alveolar surfactant. In vitro, P. carinii cells synthesized phospholipids from simple radiolabeled precursors; disaturated phosphatidylcholine was not formed. However, washed P. carinii cells avidly adsorbed radiolabeled rat surfactant, a process that appeared to be saturable, not dependent on viability of the organisms, and abolished by incubation at 4 degrees C. The surfactant was neither harmful nor beneficial to in vitro survival of the organisms. With the exception of high concentrations of arachidonic acid, fatty acids found in rat alveolar lining material were also not toxic. In addition, cultures consisting primarily of rat type II alveolar epithelial cells were toxic to P. carinii when the organisms were added to monolayers of type II cells at less than or equal to 10:1 multiplicity. At higher multiplicities, the parasite survived (but did not increase in numbers), and the type II cells deteriorated. The mechanism for this effect has not been determined.
The hyaluronidase associated with Treponema pallidum facilitates treponemal dissemination
Authors: Fitzgerald T J , Repesh L A
Published: January 2020

Treponema pallidum contains hyaluronidase (Hase) associated with its surface. Experiments were performed to determine the functional role of this enzyme in syphilitic infection. The effects of incubating organisms with rabbit anti-bovine Hase or normal or immune sera were compared. Preincubation of treponemes with anti-Hase resulted in inhibition of treponemal degradation of hyaluronic acid, indicating that these antisera did in fact retard enzyme activity. Anti-Hase did not immobilize or neutralize T. pallidum. In addition, rabbits were immunized with bovine Hase and then challenged intradermally with organisms; subsequent lesion development was not affected. Anti-Hase did not block treponemal attachment to cultured testicular fibroblasts but did inhibit attachment to isolated capillaries. Rabbit amnions were used as an in vitro model for dissemination of T. pallidum. Anti-Hase retarded the penetration of organisms through the amnions. This inhibitory effect was dependent on the presence of amniotic hyaluronic acid. When this glycosaminoglycan was selectively removed, the anti-Hase lost its ability to inhibit treponemal penetration. When exogenous hyaluronic acid was added back to treated amnions, the inhibitory effect of anti-Hase was restored. Evans blue experiments were used to characterize treponeme-induced vascular leakage following intradermal inoculation of T. pallidum. Prior treatment of organisms with anti-Hase reduced dermal leakage of the dye, indicating the involvement of the treponemal Hase in causing vessel leakage. Finally, rabbit testicular infections were used as an in vivo model for dissemination; one testis was infected, and after 10 to 13 days, treponemes in the opposite testis were quantitated. The anti-Hase restricted dissemination of organisms. These findings point to the functional role of the treponemal Hase in facilitating disseminated syphilis.
Stimulation of complement component C3 synthesis in macrophagelike cell lines by group B streptococci
Authors: Goodrum K J
Published: January 2020

Complement levels and complement activation are key determinants in streptococcus-induced inflammatory responses. Activation of macrophage functions, such as complement synthesis, by group B streptococci (GBS) was examined as a possible component of GBS-induced chronic inflammation. Using an enzyme-linked immunosorbent assay, secreted C3 from mouse macrophagelike cell lines (PU5-1.8 and J774A.1) was monitored after cultivation with GBS. Whole, heat-killed GBS (1 to 10 CFU per macrophage) of both type Ia and III strains induced 25 to 300% increases in secreted C3 in both cell lines after a 24-h cultivation. GBS-treated cell lines exhibited increases in secreted lysozyme (10%) and in cellular protein (25 to 50%). Inhibition of macrophage phagocytosis by cytochalasin B inhibited GBS stimulation of C3. Purified cell walls of GBS type III strain 603-79 (1 to 10 micrograms/ml) also enhanced C3 synthesis. Local enhancement of macrophage C3 production by ingested streptococci or by persistent cell wall antigens may serve to promote chronic inflammatory responses.
Protective efficacy in humans of killed whole-vibrio oral cholera vaccine with and without the B subunit of cholera toxin
Authors: Black R E , Levine M M , Clements M L , Young C R , Svennerholm A M , Holmgren J
Published: January 2020

Natural protection from cholera is associated with local intestinal antibacterial and antitoxic antibodies, which appear to act synergistically. Although current parenteral cholera vaccines offer insufficient protection, new vaccines administered orally have more promise. Killed Vibrio cholerae, alone or given with the B subunit of cholera toxin, was evaluated in adult volunteers. Vaccinees, who received three doses of either vaccine, and unvaccinated controls ingested 10(6) V. cholerae organisms to determine the protective efficacy of the vaccines. The combination vaccine provided 64% protection, and the whole vibrio vaccine given alone provided 56% protection. In addition, illnesses in vaccines were milder than those in controls, and both vaccines gave complete protection against more severe disease. This substantial level of protection against a dose of V. cholerae that caused cholera in nearly 90% of controls suggests that these vaccines might provide at least as high a level of protection if given to the population of an endemic area. Indeed, a field efficacy trial is underway in Bangladesh, and preliminary data indicate a protective efficacy of 85% for a killed whole vibrio plus B subunit vaccine similar to that tested in volunteers and an efficacy of 58% for the killed whole vibrio vaccine alone. Thus, the studies in human volunteers were successful in predicting the substantial protection afforded by the vaccines in a cholera endemic area.
Comparative virulence of intra- and interstrain lipopolysaccharide variants of Coxiella burnetii in the guinea pig model
Authors: Moos A , Hackstadt T
Published: January 2020

We compared the relative infectivity and virulence of lipopolysaccharide (LPS) variants of the Nine Mile strain of Coxiella burnetii with those of the Priscilla strain, a representative of endocarditis-type strains. In agreement with results of previous studies, Nine Mile phase I (9mi/I) organisms were highly infectious, eliciting seroconversion and fever with inocula containing as few as four organisms. Viable 9mi/I was recovered from the spleens of infected animals 30 days postinfection. Nine Mile phase II (9mi/II) organisms did not elicit fever or seroconversion except with very large inocula, and viable organisms could not be recovered at 30 days postinfection. The Nine Mile/Crazy variant, bearing the intermediate-type LPS, was also highly infectious, as determined by fever response and seroconversion, although, as with 9mi/II, viable organisms could not be recovered 30 days postinfection. The Priscilla strain in phase I (Pris/I) was as infectious as 9mi/I, as determined by seroconversion and its presence in the spleen 30 days postinfection; but in contrast to 9mi/I, more than 10(5) Pris/I isolates were required to induce fever. The temporal appearances of anti-phase I and II antibodies were similar for the two strains. A variety of serological techniques measuring antibody response against whole-cell and purified LPS antigens in agglutination, immunofluorescence, enzyme-linked immunosorbent, and immunoblot assays did not demonstrate sufficient specificity to distinguish between 9mi/I and Pris/I infections. Results of vaccine cross-challenge experiments showed a significant degree of protection between homologous and heterologous challenge strains. protection between homologous
Characterization of lectinlike surface components on Capnocytophaga ochracea ATCC 33596 that mediate coaggregation with gram-positive oral bacteria
Authors: Weiss E I , London J , Kolenbrander P E , Kagermeier A S , Andersen R N
Published: January 2020

The interactions between Capnocytophaga ochracea ATCC 33596 and Streptococcus sanguis H1, Actinomyces naeslundii PK984, or Actinomyces israelii PK16 are dependent on specific recognitions between heat-sensitive adhesins on C. ochracea and heat-stable structures (probably carbohydrate-containing receptors) on the surfaces of these gram-positive coaggregation partners. The coaggregation of C. ochracea with each of these three organisms was inhibited by L-rhamnose and D-fucose and to a lesser extent by beta-methyl-galactoside. The reaction with S. sanguis was the most sensitive, while the coaggregation with A. israelii was the least sensitive and was only partially inhibited by each of the sugars that were considered to be effective inhibitors. A more effective inhibition of the coaggregation between C. ochracea and A. israelii was achieved by adding a combination of the 6-deoxysugars and N-acetylneuraminic acid. To further characterize the coaggregations, naturally occurring coaggregation-defective (Cog-) mutants of C. ochracea were obtained from several different selections. Three phenotypically distinct groups of mutants were were isolated. Type 1 mutants failed to coaggregate with S. sanguis only. Type 2 mutants lost ability to interact with both S. sanguis and A. naeslundii. Type 3 mutants failed to coaggregate with all three coaggregation partners. Characterization of the Cog- mutants by sugar inhibition studies made it possible to distinguish three classes of adhesin activity.
Detection of nucleoside triphosphate hydrolase as a circulating antigen in sera of mice infected with Toxoplasma gondii
Authors: Asai T , Kim T J , Kobayashi M , Kojima S
Published: January 2020

The nucleoside triphosphate hydrolase, which is present in the tachyzoite form of Toxoplasma gondii, was detected as a circulating antigen in sera of mice infected with a virulent (RH) or an avirulent (Beverly) strain of T. gondii. The enzyme was detected with a monoclonal antibody incorporated into an enzyme-linked immunosorbent assay. The lower limit of sensitivity of the assay was about 0.3 ng/ml, and standard assays provided a linear plot of nucleoside triphosphate hydrolase concentration over a range of 0.3 to 12 ng/ml. In mice inoculated intraperitoneally with tachyzoites of the RH strain, nucleoside triphosphate hydrolase emerged in the serum 1 day after injection, and then the concentration increased and reached a value of 30 micrograms/ml on day 5. In mice inoculated intraperitoneally with cysts of the Beverly strain, nucleoside triphosphate hydrolase was detected at day 3 after injection, and a peak concentration of 89 ng/ml was seen on day 10. The concentration of enzyme decreased thereafter, and the enzyme disappeared from the circulation on day 56.
Acetoin production by wild-type strains and a lactate dehydrogenase-deficient mutant of Streptococcus mutans
Authors: Hillman J D , Andrews S W , Dzuback A L
Published: January 2020

Eleven different laboratory strains of Streptococcus mutans representing the various serogroups were found to produce an average of 6.0 +/- 4.8 mM acetoin when grown in glucose-containing medium under aerobic conditions. None of the strains produced detectable acetoin when grown anaerobically. A lactate dehydrogenase-deficient mutant produced acetoin both aerobically and anaerobically and in substantially greater amounts than the wild-type strains did. Substitution of mannitol for glucose resulted in decreased acetoin production by wild-type strains and the lactate dehydrogenase-deficient mutant, indicating a role for NADH2 in the regulation of the acetoin pathway. Pyruvate incorporated into the growth medium of a wild-type strain caused acetoin to be produced anaerobically and stimulated acetoin production aerobically. Cell extracts of a wild-type S. mutans strain were capable of producing acetoin from pyruvate and were (partly) dependent on thiamine PPi. Extracts prepared from aerobically grown cells had approximately twice the acetoin-producing activity as did extracts prepared from anaerobically grown cells. The results indicate that acetoin production by S. mutans may represent an auxiliary reaction of pyruvate dehydrogenase in this organism.
Injury to endothelial cells by phagocytosing polymorphonuclear leukocytes and modulatory role of lipoxygenase products
Authors: Vandenbroucke-Grauls C M , Thijssen H M , van Kessel K P , van Asbeck B S , Verhoef J
Published: January 2020

Phagocytosis of microorganisms by polymorphonuclear leukocytes (PMN) is accompanied by inadvertent extracellular release of microbicidal products; this could result in tissue damage. We investigated whether PMN damages endothelial cells when phagocytosis of Staphylococcus aureus occurs on the endothelial surface and how this damage might be modulated. Damage was assayed by the measurement of cell detachment or cell lysis of cultured endothelial cells that were radiolabeled with 51Cr. Uptake of bacteria was accompanied by nonlytic detachment of endothelial cells from the monolayer. This effect was inhibited by alpha-1-antitrypsin but remained unaffected by scavengers of toxic oxygen species. During phagocytosis, PMN adhered to the endothelial cells. Adherence could be prevented by inhibition of the lipoxygenase pathway of arachidonic acid metabolism of the PMN with nordihydroguaiaretic acid. This inhibition also resulted in a marked decrease of the detaching activity of the PMN. The addition of exogenous leukotriene B4 during phagocytosis greatly enhanced the damage to the endothelial monolayer. These results indicate that phagocytosis of staphylococci by PMN is accompanied by injury to endothelial cell monolayers due to released lysosomal proteases and that products of the lipoxygenase pathway of PMN play a modulatory role in this injury.
Cytotoxicity of Shiga toxin for primary cultures of human colonic and ileal epithelial cells
Authors: Moyer M P , Dixon P S , Rothman S W , Brown J E
Published: January 2020

Shiga toxin purified from Shigella dysenteriae 1 was cytotoxic to cultured epithelial cells from human colon and ileum. The cytotoxicity, which affected only about 50% of treated cells, was neutralized by rabbit antiserum monospecific for Shiga toxin and mediated by protein synthesis inhibition.
Factors affecting colonization and dissemination of Candida albicans from the gastrointestinal tract of mice
Authors: Ekenna O , Sherertz R J
Published: January 2020

Male ICR Swiss mice (2 to 3 months old) were fed Candida albicans in their drinking water for 3 days, followed by no treatment, antibiotics in their drinking water (daily), or immunosuppressants given by intraperitoneal injection (two to three times weekly) over a 3- to 4-week period. The organs of animals were processed to determine the numbers of C. albicans and total aerobic bacteria per g of tissue. Untreated animals had mean Candida counts during the 1-month period of 10(2.3) CFU/g of cecum. Animals in six of eight antibiotic-treated groups had mean cecal Candida counts higher than those of control animals (P less than 0.05), with clindamycin-gentamicin producing the highest counts (10(4.7) CFU/g). Cyclophosphamide produced counts (10(4.3) CFU/g) which were higher (P less than 0.05) than those resulting from methotrexate (10(3.0) CFU/g) or steroid (10(2.7) CFU/g) treatment. Cyclophosphamide-clindamycin-gentamicin treatment was associated with the highest (P less than 0.05) levels of Candida colonization (10(6.5) CFU/g). Mice receiving immunosuppressants plus clindamycin-gentamicin were more likely to disseminate C. albicans than were mice receiving antibiotics alone (P less than 0.001). Our findings suggest that colonization of the guts of mice by C. albicans can be facilitated by manipulating the aerobic, anaerobic, or both types of gut flora. The combined effect of immunosuppressants on both Candida gut colonization and dissemination appears multifactorial and deserves further investigation.
Immunotype epitopes of Neisseria meningitidis lipooligosaccharide types 1 through 8
Authors: Tsai C M , Mocca L F , Frasch C E
Published: January 2020

Lipooligosaccharides (LOS) of the eight immunotypes found in serogroup B Neisseria meningitidis were purified from their prototype strains grown in tryptic soy broth. Rabbit antisera to these LOS were prepared. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by silver staining revealed that most of the LOS antigens contained two major components; the larger components had apparent molecular weights (Mrs) in the range of 4,800 +/- 300, and the smaller components had an apparent Mr of 4,300. Immunoblot analysis showed that the larger major component of an LOS, in general, was much more immunogenic because the rabbits produced antibodies exclusively or primarily to this component even though the LOS immunogen contained both large and small major components. Antibodies to the smaller 4,300-Mr components were infrequently observed but, when present, were cross-reactive with the same-size components of all heterologous LOS. Hence, the immunotype epitopes reside in the larger major components of all immunotypes except type 5, in which a smaller major component having an apparent Mr of 4,400 carries the epitope. Rabbit antisera to types 1, 5, and 6 were immunotype specific. Antisera to other types had cross-reactivities with some heterologous LOS, and the larger components, but not the 4,300-Mr components, of the LOS were primarily responsible for the cross-reactivities. This finding suggests that the larger components of cross-reactive LOS have a similar structure in addition to their type-specific sugar moieties. The LOS of N. meningitidis M986, a strain used for the production of a serotype 2a vaccine, was found to contain the immunotype 7 epitope.
Structural and antigenic properties of lipopolysaccharides from serotype reference strains of Campylobacter jejuni
Authors: Preston M A , Penner J L
Published: January 2020

To investigate the molecular basis for heat-stable antigenic diversity in Campylobacter jejuni, lipopolysaccharides (LPSs) from serotype reference strains and serotyped isolates were examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis coupled with silver staining and immunoblotting. By silver staining, only low-Mr components, consisting of one major band and as many as three minor bands ranging in Mr from 4,500 to 5,000, were detected. However, by immunoblotting with homologous antisera, 10 of 34 strains were shown to have a series of high-Mr LPS components characteristic of molecules with O side chains of various lengths. Isolates of the same serotype as the reference strain that had high-Mr LPS molecules were also found to have high-Mr LPS and in one case of cross-reacting strains it was found that the cross-reaction was associated with antibodies against high-Mr LPS. The reactions of LPSs with homologous and heterologous antisera indicated that both high- and low-Mr-type LPSs were strain-specific antigens, but in some cases cross-reactions were noted. Evidently, all C. jejuni strains possess low-Mr LPS that is readily detectable by silver staining, but some serotypes also possess high-Mr LPS components that can be visualized by immunoblotting.
Analysis of colony-stimulating factors and macrophage progenitor cells in mice immunized against Listeria monocytogenes by adoptive transfer
Authors: Wing E J , Magee D M , Barczynski L K
Published: January 2020

Experiments were performed to elucidate the role of colony-stimulating factors in host defenses to the intracellular pathogen Listeria monocytogenes. Mice were protected against Listeria sp. by adoptive transfer of immune spleen cells and were then challenged with listeriae intravenously. Control mice were injected with spleen cells from uninfected mice. Adoptively immunized (immune) mice had significantly fewer listeriae in spleens and livers 2 and 4 days after Listeria challenge than did control mice. During acute infection, colony-stimulating activity in serum was increased earlier (10 h) in immune mice than in controls. Concentrations of colony-stimulating activity were equal at 24 h. By 48 h, values were decreased in immune mice, but were elevated in control mice. Similar changes were noted when a specific colony-stimulating factor, macrophage colony-stimulating factor, was measured in serum by using a radioimmunoassay. The changes in serum colony-stimulating activity in mice adoptively immunized with immune spleen cells were eliminated if spleen cells were first treated with anti-Thy-1.2 monoclonal antibodies. The number of macrophage progenitor cells in bone marrow and spleen were also determined as measures of the hemopoietic potential in these organs. The number of macrophage progenitor cells in bone marrow was higher in immune animals than control animals at 1, 2, and 4 days of infection. Similarly, the number of these cells in spleens was higher during the early stages of infection in immune mice. These results indicate that both the regulation of leukocyte production and the transfer of specific cellular immunity by spleen cells are associated, and they therefore suggest that hemopoietic regulatory factors play a role in immune host defenses.
Identification of a specific receptor for plasmin on a group A streptococcus
Authors: Lottenberg R , Broder C C , Boyle M D
Published: January 2020

Certain group A streptococci demonstrate surface receptors that bind selectively to the key fibrinolytic enzyme, plasmin. These bacteria show no reactivity with the zymogen protein plasminogen or with other serine class proteases, such as trypsin or urokinase. Bacterium-bound plasmin retains its ability to cleave synthetic substrates and its ability to hydrolyze a fibrin clot. The bacterium-bound plasmin is not effectively regulated by its physiological regulator, alpha 2-plasmin inhibitor. This study is the first report of a bacterium-associated receptor for plasmin.
Resistance to meningococcemia apparently conferred by anti-H.8 monoclonal antibody is due to contaminating endotoxin and not to specific immunoprotection
Authors: Woods J P , Black J R , Barritt D S , Connell T D , Cannon J G
Published: January 2020

We evaluated the ability of a monoclonal antibody directed against the common H.8 antigen of pathogenic Neisseria sp. to confer passive protection against meningococcal disease in mice. The apparent protection conferred by antibody purified from tissue culture supernatant was actually the result of endotoxin contamination of buffers and tissue culture media. Endotoxin-free anti-H.8 antibody was not protective. The possibility of endotoxin contamination should be considered when evaluating immunity conferred by passively administered antibody in animal models.
In strains of Escherichia coli O167 a single plasmid encodes for the coli surface antigens CS5 and CS6 of putative colonization factor PCF8775, heat-stable enterotoxin, and colicin Ia
Authors: Thomas L V , Rowe B , McConnell M M
Published: January 2020

Twenty Escherichia coli strains of serogroup O167 were examined. They all produced the two surface antigens CS5 and CS6 of the putative colonization factor PCF8775, together with heat-stable enterotoxin and colicin Ia. A plasmid coding for CS5, CS6, heat-stable enterotoxin, and colicin Ia was demonstrated in each strain.
Dissociation of virulence and protection from infection by mutant analysis in Haemophilus influenzae type b
Authors: Yogev R , Hansen E J
Published: January 2020

A Haemophilus influenzae type b (Hib) outer membrane protein with an apparent molecular weight of 98,000 (98K) previously has been shown to react with antibodies that protect against experimental Hib disease. A mutant lacking the ability to synthesize this 98K protein was produced by chemical mutagenesis and identified in a colony blot-radioimmunoassay by its failure to react with a 98K protein-specific monoclonal antibody. DNA from this mutant was used to produce a 98K protein-negative transformant of the wild-type parental strain. Comparison of the relative degree of virulence of the parental strain and the 98K protein-negative transformant in an animal model system revealed no differences in the abilities of these two strains to produce bacteremia after intranasal challenge. These results indicate that the Hib surface-exposed 98K outer membrane protein that reacts with protective antibodies plays no detectable role in the expression of virulence by Hib in an animal model system.
Influence of 1,25-(OH)2 vitamin D3 and gamma interferon on the phenotype of human peripheral blood monocyte-derived macrophages
Authors: Poulter L W , Rook G A , Steele J , Condez A
Published: January 2020

The effects of 1,25-dihydroxy vitamin D3 and gamma interferon on the phenotypic changes associated with monocyte maturation in vitro were investigated. Human monocytes separated from peripheral blood mononuclear cell populations by adherence to plastic were cultured for 7 days on glass. Immunocytological analysis was performed on monolayers fixed at various times by using monoclonal antibodies specific for mature macrophages (RFD7), interdigitating (dendritic) cells (RFD1), and class II major histocompatibility complex antigen (RFDR1). Without any addition to the culture medium, proportions of these monocytes (normally RFD1 and RFD7 negative) developed either RFD1 positivity (23%) or RFD7 positivity (49%) over 7 days of culturing. The addition of gamma interferon to these cultures markedly reduced the proportion of RFD7-positive cells (less than 10%) but increased the proportion of RFD1-positive cells (40 to 60%). In contrast, 1,25-dihydroxy vitamin D3 reduced the expression of both RFD1 and RFD7. Both of these effects were dose dependent and required at least 3 days of contact with the cells. The possibility that RFD1- and RFD7-positive cells represent functionally distinct subsets makes these effects of significance in our understanding of the role of these mediators in controlling the immunocompetence of nonlymphoid accessory cell populations and in macrophage-associated antimicrobial activity.
Protein and RNA synthesis by isolated Rickettsia prowazekii
Authors: Winkler H H
Published: January 2020

Purified host-cell-free Rickettsia prowazekii synthesized protein from leucine, lysine, or proline as radioactive precursor. Most, if not all, of the rickettsial proteins were synthesized. The specific activities of the newly synthesized proteins in the cytoplasmic fraction and in the envelope fraction were not significantly different, indicating that both soluble and envelope proteins were synthesized and the latter were inserted into the envelope. The synthesis of protein was sensitive to chloramphenicol but insensitive to the eucaryotic inhibitors cycloheximide and atractyloside. When added before the initiation of synthetic activity, rifampin, a transcriptional inhibitor, inhibited protein synthesis as quickly and extensively as did chloramphenicol, a translational inhibitor. However, when added during synthetic activity, rifampin, unlike chloramphenicol, allowed protein synthesis to continue for several minutes. These results indicate that de novo transcription is necessary for de novo translation and that no stable mRNA pool exists in these rickettsiae. UMP was incorporated into both rRNA and mRNA. The minimal conditions necessary for RNA synthesis contrasted with the complex medium required for protein synthesis. The half-life of the labeled RNA synthesized in these rickettsiae was approximately 16 min. It was concluded that all the rickettsial mRNA that had existed in the growing intracellular rickettsiae was degraded during their lengthy purification.
Oxidative killing of Aspergillus fumigatus proceeds by parallel myeloperoxidase-dependent and -independent pathways
Authors: Washburn R G , Gallin J I , Bennett J E
Published: January 2020

The relative importance of several oxygen intermediates in fungicidal action against opsonized Aspergillus fumigatus conidia was investigated with monocytes from normal volunteers and patients with either chronic granulomatous disease or myeloperoxidase (MPO) deficiency. Results from experiments in which catalase, taurine, mannitol, or glucose-glucose oxidase were added to these phagocytes indicated that the MPO-hydrogen peroxide-halide system and an MPO-independent oxidative system exerted comparable conidiacidal activity. These findings offer a plausible explanation for the susceptibility of patients with chronic granulomatous disease to invasive Aspergillus infections; their phagocytes fail to generate hydrogen peroxide, a substrate necessary for both systems. Patients with MPO deficiency are not known to be predisposed to invasive aspergillosis, suggesting that an MPO-independent oxidative system may provide an alternative mechanism for the oxidative killing of Aspergillus spp.
Major fragment of soluble peptidoglycan released from growing Bordetella pertussis is tracheal cytotoxin
Authors: Rosenthal R S , Nogami W , Cookson B T , Goldman W E , Folkening W J
Published: January 2020

Bordetella pertussis is known to release a factor which promotes the loss of ciliated respiratory epithelium and copurifies with a soluble peptidoglycan (PG) fragment termed tracheal cytotoxin (TCT). The objective of this study was to determine whether pertussis organisms turn over and release PG derivatives in addition to TCT. B. pertussis Tohama (phase III) was grown in liquid Stainer-Scholte medium containing [3H]diaminopimelic acid (DAP) to label PG specifically, washed to remove free label, and suspended in fresh medium without [3H]DAP. Molecular sieve chromatography of supernatants obtained from such cultures revealed a single included peak of 3H, the elution volume of which corresponded roughly to a disaccharide peptide monomer standard (ca. 10(3) daltons). This material (i) contained [3H]DAP in acid-hydrolyzable linkage, (ii) comigrated with 1,6-anhydro-N-acetylmuramic acid-containing disaccharide peptides on paper chromatography, (iii) was resistant to degradation by mild alkali, and (iv) was indistinguishable from authentic TCT by high-voltage paper electrophoresis and two reversed-phase high-performance liquid chromatography systems. Together, the data suggest that B. pertussis releases a markedly homogeneous set of PG fragments, consisting principally of TCT, and that TCT is possibly a nonreducing, anhydromuramic acid-containing fragment or a cyclic PG derivative.
Recombinant human interleukin-2 reverses in vitro-deficient cell-mediated immune responses to tuberculin purified protein derivative by lymphocytes of tuberculous patients
Authors: Shiratsuchi H , Okuda Y , Tsuyuguchi I
Published: January 2020

In vitro lymphocyte proliferative response to purified protein derivative of tuberculin (PPD) was investigated in patients with tuberculosis. Peripheral blood lymphocytes (PBL) from patients with advanced, refractory tuberculosis showed a significantly depressed response compared with the response of PBL from patients with newly diagnosed tuberculosis (P less than 0.01). A further characterization of this low responsiveness to PPD revealed that PBL from these advanced tuberculous patients failed to generate interleukin-2 (IL-2) in response to PPD stimulation. IL-2 receptor (Tac antigen) expression on the surface of T cells after PPD stimulation was also impaired, although to a lesser extent, in the patients with advanced, refractory tuberculosis. We attempted to overcome the depressed in vitro response observed in PBL from patients with advanced, refractory tuberculosis and found that the addition of exogenous, recombinant IL-2 returned the depressed PPD-induced PBL proliferation in these patients to the level of response observed in PBL from patients with newly diagnosed tuberculosis. The addition of recombinant IL-2 also had a restorative effect (up regulation) in vitro on the partly impaired PPD-induced IL-2 receptor expression by PBL from the patients with advanced, refractory tuberculosis. Our results suggest that recombinant IL-2 may offer a novel approach to the therapy of advanced, drug-resistant tuberculosis.
Bacterial antigens induce collagenase and prostaglandin E2 synthesis in human gingival fibroblasts through a primary effect on circulating mononuclear cells
Authors: Heath J K , Atkinson S J , Hembry R M , Reynolds J J , Meikle M C
Published: January 2020

Our previous work suggests that one mechanism through which connective tissue breakdown might occur in periodontal diseases is the production of metalloproteinases, including collagenase, by gingival fibroblasts. In this study we investigated whether highly purified preparations of lipopolysaccharide (LPS) and lipoteichoic acid (LTA) from a number of putative periodontal pathogens could induce monolayer cultures of human gingival fibroblasts to synthesize collagenase and prostaglandin E2. Using both biochemical assays and immunocytochemical techniques, we found that cells synthesized only very small amounts of collagenase in direct response to LPS or LTA (0.1 to 20.0 micrograms/ml). At the highest dose of both antigens, prostaglandin E2 production was increased. We then studied whether LPS and LTA could signal collagenase production by interacting primarily with a different cell type. Our results show that LPS and LTA were each able to stimulate cultures of human blood mononuclear cells (greater than 95% monocytes) to release collagenase-inducing cytokines. By indirect immunocytochemistry, we found that a large proportion of human gingival fibroblasts was activated to produce collagenase by supernatants from LPS- and LTA-stimulated mononuclear cells, whereas gingival fibroblasts cultured with supernatants from unstimulated mononuclear cells were not. Furthermore, in a population of activated fibroblasts we demonstrated, using a double-labeling technique, that some cells made collagenase and the specific tissue inhibitor of metalloproteinases (TIMP) simultaneously. As yet, the collagenase-inducing signals remain poorly characterized but the interleukins-1 and tumor necrosis factors seem likely candidates.
Ingestion of Staphylococcus aureus by bovine endothelial cells results in time- and inoculum-dependent damage to endothelial cell monolayers
Authors: Vann J M , Proctor R A
Published: January 2020

Cultured endothelial cells phagocytize Staphylococcus aureus, but the resultant effects are unknown. Monolayers of cultured bovine endothelial cells with or without [3H]adenine label were exposed to 100, 10, or 1 S. aureus organism per endothelial cell for 3.5 h. Lysostaphin was then applied to all cultures to destroy extracellular but not phagocytized S. aureus. In cultures treated for only 20 min with lysostaphin, S. aureus multiplied exponentially after a 9- to 12-h lag period. In cultures treated continuously with lysostaphin, numbers of S. aureus remained constant or decreased. These results indicate that S. aureus became extracellular and multiplied but did not multiply intracellularly. In parallel experiments, the release of 3H-adenine from prelabeled endothelial cell monolayers was assayed to indicate cytotoxicity. Results indicated that the loss of 3H-adenine from endothelial cell monolayers depended on the following: (i) the size of the S. aureus inoculum, (ii) the strain of S. aureus, and (iii) the length of time after exposure to S. aureus. S. aureus endocarditis and persistent septicemia could arise, at least in part, from ingestion of S. aureus by host endothelium. The intracellular location would afford S. aureus protection from host defenses and antibiotics. Eventual damage to endothelial cells could expose collagen, thus resulting in platelet adherence and vegetation formation. Intracellular S. aureus would be continuously released into the circulation, possibly accounting for the persistent bacteremia that is found in S. aureus endovascular infections.
In vivo and in vitro characterization of murine T-cell clones reactive to Mycobacterium tuberculosis
Authors: Boom W H , Husson R N , Young R A , David J R , Piessens W F
Published: January 2020

Seventeen helper T-cell clones were derived by stimulating lymph node cells from sensitized C57BL/6 mice with Mycobacterium tuberculosis H37Ra, M. tuberculosis H37Rv, or purified protein derivative. Most clones cross-reacted with Mycobacterium bovis BCG, H37Ra, H37Rv, and purified protein derivative. However, four clones were able to differentiate H37Rv from H37Ra, or BCG from H37Ra and H37Rv. In addition, four other T-cell clones recognized recombinant antigens of 19 and 65 kilodaltons isolated from a genomic expression library of M. tuberculosis by using monoclonal antibodies. All clones were Ia restricted and had the Thy-1.2+ Lyt-1+ L3T4+ Lyt-2- phenotype. On stimulation with antigen, all of the clones tested secreted interleukin-2 and gamma interferon but not B-cell stimulatory factor 1. All of the clones tested induced an antigen-specific delayed-type hypersensitivity response upon local cell transfer, although the magnitude of this response differed markedly among clones.