bluuebunny/crossref_metadata_2025 huggingface dataset:
Evidence of mitogenic activity in periodontitis-associated bacteria
Authors: Donaldson S L , Ranney R R , Tew J G
Published: January 2020

This study examines several periodontitis-associated bacterial isolates for the presence of mitogenic activity, as indicated by their capacity to stimulate unsensitized lymphocytes to undergo blastogenesis. Germfree mouse spleen cells responded vigorously to all of the bacterial sonic extracts tested. The kinetics and dose responses to these activators in germfree mouse spleen cell cultures paralleled those seen with the standard murine B-cell mitogen, Escherichia coli lipopolysaccharide. In contrast, Streptokinase-Streptodornase (Varidase; Lederle Laboratories) antigen elicited no response. Human cord blood lymphocytes also responded upon stimulation with these same bacterial isolates but failed to respond to Streptokinase-Streptodornase. The frequency, magnitude, and kinetics of these cord blood lymphocyte responses were remarkably similar to those seen with adult peripheral blood lymphocytes. However, in this and previous studies, individuals with unresponsive peripheral blood lymphocytes have been observed. Studies were initiated to determine whether these unresponsive leukocyte preparations truly lacked the capacity to respond to these bacteria or whether unresponsiveness reflected the presence of a regulatory cell population in these cultures. After the removal of the adherent cells from unresponsive peripheral blood lymphocyte cultures, the nonadherent cells were found to be responsive. Therefore, peripheral blood lymphocyte responsiveness appears to be regulated via an adherent cell population. The removal of adherent cells from unresponsive cord blood lymphocyte preparations resulted in a less consistent alteration to responsiveness. However, cord blood lymphocyte preparations unresponsive at a standard cell density were shown to be responsive at altered cell densities.
Production and characterization of monoclonal antibodies to tetanus toxin
Authors: Sheppard A J , Cussell D , Hughes M
Published: January 2020

Monoclonal antibodies against tetanus toxin were produced to obtain highly specific antisera. Ten hybridoma cell lines producing monoclonal antibodies were derived from the fusion of rat myeloma cells and spleen cells from rats immunized with tetanus toxoid. Eight produced monoclonal antibodies specific for determinants on toxin and toxoid, whereas two were specific only for determinants on the toxoid. The antibodies produced by hybridomas were characterized by determination of the class of light and heavy chain components, epitope specificity, toxin neutralization, and subunit specificity. All of the antibodies contained kappa light chain, eight contained the gamma 1 heavy chain, and the remaining two contained the gamma 2a heavy chain. Five distinct epitopes were indicated by competition assay of paired monoclonal antibodies, and 4 of the 10 monoclonal antibodies neutralized the in vivo activity of tetanus toxin. The four neutralizing monoclonal antibodies and one other were specific for the C fragment of the heavy chain of the toxin molecule.
Effect of cyclophosphamide on the immune responsiveness of jirds infected with Brugia pahangi
Authors: Katz S P , Lammie P J
Published: January 2020

The in vitro immune responsiveness of lymphocytes from Brugia pahangi-infected jirds was examined after serial administration of cyclophosphamide (20 mg/kg). Cyclophosphamide had no effect on parasite burdens, anti-B. pahangi antibody titers, or suppressed spleen cell reactivity to B. pahangi antigens. Cyclophosphamide restored cellular responsiveness to the mitogens phytohemagglutinin, concanavalin A, and pokeweed mitogen.
Protection from respiratory syncytial virus infection in cotton rats by passive transfer of monoclonal antibodies
Authors: Walsh E E , Schlesinger J J , Brandriss M W
Published: January 2020

The effects of passive administration of neutralizing monoclonal antibodies (MAB) to respiratory syncytial virus glycoproteins (GP90 and VP70) was evaluated in cotton rats challenged with respiratory syncytial virus. Animals injected with MAB to VP70 had lower mean viral titers in lung tissues than did controls (log10 2.5 versus 5.4 PFU/g; P less than 0.001), as did cotton rats given MAB to GP90 (log10 2.1 versus 5.0 PFU/g; P less than 0.001). Fifty percent of animals given either MAB had no detectable virus in lung tissues, whereas virus was detected in the lungs of all controls. Virus growth in nasal turbinates was decreased but not eliminated in recipients of either MAB.
Carbohydrate-binding sites of the mannose-specific fimbrial lectins of enterobacteria
Authors: Firon N , Ofek I , Sharon N
Published: January 2020

The combining sites of type 1 fimbrial lectins of various species of enterobacteria were studied by measuring the inhibitory activity of linear and branched oligosaccharides and several glycosides of D-mannose on the agglutination of yeast cells by the organisms. The results showed that all five strains of Escherichia coli tested possessed an elongated combining site best fitting a trisaccharide and including a hydrophobic region. Similar results were obtained with Klebsiella pneumoniae. Within the Salmonella genus, the combining sites of the six species tested were similar, but all differed significantly from those of the E. coli strains. The combining sites of Enterobacter cloacae and Enterobacter agglomerans were different from each other and from those of Salmonella sp. and E. coli. The results suggest that although classified under the general term "mannose-specific," bacterial lectins in the form of type 1 fimbriae on different genera exhibit differences in sugar specificities.
Transport of glucose and mannose by a common phosphoenolpyruvate-dependent phosphotransferase system in Streptococcus mutans GS5
Authors: Liberman E S , Bleiweis A S
Published: January 2020

Decryptified cells of Streptococcus mutans GS5 transport glucose, mannose, and fructose by constitutive phosphoenolpyruvate-dependent phosphotransferase systems (PTSs). Although the non-metabolizable glucose analog 2-deoxyglucose is transported by a PTS, alpha-methylglucose is not taken up by strain GS5. The transport of [14C]mannose and [14C]glucose was almost totally blocked by the heterologous sugars, indicating that these substrates may share a common PTS permease. [14C]fructose transport, however, was not inhibited by large excesses of glucose, indicating the existence of a separate fructose PTS. All "tight" glucose PTS- mutant clones studied were also unable to transport mannose, whereas some "leaky" glucose PTS- clones also were leaky for mannose phosphorylation. Fructose transport in most of these mutant strains was unimpaired, indicating that genetic lesions did not involve soluble (cytoplasmic) PTS components.
Immunochemical characterization of rough Brucella lipopolysaccharides
Authors: Moreno E , Jones L M , Berman D T
Published: January 2020

Lipopolysaccharides (LPS) were extracted from rough strains of Brucella abortus and Brucella melitensis and from strains of the naturally occurring rough species Brucella ovis and Brucella canis. Brucella rough lipopolysaccharides (R-LPS) were readily distinguished from Brucella smooth lipopolysaccharides (S-LPS) and enterobacterial R-LPS, by their chemical, physical, and serological characteristics. B. ovis R-LPS was differentiated from B. abortus, B. melitensis, and B. canis R-LPS by its reaction of partial identity in immunodiffusion. Monospecific mouse sera against B. ovis R-LPS agglutinated only the homologous bacteria but not R cells of other species of Brucella. B. ovis R-LPS contained more 2-keto, 3-deoxyoctonate, and glucosamine as a percentage of dry weight than any other R-LPS tested. B. abortus R-LPS was identified by the absence of an unidentified sugar present in the other R-LPS molecules, and B. melitensis R-LPS could be differentiated from B. canis R-LPS by its higher content of fatty acids. In contrast to S-LPS, all of the R-LPS studied lacked quinovosamine. In electron micrographs, Brucella R-LPS had a granular appearance, in contrast to typical lamellar structures formed by Brucella S-LPS and Escherichia coli R-LPS.
Oral implantation in humans of Streptococcus mutans strains with different degrees of hydrophobicity
Authors: Svanberg M , Westergren G , Olsson J
Published: January 2020

The more hydrophobic, rough-colony-forming, streptomycin-resistant Streptococcus mutans parent strains GW Smr and LK Smr and the less hydrophobic, smooth-colony-forming, streptomycin-resistant variant strains GW36 Smr and LK36 Smr were implanted in oral cavities. Strains GW Smr and LK Smr implanted significantly better than strains GW36 Smr and LK36 Smr. The hydrophobicity of and the colony morphology formed by the different S. mutans strains did not seem to be affected throughout the experiment.
Reactivity to spherule-derived coccidioidin in the southeastern United States
Authors: Woodruff W W , Buckley C E , Gallis H A , Cohn J R , Wheat R W
Published: January 2020

Delayed hypersensitivity skin tests with mycelium-derived (coccidioidin) or spherule-derived (spherulin) antigens (or both) can be used to identify patients who have been sensitized to the dimorphic fungus Coccidioides immitis. Prior studies suggest that coccidioidin and spherulin skin test antigens detect comparable numbers of reactors among exposed subjects. Studies in subjects residing in areas outside the United States where C. immitis is not endemic suggest that both antigens are specific for the fungus. The specificity and reactivity of coccidioidin and spherulin have not been compared in nonendemic regions of the United States in which the skin test antigens and an appropriate travel or exposure history are used to identify patients with possible C. immitis infection. A review of delayed cutaneous reactions to coccidioidin in 6,375 patients tested between 1970 and 1979 in the southeastern United States revealed 958 (15.0%) and 234 (5.7%) positive reactions (greater than or equal to 5 mm), respectively, at 24 and 48 h. Subsequent tests with spherulin in 2,775 patients tested in 1980 and 1981 revealed 866 (31.2%) and 288 (10.3%) positive reactions, respectively, at 24 and 48 h. False-positive immediate hypersensitivity reactions contributed to the large number of spherulin reactors at 24 h. Differences among the patients sampled, work exposure, and travel history were excluded as causes of this surprising and highly significant (P less than or equal to 0.0001) difference in the 48-h delayed cutaneous reaction. These observations suggest two possibilities: (i) spherulin is less specific than coccidioidin, or (ii) a surprising prevalence of C. immitis sensitization exists among patients in nonendemic regions of the United States.
Comparison of immunomodulatory activities in mice and guinea pigs of a synthetic desmuramyl peptidolipid triglymyc
Authors: Leclerc C D , Audibert F M , Chedid L A , Deriaud E J , Masihi N K , Lederer E
Published: January 2020

A nonpyrogenic desmuramyl peptidolipid, 1-O-(L-alanyl-D-isoglutaminyl-L-alanyl-glycerol-mycolate), had previously been shown to be inactive as adjuvant in guinea pigs, but to be very active in stimulating nonspecific resistance. We now show that 1-O-(L-alanyl-D-isoglutaminyl-L-alanyl-glycerol-mycolate) is capable of enhancing or suppressing the immune responses in mice when injected with or before an antigen. In vivo suppression of the immune response to sheep erythrocytes was also observed with high doses of murabutide, a nonpyrogenic adjuvant-active N-acetylmuramyl-L-alanyl-D-isoglutamine analog. Chemiluminescence measurements with mouse spleen cells show a very strong activity of 1-O-(L-alanyl-D-isoglutaminyl-L-alanyl-glycerol-mycolate) by far superior to the effect obtained with the corresponding muramyl peptide, N-acetylmuramyl-L-alanyl-D-isoglutaminyl-L-alanyl-glycerol-myco late.
Isolation and some properties of an enterotoxin produced by Bacillus cereus
Authors: Thompson N E , Ketterhagen M J , Bergdoll M S , Schantz E J
Published: January 2020

Extracellular proteins produced by Bacillus cereus B-4ac were separated by chromatography on Amberlite CG-400, QAE-Sephadex, Sephadex G-75, and hydroxylapatite. A fraction, containing three detectable antigens, obtained from chromatography on hydroxylapatite caused fluid accumulation in ligated rabbit ileal loops, was dermonecrotic to rabbit skin, was cytotoxic to cultured cells, and was lethal to mice after intravenous injection. Two other fractions obtained from chromatography on hydroxylapatite showed essentially no toxic activity when tested individually. Each nontoxic fraction contained two of the three proteins present in the toxic material. When the two nontoxic fractions were combined, activity in all of the biological assays was observed. Antiserum against either of the nontoxic fractions neutralized the dermonecrotic response of the combined material. These results suggest that all of these biological activities probably are due to a single entity and that more than one component probably comprise the toxic entity.
In vivo comparison of avirulent Vwa- and Pgm- or Pstr phenotypes of yersiniae
Authors: Une T , Brubaker R R
Published: January 2020

The abilities of Yersinia pestis to undergo restriction in Ca2+-deficient medium with concomitant production of V and W antigens (Vwa+) and to absorb exogenous pigments (Pgm+) are established virulence factors. Mutation of Y. pestis to Pgm- is known to promote resistance to pesticin (Pstr) and reduced lethality by peripheral routes of injection. Vwa+ Pgm- isolates of Y. pestis were shown in this study to retain virulence in mice when injected intravenously. Although Pgm- in appearance, wild-type cells of Yersinia pseudotuberculosis and Yersinia enterocolitica may also be sensitive to pesticin. Pstr mutants of Vwa+ strains of these species were similarly of reduced virulence, especially by peripheral routes of injection. The consequences of mutation to Vwa- and Pgm- or Pstr on growth and persistence in vivo were determined. After intravenous injection, Vwa+ yersiniae of all species exhibited sustained growth in mouse spleen, liver, and lung and accumulated in blood. Septicemia was not observed after similar injection of Vwa- mutants which were unable to maintain comparable rates of net increase in tissues. Mutation to Pgm- or Pstr did not influence proliferation but resulted in enhanced clearance from organs. It is known that reticuloendothelial cells serve as favored sites of replication for all wild-type yersiniae. Our results are consistent with the hypothesis that the Vwa+ phenotype favors growth within macrophages and that the Pgm+ and pesticin-sensitive phenotypes permit long-term, probably extracellular, retention within organs. Virulence in standard animal models (mice, rats, and guinea pigs) was not correlated with resistance to the bactericidal action of serum.
Effects of environmental and dietary factors on human rotavirus infection in gnotobiotic piglets
Authors: Steel R B , Torres-Medina A
Published: January 2020

The addition of proteolytic enzyme to diets fed to newborn gnotobiotic piglets exacerbated their diarrheal response after oral infection with human rotaviruses. Supplementation of diets with proteolytic enzyme and reduced ambient temperature were evaluated for effects upon the clinical response of gnotobiotic piglets infected with human rotavirus Wa strain, type 2. Piglets were divided into four treatment groups combining two variables: ambient temperature of 35 or 26 degrees C, with and without proteolytic enzyme supplementation of the diet. Infected piglets maintained at 26 degrees C with and without enzyme supplementation had 90 and 70% mortality, respectively. No mortality was observed in infected piglets maintained at 35 degrees C. Protease supplementation of diets fed to piglets kept at 35 degrees C resulted in more uniform onset of diarrhea of greater severity than in littermates fed diets without the supplementation.
Antigenic analysis of Japanese encephalitis virus by using monoclonal antibodies
Authors: Kobayashi Y , Hasegawa H , Oyama T , Tamai T , Kusaba T
Published: January 2020

Hybridoma cells were produced by fusing P3X63Ag8.653 mouse myeloma cells with spleen cells from BALB/c mice immunized with Japanese encephalitis (JE) virus, Nakayama-RFVL strain. The resulting 26 clones produced hemagglutination inhibition antibodies against the homologous strain. The hemagglutination inhibition reactivity of each clone was tested against six flaviviruses: JE, Murray Valley encephalitis (MVE), Egypt 101 strain of West Nile (WN), St. Louis encephalitis (SLE), Russian spring summer encephalitis, and dengue type 1. The 26 monoclonal antibodies fell into four groups: 14 JE species-specific antibodies, 6 antibodies reactive to JE and MVE viruses, 3 antibodies to three or four viruses in the JE-MVE-WN-SLE subgroup, and 3 antibodies to all six flaviviruses. Furthermore, antigenic comparison of 27 strains of JE virus was carried out by using five JE species-specific monoclonal antibodies. Of these, 24 strains were isolated in various parts of Japan, and 3 strains came from Southeast Asia. In reactivity, the 27 strains were classified into at least four antigenic groups. The results showed that the Nakayama-Yakken strain is a mutant strain which lacks the Nakayama strain-specific antigen and that the recently isolated strains are immunologically different from Nakayama and JaGAr 01 strains. One clone (NARMA 13) produced a JE species-specific antibody which showed almost the same titer against 26 JE virus strains, whereas one clone (NARMA 5) produced a Nakayama strain-specific antibody which reacted only to the Nakayama-RFVL and Nakayama-Yoken strains.
Evidence for penetration of diphtheria toxin to the cytosol through a prelysosomal membrane
Authors: Marnell M H , Shia S P , Stookey M , Draper R K
Published: January 2020

To kill mammalian cells, diphtheria toxin must be endocytosed and encounter a low pH within intracellular vesicles. The low pH initiates penetration of the catalytically active A fragment of the toxin through a membrane and into the cytosol where the A fragment arrests protein synthesis. To investigate whether penetration occurred through a prelysosomal or a lysosomal membrane, we studied the effect of low temperature on the entry of the toxin into the cytosolic and lysosomal compartments. The toxin arrested protein synthesis at 15 degrees C, indicating entry into the cytosol; however, access to lysosomes was apparently blocked at 15 degrees C, suggesting that the toxin had encountered a low pH before reaching lysosomes and had penetrated a prelysosomal membrane. To further investigate the possibility of prelysosomal acidification, we measured the time required for the toxin to encounter a low pH after endocytosis. Acidification occurred within 3 to 4 min after the toxin was internalized into vesicles. This interval is consistent with prelysosomal acidification since the entry of endocytosed ligands into secondary lysosomes usually takes more than 3 to 4 min.
Virulent and nonvirulent forms of Plasmodium yoelii are not restricted to growth within a single erythrocyte type
Authors: Fahey J R , Spitalny G L
Published: January 2020

The present studies were designed to investigate whether the erythrocyte preferences displayed by both virulent and nonvirulent forms of Plasmodium yoelii were fastidious growth requirements of these parasites. When inoculated into mice depleted of reticulocytes by lethal irradiation (900 rad), virulent parasites, which have been reported to grow predominantly in mature erythrocytes, gave rise to high parasitemias which were equivalent to those seen in unirradiated, normal mice. In addition, virulent parasites serially passaged in lethally irradiated mice showed properties of enhanced virulence upon inoculation back into normal mice. When inoculated into lethally irradiated mice, nonvirulent P. yoelii, which were reported to preferentially invade reticulocytes, invaded mature erythrocytes, and the infection progressed at a higher level of parasitemia than in unirradiated, normal mice. The inoculation of virulent parasites into mice made reticulocytemic by pretreatment with phenylhydrazine produced infections marked by the invasion of reticulocytes rather than mature erythrocytes, yet these infections remained lethal for the murine host. When nonvirulent parasites were inoculated into reticulocytemic mice, lethal infections resulted in which the parasites predominantly invaded reticulocytes. These results indicate that both the virulent and nonvirulent forms of P. yoelii possess the ability to invade and proliferate within more than one erythrocyte type and that their apparent erythrocyte preferences are not strict growth requirements.
Porins of Brucella species
Authors: Douglas J T , Rosenberg E Y , Nikaido H , Verstreate D R , Winter A J
Published: January 2020

The outer membrane of Brucella species contains two major proteins, denoted as group 2 and group 3 proteins (Verstreate et al., Infect. Immun. 35:979-989, 1982). We reconstituted proteoliposomes from the purified proteins and egg phosphatidylcholine and showed that group 2 proteins, but not a group 3 protein, had the porin activity. The influx rates of sugars of various sizes into the proteoliposomes indicated that the porin channels had apparent diameters in a range comparable to that of Escherichia coli OmpF porin and that the channels were predominantly open. Among different Brucella species, there were small but detectable differences in the channel diameter, and it was possible to explain the differential sensitivity of several Brucella species to diagnostic dyes on the basis of these observed differences.
Stimulation of cell-mediated immunity by bestatin correlates with reduction of bacterial persistence in experimental chronic Salmonella typhimurium infection
Authors: Dickneite G , Kaspereit F , Sedlacek H H
Published: January 2020

The effect of bestatin, a low-molecular-weight immunomodulating drug isolated from Streptomyces olivoreticuli, on Salmonella typhimurium infection was elaborated. Bestatin enhanced the delayed-type hypersensitivity reaction against S. typhimurium in a dose- and time-dependent manner. Parallel to the activation of delayed-type hypersensitivity reaction, bestatin reduced the amount of persistent bacteria in livers and spleens as well as the amount of necrotic foci found in these organs. This was shown when bestatin was given either prophylactically or therapeutically. The therapeutic effect of bestatin was even seen when the drug was given in the chronic phase of the infection, i.e., 6 days after inoculation of the animals with the infectious agent. No influence of bestatin, however, could be observed on the initial multiplication rate of S. typhimurium and concomitantly on the initial mortality rate of the infected mice. As bestatin has no direct antibiotic effect on S. typhimurium, it must be concluded that the therapeutic effects of the drug on chronic infection must be solely contributed to elevation of the host's own defense mechanisms.
Specific immunoglobulin A antibodies to a peptide subunit sequence of bacterial cell wall peptidoglycan
Authors: Franken N , Seidl P H , Kuchenbauer T , Kolb H J , Schleifer K H , Weiss L , Tympner K D
Published: January 2020

Immunoglobulin A (IgA) antibodies in human sera with binding specificity for the C-terminal R-D-Ala-D-Ala sequence of the precursor peptide from bacterial cell wall peptidoglycan were detected by an enzyme-linked immunosorbent assay (ELISA). Specificity of the test system was proved by comparing the high binding of specific IgA to albumin-(D-Ala3) as an antigen with the failure to bind to albumin-(L-Ala3), by binding inhibition studies with L-Ala3, D-Ala3, or peptides with structural analogy to peptidoglycan peptide subunit peptides as inhibitors, and by excluding binding of peroxidase-labeled anti-human IgA to immunoglobulin classes others than IgA. Interference of rheumatoid factors of IgA class was excluded by an ELISA for assaying IgA-rheumatoid factor and by the fact that an IgA fraction essentially free of IgG and IgM was isolated from a serum reacting strongly positive in the ELISA for measuring specific IgA to the peptide subunit of peptidoglycan. This isolated IgA again exhibited binding specificity in the ELISA, thus corroborating the existence of specific IgA in human serum to the C-terminal R-D-Ala-D-Ala sequence of peptidoglycan precursor peptide. The existence of IgA antibodies with specificity for bacterial peptidoglycan was further proved by preadsorption of serum to peptidoglycans and subsequent measurement of specific IgA in the ELISA. Screening of human sera for IgA antibodies with specificity for R-D-Ala-D-Ala peptides revealed that specific antibodies directed against this sequence of bacterial cell wall peptidoglycan may be detected in several human sera.
Purification and characterization of the serotype c antigen from Actinobacillus actinomycetemcomitans
Authors: Zambon J J , Slots J , Miyasaki K , Linzer R , Cohen R , Levine M , Genco R J
Published: January 2020

The serotype c antigen from Actinobacillus actinomycetemcomitans was purified with fractional ethanol precipitation of cell-free culture supernatant, sequential ion-exchange chromatography, and gel filtration chromatography. The preparation obtained demonstrated a single precipitin line in immunodiffusion, immunoelectrophoresis, and crossed immunoelectrophoresis when rabbit antisera to serotype c whole bacterial cells were used. No immunological reaction was detected with antisera to serotype c lipopolysaccharide, indicating that lipopolysaccharide was not present in the preparation. The serotype c antigen was composed of 95% carbohydrate, 2% protein, and 3.1% phosphate. Gas chromatographic analysis of the antigen obtained from growth in either complex or chemically defined media revealed that the carbohydrate constituent was composed of 84 to 90.1% mannose, 4.8 to 16% glucose, 1.9% N-acetylglucosamine, 1.4% fucose, and 0.2% galactose. The present data suggest that A. actinomycetemcomitans serotype c antigen is predominantly a mannose-containing carbohydrate suggestive of a mannan.
Use of a purified outer membrane protein F (porin) preparation of Pseudomonas aeruginosa as a protective vaccine in mice
Authors: Gilleland H E , Parker M G , Matthews J M , Berg R D
Published: January 2020

The outer membrane protein F (porin) from the PAO1 strain of Pseudomonas aeruginosa was purified by two different methods. One procedure involved separation by column chromatography of proteins extracted from isolated outer membranes, whereas the other involved extraction from gels after slab polyacrylamide gel electrophoresis of proteins extracted from cell envelopes. Both procedures yielded protein F preparations which successfully immunized mice from subsequent challenge with the PAO1 strain. The protein F preparations contained small quantities of lipopolysaccharide (LPS). This level of LPS contamination protected immunized mice from challenge with the homologous LPS serotype strain. However, immunization of mice with protein F preparations from the PAO1 strain also afforded protection against challenge with two different LPS serotype strains. This protective ability was lost when the protein F preparation was treated with papain before use as a vaccine. These observations support the conclusion that protein F has protective ability, which is not due to LPS contamination, when given as a vaccine. After immunization with the protein F preparation, mice showed an increase in antibody titer to the purified protein F preparation by enzyme-linked immunosorbent assay. Mice were protected passively by administration of rabbit antisera raised to the protein F preparation. These results indicate that the protein F preparation elicits a specific humoral antibody response in immunized animals. Our results suggest that purified protein F has potential as an effective vaccine for P. aeruginosa.
Immune response in mice infected in the genital tract with mouse pneumonitis agent (Chlamydia trachomatis biovar)
Authors: Barron A L , Rank R G , Moses E B
Published: January 2020

Female Swiss-Webster mice were inoculated intravaginally with mouse pneumonitis agent (MoPn), a Chlamydia trachomatis biovar. Inoculation with 3.5 X 10(5) egg lethal doses per mouse resulted in shedding of the agent from the genital tract for as long as 21 days. Immunoglobulin M antibodies to MoPn were detected in plasma by day 7 post-inoculation, and immunoglobulin G antibodies were detected by day 14. Antibodies were detected in genital secretions by day 20, and titers in plasma and secretions were still considerable on day 56. Delayed-type hypersensitivity tests, determined by footpad swelling, were not positive in appreciable numbers of animals until after day 25. Delayed-type hypersensitivity reactions were maximal 24 h after testing and were preceded by an Arthus-like reaction, which appeared within 3 h and declined by 12 h. Convalescent animals were rechallenged by intravaginal inoculation and were found to be solidly immune.
Rickettsial interactions with human endothelial cells in vitro: adherence and entry
Authors: Walker T S
Published: January 2020

Rickettsia prowazekii, Madrid E strain, was assessed for its ability to enter endothelial cells derived from the veins of human umbilical cord in vitro. Rickettsial entry increased linearly with multiplicity of infection up to a multiplicity of 500; thereafter, additional rickettsiae adhered, but without a concomitant increase in the number of intracellular rickettsiae. Rickettsial entry required participation both of rickettsiae and endothelial cells; inactivation of rickettsiae with N-ethylmaleimide or Formalin, or of endothelial cells with cytochalasin B or D or NaF greatly reduced rickettsial entry. Because rickettsiae adhered to inactivated endothelial cells, adherence could be examined in the absence of entry. Rickettsial adherence was inhibited by poisons that inhibited rickettsial hemolysis. Calcium ionophore A23187, which did not inhibit endothelial pinocytosis, stimulated rickettsial adherence to endothelial cells, but inhibited rickettsial entry. These results indicated that typhus rickettsiae entered endothelial cells via induced phagocytosis, and that the signal for entry, which was dependent upon rickettsial energy, probably involved formation of a calcium gradient.
Bacteroides fragilis: a possible cause of acute diarrheal disease in newborn lambs
Authors: Myers L L , Firehammer B D , Shoop D S , Border M M
Published: January 2020

Bacteroides fragilis with enterotoxin-like activity (BFEL) was isolated from the feces of 24- to 48-h-old lambs with acute diarrheal disease on three different sheep ranches in the Northern Rocky Mountain region of the United States. The lamb intestinal loop test was used to enrich for the bacterium before its initial isolation. Pure cultures of B. fragilis from the feces of diarrheic lambs caused fluid accumulation in five of five lamb intestinal loop tests and in three of three calf intestinal loop tests. Two different serogroups of BFEL were found in diarrheic lambs, and a third serogroups was found in the feces of a diarrheic ewe. Enteric disease characterized by diarrhea, depression, and inappetence occurred in three of eight newborn, colostrum-fed lambs after oral challenge inoculation with viable BFEL. One of the three lambs died 32 h after challenge. Clinical signs of disease were similar to those observed in the naturally occurring disease. B. fragilis is an obligately anaerobic bacterium found in the intestinal tract of most animals and humans. It is a serious cause of extraintestinal infection in humans. The bacterium has not been reported to cause fluid accumulation in the intestine or to cause diarrhea in any species of animal.
Production and characterization of monoclonal antibodies to exotoxin A from Pseudomonas aeruginosa
Authors: Galloway D R , Hedstrom R C , Pavlovskis O R
Published: January 2020

Hybridomas secreting monoclonal antibodies specific for exotoxin A from Pseudomonas aeruginosa strain PA103 were derived from the fusion of spleen cells from mice immunized with: (i) purified exotoxin A, (ii) Formalin-treated exotoxin A, (iii) exotoxin A covalently coupled to Sepharose 4B, or (iv) P. aeruginosa-infected mice. All hybridomas were screened and selected by using an enzyme-linked immuno-adsorbent assay. All antibody isotypes were represented (immunoglobulins G, A, and M) as determined by enzyme-linked immunoadsorbent assay. The most productive fusions resulted from immunization with antigens coupled to an insoluble matrix, such as Sepharose 4B, or by infection of mice. Several hybridomas were selected and cloned by limiting dilution. The specificity of the monoclonal antibodies for exotoxin A was demonstrated by indirect immunoprecipitation of 125I-labeled exotoxin A followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and by the immunoblotting technique. The protective ability of certain monoclonal antibodies was demonstrated in vitro by toxin neutralization in tissue culture and in vivo by prolonged survival time in the burned mouse infection model, after passive immunization. One monoclonal antitoxin displayed specificity for PA103-derived exotoxin yet failed to react with exotoxin purified from PAO-PR1 or PAO1, suggesting that structural differences exist between these exotoxins.
Properties of cross-linked toxoid vaccines made with hyperantigenic forms of synthetic Escherichia coli heat-stable toxin
Authors: Klipstein F A , Engert R F , Houghten R A
Published: January 2020

The ability of hyperantigenic preparations of synthetically produced Escherichia coli heat-stable toxin (ST) to provide an immunogenically more potent vaccine when cross-linked by the glutaraldehyde reaction to the heat-labile toxin B subunit was assessed. Three synthetic ST preparations were evaluated: ST(S) had the same antigenicity and toxicity (secretory potency in the suckling mouse assay) as native ST, ST 1056 had 3.5-fold more antigenicity and 1% toxicity, and ST(C) had 15-fold greater antigenicity and 31% toxicity. Vaccines that contained equal antigenic proportions of ST and B subunit, as determined by enzyme-linked immunosorbent assays, consisted by weight of 52% ST(S), 25% ST 1056, and 9% ST(C). The initially lower toxicity and smaller proportions by weight of hyperantigenic ST preparations yielded vaccines that had nearly 10-fold less residual ST toxicity than the ST(S) vaccine. Immunization of rats with graded dosages of vaccines containing 9% ST(C) and 51% ST(S) by weight, but equal amounts of ST(S) antigenicity, raised to the same degree dose-dependent increases in mucosal immunoglobulin A antitoxin titers to ST(S) which correlated with the amount of protection against challenge with a viable LT-/ST+ strain. These observations indicate that hyperantigenic synthetic ST preparations provide immunologically more potent vaccines than those obtained with the previously used synthetic ST(S) preparation, which has the same biological properties as native ST.
Mechanism of interaction of Salmonella and Schistosoma species
Authors: Melhem R F , LoVerde P T
Published: January 2020

In endemic areas where Salmonella and Schistosoma species co-occur, several lines of evidence suggest a synergistic bacteria-parasite interaction that results in a protracted course for the salmonella infection that has proven difficult to diagnose and therapeutically remedy. In an in vitro system using a pilus-negative and a pilus-producing transductant strain of Salmonella typhimurium we show that pili are the ligands for bacterial adherence to the schistosome surface tegument. Antipili antibodies produced in rabbits against purified pili, purified and digested to monovalent (Fab) fragments, blocked the association of Salmonella sp. to the surface tegument of Schistosoma sp., further demonstrating that pili are the appendages necessary for bacteria-parasite surface interaction. The use of carbohydrates, lectins, and enzymes demonstrated that the bacteria-parasite surface interaction was specific, mediated by pili that specifically recognize and bind to mannose-like receptors, probably glycolipids, on the surface of the worms. We suggest that prolonged salmonellosis in schistosome-infected patients is due to an association of Salmonella sp. with the schistosome worms themselves and further that the schistosome worms provide a multiplication focus for these bacteria in the portal mesenteric system, with a persisting bacteremia following.
Mode of inhibitory action of a bacteriocin produced by Streptococcus mutans C3603
Authors: Takada K , Ikeda T , Mitsui I , Shiota T
Published: January 2020

The basis for the lethal activity of a bacteriocin produced by Streptococcus mutans C3603 (serotype c) was studied. Bacteriocin C3603 was found to adsorb to cells of representative strains of the seven serotypes of S. mutans. S. mutans BHT (serotype b) was used to study the adsorption and the lethal properties of bacteriocin C3603. The adsorption of bacteriocin to cells of S. mutans BHT was inhibited by treatment of cells with protease and beta-glucosidase and by such ligands as poly-L-lysine, poly-L-arginine, L-aspartic acid, L-glutamic acid, glutathione, oxidized glutathione, poly-L-aspartic acid, and poly-L-glutamic acid. The adsorption to cells was also inhibited by oligosaccharides and glucosamine. Mixtures of anionic and cationic amino acids or polyamino acids did not greatly enhance or antagonize the inhibition of adsorption of bacteriocin C3603 to cells. Sodium hydroxide extracts of cell walls and cell wall-membranes contained carbohydrates and proteins; however, only proteins were found to bind to bacteriocin or to a bacteriocin affinity column. The sodium hydroxide extracts contained about 35 protein bands as determined by sodium dodecyl sulfate-polyacrylamide disc gel electrophoresis. Bacteriocin C3603 was found to immediately inhibit the synthesis of proteins, DNA, and RNA of cells and to slowly release DNA from cells of S. mutans BHT.
Coli surface antigens 1 and 3 of colonization factor antigen II-positive enterotoxigenic Escherichia coli: morphology, purification, and immune responses in humans
Authors: Levine M M , Ristaino P , Marley G , Smyth C , Knutton S , Boedeker E , Black R , Young C , Clements M L , Cheney C
Published: January 2020

Enterotoxigenic Escherichia coli (ETEC) of serotype O6:H16, biotype A, bearing colonization factor antigen II (CFA/II) possesses two distinct coli surface antigens, CS1 and CS3, whereas CFA/II-positive ETEC of serotype O8:H9 manifests only CS3. CS1 has been shown to be fimbrial in nature, but heretofore the morphology of CS3 has not been described. Accordingly, by immune electron microscopy we investigated the morphological characteristics of CS3 on bacterial cells and after purification. CS3 was found to consist of thin (2-nm), flexible, wiry, "fibrillar" fimbriae, visible both on bacteria (O6:H16, biotype A, and O8:H9 strains) and in the pure state. In contrast, CS1 exists as wider (6-nm), rigid fimbriae on the surface of O6:H16, biotype A, strains. By the use of antisera to CS1 and CS3 in immune electron microscopy, immunodiffusion in gel, and immunoblotting techniques, CS1 and CS3 were found to be immunologically as well as morphologically distinct. Six of nine volunteers who developed diarrhea after challenge with an O139:H28 ETEC strain bearing CS1 and CS3 had significant serological rises to purified CS1 and CS3 antigens, suggesting that both antigens are elaborated in vivo, play a role in pathogenesis, and stimulate an immune response.
Role of disulfide bonding in outer membrane structure and permeability in Chlamydia trachomatis
Authors: Bavoil P , Ohlin A , Schachter J
Published: January 2020

The outer membrane of Chlamydia trachomatis can be efficiently solubilized by a variety of mild detergents in the presence of the reducing agent dithiothreitol. This allows purification of the chlamydial major outer membrane protein at high yield in very gentle conditions by using its differential solubility in Sarkosyl and octylglucoside in the presence of dithiothreitol. The major outer membrane protein of the L2 serovar is an acidic protein with a pI of ca. 5. It contains three cysteine residues that allow it to form a disulfide-linked proteinaceous network responsible for the characteristic rigid outer membrane of the elementary body. By the use of an in vitro reconstitution assay developed by Nikaido and his co-workers, it was shown that the outer membrane contains pores with an "exclusion limit" between molecular weights 850 and 2,250. In addition, the "opening-closing" of the pores was shown to be controlled through a simple reduction-oxidation mechanism. A model that outlines the role of disulfide bonding in the physiology of chlamydial development is presented.
Characterization of the mechanism of action of Escherichia coli heat-stable enterotoxin
Authors: Dreyfus L A , Jaso-Friedmann L , Robertson D C
Published: January 2020

The mechanism of activation of intestinal guanylate cyclase by Escherichia coli heat-stable enterotoxin (STa) has been studied by using isolated rat intestinal epithelial cells and purified brush border membrane (BBM) preparations. Inhibitors of prostaglandin biosynthesis, quinacrine and 5,8,11,14-eicosatetraynoic acid (ETYA), significantly reduced intracellular levels of cyclic guanosine 3', 5'-monophosphate in isolated cells treated with STa. Although these data suggested that activation of phospholipase A2 and metabolism of arachidonic acid are involved in the mechanism of action of STa, other data ruled out such a mechanism. (i) The rate of release of [3H]arachidonic acid by prelabeled intestinal cells incubated with STa was the same as control cells not treated with STa. (ii) Thin-layer chromatography of lipid extracts of intestinal cells treated with STa and untreated cells did not reveal any quantitative or qualitative differences in free fatty acids, neutral lipids, and phospholipids. (iii) Amounts of prostaglandin PGE2, prostaglandin PGF2 alpha, and thromboxane B2 in intestinal cells and BBM incubated with STa did not increase compared with controls not incubated with STa. When purified BBM preparations were incubated with phospholipase A2 inhibitors (p-bromophenacyl bromide and quinacrine) or cyclooxygenase inhibitors (ETYA and indomethacin), basal and STa-induced guanylate cyclase activities were significantly reduced. Inhibitors of calcium-calmodulin-mediated reactions (EGTA [ethylene glycol-bis(beta-aminoethyl ether)-N,N-tetraacetic acid], trifluoperazine, and chlorpromazine) and calcium channel blockers (verapamil and nifedipine) also nonspecifically inhibited both basal and STa-stimulated guanylate cyclase in BBM preparations. Lanthanum, a competitive inhibitor of membrane-bound calcium, did not affect either basal or STa-stimulated guanylate cyclase of BBM preparations.(ABSTRACT TRUNCATED AT 250 WORDS)
Ultrastructural study of adhesion of enterotoxigenic Escherichia coli to erythrocytes and human intestinal epithelial cells
Authors: Knutton S , Lloyd D R , Candy D C , McNeish A S
Published: January 2020

The adhesion to erythrocytes and human intestinal epithelial cells of enterotoxigenic Escherichia coli strains H10407, B2C, and H10407P, expressing colonization factor antigen I (CFA/I), CFA/II, and type 1 fimbriae, respectively, was examined by electron microscopy. CFA and type 1 fimbriae were visualized by negative staining in thin sections after en bloc staining with ruthenium red and by immune labeling with antisera raised against purified fimbriae. By negative and ruthenium red staining, CFA/I, CFA/II, and type 1 fimbriae were indistinguishable and appeared as approximately 7-nm-diameter hollow cylindrical structures up to 1.5 micron in length; strain B2C also produced 2- to 3-nm-diameter flexible fibrillar fimbriae. Bacteria producing CFA/I, CFA/II, and type 1 fimbriae adhered to and agglutinated human, bovine, and guinea pig erythrocytes, respectively; CFA/I and CFA/II also mediated attachment of bacteria to the brush border of isolated human duodenal enterocytes. Electron microscopy of agglutinated erythrocytes and enterocytes with adherent bacteria showed, in each case, that bacterial adhesion involved the formation of many interactions between the tips of fimbriae and receptors on the erythrocyte or enterocyte brush border membrane. Immune labeling allowed different fimbrial antigens mediating bacterial attachment to human enterocytes to be identified.
Heat shock and heat stroke proteins observed during germination of the blastoconidia of Candida albicans
Authors: Dabrowa N , Howard D H
Published: January 2020

Cytoplasmic proteins extracted from germinating yeast cells of Candida albicans were analyzed by sodium dodecyl sulfate-polyacrylamide slab gel electrophoresis. Similar extracts from a recently isolated nongerminating variant were compared with those from the parent. Five proteins (18, 22, 40, 68, and 70 kilodaltons [kd]) behaved as heat-shock proteins in that they appeared or were greatly increased in amount within 20 min of a temperature shift from 23 to 37 degrees C. Three of the five (40, 68, and 70 kd) were undetected in cells incubated at 23 degrees C, appeared within 20 min of temperature shift, and were no longer detected after 120 min at 37 degrees C, whereas two of the five (18 and 22 kd) were present in small amounts at 23 degrees C, increased greatly after shift, and persisted for 120 min at the elevated temperature. Two temperature-repressed (heat-stroke) proteins (30 and 88 kd) were also observed. The same heat-shock and heat-stroke proteins were also found in the nongerminating variant. The differences in proteins expressed by blastoconidia and by germlings appeared to be related to the heat-shock response.
Effect of bacterial secondary infection in an animal model of trachoma
Authors: Taylor H R , Kolarczyk R A , Johnson S L , Schachter J , Prendergast R A
Published: January 2020

In trachoma the interaction between chronic chlamydial and acute bacterial conjuntivitis has been suggested as important in determining the severity of disease and, therefore, blindness. We investigated the effect of acute conjunctival infection with each of three common human pathogens, Haemophilus influenzae, Haemophilus aegyptius , and Streptococcus pneumoniae, in a model of trachoma established in cynomolgus monkeys. Although acute conjunctivitis developed, animals with trachoma were not more susceptible to infection than other monkeys, nor did they develop more severe disease as a result of the bacterial conjunctivitis. The failure of bacterial conjunctivitis to exacerbate the experimental trachoma indicates that, in this model at least, chronically maintained chlamydial infection alone is sufficient to produce the changes characteristic of trachoma.
Recombinant cold-adapted attenuated influenza A vaccines for use in children: reactogenicity and antigenic activity of cold-adapted recombinants and analysis of isolates from the vaccinees
Authors: Alexandrova G I , Polezhaev F I , Budilovsky G N , Garmashova L M , Topuria N A , Egorov A Y , Romejko-Gurko Y R , Koval T A , Lisovskaya K V , Klimov A I
Published: January 2020

Reactogenicity and antigenic activity of recombinants obtained by crossing cold-adapted donor of attenuation A/Leningrad/134/47/57 with wild-type influenza virus strains A/Leningrad/322/79(H1N1) and A/Bangkok/1/79(H3N2) were studied. The recombinants were areactogenic when administered as an intranasal spray to children aged 3 to 15, including those who lacked or had only low titers of pre-existing anti-hemagglutinin and anti-neuraminidase antibody in their blood. After two administrations of vaccines at a 3-week interval, both strains induced antibody in 75 to 95% of the children. On coinfection of chicken embryos with both recombinants, only weak interference was observed. Administration to children of the bivalent vaccine containing H1N1 and H3N2 recombinants induced efficient production of antibody to H1 and H3 hemagglutinins and N1 and N2 neuraminidases without adverse reactions. The recombinants studied were genetically stable as judged by retention of the temperature-sensitive phenotypes and a lack of reversion of the genes carrying temperature-sensitive mutations in all of the reisolates from vaccinated children.