bluuebunny/crossref_metadata_2025 huggingface dataset:
Development of monoclonal antibodies that recognize Treponema pallidum
Authors: Saunders J M , Folds J D
Published: January 2020

We developed a panel of monoclonal antibodies to Treponema pallidum (Nichols) antigens, some of which recognize treponemal antigens on T. pallidum (Nichols), T. pallidum strain 14, and Treponema phagedenis biotype Reiter. The antibodies were detected by either an enzyme-linked immunosorbent assay or a radioimmunoassay.
Immunotypes of Chlamydia trachomatis isolates in Seattle, Washington
Authors: Kuo C C , Wang S P , Holmes K K , Grayston J T
Published: January 2020

The immunotype distribution of 493 Chlamydia trachomatis isolates from 467 adults and 26 infants in the Seattle area from 1965 to 1982 is presented. All except one of the isolates from adults were from the genitourinary tract, rectum, or bubo. The proportions of each immunotype were ED, 46.5%; GF, 24.6%; H, I, J/CJ, and K, 5 to 7% each; B, 3.5%; and L2, 1%.
Characterization of Candida albicans adherence to human vaginal epithelial cells in vitro
Authors: Lee J C , King R D
Published: January 2020

Certain environmental, physical, and biochemical aspects of Candida albicans adherence to human vaginal epithelial cells were characterized by using an in vitro radiometric adherence assay. Blastospores harvested from cultures grown at 25 degrees C adhered to vaginal epithelial cells in significantly greater numbers than did blastospores isolated from cultures grown at 37 degrees C. C. albicans viability was not essential for adherence, but severe methods used to kill the blastospores did reduce their attachment. The addition of sodium chloride, divalent cations, sugars, mannan, or mannoprotein to the assay had no effect on attachment. Pretreatment of the blastospores with detergents, salts, urea, glycosidases, lipase, or pepsin did not affect adherence, but treatment with reducing agents or five proteolytic enzymes did render C. albicans nonadherent. Cell wall fragments prepared from C. albicans, but not from Candida krusei, adhered to vaginal epithelial cells. Loss of adherence after the cell walls were treated with alpha-mannosidase or papain suggests that cell wall mannoprotein is an essential component of the C. albicans adhesin.
Synthesis of plasmid-coded heat-labile enterotoxin in wild-type and hypertoxinogenic strains of Escherichia coli and in other genera of Enterobacteriaceae
Authors: Neill R J , Twiddy E M , Holmes R K
Published: January 2020

The effect of host determinants on expression of plasmid-coded heat-labile enterotoxin (LT) was examined. A collection of LT plasmids was introduced into isogenic strains of Escherichia coli K-12 strains containing the wild type or hypertoxinogenic (htx-2) allele. For each plasmid tested, production of LT increased by approximately 1.5- to 3-fold in the host containing htx-2, indicating that the htx-2 allele affects a regulatory function for LT production that is common to many different enterotoxin plasmids. LT plasmids from E. coli were also introduced into strains of Shigella flexneri, Shigella sonnei, Citrobacter freundii, Enterobacter cloacae, Klebsiella pneumoniae, and Salmonella typhimurium. The plasmids were stably maintained and determined production of LT in those genera, although the amounts of LT produced varied by more than 50-fold. These observations demonstrate that host factors have an important role in determining the level of expression of plasmid-coded LT genes and support the hypothesis that interspecific, conjugal transfer of enterotoxin plasmids may confer enterotoxigenicity to a wide variety of potentially pathogenic enteric bacteria.
Cloning, expression, and mapping of the Staphylococcus aureus alpha-hemolysin determinant in Escherichia coli K-12
Authors: Kehoe M , Duncan J , Foster T , Fairweather N , Dougan G
Published: January 2020

A fragment of Staphylococcus aureus DNA encoding the alpha-hemolysin determinant was cloned from strain Wood 46 by inserting Sau3A-generated genomic DNA fragments between the BamHI sites of the lambda replacement vector L47.1. Phages expressing alpha-hemolysin were detected by overlaying plaques formed from several thousand independent recombinant phage with erythrocytes and looking for zones of hemolysis. One phage expressing alpha-hemolysin was purified and named lambda w alpha 3. This was subsequently shown to contain a 10.2-kilobase pair insert of S. aureus DNA. A 7.6-kilobase pair HindIII fragment encoding the alpha-hemolysin was subcloned from lambda w alpha 3 into the plasmid vector pACYC184 to form the hybrid plasmid pDU1148. Escherichia coli K-12 cells harboring pDU1148 synthesized a low level of alpha-hemolysin which remained associated with the cells and was not secreted into culture supernatants. When the same strain was stabbed onto blood agar plates, no zones of hemolysis were detected after overnight growth at 37 degrees C but hemolysis developed if the plates were left at room temperature for 48 h. By introducing specific deletions or Tn5 insertions into plasmid pDU1148, the alpha-hemolysin gene was mapped to a region within a 3.3-kilobase pair EcoRI-HindIII fragment which was subcloned onto the vector plasmid pBR322. A specific enzyme-linked immunosorbent assay with peroxidase-labeled rabbit anti-alpha-hemolysin antibodies was used to measure the levels of alpha-hemolysin antigen expressed in E. coli K-12 cells harboring pDU1148 or a variety of pDU1148::Tn5 and pDU1148 deletion mutants.
Expression of a cloned Staphylococcus aureus alpha-hemolysin determinant in Bacillus subtilis and Staphylococcus aureus
Authors: Fairweather N , Kennedy S , Foster T J , Kehoe M , Dougan G
Published: January 2020

A DNA sequence encoding Staphylococcus aureus alpha-hemolysin, which had been previously cloned and mapped in Escherichia coli K-12, was introduced into Bacillus subtilis BD170 and several strains of S. aureus by using plasmid vectors, some of which could replicate in all three organisms. The determinant was cloned on a 3.3-kilobase pair DNA fragment into B. subtilis by using the vector plasmid pXZ105 to form the hybrid plasmid pXZ111. B. subtilis cells harboring pXZ111 produced large zones of alpha-hemolysis after 18 h of growth at 37 degrees C on rabbit blood agar plates, and alpha-hemolysin activity was detected in supernatants prepared from growing cultures of this strain. The alpha-hemolysin was apparently secreted across the B. subtilis cell envelope. Polypeptides of molecular weights 34,000 and 33,000 were precipitated with anti-alpha-hemolysin serum from lysates prepared from BD170 cells harboring pXZ111. A hybrid replicon which could replicate in both E. coli and S. aureus was constructed in E. coli by ligating a HindIII fragment encoding the replication functions and chloramphenicol resistance genes of S. aureus plasmid pCW59 to the pBR322 alpha-hemolysin hybrid plasmid pDU1150. The DNA of this plasmid, pDU1212, was prepared in E. coli and used to transform protoplasts prepared from a non-alpha-hemolytic, nonrestricting strain of S. aureus RN4220. Some of the transformants contained plasmids which had suffered extensive deletions. Some plasmids, however, were transformed intact into RN4220. Such plasmids were subsequently maintained in a stable manner. pDU1212 DNA was prepared from RN4220 and transformed into alpha-hemolytic S. aureus 8325-4 and two mutant derivatives defective in alpha-hemolysin synthesis. All three strains expressed alpha-hemolysin when harboring pDU1212.
Cellular immunity to Legionella pneumophila in guinea pigs assessed by direct and indirect migration inhibition reactions in vitro
Authors: Friedman H , Widen R , Lee I , Klein T
Published: January 2020

Spleen cell cultures from guinea pigs given legionella pneumophila vaccine in complete Freund adjuvant or as a sublethal infection were inhibited in their migration activity in vitro when incubated with specific antigen. Both direct and indirect migration inhibition assays revealed sensitization of the guinea pigs to the bacterium, with demonstrable reactivity 25 to 40 days or more after sensitization. No consistent reactions occurred when the guinea pigs were given the killed Legionella vaccine in incomplete Freund adjuvant in saline. However, spleen cells from guinea pigs injected with sublethal doses of the Legionella vaccine 3 to 4 weeks earlier showed positive migration inhibition factor reactivity. Cutaneous hypersensitivity and lymphocyte blastogenic responsiveness in vitro also developed in guinea pigs sensitized with killed Legionella vaccine in complete adjuvant or given a sublethal infection with the bacterium. These results indicate that in vitro assays for migration inhibitory activity may be utilized to monitor the development of the sensitization of guinea pigs to L. pneumophila, and such reactions correlate with skin reactivity and in vitro lymphocyte blastogenic responses.
Effect of heptakis (2,6-O-dimethyl) beta-cyclodextrin on the production of pertussis toxin by Bordetella pertussis
Authors: Imaizumi A , Suzuki Y , Ono S , Sato H , Sato Y
Published: January 2020

The effect of heptakis (2,6-O-dimethyl) beta-cyclodextrin (Me beta CD) on the production of pertussis toxin was evaluated. The addition of Me beta CD to the medium stimulated cell growth and pertussis toxin production. Me beta CD enhanced pertussis toxin production 100 times more in synthetic media, such as Stainer-Scholte medium (D. W. Stainer and M. J. Scholte, J. Gen. Microbiol. 63:211-220), than in Me beta CD-free medium in 2-day shake cultures. Maximum production of pertussis toxin was estimated as 50 mg of protein per liter of culture broth both by in vitro and in vivo assays. Purified toxin was demonstrated to be biochemically and biologically identical to the toxin produced in Me beta CD-free static cultures.
Serological response to filamentous hemagglutinin and lymphocytosis-promoting toxin of Bordetella pertussis
Authors: Burstyn D G , Baraff L J , Peppler M S , Leake R D , St Geme J , Manclark C R
Published: January 2020

Serum antibody responses to the filamentous hemagglutinin and the lymphocytosis-promoting toxin of Bordetella pertussis after vaccination with diphtheria and tetanus toxoids and pertussis vaccine, adsorbed, were assayed by using the enzyme-linked immunosorbent assay. The effect of early immunization, during the first week of life, on the antibody response also was determined. After vaccination, immunoglobulin G (IgG) and IgM directed against both the filamentous hemagglutinin and the lymphocytosis-promoting toxin were detected. Generally, antibody titers increased with subsequent injections and the age of the children. Maternal antibodies against filamentous hemagglutinin and lymphocytosis-promoting toxin were detected in cord blood. The ability of an infant to produce serum IgG anti-lymphocytosis-promoting toxin after vaccination with pertussis vaccine was inversely related to the cord blood serum IgG anti-lymphocytosis-promoting toxin titer at birth. A good antibody response was observed in infants with low cord blood titers, and a poor antibody response was seen in infants with high cord blood values. The IgM anti-lymphocytosis-promoting toxin response was good in groups with both low and high cord blood titer, with no significant difference observed between the two groups. No IgA anti-lymphocytosis-promoting toxin or IgA anti-filamentous hemagglutinin titers were observed in vaccines. IgA antibodies were observed in convalescent sera from two adults and may be presumptive evidence of infection with B. pertussis.
Colonization of the rabbit small intestine by clinical and environmental isolates of non-O1 Vibrio cholerae and Vibrio mimicus
Authors: Spira W M , Fedorka-Cray P J , Pettebone P
Published: January 2020

We examined the capability of 12 isolates of non-cholera toxin-producing O1 and non-O1 Vibrio cholerae to colonize the small intestine of adult rabbits and cause diarrhea. Using the removable intestinal tie-adult rabbit diarrhea model, we found that eight environmental isolates that showed no or marginal biological activity in other diarrhea models (rabbit ileal loop, infant rabbit, and suckling mouse) appeared to be incapable of attaching to and colonizing, even transiently, the small intestinal mucosa of animals with normal clearance mechanisms. In contrast, three clinical isolates attached, proliferated rapidly, and colonized mucosal surfaces of the entire small intestine within 8 h of challenge. This led to diarrhea with strikingly high rates of mortality compared with that of rabbits given similar challenges doses with strains of O1 V. cholerae that produce cholera toxin and Vibrio mimicus, which produces a toxin similar to cholera toxin. We have further demonstrated that multiple exposures to enteric infection by these strains elicited local and serum antibodies that reacted strongly with cell surface antigens of the homologous strain and showed a high degree of cross-reactivity against the cell surface antigens of the two heterologous strains. The enteric infections appeared to engender protection against subsequent infection as well, as evidenced by reduced incidence of diarrhea and duration of fecal shedding of the challenge organism upon subsequent challenges.
Effect of cyclosporine on the response of normal human lymphocytes to cytomegalovirus in vitro
Authors: Converse P J , Hess A D , Tutschka P J , Santos G W
Published: January 2020

Lymphocytes from healthy volunteers seropositive for cytomegalovirus (CMV) demonstrated strong lymphoproliferative responses to CMV-infected and glutaraldehyde-fixed foreskin fibroblasts (CMVFFx) when cocultured for 6 days. Addition of the immunosuppressive drug cyclosporine (CsA) to the cultures resulted in a 10-fold reduction (P less than 0.001) in counts per minute of [3H]thymidine uptake. The proliferative response to noninfected fixed fibroblasts was also reduced 10-fold. Kinetic studies showed an inhibition of the lymphoproliferative response and not an alteration in the time course kinetics in CsA-treated cultures. Cytotoxicity to CMV-infected and unfixed fibroblasts by lymphocytes primed by CMVFFx in the presence of 0.5 micrograms of CsA per ml was significantly reduced (P less than 0.01) as compared with untreated cultures but remained significantly above background level (P less than 0.01). The cytotoxic response was still present but reduced at concentrations of greater than or equal to 1.0 micrograms/ml. Cytotoxicity to noninfected fibroblasts by CMVFFx-primed lymphocytes could be eliminated by as little as 0.5 micrograms of CsA per ml. Stimulation of lymphocytes by CMVFFx but not by noninfected fixed fibroblasts resulted in the in vitro generation of suppressor cells. CsA at a final concentration of 1.0 or 2.5 micrograms/ml did not significantly impair the induction of cells capable of suppressing the lymphoproliferative response of fresh autologous mononuclear cells to CMVFFx. The findings described above may have important clinical implications in that a degree of protective immunity to CMV-infected cells is maintained even in the presence of CsA.
Fungistatic mechanism of human transferrin for Rhizopus oryzae and Trichophyton mentagrophytes: alternative to simple iron deprivation
Authors: Artis W M , Patrusky E , Rastinejad F , Duncan R L
Published: January 2020

Human serum, human transferrin (TF), and the iron chelator 1,10-phenanthroline (OP) produce iron-reversible fungistatic activity which has been attributed to simple iron deprivation. In this study, the influence of the size of the inoculum on the inhibitory activity of serum, TF, and OP prepared with the same iron-binding capacity (2.5 micrograms/ml) for Rhizopus oryzae and Trichophyton mentagrophytes was examined. Inhibition was monitored in liquid microcultures maintained at 37 degrees C and pH 7.4 to 7.5 by measuring the change in absorbance density. Increasing the number of spores in the inoculum disrupted the fungistatic activity of serum and TF, but not that of OP. The dilution at which OP lost fungistatic activity was not affected by the number of spores in the inoculum and was the same for both fungi. The dilution at which TF and serum lost fungistatic activity was dependent upon both the quantity of the inoculum and the species of fungus. The number of viable spores, rather than the total number of spores in the inoculum, was determined to be important in overcoming the inhibition of fungal growth by serum and TF. The fungistatic activity of serum and TF could be diminished by the preexposure of the serum to viable but nongrowing spores. Direct and indirect fluorescence studies indicated that both T. mentagrophytes and R. oryzae absorbed TF. Glucose uptake by R. oryzae was inhibited by a 4-h exposure to 5.0 to 0.15 mg of apotransferrin per ml. These results suggest that the fungistatic activity of TF for R. oryzae and T. mentagrophytes may not be attributable to simple iron deprivation and raise the possibility of a requirement for a direct interaction.
In vivo replication of pathogenic and attenuated strains of Junin virus in different cell populations of lymphatic tissue
Authors: Laguens M , Chambó J G , Laguens R P
Published: January 2020

Lymphatic tissue is one of the main sites for replication of Junin virus. To characterize which cells are involved in that replication, the presence of Junin virus in purified populations of macrophages and dendritic cells from the spleens of guinea pigs infected with pathogenic and attenuated strains was investigated by immunofluorescence and intracerebral inoculation into newborn mice. The pathogenic strain was present both in macrophages and in dendritic cells, but the attenuated strain selectively infected dendritic cells. These observations suggest that the pathogenic behavior and replication efficiency of these two strains of Junin virus may be related to a difference in cell targets.
Beta-hemolytic activity of Trichomonas vaginalis correlates with virulence
Authors: Krieger J N , Poisson M A , Rein M F
Published: January 2020

The reasons that some women develop symptomatic trichomonal vaginitis, whereas many other infected women remain asymptomatic, are unclear, but it has been suggested that Trichomonas vaginalis strains vary in their intrinsic virulence. We describe beta-hemolytic activity in T. vaginalis which correlates with virulence in patients as well as in an animal model and in tissue culture. Fresh T. vaginalis isolates from four women with severe, symptomatic trichomoniasis had high-level (86.3 +/- 6.6%) hemolytic activity, whereas isolates from three completely asymptomatic women had low-level (45.3 +/- 8.4%) hemolytic activity (P less than 0.001). Hemolytic activity also correlated with the production of subcutaneous abscesses in mice (r = 0.74) and with destruction of CHO cell monolayers (r = 0.94). All of the 20 clinical isolates of T. vaginalis tested possessed hemolytic activity. The beta-hemolysin may be a virulence factor for T. vaginalis.
Attaching and effacing activities of rabbit and human enteropathogenic Escherichia coli in pig and rabbit intestines
Authors: Moon H W , Whipp S C , Argenzio R A , Levine M M , Giannella R A
Published: January 2020

Three strains of enteropathogenic Escherichia coli (EPEC), originally isolated from humans and previously shown to cause diarrhea in human volunteers by unknown mechanisms, and one rabbit EPEC strain were shown to attach intimately to and efface microvilli and cytoplasm from intestinal epithelial cells in both the pig and rabbit intestine. The attaching and effacing activities of these EPEC were demonstrable by light microscopic examination of routine histological sections and by transmission electron microscopy. It was suggested that intact colostrum-deprived newborn pigs and ligated intestinal loops in pigs and rabbits may be useful systems to detect EPEC that have attaching and effacing activities and for studying the pathogenesis of such infections. The lesions (attachment and effacement) produced by EPEC in these systems were multifocal, with considerable animal-to-animal variation in response to the same strain of EPEC. The EPEC strains also varied in the frequency and extent of lesion production. For example, three human EPEC strains usually caused extensive lesions in rabbit intestinal loops, whereas two other human EPEC strains usually did not produce lesions in this system.
Effect of sodium acetate on expression of K99 pili by Escherichia coli
Authors: Francis D H , Ryan C J , Fritzemeier J D
Published: January 2020

Sodium acetate suppressed K99 production in Escherichia coli strains cultured on a minimal medium, as determined by seroagglutination and enzyme-linked immunosorbent assay. The greatest suppression occurred when the medium contained both sodium acetate and glucose. Glucose alone did not suppress K99 production.
Prevalence of transformable Streptococcus mutans in human dental plaque
Authors: Westergren G , Emilson C G
Published: January 2020

A total of 100 strains of Streptococcus mutans serotypes c/e/f and d/g, freshly isolated from dental plaque, were screened for their ability to undergo genetic transformation to streptomycin resistance. Of the serotype c/e/f strains, 28% were found to be transformable, whereas none of the serotype d/g strains could be transformed by donor DNA from streptomycin-resistant S. mutans strains of either serotype c or d/g. Two of the transformable serotype c/e/f strains were transformed by DNAs from a variety of oral streptococcal species commonly found in the microflora.
Mechanisms determining innate resistance to ectromelia virus infection in C57BL mice
Authors: O'Neill H C , Blanden R V
Published: January 2020

The mechanism for innate resistance to ectromelia virus, which is controlled by a single gene in C57BL mice, was investigated. The cells or factors involved appear to be radioresistant and to impose an early barrier to viral penetration and spread via lymphatics or blood to the target organs, i.e., liver and spleen.
Susceptibility of Blastomyces dermatitidis strains to products of oxidative metabolism
Authors: Sugar A M , Chahal R S , Brummer E , Stevens D A
Published: January 2020

Three strains of Blastomyces dermatitidis which differ in their virulence for mice were exposed in their yeast form to various components of the peroxidase-hydrogen peroxide-halide system. Susceptibility to H2O2 alone correlated with virulence, with the most virulent strain (ATCC 26199) least susceptible (50% lethal dose, greater than 50 mM) and an avirulent strain (ATCC 26197) most susceptible (50% lethal dose less than 3.3 mM). A strain of intermediate virulence (ATCC 26198) was of intermediate susceptibility (50% lethal dose, 11.5 mM). The addition of a nontoxic concentration of KI (5 X 10(-4) M) did not increase H2O2 toxicity. However, the addition of either myeloperoxidase or horseradish peroxidase and KI markedly decreased the amount of H2O2 required to kill the organisms, with 100 +/- 0% of all strains killed at 5 X 10(-5) M H2O2 and 97 +/- 4, 100 +/- 0, and 94 +/- 8% of ATCC 26199, ATCC 26198, and ATCC 26197 killed, respectively, at 5 X 10(-6) M H2O2. Kinetic studies with H2O2 alone revealed a delayed onset of killing, but virtually 100% of organisms were killed by 120 min of exposure in all strains. By comparison, the peroxidase-hydrogen peroxide-halide system was 100% lethal for all strains at 1 min. The relatively high concentrations of H2O2 required to kill the yeast phase of B. dermatitidis suggest that H2O2 alone does not account for host resistance to the organism. However, the rapidly lethal effect of the peroxidase-hydrogen peroxide-halide system at physiologically relevant concentrations suggests that this may be one mechanism of host defense to B. dermatitidis.
Stimulation of calcium uptake and cyclic GMP synthesis in rat basophilic leukemia cells by Escherichia coli heat-stable enterotoxin
Authors: Knoop F C , Thomas D D
Published: January 2020

The effect of Escherichia coli heat-stable (ST) enterotoxin on calcium and cyclic nucleotide metabolism in rat basophilic leukemia cell cultures was investigated. Addition of ST enterotoxin to rat basophilic leukemia cell cultures resulted in dose- and time-dependent stimulation of calcium uptake and elevation of the intracellular cyclic GMP (cGMP) concentration. The effect of ST enterotoxin on calcium uptake (P less than 0.02) and cGMP synthesis (P less than 0.02) was demonstrated after 5 and 30 min of incubation at 37 degrees C, respectively. In further studies ST enterotoxin did not enhance calcium release or the intracellular concentration of cyclic AMP. The stimulation of calcium uptake and cGMP synthesis by ST enterotoxin was inhibited by pharmacological and chemical agents which block cellular calcium entry and prostaglandin synthesis. These results demonstrate that ST enterotoxin induces calcium uptake and cGMP synthesis in rat basophilic leukemia cell cultures.
Antibacterial activity of microbicidal cationic proteins 1 and 2, natural peptide antibiotics of rabbit lung macrophages
Authors: Lehrer R I , Selsted M E , Szklarek D , Fleischmann J
Published: January 2020

Microbicidal cationic proteins 1 and 2, peptides derived from rabbit lung macrophages, were tested for bactericidal activity against various bacterial species. Both were highly active against diverse gram-positive and gram-negative organisms under conditions of near-neutral pH (between 7 and 8) and relatively low ionic strength. Susceptible species included Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pneumoniae, Listeria monocytogenes, Pseudomonas aeruginosa, Klebsiella pneumoniae, Haemophilus influenzae, Escherichia coli, and Serratia marcescens. Streptococcus agalactiae, type 1A, was less susceptible than the aforementioned organisms or S. agalactiae, type 3. Bordetella bronchiseptica, a common commensal and pathogen of the rabbit respiratory tract, was completely resistant to both peptides.
Mapping of the antigenic determinants of Pseudomonas aeruginosa PAK polar pili
Authors: Watts T H , Sastry P A , Hodges R S , Paranchych W
Published: January 2020

The polar pili of Pseudomonas aeruginosa are flexible filaments 5.2 nm in diameter and 2.5 microns in average length. They consist of a single subunit, pilin, which is a 144-residue polypeptide containing a hydrophobic N-terminal region (residues 1 to 30) and eight hydrophilic regions distributed throughout the remainder of the molecule. To delineate the antigenic regions of pilin, we cleaved the protein at Arg30, Arg53, and Arg120 to produce peptides TCI (residues 1 to 30), TCII (31 to 53), TCIII (54 to 120), and TCIV (121 to 144). TCIII and TCIV were further cleaved into several subfragments. The purified peptides were coupled to bovine serum albumin by using the N-hydroxysuccinimide ester of 4-azidobenzoic acid and were then subjected to immunological analysis, using the enzyme-linked immunosorbent assay and immunoblot procedures with polyclonal antiserum. Four antigenic regions were identified; one in TCI was found to be common to both PAK and PAO pilin. The remaining three were found to be specific to PAK pilin. Two of these were subfragments of TCIII, whereas the third was located close to the C-terminus of the molecule, most likely between Cys129 and Cys142. Modification of these cysteines by reduction and carboxymethylation of the disulfide linkage did not abolish the antigenicity of the C-terminal type-specific antigenic determinant.
Pseudomonas aeruginosa isolates from patients with cystic fibrosis: a class of serum-sensitive, nontypable strains deficient in lipopolysaccharide O side chains
Authors: Hancock R E , Mutharia L M , Chan L , Darveau R P , Speert D P , Pier G B
Published: January 2020

Twenty-six Pseudomonas aeruginosa strains from patients with cystic fibrosis were typed by the Fisher immunotyping scheme. Only 6 strains were agglutinated by a single typing serum, whereas 15 strains were agglutinated with more than one serum and 5 were not agglutinated by any serum. Neither the polyagglutinable nor the nonagglutinable strains were typable by hemagglutination inhibition or immunodiffusion, suggesting that these polyagglutinable strains did not express multiple serotype antigens, but were instead being agglutinated by antibody to nonserotype determinants. Four typable isolates were resistant to pooled normal human serum, whereas the 12 polyagglutinable and nonagglutinable isolates studied were very sensitive to normal human serum. The outer membranes of 16 strains were isolated and characterized. The data suggested, in general, strong conservation of outer membrane protein patterns. Lipopolysaccharides (LPS) were purified by a new technique which allowed isolation of both rough and smooth LPS in high yields. Three of four typable, serum-resistant strains examined had amounts of smooth, O-antigen-containing LPS equivalent to our laboratory wild type, P. aeruginosa PAO1 strain H103. In contrast, 10 of 12 polyagglutinable or nonagglutinable, serum-sensitive strains had very little or no smooth, O-antigen-containing LPS, and the other two contained less smooth LPS than our wild-type strain H103. In agreement with this data, five independent, rough, LPS O-antigen-deficient mutants of strain H103 were nontypable and serum sensitive. We suggest that the LPS defects described here represent a significant new property of many P. aeruginosa strains associated with cystic fibrosis.
Molecular cloning and physical characterization of a chromosomal hemolysin from Escherichia coli
Authors: Welch R A , Hull R , Falkow S
Published: January 2020

A chromosomal hemolysin determinant was isolated in the form of recombinant plasmid pSF4000 from a human urinary tract isolate. Escherichia coli J96. A restriction endonuclease fragment map of pSF4000 was constructed. The ampicilin resistance transposon Tn/ was used as a site-specific mutagen in conjunction with in vitro derived deletions to localize the hemolysin determinant to a 7.0-kilobase region of pSF4000. Seventeen hemolytic E. coli isolates from human urinary tract infections were found to share similar DNA sequences with J96 hemolysin sequences, using a hemolysin-specific restriction endonuclease fragment as a hybridization probe. In all 17 hemolytic E. coli strains the hemolysin gene sequences were localized to the bacterial chromosome. The hemolysin-specific gene sequences can be found only in hemolytic strains by colony blot hybridization, suggesting that in the evolutionary sense the hemolysin-specific genetic sequences have only recently been introduced into the E. coli chromosome. Evidence is also presented that in the regions neighboring the J96 chromosomal hemolysin there are additional unique DNA sequences not commonly found in other E. coli isolates.
Longitudinal study of immune response to Pseudomonas aeruginosa antigens in cystic fibrosis
Authors: Döring G , Høiby N
Published: January 2020

During a 10-year period, the clinical states of 10 cystic fibrosis patients were evaluated on the basis of monthly measurement of lung function and weight; serum antibody titers to alkaline protease and elastase and the number of precipitins to Pseudomonas aeruginosa standard antigen were determined by radioimmunoassay and crossed immunoelectrophoresis. Alkaline protease and elastase concentrations of the P. aeruginosa strains from the patients were measured in vitro. The immune response increased in nearly all patients after the onset of chronic P. aeruginosa lung infection over years, suggesting unimpaired production of these antigens during P. aeruginosa lung infection, whereas the clinical states declined. The mean time for immune response was 15 months for alkaline protease, 11 months for elastase, and 6 months for standard antigen.
Virus-specific antibody activity of different subclasses of immunoglobulins G and A in cytomegalovirus infections
Authors: Linde G A , Hammarström L , Persson M A , Smith C I , Sundqvist V A , Wahren B
Published: January 2020

Healthy donors and patients with primary or reactivated cytomegalovirus infections were examined for antiviral antibodies of the immunoglobulin G (IgG) and IgA subclasses. For this purpose, monoclonal antibodies against the four IgG and the two IgA subclasses were used in enzyme-linked immunosorbent assays with cytomegalovirus antigen. IgG1 and IgG3 were the principal anti-cytomegalovirus subclasses in serum samples from healthy donors and patients. IgG1 was higher in reactivated than in primary disease. Cytomegalovirus-specific IgG2 was detected only in serum samples with high total cytomegalovirus IgG. A few patients with high total serum IgG4 also had cytomegalovirus-specific IgG4 antibodies. During convalescence, there was no significant increase in total IgG of any subclass. Both IgA1 and IgA2 antibodies with cytomegalovirus specificity were detected in patients who had cytomegalovirus antibodies of the IgA class.
Identification and characterization of Campylobacter jejuni outer membrane proteins
Authors: Blaser M J , Hopkins J A , Berka R M , Vasil M L , Wang W L
Published: January 2020

Outer membrane proteins from isolates of Campylobacter jejuni were examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Sarcosinate-insoluble membrane preparations were outer membrane enriched based on increased ketodeoxyoctonate concentrations, the presence of surface-exposed 125I-labeled proteins that were hydrophobic, and similarity to membrane vesicle (bleb) sodium dodecyl sulfate-polyacrylamide gel electrophoresis profiles. Most isolates contained a single major band with molecular weight of 41,000 to 45,000. Profiles of C. jejuni and Campylobacter coli isolates were indistinguishable, but either could be easily differentiated from Campylobacter fetus and Campylobacter faecalis. The profiles were stable for strains under a variety of growth, incubation and passage conditions. We classified 110 isolates from patients with sporadic campylobacter enteritis into nine subtypes based on differences in outer membrane sodium dodecyl sulfate-polyacrylamide gel electrophoresis profiles. Two categories accounted for 76% of the isolates. Complete concordance was observed in subtypes of strains obtained from epidemiologically related cases. Thus, comparison of the major outer membrane proteins of C. jejuni is a useful technique for investigating the transmission of this organism and may provide a basis for immunological characterization of the outer membrane proteins.
Legionella pneumophila-induced suppression of macrophage spreading in vitro
Authors: Friedman M , Klein T W , Friedman H
Published: January 2020

Incubation of normal mouse resident peritoneal cell suspensions rich in macrophages with Legionella pneumophila whole cell vaccine or soluble preparations thereof resulted in marked inhibition of the ability of the cells to spread on glass surfaces during a 24-h period. This inhibition, however, was transient in that by day 2 to 3 after culture initiation only partial inhibition was evident, and by day 4 to 5 thereafter most of the treated macrophage cultures showed normal spreading activity. Suppression of macrophage spreading was evident not only with intact Legionella preparations and the sonic extract but also with a lipopolysaccharide-rich somatic antigen preparation and flagella. The suppressive effects of Legionella preparations on a functional activity of normal macrophages in vitro indicate that these bacteria may have a detrimental effect on an important activity of cells involved in the immune defense system.
Stickland reactions of dental plaque
Authors: Curtis M A , Kemp C W , Robrish S A , Bowen W H
Published: January 2020

Dental plaque samples from monkeys (Macaca fascicularis) were shown to contain proline reduction activity in coupled Stickland reactions with other amino acids and also with certain end products of bacterial glucose metabolism. The unusually high concentration of bound and free proline in the oral environment may be of importance in both the production of base and in the removal of acid from the tooth surface after dietary carbohydrate ingestion.
Effects of antiinflammatory agents on chronic Salmonella typhimurium infection in a mouse model
Authors: Plant J E , Higgs G A , Easmon C S
Published: January 2020

Anti-inflammatory corticosteroids have been demonstrated to lower the resistance of the host to a range of infections in several different animal models. We have adapted the mouse model for Salmonella typhimurium infection, using a subcutaneous inoculation of 10(4) organisms in CBA/Ca mice. The chronic infection was potentiated by treatment with hydrocortisone given orally in the diet for up to 4 weeks, beginning on day 30 after infection. A dose of 25 mg/kg of body weight resulted in 100% mortality from Salmonella infection within 24 days of beginning hydrocortisone treatment. Nonsteroid drugs, indomethacin and a trial agent BW 755C, were compared with the corticosteroid in this model. No mortality was recorded during the 4 weeks of drug treatment in these groups. Fecal samples were monitored weekly, and drug-related effects on the normal fecal flora were observed. The kinetics of S. typhimurium infection were followed in both fecal and organ samples for the duration of the experiments. The model is useful for the determination of the lowered resistance of the host to infection after drug treatment and specifically demonstrated here the potential of the trial drug BW 755C as an active anti-inflammatory agent without one of the undesirable side effects of corticosteroids.
Antigenic analysis of H1N1 viruses isolated in the Houston metropolitan area during four successive seasons
Authors: Six H R , Webster R G , Kendal A P , Glezen W P , Griffis C , Couch R B
Published: January 2020

We evaluated the antigenic stability of viruses within the H1N1 subtype, using 307 isolates spanning four winter seasons. The isolates from each seasonal period appeared relatively homogenous in hemagglutination inhibition tests employing ferret antisera. However, a panel of seven monoclonal antibodies to A/USSR/90/77 detected extensive micro-heterogeneity. Antigenic variation occurred in the hemagglutinin antigen within a few weeks after the initial appearance of A/USSR/90/77-like strains in 1977-78 season as evidenced by low reactivity to one or several of the monoclonal antibodies. By the end of the season, few of the isolates reacted with the complete panel. Antigenic heterogeneity of the hemagglutinin was detected among H1N1 viruses throughout each epidemic interval. Some of the antigenic changes detected by monoclonal antibodies appeared on isolates in succeeding years, but others were restricted to a few strains isolated during a given year. The cumulative nature of some of these antigenic changes was indicated by the observation that 20 (19%) of 107 isolates examined in the fourth season failed to react with five of the monoclonal antibodies. Strains similar to most reference variants from around the world circulated in the Houston community.
Human antibody response to outer membrane proteins of Campylobacter jejuni during infection
Authors: Mills S D , Bradbury W C
Published: January 2020

Two techniques were used to isolate outer membrane proteins from Campylobacter jejuni, EDTA-lysozyme extraction and sodium-N-lauroylsarcosinate (Sarkosyl) solubilization. The protein profiles of the two preparations were similar, with a few additional bands in the EDTA-lysozyme preparations. The major outer membrane protein was 43,000 (43K) daltons, and there were 8 to 10 minor bands ranging from 92K to 14K daltons. There was no difference in the protein profile of a strain causing an infection (strain 17) and the resulting stool isolate (strain 17J). Sera collected before the infection and during the acute and convalescent stages were used with Western blotting and immunoautoradiographic techniques to determine the antigenicity of outer membrane proteins. A number of antigenic proteins were detected before the infection by their reaction with preinfection serum (61K, 51K, 43K, 40K, 34K, and 31K daltons), and three additional bands appeared during the infection when acute and convalescent sera were used (92K, 56K, and 19K daltons). Furthermore, an area of the gel at less than 14.4K daltons that did not stain with Coomassie brilliant blue became visible in the immune blots when the convalescent serum was used.
Chemiluminescent response to pathogenic organisms: normal human polymorphonuclear leukocytes
Authors: Robinson P , Wakefield D , Breit S N , Easter J F , Penny R
Published: January 2020

Chemiluminescence (CL) is a sensitive indicator of phagocytosis and intracellular killing; however, little is known of the normal CL response by human polymorphonuclear leukocytes to different pathogenic microorganisms. We investigated the luminol-enhanced CL response of normal polymorphonuclear leukocytes to a number of common bacterial pathogens and two yeasts. We analyzed the CL response to viable and heat-killed microorganisms at 25 and 37 degrees C. The CL response to all microorganisms was greater and more rapid at 37 degrees C. Variable responses were observed with viable and heat-killed microorganisms; some were unaffected, whereas other demonstrated reduced CL. Each microorganism caused a reproducible response pattern, which could be placed into two general categories. In the first category were those which caused a rapid exponential rise and decay in CL: Enterobacter cloacae, Salmonella typhimurium, Shigella flexneri, Staphylococcus aureus, Candida albicans, and zymosan. In the second category were those which rose slowly over a longer time course to a poorly defined peak: Pseudomonas aeruginosa, Klebsiella pneumoniae, Proteus mirabilis, and Streptococcus pyogenes. The CL response also reflected serum opsonic activity. The effect of inactivated complement, factor B, and removal of specific antibody were investigated. Increasing the concentration of zymosan gave a proportional rise in peak CL; however, a strain of E. coli caused a variation in peak time rather than peak height. Different CL kinetics were shown for three strains of K. pneumoniae, possibly a result of each having different membrane or cell wall characteristics. This study defines the nature and factors affecting the normal CL response to a variety of common pathogenic microorganisms.
Alterations in production of immunoglobulin classes and subclasses during experimental Trypanosoma cruzi infection
Authors: Jeng G K , Kierszenbaum F
Published: January 2020

The spleens of mice infected with Trypanosoma cruzi were tested for their contents of cells producing IgM, IgG1, IgG2a, IgG2b, and IgG3 specific for the trinitrophenyl hapten after immunization with trinitrophenyl-Ficoll and trinitrophenyl-bovine serum albumin at various times during the acute and chronic phases of the disease. Reduced splenic contents of all of these cells was the characteristic of the acute period, and a return to normal levels occurred during the chronic stage. However, the density of IgG2a- and IgG2b-producing plaque-forming cells in the spleen was not restored to normality until 130 days postinfection, i.e., long after the chronic phase had been attained, reflecting a diluting effect of splenomegaly on cells that produce immunoglobulin isotypes known to be cytotoxic for the blood form of the parasite.
Restriction endonuclease cleavage analysis of herpes simplex virus isolates obtained from three pairs of siblings
Authors: Sakaoka H , Aomori T , Ozaki I , Ishida S , Fujinaga K
Published: January 2020

Cleavage of herpes simplex virus type 1-infected cell DNA with restriction endonucleases showed that profiles of paired isolates obtained from three groups of siblings were essentially identical. This finding is different from that for profiles of other paired isolates that are epidemiologically unrelated. One set of paired isolates of the three had completely identical profiles. However, two other sets of paired isolates yielded minor heterogeneity in fragments derived from the regions spanning the unique repeat junction and repeated regions of the herpes simplex virus type 1 genome. This suggests that the regions of the herpes simplex virus genome are variable during viral DNA replication. This finding also supports the hypothesis that the same variants tend to be transmitted and thereafter perpetuate within a family cluster.
Toxoids of Pseudomonas aeruginosa exotoxin-A: photoaffinity inactivation of purified toxin and purified toxin derivatives
Authors: Callahan L T , Martinez D , Marburg S , Tolman R L , Galloway D R
Published: January 2020

For the preparation of greatly detoxified but highly immunogenic toxoids, two enzymatically active, low-toxicity derivatives of Pseudomonas aeruginosa exotoxin-A were further inactivated by photoaffinity labeling. These derivatives were formed during toxin purification, when a relatively crude toxin preparation was concentrated by ammonium sulfate precipitation and subsequently dialyzed. These derivatives, designated peak-1 protein (PK-1) and peak-2 protein (PK-2) were antigenically indistinguishable from native toxin, but had isoelectric points (5.00 and 4.90, respectively) that were different from that of the native toxin (4.95). Although the enzymatic activities and molecular weights of PK-1 and PK-2 were similar to those of native toxin, their toxicities were greatly reduced (ca. 500-fold). Photoaffinity labeling of fully active toxin-A, purified by a process which limits the formation of these derivatives, decreased its enzymatic activity (ca. 30-fold) and toxicity (ca. 100-fold). Likewise, photoaffinity labeling of purified PK-1 and PK-2 decreased their enzymatic activities and toxicities (ca. 30-fold and 100-fold, respectively) and, thus, yielded toxoids that were ca. 50,000-fold less toxic than unpurified native toxin. These toxoids were irreversibly detoxified and highly immunogenic during 9 months of storage at 4 degrees C.
Immunoenzymatic detection of three kinds of 43,000-molecular-weight antigens by monoclonal antibodies in the insoluble fraction of Toxoplasma gondii
Authors: Ogata K , Kasahara T , Shioiri-Nakano K , Igarashi I , Suzuki M
Published: January 2020

Monoclonal antibodies (TpM 3, TpM 6, and TpM 19) against Toxoplasma gondii insoluble antigens were produced by the hybridization of NS-1, a mouse myeloma cell line, with spleen cells from mice immunized with T. gondii insoluble antigens. TpM 3, TpM 6, and TpM 19 were characterized by the dye test, the latex agglutination test, two types of enzyme-linked immunosorbent assays, using either T. gondii supernatant antigens or T. gondii insoluble antigens, and immunoperoxidase staining. TpM 3, TpM 6, and TpM 19 antigens were analyzed by the immunoblotting method, using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and electrophoretic transfer of the antigens to nitrocellulose sheets. TpM 3, TpM 6, and TpM 19 were all negative by the dye test, the latex agglutination test, and the enzyme-linked immunosorbent assay, using T. gondii supernatant antigens but positive by the enzyme-linked immunosorbent assay, using T. gondii insoluble antigens. Each antibody gave unique spot patterns in the cytoplasm of tachyzoites, and each detected antigens of ca. 43,000 molecular weight with different electrophoretic patterns. Purified TpM 3 antigen bound only to TpM 3 but not to TpM 6 or TpM 19. Comparable results were obtained with TpM 6 and TpM 19 antigens. Rabbit anti-T. gondii sera reacted with both TpM 3 and TpM 19 antigens. However, human anti-T. gondii sera reacted only with TpM 3 antigen. These results indicate that T. gondii insoluble antigens contained at least three types of 43,000-molecular-weight antigens that have been revealed for the first time in this paper.
Functions of human neutrophilic granulocytes after in vivo exposure to interferon alpha
Authors: Einhorn S , Jarstrand C
Published: January 2020

The ability of neutrophilic granulocytes to phagocytize yeast particles and to reduce Nitro Blue Tetrazolium at rest and on activation with bacterial stimuli was monitored in 32 patients receiving treatment with human interferon alpha. The ability of these cells to attach to and ingest yeast particles was not altered to any major extent during 1 year of interferon treatment. In most patients, the Nitro Blue Tetrazolium-reducing activity increased after the first injection of interferon. During prolonged treatment with interferon alpha, 1 week to 1 year, granulocytes activated with bacteria exhibited a reduced Nitro Blue Tetrazolium activity in most patients.