bluuebunny/crossref_metadata_2025 huggingface dataset:
Influence of endotoxin-protein in immunoglobulin G isotype responses of mice to Brucella abortus lipopolysaccharide
Authors: Kurtz R S , Berman D T
Published: January 2020

Brucella abortus endotoxin preparations, containing approximately 5 to 6% protein, induce strong immune and adjuvant immunoglobulin G (IgG) responses as compared with Escherichia coli endotoxin preparations, with equivalent amounts of protein, which induce responses in which IgM antibody predominates. Using an enzyme-linked immunoassay with isotype-specific conjugates, we found that antibody of all four subclasses of IgG were evoked during the course of the immune responses of C3H/HeAu mice to B. abortus endotoxin. Secondary responses of endotoxin-hyporesponsive C3H/HeJ mice were similar to those seen in C3H/HeAu mice, although lower levels of antibody were produced during their primary responses. The primary responses of BALB/c athymic mice consisted almost entirely of IgG3, and IgG1 appeared following a second injection. The effects of lipopolysaccharide (LPS)-associated protein on the immunogenic properties of B. abortus endotoxin were examined by comparing responses to endotoxin with those to a purified B. abortus LPS containing less than 1% protein. The endotoxin evoked strong primary and secondary responses in which antibody directed to LPS determinants consisted mainly of IgG3 and those to the protein determinants were largely IgG1 antibody. Primary and secondary responses to purified LPS consisted mainly of IgG3 antibody. The potential mechanism of the contribution of protein to the immunogenic properties of the endotoxin as well as possible immune mechanisms involved in these responses are discussed.
Binding of purified and radioiodinated capsular polysaccharides from Cryptococcus neoformans serotype A strains to capsule-free mutants
Authors: Small J M , Mitchell T G
Published: January 2020

Strains 6, 15, 98, 110, and 145 of Cryptococcus neoformans serotype A vary in capsule size, animal virulence, and susceptibility to in vitro phagocytosis. The isolated capsular polysaccharides (CPSs) differ in monosaccharide composition ratios and molecular size, as determined by gel filtration. The purpose of this investigation was to characterize the binding of CPSs to capsule-free mutants of C. neoformans and to examine CPSs from these strains for differences in their ability to bind, to determine whether such differences might explain the variation in the pathobiology of these strains. CPSs were partially periodate oxidized, tyraminated, iodinated with 125I, and used in binding studies with two capsule-free mutants of C. neoformans, strain 602 and Cap59. Binding was specific for yeast species and for polysaccharide and was saturable, which is consistent with a receptor-mediated mechanism of attachment. Binding occurred rapidly and was only slowly reversible. Binding was also independent of pH from pH 5.5 to 8, of cation concentrations, and of competition by sugars up to 1.0 M concentrations. Only a portion of CPS was capable of binding, and strains varied in the extent to which their CPS bound. CPS-15-IV (peak IV was the major polysaccharide peak on DEAE-cellulose chromatography of CPS from strain 15) had the highest proportion of binding (40%), followed by CPS from strains 98, 6, 145, 110, and 15-III (peak III was an earlier eluting fraction of CPS from strain 15). The CPSs differed similarly in their ability to competitively inhibit binding. Treatment of CPS, but not yeast cells, with proteinase XIV abolished binding without altering the CPS gross structure. Treatment of yeast cells with proteases, heat, or formaldehyde did not alter binding, and both strain 602 and Cap59 bound CPS similarly. Binding to encapsulated yeast cells was minimal.
Characterization of neuraminidases produced by various serotypes of group B streptococci
Authors: Brown J G , Straus D C
Published: January 2020

Neuraminidase produced by 11 strains of group B streptococci (GBS), from serotypes Ia, Ib, Ic, II, and III, were characterized according to molecular weight, antigenic identity, and substrate specificity. Following growth in a chemically defined medium, ammonium sulfate-concentrated culture supernatants were assayed for activity with bovine submaxillary mucin as substrate. Neuraminidase produced by GBS strain 122 (serotype III) was purified by a combination of salt fractionation, affinity chromatography with Affi-Gel Blue, ion-exchange chromatography with DEAE-cellulose, and gel filtration on Sephadex G-200. Purified neuraminidase was used to immunize rabbits, and the resultant antiserum reduced the activity of purified neuraminidase from strain 122 by 87.7%. The antiserum also reduced the activity of neuraminidases produced by the other four serotypes by between 78.3 and 90%. Molecular weight estimates of the neuraminidases produced by the various serotypes were obtained by gel filtration chromatography on Sephadex G-200. The molecular weights obtained for the neuraminidases from the representative strains of each serotype ranged from 110,000 to 180,000. In addition, all of the GBS neuraminidases examined (regardless of the producing serotype) were active only on bovine submaxillary mucin. On the basis of these results, it appears that the neuraminidases produced by different GBS serotypes are quite similar.
Bordetella pertussis adenylate cyclase: effects of affinity-purified adenylate cyclase on human polymorphonuclear leukocyte functions
Authors: Friedman R L , Fiederlein R L , Glasser L , Galgiani J N
Published: January 2020

Affinity-purified adenylate cyclase (AC) of Bordetella pertussis, free of contaminating pertussis toxin, was demonstrated to have biological effects on human polymorphonuclear leukocytes (PMN). AC at doses of 25 and 50 micrograms/ml increased intracellular cAMP levels in the phagocytes 7.6- to 23.5-fold, respectively, above basal levels. AC inhibited PMN chemiluminescence, chemotaxis, and superoxide production in a dose-dependent manner. The 50% inhibitory dose for chemotaxis and chemiluminescence was 36.5 micrograms/ml; for superoxide generation it was 71.0 micrograms/ml. Although these PMN metabolic functions were impaired, no effect on phagocytic activity was observed.
Induction of mucosal immunoglobulin A immune response by preparations of Neisseria gonorrhoeae porin proteins
Authors: Jeurissen S H , Sminia T , Beuvery E C
Published: January 2020

The aim of this study was to develop an immunization scheme appropriate for the induction of an immunoglobulin A (IgA) response against Neisseria gonorrhoeae at the mucosae. For several reasons, the major outer membrane protein of N. gonorrhoeae (gonococcal PI) is attractive as a component of a gonococcal vaccine. Purified PI obtained from strain B2 was used in Zwittergent 3-14 for immunization. Rats received the antigen subcutaneously, intraintestinally, or directly in Peyer's patches with or without the adjuvant N-acetylmuramyl dipeptide (MDP) or AlPO4. The immune response was studied in situ by a newly developed antigen-specific three-step immunoperoxidase method, whereas specific antibodies in the serum were measured by an enzyme-linked immunosorbent assay procedure. Subcutaneous immunizations triggered peripheral lymphoid organs, whereas intraintestinal and intra-Peyer's patch immunizations triggered mucosa-associated lymphoid organs. This was reflected not only in the lymph nodes involved, popliteal versus mandibular and mesenteric lymph nodes, but also in the isotypes of the produced anti-PI antibodies, IgG versus IgA. The adjuvants AlPO4 and MDP appeared to act differently during subcutaneous and intraintestinal immunizations. AlPO4 augmented subcutaneous immune responses, whereas MDP had no effects. In contrast, intraintestinal immune responses increased most with the adjuvant MDP. In summary, we concluded that PI is capable of inducing a mucosa-associated IgA response when administered intraintestinally and that this response can be augmented by the adjuvant MDP.
Role of plasmid-encoded adherence factors in adhesion of enteropathogenic Escherichia coli to HEp-2 cells
Authors: Knutton S , Baldini M M , Kaper J B , McNeish A S
Published: January 2020

Plasmid-encoded adherence factors have been shown to be important for the full expression of enteropathogenic Escherichia coli (EPEC) pathogenicity and for EPEC adhesion to cultured HEp-2 cells. EPEC strain E2348 (O127) shows localized HEp-2 cell adhesion and possesses a 60-megadalton plasmid, pMAR2. When E2348 is cured of pMAR2 it loses the ability to adhere to HEp-2 cells, while nonadherent E. coli K-12 strains P678-54 and HB101 acquire HEp-2 adhesiveness after they gain the plasmid. By electron microscopy, E2348 was seen to adhere to HEp-2 cells in a manner that closely resembled EPEC adhesion to intestinal mucosa; bacteria were intimately attached to projections of the apical HEp-2 cell membrane and caused localized destruction of microvilli. The plasmid-containing K-12 strains, on the other hand, did not show intimate attachment and there was no modification of cell surface architecture. It is concluded that plasmid pMAR2 codes for an adhesin, possibly fimbrial in nature, that promotes HEp-2 adhesion but that other chromosomally encoded factors are required for EPEC to achieve the characteristic mode of intimate cell attachment.
Recognition of the galactose- or N-acetylgalactosamine-binding lectin of Entamoeba histolytica by human immune sera
Authors: Petri W A , Joyce M P , Broman J , Smith R D , Murphy C F , Ravdin J I
Published: January 2020

Cure of amebic liver abscess is associated with resistance to recurrent invasive amebiasis and the development of a humoral and cell-mediated immune response. We determined whether human immune sera contain blocking antibody for the 170-kilodalton (kDa) galactose or N-acetylgalactosamine (Gal/GalNAc)-binding lectin of Entamoeba histolytica. By Western blot (immunoblot) of whole amebae subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, all eight immune sera studied here prominently recognized a 170-kDa amebic protein. Western blot of the purified Gal/GalNAc lectin with pooled human immune sera (PHIS) confirmed that the 170-kDa band was the adherence lectin. Immunoprecipitation of [35S]methionine-metabolically-labeled amebae with the antilectin monoclonal antibody H8-5 and with PHIS demonstrated that the 170-kDa lectin was the major antigen recognized by PHIS. The in vitro adherence of E. histolytica trophozoites to CHO cells at 4 degrees C was inhibited by prior exposure of amebae to greater than or equal to 1.0% PHIS. The humoral response to the Gal/GalNAc-binding lectin of the parasite may contribute to the development of protective immunity against invasive amebiasis.
Identification and characterization of the Pasteurella haemolytica leukotoxin
Authors: Chang Y F , Young R , Post D , Struck D K
Published: January 2020

The identification and chromatographic characterization of the leukotoxin of Pasteurella haemolytica is described. The toxin, which has an apparent native molecular weight of greater than 400,000 as judged by gel exclusion chromatography, has a 105-kilodalton (105K) polypeptide as its major protein component. The proteolytic degradation of the 105K polypeptide could be correlated with the loss of toxin activity in aging cultures of P. haemolytica. Antisera raised against purified 105K polypeptide neutralized toxin activity. A 3.9-kilobase-pair fragment of the P. haemolytica genome cloned into a plasmid vector resulted in the production of intracellular toxin in Escherichia coli host cells. The restriction map of this clone shows significant overlap with the map of a previously reported leukotoxin clone (R. Y. C. Lo, P. E. Shewen, C. A. Strathdee, and C. N. Greer, Infect. Immun. 50:667-671, 1985). Finally, antisera raised against the 105K species labeled the P. haemolytica cell surface in a nonuniform, punctate manner.
Hypochlorous acid-promoted loss of metabolic energy in Escherichia coli
Authors: Barrette W C , Albrich J M , Hurst J K
Published: January 2020

Oxidation of Escherichia coli by hypochlorous acid (HOCl) or chloramine (NH2Cl) gives rise to massive hydrolysis of cytosolic nucleotide phosphoanhydride bonds, although no immediate change occurs in either the nucleotide pool size or the concentrations of extracellular end products of AMP catabolism. Titrimetric curves of the extent of hydrolysis coincide with curves for loss of cell viability, e.g., reduction in the adenylate energy charge from 0.8 to 0.1-0.2 accompanies loss of 99% of the bacterial CFU. The oxidative damage caused by HOCl is irreversible within 100 ms of exposure of the organism, although nucleotide phosphate bond hydrolysis requires several minutes to reach completion. Neither HOCl nor NH2Cl reacts directly with nucleotides to hydrolyze phosphoanhydride bonds. Loss of viability is also accompanied by inhibition of induction of beta-galactosidase. The proton motive force, determined from the distribution of 14C-radiolabeled lipophilic ions, declines with incremental addition of HOCl after loss of respiratory function; severalfold more oxidant is required for the dissipation of the proton motive force than for loss of viability. These observations establish a causal link between loss of metabolic energy and cellular death and indicate that the mechanisms of oxidant-induced nucleotide phosphate bond hydrolysis are indirect and that they probably involve damage to the energy-transducing and transport proteins located in the bacterial plasma membrane.
Role of cell-bound hemolysin as a pathogenicity factor for Serratia infections
Authors: König W , Faltin Y , Scheffer J , Schöffler H , Braun V
Published: January 2020

The hemolytic activities of clinical isolates of Serratia marcescens, of Serratia liquefaciens, and of Escherichia coli strains containing a cloned hemolysin gene of S. marcescens were determined. Hemolysis was induced only by cells and not by spent media. The hemolytically active bacteria induced the release of the leukotriene C4 and of much less leukotriene B4 from polymorphonuclear leukocytes, the release of histamine from rat mast cells, and chemoluminescence of neutrophils. The hemolytic activity was correlated with the response of the leukocytes, but quantitative differences were recorded with regard to the release of the inflammatory mediators. Therefore, other factors in addition to the hemolysin contribute to the stimulation of leukotriene generation and histamine release. It is concluded that the hemolysin via these inflammatory mediators can increase vascular permeability, edema formation, and granulocyte accumulation and thus contributes to the pathogenicity of Serratia species.
Protection of infant rats from Haemophilus influenzae type b infection by antiserum to purified outer membrane protein a
Authors: Loeb M R
Published: January 2020

Protein a (46,000 molecular weight [46K]) was purified from outer membranes of Haemophilus influenzae type b by a relatively simple procedure. Spontaneously shed outer membranes from a 24-h, 12-liter culture of an unencapsulated variant of strain Eag were combined with outer membranes released from the cells by Tris buffer and extracted with the nonionic detergent octylpolyoxyethylene. The extract was then subjected to open column chromatography on Sephacryl S-200 and Trisacryl-carboxymethyl to yield 7.5 mg of protein a from 180 mg of outer membrane protein. Approximately 99% of the protein in this preparation was protein a; in addition, the preparation contained 1.25% (wt/wt) lipopolysaccharide and had a residual detergent/protein ratio of 1.6:1 (wt/wt). Antibodies to the preparation were induced in rabbits by using alum as an adjuvant. As determined by immunoblotting, the great preponderance of antibodies induced were specific for protein a. However, very low levels of antibodies to several other outer membrane components, which were not apparent on gels of the pure preparation of protein a, were also induced. Preimmune and postimmune sera, after depletion of antibodies to capsular polysaccharide and lipopolysaccharide, were tested for biological activity against H. influenzae type b. Compared with preimmune serum, postimmune serum was bactericidal in vitro against strain Eag (the only strain tested) and offered significant protection (P less than 0.01) to infant rats against infection by all four strains tested, two of which had a protein a that was larger (47K) than the 46K protein a in the preparation. These results indicate that protein a should be considered as a vaccine to prevent H. influenzae type b disease.
Establishment and characterization of an antigen-specific T-cell line from liver granulomas of Schistosoma mansoni-infected mice
Authors: Ragheb S , Mathew R C , Boros D L
Published: January 2020

Granulomatous inflammations in schistosomiasis mansoni are the result of T-cell-mediated reactions to soluble egg antigens (SEA) secreted by parasite ova. To study TDH effector cell function, a granuloma T-cell line was established from collagenase-digested liver granulomas of acutely infected CBA/J mice. Dispersed nonadherent granuloma cells were cultured with feeder layer cells and SEA or with feeder layer cells alone in alternate cycles for 32 weeks. The granuloma T-cell line was L3T4+ Lyt-1+. In vitro, the SEA-stimulated T cells showed proliferation and interleukin 2 production. One million T cells adoptively transferred SEA-specific footpad swelling, and 7.5 X 10(6) T cells adoptively transferred granulomatous hypersensitivity to injected ova or SEA-coated beads. Anti-L3T4 monoclonal antibody blocked the SEA-specific cell proliferation. Depletion of L3T4+ cells abrogated, while that of Lyt-1+ cells diminished the adoptive transfer of SEA-specific footpad swelling. These experiments demonstrate that the granuloma T-lymphocyte population contains TDH-type effector cells. Establishment of an SEA-specific granuloma T-cell line will allow the study of the effector functions of the hitherto uncharacterized intralesional granuloma T lymphocyte.
Contribution of capsular polysaccharide and surface properties to virulence of Escherichia coli K1
Authors: Allen P M , Roberts I , Boulnois G J , Saunders J R , Hart C A
Published: January 2020

We examined the surface properties, susceptibility to the bactericidal activity of serum, and susceptibility to phagocytosis of Escherichia coli K1, a laboratory strain of E. coli (LE392), and strain LE392 carrying a plasmid (pKT274) incorporating a 17-kilobase insert of DNA that encodes the ability to produce surface K1 antigen. As determined by electron microscopy, LE392 was nonencapsulated but both E. coli K1 and LE392(pKT274) possessed a thin capsule. By using charged aqueous two-phase polymer systems, both E. coli K1 and LE392(pKT274) were shown to be significantly more negatively charged than LE392. E. coli K1 was resistant to the bactericidal activity of serum, but both LE392 and LE392(pKT274) were completely inhibited by neonatal serum at a concentration of 20% (vol/vol). As measured by counting endocytosed and nonendocytosed bacteria and by chemiluminescent response, E. coli K1 was highly resistant to phagocytic uptake by polymorphonuclear leukocytes, whereas LE392 was rapidly taken up by such cells; LE392(pKT274) was resistant to endocytosis, although less so than E. coli K1. Most intraphagocytic E. coli LK1 remained structurally intact for up to 60 min, whereas both LE392 and LE392(pKT274) were rapidly degraded.
Characterization of cyclophosphamide-rat model of cryptosporidiosis
Authors: Rehg J E , Hancock M L , Woodmansee D B
Published: January 2020

A cyclophosphamide-treated-rat model for cryptosporidiosis is described. The optimal conditions for inducing a cryptosporidial infection in rats were a 50 mg/kg dose of cyclophosphamide per day in the drinking water for a minimum of 14 days before inoculation with 10(4) or more infectious oocysts. Eighteen days after inoculation, 80% or more of the animals were infected. Cryptosporidia were attached to the terminal 24 cm of the small intestine, and the immunosuppressed animals remained infected as long as they received cyclophosphamide. However, the intestinal infection cleared within 7 to 10 days after cyclophosphamide was withdrawn. Oocysts continued to be detected in the feces for an additional 5 to 7 days after cryptosporidia were no longer visibly attached to the ideal enterocytes. This study characterized an animal model that can be used to investigate the pathogenesis of and evaluate therapeutic regimens for cryptosporidiosis in the immunocompromised host.
Influence of growth temperature on virulence of Legionella pneumophila
Authors: Edelstein P H , Beer K B , DeBoynton E D
Published: January 2020

The effect of growth temperature on the virulence of a strain of broth-grown serogroup 1 Legionella pneumophila (Wadsworth F889) was examined by growing the bacterium at different temperatures and then infecting guinea pigs (by intratracheal injection) and guinea pig alveolar macrophages. The 50% lethal dose for guinea pigs infected with 25 degrees C-grown F889 was log10 5.0 CFU and that for 41 degrees C-grown F889 was log10 5.7 CFU, or a fivefold difference. Guinea pig alveolar macrophages were infected in quadruplicate with log10 3.8 CFU of F889 cells grown at either 25 or 41 degrees C. Counts of F889 in the alveolar macrophages infected with 25 degrees C-grown bacteria were 40% greater after 1 day of incubation (P = 2 X 10(-4)) than were counts in the alveolar macrophage suspensions inoculated with 41 degrees C-grown bacteria. However, the counts were not significantly different after 3 days of incubation. Examination of cover slip cultures of guinea pig alveolar macrophages infected with 25 degrees C-grown or 41 degrees C-grown bacteria showed that the bacteria grown at the lower temperature were twice as likely to be macrophage-associated after 1 h of incubation than were the bacteria grown at the higher temperature. Growth at the lower temperature was also associated with a change in reactivity with monoclonal antibodies, but not with a change in plasmid content. Thus, environmental temperature may play an important role in modulating the virulence of L. pneumophila, possibly by affecting bacterial adherence to host cells.
Role of lipopolysaccharide and complement in susceptibility of Klebsiella pneumoniae to nonimmune serum
Authors: Ciurana B , Tomás J M
Published: January 2020

The role of lipopolysaccharide (LPS) in the susceptibility of Klebsiella pneumoniae to serum and the mechanism of complement activation by serum-susceptible (SerS) strains were investigated. The classical and alternative complement pathways are involved in serum killing of susceptible K. pneumoniae strains. The LPS composition seems to play a very important role in the serum bactericidal reaction, while capsular polysaccharide from this bacterium does not play any role. High-molecular-weight LPS from serum-resistant (Serr) K. pneumoniae strains was able to inhibit completely the serum bactericidal activity. LPS from SerS K. pneumoniae strains was not able to inhibit completely the serum bactericidal activity; low-molecular-weight LPS from Serr K. pneumoniae strains could not either. All these findings suggested that LPS composition, especially the O-antigen polysaccharide chains, contributes to the susceptibility of K. pneumoniae strains to complement-mediated serum bactericidal activity.
Intrapulmonary growth of Staphylococcus aureus in rats during induced atelectasis
Authors: Frederick D , Pesanti E L
Published: January 2020

Intrinsic pulmonary antibacterial defenses are mediated by alveolar macrophages and by noncellular factors. Mechanical ventilation in the resting tidal volume range leads to alterations in the physical characteristics of alveolar surfactant, alveolar instability, regional hypoxia, and systemic hypoxemia. While a number of experimental manipulations diminish the activity of the intrinsic antibacterial defense system, the effects of mechanical ventilation per se have not been systematically evaluated previously. We found that normal rats ventilated without sighing (periodic large breaths) manifested severe defects in pulmonary clearance of Staphylococcus aureus during 6-h experiments, such that growth of the inoculum occurred. Addition of a timer-controlled mechanism to cause the animals to sigh every 2 min, without other modifications in the experimental conditions, caused significant improvement in clearance. Analysis of cellular response, compartmentalization of viable bacteria, surfactant quantities and sedimentation characteristics, and protein influx indicated that the defect in clearance paralleled alterations in the physical state of surfactant and alveolar stability but was not strongly correlated with alterations in the other parameters we measured. The data show that defective pulmonary bacterial clearance is rapidly induced by measures which alter alveolar stability and suggest that intrinsic pulmonary defenses require maintenance of normal air-liquid interfaces for optimal function.
Metacyclogenesis is a major determinant of Leishmania promastigote virulence and attenuation
Authors: da Silva R , Sacks D L
Published: January 2020

The in vivo virulence patterns of promastigote populations defined on the basis of agglutination by the lectin peanut agglutinin (PNA) were studied for various cloned lines of Leishmania major. Promastigotes derived from logarithmic-phase cultures, which were routinely 100% agglutinated at 100 micrograms of PNA per ml, were relatively avirulent for BALB/c mice. The relative virulence of stationary-phase promastigotes appeared to be attributable to the proportion of nonagglutinable (PNA-) promastigotes contained within these populations. Purification of PNA- organisms from stationary cultures provided for each clone the most virulent inoculum, supporting the view that this change in lectin binding accurately reflects the development of infective metacyclic stage promastigotes. By studying this marker, we found that there was considerable variation in the degree to which different strains and clones underwent metacyclogenesis during growth. Examination of a reportedly avirulent L. major clone revealed that metacyclogenesis was unusually delayed and inefficient for this clone, but that those PNA- promastigotes which could be recovered from late-stationary-phase cultures were virulent for BALB/c mice. The loss of virulence associated with frequent subculture could also be attributed to a drastic diminution in metacyclogenesis potential over time. A clone which yielded over 90% PNA- promastigotes during growth within passage 1 generated fewer than 10% PNA- promastigotes during growth by passage 94. Subcloning of late-passage attenuated promastigotes yielded a clone for which no PNA- promastigotes could be generated during growth, and an infective population could not be derived from this clone. Thus, metacyclogenesis does not appear to be stable for even cloned lines of Leishmania promastigotes, and virulence comparisons between different strains and clones can be meaningfully made only if the metacyclic populations contained within the respective inocula are determined.
Isolation and function of a Clostridium perfringens enterotoxin fragment
Authors: Horiguchi Y , Akai T , Sakaguchi G
Published: January 2020

A fragment was obtained by treating Clostridium perfringens enterotoxin with 2-nitro-5-thiocyanobenzoic acid, a reagent which specifically cleaves the amino-terminal peptide bond of cysteine residues. The fragment (molecular weight, 15,000) was purified by high-performance liquid chromatography. The fragment had no cytotoxic effect on Vero cells but competitively inhibited enterotoxin-induced 51Cr release. Binding of 125I-labeled fragment to Vero cells was comparable to that of enterotoxin. Moreover, 125I-labeled fragment did not bind to FL cells, which lack receptor for enterotoxin. We conclude that the fragment contains the binding domain of enterotoxin. The amino acid composition of the fragment suggests that it is located on the carboxyl-terminal part of enterotoxin.
Mycobacterium leprae surface components intervene in the early phagosome-lysosome fusion inhibition event
Authors: Frehel C , Rastogi N
Published: January 2020

Bone marrow-derived cultured macrophages were infected with Mycobacterium leprae. The bacteria were either used as freshly isolated organisms or incubated with M. leprae antiserum (1:5) for 30 min prior to phagocytosis. Immediately after inoculation (1 to 4 h) and at 1 to 8 days later, macrophages were stained for acid phosphatase activity to assess fusions between phagosomes and lysosomes. Inhibition of fusions was essentially apparent as an early event, which was partially reversed by antiserum treatment of the bacteria, suggesting a role for M. leprae immunogenic surface components in this early phenomenon. Later incubation times (1 to 8 days) did not show any considerable difference between antiserum-treated and nontreated bacteria. The formation of an electron-transparent zone around phagocytized bacteria and its role in phagosome-lysosome fusion was investigated, and a direct relationship could not be established.
Localization of circumsporozoite protein of Plasmodium ovale in midgut oocysts
Authors: Nagasawa H , Procell P M , Atkinson C T , Campbell G H , Collins W E , Aikawa M
Published: January 2020

Circumsporozoite (CS) proteins are the major proteins found on the surface of salivary gland sporozoites and are the protective antigens of several species of malaria parasites. Little is known about the distribution of CS proteins in developing oocysts, however. Immunoelectron microscopy with protein A-gold and a monoclonal antibody specific for the CS protein of Plasmodium ovale was performed to investigate the distribution of CS protein within developing P. ovale oocysts. There was an almost complete absence of label in immature oocysts prior to the development of sporoblasts. In contrast, sporoblasts and budding and free sporozoites in mature oocysts were labeled uniformly on the outer surfaces of their plasma membranes, indicating a uniform distribution of CS protein on these membranes. Gold particles were frequently associated with the cytoplasm of sporoblasts and sporozoites, as well as with the inner surface of the oocyst capsule. This is the first evidence that CS protein is present in oocyst sporozoites and sporoblasts of P. ovale.
Influence of Staphylococcus aureus antibody on experimental endocarditis in rabbits
Authors: Greenberg D P , Ward J I , Bayer A S
Published: January 2020

To evaluate the potential protective benefit of antibody to whole cells of Staphylococcal aureus for the prevention of endocarditis, the rabbit endocarditis model was used. Methicillin-sensitive (17A) and methicillin-resistant (173) S. aureus strains were evaluated in rabbits with or without indwelling intracardiac catheters. All immunized rabbits developed significant homologous agglutinating antibody titers (the mean reciprocal titers were 15,300 to strain 17A and 1,150 to strain 173). After challenge, virtually no significant differences were observed between immunized and unimmunized animals with respect to (i) incidence of endocarditis, (ii) concentration of bacteria in infected vegetations, (iii) incidence of metastatic renal abscesses, or (iv) concentrations of bacteria in infected kidneys. The clearance of homologous S. aureus strains from blood cultures was similar for immunized and unimmunized animals at 10 to 90 min after intravenous challenge. In vivo adherence of homologous S. aureus strains to aortic valves and vegetations was similar in immunized and unimmunized animals when evaluated at 30 and 90 min postchallenge. Even without catheterization, the incidence of bacteremia and renal abscesses was the same in immunized and unimmunized rabbits. Whole-cell-induced S. aureus antibody did not prevent or modify any stage in the development of endocarditis in rabbits.
Differential binding kinetics of cholera toxin to intestinal microvillus membrane during development
Authors: Lencer W I , Chu S H , Walker W A
Published: January 2020

A complete randomized block design was used to compare the binding kinetics of cholera toxin to developing rat enterocyte microvillus membranes prepared from newborn, 2-week-old, 4-week-old, and adult animals. Saturation-binding isotherms were generated on 16 independent samples (four blocks) under steady-state and reversible conditions. Scatchard analyses suggested positive cooperative binding to a single class of receptors, and the isotherms were analyzed by both the Hill-Waud and Michaelis-Menten functions. Receptor density varied significantly with age (P = 0.013). An abrupt rise in receptor density occurred after the neonatal period and normalized in the adult animal. The half-dissociation constant also varied significantly with age (P = 0.019). Microvillus membranes from suckling animals had a slightly higher apparent affinity than those from weaned animals. Neither receptor concentration nor membrane purity confounded these observations. Whereas age-related changes in apparent affinity correlated with cellular responses, changes in receptor density did not. This study suggests that developmental changes in membrane structure which influence binding affinity but not receptor density may, in part, contribute to the increased sensitivity of suckling rats to cholera toxin exposure.
Genetic control of the immune responsiveness to Streptococcus mutans by the major histocompatibility complex of the rat (RT1)
Authors: Niiyama T , Kojima H , Mizuno K , Matsuno Y , Fujii H , Misonou J , Natori T , Aizawa M , Oikawa K
Published: January 2020

The lymph node cells from 11 strains of rats, differing in the genotype of the major histocompatibility complex of the rat (RT1), were examined on the basis of their proliferative response to the cell wall antigen of Streptococcus mutans. The 11 rat strains fell into three groups: high, intermediate, and low responders. To demonstrate the influence of the major histocompatibility complex on immune responsiveness to S. mutans, further experiments were performed using the RT1-congenic rat strains WKAH.1L(LEW), WKAH. 1AV1(ACI), and WKAH.1J(LEJ), which differ only in the genotype of the RT1 region. Although the background genes of each strain were of WKAH origin, WKAH.1L(LEW) and WKAH.1AV1(ACI) rats showed a low response whereas WKAH.1J(LEJ) rats showed a moderate response to the S. mutans cell wall antigen. The results indicate that the immune response is controlled by the class II gene(s) in RT1. Furthermore, the RT1.D locus products were shown to play an important role in the restriction molecule, since a monoclonal antibody, HOK7, directed to the RT1.Dk locus products reduced the proliferative response of lymph node cells.
Polyclonal B-cell-activating factors produced by spleen cells of mice stimulated with a cell homogenate of Trypanosoma gambiense
Authors: Oka M , Ito Y
Published: January 2020

The effect of injection of a cell homogenate of Trypanosoma gambiense into mice on the production of soluble factors responsible for the induction of polyclonal B-cell activation (PBA) by their spleen cells was examined. PBA was induced by injection of the cell homogenate and was also detected in mice treated with either the serum or the spleen cells of mice treated with the cell homogenate. PBA-inducing activity became detectable in the serum and spleen cells as early as 12 h after injection of the cell homogenate, reached a peak on day 2, and then decreased. This activity was also detected in the culture medium of spleen cells obtained 2 days after injection of the cell homogenate. For determination of the type of spleen cells producing the PBA-inducing factor, the day 2 spleen cells were fractionated on the basis of differences in their adhesive properties. Spleen cells in the effluents from a Sephadex G-10 column (T and B cells) and a nylon wool column (T cells) and those adhering to a plastic flask (macrophages) all produced the PBA-inducing factor. The production of PBA-inducing factor by whole spleen cells and by cells adherent to the plastic flask was not affected by treatment with anti-Thy-1.2 antibody and complement. These data suggest that soluble factors derived from macrophages and T cells could contribute to the induction of PBA. The PBA-inducing activity in the conditioned medium was completely inactivated by treatment at pH 2.0, heating at 56 degrees C for 30 min, or exposure to 0.1% sodium dodecyl sulfate. The results are discussed in relation to cytokines that could affect B-cell activation.
Distribution of type 1 and P pili on uropathogenic Escherichia coli O6
Authors: Gander R M , Thomas V L
Published: January 2020

The distribution of type 1 and P pili on individual cells of an O6 uropathogenic Escherichia coli strain, 6260, was determined immunologically with pilus-specific monoclonal antibodies by indirect immunofluorescence and immunogold electron microscopy. Variations in pilus expression under different culture conditions were monitored with an indirect immunofluorescence assay; 63% of piliated cells expressed type 1 pili when grown on agar at 37 degrees C versus 14 to 38% when grown in broth at 37 degrees C. In contrast, generally fewer cells with P pili (18 to 44%) were detected on agar than when grown in broth (up to 86%). Both type 1 and P pili were absent from cells cultured at 20 degrees C. Immunogold and immunofluorescence double labeling techniques with monoclonal antibodies 11-2 and 91-1 were used to study subpopulations of cells with type 1 and P pili; 39 to 41% of the piliated cells demonstrated only type 1 pili, and 12 to 16% of the cells showed only P pili. The immunogold method proved more sensitive than the immunofluorescence technique for detecting subpopulations expressing both pili types simultaneously, 19 versus 7%. We observed variations between type 1 and P pili, both in expression on individual cells and in the distribution of subpopulations of cells.
Erythrocytes as a source of essential lipids for Treponema hyodysenteriae
Authors: Stanton T B , Cornell C P
Published: January 2020

Mammalian erythrocytes were tested as a nutrient source for the spirochete Treponema hyodysenteriae, the etiologic agent of swine dysentery. Brain heart infusion (BHI) broth did not support growth of T. hyodysenteriae B204 or B78. However, BHI broth supplemented with washed bovine erythrocytes, erythrocyte membranes, or cholesterol was an excellent culture medium for these strains (4 X 10(8) to 10(9) cells per ml, final population densities). Small amounts of cholesterol (3 to 10 micrograms/ml of medium) were required for maximum cell yields of strain B204. Of various sterols and sterol-like compounds tested, cholestanol and sitosterol could substitute for cholesterol. BHI-dehydrated medium extracted with CHCl3-CH3OH (2:1, vol/vol) to remove lipids was used to prepare lipid-depleted culture broth. Lipid-depleted broth supplemented only with cholesterol did not support T. hyodysenteriae B204 growth. However, this same broth supplemented either with vesicles made from cholesterol and phosphatidylcholine or with washed erythrocytes was a good culture medium for the spirochete. Thus, both cholesterol and a phospholipid are needed for T. hyodysenteriae growth in lipid-depleted broth. Mammalian erythrocytes can be used by the spirochete as a source of these lipids.
Malaria transmission-blocking immunity induced by natural infections of Plasmodium vivax in humans
Authors: Mendis K N , Munesinghe Y D , de Silva Y N , Keragalla I , Carter R
Published: January 2020

Immunity to malarial infections in human populations is known to affect the development of the asexual blood stages of the parasites in the human host and to be capable of conferring significant protection against morbidity and mortality due to the disease. In this study we show that during acute infection with Plasmodium vivax malaria, one of the two main malarial pathogens of humans, most individuals also develop immunity that suppresses the infectivity of the sexual stages of the parasite to mosquitoes. The immunity is antibody mediated and is directed against the parasites in the mosquito midgut shortly after ingestion of blood by a mosquito. This immunity could be expected to have significant effects on the natural transmission of P. vivax malaria.
Induction of tumor necrosis factor by Legionella pneumophila
Authors: Blanchard D K , Djeu J Y , Klein T W , Friedman H , Stewart W E
Published: January 2020

Mice were inoculated with Legionella pneumophila via an intratracheal route to establish an experimental model of infection. Lung lavage fluid obtained from infected mice contained a cytolytic factor identified as tumor necrosis factor (TNF). Peak levels of TNF were produced at about 24 h postinfection and rapidly declined thereafter. Treatment of the mice with dextran sulfate before inoculation with the bacteria resulted in lowered amounts of TNF in the lung lavage fluid, suggesting that macrophages were responsible for production of the cytokine. Furthermore, cultures of adherent lung leukocytes and a macrophage cell line, PU 5-1.8, were stimulated to produce TNF by exposure to Legionella antigens. In addition, adherent lung leukocytes from Legionella-infected mice spontaneously released TNF into the culture supernatant. Inoculation of mice with saline or latex particles failed to induce TNF in vivo, indicating that bacterial antigens or products were the stimulating signals. Since there was no detectable TNF activity in sera at any time after intratracheal inoculation, TNF production appeared to be confined to the site of infection. Pretreatment of PU 5-1.8 cultures with gamma interferon, which was detected in the lung lavage fluid before TNF, resulted in augmented TNF production, suggesting cooperativity may exist between the two cytokines, either in the pathogenicity of the bacterium or in a possible immunomodulatory function of TNF and interferon during infection.
Mycobacterium leprae-burdened macrophages are refractory to activation by gamma interferon
Authors: Sibley L D , Krahenbuhl J L
Published: January 2020

Mycobacterium leprae grows to enormous numbers in the nu/nu mouse footpad, producing granulomas resembling those of lepromatous leprosy in humans. Footpad granuloma cells gorged with M. leprae were established in primary cell culture to examine their functional capabilities. These cells were classified as macrophages by the following criteria: positive staining for nonspecific esterase, reduction of Nitro Blue Tetrazolium during phagocytosis of Candida albicans, possession of Fc receptors, and possession of Mac-1 antigen. Footpad macrophages also phagocytized and supported the intracellular growth of Toxoplasma gondii. However, unlike peritoneal macrophages, footpad macrophages could not be activated to kill or inhibit T. gondii by macrophage-activating factor produced by mitogen-stimulated spleen cells or by recombinant gamma interferon. Thus, although the lepromatous macrophages appeared to be normal in many of their functions, they were defective in response to macrophage-activating signals.
Aromatic alpha-glycosides of mannose are powerful inhibitors of the adherence of type 1 fimbriated Escherichia coli to yeast and intestinal epithelial cells
Authors: Firon N , Ashkenazi S , Mirelman D , Ofek I , Sharon N
Published: January 2020

Adherence of bacteria via their surface lectins to host epithelial cells is considered an important initial event in bacterial pathogenesis. Mannose-specific (type 1) fimbriae are among the most commonly found lectins in enterobacteria. We studied the effect of aromatic alpha-glycosides of mannose on the agglutination of mannan-containing yeasts by different strains of Escherichia coli and on the adherence of the bacteria to guinea pig ileal epithelial cells. In both systems these compounds were considerably more effective inhibitors than methyl alpha-mannoside, with 4-methylumbelliferyl alpha-mannoside and p-nitro-o-chlorophenyl alpha-mannoside being the strongest inhibitors. Both compounds were approximately 400-times stronger inhibitors of yeast agglutination by E. coli O128 than was methyl alpha-mannoside and 1,000- and 470-fold stronger, respectively, than was methyl alpha-mannoside in inhibiting the adherence of the bacteria to ileal epithelial cells. 4-Methylumbelliferyl alpha-mannoside was 540 to 1,000 times more effective in inhibiting yeast agglutination by four additional strains of mannose-specific E. coli. It was also more efficient than methyl alpha-mannoside in removing adherent E. coli O128 from ileal epithelial cells. Our results provide further evidence that type 1 fimbriae of E. coli possess a hydrophobic region next to the mannose-binding site. The results suggest that 4-methylumbelliferyl alpha-mannoside and p-nitro-o-chlorophenyl alpha-mannoside are good candidates for the design of therapeutic agents that may prevent adherence in vivo and infection by E. coli strains that express type 1 fimbriae.
Nucleotide sequence homology to pertussis toxin gene in Bordetella bronchiseptica and Bordetella parapertussis
Authors: Marchitto K S , Smith S G , Locht C , Keith J M
Published: January 2020

Multiple strains of Bordetella parapertussis and B. bronchiseptica were examined for the presence of nucleotide sequences which hybridized with a cloned 4.5-kilobase (kb) fragment of B. pertussis DNA containing the genes responsible for pertussis toxin expression. All six B. parapertussis strains tested had nucleic acid sequences that hybridized with the cloned 4.5-kb fragment in Southern blot analyses. When the B. parapertussis DNA was digested with restriction endonuclease PstI, the pattern of hybridization was identical to that obtained with B. pertussis. Only five of the seven B. bronchiseptica strains tested had sequences that hybridized with the 4.5-kb fragment. Three of these B. bronchiseptica strains had a hybridization pattern identical to B. pertussis upon PstI digestion and Southern blot analyses. Two B. bronchiseptica strains were shown to lack a PstI cleavage site downstream from the region analogous to that coding for the pertussis toxin structural genes. Monoclonal antibody analyses were unable to detect pertussis toxin subunits S1 and S2 in Western blots with cellular material or culture supernatant from several B. bronchiseptica and B. parapertussis strains that possessed the DNA homologies. In addition, preliminary Northern hybridizations with RNA isolated from B. bronchiseptica and B. parapertussis strains suggested that the homologous regions were not transcribed. The data show that the gene coding for the toxic component of B. pertussis is common in other Bordetella species, though the gene probably is not expressed.
F41 pili as protective antigens of enterotoxigenic Escherichia coli that produce F41, K99, or both pilus antigens
Authors: Runnels P L , Moseley S L , Moon H W
Published: January 2020

Pigs suckling dams that have been vaccinated with pilus antigen are protected against challenge with enterotoxigenic Escherichia coli (ETEC) strains that express the same pilus antigen. However, some ETEC strains express more than one pilus antigen. Pregnant swine were vaccinated either with E. coli HB101 that harbored a recombinant plasmid coding for F41 expression (F41+) or with the HB101 parent strain that carries the pHC79 vector (F41-). Suckling pigs born to vaccinated dams were challenged with ETEC that expressed either K99, F41, or both pilus antigens. Production of F41 in vivo was demonstrated by immunofluorescence assay of sections of ileum and by seroconversion against F41 antigen by pigs challenged with F41+ and K99+ F41+ ETEC strains. The F41+ vaccine protected against challenge with an F41+ ETEC strain. In contrast, F41+ vaccination did not protect against challenge with K99+ or K99+ F41+ ETEC strains. The F41- vaccine did not protect against challenge with any strain used. The results indicate that K99+ F41+ ETEC strains produce F41 antigen in the small intestine during disease and that F41+ vaccination can be a protective antigen if the challenge strain expresses only F41 antigen, but that F41+ vaccination may not protect against strains that produce both K99 and F41 antigens.