bluuebunny/crossref_metadata_2025 huggingface dataset:
Growth characteristics and pathogenesis of experimental Pneumocystis carinii pneumonia
Authors: Walzer P D , Powell R D , Yoneda K , Rutledge M E , Milder J E
Published: January 2020

Pneumocystis carinii pneumonia was produced in two groups of rats by the administration of corticosteroids, a low-protein (8%) diet, and tetracycline in the drinking water. A third group not on corticosteroids or a low-protein diet served as controls. Members of the first group were sacrificed weekly for 8 weeks, and lungs were examined. A highly significant correlation was found between the histopathological assessment of the intensity of P. carinii infection and the number of cysts counted in enzyme-digested lungs. P. carinii progressively filled alveoli, and cyst counts increased from less than or equal to 10(4) to 10(9) cysts/g of lung at peak intensity of infection at 7 to 8 weeks. The second group of rats was placed on a regular diet and tapering doses of corticosteroids after week 4, and they were sacrificed at varying intervals for up to 21 weeks. P. carinii was not cleared from the lungs until after week 13 (more than 6 weeks after discontinuation of all steroids). Histologically, there was an increased prominence of alveolar macrophages and the progressive development of interstitial mononuclear cell infiltrate and fibrosis. Thus, P. carinii grows grows slowly in vivo and interacts with specific host cells. The resulting changes may be important in the pathogenesis of the infection and in the clearance of the organism from the lung after immunocompetence has been restored.
Alveolar Macrophage Ingestion and Phagosome-Lysosome Fusion Defect Associated with Virus Pneumonia
Authors: Jakab George J. , Warr Glenn A. , Sannes Philip L.
Published: January 2020

Virus-induced suppression of pulmonary phagocytic defenses is associated with defects in the intracellular processing of bacteria by alveolar macrophages. To determine whether the intracellular defect is related to a failure in phagosomelysosome fusion, mice were infected with a sublethal dose of Sendai virus, and the capacity of phagocytic cells, obtained by lung lavage, to exhibit phagosomelysosome fusion was quantitated during the course of the viral infection. Lysosomes of alveolar macrophages were prelabeled with acridine orange, the cells were challenged with
Candida krusei
, and fusion was determined with fluorescence microscopy by the discharge of the dye into the yeast-containing phagosome. Ultrastructural cytochemical studies verified the validity of the fluorescent fusion assay. Simultaneous experiments were performed to determine whether the viral infection also suppressed phagocytic ingestion by alveolar macrophages. Phagosome-lysosome fusion was progressively inhibited during the viral infection, reaching a low at day 7 when only 13 ± 3% of the phagocytic cells fused as compared with 97 ± 3% in cells from uninfected control animals; respectively, 55 ± 5% as compared with 74 ± 2% of the phagocytic cells contained yeasts. Thereafter, phagosome-lysosome fusion progressively increased reaching near normal levels (92 ± 3%) on day 17 of the infection. At the same time period, phagocytic uptake was enhanced to a level where 97 ± 3% of the cells contained yeasts. These data demonstrated that virus-induced suppression of intrapulmonary killing of bacteria involves functional lesions that retard the ingestion of inhaled organisms by alveolar macrophages and inhibit intracellular processing by degradative lysosomal enzymes by interfering with phagosome-lysosome fusion.
Role of antibody and complement in the control of Encephalitozoon cuniculi infections by rabbit macrophages
Authors: Niederkorn J Y , Shadduck J A
Published: January 2020

The capacity of mononuclear peritoneal macrophages to phagocytose Encephalitozoon cuniculi was tested in vitro. Normal rabbit serum or cell culture medium had little effect on the rate of removal of organisms by rabbit peritoneal macrophages. Treatment with immune rabbit serum or immune rabbit immunoglobulin G significantly (P less than 0.001) increased phagocytosis of E. cuniculi. Guinea pig complement was found to significantly (P less than 0.001) enhance the phagocytosis of antibody-treated E. cuniculi. With few exceptions, induced (peritoneal exudate) macrophages were no more effective than unstimulated (resident) macrophages in the phagocytosis of E. cuniculi. Secondary lysosomes labeled with ferritin were seen fusing with phagosomes containing immune rabbit serum-treated parasites. Phagosome-lysosome fusion was not observed when parasites were treated with either normal rabbit serum or culture medium. The results of the present study suggest a role for antibody enhancement of phagocytosis and intracellular killing as a mechanism of resistance to encephalitozoonosis in rabbits.
Enhancement of cellular protein synthesis sensitivity to diphtheria toxin by interferon
Authors: Aboud M , Michalski-Stern T , Nitzan Y , Salzberg S
Published: January 2020

In attempts to determine whether, by analogy to cholera and tetanus toxins, diphtheria toxin (DT) can also relieve the antiviral effect of interferon (IF), we found that it rather enhanced the inhibitory effect of IF on the replication of murine leukemia virus in chronically infected NIH/3T3 cells. This enhancement was found to be a consequence of an increased sensitivity to DT of cellular protein synthesis in IF-treated cells. IF stimulated the anti-protein synthesis activity of DT in both mouse cells that are known to be highly resistant to this toxin and in human HeLa cells that are highly sensitive to this toxin. This stimulation was dependent on IF dose. The effect of IF on DT action was strictly species specific, indicating that it was not a consequence of the mere binding of IF to the cell membrane, but rather reflected the cellular changes that followed this initial binding. IF was found to be capable of potentiating intact DT, but could not potentiate its fragments in any combination. IF did not have any effect on the in vitro nicotinamide adenine dinucleotide glycohydrolase activity of DT, suggesting that the effect of IF is not due to molecular modification of the toxin.
Different polypeptide composition of two human rotavirus types
Authors: Espejo R , Martínez E , López S , Muñoz O
Published: January 2020

Human rotaviruses, which are placed into two groups according to their ribonucleic acid patterns obtained by gel electrophoresis, were characterized both by polypeptide components from purified virions and by polypeptides translated from their denatured ribonucleic acids in rabbit reticulocyte lysates. Viruses assigned to different groups differed in the electrophoretic migration of the second largest of the polypeptides which compose the inner shell; polypeptides that had been synthetized in vitro from ribonucleic acid from each group showed this same difference, thus indicating that this is due to the genomic composition. This study suggests that there are differences in the third largest polypeptide of the inner shell and also in the three smaller polypeptides composing the outer shell. We also demonstrated that there are differences in genomic and polypeptide compositions between simian (SA11) and calf (Nebraska calf diarrhea virus) rotaviruses grown in tissue culture and human rotaviruses.
Transfer of group A streptococcal pyrogenic exotoxin production to nontoxigenic strains of lysogenic conversion
Authors: Johnson L P , Schlievert P M , Watson D W
Published: January 2020

Production of group A streptococcal pyrogenic exotoxins (SPE) type A and C was transferred from toxigenic streptococcal strains to nontoxigenic strains by lysogeny. Lysogens were tested for SPE with Ouchterlony immunodiffusion on Todd-Hewitt agar plates; toxin diffusing from isolated colonies reacted with specific hyperimmune antisera to SPE. Phage prepared from strains T25(3) (T12gl) and 3GL16, both yielding SPE type A, formed plaques on T25(3) (NONLYSOGENIC) lawns. Over 90% of the colonies picked from the plaque centers yielded A toxin, suggesting SPE type A was transferred by lysogenic conversion. SPE type C formation was transferred to nontoxigenic strains T25(3) and K56 with supernatant fluids from mitomycin C-induced cultures of CS112, producing SPE types B and C. All lysogens tested were positive for SPE type C, indicating that C toxin induction also was transferred by lysogenic conversion. SPE type B formation was not transferable by lysogeny with the strains tested.
Clostridium difficile in gnotobiotic mice
Authors: Onderdonk A B , Cisneros R L , Bartlett J G
Published: January 2020

Germfree mice associated with Clostridium difficile developed intestinal disease characterized by polymorphonuclear cell infiltration of the lamina propria, diarrhea, and cecal cytotoxin concentrations positive at a 10(-6) dilution. The numbers of viable bacteria never exceeded 10(10) colony-forming units per g (dry weight). Despite the high toxin levels and chronic inflammation over a 30-day period, the mortality rate was low (less than 2%). Daily treatment of these animals with two oral doses of 2 mg of vancomycin resulted in stool levels of greater than 200 micrograms/ml, well in excess of the minimum inhibitory concentration for C. difficile. This therapy decreased viable cell density by 2 to 3 logs and increased the spore counts from 10(5.8) to 10(7.8) colony-forming units per g (dry weight) by day 7, and animals were free of detectable toxin. However, once therapy was stopped, viable bacteria and spore counts and cytotoxin concentrations returned to previous levels. Treatment of mice with concentrations of clindamycin shown to be inhibitory in vitro had no effect on C. difficile toxin titers or bacterial counts, although the appearance of a clindamycin-resistant population was noted. These data indicate that vancomycin, given orally, decreases the concentration of toxin, but C. difficile survive as spores. By contrast, large populations of vegetative cells and high cytotoxin levels persist when clindamycin is used, even at an inhibitory concentration.
Mitogenic Activity of Water-Soluble Adjuvant Obtained from
Bacterionema matruchotii
Authors: Nitta Toshimasa , Okumura Seiichi , Nakano Masayasu
Published: January 2020

A butanol-extracted water-soluble adjuvant (Bu-WSA) obtained from
Bacterionema matruchotii
, a gram-positive oral bacterium, is a potent B-cell mitogen for murine lymphocytes in vitro. [
3
H]thymidine uptake of cultured spleen cells of BALB/c mice and nude athymic mice was greatly enhanced by the presence of Bu-WSA. Spleen cells which had been treated previously with rabbit anti-mouse thymocyte serum and guinea pig complement responded to Bu-WSA, whereas thymocytes and nylon wool column-filtered spleen cells were unresponsive to it. It was necessary for the adjuvant to be in the culture for at least 24 h in order to obtain significant lymphocyte activation. As macrophage-depleted spleen cells still responded to Bu-WSA, the proliferative response of lymphocytes seems to be independent of the presence of macrophages. The degree of response of spleen cells stimulated by a mixture of Bu-WSA and lipopolysaccharide (LPS) was very close to the sum of the responses stimulated by Bu-WSA and LPS individually. Spleen cells of LPS-injected mice, which were refractory to LPS, responded to Bu-WSA. On the other hand, the spleen cells of Bu-WSA-injected mice responded to LPS but not to Bu-WSA. Furthermore, the cells obtained from the spleens of mice which had been heavily cobalt 60 irradiated and bone marrow reconstituted 3 weeks in advance responded to LPS but not to Bu-WSA. Therefore, it appears that the cells responding to Bu-WSA were not identical to those responding to LPS.
Participation of three major glycoprotein antigens of herpes simplex virus type 1 early in the infectious cycle as determined by antibody-dependent cell-mediated cytotoxicity
Authors: Norrild B , Shore S L , Cromeans T L , Nahmias A J
Published: January 2020

Tissue culture cells infected with herpes simplex virus type 1 synthesize three major glycoprotein antigens (Ag-11, Ag-8, and Ag-6), which have been characterized by crossed immunoelectrophoresis. The three viral antigens have been identified as a mixture of gA and gB (Ag-11), gD (Ag-8), and gC (Ag-6). Recent findings have shown that antibodies directed to each of the three antigens individually are able to mediate antibody-dependent cell-mediated cytotoxicity when tissue culture cells late in the infectious cycle (18 h postinfection) are used. In this work, antibody-dependent cell-mediated cytotoxicity was applied to study the time postinfection at which the individual viral antigens first made their appearance at the cell surface. All three viral antigens (Ag-11, Ag-8, and Ag-6) could be demonstrated as newly synthesized from 3 to 4 h postinfection, and the quantities of the antigens at the surfaces of the infected cells increased with time postinfection. The use of cycloheximide and ultraviolet-inactivated virus demonstrated that input virus could be detected by antibody-dependent cell-mediated cytotoxicity during the first 2 h postinfection, but the cytotoxicity caused by input virus remained constant with time postinfection. In conclusion, these observations demonstrate the participation of individual herpes simplex virus surface antigens in antibody-dependent cell-mediated cytotoxicity attack on cells early in infection.
Inhibition by interferon of Sendai virus cytolysis
Authors: Sato T , Tomita Y , Kuwata T
Published: January 2020

Treatment of HeLa cells with human interferons inhibited 51Cr release from cells induced by ultraviolet-inactivated Sendai virus. The inhibitory effect became apparent about 6 h after interferon treatment and persisted for 24 to 48 h. In the interferon-treated cells, the cytolysis was inhibited within 10 min after adding virus and the inhibitory action was suppressed by the treatment of the cells with cycloheximide. Mock interferon and mouse interferon did not inhibit the cytolysis and antiinterferon serum neutralized the effect of interferon. All these findings indicate that Sendai virus-induced cytolysis is inhibited by interferon per se. However, interferon did not have any influence on Sendai virus hemolysis.
Stepwise degradation of membrane sphingomyelin by corynebacterial phospholipases
Authors: Bernheimer A W , Linder R , Avigad L S
Published: January 2020

The mechanism of in vitro synergistic lysis of sheep erythrocytes by Corynebacterium ovis and Corynebacterium equi was investigated. Hemolysis required (i) the action of phospholipase D from C. ovis, (ii) the action of an extracellular protein of C. equi, and (iii) Mg2+. Maximum lysis required imposition on the system of a fourth condition (step iv), such as chilling. Steps i, ii, and iv occur sequentially and in that order. Mg2+ functions in steps i and ii. The extracellular protein C. equi was purified to homogeneity and found to be a phospholipase C capable of hydrolyzing ceramide phosphate, phosphatidic acid, and all of the isolated major phospholipids of mammalian erythrocyte membranes. The principal features of the synergistic hemolytic system could be reproduced in experiments involving liposomes containing either sphingomyelin or ceramide phosphate and trapped [14C]glucose. We inferred that sphingomyelin of sheep erythrocytes is first converted to ceramide phosphate by C. ovis phospholipase D. On the basis of results with liposomes, we propose that the ceramide phosphate is then converted to ceramide by C. equi phospholipase C. We believe that the resulting in situ ceramide then undergoes dislocation by chilling and perhaps also by virtue of an affinity between ceramide and C. equi phospholipase C. The dislocation of ceramide presumably disorganizes the lipid bilayer sufficiently to result in cell lysis.
Kinetics of Changes Induced by Indigenous Microbiota in the Activity Levels of Alkaline Phosphatase and Disaccharidases in Small Intestinal Enterocytes in Mice
Authors: Whitt Dixie D. , Savage Dwayne C.
Published: January 2020

The concentration of protein and the activities of alkaline phosphatase, maltase, and sucrase were measured in saline extracts of the proximal small intestine of germfree and ex-germfree mice colonized with an indigenous microflora. The two populations of animals were maintained in plastic film isolators under tightly controlled environmental and nutritional conditions. Samples were taken at 0, 4, 8, 12, and 17 days and at 3, 4, 5, 6, 7, and 8 weeks after association. The activities were expressed as specific activities and as total units per segment of small intestine. Enzymatic activities expressed in both ways fluctuated considerably in the samples taken from one time to the next in animals of both types. The activities expressed as total units per segment of bowel of all three enzymes had decreased from levels in germfree animals by as early as 4 days after association. The total units of activity per segment of bowel tested continued to decrease for approximately 3 weeks in the associated animals to levels two- to fivefold lower than those of germfree animals. However, the specific activities of the three enzymes in the animals of the two types became less disparate at later sample times. This latter result is predictable because the concentration of protein extractable from the small intestines of the mice of the two types was the same at the beginning of the experiment, but by the later sampling times, the concentration of protein extractable from small bowels of ex-germfree mice was significantly lower than that from germfree mice. The fluctuations in levels of the enzymatic activities, even under controlled environmental and nutritional conditions, point to the necessity of using such conditions and a kinetic approach in studies of the effects of the microbiota on the activities of enzymes in the microvillous membranes of small bowel enterocytes. The changes in protein concentrations suggest that such activities and the amounts of protein extractable from the mucosa are influenced by different properties of the microflora. Thus, studies in which the enzymes are extracted from the entire mucosa and the activities are expressed as units per weight of extractable protein may give misleading results concerning the influence of the microbiota on the enterocyte membranes.
Mannose-Sensitive and Mannose-Resistant Adherence to Human Uroepithelial Cells and Urinary Virulence of
Escherichia coli
Authors: Van Den Bosch Johannes F. , Verboom-Sohmer Ursula , Postma Peter , De Graaff Johannes , MacLaren David M.
Published: January 2020

The adherence to human uroepithelial cells of 23
Escherichia coli
strains belonging to three groups with different levels of virulence was investigated, and the mechanism of adherence was studied. It was found that strains belonging to the most virulent group adhered better to human uroepithelial cells than did avirulent strains. Adherence of loss virulent but supposedly nephropathogenic strains was more variable. These results suggest that adherence is an important virulence factor, especially in the group of strains with the highest but a more general virulence. Piliated strains adhered better than did nonpiliated strains. We found strong evidence for the existence of at least two different mechanisms of adherence: (i) mannose-sensitive adherence by piliated strains, very likely mediated by type I pili because this mannose-sensitive adherence was associated with mannose-sensitive hemagglutination of guinea pig erythrocytes by broth cultures of the strains; (ii) mannose-resistant adherence by piliated strains, very likely mediated by non-type I pili because this mannose-resistant adherence was invariably associated with mannose-resistant hemagglutination of human group A erythrocytes by the strains, whether grown in broth or on plates. Additionally, one strain without pili and without hemagglutinating activity adhered well. Thus in most cases adherence seemed to be mediated by bacterial pili, although different types might be involved.
Induction of L-phase variants of Nocardia caviae within intact murine lungs
Authors: Beaman B L
Published: January 2020

The data presented show that cells of Nocardia caviae 112 were converted to cell wall-deficient microbial variants within the intact murine lung after intranasal administration. At the time that these L-phase variants were recovered in large numbers from the lung, there was a correspondingly enhanced inflammation leading to alveolar consolidation and animal death. During the peak of this response (at 1 week after infection), normal nocardial cells were neither isolated from nor seen within the lung. It is suggested that the conversion of these normal nocardial cells to their L-phase variant leads to this extensive pulmonary damage. Furthermore, the L-phase organisms appear to play an active role in this pathological effect since introduction of similar amounts of killed nocardial cells into the lungs of the mice failed to produce a similar response.
Candida albicans
Ultrastructure: Colonization and Invasion of Oral Epithelium
Authors: Howlett Julie A. , Squier Christopher A.
Published: January 2020

The colonization and invasion of various animal oral mucosae by
Candida albicans
were examined in an organ culture model. Scanning and transmission electron microscopy of the oral epithelium between 12 and 30 h after inoculation with the fungus revealed the morphological relationships between host and parasite. Examination of the fungi in thin sections showed five distinct layers in the cell wall of
C. albicans
within the epithelium, but changes were evident in the organization and definition of the outer cell wall layers in budding hyphae and in hyphae participating in colonization and invasion of the epithelial cells. Adherence of the fungus to the superficial cells of the oral mucosa appeared to involve intimate contact between the epithelial cell surface and the deeper layers of the fungal cell wall. During invasion a close seal was maintained between the invading hyphae and the surrounding epithelial cell envelope, there being no other evidence of damage to the host cell surface except at the site of entry. Within the epithelial cells there was only occasional loss of cytoplasmic components in the vicinity of the invading hyphae. These findings would suggest that enzymatic lysis associated with the invasive process is localized and that the mechanical support provided by surface adherence and the intimate association between the fungus and the epithelial cell envelope may permit growth of
Candida
on through the epithelium.
Choriogonadotropin-like antigen in a strain of Streptococcus faecalis and a strain of Staphylococcus simulans: detection, identification, and characterization
Authors: Acevedo H F , Koide S S , Slifkin M , Maruo T , Campbell-Acevedo E A
Published: January 2020

The presence of choriogonadotropin- and alpha-subunit-like materials in two species of bacteria identified as Staphylococcus simulans and Streptococcus faecalis have been demonstrated by the indirect fluorescein-labeled and the indirect peroxidase-labeled immunocytochemical techniques, utilizing antiserum for human choriogonadotropin, for its alpha and beta-subunits and the bera-subunit COOH-terminal peptide. The bacteria were originally isolated from the urine of two patients with advanced forms of cancer. Chromatography done on the water-soluble extract of acetone powder preparations of the bacterial cultures revealed the presence of a material similar to the complete trophoblastic hormone and to its beta-subunit in the culture media of S. simulans, and to the beta-subunit in the media of S. faecalis. No free alpha-subunit was detectable. Furthermore, the choriogonadotropin-like factor demonstrated biological activity in in vivo assay systems. From the present results, it can be concluded that some species of "cancer-associated" bacteria can synthesize a human trophoblastic hormone-like glycoprotein with physicochemical properties similar to those of the human trophoblastic hormone that is biologically active and that is either released complete or as one of its subunits in the culture media.
Production of heat-labile or heat-stable enterotoxins by strains of Escherichia coli belonging to serogroups O44, O114, and O128
Authors: Scotland S M , Day N P , Cravioto A , Thomas L V , Rowe B
Published: January 2020

Strains of Escherichia coli which belong to enteropathogenic serogroups usually fail to produce heat-labile or heat-stable enterotoxins. However, 1 of 34 strains of E. coli O44, 9 of 45 strains of E. coli O114, and 18 of 82 strains of E. coli O128 produced heat-labile or heat-stable enterotoxins. Most enterotoxigenic isolates were from tropical or developing countries; all three enterotoxigenic strains isolated in Britain were from patients returned from abroad. Enterotoxigenic strains were of many different flagellar types. Certain enterotoxigenic strains of E. coli O114 and O128 possessed colonization factor antigen I.
Trypanosoma cruzi infection in B-cell-deficient rats
Authors: Rodriguez A M , Santoro F , Afchain D , Bazin H , Capron A
Published: January 2020

The effect of neonatally initiated injections of anti-mu rabbit antiserum on immunity of rats against Trypanosoma cruzi infection was investigated in vivo. Anti-mu treatment resulted in a loss of immunoglobulin M (IgM) and IgG2a synthesis and, subsequently, of antibody production. These rats so treated were shown to be significantly more susceptible to the acute phase of the infection than the control rats treated with normal rabbit serum, as measured by increased parasitemia and mortality. These results indicate the essential role of antibodies, probably in association with complement or effector cells or both, in immunity to acute Chagas' disease.
Partial purification and characterization of heat-stable enterotoxin produced by Yersinia enterocolitica
Authors: Okamoto K , Inoue T , Ichikawa H , Kawamoto Y , Miyama A
Published: January 2020

By using a suckling mouse assay, heat-stable enterotoxin (ST) was purified from the culture filtrate of Yersinia enterocolitica isolated from a diarrheal patient. The purification procedures involve ultrafiltration with an Amicon HIP-10 hollow fiber, ethanol fractionation, protamine sulfate treatment, diethylaminoethyl-Sephacel and hydroxylapatite column chromatographies, and Sephacryl S-200 superfine gel filtration. About 408-fold purification was achieved, with a yield of 12.0%. The minimal effective dose of purified ST was about 110 ng in the suckling mouse assay. The molecular weight of purified ST was 9,000 by Sephadex G-100 superfine gel filtration. The purified ST was stable to heating (100 degrees C for 20 min, 121 degrees C for 20 min) and did not lose its toxicity after treatment with protease, trypsin, lipase, phospholipase C, ribonuclease, deoxyribonuclease, beta-glucosidase, and neuraminidase. The purified ST was separated by isoelectric focusing into two active fractions, with pI's of 3.29 (ST-1) and 3.00 (ST-2), respectively. Antiserum from guinea pigs immunized with the purified ST neutralized the activity of both Y. enterocolitica ST and Escherichia coli ST.
Common polysaccharide antigens from the cell envelope of Clostridium perfringens type A
Authors: Dayalu K I , Cherniak R , Hatheway C L
Published: January 2020

Soluble antigens were obtained by extracting five serotype strains of Clostridium perfringens type A with water at 100 degrees C. The type-specific polysaccharides were precipitated with ethanol, and the common antigens were recovered from the ethanol supernatants by concentration, dialysis, and lyophilization. Refluxing the water-extracted cell residues with 1% acetic acid followed by concentration, dialysis, and lyophilization gave additional common antigen fractions. A comprehensive, side-by-side comparison of the antigen fractions, the ethanol precipitate, the ethanol supernatant, and the acetic acid supernatant, revealed that common antigens were recovered in all three fractions, and that three distinct entities were responsible for the formation of the observed common immunoprecipitin lines; whereas many fractions possessed all three immunoprecipitin lines, others contained only one or two. The serological homology observed between the various antigen fractions was apparently a consequence of N-acetylglucosamine- and N-acetylmannosamine-containing polymers. The common antigens were presumably associated with the cell envelope and may be the type of markers sought previously by others for the serological identification of C. perfringens.
Effect of herpes simplex virus infection on murine antibody-dependent cellular cytotoxicity and natural killer cytotoxicity
Authors: Kohl S , Lawman M J , Rouse B T , Cahall D L
Published: January 2020

Mice intraperitoneally inoculated with a sublethal dose of herpes simplex virus (HSV) produced immunoglobulin G antibody-dependent cellular cytotoxicity (ADCC) and radioimmunoassay (RIA) antibody as early as 3 days after infection. There was a rise in natural killer cytotoxicity (NKC) to infected and uninfected target cells 1 to 3 days postinfection mediated by nonadherent peritoneal cells (PC) in mice inoculated with HSV, but also with other substances commonly used in tissue culture media. HSV caused the highest and most consistent increase in NKC. PC-NKC, as ADCC, was inhibited by latex and silica, both macrophage inhibitors. PC-ADCC markedly declined 3 to 8 days after HSV inoculation. This was not due to a soluble or cellular suppressor factor, was not reversed by incubation or trypsin treatment of PC, was not associated with a change in PC Fc receptors, adherence, or acridine orange staining characteristics, and could not be induced by inactivated HSV. In vitro inoculation of PC with HSV similarly caused a reduction in the ability of PC to mediate ADCC to HSV-infected target cells. These data demonstrate the complex stimulatory and inhibitory interactions between virus and host defense mechanisms.
Mutanolysin-induced spheroplasts of Streptococcus mutants are true protoplasts
Authors: Siegel J L , Hurst S F , Liberman E S , Coleman S E , Bleiweis A S
Published: January 2020

A method is described for the preparation of protoplasts of Streptococcus mutans BHT. The muralytic enzyme mutanolysin was prepared free of contaminating proteinases and shown to completely dissolve cell walls of this strain. Whole cells were converted to stabilizable protoplasts by using the enzyme in an isotonic medium containing 40% raffinose. Experiments using [3H]thymidine and [14C]leucine as cytoplasmic pool markers revealed only minimal (10%) leakage during a 1-h incubation. Examination by electron microscopy revealed the apparent absence of structural cell wall on the enlarged spherical bodies. Quantitative chemical analyses of membranes prepared by lysing protoplasts demonstrated only very small amounts of rhamnose and trace amounts of galactose. These sugars are the principal components of the BHT cell wall polysaccharide. Also, there were only small amounts of peptidoglycan components (e.g., N-acetylglucosamine) in the purified membranes obtained by this method.
Synergistic effects of lipopolysaccharide on phytohemagglutinin- and concanavalin A-induced deoxyribonucleic acid synthesis in human peripheral blood lymphocytes: participation of T lymphocytes
Authors: Hatfield S M , Schmidtke J R
Published: January 2020

Lipopolysaccharide induces a synergistic uptake of tritiated thymidine in peripheral blood lymphocytes (PBL) when cultured with phytohemagglutinin or concanavalin A as compared to PBL incubated only with phytohemagglutinin or concanavalin A. In this study we investigated which subpopulations(s) of PBL is involved in this synergistic increase in deoxyribonucleic acid synthesis. Separation of PBL into sheep erythrocyte rosette-forming cells (T cells) and non-rosette-forming cells (B cells) showed that the T cells were responsible for the increased uptake of radiolabel. PBL and T cells had similar dose-response profiles and kinetic patterns. Paralleling this augmented tritiated thymidine uptake was an increase in the number of cells undergoing blast transformation. Delayed-addition experiments showed that the two mitogens must be added within 12 h of each other for maximal augmentation to occur. Finally, preincubation of T cells with lipopolysaccharide had no demonstrable effect on the amount of concanavalin A uptake by these cells. This model may provide unique information about the activation of human peripheral blood T cells compared to activation of these cells by one mitogen.
Characterization of the Streptococcus mutans plasmid pva318 cloned into Escherichia coli
Authors: Hansen J B , Abiko Y , Curtiss R
Published: January 2020

We further characterized the cryptic plasmid pVA318 of Streptococcus mutans. It had a contour length of 5.64 +/- 0.26 kilobases and a guanine-plus-cytosine content of 32 to 34 mol %. Upon cloning the pVA318 plasmid into the vector pBR322 in Escherichia coli, we made the following observations. The expression of tetracycline resistance by HindIII-cloned chimeras, where the insert was in the tetracycline resistance promotor, depended on the orientation of the pVA318 insert. Both HindIII-cloned chimeras segregated from polA(Ts) cells at a nonpermissive temperature. Chimeric molecules cloned with PstI initially showed much instability; the reason for this is unknown, although stable variants were obtained. Both HindIII-cloned variants and a PstI-cloned chimera produced a pVA318-specific protein of approximately 20,000 molecular weight in E. coli minicells. The biological function of this protein is not known; it had no bacteriocin activity against S. mutans or group A Streptococcus indicator strains, and it did not appear in the E. coli periplasm. We constructed a map of pVA318 for restriction endonucleases HindIII, HpaI, PstI, and HaeIII. A previously reported BamHI site in pVA318 did not appear in the pVA318 portion of any of our chimeric clones.
Serological and biological activities of anti-Haemophilus influenzae ribosomal serum
Authors: Katz M , Lynn M , Solotorovsky M
Published: January 2020

The antibody content in serum from rabbits immunized with ribosomes from Haemophilus influenzae type b was determined by passive hemagglutination, enzyme-linked immunosorbent assay, and complement fixation. Attempts to use passive hemagglutination to assay anti-ribosomal antibodies were unsuccessful. In the enzyme-linked immunosorbent assay tests, rabbit antiserum was allowed to react with ribosomes that adhered to microtiter plates. The enzyme-linked immunosorbent assay method detected, in two ribosome-immunized rabbits and by 3 days postimmunization, titers which rose to plateaus on days 24 to 31 and declined thereafter. With the complement fixation method, the serum from one immunized rabbit also showed a titer on day 3 and reached a plateau on days 20 to 31. Serum from the other immunized rabbit did not develop a titer until day 11; it reached a lower plateau on days 20 to 24 and then declined on days 27 to 55. Although there were no apparent differences between the two immunized rabbits by the enzyme-linked immunosorbent assay, there were differences between the complement fixation antibodies in these rabbits. Passive protection experiments were performed with these sera. Maximal passive protection was achieved when mice were challenged intraperitoneally with 100 50% lethal doses of H. influenzae and immunized intravenously 1 h later with rabbit serum collected 27 days postimmunization. Rabbit anti-ribosomal sera were evaluated for bactericidal activity. Undiluted immune sera showed bactericidal activity; however, when diluted 1:10, activity was lost. Although bactericidal activities of immune sera were correlated to passive protection activities, it is unlikely that such protection was due to bactericidal antibodies. Immune serum had opsonizing activity since it enhanced phagocytosis of H. influenzae by mouse leukocytes.
Analysis of human anti-diphtheria antibodies by isoelectric focusing: evidence for restricted clonal heterogeneity of anti-fragment A antibodies
Authors: Morrow C D , Macy E M , Stevens R H
Published: January 2020

The in vivo human humoral response to diphtheria toxoid-tetanus toxoid booster immunization was studied by isoelectric focusing analysis of sera obtained after immunization. The anti-diphtheria toxoid (immunoglobulin G [IgG]-Dip), anti-fragment A (IgG-Frag A), and anti-tetanus toxoid antibodies from 20 donors post-booster immunization were focused by using agarose isoelectric focusing and visualized by development with radiolabeled antigens. The quantities of the IgG-Dip and IgG-Frag A antibodies correlated with the number of bands seen on the isoelectric focusing pattern in that more bands were found in the spectrotypes of donors with high serum levels of antibody. No difference was apparent in the antibody spectrotypes obtained from sera of donors at successive times post-booster immunization. Individual heterogeneity of the different donors' spectrotypes was often found for IgG-Frag A antibodies, but a close comparison of several different donors revealed antibodies with the same spectrotype patterns. Thus, individual clones of antibody were revealed in humans after in vivo immunization, particularly when antibodies against antigens of restricted epitope size were analyzed. Additionally, the sharing of certain antibody spectrotypes among several individuals raised the possibility that certain antibody clones may be preferentially expressed in the human population.
Differential requirements for enriched atmospheric carbon dioxide content for intracellular growth in cell culture among selected members of the genus Rickettsia
Authors: Kopmans-Gargantiel A I , Wisseman C L
Published: January 2020

In an in vitro chicken embryo cell culture system, strains of Rickettsia prowazekii, R. mooseri, and R. rickettsii, but not of R. tsutsugamushi, required an atmosphere enriched in CO2 for intracellular growth.
Antibody to virus components in volunteers experimentally infected with human coronavirus 229E group viruses
Authors: Macnaughton M R , Hasony H J , Madge M H , Reed S E
Published: January 2020

Antibody rises to various virus subcomponents were measured by enzyme-linked immunosorbent assay in the paired sera of volunteers experimentally infected with human coronavirus 229E group viruses. Most of the antibody made during infection was directed against the virus surface projections, with only small amounts of antibody made against membrane or ribonucleoprotein components.
Mechanism of immune interferon production in vitro: interaction between immune interferon-producing cells and antigenic cells
Authors: Ito Y , Aoki H , Kimura Y , Shimokata K , Maeno K
Published: January 2020

Various processes of in vitro immune interferon production by sensitized spleen cells stimulated with allogeneic cells were investigated. When L cells, an interferon-inducing antigen, were fixed with methyl alcohol or paraformaldehyde, the ability to induce immune interferon disappeared. In this immune interferon production system, the majority of sensitized spleen cells adhered to target cells within 1 h of cocultivation. Adherence of immune interferon-producing cells to target cells was observed only when L cell-sensitized spleen cells were cocultured with L cells or with mouse embryo cells derived from C3H mice. Fixation of antigenic cells with methyl alcohol or paraformaldehyde significantly reduced cell adherence. When L cells alone or sensitized spleen cells alone were pretreated separately with cytochalasin D, neither cell type could bind to partner cells. Specific adherence did not take place at 4 degrees C, nor in the presence of dinitrophenol or sodium azide. Continuous protein synthesis in both cells was not required for immune cell adherence. Divalent cations, Ca2+ or Mg2+, were required for this immune specific adherence to take place. However, once stable adherence was established, treatment with cytochalasin D, ethylenediaminetetraacetic acid, or sodium azide, or simple reduction of temperature, did not disrupt the binding. Interaction between immune interferon-producing cells and antigenic cells can be subdivided into two phases according to the requirement for divalent cations: (i) lymphocytes and antigenic cells interact transiently, and divalent cations are required to maintain the binding; (ii) lymphocytes and antigenic cells form a stable interaction, and deprivation of divalent cations does not disrupt the binding. Colchicine showed an inhibitory effect in the period after cell-to-cell adherence. Colchicine did not inhibit the release of interferon. On the other hand, vinblastine, another antimicrotubule agent inhibited the secretion of immune interferon. Since interferon synthesis was not stopped immediately after addition of cycloheximide, continued protein synthesis of sensitized spleen cells was not required for interferon secretion. The present study showed that adherence of immune interferon-producing cells to antigenic cells was a complex phenomenon involving a series of successive events.
Staphylococcal enterotoxins fail to disrupt membrane integrity or synthetic functions of Henle 407 intestinal cells
Authors: Buxser S , Bonventre P F
Published: January 2020

The potential cytotoxic activity of purified staphylococcal enterotoxins for mammalian cells was evaluated. The effects of staphylococcal enterotoxins A (SEA) and B (SEB) on cell membrane integrity as measured by leakage of labeled cytoplasmic constituents ([3H]uridine), amino acid transport (lysine and aminoisobutyric acid), and macromolecular synthesis (protein, ribonucleic acid, and deoxyribonucleic acid) was evaluated for a human intestinal epithelial cell (Henle 407). No evidence of cytotoxicity by any of these criteria could be detected for cell monolayers incubated with SEA for periods of between 30 min and 24 h. Purified staphylococcal hemolysins (alpha- and delta-toxins) were shown to exert cytotoxicity by the leakage and amino acid uptake assays. In efforts to detect synergistic effects between enterotoxin and the staphylococcal cytotoxins, membrane functions were evaluated after sequential or combined treatment with enterotoxin and alpha-toxin or with enterotoxin and delta-toxin. In no instance could a contribution to cytotoxicity by the staphylococcal enterotoxin be detected. That the assays were sufficiently sensitive to detect synergistic effects was shown by the greater than additive effects achieved with a combination of alpha- and delta-toxins. The data, contrary to previous reports, showed that staphylococcal enterotoxins did not behave as bacterial cytotoxins.
Activation of human lymphocytes in vitro by membrane-damaging toxins from Staphylococcus aureus
Authors: Petrini B , Möllby R
Published: January 2020

Purified staphylococcal alpha-, gamma-, and delta-toxin were shown to cause activation of lymphoid cells from adult human donors and of cord cells in vitro as measured by [14C]thymidine incorporation after 7 days of incubation. T cell-enriched and T cell-depleted lymphocyte suspensions were activated in a similar fashion. Beta-toxin, on the other hand, exerted no valid stimulation of the various lymphocyte preparations. The lymphocyte-activating properties of alpha- and gamma-toxin were shown to be independent of their hemolytic capacity. The results probably reflect unspecific mitogen effects, but a component of specific reactivity cannot be excluded. We suggest that the unspecific triggering of lymphocytes in vitro is caused by surface-active properties of the toxins.
Evidence from a carbohydrate incorporation assay for direct activation of bone marrow myelopoietic precursor cells by bacterial cell wall constitutents
Authors: Monner D A , Gmeiner J , Mühlradt P F
Published: January 2020

The stimulation of incorporation of [3H]galactose into membrane glycoconjugates, measured in a precipitation test, was used as a criterion for activation of bone marrow cells. In this assay, purified bacterial lipopolysaccharide, lipoprotein, and murein monomer and dimer fragments all activated rat bone marrow cells in vitro. The response was dose dependent, followed a defined time course, and was not serum dependent. O-Acetylated murein dimer fragments from Proteus mirabilis were much less active than their unsubstituted counterparts, indicating a structural specificity for murein activation. Removal of adherent and phagocytizing cells from the marrow suspensions did not alter these results. The labeled, activated cells constituted a distinct population of buoyant density 1.064 to 1.069 g/cm3 when centrifuged on a continuous gradient of Percoll. Enrichment of the target cell population was achieved by a combination of adherent cell removal and discontinuous density gradient centrifugation to remove granulocytes and erythropoietic cells. It was concluded that a population of myelopoietic precursors could be activated by direct contact with bacterial cell wall constituents. The stimulation of galactose incorporation was not coupled to active deoxyribonucleic acid synthesis in the marrow cells. Thus, the activation was interpreted as an induction of differentiation rather than a mitotic event.
Alveolar bone loss in rats after immunization with Actinomyces viscosus
Authors: Burckhardt J J , Gaegauf-Zollinger R , Schmid R , Guggenheim B
Published: January 2020

We investigated a possible cause-and-effect relationship between sensitization against Actinomyces viscosus Nyl and destructive periodontal disease in RIC-Sprague-Dawley rats. Germfree rats (66) were either immunized with A. viscosus Nyl (day 20) or orally infected with A. viscosus Nyl (days 38 and 39) or both. We measured alveolar bone loss in maxillary and mandibular molars, in vitro T-lymphocyte responsiveness, and serum antibody titers. In immunized and monoassociated rats bone loss in both jaws progressed rapidly between days 37 and 72, whereas the rate of further resorption decreased until day 100. In monoassociated rats, development of bone loss was much slower, and the maximum resorption measured was, at best, half of the bone loss compared with the former group. However, no amplification of bone loss by immunization was observed in a second experiment using 63 conventional rats kept in relative gnotobiosis. Antibody titers to A. viscosus Nyl in gnotobiotic monoassociated rats were higher in immunized animals, whereas no difference was found in the respective groups of the relative gnotobiotic experiment. The fact that immunization more than doubled alveolar bone loss in gnotobiotic monoassociated rats confirms the allergic nature of the disease. The lack of such an effect under conventional conditions points to suppressor mechanisms whose decrease might convert stable periodontal lesions into progressive ones.
Characterization of hemadsorption-negative mutants of Mycoplasma pneumoniae
Authors: Hansen E J , Wilson R M , Clyde W A , Baseman J B
Published: January 2020

Previously isolated mutants of Mycoplasma pneumoniae incapable of hemadsorption were characterized with respect to specific protein content, tracheal ring attachment capability, and virulence for both in vitro and in vivo model systems. Two-dimensional gel electrophoresis revealed both quantitative and qualitative differences between the protein complements of two different mutant strains and that of the virulent parent strain. Studies of mycoplasma attachment to hamster tracheal rings in vitro demonstrated that only one of these mutant strains still possessed the ability to attach to the respiratory epithelium via neuraminidase-sensitive receptors. Measurement of [3H]orotic acid uptake in mycoplasma-infected tracheal rings indicated that infection with the hemadsorption-negative mutants resulted in only slight reductions of ribonucleic acid synthesis, similar to levels observed for tracheal rings infected with an avirulent strain of M. pneumoniae. The virulence potential of the two mutant strains was further investigated by utilizing the hamster model system. Both mutant strains were rapidly cleared from the lungs of infected animals and produced little or no microscopic pneumonia.