bluuebunny/crossref_metadata_2025 huggingface dataset:
Isolation of a cytopathic factor from Mycoplasma hyopneumoniae
Authors: Geary S J , Walczak E M
Published: January 2020

A cytopathic factor was isolated from M. hyopneumoniae VPP11 membranes. This factor had an isoelectric point of 6.2 and was capable of inducing cytopathic effect in MRC-5 human lung fibroblast culture at a concentration of 250 ng of protein per ml. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of this cytopathic factor showed a major band with a molecular weight of approximately 54,000.
Protection against Escherichia coli-induced urinary tract infections with hybridoma antibodies directed against type 1 fimbriae or complementary D-mannose receptors
Authors: Abraham S N , Babu J P , Giampapa C S , Hasty D L , Simpson W A , Beachey E H
Published: January 2020

Hybridoma antibodies directed against quaternary structural epitopes of the type 1 fimbrial adhesin of Escherichia coli or against D-mannose, the sugar determinant in the complementary host cell receptor, prevented the attachment of mannose-sensitive E. coli to various eucaryotic cells. Passive intraperitoneal administration of the fimbria-specific or D-mannose-specific antibodies protected mice against retrograde colonization with mannose-sensitive E. coli instilled into their urinary bladders. Monoclonal antibodies directed against fimbrial subunits rather than quaternary structural epitopes or against N-acetylgalactosamine rather than D-mannose residues lacked protective activity. These studies provide evidence that bacterial colonization can be blocked or interrupted by antibodies directed against either the adhesin or the complementary host cell receptor of pathogenic microorganisms.
Pretreatment with lipoteichoic acid sensitizes target cells to antibody-dependent cellular cytotoxicity in the presence of anti-lipoteichoic acid antibodies
Authors: Lopatin D E , Kessler R E
Published: January 2020

This study was performed to determine whether antibody-dependent cellular cytotoxicity (ADCC) could be directed against mammalian cells sensitized with spontaneously adhering bacterial substances. 51Cr-labeled SB leukemia cells were incubated with purified S43 group A streptococcal lipoteichoic acid (LTA; 0.001 to 100 micrograms/ml). Purified leukocyte ADCC effector cells were added to the LTA-coated target cells at various effector-to-target ratios (100:1 to 12:1), followed by the addition of rabbit anti-LTA. After incubation for 4 h, target cell lysis was calculated based on the release of label into the medium. As little as 1 ng of LTA per ml was sufficient to sensitize the target cells to ADCC lysis (12%); however, concentrations above 0.1 micrograms/ml generally resulted in 60 to 80% lysis. LTA alone was not cytotoxic to these target cells. Targeting did not occur if effector cells were sensitized or if free LTA was added to the medium. Specificity was demonstrated by cold-target inhibition, which showed that anti-LTA cytotoxicity could be inhibited only by unlabeled, LTA-treated target cells but not by cold SB cells alone. The findings indicate that certain soluble bacterial components, when bound to mammalian cells in the presence of specific antibody, can target ADCC effectors to these cells. This mechanism may be an important factor in the delayed sequelae of bacterial infections.
A serratial protease causes vascular permeability reaction by activation of the Hageman factor-dependent pathway in guinea pigs
Authors: Kamata R , Yamamoto T , Matsumoto K , Maeda H
Published: January 2020

The 56-kilodalton (56K) protease isolated from a culture filtrate of Serratia marcescens caused vascular permeability enhancement followed by edema formation when injected into guinea pig peripheral corneas and subconjunctival space or skin. The character and the mechanism of permeability enhancement were analyzed in vivo. The enhancement was maximum at 5 to 10 min. The permeability reaction increased exponentially by the amount of enzyme used. The enhancement of permeability induced by the 56K protease was not affected by treatment with an antihistamine but was greatly augmented by simultaneous injection of a kinin potentiator, Glu-Trp-Pro-Arg-Pro-Gln-Ile-Pro-Pro-OH (SQ 20,881). Furthermore, the permeability activity of the protease, but not the amidolytic activity, was inhibited by soybean trypsin inhibitor, a well-known inhibitor of plasma kallikrein, as well as by corn trypsin inhibitor, the best inhibitor of activated Hageman factor. Results of these in vivo studies indicate that the permeability-enhancing reaction induced by the 56K protease is caused by activation of the Hageman factor-dependent pathway in the tissue. The permeability-increasing activity of the 56K protease was parallel with the enzyme activity. Serratial lipopolysaccharide did not produce a permeability enhancement reaction within 30 min when injected into guinea pig skin. These results are consistent with the results of recent in vitro experiments in which activation of the purified Hageman factor but not of prekallikrein by the 56K protease was elucidated (Matsumoto et al., J. Biochem. (Tokyo) 96:739-749, 1984). Thus, the molecular mechanism described above appears to be operative in the pathogenesis of corneal edema and chemosis, which is induced by S. marcescens, in addition to the direct tissue destruction by the protease.
Effects of natural or synthetic microbial adjuvants on induction of autoimmune thyroiditis
Authors: Kong Y C , Audibert F , Giraldo A A , Rose N R , Chedid L
Published: January 2020

Natural and synthetic adjuvants of microbial origin were compared for their capacity to potentiate the induction of experimental autoimmune thyroiditis (EAT) with the autoantigen mouse thyroglobulin (MTg). Regardless of the immunomodulator used, severe thyroiditis was observed only in EAT-susceptible strains of the k haplotype and not in EAT-resistant strains of the d haplotype. Compared to phenol-extracted lipopolysaccharide, a potent adjuvant for enhancing EAT induction, phthalyl-substituted, detoxified lipopolysaccharide, even at doses 15- to 50-fold greater, led to only low anti-mouse thyroglobulin titers and mild thyroid infiltration. The synthetic adjuvant N-acetylmuramyl-L-alanyl-D-isoglutamine (MDP) and three of its analogs, N-acetylmuramyl-L-alanyl-D-isoglutamine-L-alanyl-D-glycerol mycolate (MDP-L-Ala-Glyc-Myc), N-acetylmuramyl-L-alanyl-D-glutamyl-(decyl)methyl ester [MDP(decyl)methyl], and N-acetylmuramyl-L-alanyl-D-glutamine-alpha n-butyl ester [MDP-(Gln)-OnBu], designated murabutide, were tested in incomplete Freund adjuvant or in saline. In incomplete Freund adjuvant, MDP-L-Ala-Glyc-Myc was inefficient in inducing EAT, murabutide induced very mild involvement, and MDP and, more so, MDP(decyl)methyl were active but to a lesser degree than CFA. When saline was used, low levels of thyroid infiltration were observed in a few of the MDP-treated animals in only one experiment, whereas no lesions were observed when murabutide was used.
Immunochemical characterization of a protein associated with Mycobacterium leprae cell wall
Authors: Gillis T P , Miller R A , Young D B , Khanolkar S R , Buchanan T M
Published: January 2020

A panel of nine monoclonal antibodies to Mycobacterium leprae were used to characterize a protein antigen of the bacillus. Two monoclonal antibodies (IVD8 and IIIE9) were specific for M. leprae and reacted with an epitope (CWPa) present on a protein molecule associated with the cell wall fraction of M. leprae. This protein, designated cell wall-associated protein (CWP), lost its immunoreactivity upon treatment with trypsin and had an apparent molecular weight of 65,000, though additional lower-molecular-weight forms of the protein were observed by immunoblotting. Four other cross-reactive epitopes (CWPb, CWPc, CWPd, and CWPe) were defined on the same molecule using seven independent monoclonal antibodies. Therefore, M. leprae possesses a trypsin-sensitive, heat-stable protein associated with the cell wall which contains at least one species-specific and four cross-reactive antigenic determinants.
Oxidative metabolic response and microbicidal activity of human milk macrophages: effect of lipopolysaccharide and muramyl dipeptide
Authors: Cummings N P , Neifert M R , Pabst M J , Johnston R B
Published: January 2020

Mouse macrophages can be primed by exposure in vitro to the bacterial products lipopolysaccharide and muramyl dipeptide (MDP) or in vivo by injection of MDP, so that they produce more of the bactericidal agent superoxide anion (O2-) when stimulated by phagocytosis or by contact with phorbol myristate acetate (PMA). Because little is known about the physiology of human tissue macrophages, we examined release of O2- by milk macrophages obtained from 45 normal women for the ability to undergo priming for greater O2- release. In samples from the same individuals, PMA-stimulated O2- release was similar from colostrum (0 to 3 days postpartum) or from transitional milk (5 to 8 days). Release of O2- by milk macrophages was almost identical to that by blood monocytes from the same women. Milk macrophages phagocytized and killed Candida albicans relatively effectively. Incubation with lipopolysaccharide activated the macrophages in that they were primed for greater PMA-stimulated O2- release. Incubation with the adjuvant MDP or its analog 6-O-(2-tetradecylhexadecanoyl)-MDP did not prime, but incubation with a second analog, 6-O-(stearoyl)-MDP, primed the macrophage for greater O2- release. These results indicated that human tissue macrophages can be primed for greater oxidative response by exposure to bacterial products. Potential exists for the therapeutic use of such immunomodulating agents in the enhancement of host defense.
Killed Escherichia coli stimulates macrophage-mediated alterations in hepatocellular function during in vitro coculture: a mechanism of altered liver function in sepsis
Authors: West M A , Keller G A , Cerra F B , Simmons R L
Published: January 2020

Hepatic dysfunction is a poorly understood and highly lethal component of multiple-system organ failure. Both in vivo and in vitro studies of "liver" function have generally neglected hepatocyte-Kupffer cell interactions. In the following experiments, isolated hepatocytes were cocultivated with unstimulated peritoneal cells, predominately macrophages, which served as a readily available Kupffer cell analog. Coculture of hepatocytes with peritoneal cells resulted in little or no change in [3H]leucine incorporation into hepatocyte protein. When gentamicin-killed Escherichia coli cells (GKEC) were added to coculture, there was a marked decrease in hepatocyte [3H]leucine incorporation. In contrast, GKEC added to hepatocytes alone had no effect. Kinetic data revealed an 8-h delay before any significant decrease in leucine incorporation into hepatocyte protein after the addition of GKEC to the coculture. The maximal decrease in hepatocyte [3H]leucine incorporation occurred 24 h after GKEC were added. The decrease observed 24 h after GKEC were added disappeared almost completely after 48 h of coculture. Similar alterations in cocultured hepatocyte protein synthesis were observed after the addition of phorbol myristate acetate, lipopolysaccharide, or muramyl dipeptide, a component of bacterial peptidoglycan. Hepatocyte viability by trypan blue exclusion was unchanged, and gross morphology by light or electron microscopy was unaffected. We propose that during sepsis, macrophages (Kupffer cells) respond to circulating microbial products and mediate alterations in hepatocyte function. These experiments underscore the important role of Kupffer cell function in attempts to understand hepatic malfunction in multiple-system organ failure.
Streptococcus pneumoniae cocultured with fibroblasts enhances both interferon production and cytotoxic activity by lymphocytes
Authors: Weigent D A , Baron S , Stanton G J
Published: January 2020

Cell-mediated cytotoxicity against normal human fibroblasts was dependent on treatment of the fibroblasts with Streptococcus pneumoniae. Both spontaneous and interferon (IFN)-enhanced lymphocytes killed human foreskin (HFS) or skin muscle cells cocultured with S. pneumoniae five- to eightfold more than control nontreated cells. Based on Percoll gradient centrifugation, the cytotoxic effector cell migrated like a large granular lymphocyte. The human IFN produced from mixtures of HFS cells, lymphocytes, and S. pneumoniae was observed to be both a mixture of IFN-alpha and IFN-gamma and in an amount 500 times greater than that observed with lymphocytes on HFS cells alone, and it was in an amount 12 times greater than when lymphocytes and bacteria were cultured together. A mixture of antibodies to IFN-alpha and -gamma added to cocultures of fibroblasts and bacteria blocked the killing of fibroblast targets by lymphocytes (47 versus 13%). Thus, endogenously produced IFN was essential for the effective killing of the fibroblasts. Treatment of HFS cells with IFN before bacterial treatment protected the HFS cells from lysis by lymphocytes. The observation that normal diploid cells exposed to bacteria can be killed by lymphocytes suggests that natural cytotoxic cells are active at the site of bacterial infection and conceivably play roles in defense or pathogenesis.
Enzymatic release of germ tube-specific antigens from cell walls of Candida albicans
Authors: Sundstrom P M , Kenny G E
Published: January 2020

The differences between blastospores and germ tubes of Candida albicans, as previously shown by immunofluorescence, were further studied by comparing digests of cell walls of both growth forms. Organisms were surface labeled with 125I, and cell walls were digested enzymatically. When Zymolase digests were treated with polyclonal, polyspecific antiserum to C. albicans 441B, which stains only germ tubes in immunofluorescence assays, components of molecular weights 200,000 and 155,000 were immunoprecipitated from digests of germ tubes of strain B311, but nothing was recovered from blastospores. Whereas the 200,000-molecular-weight component was found in the three strains tested, the 155,000-molecular-weight component was found only in strain B311. When Zymolase digests were treated with unadsorbed antiserum, which stains both blastospores and germ tubes in immunofluorescence assays, an additional component was precipitated from digests of both growth forms with a molecular weight greater than the 200,000-molecular-weight marker. All three antigens were mannoproteins, as was shown by their abilities to bind concanavalin A and to be labeled by 125I. Also, all antigens were located on the cell surface, as was shown by the following criteria: adsorption of antisera with live organisms removed antibody to these components, and antibody eluted from surfaces of whole organisms precipitated all components. Components common to both growth forms, as well as germ tube-specific components, were detected in trypsin and chymotrypsin digests, but their molecular weights differed from those of Zymolase digests. Thus, germ tube-specific surface determinants as well as determinants common to both growth forms were detected on enzymatically released cell wall components.
Vibrio vulnificus resists phagocytosis in the absence of serum opsonins
Authors: Tamplin M L , Specter S , Rodrick G E , Friedman H
Published: January 2020

Invasive disease caused by Vibrio vulnificus may result partially from resistance to phagocytic host defense mechanisms. The present studies show that V. vulnificus resists phagocytosis by murine peritoneal macrophages in the absence of serum opsonins and extracellular bacterial products, apparently through the anti-phagocytic properties of the bacterial surface.
Increased local complement levels upon intraperitoneal injection of mice with Listeria monocytogenes and regulation by polyanions
Authors: Klerx J P , Beukelman C J , van Dijk H , van Overveld F J , van der Maaden W J , Willers J M
Published: January 2020

Alternative complement pathway activity in mouse peritoneal fluid was determined by a sensitive microtiter assay with rabbit erythrocytes as target cells and cobra venom factor as the inducer of bystander hemolysis. The intra- and interstrain variations in five mouse strains were 30% and, maximally, 64%, respectively. No activity was detected in genetically C5-deficient mice. In C5-sufficient mice, intraperitoneal injection of heat-inactivated Listeria monocytogenes resulted in a 250% increase in local complement activity within 30 min. This effect was reversed by simultaneous injection of dextran sulfate but not heparin. These results are discussed in relation to the ability of listeriae to activate the mouse alternative complement pathway in vitro and the effectiveness of dextran sulfate and the failure of heparin in functioning as an adjuvant in the induction of a protective response in mice by a heat-inactivated listeria vaccine.
Changes in the Mouse Intestinal Microflora During Weaning: Role of Volatile Fatty Acids
Authors: Lee Adrian , Gemmell Erica
Published: January 2020

The influence of volatile fatty acids on the ecology of the bacterial flora of the mouse intestinal tract has been studied in three situations where large fluctuations in the composition of the microflora have been observed. Young mice were shown to ingest solid food particles when 11 days old; this correlated with the appearance of strictly anaerobic fusiform bacilli in the intestinal lumen and a 10,000-fold decrease in numbers of coliform bacilli. Over the same period, volatile fatty acids were shown by gas-liquid chromatography to appear in the intestinal content. It is suggested that the fusiform bacilli are responsible for the presence of the volatile acids (especially butyric acid) which exert an inhibitory effect on the coliform bacteria, resulting in the decline in numbers. When germ-free mice are placed in a specific pathogen-free mouse colony, changes in the intestinal flora occurred which were similar to those observed in the young mice approaching weaning. Once again, the decline in the coliform population correlated with the appearance of significant levels of butyric acid in the large intestine. In a further series of experiments, mice were fed penicillin and levels of the intestinal fatty acids were measured. The antibiotic eliminated the anaerobic fusiforms from the intestine, resulting in the disappearance of significant levels of butyric acid and a million-fold increase in the numbers of coliform bacilli.
Binucleate Classical Mycoplasmas Pathogenic for Goats
Authors: Furness Geoffrey , De Maggio Maria
Published: January 2020

The growth of three pathogenic goat mycoplasmas, strains Y, KH1 and
Mycoplasma mycoides
var.
capri
(PG3), was studied. They formed classical colonies on agar containing 1/500 thallium acetate. They were inactivated during storage at 2 to 4 C and by freezing and thawing but not by shaking. Only KH1 was killed by sonic treatment. Ultraviolet inactivation curves showed that their colony-forming units were single binucleate cells. Details of their growth curves are given. Filtration through 0.45- or 0.3-μm membrane filters removed up to 97% of the cells. Less than 0.003% passed 0.22-μm membranes. In electron micrographs, the cells were seen replicating by budding and most were 0.6 to 0.9 μm in diameter; but cells between 0.1 and 0.2 μm reproduced. They usually multiplied by producing one bud, a form of binary fission. However, two buds were produced by some synchronized cells, indicating that both nuclei had divided simultaneously to form progeny, an alternate method of multiplication.
Contribution of Humoral and Cellular Factors to the Resistance to Experimental Infection by
Pseudomonas aeruginosa
in Mice II. Opsonic, Agglutinative, and Protective Capacities of Immunoglobulin G Anti-
Pseudomonas
Antibodies
Authors: Bjornson Ann B. , Michael J. Gabriel
Published: January 2020

The opsonic, agglutinative, and mouse protective capacities of anti-
Pseudomonas
antibodies in immune and normal human immunoglobulin G (IgG) preparations were investigated. Opsonic activities of the immune IgG preparations correlated well with their protective activities. Antibodies present in normal IgG showed a substantial agglutinative activity but were poorly protective. Anti-
Pseudomonas
antibodies of both normal and immune IgG preparations were directed against the same serotype antigen as demonstrated by absorption experiments. Immune anti-
Pseudomonas
IgG antibodies fixed complement very efficiently as demonstrated by opsonophagocytic and hemolytic tests. Natural anti-
Pseudomonas
IgG antibodies fixed complement very poorly although they promoted phagocytic killing of bacteria only in the presence of heat-labile serum factors. It was concluded that, although agglutination can be used for qualitative measurement of antibacterial antibodies, it fails to measure their functional capacities.
Alteration in Antibody Response Induced by Carbon Inhalation: a Model System
Authors: Zarkower A. , Morges W.
Published: January 2020

The immune response in mice to antigens given by intraperitoneal, intravenous, and inhalation routes was compared in control and carbon-dust exposed mice. The most pronounced effect, a decrease in antibody-forming cells and serum agglutinin titers, was observed when the carbon-treated animals were immunized with antigens in the form of aerosol. Similar results were obtained when the antigens were administered by intravenous or intraperitoneal route. In contrast 4-days of pre-exposure to carbon dust increased the number of antibody-forming cells in the mediastinal lymph nodes. However, this effect was transient and was not significant when the carbon pre-exposure was continued for 15 days.
Ribosomal Vaccines I. Immunogenicity of Ribosomal Fractions Isolated from
Salmonella typhimurium
and
Yersinia pestis
Authors: Johnson William
Published: January 2020

The immunogenicity of ribosomes and ribosomal subfractions isolated from
Yersina pestis
and
Salmonella typhimurium
has been studied. Ribosomes and ribosomal protein isolated from
S. typhimurium
protected mice against lethal challenge. Ribosomal ribonucleic acid isolated by phenol extraction failed to induce any significant level of protection in mice. None of the ribosomes or ribosomal subfractions isolated from
Y. pestis
were effective in inducing immunity to lethal challenge. These results suggest that the immunogen of the ribosomal vaccine is protein.
Anti-Teichoic Acid Antibodies and Non-treponemal Serological Tests for Syphilis
Authors: Wicken A. J. , Gibbens Janet W. , Knox K. W.
Published: January 2020

Antibodies specific for the polyglycerophosphate “backbone” of glycerol teichoic acids act as reagin in the Kolmer complement fixation test for syphilis.
An early effect of the S component of staphylococcal leukocidin on methylation of phospholipid in various leukocytes
Authors: Noda M , Hirayama T , Matsuda F , Kato I
Published: January 2020

On incubation of rabbit polymorphonuclear leukocytes with the S component of staphylococcal leukocidin at 37 degrees C, the 3H-labeled methyl group of S-adenosyl[methyl-3H]methionine was rapidly incorporated into phospholipid. Subsequently, the methylated phosphatidylcholine was degraded by activated phospholipase A2. Complete blockage of the methylation of phospholipid by a mixture of erythro-9-[2-hydroxy-3-nonyl]adenine, adenosine, and L-homocysteine thiolactone markedly inhibited the activation of phospholipase A2 by the S component. It also inhibited the binding of 125I-labeled F component to the cells, but not that of the labeled S component. These results suggest that methylation of phospholipid in the cell membranes by the S component results in activation of phospholipase A2, which induces the binding of the F component to the cells.
Depressed chemiluminescence response by influenza virus is enhanced after conjugation of viral subunits to muramyl dipeptide
Authors: Masihi K N , Lange W , Rohde-Schulz B , Chedid L , Jolivet M
Published: January 2020

The effect on respiratory burst of murine spleen cells after in vitro exposure to influenza virus, subunits, or subunits conjugated to muramyl dipeptide (MDP) was studied by luminol-dependent chemiluminescence (CL) in response to stimulation by zymosan. CL induced by infectious influenza A virus was depressed but could be elevated to normal levels when MDP was added together with a low, but not with a high, dose of the virus. Profound depression of CL was induced by high doses of influenza A/Brazil, A/Bangkok, and B/Singapore subunits. The same amounts of viral subunits conjugated to MDP restored or even enhanced the CL responses of spleen cells from BALB/c and C57BL/6 mice. Splenic cells from BALB/c mice generated higher levels of CL than did cells from C57BL/6 mice.
Colonization of the mouse intestine by an avirulent Salmonella typhimurium strain and its lipopolysaccharide-defective mutants
Authors: Nevola J J , Stocker B A , Laux D C , Cohen P S
Published: January 2020

For study of the role of lipopolysaccharide (LPS) character in colonization of the mouse large intestine, use was made of S. typhimurium strain SL5316, which is streptomycin resistant and smooth (wild-type LPS) but nonvirulent because it is Aro- (aromatic dependent). Several rough variants of different LPS chemotype derived from strain SL5316 comprised: an rfb deletion transductant making type Ra (complete core) LPS; an rfaJ mutant making incomplete core of type Rb2; and an rfa-990 mutant making LPS core less complete than chemotype Rb2. We tested these strains for colon-colonizing ability by feeding them to male CD-1 mice receiving streptomycin sulfate (5 g/liter) in their drinking water. Each strain, if fed alone, was found in the feces throughout the 15 days of the experiment at about 10(8) CFU/g for the smooth strain or 10(7) CFU/g for each of its rough derivatives. However, when mice were fed equal numbers of two strains (with differentiating antibiotic resistance characters), the strain with the more complete LPS was found in the feces in great excess, 1000- to 100,000-fold, according to the pair. Thus, when strains were placed in direct competition with one another, their relevant colon-colonizing abilities were found to be wild type greater than rfb much greater than rfaJ greater than rfa-990, showing that the ability of a Salmonella strain to colonize the mouse large intestine decreases as its LPS structure becomes more defective.
Visceral leishmaniasis in congenic mice of susceptible and resistant phenotypes: T-lymphocyte-mediated immunosuppression
Authors: Nickol A D , Bonventre P F
Published: January 2020

This paper continues a comparative study (A. D. Nickol and P. F. Bonventre, Infect. Immun. 50:160-168, 1985) describing immune responses exhibited by congenic, Lshs mouse strains C57B1/10 (cure) and B10.D2 (noncure) during the course of disseminated leishmaniasis. We report that sublethal whole-body irradiation of B10.D2 mice before challenge with Leishmania donovani converted the noncuring mice to a curing phenotype. Splenic lymphocytes from L. donovani-infected B10.D2 mice failed to proliferate in response to parasite antigen stimulation in vitro. Splenic lymphocytes from irradiated, cured B10.D2 mice regained the capacity to respond to the parasite antigen stimulus. Transfer of T cells but not B cells from L. donovani-infected B10.D2 mice prevented the acquisition of immunity and recovery from infection in X-irradiated mice. In addition, a splenic T-cell population from L. donovani-infected B10.D2 mice suppressed the proliferation in vitro of parasite antigen-stimulated lymphocytes of irradiation-cured B10.D2 mice. Suppressor T cells were not demonstrable in the spleens of spontaneously cured C57B1/10 mice. Splenic lymphocytes from infected B10.D2 mice were deficient in the production of macrophage-activating factor (MAF) upon stimulation by L. donovani antigens in vitro. Deficient MAF production was specific for parasite antigen stimulation, because MAF production subsequent to concanavalin A stimulation of splenic lymphocytes from infected B10.D2 mice was not suppressed. The data suggest that a genetically based immunological defect in B10.D2 mice prevents the acquisition of effective cell-mediated immunity and subsequent elimination of L. donovani from tissue macrophages. The immunological deficit, not apparent in the curing C57B1/10, appears to be caused by the development of parasite antigen-specific suppressor T cells during the course of the disseminated leishmaniasis.
Replication rate of Pseudomonas aeruginosa in the murine lung
Authors: Sordelli D O , Cerquetti M C , Hooke A M
Published: January 2020

Using a method recently developed at our laboratory, we determined the initial rate of Pseudomonas aeruginosa replication in the lung under different experimental conditions. Mice were exposed to aerosols containing mixtures of a temperature-sensitive (ts) mutant of P. aeruginosa and its parental wild type (wt). The changes in the ratio of ts:wt were determined by quantitatively culturing homogenates of lungs from animals sacrificed over different time periods. The doubling time (DT) was calculated as the reciprocal of the slope of the linear portion of the curve generated by plotting n = (log [r0/rt])/log 2 against time where r is the ratio of ts:wt at a given time. The DTs measured in both outbred ICR mice and F1 hybrids (DBA/2J X B10.D2/nSnJ) were 32 and 30 min, respectively. These DTs were higher than that determined in the peritoneal cavities of ICR mice (20 min). The DT in the lungs of ICR mice rendered granulocytopenic by treatment with cyclophosphamide was 16 min. Experiments performed with inocula of different sizes showed that DTs tended to be higher in animals aerosolized with low doses of the ts-wt mixture.
Attachment of Staphylococcus aureus to polymethylmethacrylate increases its resistance to phagocytosis in foreign body infection
Authors: Vaudaux P E , Zulian G , Huggler E , Waldvogel F A
Published: January 2020

The mechanisms responsible for the development of a pyogenic infection (most commonly due to staphylococci) in the vicinity of an implanted foreign body have been studied recently by several investigators. Thus, we have been able to demonstrate that the phagocytic function of residential polymorphonuclear leukocytes (PMN) is deficient in the presence of a foreign body. Others have shown that in the presence of foreign surfaces, microorganisms produce extracellular amorphous material, the pathogenic role of which is still to be defined. In the present study we use a novel assay system to demonstrate that Staphylococcus aureus Wood 46, after attachment to polymethylmethacrylate (PMMA), shows increased resistance to the phagocytic-bactericidal action of normal PMN. The first step of this assay involves the reproducible attachment of [3H]thymidine-labeled bacteria to PMMA cover slips. During the second step, attached bacteria were exposed to guinea pig peritoneal exudate PMN. In the third and final step, attached S. aureus cells were removed from the cover slips using a procedure harmless to the bacteria. The extent of bacterial detachment was estimated by radioactive counts and their viability by standard colony counts. Whereas bacteria that were attached artificially and rapidly by centrifugation and immediately exposed to PMN were killed in the phagocytic assay, bacteria adhering spontaneously to the cover slips for a prolonged period of time were more resistant to the killing action of the phagocytes. The spontaneous adherence of S. aureus to PMMA renders it poorly susceptible to the killing action of PMN.
Albumin as a blocking agent in studies of streptococcal adsorption to experimental salivary pellicles
Authors: Gibbons R J , Etherden I
Published: January 2020

A procedure involving blocking with bovine albumin was useful for differentiating between streptococcal interactions with components of experimental salivary pellicles similar to those which form on teeth and potential interactions with uncoated areas of mineral.
Cloning and expression of the leukotoxin gene of Pasteurella haemolytica A1 in Escherichia coli K-12
Authors: Lo R Y , Shewen P E , Strathdee C A , Greer C N
Published: January 2020

A clone bank of Pasteurella haemolytica A1 was constructed by partial digestion of the genomic DNA with Sau3A and ligation of 5- to 10-kilobase-pair fragments into the BamHI site of the plasmid vector pBR322. After transformation into Escherichia coli K-12, a total of 4 X 10(3) recombinant clones was obtained. These were screened for the production of P. haemolytica soluble antigens by a colony enzyme-linked immunosorbent assay blot method with a rabbit antiserum raised against the soluble antigens. The clones producing P. haemolytica soluble antigens were then analyzed for the production of the leukotoxin by a cytotoxicity assay with cells from a bovine leukemia-derived B-lymphocyte cell line as the target cells. Positive clones were identified, and subsequent restriction analysis of the recombinant plasmids showed that the same 6.3 kilobase pairs of insert DNA was cloned in either of the two orientations into the plasmid vector pBR322. One of the clones was selected for further characterization of the leukotoxin as produced in E. coli. Tests for heat lability and target cell species specificity with canine, porcine, and human peripheral blood lymphocytes indicated that the activity of the cloned leukotoxin was identical to that of the P. haemolytica leukotoxin. Furthermore, the E. coli-produced leukotoxin was also neutralized by bovine or rabbit antiserum known to have antitoxic activity. When cellular proteins from the E. coli clones were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis, a 100,000-dalton protein was identified which corresponded to one of the soluble antigens found in the leukotoxic culture supernatant of P. haemolytica. These results demonstrated that the gene(s) for the P. haemolytica leukotoxin have been cloned and that the leukotoxin was expressed in E. coli.
Additive protective effects of colostral antipili antibodies in calves experimentally infected with enterotoxigenic Escherichia coli
Authors: Contrepois M G , Girardeau J P
Published: January 2020

With oral infection of calves by an enterotoxigenic Escherichia coli strain carrying K99, F41, and FY adhesins, colostrums from cows vaccinated against either K99+F41 or FY did not provide protection, but a mixture of the two colostrums did. The association of antibodies directed against the different adhesins is more effective than antibodies directed against one adhesin alone for colostral protection against enterotoxigenic E. coli carrying several adhesins.
Plasmodium berghei malaria: effects of acute-phase serum and erythrocyte-bound immunoglobulins on erythrophagocytosis by rat peritoneal macrophages
Authors: Packer B J , Kreier J P
Published: January 2020

Acute-phase serum (APS) collected from Plasmodium berghei-infected rats inhibited phagocytosis of trypsinized rat erythrocytes and of erythrocytes from P. berghei-infected rats. Macrophages (M phi) incubated with APS or heat-aggregated acute-phase serum (HAAPS) for 6 h, followed by 18 h incubation in serum-free medium, exhibited significantly higher levels of phagocytosis than M phi similarly cultured but with normal rat serum. When APS was present at the time of assay, it inhibited erythrophagocytosis by M phi which had been in culture for 0 or 24 h. M phi activation by HAAPS was inhibited by 2-deoxy-D-glucose, which suggests that activation by HAAPS is Fc-receptor mediated. Adsorption of APS with staphylococcal protein A abrogated the ability of APS to inhibit phagocytosis and that of HAAPS to effect M phi activation, suggesting that immune complexes are involved in both processes. Surface-bound immunoglobulins eluted from erythrocytes of P. berghei-infected rats promoted phagocytosis of trypsinized erythrocytes by HAAPS-activated M phi but not by resting M phi. These results indicate that the immunoglobulins which bind to infected or damaged erythrocytes during malarial infections promote erythrophagocytosis by activated M phi and that the immune complexes in serum from rats with acute malaria may inhibit erythrophagocytosis early in the infection but may, over time, induce changes in the M phi which later facilitate erythrophagocytosis.
Neutralization of lymphokine-mediated antirickettsial activity of fibroblasts and macrophages with monoclonal antibody specific for murine interferon gamma
Authors: Jerrells T R , Turco J , Winkler H H , Spitalny G L
Published: January 2020

Lymphokine-mediated inhibition of Rickettsia prowazekii multiplication in L929 fibroblasts was eliminated by treatment of the lymphokine with a monoclonal antibody specific for interferon-gamma. Soluble monoclonal antibody and antibody conjugated to Sepharose beads were equally effective. Macrophage activation to limit the multiplication of Rickettsia conorii was eliminated with antibody-conjugated beads; however, neutralization of the ability to activate macrophages with soluble antibody was not complete and required more antibody than did neutralization of antiviral activity.
Cloning of a gene involved in regulation of exotoxin A expression in Pseudomonas aeruginosa
Authors: Hedstrom R C , Funk C R , Kaper J B , Pavlovskis O R , Galloway D R
Published: January 2020

We have cloned a gene from Pseudomonas aeruginosa that stimulates the expression of exotoxin A. A recombinant library of genomic DNA from strain PA103 constructed with a broad-host-range plasmid vector containing chromosomal insert fragments generated by Sau3A was used to transform the hypotoxigenic mutant strain PA103-29. A recombinant plasmid, pFHK6, was isolated from a PA103-29 transformant which displayed increased toxin production. From pFHK6, which contained a 20-kilobase-pair chromosomal insert, a 3-kilobase-pair XhoI fragment was isolated and subcloned into the plasmid cloning vector pVK101 to give pFHK10. In toxigenic P. aeruginosa strains containing pFHK10, toxin expression was increased 10-fold and high levels of iron in the culture medium only partially inhibited the overproduction. Expression studies suggested that pFHK10 did not contain the toxin structural gene. In addition, Southern analysis with the 3-kilobase-pair XhoI fragment suggested that the putative toxin regulatory gene is common among different strains of P. aeruginosa including previously reported nontoxigenic strains.
Localization of gonococcal lipopolysaccharide and its relationship to toxic damage in human fallopian tube mucosa
Authors: Cooper M D , McGraw P A , Melly M A
Published: January 2020

An experimental model using human fallopian tubes in organ culture was used to study the localization of purified gonococcal lipopolysaccharide (LPS). LPS was visualized by light microscopy with immunoperoxidase staining. Immediately after addition to fallopian tube organ cultures, gonococcal LPS aggregated on the tips of cilia. By 1 to 2 h after exposure, LPS could be seen distributed throughout the cytoplasm of ciliated and nonciliated cells in structures resembling vesicles. By 12 h, there were sloughed, ciliated cells present in the fallopian tube lumen, which had positive LPS stain on their surfaces as well as in their cytoplasm. By 24 h, LPS was distributed throughout the cytoplasm. Control experiments with rabbit oviduct organ cultures showed that LPS failed to attach, enter, or damage mucosal cells. These studies illustrate the initial localization of LPS on human mucosal cells and its uptake into the cells, which are coincident with toxicity for ciliated epithelial cells.
Plasmids coding for colonization factor antigens I and II, heat-labile enterotoxin, and heat-stable enterotoxin A2 in Escherichia coli
Authors: Echeverria P , Seriwatana J , Taylor D N , Changchawalit S , Smyth C J , Twohig J , Rowe B
Published: January 2020

Colonization factor antigens I and II (CFA/I and CFA/II) are important in the pathogenesis of diarrhea in humans caused by some enterotoxigenic Escherichia coli (ETEC). Plasmid DNA from 16 CFA/I+ and five CFA/II+ ETEC were examined by Southern blot analysis with enterotoxin gene probes and were compared with plasmid DNA from derivatives of the same ETEC that had lost the ability to produce these colonization factors. Among the 16 CFA/I+ ETEC strains, the loss of CFA/I was accompanied by the loss of a plasmid of between 34 and 68 megadaltons (MDa) coding for heat-stable enterotoxin A2 (ST-A2) in 12 strains, by the loss of a 60-MDa plasmid coding for heat-labile enterotoxin (LT) and ST-A2 in one strain, or by deletions of a segment of DNA encoding for ST-A2 in three strains. Among five CFA/II+ ETEC strains, the loss of CFA/II was associated with the loss of a plasmid of 75 MDa coding for LT and ST-A2 in three strains, with the loss of genes coding for LT and ST-A2 from a 68-MDa plasmid in one strain, or with no discernible loss of a plasmid or DNA sequences coding for enterotoxins in the remaining strain. The loss of CFA/I and CFA/II production was associated with the loss of DNA sequences encoding for ST-A2 in 20 of 21 ETEC examined.
Effect of macrophage activation on phagocyte-Plasmodium interaction
Authors: Brown K M , Kreier J P
Published: January 2020

We investigated the effect of both immune and normal sera on the binding of free Plasmodium berghei by resident and activated macrophages. Resident macrophages bound plasmodia to a greater extent than did activated macrophages, regardless of treatment. Resident macrophages bound free plasmodia, predominantly trophozoites, in the presence of normal serum by a mechanism inhibited by N-acetylglucosamine and N-acetylmannosamine. Macrophages activated through treatment with Propionibacterium acnes ("Corynebacterium parvum"), on the other hand, did not bind free plasmodia in the presence of normal serum through systems inhibited by N-acetylmannosamine or N-acetylglucosamine. The binding of free plasmodia by activated macrophages was greatest in the presence of immune serum and could be inhibited by immune complexes but not by N-acetylmannosamine or N-acetylglucosamine. These results suggest that a receptor for a carbohydrate component of a normal serum opsonin mediates initial adherence of plasmodial antigen onto resident macrophages, triggering both the immunological cascade and macrophage activation. After activation, the macrophages no longer have the carbohydrate-specific receptor but do have functional Fc receptors which mediate the adherence of immune-serum-opsonized plasmodia.
Isolation of the outer membrane components of Bordetella pertussis which enhance the immunogenicity of Haemophilus influenzae type b capsular polysaccharide polyribosyl ribitol phosphate
Authors: Monji N , Stebbins M R , McCoy D W , Kuo J S
Published: January 2020

The outer membrane (OM) component(s) from Bordetella pertussis which potentiated the antigenicity of purified Haemophilus influenzae type b capsular polysaccharide, polyribosyl ribitol phosphate (PRP), has been isolated and partially characterized. The OM was isolated from spheroplasting medium by precipitating at pH 5.0; fractionation was carried out by dissolving the crude OM in Triton X-100 followed by selective precipitation of OM in 50% ethanol. Further purification of OM was accomplished by DEAE-Sepharose 6BCL and Sephacryl S-300 chromatography. The biochemical composition and protein profiles on sodium dodecyl sulfate-polyacrylamide gel electrophoresis of various OM preparations have been examined. Combined vaccines consisting of OM components and PRP were given to young Sprague-Dawley rats, and the serum antibody to PRP was measured by an [3H]PRP binding assay. The purified OM containing mainly a 30,000-dalton protein, but not purified lipopolysaccharide or leukocytosis-promoting toxin, exhibited strong enhancement of PRP immunogenicity.
Species specificity and lack of production of STb enterotoxin by Escherichia coli strains isolated from humans with diarrheal illness
Authors: Weikel C S , Tiemens K M , Moseley S L , Huq I M , Guerrant R L
Published: January 2020

Escherichia coli strains produce at least two heat-stable enterotoxins, STa and STb. STa is well known to be important in the pathogenesis of human diarrheal disease; the role of STb has not been defined. Fifty-two E. coli strains recovered from human diarrheal illness in northeast Brazil or Bangladesh were examined in weaned porcine ligated intestinal segments for STb activity. A total of 113 E. coli strains from human diarrheal disease in northeast Brazil and 28 E. coli strains from Bangladesh were examined for DNA hybridization to a STb gene probe. None of these strains produced STb as detected by enterotoxic activity or by the gene probe. We also examined adult human ileal mucosa for responses to STb in the Ussing chamber in vitro. In contrast to piglet jejunum, which consistently responds electrogenically to crude STb, human ileal tissue showed no response to STb but responded electrogenically to theophylline (10 mM). These results suggest that STb-producing E. coli strains are not a major cause of diarrheal illness in humans.
In vivo properties of a cloned K88 adherence antigen determinant
Authors: Dougan G , Sellwood R , Maskell D , Sweeney K , Liew F Y , Beesley J , Hormaeche C
Published: January 2020

An Escherichia coli strain of serotype O9:K36:H19 harboring the K88 recombinant plasmid pMK005 was able to efficiently colonize the small bowel of young piglets after oral infection. The strain expressed K88 antigen in vivo, and bacteria were detected in close association with the surface of the intestinal villi. Mice infected orally or intravenously with attenuated Salmonella typhimurium SL3261 harboring pMK005 were well protected against subsequent challenge with the highly virulent S. typhimurium SL1344. Anti-K88 antibodies were detected in the serum of mice immunized with SL3261(pMK005).
Early-phase endotoxin tolerance: induction by a detoxified lipid A derivative, monophosphoryl lipid A
Authors: Madonna G S , Peterson J E , Ribi E E , Vogel S N
Published: January 2020

After a sublethal exposure to lipopolysaccharide (LPS) or to lipid A, which is that portion of the LPS molecule associated with endotoxicity, a transient period ensues during which a normally responsive individual is rendered hyporesponsive to LPS-induced toxicity. This period has been defined as early-phase endotoxin tolerance. Recently, a nontoxic derivative of lipid A from Salmonella typhimurium, monophosphoryl lipid A (MPL), was isolated and purified. In this study, we assessed the ability of MPL to induce early endotoxin tolerance. Initial injection of MPL resulted in a dose-dependent stimulation of both serum colony-stimulating factor and serum interferon, indicators of in vivo LPS responsiveness. In contrast, MPL failed to induce the symptoms of endotoxicity which are normally seen after injection of even sublethal amounts of intact endotoxin or lipid A preparations. Injection of MPL on day 0 reduced significantly the amount of LPS-induced serum colony-stimulating factor and interferon produced upon challenge with Escherichia coli LPS 3 days later and also mitigated toxic manifestations, as evidenced by a marked increase in the 50% lethal dose. Like the early tolerance induced by wild-type (toxic) LPS, MPL-induced tolerance was characterized by an accompanying elevation in the number of bone marrow-derived macrophage progenitor cells and by an alteration in bone marrow cell sizing profiles. These results indicate that MPL is effective in inducing a state of LPS-hyporesponsiveness without the toxic side effects of endotoxin and that the structural component(s) necessary for induction of early-phase endotoxin tolerance is contained within MPL.