bluuebunny/crossref_metadata_2025 huggingface dataset:
Expression of protein F, the fibronectin-binding protein of Streptococcus pyogenes JRS4, in heterologous streptococcal and enterococcal strains promotes their adherence to respiratory epithelial cells
Authors: Hanski E , Horwitz P A , Caparon M G
Published: January 2020

In a previous study we reported the identification of protein F, a fibronectin-binding protein that was essential to the ability of Streptococcus pyogenes JRS4 to adhere to respiratory epithelial cells (E. Hanski and M. Caparon, Proc. Natl. Acad. Sci. USA, 89:6172-6176, 1992). To further evaluate the role of protein F in the adherence of the group A streptococci, we screened other group A streptococcal strains, including six recent clinical isolates, and one strain of Enterococcus faecalis for their capacity to bind fibronectin and for the presence of the gene encoding protein F (prtF). Seven of eight group A streptococcal strains analyzed, including all recent clinical isolates, both bound fibronectin at high affinity and contained DNA sequences that hybridized with a prtF-specific probe. One group A streptococcal isolate and the strain of E. faecalis examined neither contained a prtF-related gene nor bound fibronectin. These two strains also could not efficiently adhere to respiratory epithelial cells. However, upon the introduction of the cloned prtF gene, both of these strains gained the capacity to bind fibronectin and to adhere to respiratory epithelial cells. These results suggest that protein F is an important adhesin, which may have a general role in the virulence of the group A streptococci.
Clonal and antigenic analysis of serogroup A Neisseria meningitidis with particular reference to epidemiological features of epidemic meningitis in the People's Republic of China
Authors: Wang J F , Caugant D A , Li X , Hu X , Poolman J T , Crowe B A , Achtman M
Published: January 2020

Representative strains of serogroup A Neisseria meningitidis were chosen from all major meningitis epidemics worldwide since 1960 and subjected to analysis for the electrophoretic variation of 15 cytoplasmic allozymes and four outer membrane proteins. The 290 strains defined 84 unique electrophoretic types which were classified in nine subgroups. Tests with monoclonal antibodies specific for conserved pilin epitopes showed that the class I, IIa, and IIb epitopes were uniform within the subgroups. Similarly, the subgroups were uniform for expression of different variable regions of class 1 outer membrane protein, with a few minor exceptions. Many of the bacteria tested were isolated in the People's Republic of China, and the epidemiology of Chinese epidemics of meningococcal meningitis is described. The analysis approaches a global description of epidemic meningitis caused by serogroup A meningococci in the past 30 years.
Characterization of inhibition of Mycobacterium avium replication in macrophages by normal human serum
Authors: Douvas G S , May M H , Ross E , Crowle A J
Published: January 2020

Serum from some AIDS patients permits the rapid multiplication of Mycobacterium avium in cultured human macrophages. Serum from human immunodeficiency virus-negative individuals inhibits replication. The characteristics of the serum-induced inhibition were examined here. M. avium 7497 serovar 4 grew exponentially in macrophages when they were cultured in serumless medium. Growth was measured by determining the CFU after infected macrophages were lysed at 0 to 7 days after infection. Normal AB serum (5 to 10%) added to infected macrophages resulted in an initial 4-day lag of bacterial growth followed by rapid replication from 4 to 7 days. Serum also inhibited bacterial replication in medium without macrophages. This inhibition was not biphasic but was sustained over 7 days. Macrophage-associated M. avium became less responsive to serum inhibitor within 24 h after infection of macrophages. Within 2 days of culture, M. avium no longer responded to inhibitor. Replication of macrophage-derived M. avium (Vi) was in some instances serum inhibitable and at other times was enhanced by serum, when it was used to infect fresh macrophages. The Vi phenotype remained serum inhibitable without macrophages. Preinfection of macrophages with heat-killed M. avium did not alter serum-induced bacterial inhibition or escape from inhibition.
Isolation and expression of a gene which encodes a wall-associated proteinase of Coccidioides immitis
Authors: Cole G T , Zhu S W , Hsu L L , Kruse D , Seshan K R , Wang F
Published: January 2020

A chymotrypsinlike serine proteinase of Coccidioides immitis with an estimated molecular size of 34 kDa has been shown by immunoelectron microscopy to be associated with the walls of the parasitic cells of this human respiratory pathogen. The proteinase has been suggested to play a role in spherule development. We report the isolation of a 1.2-kb cDNA from an expression library of C. immitis constructed in the lambda ZAP II phage vector. The cDNA is suggested to encode the 34-kDa protein. We demonstrate identity between segments of the deduced amino acid sequence of the open reading frame of the 1.2-kb cDNA and three distinct sequences obtained from cyanogen bromide cleavage peptides of the purified proteinase. The occurrence of N-glycosyl linkage sites in the deduced sequence of 309 amino acids of the open reading frame (ORF) correlates with our identification of such linkage sites in the native glycosylated proteinase. A protein encoded by an 800-bp fragment of the 1.2-kb cDNA, which was produced by transformed Escherichia coli XL1-Blue, was recognized by the anti-34-kDa protein antibody in a Western blot (immunoblot). Northern (RNA) hybridization of total poly(A)-containing RNA of C. immitis with the labeled 1.2-kb cDNA clone revealed a single band of approximately 1.75 kb. Partial homology was demonstrated between the deduced amino acid sequence of the ORF (927 bp) and reported sequences of alpha-chymotrypsin and chymotrypsinogens. Expression of the proteinase gene was examined by Northern dot blot analysis of total RNA from different stages of parasitic cell development in C. immitis. Maximum levels of specific mRNA were detected during early endospore wall differentiation. The 34-kDa proteinase appears to be concentrated in walls of the parasitic cells at stages of active growth. We suggest that the enzyme may participate in wall plasticization and/or intussusception or in cell wall turnover.
Endogenous tumor necrosis factor, interleukin-6, and gamma interferon levels during Listeria monocytogenes infection in mice
Authors: Nakane A , Numata A , Minagawa T
Published: January 2020

Mice were infected intravenously with a sublethal dose of Listeria monocytogenes cells and then levels of tumor necrosis factor (TNF), interleukin-6 (IL-6), and gamma interferon (IFN-gamma) in the bloodstreams, spleens, and livers were monitored. The maximum level of TNF was detected at 72 h in the spleens and livers, but TNF was never detected in the bloodstreams. IL-6 appeared in the bloodstreams and spleens and peaked at 48 h. The maximum level of IFN-gamma could be detected in all three specimens, and the highest titer was shown in the spleens. Endogenous TNF production was suppressed by in vivo administration of anti-CD4 monoclonal antibody (MAb) or anti-asialo GM1 antibody but not by anti-CD8 MAb, whereas none of these antibodies suppressed endogenous IL-6 production. Endogenous production of neither IL-6 nor IFN-gamma was inhibited in rabbit anti-recombinant mouse TNF-alpha antibody-treated mice. Similarly, production of TNF and IL-6 did not decrease in anti-mouse IFN-gamma MAb-treated animals, but TNF production was augmented in these animals. These results suggest that the these endogenous cytokines are produced by different mechanisms in L. monocytogenes infection.
Molecular and cellular characterization of the 29-kilodalton peripheral membrane protein of Entamoeba histolytica: differentiation between pathogenic and nonpathogenic isolates
Authors: Reed S L , Flores B M , Batzer M A , Stein M A , Stroeher V L , Carlton J E , Diedrich D L , Torian B E
Published: January 2020

To further characterize the 29-kDa surface antigen of Entamoeba histolytica, we analyzed the complete nucleotide sequence and compared the immunoreactivity of this antigen in pathogenic and nonpathogenic strains. Five cDNA clones (one 1.0-kb full-length clone, designated p13, and four partial-length clones) encoding the antigen were analyzed for allelic variation. Comparison of the nucleotide sequences revealed several single-nucleotide substitutions in all five cDNAs, two of which resulted in amino acid differences. Localization of the antigen to the amebic surface in a previous report (B. E. Torian, B. M. Flores, V. L. Stroeher, F. S. Hagen, and W. E. Stamm, Proc. Natl. Acad. Sci. USA 87:6358-6362, 1990) was corroborated by transmission electron microscopy showing colloidal gold particles on the surface of the trophozoites. Computer analysis of the deduced amino acid sequence predicted that the protein encoded by p13 was a hydrophilic peripheral membrane protein, and these data were confirmed by a Triton X-114 membrane extraction showing the presence of the 29-kDa antigen primarily in the aqueous phase of the detergent partition. Monoclonal antibodies to a fusion peptide differentiated between pathogenic and nonpathogenic clinical strains of E. histolytica in immunoblots. Although no immunoreactive epitopes were detected on nonpathogenic strains, Northern (RNA) analysis and DNA-DNA hybridization with a 700-bp cDNA probe demonstrated that mRNA and the gene encoding the 29-kDa surface antigen were present in both pathogenic and nonpathogenic clinical isolates.
Human secretory and serum antibodies recognize environmentally induced antigens of Giardia lamblia
Authors: Reiner D S , Gillin F D
Published: January 2020

The variability in duration and severity of infection with Giardia lamblia is likely to be due to trophozoite interactions with immune and nonimmune components of the small intestinal milieu. Despite its potential importance, nothing is known of the isotype or the specificity of the secretory antibody response to G. lamblia. In the present study, we show that serum and secretory antibodies recognize many Giardia antigens whose expression is induced by exposure to selected intestinal conditions. Isotype-specific immunoblots of antigens from trophozoites grown at pH 7.0 without bile or at the intestinal pH of 7.8 with bile were reacted with milk or serum antibodies from subjects with or without histories of giardiasis. While the results were complex, several key observations emerged. Serum and secretory immunoglobulin A (IgA), IgM, and IgG antibodies reacted with many regulated antigens. Antigen recognition patterns varied with isotype and between milk and serum antibodies of the same isotype. Antigen recognition also differed among subjects. Antibodies from virtually every patient recognized some G. lamblia antigens. Furthermore, milk and/or serum samples from putative controls without histories of giardiasis were positive more frequently than would be predicted from published prevalence studies, suggesting either that these antibodies may be cross-reactive or that undiagnosed infections with G. lamblia may be more common than previously thought. Thus, recognition of neoantigens induced by host conditions may be due to conserved or cross-reactive epitopes which could constitute a form of immune evasion by G. lamblia.
Clustering of an outer membrane adhesin of Haemophilus parainfluenzae
Authors: Liljemark W F , Bloomquist C G , Lai C H
Published: January 2020

Haemophilus parainfluenzae synthesizes an outer membrane protein adhesin which mediates binding to oral streptococci, salivary pellicle, and neuraminidase-treated erythrocytes. An indirect gold labeling technique and immunoelectron microscopy verified the location of this outer membrane protein. Further, a clustering of gold particles was observed in irregular patches at the cell surface.
Molecular cloning of the recA gene and construction of a recA strain of Francisella novicida
Authors: Berg J M , Mdluli K E , Nano F E
Published: January 2020

A gene locus that is functionally analogous to the recA gene of Escherichia coli was molecularly cloned from Francisella novicida. The cloned gene was found to suppress the sensitivity of an E. coli strain to DNA-damaging agents and to support genetic recombination in E. coli. After transposon mutagenesis, the recA-like gene locus was returned to F. novicida and a UV-sensitive F. novicida strain was isolated. In contrast to the wild-type strain, this UV-sensitive strain could not be transformed with chromosomal DNA.
Modified lymphocyte response to mitogens induced by the lipopeptide fragment derived from Mycobacterium avium serovar-specific glycopeptidolipids
Authors: Tassell S K , Pourshafie M , Wright E L , Richmond M G , Barrow W W
Published: January 2020

The beta-elimination procedure was used to obtain two major fragments of Mycobacterium avium glycopeptidolipid antigens. The lipopeptide fragment, not the oligosaccharide, diminished the mitogen-induced blastogenic response of spleen cells at concentrations lower than those which affected viability. Electron microscopy revealed an internalization of lipopeptide and disruption of intracellular organization.
Role of superoxide dismutase in resistance of Porphyromonas gingivalis to killing by polymorphonuclear leukocytes
Authors: Amano A , Ishimoto T , Tamagawa H , Shizukuishi S
Published: January 2020

Porphyromonas gingivalis in which the synthesis of superoxide dismutase (SOD) was induced by nitrate or by aeration was rendered resistant to killing by polymorphonuclear leukocytes. SOD purified from either anaerobically maintained or aerated cells also inhibited bacterial killing when added exogenously, and no difference between the effects of the two SODs was observed. These results suggest that SOD may form part of a defense mechanism that helps protect P. gingivalis against killing by polymorphonuclear leukocytes.
Differentiation of salivary agglutinin-mediated adherence and aggregation of mutans streptococci by use of monoclonal antibodies against the major surface adhesin P1
Authors: Brady L J , Piacentini D A , Crowley P J , Oyston P C , Bleiweis A S
Published: January 2020

The ability to adhere to salivary agglutinin-coated hydroxyapatite beads and to aggregate in the presence of fluid-phase salivary agglutinin was tested by using 25 isolates of mutants streptococci representing eight serotypes. Both adherence and aggregation activity correlated with expression of the Mr-185,000 cell surface antigen P1 on Streptococcus mutans serotype c, e, and f strains. In addition, it was shown that the P1 molecule itself served as the adhesin of S. mutans serotype c, since adherence was significantly inhibited by the presence of recombinant-specified Mr-150,000 P1. The ability of S. sobrinus strains to adhere or aggregate did not correlate with expression of the P1 cross-reactive antigen SpaA. There was also evidence for interaction with salivary agglutinin, as manifested by aggregation but not adherence of S. rattus serotype b, which does not express a P1 cross-reactive antigen. To understand the interaction of P1 with salivary agglutinin at the molecular level, a panel of 11 anti-P1 monoclonal antibodies was tested for inhibitory activity in adherence and aggregation inhibition assays. Overlapping, but not identical, subsets of monoclonal antibodies were found to inhibit adherence and aggregation, indicating that the interactions of P1 with salivary agglutinin which mediate these two phenomena are different. The localization of functional domains of P1 which may mediate the aggregation and adherence reactions is discussed.
Galactoxylomannans of Cryptococcus neoformans
Authors: James P G , Cherniak R
Published: January 2020

Galactoxylomannans (GalXMs) from single isolates of Cryptococcus neoformans serotypes A, B, and D were isolated from culture supernatants and then purified by affinity, ion-exchange, and gel-filtration chromatography. GalXMs are a group of closely related complex polysaccharides. GalXMs from serotypes A (9759 A) and C (3183 C) and an acapsular mutant of serotype D (Cap67 D) have similar galactose, xylose, and mannose molar ratios, but each has some unique structural features. GalXM9759 A and GalXM 3183 C were associated with a starchlike glucan that was removed during purification. Only a trace of glucose was detected in the Cap67 D GalXM. Gas-liquid chromatography-mass spectroscopy of per-O-methylated polysaccharides and 13C nuclear magnetic resonance spectroscopy showed that GalXM is a complex branched polysaccharide. The main chain consists of mannose or galactose or alternating mannose and galactose residues. Xylose is present only as nonreducing termini. Galactofuranose occurs only in 3183 C and Cap67 D, and it is always present as nonreducing termini.
In vitro and in vivo induction of tumor necrosis factor alpha by Borrelia burgdorferi
Authors: Defosse D L , Johnson R C
Published: January 2020

Tumor necrosis factor alpha (TNF-alpha) is an immunoregulatory cytokine with many biological activities including the mediation of inflammation. We examined sera and synovial fluids from patients seropositive for infection with Borrelia burgdorferi using a radioimmunoassay specific for TNF-alpha. Significant elevation of TNF-alpha was found in the sera and synovial fluids of patients examined, while controls showed no elevation. Sera of mice infected with B. burgdorferi contained elevated levels of TNF-alpha which varied during the course of a 24-day infection. To determine whether B. burgdorferi is capable of inducing TNF-alpha production, spirochetes were added to adherent human peripheral blood mononuclear cells or mouse peritoneal exudate cells and 24 h later supernatants were assayed. TNF-alpha induction occurred in a dose-dependent manner. The maximum stimulation occurred when a ratio of 1 to 10 spirochetes per mononuclear cell was used. At optimal concentrations, induction was not diminished by inactivation of spirochetes or pretreatment with polymyxin B. These results suggest that an increase in TNF-alpha production may occur as a result of infection with B. burgdorferi.
Opsonin-independent phagocytosis of group B streptococci: role of complement receptor type three
Authors: Antal J M , Cunningham J V , Goodrum K J
Published: January 2020

The role of complement receptor type 3 (CR3) in nonopsonic recognition of group B streptococci (GBS) by macrophages was investigated. Monoclonal anti-CR3 (anti-Mac-1) inhibited phagocytosis of GBS strains by as much as 50% in serum-free cultures of both mouse peritoneal macrophages and the macrophage cell line PU5-1.8. GBS uptake was unaffected by the presence of anti-C3 or salicylhydroxamate, an inhibitor of the covalent binding reaction of C3. Soluble antibodies to LFA-1 or to the common beta-chain (CD18) of the LFA-1/CR3/p150,95 family of cell adhesion molecules did not inhibit GBS uptake. Down-modulation of surface Mac-1 on macrophages following adherence to anti-Mac-1- or anti-CD18-coated surfaces also inhibited uptake of GBS. Further evidence for GBS interaction with CR3 was demonstrated by reduction of EC3bi rosette formation in macrophages adherent to GBS-coated plates. These studies suggest that GBS can interact with macrophage CR3, promoting phagocytosis in a C3-independent fashion. In the absence of specific immunity in neonates, this recognition mechanism may be a significant virulence determinant for GBS which poorly activate the alternate complement pathway.
Chlamydia trachomatis pneumonia induces in vivo production of interleukin-1 and -6
Authors: Magee D M , Smith J G , Bleicker C A , Carter C J , Bonewald L F , Schachter J , Williams D M
Published: January 2020

Cytokine induction during Chlamydia trachomatis pneumonia may alter the pathogenesis or course of disease. We examined interleukin-1 (IL-1) and IL-6 production by measuring mRNA and bioactivity in murine lungs. mRNA and bioactivity for IL-1 alpha, IL-1 beta, and IL-6 increased after Chlamydia infection. These cytokines may be important in regulating host defenses against C. trachomatis.
Interactions between extracellular Borrelia burgdorferi proteins and non-Borrelia-directed immunoglobulin M antibodies
Authors: Dorward D W , Huguenel E D , Davis G , Garon C F
Published: January 2020

Previous work showed that outer surface protein A (OspA) and OspB of Borrelia burgdorferi may occur within an extracellular multiprotein complex, which was resolved by electrophoresis as an 83-kDa major extracellular protein band. To characterize the 83-kDa band, we sequenced the N terminus of the predominant peptide in the band and examined the interaction between the associated proteins. Peptide sequence and amino acid composition comparisons showed identity with the heavy chain of immunoglobulin M (IgM). Reduction sensitivity experiments and the recognition of the band by antibodies specific for rabbit mu chain indicated that the multiprotein complex contained pentameric IgM. Immunoelectron microscopy showed that anti-mu chain antibodies and monoclonal antibodies to OspA and OspB bound to extracellular amorphous material surrounding cells. Furthermore, the Osps coprecipitated with either nonspecific polyclonal rabbit IgM antibodies or with murine monoclonal anti-human serum albumin IgM antibodies, using insoluble anti-mu chain antibody conjugates. Although the apparent 83-kDa complex was stable under conditions of chelation and concentrated salts, it was disrupted by treatment with neuraminidase. These results indicate that extracellular B. burgdorferi proteins, including OspA and OspB, interact with IgM. The association is apparently not a classic antibody-antigen interaction but may result from other mechanisms.
Staphylococcus aureus proteins that bind to human endothelial cells
Authors: Tompkins D C , Blackwell L J , Hatcher V B , Elliott D A , O'Hagan-Sotsky C , Lowy F D
Published: January 2020

The adherence of Staphylococcus aureus to human endothelial cells is saturable in both dose- and time-dependent assays. Staphylococcal surface components which bound to endothelial cells in vitro were identified by using biotin-labeled, solubilized staphylococcal proteins. Four trypsin-sensitive components with molecular sizes of 30, 55 to 57, 70, and 85 kDa were recognized. These proteins did not label with the glycan detection system. When staphylococci were harvested during the exponential phase of growth, staphylococcal adherence to endothelial cells was significantly increased and increased expression of the S. aureus binding proteins was observed. Preincubation of endothelial cells with protein A did not reduce S. aureus adherence in an in vitro infection assay. Four S. aureus surface components whose expression is growth phase dependent adhere to human endothelial cells in vitro.
Lymphoproliferative activity of Pseudomonas exotoxin A is dependent on intracellular processing and is associated with the carboxyl-terminal portion
Authors: Legaard P K , LeGrand R D , Misfeldt M L
Published: January 2020

Pseudomonas aeruginosa exotoxin A (PE) represents a microbial superantigen that requires processing by accessory cells in order to induce the proliferation of V beta 8-bearing murine T lymphocytes. In this study, we have observed that PE requires intracellular processing by a protease in order to induce lymphoproliferation. Pepstatin A, an inhibitor of acid proteases, inhibited PE-induced lymphoproliferation, whereas leupeptin, an inhibitor of serine and thiol proteases, had no effect on PE-induced lymphoproliferation. A number of mutant forms of PE were examined for their ability to induce lymphoproliferation. The mutant form which lacks amino acids 5 to 224 of the receptor-binding domain, PE43, was capable of inducing murine thymocytes to proliferate in the presence of accessory cells. However, neither PEgly276, a mutant toxin which undergoes a different intracellular processing pattern than wild-type PE, nor PE589, a mutant toxin which lacks amino acids 590 to 613 at the carboxyl terminus, was able to induce thymocyte proliferation. In addition, the lymphoproliferation induced by the PE43 mutant form of PE could also be inhibited by pepstatin A. Therefore, our data indicate that intracellular processing by a proteolytic enzyme which is inhibited by pepstatin A is critical for PE-induced lymphoproliferation. Furthermore, the lymphoproliferative activity of PE is associated with the carboxyl-terminal portion of PE.
Importance of endogenous tumor necrosis factor alpha and gamma interferon in host resistance against Pneumocystis carinii infection
Authors: Chen W , Havell E A , Harmsen A G
Published: January 2020

C.B-17 scid/scid (SCID) mice that have acquired natural pulmonary infection with Pneumocystis carinii clear these organisms by 19 days after reconstitution with spleen cells from immunocompetent mice and therefore serve as a model for studying the pathogenesis of and immunity to P. carinii pneumonia. The present study examined the importance of endogenous tumor necrosis factor alpha (TNF-alpha) and gamma interferon (IFN-gamma) in the clearance of P. carinii by treatment of reconstituted SCID mice with anti-TNF-alpha and anti-IFN-gamma immunoglobulin G (IgG). Treatment of reconstituted mice with monospecific rabbit anti-TNF-alpha IgG almost completely inhibited the clearance of P. carinii from the lungs. In contrast, treatment with either anti-IFN-gamma antibody (polyclonal or monoclonal) or control IgG had no detectable effect on the clearance of P. carinii. The importance of endogenous TNF-alpha in the clearance of P. carinii was further supported by the finding of TNF-alpha but not IFN-gamma in lung homogenate supernatants from reconstituted SCID mice. Further study revealed that for the complete clearance of P. carinii, TNF-alpha must be present at the early stage of reconstitution, since clearance could be blocked by a single injection of anti-TNF-alpha IgG into SCID mice at day 0 but not at day 6 and/or day 12 after reconstitution. These results strongly suggest that, in reconstituted SCID mice, endogenous TNF-alpha is important in host resistance against P. carinii infection, whereas IFN-gamma appears not to play a significant role.
Effect of antigen-specific T helper cells or interleukin-2 on suppressive ability of macrophage subsets detected in spleens of Trypanosoma cruzi-infected mice as determined by limiting dilution-partition analysis
Authors: Cerrone M C , Ritter D M , Kuhn R E
Published: January 2020

Trypanosoma cruzi, a protozoan parasite and the causative agent of Chagas' disease, induces a state of lymphocyte hyporesponsiveness to both mitogenic and antigenic stimuli in mice during the acute phase of infection. Addition of spleen cells from T. cruzi-infected mice (SCinf) to microcultures of spleen cells from noninfected mice (SCn) suppresses the responsiveness of such cultures to antigenic challenge and to mitogenic stimulation. We analyzed the regulatory cell populations in SCinf by limiting dilution-partition analysis and found a complex regulatory circuit in T. cruzi-infected mice consisting of two suppressive macrophage subsets and an enhancing T-cell population. This T-cell population was able to abrogate or escape the suppressive ability of one suppressor macrophage subset, yet was suppressed by the other macrophage subset. To further study the cellular interactions of this regulatory circuit and analyze the suppressive abilities of the two suppressor macrophage subsets, we examined the effect of adding either primed T helper cells of known specificity or interleukin-2 to the limiting dilution-partition analysis microcultures. The results of these experiments suggest that one suppressor macrophage subset, which is abundant and, therefore, detected with low doses of SCinf, is able to suppress both mitogen- and primary antigen-specific responses but is unable to inhibit cells once they are already activated or primed. The other macrophage subset, which is presumably a less abundant or less active population (since high doses of SCinf are required to detect it), is able to suppress the response of activated or primed T cells by the inhibition of interleukin-2 production.
Analysis of oocyst wall and sporozoite antigens from three Cryptosporidium species
Authors: Nina J M , McDonald V , Dyson D A , Catchpole J , Uni S , Iseki M , Chiodini P L , McAdam K P
Published: January 2020

A comparison was made of the antigenic composition of oocyst walls and sporozoites from Cryptosporidium baileyi from turkeys, C. muris from rodents, and C. parvum from ruminants, employing immunoblotting and immunofluorescence. In immunoblotting, oocyst antigens were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting (immunoblotting) and detected with rabbit polyclonal anti-C. muris or -C. parvum antibodies or murine monoclonal antibodies developed against C. parvum. Immunofluorescence was used to investigate the reactivity of these monoclonal antibodies with air-dried excystation mixtures of sporozoites and oocysts of the different species. The results from both types of experiment indicated that the three Cryptosporidium species could be differentiated immunologically. In comparison, few antigenic differences were found between a number of isolates of C. parvum in immunoblotting. There was also evidence to suggest that C. parvum and C. baileyi were more closely related antigenically to one another than to C. muris.
Biochemical characterization of Porphyromonas (Bacteroides) gingivalis collagenase
Authors: Lawson D A , Meyer T F
Published: January 2020

A protease was purified from Porphyromonas gingivalis 1101, a clinical isolate, by sequential sodium dodecyl sulfate-polyacrylamide gel electrophoresis, substrate diffusion gel electrophoresis, and electroelution. The enzyme cleaved radiolabeled human basement membrane type IV collagen and the synthetic collagen peptide substrate for eukaryotic collagenases. It was inactivated by the thiol protease inhibitor N-ethylmaleimide but not by EDTA or EGTA [ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid] and activated by reducing agents such as beta-mercaptoethanol. The enzyme exists as an active precursor protein of molecular mass 94 kDa and undergoes proteolytic cleavage to 75-, 56-, and 19-kDa forms. Biotin-labeled collagen bound specifically to the 94-kDa form of the protein and to its cleavage products in ligand blots, suggesting a role for this enzyme not only in collagen degradation but also in adhesion to collagenous substrata.
Outer membrane protein patterns mark clones of Escherichia coli O2 and O78 strains that cause avian septicemia
Authors: Kapur V , White D G , Wilson R A , Whittam T S
Published: January 2020

Major outer membrane proteins were isolated from 36 Escherichia coli strains representing six common clones of the O2 and O78 serogroups implicated in avian colisepticemia. Clonal relationships among isolates were inferred from an analysis of polymorphism at 20 enzyme-encoding loci detected by multilocus enzyme electrophoresis. For isolates of these clones, there was a high concordance (greater than 90%) between identity in multilocus genotype and major outer membrane protein patterns. The results indicate that major outer membrane protein patterns discriminate among the genetically different clonal groups that constitute the heterogeneous O2 and O78 serogroups associated with avian disease.
Biochemical studies of Helicobacter mustelae fatty acid composition and flagella
Authors: Suerbaum S , Geis G , Josenhans C , Opferkuch W
Published: January 2020

The fatty acid compositions of Helicobacter mustelae whole cells, isolated phospholipids, and isolated lipopolysaccharides were analyzed by gas-liquid chromatography. Major phospholipid fatty acids were C16:0, C18:0, C18:1, and C19:0 cyc. In isolated lipopolysaccharides, 3-OH-C16:0, 3-OH-C14:0, C14:0, C16:0, and C18:0 were found. The lipid composition of H. mustelae thus showed pronounced differences from that of H. pylori. Flagella were purified by mechanical shearing and centrifugation steps. In all H. mustelae strains, the flagellin had an apparent molecular mass of 53 kDa and was thus the same size as H. pylori flagellin. The flagellin of strain NCTC 12032 was further purified and subjected to N-terminal amino acid sequence analysis. The first 10 amino acids were identical to those of H. pylori flagellin, but the next 5 were different. Significant homology was also found with flagellins of other bacteria.
Degradation of native human hemoglobin following hemolysis by Prevotella loescheii
Authors: Zwickel J , Weiss E I , Schejter A
Published: January 2020

Prevotella loescheii PK1295 can grow on native hemoglobin as a source of heme. Supernatants of P. loescheii cultures hemolysed human erythrocytes and degraded native hemoglobin. These combined activities may provide heme (or iron) for the growth of P. loescheii and other dental plaque bacteria.
Effects of endotoxin-associated protein on hematopoiesis
Authors: Porat R , Yanoov M , Johns M A , Shibolet S , Michalevicz R
Published: January 2020

Endotoxin-associated protein (EAP), a gram-negative bacterial cell wall component, was evaluated for its effects on hematopoietic colony formation in vitro. Colony-stimulating activity, induced by EAP on circulating and bone marrow progenitor cells, was found to be partially mediated by T cells and augmented by interleukin-3. The addition of anti-human interleukin-1 (IL-1) antibodies reduced EAP activity, suggesting that EAP may induce IL-1 production. However, EAP was shown to promote the growth of mature progenitor cells independently, unlike the effects of IL-1 on the hematopoietic system. These studies demonstrate that bacterial components other than lipopolysaccharide, such as EAP, may have hematopoietic activity.
Bacterial metabolism of human polymorphonuclear leukocyte-derived arachidonic acid
Authors: Sorrell T C , Muller M , Sztelma K
Published: January 2020

Evidence for transcellular bacterial metabolism of phagocyte-derived arachidonic acid was sought by exposing human blood polymorphonuclear leukocytes, prelabelled with [3H]arachidonic acid, to opsonized, stationary-phase Pseudomonas aeruginosa (bacteria-to-phagocyte ratio of 50:1) for 90 min at 37 degrees C. Control leukocytes were stimulated with the calcium ionophore A23187 (5 microM) for 5 min. Radiochromatograms of arachidonic acid metabolites, extracted from A23187-stimulated cultures and then separated by reverse-phase high-performance liquid chromatography, revealed leukotriene B4, its omega-oxidation products, and 5-hydroxy-eicosatetraenoic acid. In contrast, two major metabolite peaks, distinct from known polymorphonuclear leukocyte arachidonic acid products by high-performance liquid chromatography or by thin-layer chromatography, were identified in cultures of P. aeruginosa with [3H]arachidonic acid-labelled polymorphonuclear leukocytes. Respective chromatographic characteristics of these novel products were identical to those of two major metabolite peaks produced by incubation of stationary-phase P. aeruginosa with [3H]arachidonic acid. Production of the metabolites was dependent upon pseudomonal viability. UV spectral data were consistent with a conjugated diene structure. Metabolism of arachidonic acid by P. aeruginosa was not influenced by the presence of catalase, superoxide dismutase, nordihydroguaiaretic acid, ethanol, dimethyl sulfoxide, or ferrous ions but was inhibited by carbon monoxide, ketoconazole, and 1,2-epoxy-3,3,3-trichloropropane. Our data suggest that pseudomonal metabolism of polymorphonuclear leukocyte-derived arachidonic acid occurs during phagocytosis, probably by enzymatic epoxidation and hydroxylation via an oxygenase. By this means, potential proinflammatory effects of arachidonic acid or its metabolites may be modulated by P. aeruginosa at sites of infection in vivo.
Anaerobic growth of gonococci does not alter their Opa-mediated interactions with human neutrophils
Authors: Frangipane J V , Rest R F
Published: January 2020

Gonococci grown anaerobically (anaerobic gonococci) in the presence of nitrite induce the expression of at least three novel outer membrane proteins (PANs 1 to 3). Although PAN 1 is expressed by gonococci during gonorrhea, the function of the PAN proteins remains unknown. In the absence of serum, gonococci possessing opacity-associated (Opa, formerly PII) outer membrane proteins adhere to, stimulate, and are phagocytically killed by human neutrophils. Gonococci lacking Opa proteins demonstrate none of these activities. We investigated whether the PAN proteins, or any other characteristics of anaerobic gonococci, altered the ability of nonpiliated, Opa+ or Opa- gonococci to adhere to, stimulate, or be phagocytically killed by neutrophils. The expression of Opa4 by strain F62, as determined by its relative mobility on sodium dodecyl sulfate-polyacrylamide gels, appeared to be unaltered by anaerobic growth, as seen previously (V. L. Clark, L. A. Campbell, D. A. Palermo, T. M. Evans, and K. W. Klimpel, Infect Immun. 55:1359-1364, 1987). Anaerobic and aerobic Opa+ gonococci adhered to and stimulated neutrophils to the same extent. Similarly, anaerobic and aerobic Opa- gonococci adhered to and stimulated neutrophils equally poorly. Finally, anaerobic and aerobic Opa+ gonococci were equally sensitive to phagocytic killing by neutrophils, while anaerobic and aerobic Opa- gonococci were equally resistant to killing. Thus, the role of Opa proteins in mediating the interactions of gonococci with human neutrophils appears unaltered by anaerobic growth, and the PAN proteins, or other cryptic properties of anaerobic gonococci, do not seem to modulate or mediate these phenomena.
Listeria monocytogenes activation of human peripheral blood lymphocytes: induction of non-major histocompatibility complex-restricted cytotoxic activity and cytokine production
Authors: Guo Y , Niesel D W , Ziegler H K , Klimpel G R
Published: January 2020

Gram-negative bacteria have been shown to activate human natural killer (NK) cells. In this report, we show that the gram-positive bacterium Listeria monocytogenes can also activate human NK cells with regard to non-major histocompatibility complex (MHC)-restricted killing and the production of cytokines. Overnight incubation of peripheral blood mononuclear (PBM) cells or enriched NK cell populations with live or Formalin-fixed L. monocytogenes resulted in high levels of non-MHC-restricted cytotoxic activity. Listeria-stimulated non-MHC-restricted cytotoxic activity could be achieved with pathogenic as well as nonpathogenic Listeria strains. PBM cells also produced tumor necrosis factor alpha and different interferons (IFNs) after incubation with Listeria strains. Optimal cytokine production appeared to be dependent on nylon wool- and plastic-adherent cells. Different IFNs were produced by Listeria-stimulated PBM cells obtained from different donors. IFN-gamma was always produced but was sometimes associated with IFN-alpha and/or IFN-beta. Interleukin-2 (IL-2) activity was never detected in culture supernatants obtained from Listeria-stimulated PBM cell cultures. However, IL-2 appeared to be produced by Listeria-stimulated PBM cells, since antibody to IL-2 inhibited Listeria-stimulated NK cell cytotoxic activity. Listeria activation of NK cell cytotoxic activity was also dependent on tumor necrosis factor alpha production. Antibody to IFN-gamma, IFN-beta, or IFN-alpha had no effect on Listeria-stimulated NK cell cytotoxic activity. These results demonstrate that NK cells can be activated by Listeria strains and add further evidence that NK cells may play an important role in host defense against bacterial infections.
Detoxified exoantigens and phosphatidylinositol derivatives inhibit tumor necrosis factor induction by malarial exoantigens
Authors: Bate C A , Taverne J , Playfair J H
Published: January 2020

We have previously shown that malaria parasites liberate exoantigens which, through a phospholipid component, stimulate mouse macrophages to secrete tumor necrosis factor (TNF), which are toxic to D-galactosamine-sensitized mice, and which therefore might be involved in pathology. Plasmodium yoelii exoantigens detoxified by dephosphorylation or digestion with lipases do not induce TNF production. However, these partial structures inhibited its production in response to the exoantigens, although not to bacterial lipopolysaccharide (LPS). When pure phospholipids were tested in a macrophage assay, none stimulated the production of TNF, but phosphatidylinositol (PI) inhibited TNF induction by P. yoelii exoantigens. Moreover, inositol monophosphate (IMP) was the only one of a number of monophosphate saccharides tested which was inhibitory; inositol was not. Macrophages pretreated with PI, IMP, or detoxified exoantigens and then incubated with parasite exoantigens also yielded much less TNF. PI, IMP, and lipase-digested exoantigens of P. yoelii similarly inhibited the TNF-inducing activity of exoantigens of the human parasites Plasmodium falciparum and Plasmodium vivax. Neither PI nor IMP diminished TNF production in response to LPS, in contrast to a platelet-activating factor antagonist [1-O-hexadecyl-2-acetyl- sn-glycero-3-phospho(N,N,N-trimethyl hexanolamine)] which inhibited both exoantigen- and LPS-induced production of TNF. We conclude that at least two different parts of the molecule are involved in the induction of TNF secretion by parasite exoantigens: one requires the presence of a phosphate bound to inositol, and, since dephosphorylated exoantigens were also inhibitory, one does not. It would seem that both affect interactions between parasite-derived exoantigens and the macrophage receptors.
Penicillin treatment accelerates middle ear inflammation in experimental pneumococcal otitis media
Authors: Kawana M , Kawana C , Giebink G S
Published: January 2020

Most Streptococcus pneumoniae strains are killed by very low concentrations of penicillin and other beta-lactam antibiotics, yet middle ear inflammation and effusion persist for days to weeks after treatment in most cases of pneumococcal otitis media. To study the effect of beta-lactam antibiotic treatment on pneumococci and the middle ear inflammatory response during pneumococcal otitis media, we measured concentrations of pneumococci, inflammatory cells, and lysozyme in middle ear fluid (MEF) by using the chinchilla model. Procaine penicillin G given intramuscularly 12 and 36 h after inoculation of pneumococci into the middle ear caused a significant acceleration in the MEF inflammatory cell concentration compared with that in untreated controls, with a significant peak in the inflammatory cell concentration 24 h after pneumococcal inoculation. The lysozyme concentration in MEF also increased more rapidly in treated than in control animals. Viable pneumococci were not detected in MEF after the second dose of penicillin, but the total pneumococcal cell concentration remained unchanged for at least 45 days. Therefore, penicillin treatment accelerated middle ear inflammation while killing pneumococci, but treatment did not accelerate clearance of the nonviable pneumococcal cells from MEF. Further studies will need to define the contribution of these responses to acute and chronic tissue injury.
Interleukin-3 induces antimicrobial activity against Leishmania amazonensis and Trypanosoma cruzi and tumoricidal activity in human peripheral blood-derived macrophages
Authors: Ho J L , Reed S G , Sobel J , Arruda S , He S H , Wick E A , Grabstein K H
Published: January 2020

The ability of interleukin-3 (IL-3) to induce antimicrobial and tumoricidal activity was evaluated. Macrophages infected with two intracellular protozoa, Leishmania amazonensis or Trypanosoma cruzi, were treated with cytokines. IL-3 induced a dose-dependent enhancement of microbistasis against leishmanias, and the activity of IL-3 (100 ng/ml) was comparable to that of gamma interferon (IFN-gamma) (1,000 U/ml). In addition, IL-3 in combination with either granulocyte-macrophage colony-stimulating factor (GM-CSF) or macrophage CSF (M-CSF) or with IFN-gamma reduced infection and lowered the required dose. IL-3 similarly activated macrophages to inhibit intracellular replication of T. cruzi. Furthermore, IL-3 induced antibody-independent tumoricidal activity against melanoma cells that was dose dependent and comparable to that of lipopolysaccharide and GM-CSF. The mechanisms by which IL-3 induced antimicrobial activity may involve at least the augmentation of oxidative capacity. IL-3, at concentrations of 0.5 ng/ml or greater, led to a significantly increased oxidative burst which paralleled the inhibition of protozoan replication. The enhancement of oxidative capacity by IL-3 (5 ng/ml or higher) was comparable to that of IFN-gamma. The induction of tumoricidal activity was associated with the production of tumor necrosis factor alpha (TNF-alpha), which in this system may feed back to enhance the macrophage inhibition of leishmanias, as demonstrated by neutralization of IL-3 activation by anti-TNF-alpha antibody. Thus, peripheral blood macrophages remain responsive to IL-3, as demonstrated by enhanced antimicrobial and tumoricidal activity. IL-3 may have potential clinical applications because of these properties and its effect on myelopoiesis.
Neonatal, urogenital isolates of biotype 4 nontypeable Haemophilus influenzae express a variant P6 outer membrane protein molecule
Authors: Murphy T F , Kirkham C , Sikkema D J
Published: January 2020

The P6 outer membrane protein is a highly conserved molecule which is present on the surface of all strains of Haemophilus influenzae. Sixty strains of nontypeable H. influenzae which caused invasive disease or colonized the female urogenital tract were studied with monoclonal antibodies 7F3 and 4G4, which recognize different surface-exposed epitopes on the P6 molecule. All 60 strains expressed the epitope recognized by 4G4, whereas 47 of 60 strains expressed the epitope recognized by antibody 7F3. The 7F3-nonreactive strains were all biotype 4 and were recovered from the blood of neonates or postpartum women or from the female urogenital tract. The P6 genes from two 7F3-nonreactive strains were cloned, and the nucleotide sequences were determined. Analysis of amino acid sequences, immunoassays with synthetic peptides, and site-directed mutation of the P6 gene indicate that the epitope recognized by antibody 7F3 is conformational and that the sequence Asp-Ile-Thr is critical in maintaining the conformation of the epitope. We conclude that the unusually virulent clone family of biotype 4 strains of nontypeable H. influenzae express a variant P6 molecule which has an alteration in a highly conserved surface-exposed epitope.
Maternal immunization with gametocyte antigens as a means of providing protective immunity against Eimeria maxima in chickens
Authors: Wallach M , Halabi A , Pillemer G , Sar-Shalom O , Mencher D , Gilad M , Bendheim U , Danforth H D , Augustine P C
Published: January 2020

In the present study, we wished to demonstrate the ability of surface gametocyte antigens to induce protective immunity against Eimeria maxima infections in chickens. In order to accomplish this goal, we employed maternal immunization as a means of providing large amounts of specific antibodies to offspring chicks. Upon challenge with sporulated E. maxima oocysts, chicks from hens immunized with affinity-purified gametocyte antigens showed greatly reduced oocyst production compared with chicks from sham-immunized hens. These results suggest that maternal immunization with gametocyte antigens can be used as a means to provide transmission-blocking immunity against E. maxima infections.