bluuebunny/crossref_metadata_2025 huggingface dataset:
Glycoproteins composed of major surface immunodeterminants of Pneumocystis carinii
Authors: Tanabe K , Takasaki S , Watanabe J , Kobata A , Egawa K , Nakamura Y
Published: January 2020

Pneumocystis carinii is a pathogen which causes fatal pneumonia in patients with acquired immune deficiency syndrome. To facilitate the basic study of P. carinii, we analyzed the major surface proteins by immunochemical and biochemical methods. The major protein components of both cysts (resting form) and trophozoites (vegetative form) are part of a group of proteins called P115 with apparent masses of 105 to 120 kilodaltons. They represent an unusually large portion of the total proteins of this organism. The purified proteins exhibited six isoelectric variants when analyzed by two-dimensional gel electrophoresis. A monoclonal antibody raised against cysts recognized all six variants and reacted with epitopes that were located in the cell wall, thereby indicating that P115 is an immunoreactive surface component. Data are presented that the isoelectric variants contain identical or closely related protein components and that they are mannose-rich glycoproteins. Deglycosylated P115 migrates primarily as a single more acidic protein in two-dimensional gels, suggesting that the isoelectric variants may be due primarily to differences in glycosylation. The majority of sera tested from humans with diagnosed pneumocystosis reacted strongly with the P115 proteins.
Mutations in the hemolytic-phospholipase C operon result in decreased virulence of Pseudomonas aeruginosa PAO1 grown under phosphate-limiting conditions
Authors: Ostroff R M , Wretlind B , Vasil M L
Published: January 2020

The phospholipase C (PLC) operon of Pseudomonas aeruginosa consists of plcS, which encodes a heat-labile secreted hemolysin, and two in-phase, overlapping genes, plcR1 and plcR2, which may encode Pi-regulatory genes. A 2.8-kilobase-pair deletion mutation in this operon was constructed, and a tetracycline resistance (Tcr) cartridge replaced the deleted sequences. A deletion mutant of strain PAO1 was obtained through recombination between the flanking regions of the mutated cloned PLC operon and the homologous chromosomal regions. The deletion of the chromosomal PLC operon and its replacement by the Tcr cartridge was confirmed by Southern hybridization. The deletion strain, PLC SR, is nonhemolytic. However, it retains PLC activity when measured on a synthetic substrate. A second mutant strain, PLC R, contains a deletion in the plcR genes. This mutant is more hemolytic and produces more enzymatic activity than PAO1. The virulence of both of these mutants was compared with that of PAO1 in the mouse burn model of infection. When mice were infected with cultures grown in a high-Pi medium, there was a 10-fold increase in the 50% lethal dose of the mutants compared with PAO1. In contrast, when the inoculum originated from low-Pi cultures, there was a 200- to 10,000-fold increase in the 50% lethal dose of the mutants over PAO1.
Isolation and immunological characterization of a 55-kilodalton surface protein from Salmonella typhimurium
Authors: Foulaki K , Gruber W , Schlecht S
Published: January 2020

Surface proteins of different Salmonella R mutants were labeled selectively by treating live bacteria with cycloheptaamylose-dansylchloride. The labeled proteins were extracted from the cells with 6 M urea and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. From the urea extract a 55-kilodalton protein common to numerous Salmonella strains could be isolated by ion-exchange chromatography and gel filtration free of lipopolysaccharide. Immunization of rabbits with isolated protein led to the formation of specific antibodies. Such antiprotein antisera could be employed in Western blots for the specific identification of the 55-kilodalton protein in bacterial extracts containing mixtures of different Salmonella proteins. The importance of this antigen is emphasized by antisera against acetone-killed Salmonella bacteria, showing a preferential interaction with the 55-kilodalton protein in Western blots. Active immunization of mice with the 55-kilodalton protein afforded significant protection against experimental infection with S. typhimurium.
Regulation of immunoglobulin E production in mice immunized with an extract of Toxoplasma gondii
Authors: Watanabe N , Kobayashi A
Published: January 2020

Repeated infection with Toxoplasma gondii could not induce immunoglobulin E (IgE) antibody production. When mice were injected intraperitoneally twice over a 3-week interval with an extract of tachyzoites of T. gondii and Al(OH)3 as adjuvant, antitoxoplasma IgE antibody was produced. Antitoxoplasma IgE antibody titers were low and diminished after a short time in B10.S, BALB/c, C3H, and C57BL/6 mice. This tendency was more evident in IgE low-responder SJL mice. IgE-inducing activity of Toxoplasma antigen was weaker than those of keyhole limpet hemocyanin, ovalbumin, and an extract from Nippostrongylus brasiliensis. The antitoxoplasma IgE antibody production was enhanced by and persisted after whole-body irradiation (150 R) following secondary immunization. The enhanced antitoxoplasma IgE antibody production was suppressed by transferring spleen cells from Toxoplasma antigen-immunized mice. The suppressive effect of the spleen cells was Toxoplasma antigen specific and was removed by treatment with anti-Thy-1.2 and complement. These results indicate that the low IgE production induced by Toxoplasma antigen is the result of irradiation-sensitive and antigen-specific suppressor T cells. These findings might explain the lack of IgE antibody response in mice with Toxoplasma infection.
Heterologous protection against invasive Escherichia coli K1 disease in newborn rats by maternal immunization with purified mannose-sensitive pili
Authors: Guerina N G , Woodson K , Hirshfeld D , Goldmann D A
Published: January 2020

Heterologous protection against Escherichia coli K1 bacteremia with antibody to purified mannose-sensitive (MS) pili was demonstrated in a neonatal rat model. The serological relatedness of purified MS pili from 17 E. coli K1 clinical isolates was examined by an enzyme-linked immunosorbent assay. Five pilus serogroups were identified, with the pili in each group showing 50% or greater cross-reactivity with the typing serum of the group. The MS pili from 12 of 17 (70%) strains belonged to just two serogroups. Pregnant Sprague-Dawley rats (dams) were immunized with purified pili, and their newborns (pups) were challenged with heterologous E. coli. Bacteremia was significantly reduced when the pili used for immunization were from the same serogroup as the pili expressed by the challenge bacteria. Thus, immunization with C94 pili and challenge with E03 (71% cross-reactivity) or E04 (50% cross-reactivity) resulted in bacteremia rates of 12 of 17 (17%) versus 51 of 79 (65%) in controls and 0 of 75 (0%) versus 28 of 70 (40%) in controls, respectively (P less than 0.001 for each comparison). With lower cross-reactivity, less protection was observed (P less than 0.05 for 22 to 37% pilus serological relatedness). No protection was seen in pups suckled by dams immunized with MS pili having only 5% serological relatedness to the pili on the challenge strain.
Immunodominant antigen of Actinobacillus actinomycetemcomitans Y4 in high-responder patients
Authors: Califano J V , Schenkein H A , Tew J G
Published: January 2020

This study was undertaken to look for characteristics of the immunodominant antigen(s) of Actinobacillus actinomycetemcomitans Y4 that might help explain the high antibody titers in periodontitis patients. Radioimmunoassays (RIA) were performed on sera from 481 patients; sera from the 32 patients with the highest anti-Y4 titers (above 128,000 RIA U/ml) were further analyzed. Y4 antigen was boiled for 45 min or treated with papain, and antibody responses were analyzed by RIA and Western blotting (immunoblotting). In addition, carbohydrate was purified from Y4 and examined by Western blotting. The results indicated that the immunodominant antigen of Y4 in high responders was stable after papain treatment or boiling for 45 min. Papain or boiling eliminated protein bands but a large diffuse band persisted on Western blots. With increasing dilutions of sera, bands on Western blots corresponding to protein antigens disappeared, while the large diffuse band resembling that of carbohydrate persisted. Partially purified Y4 carbohydrate contained the large diffuse band. Double-immunodiffusion analysis indicated that rabbit serotype b-specific antiserum and patient sera recognized the same antigen. When the carbohydrate extract was passed over a lipid A-binding column to remove lipopolysaccharide, the smear corresponding to the immunodominant antigen was still present on Western blots. The immunodominant antigen of Y4 in high-responder individuals appears to be a carbohydrate and is possibly the capsular polysaccharide.
Impact of protein malnutrition on exogenous reinfection with Mycobacterium tuberculosis
Authors: McMurray D N , Bartow R A , Mintzer C L
Published: January 2020

Malnutrition may be a predisposing host factor in the development of exogenous-reinfection tuberculosis. Outbred Hartley guinea pigs were given isocaloric diets containing either 30% ovalbumin (control animals) or 10% ovalbumin (low-protein-fed [LP] animals). Equal numbers of control and LP animals were assigned to one of three infection groups: (i) primary pulmonary infection with a low-virulence, streptomycin-resistant (LVsr) isolate of Mycobacterium tuberculosis and then reinfection 6 weeks later by the same route with a high-virulence (HV) isolate; (ii) only the primary infection (LVsr isolate); and (iii) only the secondary infection (HV isolate). Each infection resulted in the development of 4 to 12 pulmonary tubercles. Guinea pigs were skin tested with purified protein derivative and killed 6 weeks after the second infection. Protein deprivation suppressed the dermal responses to purified protein derivative in all infection groups. Primary infection of well-nourished animals with the LVsr isolate induced significant protection against infection with the HV isolate in the reinfected group, based upon the numbers of viable mycobacteria in the lung and spleen. Protein malnutrition did not exacerbate disease in the reinfected group beyond that observed in malnourished animals infected with the HV isolate only, but neither did the infection with the LVsr isolate protect the LP animals against reinfection with the HV isolate. We conclude that malnutrition interferes with the protection normally afforded by primary infection but does not result in more severe disease in reinfected individuals than would be observed in singly infected subjects.
In vivo immune responses to Candida albicans modified by treatment with recombinant murine gamma interferon
Authors: Garner R E , Kuruganti U , Czarniecki C W , Chiu H H , Domer J E
Published: January 2020

The immunologic effects of in vivo administration of recombinant murine gamma interferon (rMuIFN-gamma) were determined in a murine model of candidiasis. Naive mice were given graded doses of rMuIFN-gamma and then challenged intravenously with Candida albicans. Increased morbidity and mortality were noted in four different strains of mice, viz., BALB/c, A/J, Swiss Webster, and CBA/J, providing the mice had not been immunized with C. albicans before challenge. Quantitative culture of selected organs of Swiss Webster and CBA/J mice surviving treatment with rMuIFN-gamma revealed elevated numbers of C. albicans cells, particularly in the kidneys, but also in the liver, lungs, and spleen. The lungs, livers, and spleen of female CBA/J mice were more protected from increased multiplication of the fungus than were those of males of the same species or female Swiss Webster mice. On the basis of these initial findings, the effect of treatment with 5,000 U of rMuIFN-gamma on immune responses in a gastrointestinal model of candidiasis was determined. CBA/J mice that had been colonized with C. albicans as infants were boosted with a cutaneous inoculation of the fungus when 6 to 10 weeks old; development of delayed hypersensitivity (DH), antibodies, and protective responses was assayed at intervals thereafter. Daily treatment with rMuIFN-gamma (beginning 1 day before cutaneous inoculation) suppressed weak immune responses but had little effect on responses which were strong. For example, DH and anti-C. albicans antibody production were suppressed in animals colonized with C. albicans but not boosted by cutaneous inoculation, and DH was suppressed in uncolonized animals that had been inoculated once cutaneously with the fungus as well. There was no rMuIFN-gamma-induced suppressive effect of DH in mice which had been stimulated maximally with C. albicans, i.e., colonized animals that had been boosted cutaneously with the organisms. Collectively, these data indicate that naive mice or mice with minimal levels of anti-C. albicans sensitivity, females somewhat more so than males, were sensitive to suppressive effects of in vivo treatment with rMuIFN-gamma when challenged with C. albicans. In contrast, under conditions similar to those of humans, in whom underlying immunity to C. albicans is usually present, suppression of host responses to C. albicans was not observed in immunized mice in response to treatment with rMuIFN-gamma.
Vibrio cholerae expresses cell surface antigens during intestinal infection which are not expressed during in vitro culture
Authors: Jonson G , Svennerholm A M , Holmgren J
Published: January 2020

Vibrio cholerae O1 bacteria harvested directly from ligated or nonligated intestines of rabbits with experimental cholera expressed at least 7 to 8 novel, in vivo-specific cell envelope (env) proteins that were not found on vibrios after in vitro culture in various ordinary liquid media. At the same time, several of the env proteins ordinarily expressed in vitro had disappeared or become much reduced. The infection-induced novel env protein were immunogenic. In immunoblot analyses, antisera raised against in vivo-grown vibrios and then absorbed with in vitro-grown bacteria of the same strain specifically stained at least eight infection-induced antigens ranging from 62 to approximately 200 kilodaltons; absorption with washed in vivo-grown bacteria, on the other hand, removed the antibodies reacting with these antigens, indicating that the antigens were present on the bacterial cell surface. Conversely, antiserum against in vitro-grown bacteria reacted with several env antigens in in vitro-grown bacteria that were missing in the infection-derived vibrios. These adaptational changes were strikingly similar for different strains of cholera vibrios of both classical and El Tor biotypes. Most of the in vivo-specific proteins (with apparent molecular masses of approximately 200, approximately 150, approximately 140, 92, 68, 62, 43, and 29 kilodaltons) were not induced during cultivation of bacteria in iron-depleted medium and are probably not related to the iron-regulated env proteins known to be involved in iron transport systems.
Two-dimensional gel electrophoresis and immunoblotting of Campylobacter pylori proteins
Authors: Dunn B E , Perez-Perez G I , Blaser M J
Published: January 2020

Whole-cell, outer-membrane protein, flagellum-associated antigens and partially purified urease of Campylobacter pylori were analyzed by two-dimensional gel electrophoresis. C. pylori strains were readily distinguished from strains of Campylobacter jejuni, C. coli, and C. fetus by absence of major outer membrane proteins with Mrs of 41,000 to 45,000. C. pylori strains also lacked the acidic surface-array proteins at Mr 100,000 to 149,000 identified previously in serum-resistant strains of C. fetus. Surface labeling of intact C. pylori cells with 125I revealed two common major proteins, which we have designated protein 2 (pI 5.6 to 5.8, Mr 66,000) and protein 3 (pI 5.2 to 5.5, Mr 63,000). Proteins 2 and 3 were also the major components (subunits) observed in partially purified urease. Partially purified preparations of flagella consistently contained proteins 2 and 3. Thus, urease appears to be associated with both outer membranes and flagella of C. pylori. C. pylori strains also possessed an antigen at Mr 59,000 which was cross-reactive with antiserum against flagella of C. jejuni. However, the antigen did not appear to be associated with flagella per se in C. pylori. Protein 2 was unique to C. pylori among the Campylobacter species studied. It was not recognized by antibody against whole cells of C. jejuni or C. fetus or flagella of C. jejuni. Protein 3 was cross-reactive with antiserum against whole cells of C. jejuni and C. fetus, as were several other major protein antigens. Because protein 2 is a major outer membrane protein that is apparently unique to C. pylori, development of monospecific antibodies against this antigen may be useful for the identification of C. pylori in tissues, and purified antigen may be useful for serologic tests for specific diagnosis of C. pylori infections.
Effect of a purA mutation on efficacy of Salmonella live-vaccine vectors
Authors: Sigwart D F , Stocker B A , Clements J D
Published: January 2020

We made delta aroA, delta purA, and delta aroA delta purA derivatives of a strain of Salmonella dublin and isolated a nalidixate-resistant mutant of each construct. An inoculum of each of the nearly isogenic nalidixate-resistant auxotrophs was administered to BALB/c mice by gavage. The ability of each strain to colonize, invade, persist in tissues, and evoke serum and mucosal antibody responses to the lipopolysaccharide of the parent strain was examined. Only the delta aroA strain colonized, invaded, persisted, and (more importantly) evoked sustained significant serum and mucosal antibody responses. Neither the delta purA nor the delta aroA delta purA strain showed any of these abilities. These observations demonstrate that the purA defect, which causes a requirement for adenine, reduces the live-vaccine efficacy of attenuated Salmonella strains and may limit the effectiveness of Salmonella strains as carriers of heterologous antigens. These findings may be important in the selection of attenuated S. typhi strains for use in humans either as antityphoid live vaccines or as vectors for antigens of other pathogens.
Intestinal intraepithelial and splenic natural killer cell responses to eimerian infections in inbred chickens
Authors: Lillehoj H S
Published: January 2020

Splenic and intestinal natural killer (NK) cell responses were assessed in chickens inoculated with Eimeria parasites. The NK cell activities of both splenic and intestinal intraepithelial lymphocytes (IELs) decreased to a subnormal level during the early phase of eimerian infections but returned to normal or slightly higher than normal levels at about 1 week after the primary inoculation. Lymphocytes obtained from the lamina propria did not show any detectable level of NK cell activity during or following eimerian infections. Significant increases in splenic and intestinal IEL NK cell activities were seen during the early phase of secondary infection. The increase in the IEL NK cell activity that was seen shortly following secondary eimerian infection was accompanied by a substantial increase in the number of IELs expressing the asialo-GM1 antigen. Host strain differences in both splenic and IEL NK cell responses were detected following primary eimerian infections. These results suggest that both splenic and intestinal IEL NK cells may play an important role in the host defense against intestinal protozoan infections.
Molecular and immunochemical characterization of recombinant Escherichia coli containing the spaA gene region of Streptococcus sobrinus
Authors: Ogundipe J O , Holt R G
Published: January 2020

We identified and characterized a recombinant Escherichia coli containing the entire gene for surface protein antigen A (spaA) of Streptococcus sobrinus 6715. The recombinant E. coli was isolated from a cosmid gene bank of size-fractionated S. sobrinus DNA fragments, and recombinants expressing the SpaA protein were detected immunologically. Subcloning experiments showed that the DNA sequences encoding the SpaA protein could be isolated on two contiguous EcoRI fragments, 3.7 and 3.3 kilobases (kb) in size, both contained on a 16.2-kb BglII fragment. Southern blot hybridization experiments using the EcoRI fragments to probe genomic DNAs from various serotypes of the mutans group of streptococci revealed DNA sequence homology not only to S. sobrinus 6715 (serotype g) chromosomal DNA but also to S. sobrinus serotype d DNA. Weak hybridization signals to Streptococcus mutans serotypes c, e, and f and to Streptococcus cricetus serotype a were observed with the 3.3-kb EcoRI fragment. These results suggest that the coding sequence for the spaA gene is probably conserved in S. sobrinus strains. Plasmid-encoded polypeptides made in E. coli minicells revealed that transcription of the spaA gene was initiated on the 3.7-kb EcoRI fragment and that its product size was about 210 kilodaltons. The cloned SpaA protein was purified from the periplasmic protein of E. coli, and monospecific antiserum against the cloned product was prepared in rabbits. The data obtained from various physiochemical and immunological procedures allowed us to conclude that the coding sequence for the entire spaA gene of S. sobrinus 6715 had been successfully cloned in E. coli and that faithful expression of the cloned product could be obtained.
Optimal induction of tumor necrosis factor production in human monocytes requires complete S-form lipopolysaccharide
Authors: Männel D N , Falk W
Published: January 2020

Optimal activation of human monocytes in vitro for the biosynthesis of tumor necrosis factor was achieved only with complete S-form lipopolysaccharide. Endotoxin preparations with shorter carbohydrate chains or the lipid A component of lipopolysaccharide were not able to induce release of comparable amounts of tumor necrosis factor by monocytes under the conditions described. The same differences in the level of tumor necrosis factor mRNA were observed. Moreover, addition of these agents to appropriate monocyte-activating substances inhibited the production of tumor necrosis factor. The regulatory implications of this phenomenon are discussed.
Modulation of macrophage Ia expression by lipopolysaccharide: stem cell requirements, accessory lymphocyte involvement, and IA-inducing factor production
Authors: Wentworth P A , Ziegler H K
Published: January 2020

The mechanism of induction of murine macrophage Ia expression by lipopolysaccharide (LPS) was studied. Intraperitoneal injection of 1 microgram of LPS resulted in a 3- to 10-fold increase in the number of IA-positive peritoneal macrophages (flow cytometry and immunofluorescence and a 6-to 16-fold increase by radioimmunoassay. The isolated lipid A moiety of LPS was a potent inducer of macrophage Ia expression. Ia induction required a functional myelopoietic system as indicated by the finding that the response to LPS was eliminated in irradiated (900 rads) mice and reinstated by reconstitution with bone marrow cells. Comparison of LPS-induced Ia expression in normal and LPS-primed mice revealed a faster secondary response to LPS. The memory response could be adoptively transferred to normal mice with nonadherent spleen cells prepared 60 days after LPS injection. Spleen cells prepared 5 days after LPS injection caused Ia induction in LPS-nonresponder mice; such induction was not observed in irradiated (900 rads) recipients. The cell responsible for this phenomenon was identified as a Thy-1+, immunoglobulin-negative nonadherent cell. The biosynthesis and expression of Ia were not increased by direct exposure of macrophages to LPS in vitro. Small amounts of LPS inhibited Ia induction by gamma interferon. LPS showed positive regulatory effects on Ia expression by delaying the loss of Ia expression on cultured macrophages and by stimulating the production of Ia-inducing factors. Supernatants from cultured spleen cells stimulated with LPS in vitro contained antiviral and Ia-inducing activity that was acid labile, indicating that the active factor is gamma interferon. We conclude that induction of Ia expression by LPS in vivo is a bone-marrow-dependent, radiation-sensitive process which involves the stimulation of a gamma interferon-producing accessory lymphocyte and a delay in Ia turnover.
Specific immunological unresponsiveness to bacterial lipopolysaccharides develops in a cyclic manner
Authors: Elkins K L , Stashak P W , Baker P J
Published: January 2020

Prior exposure (priming) of BALB/cByJ mice to a low dose of lipopolysaccharide derived from Escherichia coli 055 or Serratia marcescens, followed by immunization with an optimally immunogenic dose of the same lipopolysaccharide 2 to 30 days later, results in the expression of substantially reduced antibody responses. Such unresponsiveness, which is antigen specific, occurs in a cyclic manner with time after priming.
Effects of immune colostrum and orally administered antisporozoite monoclonal antibodies on the outcome of Cryptosporidium parvum infections in neonatal mice
Authors: Arrowood M J , Mead J R , Mahrt J L , Sterling C R
Published: January 2020

A neonatal BALB/c mouse model of cryptosporidiosis was used to examine the potential passive transfer of immunity via immune colostrum and oral treatment with anticryptosporidial monoclonal antibodies. Neonates suckled by dams that recovered from Cryptosporidium parvum infections were equally susceptible to infection as their control counterparts suckled by naive dams. Parasite loads among the control and immune colostrum-fed mice were indistinguishable. Neonates receiving orally administered antisporozoite monoclonal antibodies were equally susceptible to infections compared with the control and immune colostrum-fed mice. Parasite loads among the mice receiving daily oral treatment with monoclonal antibody mixtures exhibited significantly lower parasite loads compared with the control mice (P less than 0.05).
Humoral immune response to chlamydial genital infection of mice with the agent of mouse pneumonitis
Authors: Ramsey K H , Newhall W J , Rank R G
Published: January 2020

The objective of this study was to characterize the humoral immune response to chlamydial genital infection of mice with the mouse pneumonitis agent (MoPn). With an enzyme-linked immunoabsorbent assay, immunoglobulin G antibodies to MoPn were first detected in plasma by day 14. Peak plasma antibody concentrations were reached by day 49, and this response did not decline significantly throughout the 300-day monitoring period. Immunoglobulin A against MoPn could first be detected in pooled vaginal washes by day 21 after infection and had reached peak concentrations by day 28, but anti-MoPn immunoglobulin G was not consistently present in secretions. The antibody response in secretions had declined slightly by day 300. Immunoblot analysis revealed that the early phase of the plasma antibody response to MoPn as a result of genital infection was against lipopolysaccharide, the major outer membrane protein, and a 62-kilodalton (kDa) protein. In secretions, early-phase immunoglobulin A antibodies were directed to the major outer membrane protein and lipopolysaccharide. Late reactions to 15-, 22-, and 83-kDa proteins in plasma were noted. Late reactions to the 62-kDa protein in secretions were also noted. The cause of these late responses remains unexplained. When mice were challenged intravaginally with MoPn at 50-day intervals after the primary infection, it was found that mice inoculated on day 100 or after were susceptible to reinfection. Susceptibility could not be related to a decline in the antibody concentration in plasma or secretions or in the antibody response to specific components of MoPn as measured by immunoblot analysis.
Trehalose dimycolate enhances resistance to infection in neutropenic animals
Authors: Madonna G S , Ledney G D , Elliott T B , Brook I , Ulrich J T , Myers K R , Patchen M L , Walker R I
Published: January 2020

Bacterial infections are lethal complications of neutropenia, and antibiotics alone are inadequate therapy for these infections. Irradiated mice become severely neutropenic and remain susceptible to infection for 2 to 3 weeks, depending on the dose and quality of radiation. Some bacterial cell wall derivatives stimulate nonspecific host defense mechanisms against a variety of microbes which might cause postirradiation infection. In this study we determined if the cell wall glycolipid trehalose dimycolate (TDM), derived from Mycobacterium phlei, or a synthetic preparation of TDM was able to (i) enhance survival in mice when given before or after lethal doses of 60Co radiation and (ii) increase nonspecific resistance to postirradiation infection. Treatment with TDM oil-in-water emulsions and with synthetic TDM significantly enhanced survival before and after lethal doses of 60Co irradiation. This result correlated with the ability of TDM to reduce the translocation of intestinal bacteria and to stimulate hematopoiesis. With respect to nonspecific resistance to infection, TDM injected 1 h after sublethal irradiation increased resistance to a lethal Klebsiella pneumoniae challenge (10 50% lethal doses of K. pneumoniae in 30 days [LD50/30]) 4 or 14 days later. Increasing the dose of K. pneumoniae to 5,000 LD50/30 on day 4 overwhelmed the ability of TDM-treated mice to overcome infection. However, TDM treatment 1 h postirradiation combined with ceftriaxone antibiotic therapy (days 5 through 14) enhanced survival, even when the higher dose of bacteria (5,000 LD50/30) was used. These results indicate that in irradiated mice, TDM can be used to enhance survival and, as a potent stimulant of nonspecific resistance to infection in neutropenic mice, can act synergistically with antibiotic therapy to reduce sepsis and mortality.
Autoimmunity in Chagas' disease: specific inhibition of reactivity of CD4+ T cells against myosin in mice chronically infected with Trypanosoma cruzi
Authors: Rizzo L V , Cunha-Neto E , Teixeira A R
Published: January 2020

In this study, we assessed the proliferative response of T cells from mice chronically infected with Trypanosoma cruzi to actin, myosin, or T. cruzi soluble antigen (SA). We report here that CD4+ T cells from mice chronically infected with T. cruzi proliferated in response to myosin but not to actin, whereas cells from naive mice did not proliferate against any of the antigens tested. Antisera raised against myosin- or SA-activated T cells specifically inhibited respectively, the myosin or SA in vitro proliferative response, whereas the response to unrelated antigen remained unimpaired. Sera from chronically infected mice failed to show any significant inhibitory activity. The above findings suggest that autoreactive and T. cruzi-reactive T cells belong to different, perhaps nonoverlapping, compartments of the immune cell repertoire of mice chronically infected with T. cruzi. The failure of infected mice to trigger the suppressive mechanisms described here might be the primary immune defect leading to breakdown of self-tolerance and unopposed, perhaps tissue-damaging, autoimmunity in experimental Chagas' disease.
Difference between bacterial and food antigens in mucosal immunogenicity
Authors: Wold A E , Dahlgren U I , Hanson L A , Mattsby-Baltzer I , Midvetdt T
Published: January 2020

The mucosa-associated lymphoid tissue may deviate from its systemic counterpart in being able to discriminate between microbial and nonmicrobial antigens. To study this, the systemic and mucosal antibody responses to bacterial and food antigens were followed in parallel in female rats during two pregnancies and lactation periods. Germfree rats were monocolonized with an Escherichia coli O6K13H1 strain, and their diet was switched to pellets containing large amounts of ovalbumin and beta-lactoglobulin. Antibodies against O6 lipopolysaccharide already appeared in serum and bile 1 week after colonization, and those against type 1 fimbriae appeared a few weeks later. Serum immunoglobulin G antibodies against the E. coli enzyme beta-galactosidase were found in moderate titers in all rats after 16 weeks of exposure. In contrast, few rats had detectable antibody levels against the dietary proteins ovalbumin and beta-lactoglobulin in serum or bile even after 16 weeks of exposure. In the milk, antibodies against E. coli beta-galactosidase and type 1 fimbriae reached the highest titers, while moderate titers were found against the food antigens and against O6 lipopolysaccharide. The difference in immune reactivity against bacterial versus dietary antigens was not likely due to insufficient amounts of the latter reaching lymphoid tissue, since (i) uptake studies indicated that ovalbumin was more efficiently taken up than endotoxin and (ii) the same difference in antigenicity between ovalbumin and E. coli was seen after immunization directly into Peyer's patches. We therefore suggest that a prerequisite for a strong mucosal antibody response is that the antigen be encountered by the gut-associated lymphoid tissue within microorganisms capable of stimulating antigen presentation.
Cloning and nucleotide sequence of the gene (trh) encoding the hemolysin related to the thermostable direct hemolysin of Vibrio parahaemolyticus
Authors: Nishibuchi M , Taniguchi T , Misawa T , Khaeomanee-Iam V , Honda T , Miwatani T
Published: January 2020

Vibrio parahaemolyticus isolates derived from an outbreak of gastroenteritis in the Republic of Maldives did not have the genetic potential to produce the thermostable direct hemolysin, but one such isolate produced a hemolysin immunologically related to the thermostable direct hemolysin (T. Honda, Y. Ni, and T. Miwatani, Infect. Immun. 56:61-965, 1988). The Maldives isolates hybridized with the DNA probe for the gene encoding the thermostable direct hemolysin (the tdh gene) under reduced stringencies. A DNA fragment containing the probe-reactive nucleotide sequence was isolated from a selected strain and cloned into pBR322 in Escherichia coli. A clone producing the thermostable direct hemolysin-related hemolysin was obtained by screening with hemolysis assays and by an immunological assay. Nucleotide sequence analysis of the cloned DNA fragment revealed that the gene encoding the thermostable direct hemolysin-related hemolysin (the trh gene), like the tdh gene, encoded the hemolysin subunit composed of 189 amino acid residues. The trh gene had significant nucleotide sequence homology with the tdh gene (68.4% with the tdh1 gene copy and 68.6% with the tdh2 gene copy). The amino acid sequences of the hemolysin subunits deduced from the nucleotide sequences of the trh gene and tdh gene were homologous (61.9% homology with the tdh1-encoded subunit and 63.0% homology with the tdh2-encoded subunit) and contained the two cysteine residues to form an intrachain bond at the same positions, and their possible conformations appeared to be similar as determined by hydrophobicity-hydrophilicity analysis and a secondary structure prediction. The trh and tdh genes may have had a common ancestor and may have evolved by single-base changes so that they maintained the fundamental architecture of the molecules.
Effects of continuously administered murine interleukin-1 alpha: tolerance development and granuloma formation
Authors: Otterness I G , Golden H W , Brissette W H , Seymour P A , Daumy G O
Published: January 2020

Continuous infusion of murine recombinant interleukin-1 alpha (rIL-1 alpha) into rats by using intraperitoneally implanted osmotic pumps led to marked decreases in body weight, liver enzymes (serum glutamic oxalacetic transaminase, serum glutamic pyruvic transaminase, and sorbitol dehydrogenase), appetite, and mobility and increases in drinking, blood urea nitrogen, and total peripheral blood leukocytes within 3 days. Granuloma formation was found in the local area of rIL-1 alpha release. As early as day 3, a focal infiltrate of polymorphonuclear leukocytes, mononuclear leukocytes, and plasma cells filled the area; by day 6, extensive fibrosis was found. A loss of rIL-1 alpha-induced changes, with the exception of granuloma formation, occurred by day 10. A marked decrease in the response to rIL-1 alpha was also observed when animals were challenged by implantation of new pumps containing rIL-1 alpha, with monitoring of body weight, or by subcutaneous injection of rIL-1 alpha, with monitoring of serum colony-stimulating factor production. We propose that, even in the continuous presence of interleukin-1, replacement of the acute responses to interleukin-1 by restoration of more normal physiology may be advantageous upon acquisition of specific immunity.
Effects of gamma interferon on syntheses of DNA and proteins by Entamoeba histolytica
Authors: Castellanos C , Ramos C , Ortiz-Ortiz L
Published: January 2020

To define the participation of cell-mediated immunity in resistance to amebic infection through the action of soluble mediators or lymphokines (LKs), including gamma interferon (IFN-gamma), we studied their effect on Entamoeba histolytica. Supernatants from cultures of lymphoid cells, which had been stimulated in vitro with concanavalin A and were rich in lymphokines (LRSNs), and recombinant IFN-gamma were used. LRSN and recombinant IFN-gamma inhibited the growth of E. histolytica trophozoites in vitro. These LKs did not show a cytotoxic effect on the ameba, but they did inhibit rather significantly protein and DNA syntheses of the protozoa. Interestingly, LRSN incubated at 4 degrees C in the presence of trophozoites lost the ability to inhibit the replication of vesicular stomatitis virus. IFN-gamma inactivated at pH 2 had no effect on DNA synthesis by the ameba, thus suggesting that IFN-gamma is responsible for the observed inhibition of parasite growth. Furthermore, the IFN-gamma inhibitory effect was abolished by a monoclonal antibody specific for this LK. The results suggest that IFN-gamma may participate in protection against amebiasis infection through the activity of mediators released by lymphocytes during infection.
Hybridization of Escherichia coli producing Shiga-like toxin I, Shiga-like toxin II, and a variant of Shiga-like toxin II with synthetic oligonucleotide probes
Authors: Brown J E , Sethabutr O , Jackson M P , Lolekha S , Echeverria P
Published: January 2020

Synthetic oligonucleotides, constructed from the nucleotide sequences of genes coding for the A subunit of Shiga-like toxin (SLT) I and the B subunit of SLT-II, were used as probes at different degrees of stringency to identify Escherichia coli producing different types of SLTs. At 45 degrees C, the A-I oligonucleotide probe hybridized with E. coli producing SLT-I, SLT-II, and variant of SLT-II (SLT-IIv). At 53 degrees C, only SLT-I-producing E. coli hybridized with this probe. At 45 degrees C, the B-II oligonucleotide probe hybridized with SLT-II- and SLT-IIv-producing E. coli. At 53 degrees C, this probe hybridized with only SLT-II-producing E. coli. The A-I and B-II oligonucleotide probes were subsequently tested for hybridization with 73 SLT-producing E. coli and 49 non-SLT-producing E. coli isolated in Asia and Canada. At 45 degrees C, the A-I oligomer had a sensitivity of 97% and a specificity of 100% in identifying SLT-producing E. coli. At 53 degrees C, the A-I oligonucleotide probe had a sensitivity of 92% and a specificity of 91% in identifying E. coli containing genes encoding SLT-I. At 45 degrees C, the B-II oligonucleotide had a 100% sensitivity and 97% specificity in identifying E. coli that hybridized with the SLT-II probe. Of 17 E. coli that hybridized only with the SLT-II probe, 10 did not hybridize with the B-II oligonucleotide at 53 degrees C. All 10 isolates were cytotoxic to Vero cells but not to HeLa cells, confirming that the B-II oligonucleotide probe used at 53 degrees C will differentiate isolates producing SLT-II and SLT-IIv.
Hemagglutinating properties of Shigella dysenteriae type 1 and other Shigella species
Authors: Qadri F , Haq S , Ciznár I
Published: January 2020

Strains of Shigella dysenteriae type 1 cultured in Casamino Acids-yeast extract broth medium in the presence of 1 mM calcium chloride at 37 degrees C for 22 h induced hemagglutination of erythrocytes that was inhibited by N-acetylneuraminic acid, N-acetylneuramin-lactose, and alpha 1-glycoprotein. The hemagglutination was heat labile, and the absence of cell-surface appendages suggested a nonfimbrial adhesin(s). Under the same conditions, strains of Shigella flexneri (types 1a, 1b, 2a, and 2b) showed N-acetylneuraminic acid-resistant hemagglutination of erythrocytes.
Cellular coaggregation of oral Streptococcus milleri with actinomyces
Authors: Eifuku H , Yakushiji T , Mizuno J , Kudo N , Inoue M
Published: January 2020

Oral isolates of Streptococcus milleri were examined for their ability to coaggregate with actinomyces. Of the 68 S. milleri strains tested, including 3 reference strains, 40 strains coaggregated with Actinomyces naeslundii WVU45 (actinomyces coaggregation group B) and 36 strains coaggregated with Actinomyces viscosus T14V (actinomyces coaggregation group A). All S. milleri strains of serotypes b (4 strains), e (2 strains), and f (24 strains) coaggregated with both of the actinomyces. The coaggregation reactions between the S. milleri cells and A. naeslundii WVU45 cells were optimal at about pH 7.0 and were Ca2+ or Mg2+ dependent, but they were not inhibited by the presence of simple sugars or amino sugars, including lactose (up to 0.5 M). Treatment of the S. milleri cells with heat (100 degrees C, 3 min) or proteases (trypsin, 1.0 mg/ml; pronase, 0.25 mg/ml; 37 degrees C; 3 h) and of the actinomyces cells with periodate (0.01 M, 4 degrees C, 16 h) destroyed their coaggregating abilities. The coaggregations between cells of the S. milleri strains, we well as cells of the Streptococcus sanguis H1 (reference strain for streptococcus coaggregation group 2) and the actinomyces strains (WVU45 and T14V), were inhibited by AFH1 (a carbohydrate receptor on T14V cells for a lectin on H1 cells). These interactions were also inhibited by anti-AFH1 immunoglobulin G (IgG) and by anti-b, anti-e, and anti-f S. milleri IgG or anti-f IgG Fab fragments. These results suggest that S. milleri, at least strains of serotypes b, e, and f, belongs to streptococcus coaggregation group 2.
The 75-kilodalton protein of Chlamydia trachomatis: a member of the heat shock protein 70 family?
Authors: Danilition S L , Maclean I W , Peeling R , Winston S , Brunham R C
Published: January 2020

The gene encoding a 75-kilodalton (kDa) protein of Chlamydia trachomatis was cloned, expressed, and sequenced. Genomic libraries from C. trachomatis serovar D DNA were constructed in vectors pUC18 and lambda gt11 and were screened with a panel of monoclonal antibodies against C. trachomatis antigens. The only recombinants identified were those that reacted with antibody UM-13, which has specificity for a genus-specific epitope on the 75-kDa protein. The gene was localized to a 2.9-kilobase DNA fragment and sequenced. The gene consists of a long open reading frame of 1,956 nucleotides, which translates into 652 amino acids totalling 70,558 daltons in mass. Putative promoter elements and a ribosome binding site were identified within 5'-flanking sequences, and a typical rho-independent terminator was identified within 3'-flanking sequences. Screening of the GenBank nucleic acid sequence data bank revealed extensive similarity between the chlamydial 75-kDa gene and the heat shock protein 70 (hsp70) family or proteins. In particular, 71 and 69% amino acid sequence similarities were identified with hsp70 of Escherichia coli and Bacillus megaterium, respectively. Polyclonal antibodies were produced to the recombinant antigen in rabbits and detected epitopes on elementary bodies in enzyme-linked immunosorbent and indirect microimmunofluorescence assays. Antibodies reacted with an antigen of identical molecular mass in L2 and C serovars in an immunoblot assay and neutralized these serovars in cell culture. The 75-kDa protein appears to be a chlamydial homolog of hsp70, is immunoaccessible on native elementary bodies, and is a target for neutralization.
Identification of outer oocyst wall proteins of three Cryptosporidium (Apicomplexa: Cryptosporidiidae) species by 125I surface labeling
Authors: Tilley M , Upton S J , Blagburn B L , Anderson B C
Published: January 2020

Autoradiography of oocyst wall surface proteins of three Cryptosporidium spp. revealed common bands at 285 to 290, 145 to 148, 120, 57, and 32 kilodaltons (kDa). Cryptosporidium baileyi and C. muris share proteins at 180, 100, 80 to 81, 29, and 18 to 19 kDa; C. baileyi and C. parvum share one protein at 46 to 47 kDa; and C. muris and C. parvum share a protein at 67 to 69 kDa. Additional protein bands, each unique to one species, were also observed.
Pili of a Vibrio parahaemolyticus strain as a possible colonization factor
Authors: Nakasone N , Iwanaga M
Published: January 2020

Pili of Vibrio parahaemolyticus were purified from a Kanagawa phenomenon-positive strain (Ha7) that belongs to serogroup O2:K3 and is adhesive to rabbit intestine. The organisms treated with the Fab fraction of antipilus antibody failed to adhere to the intestine. Purified pili had the ability to adhere to the intestine, but the pretreatment of the intestine with purified pili did not allow adherence of the organisms to the intestine. These results suggest that pili of this V. parahaemolyticus strain play an important role in colonization.
Phorbol myristate acetate inhibits HeLa 229 invasion by Bordetella pertussis and other invasive bacterial pathogens
Authors: Ewanowich C A , Peppler M S
Published: January 2020

The microfilament inhibitors cytochalasins B and D have been traditionally used to indirectly evaluate the requirement for actin in the uptake of invasive bacterial pathogens by nonprofessional phagocytes. Through their effects on microfilaments, both cytochalasins also impart profound alterations in cellular morphology and surface topology, which likely interfere with adherence. Alterations affecting adherence would complicate interpretation of the effect of cytochalasins on entry alone. As an alternative to cytochalasins, the effect of the tumor promoter phorbol myristate acetate (PMA) was examined for its effects on uptake of several invasive bacterial pathogens by HeLa 229 cells. In this communication, PMA was shown to induce a similar change in HeLa cell actin distribution, but, in contrast to cytochalasins B and D, PMA had no significant effect on gross cell morphology. The modified actin distribution was shown to reduce internalization of Bordetella pertussis, Yersinia pseudotuberculosis, Shigella flexneri, and Salmonella hadar in a dose-dependent manner at concentrations ranging from 1 to 1,000 ng/ml. The magnitude of reduction at a PMA concentration of 1,000 ng/ml was greater than the reduction elicited by cytochalasin B at 2.5 micrograms/ml but was less than that elicited by cytochalasin D at 2.5 micrograms/ml. Mezerein, a functional analog of PMA, caused a similar dose-dependent reduction in uptake of B. pertussis, whereas an inactive analog of PMA, alpha-4-phorbol-12,13-didecanoate was without effect on invasion. Binding studies further reveal that pretreatment of HeLa cells with PMA or mezerein did not significantly impair the ability of B. pertussis to adhere, in contrast to cytochalasins B and D, which caused a marked reduction in adherence.