bluuebunny/crossref_metadata_2025 huggingface dataset:
Relative functional activity of purified human immunoglobulin G against a type III group B streptococcal strain
Authors: Kim K S , Kang J H , Concepcion N F , Anthony B F
Published: January 2020

Human immunoglobulin G (IgG) separated from whole serum by a quaternary aminoethyl-Sephadex A-50 ion exchanger was evaluated for its activity against a type III group B streptococcal strain in the newborn rat model. Separated IgG yielded approximately 70 to 80% of whole serum IgG and did not contain detectable IgM or IgA. This IgG preparation also contained similar ratios of specific type III group B streptococcal antibody to total IgG in comparison with whole serum. In vivo, 50% protection from death was achieved by 3.9 ng of type III-specific antibody per 10 g of rat body weight. This value was considerably lower than 50% protective doses obtained in our previous studies with different human IgG preparations. Further studies are needed to understand the mechanisms responsible for these differences in the functional activity of IgG antibody.
Comparison of in vivo and in vitro multiplication rates of Haemophilus influenzae type b
Authors: Rubin L G
Published: January 2020

A model of invasive Haemophilus influenzae type b disease in rats was used to compare the net in vivo and in vitro multiplication rates of this bacterium. In both the blood of rats and in vitro cultures (fresh rat blood or enriched broth) there was an exponential increase in the number of CFU per milliliter with calculated net mean generation times as follows: sham-operated rats, 82 +/- 39 min (n = 5); asplenic rats, 34 +/- 5 min (n = 13); broth, 26 +/- 4 min (n = 12); rat blood, 24 +/- 1 min (n = 14). Thus, in vivo multiplication of H. influenzae type b can be extremely efficient.
Monoclonal antibodies specific for Shigella flexneri lipopolysaccharides: clones binding to type I and type III:6,7,8 antigens, group 6 antigen, and a core epitope
Authors: Carlin N I , Lindberg A A
Published: January 2020

Monoclonal antibodies against the Shigella flexneri lipopolysaccharide (LPS) were generated in two fusions by using the myeloma cell line Sp2/0 as a fusion partner with spleen cells from BALB/c mice immunized with S. flexneri serotypes 1b and 3a bacteria. The antibodies were characterized by immunoblotting, enzyme-linked immunosorbent assay (ELISA), hemagglutination, and coagglutination. Four different types of monoclonal antibodies were isolated: antibodies specific for the core antigen of the LPS, antibodies specific for the type I O antigen, antibodies specific for the group 6 O antigen, and antibodies specific for the type III:6,7,8 O antigen. The core-specific antibodies were shown to be specific for the Escherichia coli R3 core, which all S. flexneri LPSs tested, except for S. flexneri serotype 6 LPS, have. The type I O antigen-specific antibodies were shown to bind exclusively to S. flexneri serotypes 1a and 1b in ELISA. The type III:6,7,8 O-antigen-specific antibodies were specific for S. flexneri serotype 3a in ELISA and hemagglutination. Two different group 6 O-antigen-specific antibodies were bound. One was bound in both ELISA and hemagglutination to LPSs of S. flexneri serotypes 1b, 3a, 3b, and 4b, whereas the second was bound only to LPSs of serotypes 3a, 3b, and 4b in ELISA but to LPSs of all four serotypes in hemagglutination. The specificity of the isolated I, III:6,7,8, and group 6 monoclonal antibodies was verified by coagglutination of 363 S. flexneri clinical isolates.
Killing of Actinobacillus actinomycetemcomitans by the human neutrophil myeloperoxidase-hydrogen peroxide-chloride system
Authors: Miyasaki K T , Wilson M E , Genco R J
Published: January 2020

Actinobacillus actinomycetemcomitans is a facultative gram-negative coccobacillus associated with periodontal disease and nonoral infections. This organism is resistant to serum bactericidal mechanisms but is nevertheless killed by human neutrophils under aerobic and anaerobic conditions. Most of the killing attributable to oxidative mechanisms is inhibited by sodium cyanide, which suggests that the myeloperoxidase-hydrogen peroxide-chloride (MPO-H2O2-Cl-) system may be a key factor in the oxidative killing process. In this report, we examine whether the isolated MPO-H2O2-Cl- system is bactericidal against A. actinomycetemcomitans. We found that three major chromatographic forms of MPO were capable of killing A. actinomycetemcomitans at sublethal concentrations of H2O2 and that both catalase-positive and catalase-negative strains of this organism were sensitive to killing by the MPO-H2O2-Cl- system. We conclude that the isolated MPO-H2O2-Cl- system is bactericidal for A. actinomycetemcomitans independent of other neutrophil granule constituents and may be an important component of the oxygen-dependent bactericidal activity of the neutrophil with respect to this periodontopathic organism.
Serum immune response to Shigella protein antigens in rhesus monkeys and humans infected with Shigella spp
Authors: Oaks E V , Hale T L , Formal S B
Published: January 2020

The serum antibody response to proteins encoded by the virulence-associated plasmid of Shigella flexneri was determined in monkeys challenged with virulent S. flexneri serotype 2a. With water-extractable antigen in an enzyme-linked immunosorbent assay, a significant increase in antibody titer against proteins from a plasmid-carrying, virulent strain of S. flexneri serotype 5 could be demonstrated in convalescent sera. There were minimal antibody titers against proteins from an avirulent (plasmid-free) organism. Previously identified plasmid-coded polypeptides a, b, c, and d were predominant antigens recognized by a majority of the convalescent sera in immunoblots. An additional 140-megadalton plasmid-coded polypeptide was also recognized by half of the sera. Convalescent serum from an infected monkey recognized antigens on the bacterial surface in several different plasmid-containing Shigella species and in an enteroinvasive Escherichia coli strain. A survey of sera obtained from children 5 to 10 years of age who had been infected with S. flexneri or S. sonnei revealed high enzyme-linked immunosorbent assay titers in both acute and convalescent sera against a water extract from a virulent Shigella strain. In contrast, children under 3 years of age had no antibody titer in either acute or convalescent sera against the virulence-associated shigella proteins, while 3- to 4-year-old children mounted an immune response against these proteins only in convalescence.
Acid tolerance, proton permeabilities, and membrane ATPases of oral streptococci
Authors: Bender G R , Sutton S V , Marquis R E
Published: January 2020

Differences in acid tolerance among representative oral streptococci were found to be related more closely to the dynamic permeabilities of the bacteria to protons than to differences in the sensitivities of cell membranes to gross damage caused by environmental acidification. For Streptococcus mutans GS-5, Streptococcus sanguis NCTC 10904, and Streptococcus salivarius ATCC 13419, gross membrane damage, indicated by the release of magnesium from whole cells, occurred at pH values below about 4 and was rapid and extensive at pH values of about 3 or less. A more aciduric, lactic acid bacterium, Lactobacillus casei ATCC 4646, was more resistant to environmental acidification, and gross membrane damage was evident only at pH values below 3. Assessments of the movements of protons into S. mutans cells after an acid pulse at various pH values indicated that permeability to protons was minimal at a pH value of about 5, at which the average half time for pH equilibration across the cell membrane was about 12 min. The corresponding values for the less aciduric organism S. sanguis were pH 7 and 8.2 min, and the values for the intermediate organism S. salivarius were pH 6 and 6.6 min. The ATPase inhibitor dicyclohexylcarbodiimide acted to increase markedly the permeability of each organism to protons, and this action indicated that permeability involved not only the passive inflow of protons but also active outflow through the proton-translocating membrane ATPase. Membranes were isolated from each of the bacteria, and pH profiles for ATPase activities indicated pH optima of about 7.5, 7.0, 6.0, and 5.0 for S. sanguis, S. salivarius, S. mutans, and L. casei, respectively. Thus, the pH profiles for the enzymes reflected the acid tolerances of the bacteria and the permeabilities of whole cells to protons.
Levels of antibodies to Plasmodium falciparum sporozoite surface antigens reflect malaria transmission rates and are persistent in the absence of reinfection
Authors: Druilhe P , Pradier O , Marc J P , Miltgen F , Mazier D , Parent G
Published: January 2020

Antibodies reacting with Plasmodium falciparum sporozoite surface antigens were measured by an immunofluorescence assay using wet preparations of sporozoites attached to poly-L-lysine-treated glass slides, a procedure which was found to be more specific than one using glutaraldehyde-treated and dried preparations. Subjects recovering from a first attack were found to be negative. In two African villages which differed in the level at which mosquitoes transmit the disease (1 and 100 infective bites per year and per individual), both the prevalence by age group and the levels of anti-sporozoite antibodies differed markedly, as follows. In the low-transmission area, these antibodies were not detected in subjects aged 2 to 10 years; thereafter, prevalence increased gradually with the age of the subject and reached 90% in subjects aged 50 to 80 years. In the high-transmission area, all of the subjects studied, including the younger ones, were positive. Anti-sporozoite antibody levels were independent of the levels of antibodies directed against blood stages. On average, the mean antibody titers were equal to 1/16 in the first village and 1/1,650 in the second one. These results suggest that stage-specific antibodies reflect the cumulative number of sporozoites inoculated in humans by mosquitoes and may therefore have useful epidemiological applications. In addition, the presence of stage-specific antibodies in the sera of African adults collected at different times after departure from the endemic area indicates that they may last for several years. During the course of this study, we observed a heterogeneity of immunofluorescence labeling in parasite populations prepared from mosquito salivary glands. This raises the question of possible qualitative or quantitative antigenic differences or both between one sporozoite and the other.
Localization of the streptococcal C5a peptidase to the surface of group A streptococci
Authors: O'Connor S P , Cleary P P
Published: January 2020

Immunofluorescent staining was used to determine that the streptococcal C5a peptidase (SCP) exists as a cell surface antigen on group A streptococci. The ability of hyperimmune serum to neutralize cell-associated SCP activity provided further evidence for the location of SCP. Quantification of SCP during growth in vitro by indirect enzyme-linked immunosorbent assay showed that approximately 90% of the measurable antigen is cell bound.
Common and specific epitopes of Campylobacter flagellin recognized by monoclonal antibodies
Authors: Nachamkin I , Hart A M
Published: January 2020

Murine monoclonal antibodies to Campylobacter jejuni recognized a flagellin epitope common to most Campylobacter species and an epitope restricted to C. jejuni and C. coli. These epitopes are distinct from the serotype-specific epitope recently detected on the flagellin and have not been described previously.
Resistance of C3H/HeJ mice to the effects of Haemophilus pleuropneumoniae
Authors: Fenwick B W , Osburn B I , Olander H J
Published: January 2020

Comparisons were made in the mortality associated with an inhaled dose of viable Haemophilus pleuropneumoniae type 5, strain J45, between adult C3H/HeN and C3H/HeJ mice. Mice of both strains were also challenged with Escherichia coli strains O111:B4 and J5. The 50% lethal dose (LD50) of H. pleuropneumoniae in C3H/HeN mice was calculated to be 10(6.5) CFU. At a mean dose of 10(6.7) CFU a 46% mortality rate occurred in C3H/HeN mice, whereas only 10% of the C3H/HeJ mice died (P less than 0.01). Deaths occurred significantly earlier in C3H/HeN mice (P less than 0.01). No deaths occurred later than 12 h postinfection in either group. Pulmonary lesions in the mice that died were similar to those in pigs that die during the acute phase of H. pleuropneumoniae infection. In surviving mice of both strains, a mild resolving interstitial and bronchopneumonia was present which was not typical of subacute H. pleuropneumoniae infections in swine. Quantitative bacterial isolations from the lungs, liver, and spleen indicate that H. pleuropneumoniae did not multiply in the lungs, was rapidly cleared, and did not become systemic. No deaths occurred in the mice inoculated with E. coli J5 or O111:B4 at mean doses of 10(6.3), 10(7.2), and 10(8.5) CFU, and 10(6.4), 10(7.5), and 10(8.2) CFU, respectively. The difference in the mortality rate between the C3H/HeN and C3H/HeJ mice suggests that endotoxin may be involved in acute deaths in pigs infected with H. pleuropneumonia. As indicated by the E. coli challenge, however, other factors are also likely to be involved. Because of the differences in the pathology and microbiology following H. pleuropneumoniae pulmonary infections in mice and pigs, mice do not appear to be an accurate model of the overall disease in swine.
Identification of antibody classes and Fc receptors responsible for phagocytosis of Trypanosoma musculi by mouse macrophages
Authors: Vincendeau P , Daëron M , Daulouede S
Published: January 2020

The phagocytosis of Trypanosoma musculi by macrophages in the presence of specific antibodies was investigated. In 14-day-infected mice, opsonic antibodies were detected in serum, and phagocytosis of parasites by peritoneal macrophages was observed. The mechanism of T. musculi phagocytosis was analyzed. The binding of trypanosomes to peritoneal macrophages and J774 cells was observed in the presence of serum from hyperimmune mice and from mice infected 14 or 28 days earlier, but not in the presence of control mouse serum or sera from 7-day-infected mice. Binding was partially inhibited by mouse monoclonal immunoglobulins G1 (IgG1) or IgG2a and almost completely inhibited by a mixture of both. Binding was also partially inhibited by the anti-Fc gamma 1/gamma 2b receptor monoclonal antibody 2.4G2. Binding of T. musculi was also induced by fractions of serum from 28-day-infected mice obtained by protein A-Sepharose chromatography. Only the IgG1-rich fraction eluted at pH 6.0 and the IgG2a-rich fraction eluted at pH 4.5 promoted binding which could be almost completely inhibited by monoclonal IgG1 and IgG2a. These data indicate that IgG1 and IgG2a anti-T. musculi antibodies are responsible for the phagocytosis of T. musculi by mouse macrophages and both Fc gamma 2a and Fc gamma 1/gamma 2b receptors are involved. Such a mechanism is likely to account for the elimination of parasites in T. musculi-infected mice.
Heat-modifiable envelope proteins of Bordetella pertussis
Authors: Armstrong S K , Parker C D
Published: January 2020

Several envelope proteins of Bordetella pertussis demonstrated differences in electrophoretic mobility, depending upon solubilization temperature before sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These proteins were exposed on the cell surface as judged by their accessibility to radiolabeling with 125I. Monoclonal antibodies to two of the heat-modifiable proteins (Mrs of 18,000 and 91,000) reacted with intact cells in immunofluorescence microscopy experiments, also indicating surface exposure of these two proteins. Two-dimensional gel electrophoresis revealed that two heat-modifiable proteins (a major protein with an Mr of 38,000 and one with an Mr of 18,000) migrated as higher-Mr moieties when solubilized at low temperatures (25 degrees C). Three proteins (Mrs of 91,000, 32,000, and 30,000) and possibly a fourth (31,000) migrated as lower-Mr species when solubilized at 25 degrees C, as revealed in the two-dimensional gel system; these three proteins were found only in virulent B. pertussis and were not detected in a phase IV avirulent strain nor in a strain modulated to phenotypic avirulence by growth in nicotinic acid. The 38,000 molecular-weight protein (38K protein) and a 25K protein were found to be noncovalently associated with the underlying peptidoglycan. Small amounts of the 91K and 18K proteins were also found associated with peptidoglycan.
Influence of iron on growth, morphology, outer membrane protein composition, and synthesis of siderophores in Campylobacter jejuni
Authors: Field L H , Headley V L , Payne S M , Berry L J
Published: January 2020

Three human isolates of Campylobacter jejuni were grown in a biphasic culture medium with and without the addition of a synthetic chelator to induce iron limitation. Cells grown in low-iron medium exhibited slower growth rates and altered cellular morphology. Increased numbers of longer, more filamentous forms were seen in Gram-stained smears. Three proteins, with apparent Mrs of 82,000, 76,000, and 74,000, were consistently present in the outer membrane of cells grown in low-iron medium. At least one of these proteins (76,000 to 74,000) was exposed on the cell surface. A bioassay was used to look for the production of siderophores by these and other strains of C. jejuni. Seven of 26 strains tested produced detectable amounts of siderophores. Growing strains at 42 degrees C failed to suppress siderophore synthesis or to alter the outer membrane protein profiles of iron-starved cells. The ability of three strains to utilize exogenously supplied siderophores for growth in low-iron medium was also examined. All three strains were able to utilize enterochelin and ferrichrome, but none utilized aerobactin, rhodotorulic acid, or desferrioxamine B. The effect of iron on the virulence of C. jejuni for 11-day-old chicken embryos inoculated via the chorioallantoic membrane was also determined.
Macrophage uptake and retention of radiolabeled glycopeptidolipid antigens associated with the superficial L1 layer of Mycobacterium intracellulare serovar 20
Authors: Hooper L C , Johnson M M , Khera V R , Barrow W W
Published: January 2020

Glycopeptidolipid (GPL) antigens which are associated with the superficial L1 layer of Mycobacterium intracellulare serovar 20 were labeled with radioisotopes by means of internal labeling techniques and used in macrophage uptake and retention studies. The use of tritiated alanine and phenylalanine allowed the incorporation of label into the GPL invariant fatty acyl peptide core, which is common to all members of the Mycobacterium avium-M. intracellulare complex. Radiolabeled GPL antigens were then purified by a one-step column chromatographic procedure and subsequently used to determine the maximum uptake and retention in peritoneal macrophages isolated from C57BL/6 and CBA/J mice. Maximum uptake for peritoneal macrophages from both strains of mice occurred at a concentration between 200 and 250 micrograms of antigen per ml of medium when 3.4 X 10(5) cells were pulsed. Timed experiments demonstrated that approximately 20% of the antigens remained associated with the macrophages up to 4 days after a pulse of 200 micrograms of GPL, and examination of chloroform-extractable components from both macrophages and spent medium revealed that 98% or more of the radioactivity corresponded to intact GPL components. The ability of the GPL antigens to become associated with macrophages is demonstrated by these results, which strongly suggest that these potentially important mycobacterial antigens are inert to degradation by those cells.
Immunopotentiating activities of cell walls, peptidoglycans, and teichoic acids from two strains of Listeria monocytogenes
Authors: Paquet A , Raines K M , Brownback P C
Published: January 2020

Crude cell wall (CCW), enzyme purified wall (PF), peptidoglycans (PEP), and teichoic acid fractions from two strains of Listeria monocytogenes (EGD and a human isolate) were analyzed by electron microscopy, chemical analyses, and various immunological assays. Electron microscopy and chemical analyses revealed that both strains were quite similar. The CCW, PF, and PEP fractions were mitogenic for mouse lymphocytes, could activate macrophages in vivo but not in vitro, and stimulated nonspecific immunity in vivo to Candida albicans. The PF and PEP fractions but not the CCW induced migration inhibition factor production. The CCW and PEP fractions were tested and found to elevate natural killer activity in vivo. Teichoic acid fractions had no activity in the above assays. Since most of the immunological activities were retained in the PEP fraction, we examined this fraction to determine the kinetics of the mitogenic response and the nature of the responding cell population. The peak mitogenic response occurred on day 2 over a 6-day period. This response was not enriched in T-cell-enriched populations but followed the pattern of LPS in B-cell-enriched populations.
Experimental intrauterine infection of adult BALB/c mice with Cryptosporidium sp
Authors: Liebler E M , Pohlenz J F , Woodmansee D B
Published: January 2020

Inoculation of adult, female BALB/c mice with 2 X 10(5) bleach-treated Cryptosporidium sp. oocysts isolated from calf feces resulted in infection of the uterine mucosa in more than 50% of the animals. Cryptosporidium sp. completed the entire life cycle in the uterus, and infectious oocysts were passed into the vagina. Two methods of application were used to establish intrauterine infection. The inoculum was either injected into the uterus after abdominal surgery or intracervically instilled. Mice were susceptible at all phases of the sexual cycle, but the highest infection rates were obtained during estrus and diestrus. Parasites were demonstrated as early as 5 days postinfection. Phagocytic cells in the uterine lumen and in the vagina contained Cryptosporidium sp. Phagocytosis may be an important immune response and a mechanism of parasitic clearance. These results suggest that Cryptosporidium sp. is a potential pathogen of the reproductive tract.
Isolation, characterization, and mapping of Tn5 insertions into the 140-megadalton invasion plasmid defective in the mouse Sereny test in Shigella flexneri 2a
Authors: Sasakawa C , Makino S , Kamata K , Yoshikawa M
Published: January 2020

Using Shigella flexneri 2a YSH6000, we isolated 304 independent Tn5 insertion mutants in the 230-kilobase invasion plasmid, pMYSH6000. The site of each Tn5 insertion was assigned to 23 SalI fragments on the previously made SalI cleavage map of pMYSH6000. Among the 304 insertions, 150 were negative in expression of four phenotypes examined (mouse Sereny test [Ser], invasion into epithelial cells [Inv], Congo red binding [Pcr], and inhibition of bacterial growth [Igr] ): 12 were Ser- Inv+ Pcr+ Igr+, and 142 were positive in all four phenotypes. Tn5 insertions in the avirulent mutants were distributed in two separate SalI fragments, F and G, and in four contiguous SalI fragments, B, P, H, and D. Fragment G contains a novel class of determinant(s) which is required only for Ser+ but not for Inv+, Pcr+, and Igr+. Fragment F contains the previously characterized virF locus. B, P, H, and D each contained both virulent and avirulent Tn5 insertions. This indicates that more than two gene clusters exist within this region. Both are required for expression of all four virulence phenotypes.
Identification and characterization of a uroepithelial cell adhesin from a uropathogenic isolate of Proteus mirabilis
Authors: Wray S K , Hull S I , Cook R G , Barrish J , Hull R A
Published: January 2020

Proteus mirabilis is a frequent cause of urinary tract infections in rehabilitation hospitals and among persons with structural abnormalities of the urinary tract. Adherence to uroepithelial tissues may be an important virulence determinant in these infections because most Proteus strains adhere to desquamated uroepithelial cells. To identify the adherence factor responsible for this phenomenon, we sheared outer membrane material from 35SO4-radiolabeled bacteria and allowed it to bind to uroepithelial cells. Following sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the major adherence element was a protein with an apparent molecular weight of 17,500 and was provisionally designated as the uroepithelial cell adhesin. This adhesin was purified by heat shock and gel filtration on Sepharose CL-4B. After purification, the adhesin was seen assembled as long, flexible rods by electron microscopy. The N-terminal amino acid sequence of the subunit had limited homology with that of the K99 fimbriae of Escherichia coli.
Immunogenic activity of gonococcal protein I in mice with three different lipoidal adjuvants delivered in liposomes and in complexes
Authors: Jiskoot W , Teerlink T , Van Hoof M M , Bartels K , Kanhai V , Crommelin D J , Beuvery E C
Published: January 2020

For several reasons the major outer membrane protein from Neisseria gonorrhoeae (gonococcal protein [PI]) is an attractive component for a gonococcal vaccine. This paper describes the influence of two different physical forms of PI on its immunogenic activity. To this end PI was delivered in liposomes and in protein-detergent complexes. In both forms PI was present in a multimeric form. The liposomes were composed of phosphatidylcholine and cholesterol. The effect of dicetylphosphate as a negatively charged amphiphile and three lipoidal adjuvants was investigated. Two lipoidal adjuvants (Avridine and dimethyldioctadecylammoniumbromide) were positively charged amphiphiles, whereas the third one (tridecyl N-acetylmuramyl-L-alanyl-D-isoglutaminate) was neutral. The protein-detergent complexes were also tested in the presence of the lipoidal adjuvants and in an AlPO4-adsorbed form. The liposome preparations were characterized for their size, charge, and residual amount of detergent. The immunogenic activity of PI in all forms was tested in mice. The results of the antibody assays showed that PI in the liposomes was more immunogenic than PI in the complexes. A second dose with liposomes induced only a small booster effect, whereas such a dose with the complexes produced pronounced booster effects. The incorporation of the positively charged lipoidal adjuvants in the liposomes resulted in enhanced booster effects. The highest immunogenic activity of PI after two injections, however, was observed in the complexed form adsorbed to AlPO4.
Suppression of interleukin-2 production by macrophages in genetically susceptible mice infected with Leishmania major
Authors: Cillari E , Liew F Y , Lelchuk R
Published: January 2020

Spleen cells from BALB/c mice infected with 2 X 10(7) L. major promastigotes and developing progressive disease produced significantly lower levels of interleukin-2 (IL-2) in response to concanavalin A stimulation than did spleen cells from uninfected mice. In contrast, spleen cells from sublethally irradiated and infected mice, which were able to contain lesion development, produced significantly higher levels of IL-2. The increase in IL-2 production closely paralleled lesion regression. Mice protectively immunized by four intravenous injections with lethally irradiated promastigotes also produced enhanced levels of IL-2, which were sustained after challenge infection. In contrast, spleen cells from BALB/c mice given four s.c. injections of irradiated promastigotes produced high levels of IL-2 before but not after infection. These mice eventually produced levels of IL-2 indistinguishable from those of unimmunized mice with progressive disease. There is thus an inverse relation between disease progression and the ability of spleen cells to produce IL-2. Spleen cells from mice with uncontrolled disease not only produced lower levels of IL-2 but also impaired IL-2 production by normal spleen cells. The ability to inhibit IL-2 was abrogated by passing the cells through a Sephadex G-10 column, removal of plastic adherent cells, and removal of carbonyl iron-ingesting cells. Furthermore, Sephadex G-10 column-treated and plastic adherent, nonspecific esterase-positive spleen cells from mice with progressive disease were able to suppress IL-2 production by normal splenic T cells. The suppressive activity of the adherent cells was not affected by treatment with anti-Thy-1.2 antibody and complement. In contrast, adherent spleen cells from uninfected mice were devoid of such suppressor activity. The depressed IL-2 production by spleen cells from progressively infected mice could be restored to that of normal spleen cells by the addition of indomethacin to the culture. There was however, no correlation between IL-2 production and IL-1 activity in infected or immunized BALB/c mice. Thus, it appears that the suppression of IL-2 production is mediated by prostaglandins elaborated by macrophages from chronically infected mice.
Hemolysin production and cloning of two hemolysin determinants from classical Vibrio cholerae
Authors: Richardson K , Michalski J , Kaper J B
Published: January 2020

The hemolytic activity of 20 classical and 3 El Tor strains of V. cholerae O1 was examined phenotypically and genetically. The El Tor strains lysed bovine, chicken, human, rabbit, and sheep erythrocytes (RBCs), while the classical strains lysed only chicken and rabbit RBCs. The assay was done with RBCs in Tris-NaCl buffer, since phosphate-buffered saline was found to inhibit hemolytic activity. Hemolytic activity in culture supernatants from El Tor strains was more sensitive to heat inactivation than that in supernatants from the classical strain 395. A gene library of strain 395 was examined for hemolytic activity, and two distinct hemolytic clones were identified. One clone appeared identical to the previously cloned hemolysin structural gene from El Tor V. cholerae, while the other did not hybridize to the El Tor hemolysin probe, had a unique restriction enzyme digestion pattern, and encoded a hemolysin whose activity differed from that of the El Tor hemolysin clones. We suggest that the hemolysin specified by the determinant originally cloned from an El Tor vibrio be designated hemolysin I and the second hemolysin, cloned from the classical vibrio, be designated hemolysin II.
Heterogeneous activity of immature and mature cells of the murine monocyte-macrophage lineage derived from different anatomical districts against yeast-phase Candida albicans
Authors: Decker T , Lohmann-Matthes M L , Baccarini M
Published: January 2020

Mature mononuclear phagocytes have been receiving much attention as effectors of spontaneous candidacidal activity, although with controversial results due to differences in the effector populations and the methods used in different laboratories. We here systematically compare the fungistatic activity of immature and mature cells of the murine macrophage series. The results show that nonadherent, nonphagocytic precursor cells (isolated either [90% purity] from bone marrow liquid cultures or from the organs of mice in which inflammatory conditions had been elicited in vivo) exerted a strong extracellular candidastatic activity. In contrast, mature macrophages, either obtained from different anatomical areas (spleen, liver, lung, peritoneal cavity) or matured in vitro from the precursor populations, displayed striking heterogeneity in their ability to inhibit the growth of Candida albicans, depending on the anatomical site they were derived from. Lymphokine activation did not alter the fungistatic pattern of the untreated cells. The different macrophage populations behaved very differently also in the production of reactive oxygen intermediates (ROI) in response to phagocytosis of C. albicans. The amounts of ROI generated, however, showed no correlation with candidastatic ability. Low levels of candidastatic activity exerted by resident peritoneal macrophages (good ROI producers) were inhibited by catalase, whereas high levels of growth inhibition by Kupffer cells (poor ROI producers) after 8 h of assay were hardly influenced by the enzyme. Our data suggest the existence of two different effector mechanisms in macrophage-mediated C. albicans growth inhibition, a rather inefficient ROI-dependent one, and a second, very efficient oxygen-independent mechanism. The implications of these findings are discussed.
Fimbria-specific antibodies in serum and saliva of mice immunized with Actinomyces viscosus T14V fimbriae
Authors: Crawford P C , Clark W B
Published: January 2020

Fimbria-specific antibody responses were compared in mice immunized with purified fimbrial adhesins in the region of the submandibular gland (i.e., local site) or at a remote site in the back. One hundred micrograms of fimbriae isolated from Actinomyces viscosus T14V was used as the vaccine. Four subcutaneous injections of the vaccine in the local site induced greater amounts of fimbria-specific immunoglobulin G (IgG) in serum and saliva than three injections. However, there was no difference in the response of fimbria-specific IgA in serum and saliva. Fimbria-specific IgG in serum and saliva were first detected 21 days after the primary immunization at both the local or remote sites. Fimbria-specific IgA in serum was first detected 28 days after the primary immunization at both the local or remote sites. However, fimbria-specific IgA in saliva occurred only in mice immunized with the fimbrial vaccine at the local site and was first detected 14 days after the primary immunization. Both serum and saliva from mice immunized 4 times with the fimbrial vaccine in the local site inhibited in vitro adsorption of strain T14V cells to hydroxyapatite beads pretreated with normal mouse saliva, whereas adsorption of strain T14V cells suspended in serum and saliva from sham-immunized animals was not inhibited. Collectively, these data suggest that mice immunized locally in the submandibular gland region with a vaccine composed of purified fimbrial adhesins provide a potential model for evaluating the efficacy of fimbria-specific antibodies in saliva to inhibit strain T14V colonization of tooth surfaces.
Cloning and expression of the Bacillus anthracis protective antigen gene in Bacillus subtilis
Authors: Ivins B E , Welkos S L
Published: January 2020

The gene encoding the protective antigen (PA) moiety of the tripartite exotoxin of Bacillus anthracis was cloned from the recombinant plasmid pSE36 into Bacillus subtilis 1S53 by using the plasmid vector pUB110. Two clones, designated PA1 and PA2, were identified which produced PA in liquid cultures at levels of 20.5 to 41.9 micrograms/ml. This PA was identical to B. anthracis Sterne PA with respect to migration on sodium dodecyl sulfate-polyacrylamide gels and to Western blot antigenic reactivity. Addition of lethal factor or edema factor to PA1 and PA2 supernatants generated biologically active anthrax lethal toxin or edema-producing toxin, respectively. The recombinant plasmid in PA1 (pPA101) was 7.8 kilobases, whereas the PA2 strain plasmid (pPA102) was 6.1 kilobases. Both plasmids had deletions extending into the insert sequence but not into the DNA encoding the PA protein. Immunization with the live recombinant strains protected guinea pigs from lethal challenge with virulent B. anthracis spores, and the immunization partially or completely protected rats from intravenous challenge with anthrax lethal toxin.
Production of type II heat-labile enterotoxin by Escherichia coli isolated from food and human feces
Authors: Guth B E , Pickett C L , Twiddy E M , Holmes R K , Gomes T A , Lima A A , Guerrant R L , Franco B D , Trabulsi L R
Published: January 2020

Escherichia coli strains isolated in Sao Paulo, Brazil, from feces of patients with diarrhea and from food samples produced toxin(s) that was shown to be related both immunologically and genetically to the recently characterized type II heat-labile enterotoxin of E. coli. The new isolates of type II heat-labile enterotoxin-producing E. coli belonged to five different serotypes and did not represent a single clone.
Inhibition of immediate and Arthus responses to schistosome egg antigens by T cells from Schistosoma japonicum-infected mice
Authors: Stavitsky A B , Olds G R
Published: January 2020

Schistosoma japonicum-infected mice develop hepatic granulomas, immediate hypersensitivity (IH), and delayed hypersensitivity (DH) to soluble egg antigens (SEA) released by parasite eggs trapped in liver sinusoids. All of these responses spontaneously regress after 7 to 9 weeks of infection. This study aimed to develop an in vivo system for the further dissection of cellular and humoral immune responses to SEA. C57BL/6 mice immunized subcutaneously with SEA in complete Freund adjuvant developed IH, an Arthus reaction, and DH to this antigen 5 to 9 days later. IH and the Arthus reaction, but not DH, were markedly inhibited if, 1 day before injection of SEA in complete Freund adjuvant, the mice were injected intravenously with purified T cells from the spleens of mice infected for at least 9 weeks. This in vivo model system can be used to study various aspects of cellular and humoral immune responses to SEA and their modulation. These results raise questions about the role of antibodies in the pathogenesis of granulomatous inflammation and about the mechanisms of its cellular regulation in infections with S. japonicum.
Purification and properties of hemagglutinin from culture supernatant of Bacteroides gingivalis
Authors: Okuda K , Yamamoto A , Naito Y , Takazoe I , Slots J , Genco R J
Published: January 2020

The hemagglutinating factor (hemagglutinin) of Bacteroides gingivalis was prepared from the supernatant of a 5-day diffusate broth culture by ammonium sulfate precipitation and column chromatography with a hydrophobic column of Phenyl-Sepharose CL-4B, DEAE-Sephadex A-50, and Sephadex G-100 gel filtration. The hemagglutinating activity of the preparation was 53.3 times higher than that of ammonium sulfate precipitate. In electron microphotographs, hemagglutinin appears to have a vesicle or tubelike structure. The hemagglutinating activity of intact cells was completely destroyed by heating at 100 degrees C for 10 min, but the activity of extracted hemagglutinin was heat stable. The activity of hemagglutinin was inhibited by L-arginine and L-lysine and partially inhibited by phospholipase D, but it was not affected by proteolytic enzymes, neuraminidase, hyaluronidase, lipase, phospholipase A and C, or sugars. The B. gingivalis hemagglutinin appeared to be comprised mainly of a 40,000-molecular-weight material. The Fab fragment of immunoglobulin G prepared from rabbit antiserum to whole cells of B. gingivalis and monoclonal antibody against the hemagglutinin bound to the cell surface and inhibited the hemagglutinating activity of both the cells and the purified hemagglutinin.
Purification and characterization of Clostridium perfringens iota toxin: dependence on two nonlinked proteins for biological activity
Authors: Stiles B G , Wilkins T D
Published: January 2020

Clostridium perfringens type E iota toxin, a dermonecrotic and lethal binary toxin, was purified to homogeneity. Each protein component of the toxin, iota a (ia) or iota b (ib), appeared as a single band by gradient or sodium dodecyl sulfate-polyacrylamide gel electrophoresis and yielded a single immunoprecipitin arc by crossed immunoelectrophoresis with homologous antiserum. Individually, ia (Mr 47,500) or ib (Mr 71,500) had little biological activity. However, when combined in equimolar amounts, there was a 64-fold increase in the guinea pig dermonecrotic titer. The biological activity of ia was heat stable (85 degrees C for 15 min), whereas ib was inactivated at 55 degrees C. Our results demonstrated that C. perfringens iota toxin required two different, nonlinked protein components for biological activity.
Eosinophil chemotactic lymphokine produced by egg-associated granulomas in murine schistosomiasis japonicum
Authors: Owhashi M , Maruyama H , Nawa Y
Published: January 2020

Eosinophil chemotactic factor (ECF) was detected in the culture supernatant of isolated intact egg granulomas from the livers of Schistosoma japonicum-infected mice. This factor had an apparent molecular weight of 15,000 by high-pressure liquid chromatography with an SW3000 column and bound to concanavalin A-Sepharose 4B. When cells obtained by enzymatic digestion of isolated granulomas were cultured under the presence of soluble egg antigen of S. japonicum or concanavalin A, ECF was also detected in the conditioned medium. The physicochemical nature of the ECF produced by concanavalin A-stimulated granuloma cells was similar to that produced by isolated intact granulomas. The ECF-producing activity of the cells was abolished by pretreatment with anti-Thy-1.2 or anti-Lyt-1.2 monoclonal antibody and complement but not by anti-Lyt-2.2 antibody. Furthermore, nylon wool-passed, T-enriched granuloma cells required collaboration of syngeneic macrophages to produce ECF. These results suggest that Lyt-1-positive T cells in the granuloma could, in collaboration with macrophages, produce ECF and thereby attract eosinophils to this lesion.
Antibody response of swine to outer membrane components of Haemophilus pleuropneumoniae during infection
Authors: Rapp V J , Ross R F
Published: January 2020

Sera from pigs infected with Haemophilus (Actinobacillus) pleuropneumoniae were tested for antibodies to outer membrane proteins (OMPs) of the organism by immunoblotting. Convalescent sera were produced in naturally born, colostrum-fed pigs and in cesarean-derived, colostrum-deprived pigs given H. pleuropneumoniae serotype 5 intranasally twice at 5-week intervals. Sera, collected at weekly intervals, were reacted with Sarkosyl-insoluble, OMP-enriched preparations of H. pleuropneumoniae which had been separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and electrophoretically transferred to nitrocellulose. Antibodies were detected to OMPs with an apparent molecular weight of 16,500 (16.5K OMP); to 29K, 38.5K, 43.5K, 45K, 49.5K, and 66.5K OMPs; and to several high-molecular-weight (greater than or equal to 94,000) OMPs, but not to the major 42K OMP. Antibodies to the heat-modifiable OMP (29K/43.5K) and the 38.5K OMP were detected in sera from noninfected pigs. Antibodies were also detected to two broad 54,000- and 95,000-molecular-weight bands which did not stain with Coomassie blue, stained with silver nitrate, resisted proteinase K digestion, and were eliminated by oxidation with sodium metaperiodate. This indicates that the 54,000- and 95,000-molecular-weight bands represent polysaccharide, possibly capsular or lipopolysaccharide immunogens. Adsorption of sera with cells from the homologous serotype 5 strain removed antibodies to the 45K, 49.5K, 66.5K, and greater than or equal to 94K OMPs and to the two polysaccharide bands, indicating that these antibodies were directed primarily to surface-exposed epitopes. When tested with OMP preparations from other serotype 5 strains, heterogeneity was apparent, both in the reactions with OMPs and with the polysaccharide bands. Silver staining of proteinase K-treated, whole-cell lysates from serotype 5 strains also indicated variable expression of the polysaccharide bands. Sera also reacted with OMPs from H. pleuropneumoniae serotypes 1 and 7; however, several OMPs and the lipopolysaccharide or polysaccharide determinants of these serotypes appeared to be type specific.
Siderophore production by Pasteurella multocida
Authors: Hu S P , Felice L J , Sivanandan V , Maheswaran S K
Published: January 2020

Pasteurella multocida grown under conditions of iron deprivation secreted into the culture medium a growth-enhancing factor which functioned as a siderophore. The siderophore was found to be neither a phenolate nor a hydroxamate by chemical tests and bioassays and was given the trivial name multocidin. Multocidin was partially purified and found to be a highly polar, nonaromatic, and dialyzable compound. This is the first report demonstrating the production of a siderophore by P. multocida.
Anti-L3T4 antibody treatment suppresses hepatic granuloma formation and abrogates antigen-induced interleukin-2 production in Schistosoma mansoni infection
Authors: Mathew R C , Boros D L
Published: January 2020

In murine schistosomiasis mansoni, granulomatous inflammation is an immune response that involves egg antigen presentation to T cells in the context of class II major histocompatibility complex determinants and subsequent inflammatory lymphokine production by delayed-hypersensitivity (TDH) lymphocytes. In the present study, monoclonal antibodies directed against L3T4, I-A, and Lyt-2 molecules were injected intraperitoneally into S. mansoni-infected mice to study the role of these membrane antigens in the process of granuloma formation. A dramatic suppression of the hepatic granuloma size and antigen-induced interleukin-2 (IL-2) production by spleen cells was seen in mice that received anti-L3T4 monoclonal antibody treatment. The total number of cells, especially the L3T4+ T cells, was greatly diminished in the spleens. Furthermore, histopathological study of the granulomas in stained liver sections demonstrated the paucity of eosinophils and macrophages, absence of epithelioid cells and multinucleated giant cells, and minimal collagen deposition within the lesions. Damaged hepatocytes were also seen surrounding these ill-formed granulomas. In contrast, anti-I-A monoclonal antibody treatment partially suppressed IL-2 production, although granuloma size and cellular composition remained the same. Mice that received anti-Lyt-2 monoclonal antibody did not show any changes in either IL-2 production or hepatic granulomatous inflammation. The data presented in this paper indicate a crucial role for L3T4 molecules present on a subset of class II major histocompatibility complex-restricted TDH cells in IL-2 production and the generation of the granulomatous response.
Effect of acetylation on arthropathic activity of group A streptococcal peptidoglycan-polysaccharide fragments
Authors: Stimpson S A , Lerch R A , Cleland D R , Yarnall D P , Clark R L , Cromartie W J , Schwab J H
Published: January 2020

Purified group A streptococcal peptidoglycan-polysaccharide (PG-PS) fragments were either de-O-acylated, or acetylated and then de-O-acylated to yield N-acetylated PG-PS. Native PG-PS was poorly degraded, N-acetylated PG-PS was extensively degraded, and de-O-acylated PG-PS was only slightly degraded by hen egg white lysozyme. N-acetylated PG-PS was also extensively degraded by human lysozyme and partially degraded by rat serum or rat liver extract. After a single intraperitoneal injection of rats with a sterile, aqueous suspension, all PG-PS preparations induced acute arthritis. The acute arthritis induced by N-acetylated PG-PS was significantly more severe than that induced by native PG-PS; that induced by de-O-acylated PG-PS was of intermediate severity. After the acute reaction, rats injected with native PG-PS developed chronic relapsing erosive synovitis which remained severe for the duration of the experiment (83 days). In contrast, joint inflammation induced by N-acetylated PG-PS resolved within 6 weeks with little evidence of recurrent disease. Chronic arthritis induced by de-O-acylated PG-PS was of intermediate severity. In another assay of arthropathic activity, the arthritis in all rat ankle joints, which had been injected directly with native PG-PS, could be reactivated 3 weeks later by the intravenous injection of a small dose of PG. In contrast, only 50% of the joints initially injected with de-O-acylated PG-PS and none of the joints injected with N-acetylated PG-PS could be reactivated. These studies support the concepts that the resistance of PG-PS to muralytic digestion is crucial for chronic arthropathic activity and that the nature and degree of PG acetylation are important molecular determinants of the phlogistic activities of PG-PS polymers.