bluuebunny/crossref_metadata_2025 huggingface dataset:
Serological Investigations of a Bovine Respiratory Disease (“Urner Pneumonie”) Resembling Farmer's Lung
Authors: Nicolet J. , de Haller R. , Herzog J.
Published: January 2020

The immunological response of cattle exposed to moldy hay was examined by agar gel diffusion with standard farmer's lung hay antigens. A high incidence of precipitins against
Micropolyspora faeni
(60%) and moldy hay antigen (80%) was detected in exposed but apparently healthy cattle from a region with a high incidence of bovine farmer's lung. In comparison, in the plains, a low incidence area, we found only 1 animal of 164 harboring precipitins against
M. faeni.
We further observed that many animals from exposed populations lost their precipitins during pasturing and regained them during winter housing. Thirty-nine clinical cases of bovine farmer's lung (“Urner Pneumonie”) were investigated serologically. Only 49% of these animals showed precipitins against
M. faeni
and 54% showed precipitins against moldy hay antigen. We discuss in this paper the probable causes of this apparent lack of immunological response.
Mycoplasma-Like Organism, Parasite of Red Blood Cells of an Amphibian,
Hydromantes italicus
(
Spelerpes fuscus
)
Authors: Babudieri B.
Published: January 2020

In the present investigation we studied a parasite of red blood cells in the amphibian
Hydromantes italicus.
The parasite, a globular microorganism having a diameter of 0.25 to 0.3 μm, was found in large numbers in vacuoles of red blood cells. As seen by electron microscopy, it possesses no cell wall and has the morphological appearance of a mycoplasma. The parasite has great motility by mechanisms not yet elucidated. It is possible to transmit the infection experimentally from infected to healthy amphibians. Culture of the microorganism, named
Chondrostagon haematicum
, in artificial media has not yet been successful.
Effect of Vincristine Sulfate on
Pseudomonas
Infections in Monkeys
Authors: Saslaw Samuel , Carlisle Harold N. , Moheimani Mohammad
Published: January 2020

In rhesus monkeys, intravenous challenge with 0.6 × 10
10
to 2.2 × 10
10
Pseudomonas aeruginosa
organisms caused acute illness of 4 to 5 days' duration with spontaneous recovery in 13 of 15 monkeys; blood cultures became negative 3 to 17 days after challenge. Leukocytosis was observed in all monkeys. Intravenous or intratracheal inoculation of 2.0 to 2.5 mg of vincristine sulfate was followed by leukopenia in 4 to 5 days. Intravenous inoculation of 4.2 × 10
10
to 7.8 × 10
10
pyocin type 6
Pseudomonas
organisms in monkeys given vincristine sulfate 4 days previously resulted in fatal infection in 11 of 14 monkeys, whereas none of four receiving
Pseudomonas
alone died. These studies suggest that an antimetabolite-induced leukopenia predisposes to severe
Pseudomonas
sepsis and that such monkeys may serve as a biological model for study of comparative efficacy of antimicrobial agents.
Intracellular Fate of
Cryptococcus neoformans
Authors: Tacker J. R. , Farhi F. , Bulmer G. S.
Published: January 2020

Human peripheral leukocytes were found to engulf and kill cells of
Cryptococcus neoformans.
Fewer encapsulated than nonencapsulated cells met this fate, since cryptococcal capsular polysaccharide inhibited phagocytosis. During 10 to 12 hr of incubation of nonencapsulated cells in human serum, sufficient polysaccharide was produced to inhibit phagocytosis by 50%. The polysaccharide inhibitor was found in the sera of four patients with cryptococcosis, but not on the surfaces of their leukocytes. Additional experiments indicated that serum is not essential for effective phagocytosis. However, normal human serum contains anticryptococcal activity which is not inhibited by capsular material. Preliminary findings indicate that the phagocytic index of a patient with cryptococcosis may be correlated with the severity of his disease.
Tumor Induction by
Cryptococcus neoformans
Authors: Price J. Townley , Bulmer Glenn S.
Published: January 2020

A human isolate of
Cryptococcus neoformans
strain CIA, which was originally obtained in 1963, produced fatal disease in mice. Postmortem examinations showed extensive central nervous system disease. After using this yeast for research for 7 years, we found that it began to produce large tumors after intraperitoneal inoculation in mice. The present study showed that the tumors consisted of large agglomerates of yeast cells, capillaries, and reticular stroma. In concomitant experiments, the same
C. neoformans
strain, introduced into the lungs of mice in an aerosol, produced asymptomatic infections. A definition for the term “cryptococcoma” is proposed.
Nitrogen Metabolism and Protein Synthesis During Pneumococcal Sepsis in Rats
Authors: Powanda M. C. , Wannemacher R. W. , Cockerell G. L.
Published: January 2020

Markedly increased synthesis of α
2
and β globulins and α
1
, α
2
, and β glycoglobulins occurs during pneumococcal sepsis in the rat simultaneously with decreased albumin formation, diminished tritiated leucine incorporation into muscle protein, and enhanced excretion of nitrogen. This augmented synthesis of specific serum proteins does not become evident until fever and bacteremia develop, and it appears to be a fundamental aspect of host response to a proliferating bacterial infection in that it occurs even in rats fed a protein-deficient (6% protein) diet after weaning and before exposure to
Diplococcus pneumoniae.

Although amino acid catabolism, in general, appears to be increased during infection, tryptophan degradation via the kynurenine pathway, as assessed by measuring diazotizable urinary metabolites, changes little or is, at times, significantly less than in control animals. Coincidentally, functional tryptophan oxygenase activity decreases at 16 hr after exposure. Total tryptophan oxygenase activity, however, is unchanged.
Human Immunoglobulin Specificity After Group B Arbovirus Infections
Authors: McNair Scott Robert , McCown Jack M. , Russell Philip K.
Published: January 2020

Sera obtained during a dengue-2 epidemic from individuals with primary or secondary group B arbovirus infections contained cross-reactive antibody against several viruses in this group. Hemagglutination-inhibition antibody titers of the immunoglobulin M fractions from these sera revealed a rise in antibody specific for dengue-2. The acute and convalescent sera in over half of the secondary infection group also had low-level unchanging immunoglobulin M antibody titers to yellow fever which implicated yellow fever immunization as the primary infection.
Inhibition of Protein Synthesis After Intravenous or Intramuscular Challenge with Diphtheria Toxin
Authors: Bonventre Peter F. , Saelinger Catharine Bowman
Published: January 2020

Inhibition of protein synthesis in guinea pig tissues after administration of diphtheria toxin varies significantly with the route of injection.
Herpesvirus hominis
Type 2 Infections in Rabbits: Effect of Prior Immunization with Attenuated
Mycobacterium bovis
(BCG) Cells
Authors: Larson C. L. , Ushijima R. N. , Karim R. , Baker M. B. , Baker R. E.
Published: January 2020

Angora, New Zealand, and Dutch-belted rabbits were tested for their susceptibility to graded doses of
Herpesvirus hominis
type 2 administered by vaginal instillation, intracorneal injection, and scarification of the cornea. Central nervous system involvement and death occurred after infection by the various routes employed. Prior immunization of Dutch-belted and New Zealand rabbits with viable
Mycobacterium bovis
(BCG) cells injected intravenously provided protection against subsequent infections with type 2 virus.
Immunological Conversion of
Vibrio cholerae
in Gnotobiotic Mice
Authors: Miller C. E. , Wong K. H. , Feeley J. C. , Forlines M. E.
Published: January 2020

Cholera vibrios grow readily in the intestines of gnotobiotic mice and change their antigenic structure in response to immunological pressures. These changes were progressive, with the rough form persisting as the apparent end point in the mice. Inoculation of the rough form into fresh gnotobiotic mice resulted in the recovery of smooth forms. An untypable rough strain of
Vibrio cholerae
isolated from a chronic carrier in Calcutta in 1967 was passed serially in gnotobiotic mice. Smooth strains of
V. cholerae
were recovered from the feces after selective treatment with complement and antiserum against a rough strain. They were confirmed as O group I cholera vibrios belonging to the
eltor
biotype. The ability of these strains to produce diarrheal fluid in infant rabbits and to increase capillary permeability in rabbit skin was markedly increased over that of the original human isolate. This demonstrates that rough avirulent forms of
V. cholerae
from human carriers can undergo antigenic changes and concurrently increase in virulence when placed in an appropriate environment. The many inapparent cholera infections in endemic areas may provide a mechanism by which
V. cholerae
changes its serotype away from the immunological pressures in the population.
Competence of Pneumococcal Isolates and Bacterial Transformations in Man
Authors: Ottolenghi-Nightingale Elena
Published: January 2020

A survey of pneumococci isolated from 19 healthy carriers and from 23 patients with pneumococcal disease showed that, for both groups, 25 to 30% of the isolates were competent for transformation by soluble deoxyribonucleic acid (DNA) in vitro. Untransformable type 3 and type 8 pneumococci, whose capsules had been hydrolyzed by the
Bacillus palustris
enzymes prior to exposure to DNA, remained untransformable. Thus, at least for these isolates, it was not the presence of capsule that prevented transformation. Type 9 pneumococci in a healthy human carrier were transformed by DNA released from living unencapsulated pneumococci sprayed onto the pharynx. The donor bacteria were resistant to 1,000 μg of streptomycin/ml. Two types of streptomycin-resistant bacteria were recovered from the carrier's pharynx: a type 9 pneumococcus and an alpha-hemolytic streptococcus. No streptomycin-resistant, gram-positive cocci were isolated from this individual prior to inoculation of the pharynx with the resistant organisms. It seems possible that transformations can occur in the natural environment of some gram-positive cocci.
Target-Organ Treatment of Neurotropic Virus Disease with Interferon Inducers
Authors: Allen Lois B. , Cochran Kenneth W.
Published: January 2020

Interferon inducers were used against vaccinial encephalitis to study the target-organ treatment of neurotropic disease and to correlate interferon levels and the antiviral state following such treatment. A 45-μg amount of statolon, 30 μg of polyribinosinic-polyribocytidylic acid complex (poly I·poly C), or 0.0154 HA unit of Sendai virus given intracerebrally protected 100% of mice challenged the next day with 1,000 median lethal doses (LD
50
) of vaccinia virus. Significant protection against 1,000 LD
50
of vaccinia virus persisted for 1, 4, or 3 weeks after poly I·poly C, statolon, or Sendai virus (154 HA units), respectively. These doses of poly I·poly C and statolon were also used to study postinfection treatment. Mice challenged with 1, 10, 100, or 1,000 LD
50
were treated intracerebrally with poly I·poly C or statolon 24 or 48 hr later. Significant increases in survival time were seen in mice challenged with 1 to 100 LD
50
of vaccinia virus and treated 24 hr later. At challenges of 10 or 100 LD
50
, statolon was more effective than poly I·poly C in increasing survival times. When treatment was delayed until 48 hr after infection, significant increases in survival time occurred only when the challenges were in the range of 1 to 10 LD
50
, with poly I·poly C and statolon being equally effective. Interferon was measured by Finter's dye-uptake method, with L-929 cells and Semliki Forest virus. Poly I·poly C, statolon, or Sendai virus, given intracerebrally to mice, produced serum interferon peaks of 5,120 units/ml at 2 hr, 2,560 units/ml at 12 hr, or 320 units/ml at 18 hr, respectively. Corresponding brain interferon peaks were 640 units/g at 2 hr, 640 units/g at 4 to 24 hr, and 960 units/g at 72 hr.
Isolation and Characterization of a Native Cell Wall Complex from
Neisseria meningitidis
Authors: Zollinger W. D. , Kasper D. L. , Veltri B. J. , Artenstein M. S.
Published: January 2020

A cell wall complex has been isolated by gentle methods from both the medium supernatant fluid and whole organisms of
Neissieria meningitidis
cultures. The two types of preparations have been shown to be essentially identical on the basis of chemical composition, electron microscopy, and polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS-PAGE). Four major components were identified in the complex: group-specific polysaccharide (4 to 10%), protein (45 to 65%), lipopolysaccharide (10 to 25%), and lipid (15 to 30%). The whole complex was found to be immunogenic in rabbits and to elicit production of antibody directed against the protein, the group-specific polysaccharide, and the lipopolysaccharide components. The isolated protein component was also found to be immunogenic in rabbits and to elicit production of serotype-specific antibody. The protein component was found to produce a band pattern in SDS-PAGE that is simple, reproducible, and strain dependent. The lipopolysaccharide component was found to have chemical and biological properties characteristic of bacterial endotoxin. We propose that this complex is representative of the outer trilaminar membrane of the meningococcal cell envelope in its native state.
Enhancement of Hemagglutination Inhibition by Complement
Authors: Reno Paul W. , Hoffmann Edward M.
Published: January 2020

Experiments were conducted to determine the effect of specific antibody and complement on hemagglutination inhibition of influenza virus. It was found that early immunoglobulin M antibody to influenza virus inhibited the hemagglutinating capacity of the virus. When fresh guinea pig serum was added, the inhibiting capacity of the serum was elevated from four- to eightfold When guinea pig serum was treated with known complement inhibitors, it lost its capacity to enhance hemagglutination. The use of functionally purified complement components indicated that the first, second, and fourth components were necessary and sufficient for enhancement of hemagglutination inhibition by specific antibody to influenza virus.
Chronic Rabies Virus Infection of Cell Cultures
Authors: Wiktor T. J. , Clark H F.
Published: January 2020

Exposure of both mammalian and reptilian cells in tissue culture to different strains of fixed rabies virus resulted in a carrier type of infection. No cytopathic effect was observed in either type of culture; infected cultures could be maintained by cell transfer for unlimited numbers of passages. A consistent pattern of cyclically rising and falling levels of viral infection was observed by fluorescent-antibody staining techniques and by titration of released infectious virus. Resistance to super-infection by vesicular stomatis virus and the production of an interferon-like substance by infected cells indicated that the maintenance of a carrier type of infection may be interferon-mediated. The degree of susceptibility of rabies-infected cells to immunolysis by antirabies antibody in the presence of complement was found to be correlated with the amount of virus maturation occurring by budding through the cell membrane and not with the presence of immunofluorescent antigen in the cytoplasm of infected cells.
Pneumocystis carinii induces an oxidative burst in alveolar macrophages
Authors: Hidalgo H A , Helmke R J , German V F , Mangos J A
Published: January 2020

There is evidence that alveolar macrophages (AM) play a role in the clearing of Pneumocystis carinii from the lungs. To investigate the mechanisms involved in this process, we studied in vitro the induction of an oxidative burst by P. carinii in a cell line of macrophages (NR8383) and AM from normal rats. P. carinii was added to macrophage monolayers (10(6) cells), and the H2O2 produced after 4 h of incubation was measured. Both NR8383 macrophages and normal rat AM produced H2O2 in response to P. carinii cysts and trophozoites isolated from dexamethasone-treated rats, although the amount of H2O2 induced in AM from normal rats was larger. NR8383 macrophages bound and phagocytized both P. carinii cysts and trophozoites and produced increasing amounts of H2O2 as a dose-related response to cysts and trophozoites. Opsonization of P. carinii with normal rat serum increased H2O2 production by both types of macrophages; this enhancement was decreased, but not abolished, when the serum was first depleted of complement by heat treatment. These findings demonstrate that NR8383 macrophages and normal rat AM produce an oxidative burst in response to P. carinii and that this response is enhanced by complement.
Mechanism of YadA-mediated serum resistance of Yersinia enterocolitica serotype O3
Authors: Pilz D , Vocke T , Heesemann J , Brade V
Published: January 2020

Complement activation via the alternative pathway was analyzed with isogenic strains of Yersinia enterocolitica serotype O3 differing in plasmid content (p- or p+ strains) or selective lack of YadA expression (YadA- strain). The p+ strain was serum resistant, even after antibody-enhanced complement activation. Serum sensitivity was observed with the p- and YadA- strains but was more pronounced in the p- strain. The p+ strain deposited less C5b-9(m) complexes on its surface than the p- and YadA- strains. No size difference, however, was detected with solubilized C5b-9(m) complexes obtained from resistant and sensitive strains. At the C3 level, it became evident that surface-bound C3b was degraded faster into iC3b on the p+ strain than on the p- and YadA- strains. Our results demonstrate that YadA inhibits complement activation at the C3 and C9 level. As a result, reduced amounts of C5b-9(m) are generated on the surface of YadA-bearing bacteria. In addition, YadA seems to protect against the lytic action of those C5b-9(m) complexes whose deposition could not be prevented.
Early hepatic stages of Plasmodium berghei: release of circumsporozoite protein and host cellular inflammatory response
Authors: Khan Z M , Ng C , Vanderberg J P
Published: January 2020

After injection of Plasmodium berghei sporozoites into Norway-Brown rats, we were able to localize these sporozoites and the early hepatic trophozoites developing from them in histological sections of the liver stained with a sensitive immunogold-silver procedure. Sporozoites invading hepatocytes released substantial quantities of circumsporozoite protein into the hepatocyte cytoplasm. This intrahepatic cytoplasmic distribution reached a maximal level at about 4 h post-sporozoite injection. As the hepatic parasites continued to differentiate, circumsporozoite protein became undetectable within the cytoplasm of the hepatocytes and became localized around the periphery of each parasite. There was generalized cellular inflammation within the liver of the host, which first became evident at around 4 h post-sporozoite injection and progressed to the formation of well-defined granulomas by 24 h. Such histopathological changes were not seen in rats injected with killed sporozoites, indicating that the cellular inflammation was induced by viable, infective sporozoites. We did not observe cellular infiltration specifically associated with any of the developing hepatic stages that we observed, even up to 28 h post-sporozoite inoculation. These results indicate that viable sporozoites induced rapid and generalized hepatic inflammation in host rats. However, sporozoites that successfully invaded hepatocytes and then proceeded to develop further did not appear to be the target of inflammatory cells until a period beginning at around 40 h post-sporozoite inoculation.
In vivo modulation of the murine immune response to Francisella tularensis LVS by administration of anticytokine antibodies
Authors: Leiby D A , Fortier A H , Crawford R M , Schreiber R D , Nacy C A
Published: January 2020

The role(s) of gamma interferon (IFN-gamma), tumor necrosis factor alpha (TNF-alpha), and interleukin-4 (IL-4) in establishment and maintenance of protective immunity to Francisella tularensis LVS in mice (C3H/HeN) was examined by selective removal of these cytokines in vivo with neutralizing antibodies. The 50% lethal dose (LD50) for mice infected intradermally with F. tularensis alone was 136,000 CFU; treatment of mice with anti-IFN-gamma or anti-TNF-alpha at the time of infection significantly reduced (P much less than 0.05) the LD50 to 2 and 5 CFU, respectively. Abrogation of protective immunity, however, was effective only when anti-IFN-gamma or anti-TNF-alpha was administered prior to day 3 postinfection. In contrast, the LD50 for mice treated with anti-IL-4 was repeatedly higher (555,000 CFU) than for controls; this difference, however, was not significant (P greater than 0.05). Thus, IL-4 may be detrimental, while IFN-gamma and TNF-alpha were clearly crucial to the establishment of protective immunity to F. tularensis during a primary infection. The importance of IFN-gamma and TNF-alpha during a secondary immune response to F. tularensis was also investigated. Spleen cells from immune mice passively transfer protective immunity to recipient mice in the absence of confounding antibody-mediated immunity. This passive transfer of immunity, however, was abrogated by treatment of recipient mice with anti-IFN-gamma or anti-TNF-alpha at the time of challenge infection. That anticytokines effectively abrogate protective immunity very early in the course of infection with F. tularensis suggests that T-cell-dependent activation of macrophages for microbicidal activity is unlikely. These T-cell-independent events early in the course of infection may suppress bacterial replication until a T-cell-dependent response ultimately clears the bacteria.
Characterization of a Salmonella typhimurium aro vaccine strain expressing the P.69 antigen of Bordetella pertussis
Authors: Strugnell R , Dougan G , Chatfield S , Charles I , Fairweather N , Tite J , Li J L , Beesley J , Roberts M
Published: January 2020

The P.69 Bordetella pertussis protective antigen was expressed by use of the trc promoter from the chromosome of a Salmonella typhimurium aro vaccine strain, BRD509, by integrating the prn gene, encoding the 93-kDa precursor of this protein, into the aroC locus. P.69 was detected on the cell surface of the S. typhimurium strain (BRD640) by agglutination and immunoelectron microscopy. BALB/c mice immunized orally or intravenously with BRD640 showed a significant level of protection against an aerosol challenge with virulent B. pertussis, compared with control animals. No anti-P.69 antibodies in the serum or anti-P.69 antibody-secreting cells in the lungs were detected in BRD640-vaccinated animals, although cells isolated from spleens showed a P.69-dependent cell proliferative response. In contrast, low levels of anti-P.69 antibodies in the serum and anti-P.69 antibody-secreting cells in the lungs were detected in immunized mice following a B. pertussis challenge.
Early expression of cytokine mRNA in mice infected with Listeria monocytogenes
Authors: Iizawa Y , Brown J F , Czuprynski C J
Published: January 2020

Protective immunity first becomes evident at 3 to 4 days after inoculation of mice with a sublethal dose of Listeria monocytogenes. Recent evidence suggests that production of gamma interferon (IFN-gamma) occurs earlier (within the first 24 h of infection). The purpose of this study was to define better the sequence of cytokine mRNA expression during the early stages of L. monocytogenes infection. Cytokine mRNA expression was detected by polymerase chain reaction-assisted amplification of RNA extracted from the spleen cells of individual mice euthanized at 0.5 to 120 h after L. monocytogenes challenge. By using this method, mRNAs for tumor necrosis factor alpha, interleukin-1 alpha (IL-1 alpha), IL-2, IL-4, IL-5, and IFN-gamma were detected in RNA from the spleen cells of uninfected mice. The intensity of the bands for IFN-gamma, however, was increased greatly at 16 h after intravenous injection of 5 x 10(4) CFU (nearly 1 50% lethal dose) of L. monocytogenes. IL-6 and granulocyte-macrophage colony-stimulating factor mRNAs were not detected in spleen cell RNA from uninfected mice but were induced within 30 and 60 min, respectively, after inoculation with L. monocytogenes. Increased amounts of mRNAs for IFN-gamma, IL-6, and granulocyte-macrophage colony-stimulating factor were detected after injection of viable, but not killed, L. monocytogenes. IL-3 mRNA was not detected at any time in RNA extracted from the spleen cells of uninfected or L. monocytogenes infected mice. These results suggest that infection with L. monocytogenes elicits a detectable cytokine mRNA response within the first few hours of infection.
Structural identification of an epitope of antigenic factor 5 in mannans of Candida albicans NIH B-792 (serotype B) and J-1012 (serotype A) as beta-1,2-linked oligomannosyl residues
Authors: Shibata N , Arai M , Haga E , Kikuchi T , Najima M , Satoh T , Kobayashi H , Suzuki S
Published: January 2020

In previous articles, we reported the presence of phosphate-bound beta-1,2-linked oligomannosyl residues in the mannans of strains of Candida albicans serotypes A and B and Candida stellatoidea. To identify the antigenic factor corresponding to this type of oligomannosyl residue, a relationship between chemical structure and antigenic specificity in the mannans of C. albicans NIH B-792 (serotype B, B-strain) and C. albicans J-1012 (serotype A, J-strain) was investigated by using a combination of two-dimensional 1H nuclear magnetic resonance spectroscopy of H-1, H-2, and H-5 regions in the mannans and an enzyme-linked immunosorbent assay that employed concanavalin A-coated microtiter plates. It was shown in the present 1H nuclear magnetic resonance study that an examination of chemical shifts not only in the H-1 region but also in the H-5 region was useful for the quantitative determination of the phosphate-bound beta-1,2-linked oligomannosyl residues. In the enzyme-linked immunosorbent assay using concanavalin A-coated plates, it was revealed that, of factor sera 1, 4, and 5, only factor serum 5 showed a reactivity proportional to the densities of the beta-1,2-linked oligomannosyl residues of the mannan subfractions of different phosphate contents that had been prepared from the bulk B-strain mannan by DEAE-Sephadex chromatography. The above results indicate that the phosphate-bound beta-1,2-linked oligomannosyl residues, Manp beta 1----(2Manp beta 1----)n2Man (n = 0-5), correspond to antigenic factor 5.
Staphylococcus saprophyticus hemagglutinin is a 160-kilodalton surface polypeptide
Authors: Gatermann S , Meyer H G , Wanner G
Published: January 2020

Many strains of Staphylococcus saprophyticus cause direct hemagglutination of sheep erythrocytes. For a high proportion of clinical isolates, a surface protein (Ssp) that is apparently not involved in this property has been described. In this study, S. saprophyticus CCM883, a hemagglutinating but Ssp-negative strain, was used for the identification, purification, and characterization of a 160-kDa surface polypeptide that appears to be the major component of the hemagglutinin. Expression of the protein required the addition to the growth medium of EDTA in micromolar quantities, suggesting an inhibitory role for some unidentified metal ion. The protein was purified by means of Sephacryl S-300 chromatography, and antisera were raised in rabbits. Antibody against this protein inhibited the hemagglutination of two other, unrelated strains and was used to demonstrate, by electron microscopy, the presence of the protein on the surface of the cells. In a confirmatory experiment, the purified antigen was incubated with erythrocytes and binding was detected by the Western immunoblot technique with the antibody to the 160-kDa polypeptide. These experiments indicate that this surface protein is the hemagglutinin of S. saprophyticus.
Killing of Coccidioides immitis by human peripheral blood mononuclear cells
Authors: Ampel N M , Bejarano G C , Galgiani J N
Published: January 2020

The ability of human peripheral blood mononuclear cells (MNL) obtained from healthy donors to kill the fungus Coccidioides immitis was examined in vitro with an assay that uses a single fungal particle per well. MNL killed 25.0% +/- 3.5% of a coccidioidal arthroconidial target, compared with the 4.7% +/- 2.9% killed by polymorphonuclear leukocytes obtained from the same donors (P = 0.012). Arthroconidial killing by MNL was not dependent on donor delayed dermal hypersensitivity to spherulin. Killing of another fungal target, Candida glabrata, was not significantly different between MNL and polymorphonuclear leukocytes (P = 0.783). Depletion of monocytes from MNL with Sephadex G-10 resulted in a significant reduction in arthroconidial killing (21.4% +/- 13.6% versus 2.4% +/- 3.4%; P = 0.025), while enrichment of monocytes by Percoll density gradient centrifugation or plastic adherence resulted in significantly increased arthroconidial killing compared with that by MNL (P = 0.005 and 0.001, respectively). Killing of 96-h spherules by MNL was 7.3% +/- 3.1%, significantly less than the 21.4% +/- 2.8% killing of arthroconidia in the same experiments (P = 0.016). Incubation of MNL with human recombinant gamma interferon or tumor necrosis factor alpha did not result in increased MNL killing of coccidioidal arthroconidia under various conditions. These results suggest that MNL have an inherent ability to kill coccidioidal arthroconidia in vitro which is not dependent on prior host exposure to C. immitis. This activity appears to reside in peripheral blood monocytes.
A seroreactive 120-kilodalton beta-1,3-glucanase of Coccidioides immitis which may participate in spherule morphogenesis
Authors: Kruse D , Cole G T
Published: January 2020

A beta-glucosidase of Coccidioides immitis was identified in electrophoresis gel separations of the concanavalin A-bound mycelial culture-filtrate-plus-lysate preparation. p-Nitrophenol-beta-D-glucopyranoside was used as the substrate to visualize the enzymatically active fraction in nonreducing gels. The gel-isolated, chromatographically purified enzyme has an optimal pH of 8.0 and cleaves beta-1,3-glycosyl linkages. The alkaline beta-glucosidase was further characterized by a pI of 3.8 to 4.0, optimal activity at 37 to 40 degrees C, and molecular size of 120 kDa as identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The purified beta-glucosidase is identical to a previously reported 120-kDa antigen (Ag) which reacts with immunoglobulin M (IgM) tube precipitin (TP) antibody in sera from patients with coccidioidomycosis. The TP-Ag was described as a valuable serodiagnostic reagent for detection of specific IgM in patients with early coccidioidal infections. The beta-glucosidase, like the TP-Ag, was localized in the cell wall and cytoplasmic vesicles of parasitic cells (spherules) by immunofluorescence and immunoelectron microscopy with specific antiserum raised against the purified enzyme. The boiled cell wall fraction isolated from these same young (presegmented) spherules was partially digested by the beta-glucosidase. Addition of a potent beta-glucosidase inhibitor, 1-deoxynojirimycin, to the parasitic-phase culture medium at a concentration of 200 microM blocked or retarded conversion of arthroconidia to spherules. Antibody was raised in guinea pigs against chromatographically purified 1-deoxynojirimycin which was conjugated with bovine serum albumin. The inhibitor was localized by immunofluorescence in the wall of the 1-deoxynojirimycin-treated cells. We suggest that the spherule wall-associated, alkaline hydrolase functions as a beta-1,3-glucanase to provide for wall plasticity as well as intussusception of newly synthesized wall polymers during the period of rapid diametric growth of parasitic cells of C. immitis.
Bovine helper T cell clones recognize five distinct epitopes on Babesia bovis merozoite antigens
Authors: Brown W C , Zhao S , Rice-Ficht A C , Logan K S , Woods V M
Published: January 2020

Helper T cell clones from two Babesia bovis-immune cattle were characterized for use in identification of potentially protective immunogens of B. bovis merozoites. Proliferation assays with 11 CD4+ clones revealed a differential pattern of response to soluble cytosolic antigen, membrane-enriched antigen, detergent extracts of the membrane-enriched antigen, soluble culture supernatant exoantigen, and different geographical isolates of B. bovis as well as Babesia bigemina parasites. When the data were combined, the clones could be grouped according to five different patterns of response. One group recognized only the membrane-enriched fraction of New World and Australian parasites. Four remaining groups recognized antigens found in the cytosolic as well as the membrane-enriched fraction, and clones representative of each group were used to identify cytosolic antigens fractionated by anion-exchange chromatography with the use of fast-performance liquid chromatography. One clone (C97.3C3), which responded to all B. bovis isolates and to B. bigemina, recognized a single peak of activity that eluted with 0.25 M NaCl and contained protein bands of 70 and 75 kDa. The remaining clones were stimulated by a second antigenic peak that eluted between 0.35 and 0.45 M NaCl and contained protein bands of 42, 47, 56, and 84 kDa. The majority of the clones produced interferon, whereas tumor necrosis factor alpha/tumor necrosis factor beta production was less frequent. These studies provide the basis for using helper T cell clones to identify potentially protective immunogens of B. bovis and delineate a minimum of five helper T cell epitopes recognized by two immune cattle.
Identification and characterization of epitopes shared between the mycobacterial 65-kilodalton heat shock protein and the actively secreted antigen 85 complex: their in situ expression on the cell wall surface of Mycobacterium leprae
Authors: Rambukkana A , Das P K , Burggraaf J D , Faber W R , Teeling P , Krieg S , Thole J E , Harboe M
Published: January 2020

Both mycobacterial hsp65 and the actively secreted antigen 85 complex of 30-kDa region proteins are considered to be major immune targets in mycobacterial diseases. In this study, by using a novel series of monoclonal antibodies (MAbs) directed to these antigens, we identified and partially characterized three unique epitopes (Rb2, Pe12, and A2h11) that are shared between mycobacterial hsp65 and the individual components of the antigen 85 complex. Dot blot assays with native purified proteins revealed that all three MAbs are strongly bound to hsp65 and antigens 85A (MPT44) and 85B (MPT59), while a weak reaction or no reaction was found with antigen 85C (MPT45). Immunoblotting showed that MAb Rb2 reacted strongly with both hsp65 and the antigen 85 complex proteins, whereas MAbs Pe12 and A2h11 reacted strongly with the former but weakly with the latter. Moreover, these MAbs did not react with other closely related MPT51 and MPT64 secreted proteins. Further characterization of these epitopes was performed by using recombinant fusion and truncated proteins of Mycobacterium bovis BCG hsp65 (MbaA) and the M. leprae 30- and 31-kDa antigen 85 complex fusion proteins. In hsp65, Rb2-Pe12- and A2h11-reactive epitopes were found to reside in the C-terminal region of amino acid residues 479 to 540 and 303 to 424, respectively. In the M. leprae 30- and 31-kDa antigen 85 complex, all three epitopes were located in an N-terminal region of amino acid residues 55 to 266, one of the known fibronectin-binding sites of the M. leprae antigen 85 complex. Comparison of these MAb-reactive amino acid sequence regions between mycobacterial hsp65 and the components of the antigen 85 complex revealed that these regions show certain amino acid sequence identities. Furthermore, by immunoperoxidase and immunogold ultracytochemistry, we demonstrated that Rb2-, Pe12-, and A2h11-reactive epitopes are expressed both on the cell wall surface and in the cytosol of M. leprae bacilli within the lesions of lepromatous leprosy patients and in M. leprae-infected armadillo liver tissue.
Immunochemical and biological characterization of outer membrane proteins of Porphyromonas endodontalis
Authors: Ogawa T , Kuribayashi S , Shimauchi H , Toda T , Hamada S
Published: January 2020

Outer membrane proteins (OMP) of Porphyromonas endodontalis HG 370 (ATCC 35406) were prepared from the cell envelope fraction of the organisms. The cell envelope that had been obtained by sonication of the whole cells was extracted in 2% lithium dodecyl sulfate and then successively chromatographed with Sephacryl S-200 HR and DEAE-Sepharose Fast Flow. Two OMP fractions, OMP-I and OMP-II, were obtained, and their immunochemical properties and induction of specific antibodies were examined. The OMP-I preparation consisted of a major protein with an apparent molecular mass of 31 kDa and other moderate to minor proteins of 40.3, 51.4, 67, and 71.6 kDa, while the OMP-II preparation contained 14-, 15.5-, 27-, and 44-kDa proteins as revealed by sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis. OMP-I was found to form hydrophilic diffusion pores by incorporation into artificial liposomes composed of egg yolk phosphatidylcholine and dicetylphosphate, indicating that OMP-I exhibited significant porin activity. However, the liposomes containing heat-denatured OMP-I were scarcely active. Spontaneous and antigen-specific immunoglobulin M (IgM)-, IgG-, and IgA-secreting spot-forming cells (SFC) enzymatically dissociated into single-cell suspensions from chronically inflamed periapical tissues and were enumerated by enzyme-linked immunospot assay. In patients with radicular cysts or dental granulomas, the major isotype of spontaneous SFC was IgG. In radicular cysts, the OMP-II-specific IgG SFC represented 0.13% of the total IgG SFC, while the antigen-specific IgA or IgM SFC was not observed. It was also found that none of these mononuclear cells produced antibodies specific for OMP-I or lipopolysaccharide of P. endodontalis.
Proliferative responses and gamma interferon and tumor necrosis factor production by lymphocytes isolated from tracheobroncheal lymph nodes and spleen of mice aerosol infected with Bordetella pertussis
Authors: Petersen J W , Ibsen P H , Hasløv K , Heron I
Published: January 2020

A group of mice was aerosol infected with live, virulent Bordetella pertussis bacteria. During a period of 7 weeks following the infection, with intervals of 1 week, lymphocytes were isolated from the tracheobroncheal lymph nodes (TBL) and the spleens (SPL) of the infected mice. The in vitro proliferative responses as well as the gamma interferon and tumor necrosis factor production levels of the isolated lymphocytes in response to stimulation with whole killed B. pertussis bacteria were measured as parameters for cell-mediated immunity (CMI). The course of the infection was monitored by counting of CFU in the lungs of the mice. Moreover, antibody responses in serum against a range of B. pertussis antigens were assessed. The results showed that a vigorous proliferative response of the TBL and SPL to stimulation with whole killed B. pertussis bacteria was induced by the infection. The proliferative response of the TBL was significantly higher than the response of the SPL. The proliferative responses were maximal 3 to 4 weeks after the infection and were paralleled by in vitro gamma interferon and tumor necrosis factor production upon specific stimulation. The development of the CMI was observed simultaneously with the clearance of the infection from the lungs. Antibody responses became measurable in the sera only after the infection was cleared. A specific CMI against pertussis toxin, the filamentous hemagglutinin, the 69-kDa outer membrane protein, and the agglutinogens 2 and 3, antigens which are under consideration for inclusion in future acellular pertussis vaccines, was successfully demonstrated in mice 3 weeks after the infection.
Identification of T- and B-cell epitopes of the S2 and S3 subunits of pertussis toxin by use of synthetic peptides
Authors: Chong P , Zobrist G , Sia C , Loosmore S , Klein M
Published: January 2020

To design an optimized synthetic vaccine against whooping cough, we have studied the biological and immunological properties of three peptides of the S2 subunit and nine overlapping synthetic peptides covering the entire sequence of the S3 subunit of pertussis toxin (PT). Synthetic peptides corresponding to sequences 18 to 41, 78 to 108, 134 to 154, and 149 to 176 of S3 were found to be consistently capable of stimulating the proliferation of PT-specific T-cell lines primed with pertussis toxoid in both BALB/c and A/J strains of mice. All synthetic peptides were recognized by rabbit antisera raised against PT or pertussis toxoid. Both S2 and S3 peptide-keyhole limpet hemocyanin (KLH) conjugates in the presence of complete Freund's adjuvant induced peptide-specific antibody responses in rabbits, and the antisera raised against S2(1-23), S3(18-41), S3(37-64), and S3(149-176) peptide-KLH conjugates cross-reacted with both subunits in the immunoblots. All antisera except those against S2(123-154) and S3(103-127) reacted with native PT in an enzyme-linked immunosorbent assay (ELISA) with PT directly coated onto microtiter wells. In contrast, antisera raised against S2(123-154), S3(1-23), S3(18-41), S3(37-64), S3(60-87), and S3(103-127) peptide-KLH conjugates recognized native PT in a fetuin-PT capture ELISA. S2(78-98), S3(1-23), and S3(149-176) peptide-KLH conjugates elicited good PT-neutralizing antibody responses as judged by the antitoxin CHO cell assay. Identification of these B-cell neutralization epitopes and T-cell immunodominant determinants represents a first step towards the rational design of a synthetic vaccine against whooping cough.
Purification and characterization of a peptide essential for formation of streptolysin S by Streptococcus pyogenes
Authors: Akao T , Takahashi T , Kobashi K
Published: January 2020

Peptides in a pronase digest of bovine serum albumin were required for streptolysin S formation by Streptococcus pyogenes besides maltose and a carrier (the oligonucleotide fraction obtained by treatment of Saccharomyces cerevisiae RNA with RNase A). A peptide essential for streptolysin S formation was purified to homogeneity from a pronase digest of bovine serum albumin by Sephadex G-25 column chromatography, and anion-exchange, reverse-phase, and gel filtration high-performance liquid chromatography. The purified peptide was divided into more than two peptides by HCOOOH oxidation and was composed of four residues of cysteine, three of leucine, and one each of aspartic acid and glutamic acid. Leucine and cysteine were detected as amino-terminal residues, and leucine and glutamic acid were detected as carboxyl-terminal residues, suggesting that two or three peptides are linked by a disulfide bond(s). A disulfide bond structure in the peptide seemed to be required for streptolysin S formation.
Human neutrophil azurocidin synergizes with leukocyte elastase and cathepsin G in the killing of Capnocytophaga sputigena
Authors: Miyasaki K T , Bodeau A L
Published: January 2020

Azurocidin was purified in the presence of phenylmethylsulfonyl fluoride. Electrophoresis revealed at least seven species which exhibited N-terminal sequences consistent with azurocidin. Azurocidin exhibited no bactericidal activity against Capnocytophaga sputigena or other oral bacteria but synergized the bactericidal activity of enzymatically active elastase. Azurocidin also interacted synergistically with cathepsin G.
Phagolysosomes of Coxiella burnetii-infected cell lines maintain an acidic pH during persistent infection
Authors: Maurin M , Benoliel A M , Bongrand P , Raoult D
Published: January 2020

Coxiella burnetii, the agent of Q fever, is an obligate intracellular bacterium that multiples within vacuoles of phagolysosomal origin. Persistently infected cell lines were maintained in continuous culture for months. We studied the pH of the phagolysosomes by using two murine cell lines during early propagation of the bacteria and after establishment of persistent infection. Three strains of C. burnetii were studied because of the purported propensity of each strain to cause acute or chronic disease and to be resistant or susceptible to antibiotics. The pHs were calculated from fluorescence experiments with fluoresceinated dextran as a lysosomal probe. Phagolysosomal vacuoles maintained an acidic pH during a 36-day infection. Minimal variation of the pH occurred over the duration of the experiment with strains that caused either acute or chronic disease. Phagolysosomal pH remained stable for as long as 153 days with the Nine Mile phase II isolate. Thus, neither the course of C. burnetti infection nor the diversity of antibiotic susceptibility of the strains is related to variations in the phagolysosomal pH.
Role of fibrinogen in complement inhibition by streptococcal M protein
Authors: Horstmann R D , Sievertsen H J , Leippe M , Fischetti V A
Published: January 2020

M protein, the major virulence factor of group A streptococci, has antiopsonic activity in that it inhibits activation of the alternative complement pathway on the streptococcal surface. Two properties of M protein have been claimed to account for the inhibitory activity, namely, (i) its binding affinity for complement factor H, which is an inhibitor of alternative pathway activation, and (ii) its high binding affinity for fibrinogen. We have recently shown that fibrinogen, like M protein, inhibits alternative pathway activation by possessing binding affinity for factor H. Here we report that fibrinogen effectively competes with factor H for binding to M protein but retains its own binding affinity for factor H. The presence of fibrinogen did not significantly affect alternative pathway inhibition on the streptococcal surface.
Characterization of a polysaccharide capsular antigen of septicemic Escherichia coli O115:K "V165" :F165 and evaluation of its role in pathogenicity
Authors: Ngeleka M , Harel J , Jacques M , Fairbrother J M
Published: January 2020

Escherichia coli strains of serogroup O115:K(-):F165 have been associated with septicemia in calves and piglets. These strains express a capsular antigen referred to as K"V165" which inhibits agglutination of the O antigen by anti-O115 serum. We used hybrid transposon TnphoA mutants M48, 18b, and 2, and a spontaneous O-agglutinable mutant, 5131a, to evaluate the role of K"V165" in the pathogenicity of E. coli O115. Mutant M48 was as resistant to 90% rabbit serum and as virulent in day-old chickens as the parent strain 5131, mutants 18b and 5131a were less resistant to serum and less virulent in chickens, and mutant 2 was serum sensitive and avirulent. Analysis of outer membrane protein and lipopolysaccharide profiles failed to show any difference between the transposon mutants and the parent strain. In contrast, the spontaneous O-agglutinable mutant showed additional bands in the 16-kDa region of the polysaccharide ladder-like pattern. Mutants 2 and 5131a produced significantly less K"V165" capsular antigen than the parent strain, as demonstrated by a competitive enzyme-linked immunosorbent assay with adsorbed anti-K"V165" serum. In addition, electron microscopic analysis revealed that mutants 2 and 5131a had lost the capsular layer observed in the parent strain after fixation with glutaraldehyde-lysine. This capsule contained carbohydrate compounds and resembled an O-antigen capsule since it prevented O-antigen agglutination before the bacteria were heated at 100 degrees C and induced bacterial serum resistance. The capsule-defective mutants colonized the intestinal epithelium of experimentally infected gnotobiotic pigs but failed to induce clinical signs of septicemia. We concluded that E. coli strains of serogroup O115 expressed a polysaccharide capsular antigen which induced serum resistance and consequently contributed to the pathogenicity of the bacteria.