bluuebunny/crossref_metadata_2025_split huggingface dataset:
Immunogenicity of aSalmonella typhimurium aroA aroDVaccine Expressing a Nontoxic Domain ofClostridium difficileToxin A
Authors: Ward Stephen J. , Douce Gill , Figueiredo Dayse , Dougan Gordon , Wren Brendan W. 
Published: December 2019

ABSTRACTThe C-terminal repeat domain ofClostridium difficiletoxin A harbors toxin-neutralizing epitopes and is considered to be a candidate component of a vaccine againstC. difficile-associated disease (CDAD). Fourteen of the 38 C-terminal toxin A repeats (14CDTA) were cloned into pTECH-1 in frame with the immunogenic fragment C of tetanus toxin (TETC) to generate plasmid p56TETC. Expression of the TETC-14CDTA fusion protein was driven from the anaerobically induciblenirBpromoter within attenuatedSalmonella typhimuriumBRD509 (aroA aroD). The TETC-14CDTA fusion protein was purified and shown to bind to known toxin A receptors found on the surface of rabbit erythrocytes. Intranasal (i.n.) and intragastric (i.g.) immunization with 107and 1010CFU, respectively, of BRD509(p56TETC) generated significant (P< 0.05) anti-toxin A serum responses after a single dose. Antibody titers were elevated following a boosting dose with either live vaccine or a subcutaneous injection of 0.5 μg of purified 14CDTA protein. Importantly, serum from mice immunized with BRD509(p56TETC) neutralized toxin A cytotoxicity. Both i.n. and i.g. immunizations also generated toxin A-specific immunoglobulin A on the pulmonary and intestinal mucosa, respectively. Intranasal vaccination induced consistently higher serum and mucosal anti-toxin A antibody responses. Significant anti-tetanus toxoid serum and mucosal antibodies were also generated by both immunization routes. The availability of live attenuatedSalmonella typhifor human use may allow the development of a multivalent mucosal vaccine against CDAD, tetanus, and typhoid.
Transient Transcriptional Activation of the
Vibrio cholerae
El Tor Virulence Regulator ToxT in Response to Culture Conditions
Authors: Medrano Ana I. , DiRita Victor J. , Castillo Gabriela , Sanchez Joaquin 
Published: December 2019

ABSTRACT

Vibrio cholerae
El Tor require special in vitro culture conditions, consisting of an initial static growth period followed by shift to shaking (AKI conditions), for expression of cholera toxin (CT) and toxin coregulated pili (TCP). ToxT, a regulator whose initial transcription depends on the ToxR regulator, positively modulates expression of CT and TCP. To help understand control of CT and TCP in El Tor vibrios, we monitored
ctxAB
and ToxR-dependent
toxT
transcription by time course primer extension assays. AKI conditions stimulated CT synthesis with an absence of
ctxAB
transcription during static growth followed by induction upon shaking. ToxR-dependent
toxT
transcription was induced at the end of the static growth period but was transient, stopping shortly after shaking was initiated but, interestingly, also if the static phase was prolonged. Immunoblot assays showed that ToxR protein levels were not coincidentally transient, implying a protein on/off switch mechanism for ToxR. Despite the transient activation by ToxR, transcription of
ctxAB
was maintained during shaking. This finding suggested continued
toxT
expression, possibly through relay transcription from another promoter. The 12.6-kb distant upstream
tcpA
promoter responsible for expression of the TCP operon has been proposed to provide an alternate
toxT
message by readthrough transcription. Activation of the
tcpA
promoter is supported by increased expression of TcpA protein during the shaking phase of the culture. Readthrough transcription of
toxT
from
tcpA
would be compatible with reverse transcription-PCR evidence for a
toxT
mRNA at times when ToxR-dependent transcription was no longer detectable by primer extension.
Differential Expression ofVibrio vulnificusCapsular Polysaccharide
Authors: Wright Anita C. , Powell Jan L. , Tanner Mike K. , Ensor Lynne A. , Karpas Arthur B. , Morris J. Glenn , Sztein Marcelo B. 
Published: December 2019

ABSTRACTVibrio vulnificusis a human pathogen whose virulence has been associated with the expression of capsular polysaccharide (CPS). Multiple CPS types have been described; however, virulence does not appear to correlate with a particular CPS composition. Reversible-phase variation for opaque and translucent colony morphologies is characterized by changes in CPS expression, as suggested by electron microscopy of cells stained nonspecifically with ruthenium red. Isolates with opaque colony morphologies are virulent and appear to be more thickly encapsulated than naturally occurring translucent-phase variants, which have reduced, patchy, or absent CPS. Previously, we have shown that the virulence of translucent-phase variants was intermediate between opaque-phase variants and acapsular transposon mutants, suggesting a correlation between virulence and the amount of CPS expressed. In the present study, CPS expression of phase variants and genetically defined mutants ofV. vulnificusM06-24/O was examined by using a CPS-specific monoclonal antibody with an enzyme-linked immunosorbent assay, flow cytometry, and immunoelectron microscopy. Semiquantitative analyses of CPS expression correlated well among these assays, confirming that the translucent-phase variant was intermediate in CPS expression and retained type I CPS-specific epitopes. Cell surface expression of CPS varied with the growth phase, increasing during logarithmic growth and declining in stationary culture. Significantly greater CPS expression (P= 0.026) was observed for cells grown at 30°C than for those at 37°C. These studies confirm that phase variation and virulence inV. vulnificuscorrelate with the amount of CPS expressed and demonstrate the fluidity of bacterial polysaccharide expression in response to environmental conditions.
Antibacterial Action of Extracellular Mammalian Group IIA Phospholipase A2 against Grossly Clumped
Staphylococcus aureus
Authors: Dominiecki Mary E. , Weiss Jerrold 
Published: December 2019

ABSTRACT

Fibrinogen-dependent interactions of
Staphylococcus aureus
are believed to contribute to bacterial virulence by promoting bacterial attachment to fibrinogen-coated surfaces and inducing the formation of bacterial clumps that are likely resistant to phagocytosis. Although
S. aureus
produces several fibrinogen-binding proteins, the cell wall-associated protein clumping factor (encoded by
clfA
) appears to be most important in bacterial interactions with immobilized or soluble purified fibrinogen. We have compared bacterial clumping in several strains of
S. aureus
, including isogenic ClfA
+
and ClfA
−
Newman strains, in the presence of purified rabbit fibrinogen, human plasma, and inflammatory fluid and examined the effect of clumping on bacterial sensitivity to mammalian group IIA phospholipase A2 (PLA2). This enzyme is the major extracellular bactericidal agent in inflammatory fluid active against
S. aureus
. Both ClfA-dependent and ClfA-independent bacterial clumping was observed, depending on the source and fibrinogen content of the biological fluid. In each case, clumping only partially reduced the antibacterial activity of PLA2, suggesting that this extracellular enzyme can substantially penetrate dense bacterial clumps. Bacterial clumps could be dispersed by added proteases, restoring full antibacterial activity to PLA2. Thus, the extracellular mobilization of group IIA PLA2 during inflammation may provide a mechanism by which the host can control the proliferation and survival of
S. aureus
even after bacterial clumping.
Interleukin-12 as an Adjuvant for an Antischistosome Vaccine Consisting of Adult Worm Antigens: Protection of Rats from Cercarial Challenge
Authors: Bungiro Richard D. , Goldberg Miriam , Suri Parmjeet K. , Knopf Paul M. 
Published: December 2019

ABSTRACTOur group previously demonstrated that a detergent extract (fraction S3) prepared from immature (4-week)Schistosoma mansoniparasites can induce partial, serum-transferable immunity to challenge infection in rats when administered as an alum precipitate. In the present study, we examined whether S3 prepared from adult (7-week) worms could similarly induce protection and whether immunity could be positively influenced by treatment with interleukin-12 (IL-12). IL-12 coadministered to Fischer rats and C57BL/6 mice at the time of S3 vaccination altered the prechallenge kinetics of S3-specific antibody titers in both species, ultimately leading to a stable enhancement of titers (relative to those in animals vaccinated without IL-12) in mice but not rats. Immunoblot analysis of prechallenge immune sera demonstrated that IL-12 treatment was associated with changes in the S3 antigen recognition profile in each species. Isotyping of specific antibodies in S3- plus IL-12-vaccinated mice prior to challenge infection revealed a moderate elevation in immunoglobulin G1 (IgG1) responses, strongly enhanced IgG2a and IgG2b responses, as well as diminished total serum IgE responses compared to those in mice given S3 only. In vaccinated rats, IL-12 profoundly suppressed specific IgG1 and enhanced IgG2b responses but did not affect IgG2a responses. S3- plus IL-12-vaccinated rats also produced less total IgE upon challenge infection. Enumeration of worm burdens revealed that vaccination with S3 plus IL-12 conferred 50% protection from cercarial challenge to rats, whereas rats given S3 only were not protected; mice were not protected by S3 vaccination regardless of IL-12 coadministration. The protection observed in S3- plus IL-12-vaccinated rats could not be transferred with serum, suggesting participation of an activated cellular component in the expression of immunity.
Gamma Interferon Production Is Critical for Protective Immunity to Infection with Blood-StagePlasmodium bergheiXAT but Neither NO Production nor NK Cell Activation Is Critical
Authors: Yoneto Toshihiko , Yoshimoto Takayuki , Wang Chrong-Reen , Takahama Yasuhiro , Tsuji Moriya , Waki Seiji , Nariuchi Hideo 
Published: December 2019

ABSTRACTWe have examined the roles of gamma interferon (IFN-γ), nitric oxide (NO), and natural killer (NK) cells in the host resistance to infection with the blood-stage malarial parasitePlasmodium bergheiXAT, an irradiation-induced attenuated variant of the lethal strainP. bergheiNK65. Although the infection withP. bergheiXAT enhanced NK cell lytic activity of splenocytes, depletion of NK1.1+cells caused by the treatment of mice with anti-NK1.1 antibody affected neither parasitemia nor IFN-γ production by their splenocytes. TheP. bergheiXAT infection induced a large amount of NO production by splenocytes during the first peak of parasitemia, whileP. bergheiNK65 infection induced a small amount. Unexpectedly, however, mice deficient in inducible nitric oxide synthase (iNOS−/−) clearedP. bergheiXAT after two peaks of parasitemia were observed, as occurred for wild-type control mice. Although the infected iNOS−/−mouse splenocytes did not produce a detectable level of NO, they produced an amount of IFN-γ comparable to that produced by wild-type control mouse splenocytes, and treatment of these mice with neutralizing anti-IFN-γ antibody led to the progression of parasitemia and fatal outcome. CD4−/−mice infected withP. bergheiXAT could not clear the parasite, and all these mice died with apparently reduced IFN-γ production. Furthermore, treatment with carrageenan increased the susceptibility of mice toP. bergheiXAT infection. These results suggest that neither NO production nor NK cell activation is critical for the resistance toP. bergheiXAT infection and that IFN-γ plays an important role in the elimination of malarial parasites, possibly by the enhancement of phagocytic activity of macrophages.
Pseudomonas aeruginosa
Hemolytic Phospholipase C Suppresses Neutrophil Respiratory Burst Activity
Authors: Terada Lance S. , Johansen Kristine A. , Nowbar Sogol , Vasil Adriana I. , Vasil Michael L. 
Published: December 2019

ABSTRACT

Pseudomonas aeruginosa
is a persistent pathogen in the airways of patients with cystic fibrosis or bronchiectasis from other causes and appears to have evolved strategies to survive the inflammatory response of the host. We hypothesized that the secreted hemolytic phospholipase C (PLC) of
P. aeruginosa
(PlcHR) would decrease neutrophil respiratory burst activity. We found that while intact wild-type
P. aeruginosa
cells stimulated moderate respiratory burst activity from human neutrophils, an isogenic mutant pseudomonas (ΔHR strain) containing a targeted deletion of the
plcHR
operon induced a much more robust oxidative burst from neutrophils. In contrast, a second pseudomonas mutant (ΔN) containing a disruption in the gene encoding the nonhemolytic PLC (PlcN) was not different from the wild type in stimulating neutrophil O
2
·−
production. Readdition of purified PlcHR to the ΔHR strain suppressed neutrophil O
2
·−
production to levels stimulated by wild-type bacteria. Interestingly, purified PlcHR decreased phorbol myristate acetate (PMA)- but not formyl methionyl-leucyl-proline (fMLP)-induced respiratory burst activity, suggesting interference by PlcHR with a protein kinase C (PKC)-specific signaling pathway. Accordingly, the PKC inhibitor bisindolylmaleimide inhibited the oxidative burst induced by either PMA or intact pseudomonas, but not by fMLP, whereas the p38 kinase inhibitor SB-203580 fully inhibited the respiratory burst induced by fMLP or the PlcHR-replete wild-type bacteria, but not PMA or the PlcHR-deficient ΔHR bacterial mutant. We conclude that expression of PlcHR by
P. aeruginosa
suppresses bacterium-induced neutrophil respiratory burst by interfering with a PKC-dependent, non-p38 kinase-dependent pathway.
Cellular Immune Responses toNeisseria meningitidisin Children
Authors: Pollard Andrew J. , Galassini Rachel , Rouppe van der Voort Eileene M. , Hibberd Martin , Booy Robert , Langford Paul , Nadel Simon , Ison Catherine , Kroll J. Simon , Poolman Jan , Levin Michael 
Published: December 2019

ABSTRACTThere is an urgent need for effective vaccines against serogroup BNeisseria meningitidis. Current experimental vaccines based on the outer membrane proteins (OMPs) of this organism provide a measure of protection in older children but have been ineffective in infants. We postulated that the inability of OMP vaccines to protect infants might be due to age-dependent defects in cellular immunity. We measured proliferation and in vitro production of gamma interferon (IFN-γ), tumor necrosis factor alpha, and interleukin-10 (IL-10) in response to meningococcal antigens by peripheral blood mononuclear cells (PBMCs) from children convalescing from meningococcal disease and from controls. After meningococcal infection, the balance of cytokine production by PBMCs from the youngest children was skewed towards a TH1 response (low IL-10/IFN-γ ratio), while older children produced more TH2 cytokine (higher IL-10/IFN-γ ratio). There was a trend to higher proliferative responses by PBMCs from older children. These responses were not influenced by the presence or subtype of class 1 (PorA) OMP or by the presence of class 2/3 (PorB) or class 4 OMP. Even young infants might be expected to develop adequate cellular immune responses to serogroup BN. meningitidisvaccines if a vaccine preparation can be formulated to mimic the immune stimulus of invasive disease, which may include stimulation of TH2 cytokine production.
The Internalization Time Course of a Given Lipopolysaccharide Chemotype Does Not Correspond to Its Activation Kinetics in Monocytes
Authors: Lentschat A. , El-Samalouti V. T. , Schletter J. , Kusumoto S. , Brade L. , Rietschel E. T. , Gerdes J. , Ernst M. , Flad H.-D. , Ulmer A. J. 
Published: December 2019

ABSTRACTThe prerequisites for the initiation of pathophysiological effects of endotoxin (lipopolysaccharide [LPS]) include binding to and possibly internalization by target cells. Monocytes/macrophages are prominent target cells which are activated by LPS to release various pro- and anti-inflammatory mediators. The aim of the present study was to establish a new method to determine the binding and internalization rate of different LPS chemotypes by human monocytes and to correlate these phenomena with biological activity. It was found that membrane-bound LPS disappears within hours from the surface being internalized into the cell. Further, a correlation between the kinetics of internalization and the length of the sugar chain as well as an inverse correlation between the time course of internalization and LPS hydrophobicity was revealed. Comparison of the internalization kinetics of different LPS chemotypes with kinetics of tumor necrosis factor alpha release and kinetics of oxidative burst did not reveal any correlation of these parameters. These findings suggest that cellular internalization of and activation by LPS are mechanisms which are independently regulated.
Human Opsonins Induced during Meningococcal Disease Recognize Outer Membrane Proteins PorA and PorB
Authors: Lehmann A. K. , Halstensen A. , Aaberge I. S. , Holst J. , Michaelsen T. E. , Sørnes S. , Wetzler L. M. , Guttormsen H.-K. 
Published: December 2019

ABSTRACT

Human opsonins directed against specific meningococcal outer membrane structures in sera obtained during meningococcal disease were quantified with a recently developed antigen-specific, opsonin-dependent phagocytosis and oxidative burst assay. Outer membrane vesicles (OMVs) and PorA (class 1) and PorB (class 3) proteins purified from mutants of the same strain (44/76; B:15:P1.7.16) were adsorbed to fluorescent beads, opsonized with acute- and convalescent-phase sera from 40 patients with meningococcal disease, and exposed to human leukocytes. Flow cytometric quantitation of the resulting leukocyte phagocytosis products (PPs) demonstrated that disease-induced serum opsonins recognized meningococcal OMV components and both porins. The PP
PorA
and PP
PorB
values induced by convalescent-phase sera correlated positively with the PP
OMV
values. However, the PP
PorB
values were higher than the PP
PorA
values in convalescent-phase sera (medians [ranges] of 754 [17 to 1,057] and 107 [4 to 458], respectively) (
P
< 0.0001) and correlated positively with higher levels of immunoglobulin G against PorB than against PorA as evaluated by enzyme-linked immunosorbent assay. Extensive individual variations in the anti-OMV and antiporin serum opsonic activities between patients infected by serotypes and serosubtypes homologous and heterologous to the target antigens were observed. Simultaneously measured oxidative burst activity correlated with the opsonophagocytosis, an indication that both of these important steps in the in vitro phagocytic elimination of meningococci are initiated by opsonins directed against OMV components, including PorA and PorB. In conclusion, human patient opsonins against meningococcal OMV components and in particular PorB epitopes were identified by this new method, which might facilitate selection of opsonin-inducing meningococcal antigens for inclusion in future vaccines.
The Human Cationic Antimicrobial Protein (hCAP18), a Peptide Antibiotic, Is Widely Expressed in Human Squamous Epithelia and Colocalizes with Interleukin-6
Authors: Frohm Nilsson Margareta , Sandstedt Bengt , Sørensen Ole , Weber Günther , Borregaard Niels , Ståhle-Bäckdahl Mona 
Published: December 2019

ABSTRACTPeptide antibiotics are widespread in nature and, by providing a rapid first line of defense, may be key players in the innate immune system. Although epithelia are the main barriers shielding the internal environment from microorganisms, the role for peptide antibiotics in epithelial protection is unclear. We recently reported that the human cationic antimicrobial protein hCAP18, the precursor of the antimicrobial peptide called LL-37, is not expressed by normal human keratinocytes but is induced in various inflammatory skin disorders. In the present study we demonstrate that hCAP18 is consistently expressed at both mRNA and protein levels in squamous epithelia of the mouth, tongue, esophagus, cervix, and vagina in humans. The gene for hCAP18 contains promoter elements that are potentially regulated by interleukin-6, and our data further show a colocalization between interleukin-6 and hCAP18 expression in these tissues. Our finding that hCAP18 is widely produced in squamous epithelia suggests a role for this peptide in epithelial antimicrobial defense. Furthermore, colocalization with interleukin-6 indicates a potential local mechanism for the upregulation of hCAP18 at the epithelial surfaces.
Vaccination of Guinea Pigs with Nutritionally Impaired Avirulent Mutants of
Mycobacterium bovis
Protects against Tuberculosis
Authors: de Lisle Geoffrey W. , Wilson Theresa , Collins Desmond M. , Buddle Bryce M. 
Published: December 2019

ABSTRACT

Four nutritionally impaired strains of
Mycobacterium bovis
produced by illegitimate recombination were tested for their ability to protect guinea pigs against intratracheal challenge with virulent
M. bovis
. All four strains and
M. bovis
BCG induced significant levels of protection as measured by the reduced spread of infection to the spleen and liver. In animals vaccinated with BCG or two of the other strains, the bacterial counts from the lungs were significantly lower than those of the nonvaccinated animals.
Specific Antibodies Reactive with the 22-Kilodalton Major Outer Surface Protein of
Borrelia anserina
Ni-NL Protect Chicks from Infection
Authors: Sambri Vittorio , Marangoni Antonella , Olmo Andrea , Storni Elisa , Montagnani Marco , Fabbi Massimo , Cevenini Roberto 
Published: December 2019

ABSTRACT

An outer surface lipoprotein of 22 kDa was identified in the avian pathogen
Borrelia anserina
Ni-NL by using antibody preparations reactive with bacterial surface-exposed proteins. Amino acid sequence analysis of the 22-kDa protein demonstrated 90% identity with VmpA of
B. turicatae
, suggesting that the protein belongs to the family of 20-kDa outer surface proteins of the genus
Borrelia
. All of the 60 chicks intramuscularly treated with antibodies specifically reacting with the 22-kDa protein and infected with strain Ni-NL were completely protected from infection, since no spirochetemia was detected, and from death. Control chicks were treated with immune sera raised against apathogenic strain
B. anserina
Es, which expresses a prominent 20-kDa polypeptide that is also a member of the Vmp family but does not cross-react immunologically with the 22-kDa protein of the Ni-NL strain. These animals, infected with
B. anserina
Ni-NL, showed a high degree of spirochetemia 10 days after infection, and all died between 14 and 21 days after infection. The results showed that the 22-kDa surface protein of
B. anserina
Ni-NL is a determinant of the pathogenic potential of the strain and also confirmed that only strain-specific antibodies are protective against
B. anserina
infection.