bluuebunny/crossref_metadata_2025_split huggingface dataset:
Use of an Isogenic Mutant Constructed in
Moraxella catarrhalis
To Identify a Protective Epitope of Outer Membrane Protein B1 Defined by Monoclonal Antibody 11C6
Authors: Luke Nicole R. , Russo Thomas A. , Luther Neal , Campagnari Anthony A. 
Published: December 2019

ABSTRACT

Moraxella catarrhalis
-induced otitis media continues to be a significant cause of infection in young children, prompting increased efforts at identifying effective vaccine antigens. We have previously demonstrated that
M. catarrhalis
expresses specific outer membrane proteins (OMPs) in response to iron limitation and that this organism can utilize transferrin and lactoferrin for in vitro growth. One of these proteins, which binds human transferrin, is OMP B1. As the human host presents a naturally iron-limited environment, proteins, like OMP B1, which are expressed in response to this nutritional stress are potential vaccine antigens. In this study, we have developed monoclonal antibody (MAb) 11C6, which reacts to a surface-exposed epitope of OMP B1 expressed by
M. catarrhalis
7169. This antibody was used to clone
ompB1
, and sequence analysis suggested that OMP B1 is the
M. catarrhalis
homologue to the transferrin binding protein B described for pathogenic
Neisseriaceae
,
Haemophilus influenzae
,
Actinobacillus pleuropneumoniae
, and
M. catarrhalis
. Expression of recombinant OMP B1 on the surface of
Escherichia coli
confers transferrin binding activity, confirming that this protein is likely involved in iron acquisition. In addition,
ompB1
was used to construct an isogenic mutant in
M. catarrhalis
7169. This mutant, termed 7169b12, was used as the control in bactericidal assays designed to determine if OMP B1 elicits protective antibodies. In the presence of MAb 11C6 and human complement, wild-type 7169 demonstrated a 99% decline in viability, whereas the
ompB1
isogenic mutant was resistant to this bactericidal activity. Further analysis with MAb 11C6 revealed the presence of this OMP B1 epitope on 31% of the clinical isolates tested. These data suggest that OMP B1 is a potential vaccine antigen against
M. catarrhalis
infections.
The R-Type Pyocin of
Pseudomonas aeruginosa
C Is a Bacteriophage Tail-Like Particle That Contains Single-Stranded DNA
Authors: Lee Frank K. N. , Dudas Kathleen C. , Hanson Julie A. , Nelson M. Bud , LoVerde Philip T. , Apicella Michael A. 
Published: December 2019

ABSTRACT

Pseudomonas aeruginosa
R-type pyocin particles have been described as bacteriocins that resemble bacteriophage tail-like structures. Because of their unusual structure, we reexamined whether they contained nucleic acids. Our data indicated that pyocin particles isolated from
P. aeruginosa
C (pyocin C) contain DNA. Probes generated from this DNA by the random-primer extension method hybridized to distinct bands in restriction endonuclease-digested
P. aeruginosa
C genomic DNA. These probes also hybridized to genomic DNA from 6 of 18
P. aeruginosa
strains that produced R-type pyocins. Asymmetric PCR, complementary oligonucleotide hybridization, and electron microscopy indicated that pyocin C particles contained closed circular single-stranded DNA, approximately 4.0 kb in length. Examination of total intracellular DNA from mitomycin C-induced cultures revealed the presence of two extrachromosomal DNA molecules, a double-stranded molecule and a single-stranded molecule, which hybridized to pyocin DNA. Sequence analysis of 7,480 nucleotides of
P. aeruginosa
C chromosomal DNA containing the pyocin DNA indicated the presence of pyocin open reading frames with similarities to open reading frames from filamentous phages and cryptic phage elements. We did not observe any similarities to known phage structural proteins or previously characterized pseudomonal
prt
genes expressing R-type pyocin structural proteins. These studies demonstrate that pyocin particles from
P. aeruginosa
C are defective phages that contain a novel closed circular single-stranded DNA and that this DNA was derived from the chromosome of
P. aeruginosa
C.
Monocytes That Have IngestedYersinia enterocoliticaSerotype O:3 Acquire Enhanced Capacity To Bind to Nonstimulated Vascular Endothelial Cells via P-Selectin
Authors: Wuorela Maarit , Tohka Sami , Granfors Kaisa , Jalkanen Sirpa 
Published: December 2019

ABSTRACTReactive arthritis is usually a self-limiting polyarthritis which develops after certain gastrointestinal or urogenital infections. Microbial antigens found in the inflamed joints are thought to play a key role in the development of this disease. It is not known how antigens of the pathogenic organisms migrate from the mucosal tissues into the joints. The data presented here show that mononuclear phagocytes which mediate the dissemination of several intracellular pathogens acquire an enhanced capacity to bind to nonstimulated vascular endothelial cells after phagocytosis ofYersinia enterocoliticaO:3, one of the causative organisms of reactive arthritis. The increased binding to previously nonstimulated endothelial cells was mediated by P-selectin, whose translocation to the endothelial cell surface was induced by monocytes with intracellularYersiniabacteria. These results suggest that mononuclear phagocytes may be responsible for the dissemination of bacterial antigens and the initiation of the joint inflammation in reactive arthritis.
Impact of the High-Affinity Proline Permease Gene (
putP
) on the Virulence of
Staphylococcus aureus
in Experimental Endocarditis
Authors: Bayer Arnold S. , Coulter Silvija N. , Stover C. Kendall , Schwan William R. 
Published: December 2019

ABSTRACT

Staphylococcus aureus
causes a wide variety of invasive human infections. However, delineation of the genes which are essential for the in vivo survival of this pathogen has not been accomplished to date. Using signature tag mutagenesis techniques and large mutant pool screens, previous investigators identified several major gene classes as candidate essential gene loci for in vivo survival; these include genes for amino acid transporters, oligopeptide transporters, and lantibiotic synthesis (W. R. Schwan, S. N. Coulter, E. Y. W. Ng, M. H. Langhorne, H. D. Ritchie, L. L. Brody, S. Westbrock-Wadman, A. S. Bayer, K. R. Folger, and C. K. Stover, Infect. Immun. 66:567–572, 1998). In this study, we directly compared the virulence of four such isogenic signature tag mutants with that of the parental strain (RN6390) by using a prototypical model of invasive
S. aureus
infection, experimental endocarditis (IE). The oligonucleotide signature tag (OST) mutant with insertional inactivation of the gene (
putP
) which encodes the high-affinity transporter for proline uptake exhibited significantly reduced virulence in the IE model across three challenge inocula (10
4
to 10
6
CFU) in terms of achievable intravegetation densities (
P
, <0.05). The negative impact of
putP
inactivation on in vivo survival in the IE model was confirmed by simultaneous challenge with the original
putP
mutant and the parental strain as well as by challenge with a
putP
mutant in which this genetic inactivation was transduced into a distinct parental strain (S6C). In contrast, inactivation of loci encoding an oligopeptide transporter, a purine repressor, and lantibiotic biosynthesis had no substantial impact on the capacity of OST mutants to survive within IE vegetations. Thus, genes encoding the uptake of essential amino acids may well represent novel targets for new drug development. These data also confirm the utility of the OST technique as an important screening methodology for identifying candidate genes as requisite loci for the in vivo survival of
S. aureus.
Intranasal Immunization Confers Protection against MurinePneumocystis cariniiLung Infection
Authors: Pascale Juan M. , Shaw Margaret M. , Durant Pamela J. , Amador Aytza A. , Bartlett Marilyn S. , Smith James W. , Gregory Richard L. , McLaughlin Gerald L. 
Published: December 2019

ABSTRACTTo evaluate the feasibility of mucosal immunization againstPneumocystis carinii(Pc) experimental infection, female BALB/c mice were intranasally immunized three times with soluble Pc antigens plus cholera toxin fraction B (Pc-CTB); control groups received either Pc antigen, CTB, or phosphate-buffered saline (PBS) alone. Two weeks after the last immunization, five animals from each group were sacrificed, and cellular and humoral immune responses were evaluated. The remaining five mice were CD4 depleted using a monoclonal antibody against mouse CD4 and inoculated with viable Pc. Significantly higher specific lymphoproliferative responses from tracheobronchial lymph node cells, immunoglobulin M (IgM) and IgG antibody levels in serum, and bronchoalveolar lavage (BAL)-derived IgA antibody concentrations were observed in the Pc-CTB group of mice relative to control groups (P< 0.01). Five weeks after challenge, no Pc organisms were observed in the lung smears of the Pc-CTB group, while the animals receiving antigen, adjuvant, or PBS had progressively higher numbers of Pc microorganisms. By Western blot analysis, a strongly reactive 55- to 60-kDa antigen was recognized by BAL IgA and by serum IgG. In summary, mucosal immunization elicited specific cellular and humoral immune responses and protected against Pc lung infection after immunosuppression.
Characterization of a WaaF (RfaF) Homolog Expressed by
Haemophilus ducreyi
Authors: Bauer Beth A. , Lumbley Sheryl R. , Hansen Eric J. 
Published: December 2019

ABSTRACT

Haemophilus ducreyi
lipooligosaccharide (LOS) is capable of inducing an inflammatory response in skin (A. A. Campagnari, L. M. Wild, G. Griffiths, R. J. Karalus, M. A. Wirth, and S. M. Spinola, Infect. Immun. 59:2601–2608, 1991) and likely contributes to the virulence of this sexually transmitted pathogen (B. A. Bauer, M. K. Stevens, and E. J. Hansen, Infect. Immun. 68:4290–4298, 1998). An open reading frame in
H. ducreyi
35000 was found to encode a predicted protein that was 59% identical to the protein product of the
rfaF
(
waaF
) gene of
Salmonella typhimurium
. The
H. ducreyi waaF
gene was able to complement an
S. typhimurium rfaF
(
waaF
) mutant, a result which confirmed the identity of this gene. In contrast to the
rfaF
(
waaF
) gene of enteric bacteria, the
H. ducreyi waaF
gene was not located adjacent to other genes involved in lipopolysaccharide expression. Inactivation of the
H. ducreyi waaF
gene by insertion mutagenesis resulted in expression of a LOS that migrated much faster than wild-type LOS in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The LOS of this mutant also did not bind a monoclonal antibody directed against a cell surface-exposed epitope of wild-type
H. ducreyi
LOS. Testing of the wild-type
H. ducreyi
strain and its isogenic
waaF
mutant in the temperature-dependent rabbit model for dermal lesion production by
H. ducreyi
revealed that this
waaF
mutant was less virulent than the wild-type parent strain. Complementation of the
H. ducreyi waaF
mutant with the wild-type
H. ducreyi waaF
gene resulted in expression of both wild-type LOS and wild-type virulence by this mutant.
Antigen-Specific T-Cell Responses in Humans after Intranasal Immunization with a Meningococcal Serogroup B Outer Membrane Vesicle Vaccine
Authors: Oftung Fredrik , Næss Lisbeth Meyer , Wetzler Lee M. , Korsvold Gro Ellen , Aase Audun , Høiby E. Arne , Dalseg Rolf , Holst Johan , Michaelsen Terje E. , Haneberg Bjørn 
Published: December 2019

ABSTRACTWe have studied the ability of the Norwegian group B meningococcal outer membrane vesicle (OMV) vaccine, when administered intranasally without adjuvant, to induce T-cell responses in humans. A group of 12 vaccinees was immunized with four doses of OMVs (250 μg of protein/dose) at weekly intervals, and a single booster dose was given 5 months later. In vitro T-cell proliferation in response to the OMV vaccine, purified PorA (class 1) protein, PorB (class 3) protein, and one unrelated control antigen (Mycobacterium bovisBCG) was measured by [3H]thymidine incorporation into peripheral blood mononuclear cells obtained from the vaccinees before and after the immunizations. The nasal OMV immunizations induced antigen-specific T-cell responses in the majority of the vaccinees when tested against OMVs (7 of 12) and the PorA antigen (11 of 12). None of the vaccinees showed a vaccine-induced T-cell response to the PorB antigen after the initial four doses. Although some individuals responded to all the vaccine antigens after the booster dose, this response was not significant when the vaccinees were analyzed as a group. We have also demonstrated that the PorA antigen-specific T-cell responses correlated with anti-OMV immunoglobulin A (IgA) levels in nasal secretions, with anti-OMV IgG levels in serum, and with serum bactericidal activity. In conclusion, we have shown that it is possible to induce antigen-specific T-cell responses in humans by intranasal administration of a meningococcal OMV vaccine without adjuvant.
Cloning of the Gene Encoding the
Actinobacillus actinomycetemcomitans
Serotype b OmpA-Like Outer Membrane Protein
Authors: Komatsuzawa Hitoshi , Kawai Toshihisa , Wilson Mark E. , Taubman Martin A. , Sugai Motoyuki , Suginaka Hidekazu 
Published: December 2019

ABSTRACT

The gene encoding an outer membrane protein A (OmpA)-like, heat-modifiable Omp of
Actinobacillus actinomycetemcomitans
ATCC 43718 (strain Y4, serotype b) was cloned by a PCR cloning procedure. DNA sequence analysis revealed that the gene encodes a protein of 346 amino acid residues with a molecular mass of 36.9 kDa. The protein expressed by the cloned gene reacted with a monoclonal antibody to the previously described 29-kDa Omp (Omp29) of strain Y4. This monoclonal antibody reacted specifically with Omp29 of
A. actinomycetemcomitans
(serotype b), but not with any Omp of
Escherichia coli
, including OmpA. This protein exhibited characteristic heat modifiability on sodium dodecyl sulfate-polyacrylamide gels, showing an apparent molecular mass of 29 kDa when unheated and a mass of 34 kDa when heated. The N-terminal amino acid sequence of the protein expressed in
E. coli
perfectly matched those deduced from the purified Omp29 of strain Y4. The deduced amino acid sequence of the gene coding for Omp29 from serotype b matched completely (except for valine at position 321) that of a recently reported
omp34
gene described for
A. actinomycetemcomitans
serotype c (NCTC 9710). Because of the conserved nature of the gene within these serotypes, we designated the gene described herein from serotype b as
omp34
.
Human Intestinal M Cells Display the Sialyl Lewis A Antigen
Authors: Giannasca Paul J. , Giannasca Karen T. , Leichtner Alan M. , Neutra Marian R. 
Published: December 2019

ABSTRACTThe biochemical features that distinguish human M cells from other intestinal epithelial cell types are important for understanding microbial pathogenesis and for targeting vaccines to the mucosal immune system. We applied a large panel of carbohydrate-specific monoclonal antibodies and lectins to Peyer’s patch and cecum biopsy specimens from three normal individuals and a patient with inflammatory bowel disease. The results show that human M-cell glycosylation patterns are distinct from those of other species examined and that human M cells preferentially display the sialyl Lewis A antigen. This carbohydrate epitope is also present in a small subpopulation of enterocytes in the follicle-associated epithelium and in goblet cell mucins.
Pseudomonas aeruginosaKeratitis in Knockout Mice Deficient in Intercellular Adhesion Molecule 1
Authors: Hobden Jeffery A. , Masinick-McClellan Sharon , Barrett Ronald P. , Bark Kenneth S. , Hazlett Linda D. 
Published: December 2019

ABSTRACTIn this study, the role of intercellular adhesion molecule 1 (ICAM-1) in the pathogenesis ofPseudomonas aeruginosakeratitis was examined by using inbred ICAM-1-deficient knockout mice. These mice had significantly less (P≤ 0.02) ocular disease than wild-type mice, suggesting that ICAM-1 contributes to a more severe disease response followingP. aeruginosainfection.
Salmonella typhimuriumand Lipopolysaccharide Stimulate Extracellularly Regulated Kinase Activation in Macrophages by a Mechanism Involving Phosphatidylinositol 3-Kinase and Phospholipase D as Novel Intermediates
Authors: Procyk Katarzyna J. , Kovarik Pavel , von Gabain Alexander , Baccarini Manuela 
Published: December 2019

ABSTRACTActivation of the extracellularly regulated kinase (ERK) pathway is part of the early biochemical events that follow lipopolysaccharide (LPS) treatment of macrophages or their infection by virulent and attenuatedSalmonellastrains. Phagocytosis as well as the secretion of invasion-associated proteins is dispensable for ERK activation by the pathogen. Furthermore, the pathways used bySalmonellaand LPS to stimulate ERK are identical, suggesting that kinase activation might be solely mediated by LPS. Both stimuli activate ERK by a mechanism involving herbimycin-dependent tyrosine kinase(s) and phosphatidylinositol 3-kinase. Phospholipase D activation and stimulation of protein kinase C appear to be intermediates in this novel pathway of MEK/ERK activation.
The Staphylococcal Transferrin-Binding Protein Is a Cell Wall Glyceraldehyde-3-Phosphate Dehydrogenase
Authors: Modun Belinda , Williams Paul 
Published: December 2019

ABSTRACT

Staphylococcus aureus
and
Staphylococcus epidermidis
possess a 42-kDa cell wall transferrin-binding protein (Tpn) which is involved in the acquisition of transferrin-bound iron. To characterize this protein further, cell wall fractions were subjected to two-dimensional sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis blotted, and the N-terminus of Tpn was sequenced. Comparison of the first 20 amino acid residues of Tpn with the protein databases revealed a high degree of homology to the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Analysis of staphylococcal cell wall fractions for GAPDH activity confirmed the presence of a functional enzyme which, like Tpn, is regulated by the availability of iron in the growth medium. To determine whether Tpn is responsible for this GAPDH activity, it was affinity purified with NAD
+
agarose. Both
S. epidermidis
and
S. aureus
Tpn catalyzed the conversion of glyceraldehyde-3-phosphate to 1,3-diphosphoglycerate. In contrast,
Staphylococcus saprophyticus
, which lacks a Tpn, has no cell wall-associated GAPDH activity. Native polyacrylamide gel electrophoresis of the affinity-purified Tpn revealed that it was present in the cell wall as a tetramer, consistent with the structures of all known cytoplasmic GAPDHs. Furthermore, the affinity-purified Tpn retained its ability to bind human transferrin both in its native tetrameric and SDS-denatured monomeric forms. Apart from interacting with human transferrin, Tpn, in common with the group A streptococcal cell wall GAPDH, binds human plasmin. Tpn-bound plasmin is enzymatically active and therefore may contribute to the ability of staphylococci to penetrate tissues during infections. These studies demonstrate that the staphylococcal transferrin receptor protein, Tpn, is a multifunctional cell wall GAPDH.
Identification and Characterization of a Novel Ibe10 Binding Protein That Contributes to
Escherichia coli
Invasion of Brain Microvascular Endothelial Cells
Authors: Prasadarao Nemani V. , Wass Carol A. , Huang Sheng-He , Kim Kwang Sik 
Published: December 2019

ABSTRACT

The molecular basis of
Escherichia coli
traversal of the blood-brain barrier in the development of
E. coli
meningitis is not well understood. We have previously shown that a novel Ibe10 protein found in cerebrospinal fluid isolates of
E. coli
is necessary for invasion of the brain microvascular endothelial cells (BMEC) that constitute the blood-brain barrier both in vitro and in a newborn rat model of hematogenous meningitis. Here we identified a novel Ibe10 binding molecule/receptor (Ibe10R) on both bovine BMEC (HBMEC) and human BMEC (HBMEC) that is responsible for invasion by
E. coli
. Ibe10R, an approximately 55-kDa protein, was purified from BBMEC by Ibe10-Ni-Sepharose affinity chromatography. Bovine Ibe10R, as well as polyclonal antibodies to Ibe10R, blocked
E. coli
invasion of BBMEC very effectively. The N-terminal amino acid sequence of Ibe10R showed 75% homology to serum albumin. However, the amino acid sequence of an Ibe10R fragment generated by limited enzymatic digestion did not reveal homology to any other proteins, suggesting that Ibe10R represents a novel albumin-like protein. Immunocytochemical analysis of BBMEC using anti-Ibe10R antibody suggested that only a subset of cultured BBMEC express Ibe10R on their surface. Enrichment of Ibe10R-positive BBMEC by fluorescence-activated cell sorting with anti-Ibe10R antibody resulted in enhanced invasion by
E. coli
. The anti-Ibe10R antibody raised against bovine Ibe10R also blocked
E. coli
invasion of HBMEC very effectively. Interestingly, anti-Ibe10R antibody affinity chromatography of HBMEC membrane proteins revealed a smaller protein with an approximate molecular mass of 45 kDa. These results suggest that the Ibe10 of
E. coli
interacts with a novel BMEC surface protein, Ibe10R, for invasion of both BBMEC and HBMEC.
Cloning and Expression of the
dnaK
Gene of
Campylobacter jejuni
and Antigenicity of Heat Shock Protein 70
Authors: Thies Frank L. , Karch Helge , Hartung Hans-Peter , Giegerich Gerhard 
Published: December 2019

ABSTRACT

Campylobacter jejuni
is a leading cause of infectious diarrhea throughout the world. In addition, there is growing evidence that Guillain-Barré syndrome, an inflammatory demyelinating disease of the peripheral nervous system, is frequently preceded by
C. jejuni
infection. In the present study, the
hrcA-grpE-dnaK
gene cluster of
C. jejuni
was cloned and sequenced. The
dnaK
gene consists of an open reading frame of 1,869 bp and encodes a protein with a high degree of homology to other bacterial 70-kDa heat shock proteins (HSPs). The overall percentages of identity to the HSP70 proteins of
Helicobacter pylori
,
Borrelia burgdorferi
,
Chlamydia trachomatis
, and
Bacillus subtilis
were calculated to be 78.1, 60.5, 57.2, and 53.8%, respectively. Regions similar to the
Escherichia coli
ς
70
promoter consensus sequence and to a
cis
-acting regulatory element (CIRCE) are located upstream of the
hrcA
gene. Following heat shock, a rapid increase of
dnaK
mRNA was detectable, which reached its maximum after 20 to 30 min. A 6-His-tagged recombinant DnaK protein (rCjDnaK-His) was generated in
E. coli
, after cloning of the
dnaK
coding region into pET-22b(+), and purified by affinity and gel filtration chromatography. Antibody responses to rCjDnaK-His were significantly elevated, compared to those of healthy individuals, in about one-third of the serum specimens obtained from
C. jejuni
enteritis patients.