bluuebunny/crossref_metadata_2025_split huggingface dataset:
Receptor-Mediated Recognition and Uptake of Iron from Human Transferrin by
Staphylococcus aureus
and
Staphylococcus epidermidis
Authors: Modun Belinda , Evans Robert W. , Joannou Christopher L. , Williams Paul 
Published: December 2019

ABSTRACT

Staphylococcus aureus
and
Staphylococcus epidermidis
both recognize and bind the human iron-transporting glycoprotein, transferrin, via a 42-kDa cell surface protein receptor. In an iron-deficient medium, staphylococcal growth can be promoted by the addition of human diferric transferrin but not human apotransferrin. To determine whether the staphylococcal transferrin receptor is involved in the removal of iron from transferrin, we employed 6 M urea–polyacrylamide gel electrophoresis, which separates human transferrin into four forms (diferric, monoferric N-lobe, and monoferric C-lobe transferrin and apotransferrin).
S. aureus
and
S. epidermidis
but not
Staphylococcus saprophyticus
(which lacks the transferrin receptor) converted diferric human transferrin into its apotransferrin form within 30 min. During conversion, iron was removed sequentially from the N lobe and then from the C lobe. Metabolic poisons such as sodium azide and nigericin inhibited the release of iron from human transferrin, indicating that it is an energy-requiring process. To demonstrate that this process is receptor rather than siderophore mediated, we incubated (i) washed staphylococcal cells and (ii) the staphylococcal siderophore, staphyloferrin A, with porcine transferrin, a transferrin species which does not bind to the staphylococcal receptor. While staphyloferrin A removed iron from both human and porcine transferrins, neither
S. aureus
nor
S. epidermidis
cells could promote the release of iron from porcine transferrin. In competition binding assays, both native and recombinant N-lobe fragments of human transferrin as well as a naturally occurring human transferrin variant with a mutation in the C-lobe blocked binding of
125
I-labelled transferrin. Furthermore, the staphylococci removed iron efficiently from the iron-loaded N-lobe fragment of human transferrin. These data demonstrate that the staphylococci efficiently remove iron from transferrin via a receptor-mediated process and provide evidence to suggest that there is a primary receptor recognition site on the N-lobe of human transferrin.
Site-Directed Mutagenesis of the 19-Kilodalton Lipoprotein Antigen Reveals No Essential Role for the Protein in the Growth and Virulence of
Mycobacterium intracellulare
Authors: Mahenthiralingam Eshwar , Marklund Britt-Inger , Brooks Lucy A. , Smith Debbie A. , Bancroft Gregory J. , Stokes Richard W. 
Published: December 2019

ABSTRACT

The mycobacterial 19-kilodalton antigen (19Ag) is a highly expressed, surface-associated glycolipoprotein which is immunodominant in infected patients and has little homology with other known proteins. To investigate the pathogenic significance of the 19Ag, site-directed mutagenesis of the
Mycobacterium intracellulare
19Ag gene was carried out by using a suicide vector-based strategy. Allelic replacement of the 19Ag gene of a mouse-avirulent M. intracellulare strain, 1403, was achieved by double-crossover homologous recombination with a gentamicin resistance gene-mutated allele. Unfortunately, an isogenic 19Ag was not achievable in the mouse-virulent strain, D673. However, a 19Ag mutant was successfully constructed in
M. intracellulare
FM1, a chemically mutagenized derivative of strain D673. FM1 was more amenable to genetic manipulation and susceptible to site-directed mutagenesis of the 19Ag gene yet retained the virulent phenotype of the parental strain. No deleterious effects of 19Ag gene mutation were observed during in vitro growth of
M. intracellulare
. Virulence assessment of the isogenic 19Ag mutants in a mouse infection model demonstrated that the antigen plays no essential role in the growth of
M. intracellulare
in vivo. Site-directed mutagenesis of the 19Ag gene demonstrated that it plays no essential role in growth and pathogenicity of
M. intracellulare
; however, the exact nature of its biological function remains unknown.
Regulation of
hly
Expression in
Listeria monocytogenes
by Carbon Sources and pH Occurs through Separate Mechanisms Mediated by PrfA
Authors: Behari Jaideep , Youngman Philip 
Published: December 2019

ABSTRACT

Expression of the PrfA-controlled virulence gene
hly
(encoding the pore-forming cytolysin listeriolysin) is under negative regulation by readily metabolized carbon sources in
Listeria monocytogenes
. However, the hyperhemolytic strain NCTC 7973 exhibits deregulated
hly
expression in the presence of repressing sugars, raising the possibility that a defect in carbon source regulation is responsible for its anomalous behavior. We show here that the activity of a second glucose-repressed enzyme, α-glucosidase, is 10-fold higher in NCTC 7973 than in 10403S. Using
hly-gus
fusions, we show that the
prfA
allele from NCTC 7973 causes deregulated
hly-gus
expression in the presence of sugars in either the wild-type or the NCTC 7973 background, while the 10403S
prfA
allele restores carbon source regulation. However, the
prfA
genotype does not affect the regulation of α-glucosidase activity by repressing sugars. Of the two mutational differences in PrfA, only a Gly145Ser change is important for regulation of
hly-gus
. Therefore, NCTC 7973 and 10403S have genetic differences in at least two loci: one in
prfA
that affects carbon source regulation of virulence genes and another in an unidentified gene(s) that up-regulates α-glucosidase activity. We also show that the decrease in pH associated with utilization of sugars negatively regulates
hly-gus
expression, although sugars can affect
hly-gus
expression by another mechanism that is independent of pH.
Cloning and Expression of the
Moraxella catarrhalis
Lactoferrin Receptor Genes
Authors: Du Run-Pan , Wang Qijun , Yang Yan-Ping , Schryvers Anthony B. , Chong Pele , Klein Michel H. , Loosmore Sheena M. 
Published: December 2019

ABSTRACT

The lactoferrin receptor genes from two strains of
Moraxella catarrhalis
have been cloned and sequenced. The
lfr
genes are arranged as
lbpB
followed by
lbpA
, a gene arrangement found in lactoferrin and transferrin receptor operons from several bacterial species. In addition, a third open reading frame,
orf3
, is located one nucleotide downstream of
lbpA
. The deduced lactoferrin binding protein A (LbpA) sequences from the two strains were found to be 99% identical, the LbpB sequences were 92% identical, and the ORF3 proteins were 98% identical. The
lbpB
gene was PCR amplified and sequenced from a third strain of
M. catarrhalis
, and the encoded protein was found to be 77% identical and 84% similar to the other LbpB proteins. Recombinant LbpA and LbpB proteins were expressed from
Escherichia coli
, and antisera raised to the purified proteins were used to assess antigenic conservation in a panel of
M. catarrhalis
strains. The recombinant proteins were tested for the ability to bind human lactoferrin following gel electrophoresis and electroblotting, and rLbpB, but not rLbpA, was found to bind lactoferrin. Bactericidal antibody activity was measured, and while the anti-rLbpA antiserum was not bactericidal, the anti-rLbpB antisera were found to be weakly bactericidal. Thus, LbpB may have potential as a vaccine candidate.
Characterization of the Thermal Stress Response of
Campylobacter jejuni
Authors: Konkel Michael E. , Kim Bong J. , Klena John D. , Young Colin R. , Ziprin Richard 
Published: December 2019

ABSTRACT

Campylobacter jejuni
, a microaerophilic, gram-negative bacterium, is a common cause of gastrointestinal disease in humans. Heat shock proteins are a group of highly conserved, coregulated proteins that play important roles in enabling organisms to cope with physiological stresses. The primary aim of this study was to characterize the heat shock response of
C. jejuni
. Twenty-four proteins were preferentially synthesized by
C. jejuni
immediately following heat shock. Upon immunoscreening of
Escherichia coli
transformants harboring a
Campylobacter
genomic DNA library, one recombinant plasmid that encoded a heat shock protein was isolated. The recombinant plasmid, designated pMEK20, contained an open reading frame of 1,119 bp that was capable of encoding a protein of 372 amino acids with a calculated molecular mass of 41,436 Da. The deduced amino acid sequence of the open reading frame shared similarity with that of DnaJ, which belongs to the Hsp-40 family of molecular chaperones, from a number of bacteria. An
E. coli dnaJ
mutant was successfully complemented with the pMEK20 recombinant plasmid, as judged by the ability of bacteriophage λ to form plaques, indicating that the
C. jejuni
gene encoding the 41-kDa protein is a functional homolog of the
dnaJ
gene from
E. coli
. The ability of each of two
C. jejuni dnaJ
mutants to form colonies at 46°C was severely retarded, indicating that DnaJ plays an important role in
C. jejuni
thermotolerance. Experiments revealed that a
C. jejuni
DnaJ mutant was unable to colonize newly hatched Leghorn chickens, suggesting that heat shock proteins play a role in vivo.
Regulation of Sucrose-6-Phosphate Hydrolase Activity in
Streptococcus mutans
: Characterization of the
scrR
Gene
Authors: Hiratsuka Koichi , Wang Bing , Sato Yutaka , Kuramitsu Howard 
Published: December 2019

ABSTRACT

Previous results have implicated an important role for the enzyme II
Scr
, the sucrose-specific permease, in the transport of sucrose by cariogenic
Streptococcus mutans
. The product of the
scrB
gene, sucrose-6-phosphate hydrolase (Suc-6PH), is required for the metabolism of phosphorylated sucrose. The results from the utilization of
scrB
::
lacZ
fusions in
S. mutans
GS-5 have suggested that sucrose-grown cells have higher levels of
scrB
gene expression than do cells grown with glucose or fructose. Northern blot analysis of
scrB
transcripts has also confirmed the relative strengths of expression as sucrose>glucose>fructose. Immediately downstream from the
scrB
gene, an open reading frame with homology to regulatory proteins of the GalR-LacI family as well as to ScrR proteins from several other bacteria has been identified. In addition, this gene appears to be transcribed in the same operon as
scrB
. Inactivation of this gene,
scrR
, did not alter the relative expression of the
scrB
gene in the presence of sucrose or fructose but did increase SUC-6PH levels in the presence of glucose to that observed with sucrose. Furthermore, the
S. mutans
ScrR homolog appears to bind to the
scrB
promoter region as determined from the results of gel shift assays. These results suggest that the
scrR
gene is involved in the regulation of
scrB
, and likely
scrA
, expression. However, it is not clear whether sucrose acts as an inducer of expression of these genes or, alternatively, whether glucose and fructose act as repressors.
Induction of Protective Immunity in Rabbits by Coadministration of InactivatedPasteurella multocidaToxin and Potassium Thiocyanate Extract
Authors: Jarvinen L. Z. , Hogenesch H. , Suckow M. A. , Bowersock T. L. 
Published: December 2019

ABSTRACTPasteurella multocidais a bacterial pathogen that causes rhinitis (snuffles), pneumonia, otitis media, septicemia, metritis, and death in domestic rabbits. Currently, there are no effective vaccines to prevent infection by this organism. Subcutaneous (s.c.) immunization with either exotoxin or thiocyanate extracts ofP. multocidainduces partial protection in rabbits. Since disease begins at mucosal sites, induction of local immunity may be important in preventing systemic disease. Little is known concerning the efficacy of intranasal (i.n.) administration of these antigens in inducing protective mucosal immunity toP. multocidain rabbits. The purpose of this study was twofold: (i) to investigate the effectiveness of vaccination with purifiedP. multocidatoxin (PMT) and a potassium thiocyanate extract ofP. multocida(CN) in combination and (ii) to evaluate the efficacy of administration of these antigens i.n. versus s.c. Forty-eight rabbits were randomly divided into eight different treatment groups. Rabbits received either one or both antigens by either s.c. or i.n. administration. Following vaccination, each group received an i.n. challenge ofP. multocida. Rabbits vaccinated with both antigens i.n. or s.c. had a 100% survival rate, few or no bacteria in the liver and lungs, high serum immunoglobulin G (IgG) and IgM antibody titers, and significant numbers of IgG antibody-secreting cells (ASC) in the spleen and tracheobronchial lymph node. Rabbits vaccinated i.n. had significant nasal and bronchoalveolar lavage IgA antibody levels. Rabbits vaccinated with only one antigen, either PMT or CN, had lower antibody titers, moderate to severe liver and lung infections, and fewer ASC compared to rabbits receiving both antigens. Rabbits in the control groups had moderate to severe liver and lung infections. This study indicates that i.n. immunization with both PMT and CN induces an effective response against homologousP. multocidachallenge.
Molecular Evolution of a Pathogenicity Island from Enterohemorrhagic
Escherichia coli
O157:H7
Authors: Perna Nicole T. , Mayhew George F. , Pósfai György , Elliott Simon , Donnenberg Michael S. , Kaper James B. , Blattner Frederick R. 
Published: December 2019

ABSTRACT

We report the complete 43,359-bp sequence of the locus of enterocyte effacement (LEE) from EDL933, an enterohemorrhagic
Escherichia coli
O157:H7 serovar originally isolated from contaminated hamburger implicated in an outbreak of hemorrhagic colitis. The locus was isolated from the EDL933 chromosome with a homologous-recombination-driven targeting vector. Recent completion of the LEE sequence from enteropathogenic
E. coli
(EPEC) E2348/69 afforded the opportunity for a comparative analysis of the entire pathogenicity island. We have identified a total of 54 open reading frames in the EDL933 LEE. Of these, 13 fall within a putative P4 family prophage designated 933L. The prophage is not present in E2348/69 but is found in a closely related EPEC O55:H7 serovar and other O157:H7 isolates. The remaining 41 genes are shared by the two complete LEEs, and we describe the nature and extent of variation among the two strains for each gene. The rate of divergence is heterogeneous along the locus. Most genes show greater than 95% identity between the two strains, but other genes vary more than expected for clonal divergence among
E. coli
strains. Several of these highly divergent genes encode proteins that are known to be involved in interactions with the host cell. This pattern suggests recombinational divergence coupled with natural selection and has implications for our understanding of the interaction of both pathogens with their host, for the emergence of O157:H7, and for the evolutionary history of pathogens in general.
Differential Regulation of the Interleukin-12 Receptor during the Innate Immune Response toLeishmania major
Authors: Jones Douglas , Elloso M. Merle , Showe Louise , Williams Donna , Trinchieri Giorgio , Scott Phillip 
Published: December 2019

ABSTRACTPrevious studies have shown the central role of interleukin 12 (IL-12) in the development of resistance toLeishmania majorinfection in C3H mice. We now show that during the innate immune response the lymph node cells ofL. major-infected C3H mice upregulate the IL-12 receptor on CD4+, CD8+, and B220+cells. An increase in the ability of the lymph node cells to bind IL-12 correlates with 9.3- and 4.6-fold increases in the mRNA expression levels of the IL-12Rβ1 and -β2 subunits, respectively. In contrast, BALB/c mice, which are susceptible toL. majorinfection, have no increase in the ability of the lymph node cells to bind IL-12 and correspondingly smaller increases in the mRNA expression levels of the IL-12Rβ1 and -β2 subunits of 2- and 1.5-fold, respectively. Neutralizing IL-4 and the administration of exogenous IL-12 upregulate IL-12R expression in BALB/c mice, while the neutralization of IL-12 in C3H mice blocks increased IL-12 receptor expression. These experiments reveal an important role for the regulation of the IL-12 receptor during the innate immune response after infection of mice with a pathogen.
Production of Stabilized Virulence Factor-Negative Variants by Group A Streptococci during Stationary Phase
Authors: Leonard B. A. B. , Woischnik M. , Podbielski A. 
Published: December 2019

ABSTRACT

Many of the virulence factors associated with fulminant group A streptococci (GAS) infection are expressed under in vitro exponential growth conditions. However, the survival of GAS in tissue and intracellularly, as well as colonization of asymptomatic carriers, has been reported for GAS. The bacteria associated with these niches may encounter high-density, low-nutrient-flowthrough conditions that may more closely mimic in vitro stationary-phase conditions than exponential growth. Therefore, the behavior of GAS in stationary-phase culture was examined. We observed that after 24 h in stationary phase, GAS serotypes M49 and M2 developed a unstable colony dimorphism of typical large and atypical small colonies. Between days 4 and 5, we isolated stabilized atypical small colonies which remained stable for up to nine passages (approximately 200 generations) on fresh medium before fully reverting to the large-colony phenotype. Upon analysis, the small colonies showed no difference in cell number per colony, growth rate, survival in prolonged stationary-phase culture, or antibiotic sensitivity. However, the small colonies showed decreased transcription of hyaluronic acid capsule, the global positive virulence factor regulator gene
mga
, the
mga
-regulated
emm
mRNA (M-protein structural gene), and
speB
(cysteine protease). Accordingly, the small colonies were completely sensitive in a traditional phagocytosis assay. The production of virulence factors and phagocytosis resistance of the small-colony isolates was recovered when, after several passages on fresh medium, the colony morphology began to revert.
Hemolysin-Positive Enteroaggregative and Cell-DetachingEscherichia coliStrains Cause Oncosis of Human Monocyte-Derived Macrophages and Apoptosis of Murine J774 Cells
Authors: Fernandez-Prada Carmen , Tall Ben D. , Elliott Simon E. , Hoover David L. , Nataro James P. , Venkatesan Malabi M. 
Published: December 2019

ABSTRACTInfection of human monocyte-derived macrophages (HMDM) and J774 cells (murine macrophage cell line) with several enteroaggregative and cytodetachingEscherichia coli(EAggEC and CDEC, respectively) strains demonstrated that some strains could induce macrophage cell death accompanied by release of lactate dehydrogenase activity and interleukin 1β (IL-1β) into culture supernatants. The mode of cell death differed in the two types of macrophages. Damage to macrophage plasma membrane integrity without changes in nuclear morphology resulted in cytolysis of HMDM. This mechanism of cell death has been previously described for virulentShigellainfection of HMDM and is termed oncosis. In contrast, infection of J774 cells by EAggEC and CDEC strains resulted in apoptosis. The presence of α-hemolysin (Hly) in EAggEC and CDEC strains appears to be critical for both oncosis in HMDM and apoptosis in J774 cells. Bacteria lacking Hly, including Hly−EAggEC strains as well as enterotoxigenic, enteropathogenic, and enterohemorrhagicE. colistrains, behaved like avirulentShigella flexneriin that the macrophage monolayers were intact, with no release of lactate dehydrogenase activity or IL-1β into the culture supernatants.
Heterogeneous Antibody Responses in Tuberculosis
Authors: Lyashchenko Konstantin , Colangeli Roberto , Houde Michel , Al Jahdali Hamdan , Menzies Dick , Gennaro Maria Laura 
Published: December 2019

ABSTRACT

Antibody responses during tuberculosis were analyzed by an enzyme-linked immunosorbent assay with a panel of 10 protein antigens of
Mycobacterium tuberculosis
. It was shown that serum immunoglobulin G antibodies were produced against a variety of
M. tuberculosis
antigens and that the vast majority of sera from tuberculosis patients contained antibodies against one or more
M. tuberculosis
antigens. The number and the species of serologically reactive antigens varied greatly from individual to individual. In a given serum, the level of specific antibodies also varied with the antigen irrespective of the total number of antigens recognized by that particular serum. These findings indicate that person-to-person heterogeneity of antigen recognition, rather than recognition of particular antigens, is a key attribute of the antibody response in tuberculosis.
Constitutive and Inducible Green Fluorescent Protein Expression inBartonella henselae
Authors: Lee Anthea K. , Falkow Stanley 
Published: December 2019

ABSTRACTThe green fluorescent protein (GFP) gene was expressed on a plasmid inB. henselae, and GFP-expressing bacteria were visualized by fluorescence microscopy. HEp-2 cells infected with GFP-expressing bacteria were separated from uninfected cells with a fluorescence activated cell sorter. Promoter fusions ofB. henselaechromosomal DNA togfpwere examined by flow cytometry, and aB. henselae groELpromoter fusion which induced expression at 37°C was isolated.
Binding of theHelicobacter pyloriVacuolating Cytotoxin to Target Cells
Authors: Massari Paola , Manetti Roberto , Burroni Daniela , Nuti Sandra , Norais Nathalie , Rappuoli Rino , Telford John L. 
Published: December 2019

ABSTRACTThe vacuolating cytotoxin ofHelicobacter pylori, VacA, enters the cytoplasm of target cells and causes vacuolar degeneration by interfering with late stages of endocytosis. By using indirect immunofluorescence and flow cytometry, we have demonstrated that VacA binds to specific high-affinity cell surface receptors and that this interaction is necessary for cell intoxication.
A Chimeric Influenza Virus Expressing an Epitope of Outer Membrane Protein F ofPseudomonas aeruginosaAffords Protection against Challenge withP. aeruginosain a Murine Model of Chronic Pulmonary Infection
Authors: Staczek J. , Gilleland H. E. , Gilleland L. B. , Harty R. N. , García-Sastre A. , Engelhardt O. G. , Palese P. 
Published: December 2019

ABSTRACTThe ability of a chimeric influenza virus containing, within the antigenic B site of its hemagglutinin, an 11-amino-acid (AEGRAINRRVE) insert from the peptide 10 epitope of outer membrane (OM) protein F ofPseudomonas aeruginosato serve as a protective vaccine againstP. aeruginosawas tested by using the murine chronic pulmonary infection model. Mice immunized with the chimeric virus developed antibodies that reacted in an enzyme-linked immunosorbent assay with peptide 10, with purified protein F, and with whole cells of various immunotype strains ofP. aeruginosabut failed to react with a protein F-deficient strain ofP. aeruginosa. The chimeric-virus antisera reacted specifically with protein F alone when immunoblotted against proteins extracted from cell envelopes of each of the seven Fisher-Devlin immunotype strains and had significantly greater in vitro opsonic activity forP. aeruginosathan did antisera from wild-type influenza virus-immunized mice. Subsequent to intratracheal challenge with agar-encased cells ofP. aeruginosa, chimeric-virus-immunized mice developed significantly fewer severe lung lesions than did control mice immunized with the wild-type influenza virus. Furthermore, the chimeric influenza virus-immunized group had a significantly smaller percentage of mice with >5 × 103CFU ofP. aeruginosain their lungs upon bacterial quantitation than did the control group. These data indicate that chimeric influenza viruses expressing epitopes of OM protein F warrant continued development as vaccines to prevent pulmonary infections caused byP. aeruginosa.