bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Role for Host Phosphoinositide 3-Kinase and Cytoskeletal Remodeling during
Cryptosporidium parvum
Infection
Authors: Forney John R. , DeWald Daryll B. , Yang Shiguang , Speer Clarence A. , Healey Mark C. 
Published: December 2019

ABSTRACT

Cryptosporidium parvum
preferentially infects epithelial cells lining the intestinal mucosa of mammalian hosts. Parasite development and propagation occurs within a unique intracellular but extracytoplasmic parasitophorous vacuole at the apical surface of infected cells. Parasite-induced host cell signaling events and subsequent cytoskeletal remodeling were investigated by using cultured bovine fallopian tube epithelial (BFTE) cells inoculated with
C. parvum
sporozoites. Indirect-immunofluorescence microscopy detected host tyrosine phosphorylation within 30 s of inoculation. At >30 min postinoculation, actin aggregates were detected at the site of parasite attachment by fluorescein isothiocyanate-conjugated phalloidin staining as well as by indirect immunolabeling with monoclonal anti-actin. The actin-binding protein villin was also detected in focal aggregates at the site of attachment. Host cytoskeletal rearrangement persisted for the duration of the parasitophorous vacuole and contributed to the formation of long, branched microvilli clustered around the cryptosporidial vacuole. The phosphoinositide 3-kinase inhibitor wortmannin significantly inhibited (
P
< 0.05)
C. parvum
infection when BFTE cells were pretreated for 60 min at 37°C prior to inoculation. Similarly, treatment of BFTE cells with the protein kinase inhibitors genistein and staurosporine and the cytoskeletally acting compounds 1-(5-iodonaphthalene-1-sulfonyl)-1
H
-hexahydro-1,4-diazapine, cytochalasin D, and 2,3-butanedione monoxime significantly inhibited (
P
< 0.05) in vitro infection at 24 h postinoculation. These findings demonstrate a prominent role for phosphoinositide 3-kinase activity during the early
C. parvum
infection process and suggest that manipulation of host signaling pathways results in actin rearrangement at the site of sporozoite attachment.
Lmb, a Protein with Similarities to the LraI Adhesin Family, Mediates Attachment of
Streptococcus agalactiae
to Human Laminin
Authors: Spellerberg Barbara , Rozdzinski Eva , Martin Simone , Weber-Heynemann Josephine , Schnitzler Norbert , Lütticken Rudolf , Podbielski Andreas 
Published: December 2019

ABSTRACT

Streptococcus agalactiae
is a leading cause of neonatal sepsis and meningitis. Adherence to extracellular matrix proteins is considered an important factor in the pathogenesis of infection, but the genetic determinants of this process remain largely unknown. We identified and sequenced a gene which codes for a putative lipoprotein that exhibits significant homology to the streptococcal LraI protein family. Mutants of this locus were demonstrated to have substantially reduced adherence to immobilized human laminin. The nucleotide sequence of the gene was subsequently designated
lmb
(laminin binding) and shown to be present in all of the common serotypes of
S. agalactiae
. To determine the role of Lmb in the adhesion of
S. agalactiae
wild-type strains to laminin, a recombinant Lmb protein harboring six consecutive histidine residues at the C terminus was cloned, expressed, and purified from
Escherichia coli
. Preincubation of immobilized laminin with recombinant Lmb significantly reduced adherence of the wild-type strain O90R to laminin. These results indicate that Lmb mediates the attachment of
S. agalactiae
to human laminin, which may be essential for the bacterial colonization of damaged epithelium and translocation of bacteria into the bloodstream.
rfb
Mutations in
Vibrio cholerae
Do Not Affect Surface Production of Toxin-Coregulated Pili but Still Inhibit Intestinal Colonization
Authors: Chiang Su L. , Mekalanos John J. 
Published: December 2019

ABSTRACT

The toxin-coregulated pilus (TCP) of
Vibrio cholerae
is essential for colonization. It was recently reported that
rfb
mutations in
V. cholerae
569B cause the translocation arrest of the structural subunit of TCP, raising the possibility that the colonization defects of lipopolysaccharide mutants are due to effects on TCP biogenesis. However, an
rfbB
gene disruption in either
V. cholerae
O395 or 569B has no apparent effect on surface TCP production as assessed by immunoelectron microscopy and CTX phage transduction, and an
rfbD
::Tn
5lac
mutant of O395 also shows no defect in TCP expression. We conclude that the colonization defect associated with
rfb
mutations is unrelated to defects in TCP assembly.
All Accessible Epitopes in theSalmonellaLipopolysaccharide Core Are Associated with Branch Residues
Authors: Nnalue Ndubisi Anthony 
Published: December 2019

ABSTRACTAntisera generated against each of the nine known chemotypes ofSalmonellalipopolysaccharide (LPS) core were characterized in order to delineate cross-reactive epitopes and define the bases for their accessibility. Strongly cross-reactive epitopes were associated with three chemotypes: Ra and Rb4, which recognized α-GlcNAc-1→2-α-Glc, and Rd1, which recognizedl-α-d-heptose-1→7-l-α-d-heptose. Both these disaccharides and the more weakly cross-reactive α-Gal-1→6-α-Glc terminal in Rb3LPS represent branch points along the core oligosaccharide. Therefore, branch points in endotoxin core oligosaccharides may generally be cross-reactive.
Transcutaneous Immunization with Bacterial ADP-Ribosylating Exotoxins as Antigens and Adjuvants
Authors: Glenn Gregory M. , Scharton-Kersten Tanya , Vassell Russell , Matyas Gary R. , Alving Carl R. 
Published: December 2019

ABSTRACTTranscutaneous immunization (TCI) is a new technique that uses the application of vaccine antigens in a solution on the skin to induce potent antibody responses without systemic or local toxicity. We have previously shown that cholera toxin (CT), a potent adjuvant for oral and nasal immunization, can induce both serum and mucosal immunoglobulin G (IgG) and IgA and protect against toxin-mediated mucosal disease when administered by the transcutaneous route. Additionally, CT acts as an adjuvant for coadministered antigens such as tetanus and diphtheria toxoids when applied to the skin. CT, a member of the bacterial ADP-ribosylating exotoxin (bARE) family, is most potent as an adjuvant when the A-B subunits are present and functional. We now show that TCI induces secondary antibody responses to coadministered antigens as well as to CT in response to boosting immunizations. IgG antibodies to coadministered antigens were also found in the stools and lung washes of immunized mice, suggesting that TCI may target mucosal pathogens. Mice immunized by the transcutaneous route with tetanus fragment C and CT developed anti-tetanus toxoid antibodies and were protected against systemic tetanus toxin challenge. We also show that bAREs, similarly organized as A-B subunits, as well as the B subunit of CT alone, induced antibody responses to themselves when given via TCI. Thus, TCI appears to induce potent, protective immune responses to both systemic and mucosal challenge and offers significant potential practical advantages for vaccine delivery.
Evolutionary Relationships of Pathogenic Clones of
Vibrio cholerae
by Sequence Analysis of Four Housekeeping Genes
Authors: Byun Roy , Elbourne Liam D. H. , Lan Ruiting , Reeves Peter R. 
Published: December 2019

ABSTRACT

Studies of the
Vibrio cholerae
population, using molecular typing techniques, have shown the existence of several pathogenic clones, mainly sixth-pandemic, seventh-pandemic, and U.S. Gulf Coast clones. However, the relationship of the pathogenic clones to environmental
V. cholerae
isolates remains unclear. A previous study to determine the phylogeny of
V. cholerae
by sequencing the
asd
(aspartate semialdehyde dehydrogenase) gene of
V. cholerae
showed that the sixth-pandemic, seventh-pandemic, and U.S. Gulf Coast clones had very different
asd
sequences which fell into separate lineages in the
V. cholerae
population. As gene trees drawn from a single gene may not reflect the true topology of the population, we sequenced the
mdh
(malate dehydrogenase) and
hlyA
(hemolysin A) genes from representatives of environmental and clinical isolates of
V. cholerae
and found that the
mdh
and
hlyA
sequences from the three pathogenic clones were identical, except for the previously reported 11-bp deletion in
hlyA
in the sixth-pandemic clone. Identical sequences were obtained, despite average nucleotide differences in the
mdh
and
hlyA
genes of 1.52 and 3.25%, respectively, among all the isolates, suggesting that the three pathogenic clones are closely related. To extend these observations, segments of the
recA
and
dnaE
genes were sequenced from a selection of the pathogenic isolates, where the sequences were either identical or substantially different between the clones. The results show that the three pathogenic clones are very closely related and that there has been a high level of recombination in their evolution.
Activation of Murine Macrophages by Lipoprotein and Lipooligosaccharide ofTreponema denticola
Authors: Rosen Graciela , Sela Michael N. , Naor Ronit , Halabi Amal , Barak Vivian , Shapira Lior 
Published: December 2019

ABSTRACTWe have recently demonstrated that the periodontopathogenic oral spirocheteTreponema denticolapossesses membrane-associated lipoproteins in addition to lipooligosaccharide (LOS). The aim of the present study was to test the potential of these oral spirochetal components to induce the production of inflammatory mediators by human macrophages, which in turn may stimulate tissue breakdown as observed in periodontal diseases. An enriched lipoprotein fraction (dLPP) fromT. denticolaATCC 35404 obtained upon extraction of the treponemes with Triton X-114 was found to stimulate the production of nitric oxide (NO), tumor necrosis factor alpha (TNF-α), and interleukin-1 (IL-1) by mouse macrophages in a dose-dependent manner. Induction of NO by dLPP was at 25% of the levels obtained bySalmonella typhosalipopolysaccharide (LPS) at similar concentrations, while IL-1 was produced at similar levels by both inducers. dLPP-mediated macrophage activation was unaffected by amounts of polymyxin B that neutralized the induction produced byS. typhosaLPS. dLPP also induced NO and TNF-α secretion from macrophages isolated from endotoxin-unresponsive C3H/HeJ mice to an extent similar to the stimulation produced in endotoxin-responsive mice. PurifiedT. denticolaLOS also produced a concentration-dependent activation of NO and TNF-α in LPS-responsive and -nonresponsive mouse macrophages. However, macrophage activation by LOS was inhibited by polymyxin B. These results suggest thatT. denticolalipoproteins and LOS may play a role in the inflammatory processes that characterize periodontal diseases.
Isolation and Chemical Characterization of a Capsular Polysaccharide Antigen Shared by Clinical Isolates of
Enterococcus faecalis
and Vancomycin-Resistant
Enterococcus faecium
Authors: Huebner Johannes , Wang Ying , Krueger Wolfgang A. , Madoff Lawrence C. , Martirosian Gayane , Boisot Saskia , Goldmann Donald A. , Kasper Dennis L. , Tzianabos Arthur O. , Pier Gerald B. 
Published: December 2019

ABSTRACT

Enterococci are a common cause of serious infections, especially in newborns, severely immunocompromised patients, and patients requiring intensive care. To characterize enterococcal surface antigens that are targets of opsonic antibodies, rabbits were immunized with various gentamicin-killed
Enterococcus faecalis
strains, and immune sera were tested in an opsonophagocytic assay against a selection of clinical isolates. Serum raised against one strain killed the homologous strain (12030) at a dilution of 1:5,120 and mediated opsonic killing of 33% of all strains tested. In addition, this serum killed two (28%) of seven vancomycin-resistant
Enterococcus faecium
strains. Adsorption of sera with the homologous strain eliminated killing activity. The adsorbing antigens were resistant to treatment with proteinase K and to boiling for 1 h, but were susceptible to treatment with sodium periodate, indicating that the antigen inducing opsonic activity is a polysaccharide. Antibodies in immune rabbit sera reacted with a capsule-like structure visualized by electron microscopy both on the homologous
E. faecalis
strain and on a vancomycin-resistant
E. faecium
strain. The capsular polysaccharides from
E. faecalis
12030 and
E. faecium
838970 were purified, and chemical and structural analyses indicated they were identical glycerol teichoic acid-like molecules with a carbohydrate backbone structure of 6-α-
d
-glucose-1-2 glycerol-3-PO
4
with substitution on carbon 2 of the glucose with an α-2-1-
d
-glucose residue. The purified antigen adsorbed opsonic killing activity from immune rabbit sera and elicited high titers of antibodies (when used to immunize rabbits) that both mediated opsonic killing of bacteria and bound to a capsule-like structure visualized by electron microscopy. These results indicate that approximately one-third of a sample of 15
E. faecalis
strains and 7 vancomycin-resistant
E. faecium
strains possess shared capsular polysaccharides that are targets of opsonophagocytic antibodies and therefore are potential vaccine candidates.
Enhanced Adhesion ofPasteurella multocidato Cultured Turkey Peripheral Blood Monocytes
Authors: Pruimboom Ingrid M. , Rimler Richard B. , Ackermann Mark R. 
Published: December 2019

ABSTRACTCapsular hyaluronic acid (HA) mediates adhesion of serogroup A strains ofPasteurella multocidato elicited turkey air sac macrophages (TASM). In contrast, freshly isolated turkey peripheral blood monocytes (TPBM) do not bind serogroup A strains. Following culture of TPBM for 6 days in chamber slides, adhesion of the bacteria to TPBM increased gradually. Incubation in chamber slides coated with entactin-collagen IV-laminin (ECL) attachment matrix or exposure to phorbol myristate acetate (PMA) further enhanced the adhesion ofP. multocidato TPBM. Addition of HA, but not Arg-Gly-Asp peptide, to TPBM culture inhibited bacterial adherence similarly to the inhibition previously reported for TASM. Exposure of TPBM to monoclonal antibody directed against HA-binding cell surface proteoglycan (CD44) decreased binding ofP. multocida. Collectively, these findings indicate thatP. multocidaadhesion to TPBM is mediated by capsular HA and can be increased by culture on ECL attachment matrix or PMA exposure. Additionally, the findings suggest that the capsular mucopolysaccharide of serogroup A strains ofP. multocidarecognizes an isoform of CD44 expressed on cultured TPBM.
Genetic Characterization of a New Type IV-A Pilus Gene Cluster Found in Both Classical and El Tor Biotypes of
Vibrio cholerae
Authors: Fullner Karla Jean , Mekalanos John J. 
Published: December 2019

ABSTRACT

The
Vibrio cholerae
genome contains a 5.4-kb
pil
gene cluster that resembles the
Aeromonas hydrophila tap
gene cluster and other type IV-A pilus assembly operons. The region consists of five complete open reading frames designated
pilABCD
and
yacE
, based on the nomenclature of related genes from
Pseudomonas aeruginosa
and
Escherichia coli
K-12. This cluster is present in both classical and El Tor biotypes, and the
pilA
and
pilD
genes are 100% conserved. The
pilA
gene encodes a putative type IV pilus subunit. However, deletion of
pilA
had no effect on either colonization of infant mice or adherence to HEp-2 cells, demonstrating that
pilA
does not encode the primary subunit of a pilus essential for these processes. The
pilD
gene product is similar to other type IV prepilin peptidases, proteins that process type IV signal sequences. Mutational analysis of the
pilD
gene showed that
pilD
is essential for secretion of cholera toxin and hemagglutinin-protease, mannose-sensitive hemagglutination (MSHA), production of toxin-coregulated pili, and colonization of infant mice. Defects in these functions are likely due to the lack of processing of N termini of four Eps secretion proteins, four proteins of the MSHA cluster, and TcpB, all of which contain type IV-A leader sequences. Some
pilD
mutants also showed reduced adherence to HEp-2 cells, but this defect could not be complemented in
trans
, indicating that the defect may not be directly due to a loss of
pilD
. Taken together, these data demonstrate the effectiveness of the
V. cholerae
genome project for rapid identification and characterization of potential virulence factors.
Responses of Human Intestinal Microvascular Endothelial Cells to Shiga Toxins 1 and 2 and Pathogenesis of Hemorrhagic Colitis
Authors: Jacewicz Mary S. , Acheson David W. K. , Binion David G. , West Gail A. , Lincicome Lisa L. , Fiocchi Claudio , Keusch Gerald T. 
Published: December 2019

ABSTRACTEndothelial damage is characteristic of infection with Shiga toxin (Stx)-producingEscherichia coli(STEC). Because Stx-mediated endothelial cell damage at the site of infection may lead to the characteristic hemorrhagic colitis of STEC infection, we compared the effects of Stx1 and Stx2 on primary and transformed human intestinal microvascular endothelial cells (HIMEC) to those on macrovascular endothelial cells from human saphenous vein (HSVEC). Adhesion molecule, interleukin-8 (IL-8), and Stx receptor expression, the effects of cytokine activation and Stx toxins on these responses, and Stx1 and Stx2 binding kinetics and bioactivity were measured. Adhesion molecule and IL-8 expression increased in activated HIMEC, but these responses were blunted in the presence of toxin, especially in the presence of Stx1. In contrast to HSVEC, unstimulated HIMEC constitutively expressed Stx receptor at high levels, bound large amounts of toxin, were highly sensitive to toxin, and were not further sensitized by cytokines. Although the binding capacities of HIMEC for Stx1 and Stx2 were comparable, the binding affinity of Stx1 to HIMEC was 50-fold greater than that of Stx2. Nonetheless, Stx2 was more toxic to HIMEC than an equivalent amount of Stx1. The decreased binding affinity and increased toxicity for HIMEC of Stx2 compared to those of Stx1 may be relevant to the preponderance of Stx2-producing STEC involved in the pathogenesis of hemorrhagic colitis and its systemic complications. The differences between primary and transformed HIMEC in these responses were negligible. We conclude that transformed HIMEC lines could represent a simple physiologically relevant model to study the role of Stx in the pathogenesis of hemorrhagic colitis.
In Vitro Induction of Activation-Induced Cell Death in Lymphocytes from Chronic Periodontal Lesions by Exogenous Fas Ligand
Authors: Sawa Takamasa , Nishimura Fusanori , Ohyama Hideki , Takahashi Keiso , Takashiba Shogo , Murayama Yoji 
Published: December 2019

ABSTRACTPeriodontitis is a chronic inflammatory disease which gradually destroys the supporting tissues of the teeth, leading to tooth loss in adults. The lesions are characterized by a persistence of inflammatory cells in gingival and periodontal connective tissues. To understand what mechanisms are involved in the establishment of chronic lesions, we hypothesized that infiltrating lymphocytes might be resistant to apoptosis. However, both Bcl-2 and Bcl-xL were weakly detected in lymphocytes from the lesions, compared with those from peripheral blood, suggesting that these cells are susceptible to apoptosis. Nevertheless, very few apoptotic cells were observed in tissue sections from the lesions. Lymphocytes from the lesions expressed mRNA encoding Fas, whereas Fas-ligand mRNA was very weakly expressed in lymphocytes from the lesions and in periodontal tissues. Since the results indicated that lymphocytes in the lesions might be susceptible to Fas-mediated apoptosis but lack the death signal, we next investigated if these lymphocytes actually undergo apoptosis by the addition of anti-Fas antibodies in vitro. Fas-positive lymphocytes from the lesions underwent apoptosis by these antibodies, but Fas-negative lymphocytes and Fas-positive peripheral lymphocytes did not undergo apoptosis by these antibodies. These results indicate that lymphocytes in the lesions are susceptible to activation-induced cell death and are induced to die by apoptosis after the addition of exogenous Fas ligand.
Specificity and Function of Immunogenic Peptides from the 35-Kilodalton Protein ofMycobacterium leprae
Authors: Wilkinson Robert John , Wilkinson Katalin Andrea , Jurcevic Stipo , Hills Adrian , Sinha Sudhir , Sengupta Utpal , Lockwood Diana N. J. , Katoch Kiran , Altman Daniel , Ivanyi Juraj 
Published: December 2019

ABSTRACTWe identified a T-cell determinant of the 35-kDa antigen ofMycobacterium lepraewhich is discriminatory against cross-sensitization by its closely related homologue inMycobacterium avium. From synthetic peptides covering the entire sequence, those with the highest affinity and permissive binding to purified HLA-DR molecules were evaluated for the stimulation of proliferation of peripheral blood mononuclear cells (PBMCs) from leprosy patients and healthy sensitized controls. Responses to the peptide pair 206–224, differing by four residues betweenM. lepraeandM. avium, involved both species-specific and cross-reactive T cells. Lymph node cell proliferation in HLA-DRB1*01 transgenic mice was reciprocally species specific, but only the response to theM. lepraepeptide in the context of DR1 was immunodominant. Of the cytokines in human PBMC cultures, gamma interferon production was negligible, while interleukin 10 (IL-10) responses in both patients and controls were more pronounced. IL-10 was most frequently induced by the shared 241–255 peptide, indicating that environmental cross-sensitization may skew the response toward a potentially pathogenic cytokine phenotype.
Legionella pneumophila
Utilizes the Same Genes To Multiply within
Acanthamoeba castellanii
and Human Macrophages
Authors: Segal Gil , Shuman Howard A. 
Published: December 2019

ABSTRACT

In previous reports we described a 22-kb
Legionella pneumophila
chromosomal locus containing 18 genes. Thirteen of these genes (
icmT
, -
R
, -
Q
, -
P
, -
O
, -
M
, -
L
, -
K
, -
E
, -
C
, -
D
, -
J
, and -
B
) were found to be completely required for intracellular growth and killing of human macrophages. Three genes (
icmS
, -
G
, and -
F
) were found to be partially required, and two genes (
lphA
and
tphA
) were found to be dispensable for intracellular growth and killing of human macrophages. Here, we analyzed the requirement of these genes for intracellular growth in the protozoan host
Acanthamoeba castellanii
, a well-established important environmental host of
L. pneumophila
. We found that all the genes that are completely required for intracellular growth in human macrophages are also completely required for intracellular growth in
A. castellanii
. However, the genes that are partially required for intracellular growth in human macrophages are completely required for intracellular growth in
A. castellanii
. In addition, the
lphA
gene, which was shown to be dispensable for intracellular growth in human macrophages, is partially required for intracellular growth in
A. castellanii
. Our results indicate that
L. pneumophila
utilizes the same genes to grow intracellularly in both human macrophages and amoebae.