bluuebunny/crossref_metadata_2025_split huggingface dataset:
CesD2 of Enteropathogenic
Escherichia coli
Is a Second Chaperone for the Type III Secretion Translocator Protein EspD
Authors: Neves Bianca C. , Mundy Rosanna , Petrovska Liljana , Dougan Gordon , Knutton Stuart , Frankel Gad 
Published: March 2003

ABSTRACT

Enteropathogenic
Escherichia coli
(EPEC) and enterohemorrhagic
E. coli
are extracellular pathogens that employ a type III secretion system to export translocator and effector proteins, proteins which facilitates colonization of the mucosal surface of the intestine via formation of attaching and effacing (A/E) lesions. The genes encoding the proteins for A/E lesion formation are located on a pathogenicity island, termed the locus of enterocyte effacement (LEE), which contains
eae
encoding intimin as well as the type III secretion system and effector genes. Many type III secreted proteins are stabilized and maintained in a secretion-competent conformation in the bacterial cytosol by specific chaperone proteins. Three type III chaperones have been described thus far within the EPEC LEE region: CesD, for the translocator proteins EspB and EspD; CesT, for the effector proteins Tir and Map; and CesF, for EspF. In this study we report the characterization of CesD2 (previously Orf27), a second LEE-encoded chaperone for EspD. We show specific CesD2-EspD protein interaction which appears to be necessary for proper EspD secretion in vitro and pathogenesis in vivo as demonstrated in the A/E-lesion-forming mouse pathogen
Citrobacter rodentium
.
AnEscherichia coliMG1655 Lipopolysaccharide Deep-Rough Core Mutant Grows and Survives in Mouse Cecal Mucus but Fails To Colonize the Mouse Large Intestine
Authors: Møller Annette K. , Leatham Mary P. , Conway Tyrrell , Nuijten Piet J. M. , de Haan Louise A. M. , Krogfelt Karen A. , Cohen Paul S. 
Published: March 2003

ABSTRACTThe ability ofE. colistrains to colonize the mouse large intestine has been correlated with their ability to grow in cecal and colonic mucus. In the present study, anE. coliMG1655 strain was mutagenized with a mini-Tn5Km (kanamycin) transposon, and mutants were tested for the ability to grow on agar plates with mouse cecal mucus as the sole source of carbon and nitrogen. One mutant, designated MD42 (for mucus defective), grew poorly on cecal-mucus agar plates but grew well on Luria agar plates and on glucose minimal-agar plates. Sequencing revealed that the insertion in MD42 was in thewaaQgene, which is involved in lipopolysaccharide (LPS) core biosynthesis. Like “deep-rough”E. colimutants, MD42 was hypersensitive to sodium dodecyl sulfate (SDS), bile salts, and the hydrophobic antibiotic novobiocin. Furthermore, its LPS core oligosaccharide was truncated, like that of a deep-rough mutant. MD42 initially grew in the large intestines of streptomycin-treated mice but then failed to colonize (<102CFU per g of feces), whereas its parent colonized at levels between 107and 108CFU per g of feces. When mouse cecal mucosal sections were hybridized with anE. coli-specific rRNA probe, MD42 was observed in cecal mucus as clumps 24 h postfeeding, whereas its parent was present almost exclusively as single cells, suggesting that clumping may play a role in preventing MD42 colonization. Surprisingly, MD42 grew nearly as well as its parent during growth in undiluted, highly viscous cecal mucus isolated directly from the mouse cecum and, like its parent, survived well after reaching stationary phase, suggesting that there are no antimicrobials in mucus that prevent MD42 colonization. After mini-marinertransposon mutagenesis, an SDS-resistant suppressor mutant of MD42 was isolated. The mini-marinerinsertion was shown to be in thebipAgene, a known regulator ofE. colisurface components. When grown in Luria broth, the LPS core of the suppressor mutant remained truncated; however, the LPS core was not truncated when the suppressor mutant was grown in the presence of SDS. Moreover, when the suppressor mutant was grown in the presence of SDS and fed to mice, it colonized the mouse large intestine. Collectively, the data presented here suggest that BipA may play a role inE. coliMG1655 LPS core biosynthesis and that because MD42 forms clumps in intestinal mucus, it is unable to colonize the mouse large intestine.
Involvement of Myeloid Dendritic Cells in the Development of Gastric Secondary Lymphoid Follicles inHelicobacter pylori-Infected Neonatally Thymectomized BALB/c Mice
Authors: Nishi Toshiki , Okazaki Kazuichi , Kawasaki Kimio , Fukui Toshiro , Tamaki Hiroyuki , Matsuura Minoru , Asada Masanori , Watanabe Tomohiro , Uchida Kazushige , Watanabe Norihiko , Nakase Hiroshi , Ohana Masaya , Hiai Hiroshi , Chiba Tsutomu 
Published: March 2003

ABSTRACTWe previously described an animal model ofHelicobacter pylori-induced follicular gastritis in neonatally thymectomized (nTx) mice. However, it is still not clear whether antigen-presenting dendritic cells (DCs) in the stomach have a role in the development of secondary follicles inH. pylori-infected nTx mice. We investigated the distribution of DC subsets using this model and examined their roles. To identify lymphoid and myeloid DCs, sections were stained with anti-CD11c (pan-DC marker) in combination with anti-CD8α (lymphoid DC marker) or anti-CD11b (myeloid DC marker) and were examined with a confocal microscope. Expression of macrophage inflammatory protein 3α (MIP-3α), which chemoattracts immature DCs, was analyzed by real-time PCR and immunohistochemistry. Follicular dendritic cells (FDCs) were stained with anti-SKY28 antibodies. In noninfected nTx mice, a few myeloid and lymphoid DCs were observed in the bottom portion of the lamina propria, whereas inH. pylori-infected nTx mice, there was an increased influx of myeloid DCs throughout the lamina propria. FDC staining was also observed in the stomachs of members of the infected group. MIP-3α gene expression was upregulated in the infected nTx group, and the immunohistochemistry analysis revealed MIP-3α-positive epithelial cells. These data suggest thatH. pyloriinfection upregulates MIP-3α gene expression in gastric epithelial cells and induces an influx of myeloid DCs in the lamina propria of the gastric mucosa in nTx mice. Myeloid DCs and FDCs might contribute to the development of gastric secondary lymphoid follicles inH. pylori-infected nTx mice.
Mapping the Binding Domain of the F18 Fimbrial Adhesin
Authors: Smeds A. , Pertovaara M. , Timonen T. , Pohjanvirta T. , Pelkonen S. , Palva A. 
Published: March 2003

ABSTRACT

F18 fimbrial
Esherichia coli
strains are associated with porcine postweaning diarrhea and pig edema disease. Recently, the FedF subunit was identified as the adhesin of the F18 fimbriae. In this study, adhesion domains of FedF were further studied by constructing deletions within the
fedF
gene and expressing FedF proteins with deletions either together with the other F18 fimbrial subunits or as fusion proteins tagged with maltose binding protein. The region essential for adhesion to porcine intestinal epithelial cells was mapped between amino acid residues 60 and 109 of FedF. To map the binding domain even more closely, all eight charged amino acid residues within this region were independently replaced by alanine. Three of these single point mutants expressing F18 fimbriae exhibited significantly diminished capabilities to adhere to porcine epithelial cells in vitro. In addition, a triple point mutation and a double point mutation completely abolished receptor adhesiveness. The result further confirmed that the region between amino acid residues 60 and 109 is essential for the binding of F18 fimbriae to their receptor. In addition, the adhesion capability of the binding domain was eliminated after treatment with iodoacetamide, suggesting the formation of a disulfide bridge between Cys-63 and Cys-83, whereas Cys-111 and Cys-116 could be deleted without affecting the binding ability of FedF.
Identification and Characterization of a Conserved, Stage-Specific Gene Product of
Plasmodium falciparum
Recognized by Parasite Growth Inhibitory Antibodies
Authors: Daubenberger Claudia A. , Diaz Diana , Curcic Marija , Mueller Markus S. , Spielmann Tobias , Certa Ulrich , Lipp Joachim , Pluschke Gerd 
Published: March 2003

ABSTRACT

We have identified a novel conserved protein of
Plasmodium falciparum
, designated D13, that is stage-specifically expressed in asexual blood stages of the parasite. The predicted open reading frame (ORF) D13 contains 863 amino acids with a calculated molecular mass of 99.7 kDa and displays a repeat region composed of pentapeptide motives. Northern blot analysis with lysates of synchronized blood stage parasites showed that D13 is highly expressed at the mRNA level during schizogony. The first N′-terminal 138 amino acids of D13 were expressed in
Escherichia coli
and the purified protein was used to generate anti-D13 monoclonal antibodies (MAbs). Using total lysates of blood stage parasites and Western blot analysis, these MAbs stained one single band of ∼100 kDa, corresponding to the predicted molecular mass of ORF D13. Western blot analysis demonstrated further that D13 is expressed during schizogony, declines rapidly in early ring stages and is undetectable in trophozoites. D13 protein is localized in individual merozoites in a distinct area, as demonstrated by indirect immunofluorescence analysis. After subcellular fractionation, D13 was confined to the pelleted fraction of the parasite lysate and its extraction by alkaline carbonate buffer treatment indicated that D13 is not a membrane-integral protein. Inclusion of certain anti-D13 MAbs into in vitro cultures of blood stage parasites resulted in considerable reduction in parasite growth. The N′-terminal domain encompassing 158 amino acids is 94 and 95%, respectively, identical at the amino acid level between
Plasmodium knowlesi
,
Plasmodium yoelii
, and
P. falciparum
. In summary, we describe a novel stage-specifically expressed, highly conserved gene product of
P. falciparum
that is recognized by parasite growth inhibitory antibodies.
Rapid Protection of Gnotobiotic Pigs against Experimental Salmonellosis following Induction of Polymorphonuclear Leukocytes by AvirulentSalmonella enterica
Authors: Foster N. , Lovell M. A. , Marston K. L. , Hulme S. D. , Frost A. J. , Bland P. , Barrow P. A. 
Published: March 2003

ABSTRACTOral inoculation of 5-day-old gnotobiotic pigs withSalmonella entericaserovar Typhimurium strain F98 resulted in severe enteritis and invasive disease. Preinoculation 24 h earlier with an avirulent mutant ofSalmonella entericaserovar Infantis (1326/28) completely prevented disease for up to 14 days (when the experiment was terminated).S. entericaserovar Infantis colonized the alimentary tract well, with high bacterial counts in the intestinal lumen but with almost no invasion into the tissues. Unprotected pigs had highS. entericaserovar Typhimurium counts in the intestines, blood, and major nonintestinal organs. Recovery of this strain from the blood and major organs inS. entericaserovar Infantis-protected pigs was substantially reduced despite the fact that intestinal counts were also very high. Protection against disease thus did not involve a colonization exclusion phenomenon. Significant (P< 0.05) infiltration of monocytes/macrophages was observed in the submucosal regions of the intestines of bothS. entericaserovar Infantis-protectedS. entericaserovar Typhimurium-challenged pigs and unprotectedS. entericaserovar Typhimurium-challenged pigs. However, only polymorphonuclear neutrophils (PMNs) were observed throughout the villus, where significant (P< 0.05) numbers infiltrated the lamina propria and the subnuclear and supranuclear regions of the epithelia, indicating that PMN induction and positioning followingS. entericaserovar Infantis inoculation was consistent with rapid protection against the challenge strain. Similarly, in vitro experiments using a human fetal intestinal epithelial cell line (INT 407) demonstrated that, although significantly (P< 0.05) fewerS. entericaserovar Infantis thanS. entericaserovar Typhimurium organisms invaded the monolayers,S. entericaserovar Infantis induced an NF-κB response and significantly (P< 0.05) raised interleukin 8 levels and transmigration of porcine PMN. The results of this study suggest that attenuatedSalmonellastrains can protect the immature intestine against clinical salmonellosis by PMN induction. They also demonstrate that PMN induction is not necessarily associated with clinical symptoms and/or intestinal pathology.
Pulmonary Necrosis Resulting from DNA Vaccination against Tuberculosis
Authors: Taylor Jennifer L. , Turner Oliver C. , Basaraba Randall J. , Belisle John T. , Huygen Kris , Orme Ian M. 
Published: March 2003

ABSTRACTThe use of DNA constructs encoding mycobacterial proteins is a promising new approach to vaccination against tuberculosis. A DNA vaccine encoding the hsp60 molecule ofMycobacterium lepraehas previously been shown to protect against intravenous infection of mice withMycobacterium tuberculosisin both the prophylactic and immunotherapeutic modes. It is shown here, however, that this vaccine was not effective in a more realistic aerosol infection model or in a model of latent tuberculosis in the lungs. Moreover, when given in an immunotherapeutic model the immunized mice developed classical Koch reactions characterized by multifocal discrete regions of cellular necrosis throughout the lung granulomas. Similar and equally severe reactions were seen in mice given a vaccine with DNA coding for the Ag85 antigen ofM. tuberculosis. This previously unanticipated safety problem indicates that DNA vaccines should be used with caution in individuals who may have already been exposed to tuberculosis.
Group AStreptococcusGene Expression in Humans and Cynomolgus Macaques with Acute Pharyngitis
Authors: Virtaneva Kimmo , Graham Morag R. , Porcella Stephen F. , Hoe Nancy P. , Su Hua , Graviss Edward A. , Gardner Tracie J. , Allison James E. , Lemon William J. , Bailey John R. , Parnell Michael J. , Musser James M. 
Published: March 2003

ABSTRACTThe molecular mechanisms used by group AStreptococcus(GAS) to survive on the host mucosal surface and cause acute pharyngitis are poorly understood. To provide new information about GAS host-pathogen interactions, we used real-time reverse transcription-PCR (RT-PCR) to analyze transcripts of 17 GAS genes in throat swab specimens taken from 18 pediatric patients with pharyngitis. The expression of known and putative virulence genes and regulatory genes (including genes in seven two-component regulatory systems) was studied. Several known and previously uncharacterized GAS virulence gene regulators were highly expressed compared to the constitutively expressed control geneproS. To examine in vivo gene transcription in a controlled setting, three cynomolgus macaques were infected with strain MGAS5005, an organism that is genetically representative of most serotype M1 strains recovered from pharyngitis and invasive disease episodes in North America and Western Europe. These three animals developed clinical signs and symptoms of GAS pharyngitis and seroconverted to several GAS extracellular proteins. Real-time RT-PCR analysis of throat swab material collected at intervals throughout a 12-day infection protocol indicated that expression profiles of a subset of GAS genes accurately reflected the profiles observed in the human pediatric patients. The results of our study demonstrate that analysis of in vivo GAS gene expression is feasible in throat swab specimens obtained from infected human and nonhuman primates. In addition, we conclude that the cynomolgus macaque is a useful nonhuman primate model for the study of molecular events contributing to acute pharyngitis caused by GAS.
Caenorhabditis elegansas a Model Host forStaphylococcus aureusPathogenesis
Authors: Sifri Costi D. , Begun Jakob , Ausubel Frederick M. , Calderwood Stephen B. 
Published: March 2003

ABSTRACTStaphylococcus aureus, an important pathogen of humans and other warm-blooded animals, is also capable of killing the nematodeCaenorhabditis elegans. Here, we show thatC. elegansorganisms that are fedS. aureusdie over the course of several days in a process that is correlated with the accumulation of bacteria within the nematode digestive tract. SeveralS. aureusvirulence determinants known or speculated to be important in mammalian pathogenesis, including the quorum-sensing global virulence regulatory systemagrand the global virulence regulatorsarA, the alternative sigma factor σB, alpha-hemolysin, and V8 serine protease, are required for full pathogenicity in nematodes. In addition, several definedC. elegansmutants were examined for susceptibility toS. aureusinfection. Enhanced susceptibility toS. aureuskilling was observed with loss-of-function mutations in theC. elegansgenesesp-2/sek-1andesp-8/nsy-1, which encode components of a conserved p38 MAP kinase signaling pathway involved in nematode defense against multiple pathogens. These results suggest that key aspects ofS. aureuspathogenesis have been conserved, irrespective of the host, and that specificC. eleganshost factors can alter susceptibility to this gram-positive human pathogen.
Cloning and Characterization of anEhrlichia canisGene Encoding a Protein Localized to the Morula Membrane
Authors: Teng Ching-Hao , Palaniappan Raghavan U. M. , Chang Yung-Fu 
Published: March 2003

ABSTRACTA gene encoding a 23.5-kDa ehrlichial morula membrane protein designated MmpA was cloned by screening anEhrlichia canisexpression library with convalescent dog sera, which resulted in three positive clones. Sequence analysis of the insert DNAs from all three clones indicated an open reading frame with a size of 666 bp that encodes MmpA. The structural analysis of MmpA indicated that it is a transmembrane protein with extreme hydrophobicity. Southern blot analysis of theHindIII-digested chromosomal DNA demonstrated the presence of a single copy of themmpAgene inE. canisandEhrlichia chaffeensisbut not in the human granulocytic ehrlichiosis agent. ThemmpAgene was amplified, cloned, and expressed as a fusion protein. Polyclonal antibodies to the recombinant protein (rMmpA) were raised in rabbits. Western blot analysis ofE. canisandE. chaffeensislysates with the anti-rMmpA serum resulted in the presence of an MmpA band only inE. canis, not inE. chaffeenesis. Sera from dogs which were either naturally or experimentally infected withE. canisrecognized the recombinant protein. Double immunofluorescence confocal microscopy studies demonstrated that MmpA was localized mainly on the morula membrane ofE. canis. Since the morula membrane is the interface between the ehrlichial growing environment and the host cytoplasm, MmpA may play a role in bacterium-host cell interactions.
Passive Immunity in
Helicobacter
-Challenged Neonatal Mice Conferred by Immunized Dams Lasts until Weaning
Authors: Corthésy-Theulaz Irène , Corthésy Blaise , Bachmann Daniel , Velin Dominique , Kraehenbuhl Jean-Pierre 
Published: March 2003

ABSTRACT

The objective of this study was to examine the effect of breast-feeding by immunized dams on
Helicobacter
colonization in newborns. Urease-based immunization regimens failed to protect nursing pups against
H. felis
, whereas
H. felis
lysate-cholera toxin resulted in protection. This observation correlated with a high recognition of cell surface-expressed bacterial antigens by milk antibodies. Protection lasted until weaning, indicating that infection is maintained at undetectable levels by passive immunity but then resumes when breast-feeding stops.
Protein Binding between PcrG-PcrV and PcrH-PopB/PopD Encoded by thepcrGVH-popBDOperon of thePseudomonas aeruginosaType III Secretion System
Authors: Allmond Leonard R. , Karaca Timur J. , Nguyen Vinh N. , Nguyen Thong , Wiener-Kronish Jeanine P. , Sawa Teiji 
Published: March 2003

ABSTRACTOf the proteins encoded by thepcrGVH-popBDoperon of thePseudomonas aeruginosatype III secretion system, PcrG bound to PcrV and PcrH bound to PopB/PopD. In addition,YersiniaLcrG bound to PcrV, andYersiniaLcrH bound to PopD. The results imply a highly functional conservation of type III secretion betweenP.aeruginosaandYersiniaspecies.
Synergistic Protection of Mice against Plague with Monoclonal Antibodies Specific for the F1 and V Antigens of
Yersinia pestis
Authors: Hill Jim , Copse Catherine , Leary Sophie , Stagg Anthony J. , Williamson E. Diane , Titball Richard W. 
Published: March 2003

ABSTRACT

Monoclonal antibodies specific for
Yersinia pestis
V antigen and F1 antigen, administered singly or in combination, protected mice in models of bubonic and pneumonic plague. Antibodies showed synergy when administered prophylactically and as a therapy 48 h postinfection. Monoclonal antibodies therefore have potential as a treatment for plague.
Enhanced Murine Antigen-Specific Gamma Interferon and Immunoglobulin G2a Responses by Using Mycobacterial ESAT-6 Sequences in DNA Vaccines
Authors: Minion F. Chris , Menon Sreekumar A. , Mahairas Gregory G. , Wannemuehler M. J. 
Published: March 2003

ABSTRACTTheMycobacterium tuberculosisprotein ESAT-6 has unusual immune stimulating activities, has been implicated in the recall of long-lived immunity, and induces protection against tuberculosis in mice. For many diseases caused by bacterial or viral pathogens, a strong cell-mediated immune (i.e., type 1) response is often required for recovery or protection. Therefore, it is important to design immunization regimens that induce agent-specific type 1 immunity. We have shown in previous studies that ESAT-6 could enhance antigen-specific type 1 immune responses in BALB/c mice against a second antigen when presented as a purified fusion protein. It was also of interest to determine if ESAT-6 could enhance the type 1 response against a second antigen beyond that afforded by DNA vaccination through CpG motifs. This was tested by using gene fusions of ESAT-6 and theMycoplasma hyopneumoniaesurface antigen P71. Modified P71 gene sequences were cloned with or without ESAT-6 sequences into a DNA vaccine vector and were used to immunize mice. Splenic lymphocytes from vaccinated mice were tested for gamma interferon (IFN-γ) and interleukin-10 (IL-10) secretion. Serum antibodies were examined for P71 antigen-specific isotype responses. When stimulated in vitro with purified P71 antigen, splenocytes from the ESAT-6:P71 vaccinates secreted higher levels of IFN-γ and lower levels of IL-10 compared to those of vaccinates receiving the P71 construct alone. Furthermore, the immunoglobulin G2a serum antibody levels were significantly higher in the ESAT-6:P71 vaccinates compared to those of the vaccinates receiving P71 alone. In conclusion, ESAT-6 was shown to enhance antigen-specific type 1 immune responses in BALB/c mice when used in DNA vaccines.
Th2 Responses Predominate during the Early Phases of Infection in Patients with Localized Cutaneous Leishmaniasis and Precede the Development of Th1 Responses
Authors: Bourreau Eliane , Gardon Jacques , Pradinaud Roger , Pascalis Hervé , Prévot-Linguet Ghislaine , Kariminia Amina , Pascal Launois 
Published: March 2003

ABSTRACT

Intralesional Th2 responses preceded the development of Th1 responses in localized cutaneous leishmaniasis due to
Leishmania guyanensis
. Although the number of parasites increased in Th2 lesions, no correlation was found between the levels of cytokine expression and the number of parasites. In contrast, the decreased number of parasites in Th1 lesions is negatively correlated to gamma interferon expression.
Molecular Characterization of the Prototrophic
Salmonella
Mutants Defective for Intraepithelial Replication
Authors: Suvarnapunya Akamol E. , Zurawski Daniel V. , Guy Rebecca L. , Stein Murry A. 
Published: March 2003

ABSTRACT

Three Mu
d
J prototrophs demonstrated that intracellular replication is a
Salmonella
virulence trait (K. Y. Leung and B. B. Finlay, Proc. Natl. Acad. Sci. USA,
88:
11470-11474, 1991).
mutS
and
mutH
are disrupted in mutants 3-11 and 12-23, and
ssaQ
is disrupted in mutant 17-21. Further analysis revealed that loss of
Salmonella
pathogenicity island 2 function underlies the intracellular replication defect of 3-11 and 17-21.