bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Thermophilic, Homohexameric Aminopeptidase ofBorrelia burgdorferiIs a Member of the M29 Family of Metallopeptidases
Authors: Bertin Patrícia B. , Lozzi Silene P. , Howell Jerrilyn K. , Restrepo-Cadavid Glória , Neves David , Teixeira Antonio R. L. , de Sousa Marcelo V. , Norris Steven J. , Santana Jaime M. 
Published: March 2005

ABSTRACTProteases are implicated in several aspects of the physiology of microorganisms, as well as in host-pathogen interactions. Aminopeptidases are also emerging as novel drug targets in infectious agents. In this study, we have characterized an aminopeptidase from the spirocheteBorrelia burgdorferi, the causative agent of Lyme disease. The aminopeptidolytic activity was identified in cell extracts fromB. burgdorferiby using the substrate leucine-7-amido-4-methylcoumarin. A protein displaying this activity was purified fromB. burgdorferiby a two-step chromatographic procedure, yielding a ∼300-kDa homo-oligomeric enzyme formed by monomers of ∼50 kDa. Gel enzymography experiments showed that enzymatic activity depends on the oligomeric structure of the protease but does not involve interchain disulfide bonds. The enzyme was identified by peptide mass fingerprinting as the putative aminopeptidase II ofB. burgdorferi, encoded by the gene BB0069. It shares significant identity to members of the M29/T family of metallopeptidase, is sensitive to bestatin, has a neutral pH optimum, and displays maximal activity at 60°C. Its activity is 1.75-fold higher at the temperature of the mammalian host than at that of the insect host of the pathogen. The activity of this thermophilic aminopeptidase ofB. burgdorferi(TAPBb) depends on Zn2+, and temperatures over 70°C promoted its inactivation through a transition from the hexameric state to the monomeric state. SinceB. burgdorferiis deficient in pathways for amino acid synthesis, TAPBbcould play a role in supplying required amino acids. Alternatively, the enzyme could be involved in peptide and/or protein processing.
Virulence ofPseudomonas aeruginosain a Murine Model of Gastrointestinal Colonization and Dissemination in Neutropenia
Authors: Koh Andrew Y. , Priebe Gregory P. , Pier Gerald B. 
Published: March 2005

ABSTRACTPseudomonas aeruginosabacteremia in cancer patients develops from initial gastrointestinal (GI) colonization with translocation into the bloodstream in the setting of chemotherapy-induced neutropenia and GI mucosal damage. We established a reproducible mouse model ofP. aeruginosaGI colonization and systemic spread during neutropenia. Mice received 2 mg of streptomycin/ml of drinking water and 1,500 U of penicillin G/ml for 4 days and then ingested 107CFU ofP. aeruginosaper ml of drinking water for 5 days. After GI colonization levels were determined, cyclophosphamide (Cy) was then injected intraperitoneally (i.p.) three times every other day or an antineutrophil monoclonal antibody, RB6-8C5, was injected i.p. once. Dissemination was defined by the presence ofP. aeruginosain spleens of moribund or dead mice. In this mouse model,P. aeruginosacolonizes the GI tract and then disseminates systemically once Cy or RB6-8C5 is administered. The duration and intensity of neutropenia, related to Cy dose, was found to be a means to compare the virulence of differentP. aeruginosastrains, as exhibited by comparisons of strains lacking or producing the virulence-enhancing ExoU cytotoxin. The lipopolysaccharide outer core polysaccharide and O side chains were critical in establishing GI colonization, andP. aeruginosamutants lacking thearoAgene (necessary for synthesizing aromatic amino acids) were able to establish GI colonization but unable to disseminate. Both the colonization and dissemination phases ofP. aeruginosapathogenesis can be studied in this model, which should prove useful for evaluating pathogenesis, therapies, and associated means to controlP. aeruginosanosocomial infections.
Immunoglobulin G Isotype Responses to Erythrocyte Surface-Expressed Variant Antigens of
Plasmodium falciparum
Predict Protection from Malaria in African Children
Authors: Yone Clarisse L. R. P. , Kremsner Peter G. , Luty Adrian J. F. 
Published: March 2005

ABSTRACT

We assessed immunoglobulin G (IgG) isotype responses to variant surface antigens (VSA) expressed on parasite-infected erythrocytes of a panel of heterologous isolates during and after acute episodes in groups of Gabonese children presenting with either mild or severe
Plasmodium falciparum
malaria. In the acute and convalescent phases IgG3 and IgG1 anti-VSA antibodies, respectively, predominated. In the absence of infection, the levels of both cytophilic isotypes waned, while those of IgG4 increased, particularly in those admitted with severe malaria. Prospective analyses showed significantly longer delays between malaria attacks associated both (i) with increasing IgG1 responses with specificity for VSA of isolates from children with mild malaria and (ii) with increasing IgG4 responses with specificity for VSA of isolates from children with severe malaria. These findings imply that the predictive value of prospectively measured cross-reactive VSA-specific IgG antibodies with respect to protection against malaria in African children depends both on their isotype and on their fine specificity.
DNA Immunization with Na
+
-K
+
ATPase (
Sseat-6
) Induces Protective Immunity to Larval
Strongyloides stercoralis
in Mice
Authors: Kerepesi Laura A. , Keiser Paul B. , Nolan Thomas J. , Schad Gerhard A. , Abraham David , Nutman Thomas B. 
Published: March 2005

ABSTRACT

Strongyloides stercoralis
causes chronic asymptomatic infections which can be maintained in the human host for many decades. Identification and treatment of S.
stercoralis
-infected individuals is required because immunosuppression can lead to fatal hyperinfection. In this study, human immunoglobulin G (IgG) that had previously been shown to transfer protective immunity to mice was used to identify potential protective antigens. Three antigens or genes from
S. stercoralis
larvae were identified as tropomyosin (
Sstmy-1
), Na
+
-K
+
ATPase (
Sseat-6
), and LEC-5 (
Sslec-5
). The genes were cloned into plasmids for DNA immunization, and mice were immunized intradermally with the three plasmids individually in combination with a plasmid containing murine granulocyte-macrophage colony-stimulating factor. Only Na
+
-K
+
ATPase induced a significant reduction in larval survival after DNA immunization. Immunization with a combination of all three plasmids, including Na
+
-K
+
ATPase, did not induce protective immunity. Serum from mice immunized with DNA encoding Na
+
-K
+
ATPase was transferred to naïve mice and resulted in partial protective immunity. Therefore, DNA immunization with Na
+
-K
+
ATPase induces protective immunity in mice, and it is the first identified vaccine candidate against infection with larval
S. stercoralis
.
Processing and Major Histocompatibility Complex Class II Presentation ofLegionella pneumophilaAntigens by Infected Macrophages
Authors: Neild Annie , Murata Takahiro , Roy Craig R. 
Published: March 2005

ABSTRACTTo better understand interactions between the intracellular pathogenLegionella pneumophilaand macrophages (Mφs), host and bacterial determinants important for presentation of antigens on major histocompatibility complex class II molecules (MHC-II) were investigated. It was determined that immune CD4 T-cell responses to murine bone marrow-derived Mφs (BMφs) infected with wild-typeL. pneumophilawere higher than the responses to avirulentdotAmutant bacteria. Although this enhanced response by immune T cells required modulation of vacuole transport mediated by the Dot/Icm system, it did not require intracellular replication ofL. pneumophila. Intracellular cytokine staining identified a population of immune CD4 T cells that produced gamma interferon upon incubation with BMφs infected with wild-typeL. pneumophilathat did not respond to Mφ infection withdotAmutant bacteria. Endocytic processing was required for presentation ofL. pneumophilaantigens on MHC-II as determined by a defect in CD4 T-cell responses when the pH of BMφ endosomes was neutralized with chloroquine. Investigation of MHC-II presentation of antigens by BMφs infected withL. pneumophila icmR,icmW, andicmSmutants indicated that these mutants have an intermediate presentation phenotype relative to those of wild-type anddotAmutant bacteria. In addition, it was found that antigens fromdotandicmmutants are presented earlier than antigens from wild-typeL. pneumophila. Although immune CD4 T-cell responses to proteins secreted by theL. pneumophilaLsp system were not detected, it was found that the Lsp system is important for primingL. pneumophila-specific T cells in vivo. These data indicate that optimal antigen processing and MHC-II presentation to immune CD4 T cells involves synthesis ofL. pneumophilaproteins in an endoplasmic reticulum-derived compartment followed by transport to lysosomes.
Identification and Functional Characterization of Chicken Toll-Like Receptor 5 Reveals a Fundamental Role in the Biology of Infection with
Salmonella enterica
Serovar Typhimurium
Authors: Iqbal Muhammad , Philbin Victoria J. , Withanage G. S. K. , Wigley Paul , Beal Richard K. , Goodchild Marianne J. , Barrow Paul , McConnell Ian , Maskell Duncan J. , Young John , Bumstead Nat , Boyd Yvonne , Smith Adrian L. 
Published: March 2005

ABSTRACT

Toll-like receptors (TLRs) are a major component of the pattern recognition receptor repertoire that detect invading microorganisms and direct the vertebrate immune system to eliminate infection. In chickens, the differential biology of
Salmonella
serovars (systemic versus gut-restricted localization) correlates with the presence or absence of flagella, a known TLR5 agonist. Chicken TLR5 (chTLR5) exhibits conserved sequence and structural similarity with mammalian TLR5 and is expressed in tissues and cell populations of immunological and stromal origin. Exposure of chTLR5
+
cells to flagellin induced elevated levels of chicken interleukin-1β (chIL-1β) but little upregulation of chIL-6 mRNA. Consistent with the flagellin-TLR5 hypothesis, an aflagellar
Salmonella enterica
serovar Typhimurium
fliM
mutant exhibited an enhanced ability to establish systemic infection. During the early stages of infection, the
fliM
mutant induced less IL-1β mRNA and polymorphonuclear cell infiltration of the gut. Collectively, the data represent the identification and functional characterization of a nonmammalian TLR5 and indicate a role in restricting the entry of flagellated
Salmonella
into systemic sites of the chicken.
Identification and Functional Characterization of Complement Regulator-Acquiring Surface Protein 1 of the Lyme Disease SpirochetesBorrelia afzeliiandBorrelia garinii
Authors: Wallich Reinhard , Pattathu Joseph , Kitiratschky Veronique , Brenner Christiane , Zipfel Peter F. , Brade Volker , Simon Markus M. , Kraiczy Peter 
Published: March 2005

ABSTRACTComplement regulator-acquiring surface protein 1 (CRASP-1) is the dominant factor-H-like protein 1 (FHL-1)- and factor-H-binding protein ofBorrelia burgdorferiand is suggested to contribute to persistence of the pathogen. The prototype CRASP-1 ofB. burgdorferisensu stricto (CRASP-1Bb) has been formerly characterized. As shown recently, serum-resistantBorrelia afzeliistrains express a unique FHL-1 and factor H-binding protein, designated CRASP-1Ba. Here, we describe for the first time the isolation and functional characterization of the gene encoding the full-length CRASP-1Baof 28 kDa, which, upon processing, is predicted to be 26.4 kDa. CPASP-1BaofB. afzeliispirochetes is associated with a genetic locus encoding the orthologous gbb54 gene family that maps to the linear plasmid of approximately 54 kb. Ligand affinity blotting techniques demonstrate that both native and recombinant CRASP-1Bamolecules strongly bind to FHL-1 and much more weakly to factor H. The FHL-1 and factor-H-binding site in CRASP-1Bais shown to be localized to a 12-amino-acid residue domain at the C terminus of the protein. For comparison, the correspondingcspA-like gene(s) of a serum-sensitiveBorrelia gariniistrain has also been cloned and characterized. Most notably, two CRASP-1-relatedB. gariniiproteins were identified; however, both molecules bind only weakly to FHL-1 and not at all to factor H. The present identification of the binding site of CRASP-1Barepresents an important step forward in our understanding of the pathogenesis of Lyme disease and may be helpful to design therapeutic regimens to interfere with complement evasion strategies of human pathogenicBorreliastrains.
Clinical and Microbiological Responses of Volunteers to Combined Intranasal and Oral Inoculation with a
Streptococcus gordonii
Carrier Strain Intended for Future Use as a Group A Streptococcus Vaccine
Authors: Kotloff Karen L. , Wasserman Steven S. , Jones Kevin F. , Livio Sofie , Hruby Dennis E. , Franke Christine A. , Fischetti Vincent A. 
Published: March 2005

ABSTRACT

Streptococcus gordonii
shows promise as a live mucosal vaccine vector for immunization against respiratory pathogens. In preparation for clinical trials to evaluate
S. gordonii
engineered to express group A streptococcal M protein antigens, we characterized the responses of 150 healthy volunteers to combined nasal and oral inoculation with approximately 1.5 × 10
9
CFU of SP204(1-1), an
S. gordonii
strain not bearing vaccine antigens. SP204(1-1) was selected for resistance to streptomycin and 5-fluoro-2-deoxyuridine to distinguish it from indigenous flora. In two antibiotic treatment studies, we performed serial culturing of nose, mouth, and saliva samples from 120 subjects treated with azithromycin beginning 5 days after inoculation to determine whether SP204(1-1) could be rapidly eliminated should safety concerns arise. A natural history study was performed to assess the time until spontaneous eradication in the remaining 30 subjects, who did not receive the antibiotic and who were monitored with repeated culturing for 14 weeks after inoculation. SP204(1-1) was generally well tolerated. Symptoms reported most often within 5 days of inoculation were nasal congestion (36%), headache (30%), and sore throat (19%). The strain was detected by culturing in 98% of subjects. A single dose of azithromycin eliminated colonization in 95% of subjects; all subjects receiving a 5-day course of an antibiotic showed clearance by day 11. Without the antibiotic, 82% of subjects showed spontaneous eradication of the implanted strain within 7 days, and all showed clearance by 35 days. The results of these clinical trials provide encouragement that the use of
S. gordonii
as a live mucosal vaccine vector is a feasible strategy.
Involvement of theEscherichia coliO157:H7(pO157)ecfOperon and Lipid A Myristoyl Transferase Activity in Bacterial Survival in the Bovine Gastrointestinal Tract and Bacterial Persistence in Farm Water Troughs
Authors: Yoon Jang W. , Lim Ji Youn , Park Yong H. , Hovde Carolyn J. 
Published: March 2005

ABSTRACTEscherichia coliO157:H7 is an important food-borne pathogen that causes hemorrhagic colitis and the hemolytic-uremic syndrome in humans. Recently, we reported that the pO157ecf(E. coliattaching and effacing gene-positive conserved fragments) operon is thermoregulated by an intrinsically curved DNA and contains the genes for bacterial surface-associated proteins, including a second copy of lipid A myristoyl transferase, whose chromosomal copy is thelpxMgene product.E. coliO157:H7 survives and persists well in diverse environments from the human and bovine gastrointestinal tracts (GIT) to nutrient-dilute farm water troughs. Transcriptional regulation of theecfoperon by intrinsic DNA curvature and the genetic redundancy oflpxMthat is associated with lipid A modification led us to hypothesize that the pO157ecfoperon andlpxMare associated with bacterial survival and persistence in various in vivo and ex vivo environments by optimizing bacterial membrane structure and/or integrity. To test this hypothesis, three isogenicecfoperon and/orlpxMdeletion mutants ofE. coliO157:H7 ATCC 43894 were constructed and analyzed in vitro and in vivo. The results showed that a double mutant carrying deletions in theecfandlpxMgenes had an altered lipid A structure and membrane fatty acid composition, did not survive passage through the bovine GIT, did not persist well in farm water troughs, had increased susceptibility to a broad spectrum of antibiotics and detergents, and had impaired motility. Electron microscopic analyses showed gross changes in bacterial membrane structure.
Haemophilus ducreyi
Outer Membrane Determinants, Including DsrA, Define Two Clonal Populations
Authors: White Catherine Dinitra , Leduc Isabelle , Olsen Bonnie , Jeter Chrystina , Harris Chavala , Elkins Christopher 
Published: March 2005

ABSTRACT

The
Haemophilus ducreyi
outer membrane component DsrA (for ducreyi serum resistance A) is necessary for complete resistance to normal human serum (NHS). When DsrA expression in 19 temporally and geographically diverse clinical isolates of
H. ducreyi
was examined by Western blotting, 5 of the strains expressed a different immunotype of the DsrA protein (DsrA
II
) than the well-characterized prototypical strain 35000HP (DsrA
I
). The predicted DsrA proteins expressed by the DsrA
II
strains were 100% identical to each other but only 48% identical to that of strain 35000HP. In addition to the DsrA
II
protein, class II strains also expressed variant forms of other outer membrane proteins (OMPs) including NcaA (necessary for collagen adhesion A), DltA (ducreyi lectin A), Hlp (
H. ducreyi
lipoprotein), major OMP, and/or OmpA2 (for OMP A2) and synthesized a distinct, faster-migrating lipooligosaccharide. Based on these data, strains expressing DsrA
I
were termed class I, and those expressing DsrA
II
were termed class II. Expression of
dsrA
II
from strain CIP 542 ATCC in the class I
dsrA
I
mutant FX517 (35000HP background), which does not express a DsrA protein, rendered this strain resistant to 50% NHS. This demonstrates that DsrA
II
protein is also critical to serum resistance. Taken together, these results indicate that there are two clonal populations of
H. ducreyi
. The implications of two classes of
H. ducreyi
strains differing in important antigenic outer membrane components are discussed.
Staphylococcus aureusDeficient in Lipidation of Prelipoproteins Is Attenuated in Growth and Immune Activation
Authors: Stoll Hartmut , Dengjel Jörn , Nerz Christiane , Götz Friedrich 
Published: March 2005

ABSTRACTA lipoprotein diacylglyceryl transferase (lgt) deletion mutant ofStaphylococcus aureusSA113 was constructed. The lipoprotein and prelipoprotein expression, the growth behavior, and the ability of the mutant to elicit an immune response in various host cells were studied. In the wild type, the majority of [14C]palmitate-labeled lipoproteins were located in the membrane fraction, although some lipoproteins were also present on the cell surface and in the culture supernatant. Thelgtmutant completely lacked palmitate-labeled lipoproteins and released high amounts of some unmodified prelipoproteins, e.g., the oligopeptide-binding protein OppA, the peptidyl-prolylcis-transisomerase PrsA, and the staphylococcal iron transporter SitC, into the culture supernatant. The growth of thelgtmutant was hardly affected in rich medium but was retarded under nutrient limitation. Thelgtmutant and its crude lysate induced much fewer proinflammatory cytokines and chemokines in human monocytic (MonoMac6), epithelial (pulmonary A549), and endothelial (human umbilical vein endothelial) cells than the wild type. However, in whole blood samples, the culture supernatant of thelgtmutant was equal or even superior to the wild-type supernatant in tumor necrosis factor alpha induction. Lipoprotein fractionation experiments provided evidence that a small proportion of the mature lipoproteins are released by theS. aureuswild type despite the lipid anchor and are trapped in part by the cell wall, thereby exposing the immune-activating lipid structure on the cell surface. Bacterial lipoproteins appear to be essential for a complete immune stimulation by gram-positive bacteria.
Genetic Diversity and Carriage Dynamics ofNeisseria lactamicain Infants
Authors: Bennett Julia S. , Griffiths David T. , McCarthy Noel D. , Sleeman Karen L. , Jolley Keith A. , Crook Derrick W. , Maiden Martin C. J. 
Published: March 2005

ABSTRACTNeisseria lactamica, a harmless human commensal found predominantly in the upper respiratory tracts of infants, is closely related toNeisseria meningitidis, a pathogen of global significance. Colonization withN. lactamicamay be responsible for the increase in immunity to meningococcal disease that occurs during childhood, when rates of meningococcal carriage are low. This observation has led to the suggestion thatN. lactamicawhole cells or components are potential constituents of novel meningococcal vaccines. However, the dynamics of carriage and population diversity ofN. lactamicain children are poorly understood, presenting difficulties for the choice of representative isolates for use in vaccine development. This problem was addressed by the multilocus sequence typing ofN. lactamicaisolates from two longitudinal studies of bacterial carriage in infants. The studies comprised 100 and 216 subjects, withN. lactamicacarriage monitored from age 4 weeks until age 96 weeks and from age 2 weeks until age 24 weeks, respectively. The maximum observed carriage rate was 44% at 56 weeks of age, with isolates obtained on multiple visits for the majority (54 of 75, 72%) of carriers. TheN. lactamicaisolates were genetically diverse, with 69 distinct genotypes recovered from the 75 infants. Carriage was generally long-lived, with an average rate of loss of under 1% per week during the 28 weeks following acquisition. Only 11 of the 75 infants carried more than one genotypically unique isolate during the course of the study. Some participants shared identical isolates with siblings, but none shared identical isolates with their parents. These findings have implications for the design of vaccines based on this organism.
Role of Predicted Transmembrane Domains for Type III Translocation, Pore Formation, and Signaling by the
Yersinia pseudotuberculosis
YopB Protein
Authors: Ryndak Michelle B. , Chung Hachung , London Erwin , Bliska James B. 
Published: March 2005

ABSTRACT

YopB is a 401-amino-acid protein that is secreted by a plasmid-encoded type III secretion system in pathogenic
Yersinia
species. YopB is required for
Yersinia
spp. to translocate across the host plasma membrane a set of secreted effector proteins that function to counteract immune signaling responses and to induce apoptosis. YopB contains two predicted transmembrane helices (residues 166 to 188 and 228 to 250) that are thought to insert into the host plasma membrane during translocation. YopB is also required for pore formation and host-cell-signaling responses to the type III machinery, and these functions of YopB may also require membrane insertion. To elucidate the importance of membrane insertion for YopB function, YopB proteins containing helix-disrupting double consecutive proline substitutions in the center of each transmembrane domain were constructed.
Yersinia pseudotuberculosis
strains expressing the mutant YopB proteins were used to infect macrophages or epithelial cells. Effector translocation, pore formation, and host-cell-signaling responses were studied. Introduction of helix-disrupting substitutions into the second transmembrane domain of YopB resulted in a nonfunctional protein that was not secreted by the type III machinery. Introduction of helix-disrupting substitutions into the first transmembrane domain of YopB resulted in a protein that was fully functional for secretion and for interaction with YopD, another component of the translocation machinery. However, the YopB protein with helix-disrupting substitutions in the first transmembrane domain was partially defective for translocation, pore formation, and signaling, suggesting that all three functions of YopB involve insertion into host membrane.