bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mimotopes of Apical Membrane Antigen 1: Structures of Phage-Derived Peptides Recognized by the Inhibitory Monoclonal Antibody 4G2dc1 and Design of a More Active Analogue
Authors: Sabo Jennifer K. , Keizer David W. , Feng Zhi-Ping , Casey Joanne L. , Parisi Kathy , Coley Andrew M. , Foley Michael , Norton Raymond S. 
Published: December 2006

ABSTRACT

Apical membrane antigen 1 (AMA1) of the malaria parasite
Plasmodium falciparum
is an integral membrane protein that plays a key role in merozoite invasion of host erythrocytes. A monoclonal antibody, 4G2dc1, recognizes correctly folded AMA1 and blocks merozoite invasion. Phage display was used to identify peptides that bind to 4G2dc1 and mimic an important epitope of AMA1. Three of the highest-affinity binders—J1, J3, and J7—were chosen for antigenicity and immunogenicity studies. J1 and J7 were found to be true antigen mimics since both peptides generated inhibitory antibodies in rabbits (J. L. Casey et al., Infect. Immun. 72:1126-1134, 2004). In the present study, the solution structures of all three mimotopes were investigated by nuclear magnetic resonance spectroscopy. J1 adopted a well-defined region of structure, which can be attributed in part to the interactions of Trp11 with surrounding residues. In contrast, J3 and J7 did not adopt an ordered conformation over the majority of residues, although they share a region of local structure across their consensus sequence. Since J1 was the most structured of the peptides, it provided a template for the design of a constrained analogue, J1cc, which shares a structure similar to that of J1 and has a disulfide-stabilized conformation around the Trp11 region. J1cc binds with greater affinity to 4G2dc1 than does J1. These peptide structures provide the foundation for a better understanding of the complex conformational nature of inhibitory epitopes on AMA1. With its greater conformational stability and higher affinity for AMA1, J1cc may be a better in vitro correlate of immunity than the peptides identified by phage display.
Resistance of Primary Murine CD4
+
T Cells to
Helicobacter pylori
Vacuolating Cytotoxin
Authors: Algood Holly M. Scott , Torres Victor J. , Unutmaz Derya , Cover Timothy L. 
Published: December 2006

ABSTRACT

Persistent colonization of the human stomach by
Helicobacter pylori
is a risk factor for the development of gastric cancer and peptic ulcer disease.
H. pylori
secretes a toxin, VacA, that targets human gastric epithelial cells and T lymphocytes and enhances the ability of
H. pylori
to colonize the stomach in a mouse model. To examine how VacA contributes to
H. pylori
colonization of the mouse stomach, we investigated whether murine T lymphocytes were susceptible to VacA activity. VacA inhibited interleukin-2 (IL-2) production by a murine T-cell line (LBRM-33), similar to its effects on a human T-cell line (Jurkat), but did not inhibit IL-2 production by primary murine splenocytes or CD4
+
T cells. VacA inhibited activation-induced proliferation of primary human CD4
+
T cells but did not inhibit the proliferation of primary murine CD4
+
T cells. Flow cytometry studies indicated that the levels of VacA binding to primary murine CD4
+
T cells were significantly lower than levels of VacA binding to human CD4
+
T cells. This suggests that the resistance of primary murine CD4
+
T cells to VacA is attributable, at least in part, to impaired VacA binding to these cells.
The Immunologically Distinct O Antigens fromFrancisella tularensisSubspeciestularensisandFrancisella novicidaAre both Virulence Determinants and Protective Antigens
Authors: Thomas Rebecca M. , Titball Richard W. , Oyston Petra C. F. , Griffin Kate , Waters Emma , Hitchen Paul G. , Michell Stephen L. , Grice I. Darren , Wilson Jennifer C. , Prior Joann L. 
Published: December 2006

ABSTRACTWe have determined the sequence of the gene cluster encoding the O antigen inFrancisella novicidaand compared it to the previously reported O-antigen cluster inFrancisella tularensissubsp.tularensis. Immunization with purified lipopolysaccharide (LPS) fromF. tularensissubsp.tularensisorF. novicidaprotected against challenge withFrancisella tularensissubsp.holarcticaandF. novicida, respectively. The LPS fromF. tularensissubsp.tularensisdid not confer protection against challenge withF. novicida, and the LPS fromF. novicidadid not confer protection against challenge withF. tularensissubsp.holarctica. Allelic replacement mutants ofF. tularensissubsp.tularensisorF. novicidawhich failed to produce O antigen were attenuated, but exposure to these mutants did not induce a protective immune response. The O antigen ofF. tularensissubsp.tularensisappeared to be important for intracellular survival whereas the O antigen ofF. novicidaappeared to be critical for serum resistance and less important for intracellular survival.
The Combined CTA1-DD/ISCOMs Vector Is an Effective Intranasal Adjuvant for Boosting PriorMycobacterium bovisBCG Immunity toMycobacterium tuberculosis
Authors: Andersen Claire Swetman , Dietrich Jes , Agger Else Marie , Lycke Nils Y. , Lövgren Karin , Andersen Peter 
Published: December 2006

ABSTRACTInfection withMycobacterium tuberculosis, the causative agent of tuberculosis (TB), remains one of the leading causes of mortality worldwide. The current “gold standard” vaccineMycobacterium bovisBCG has a limited efficacy that wanes over time. The development of a vaccine to boost BCG-induced immunity is therefore a highly active area of research. Mucosal administration of vaccines is believed to provide better protection against pathogens, such asM. tuberculosis, that invade the host via mucosal surfaces. In this study we demonstrate that an intranasal vaccine, comprising the antigenic fusion protein Ag85B-ESAT-6 and the mucosal combined adjuvant vector CTA1-DD/ISCOMs, strongly promotes a Th1-specific immune response, dominated by gamma interferon-secreting CD4-positive T cells. Mucosal administration of Ag85B-ESAT-6 mixed with CTA1-DD/ISCOMs strongly boosted prior BCG immunity, leading to a highly increased recruitment of antigen-specific cells to the site of infection. Most importantly, we observed a significantly (P< 0.001) reduced bacterial burden in the lung compared to nonboosted control animals. Thus, the results demonstrate the effectiveness of mucosal vaccination with Ag85B-ESAT-6 mixed with CTA1-DD/ISCOMs as adjuvant for stimulating TB-specific protective immunity in the lung.
The TolC Homologue of
Brucella suis
Is Involved in Resistance toAntimicrobial Compounds and Virulence
Authors: Posadas Diana M. , Martín Fernando A. , Sabio y García Julia V. , Spera Juan M. , Delpino M. Victoria , Baldi Pablo , Campos Eleonora , Cravero Silvio L. , Zorreguieta Angeles 
Published: December 2006

ABSTRACT

Brucella
spp., like other pathogens, must cope with the environment of diverse
host niches during the infection process. In doing this, pathogens
evolved different type of transport systems to help them survive and
disseminate within the host. Members of the TolC family have been shown
to be involved in the export of chemically diverse molecules ranging
from large protein toxins to small toxic compounds. The role of
proteins from the TolC family in
Brucella
and otherα
-2-proteobacteria has been explored little. The gene encoding
the unique member of the TolC family from
Brucella suis
(BepC)
was cloned and expressed in an
Escherichia coli
mutant
disrupted in the gene encoding TolC, which has the peculiarity of being
involved in diverse transport functions. BepC fully complemented the
resistance to drugs such as chloramphenicol and acriflavine but was
incapable of restoring hemolysin secretion in the
tolC
mutant
of
E. coli
. An insertional mutation in the
bepC
gene
strongly affected the resistance phenotype of
B. suis
to bile
salts and toxic chemicals such as ethidium bromide and rhodamine and
significantly decreased the resistance to antibiotics such as
erythromycin, ampicillin, tetracycline, and norfloxacin. Moreover, the
B. suis bepC
mutant was attenuated in the mouse model of
infection. Taken together, these results suggest that BepC-dependent
efflux processes of toxic compounds contribute to
B. suis
survival inside the
host.
Identification of a Glycosylated
Ehrlichia canis
19-Kilodalton Major Immunoreactive Protein with a Species-Specific Serine-Rich Glycopeptide Epitope
Authors: McBride Jere W. , Doyle C. Kuyler , Zhang Xiaofeng , Cardenas Ana Maria , Popov Vsevolod L. , Nethery Kimberly A. , Woods Michael E. 
Published: December 2006

ABSTRACT

Ehrlichia canis
has a small subset of major immunoreactive proteins that includes a 19-kDa protein that elicits an early
Ehrlichia
-specific antibody response in infected dogs. We report herein the identification and molecular characterization of this highly conserved 19-kDa major immunoreactive glycoprotein (gp19) ortholog of the
Ehrlichia chaffeensis
variable-length PCR target (VLPT) protein.
E. canis
gp19 has substantial carboxyl-terminal amino acid homology (59%) with
E. chaffeensis
VLPT and the same chromosomal location; however, the
E. chaffeensis
VLPT gene (594 bp) has tandem repeats that are not present in the
E. canis
gp19 gene (414 bp). Consistent with other ehrlichial glycoproteins, the gp19 protein exhibited a larger-than-predicted mass (∼3 kDa), O-linked glycosylation sites were predicted in an amino-terminal serine/threonine/glutamate (STE)-rich patch (26 amino acids), carbohydrate was detected on the recombinant gp19 protein, and the neutral sugars glucose and galactose were detected on the recombinant amino-terminal polypeptide.
E. canis
gp19 composition consists of five predominant amino acids, cysteine, glutamate, tyrosine, serine, and threonine, concentrated in the STE-rich patch and a carboxyl-terminal domain predominated by cysteine and tyrosine (55%). The amino-terminal STE-rich patch contained a major species-specific antibody epitope strongly recognized by serum from an
E. canis-
infected dog. The recombinant glycopeptide epitope was substantially more reactive with antibody than the synthetic (nonglycosylated) peptide, and periodate treatment of the recombinant glycopeptide epitope reduced its immunoreactivity, demonstrating the importance of a carbohydrate immunodeterminant(s). The gp19 protein was present on reticulate and dense-cored cells, and it was found extracellularly in the fibrillar matrix and associated with the morula membrane, the host cell cytoplasm, and the nucleus.
A Genotype-Independent Real-Time PCR Assay for Quantification of Hepatitis B Virus DNA
Authors: Liu Ying , Hussain Munira , Wong Stephen , Fung Scott K. , Yim Hyung Joon , Lok Anna S. F. 
Published: December 2006

ABSTRACT

Accurate quantification of hepatitis B virus (HBV) DNA levels is important for monitoring patients with chronic HBV infection and for assessing their responses to antiviral therapy. This study aimed to develop a real-time PCR assay that is sensitive and can accurately quantify a wide range of HBV DNA levels across the known HBV genotypes. An “in-house” real-time PCR assay using primers and a TaqMan probe in a highly conserved region of the HBV surface gene was designed. The assay was standardized against a WHO standard and validated against plasmids of HBV genotypes A through H. The linear quantification range was approximately 5 × 10
0
to 2.0 × 10
9
IU/ml. Results of samples from patients infected with HBV genotypes A through H tested using our real-time “in-house” PCR assay showed an excellent correlation with those of the Cobas Amplicor HBV Monitor (
R
2
= 0.9435) and the Cobas TaqMan HBV (
R
2
= 0.9873) tests. We have established a real-time PCR assay that is genotype independent and can accurately quantify a wide range of HBV DNA levels. Further studies of additional samples are ongoing to validate the genotype independence of our assay.
Comparative Study of Kaposi's Sarcoma-Associated Herpesvirus Serological Assays Using Clinically and Serologically Defined Reference Standards and Latent Class Analysis
Authors: Nascimento Maria Claudia , de Souza Vanda Akico , Sumita Laura Masami , Freire Wilton , Munoz Fernando , Kim Joseph , Pannuti Claudio S. , Mayaud Philippe 
Published: December 2006

ABSTRACT
Accurate determination of infection with Kaposi's sarcoma-associated herpesvirus (KSHV) has been hindered by the lack of a “gold standard” for comparison of serological assays used to estimate KSHV prevalence in serosurveys conducted in different settings. We have evaluated the performance of five in-house (developed at University College London [UCL], United Kingdom, and at the virology laboratory of the Instituto de Medicine Tropical [IMT] in Sao Paulo, Brazil) and two commercial (ABI and DIAVIR) serological assays to detect antibodies to latency-associated nuclear antigen (LANA) and to lytic KSHV antigens. We used a variety of serum samples assembled to represent populations likely to be at high, intermediate, and low risk of KSHV infection in Brazil. Composite reference standard panels were prepared based on clinical and serological parameters, against which assay performances were assessed using conventional Bayesian statistics and latent class analysis (LCA). Against the clinical reference standard, in-house immunofluorescence assays to detect anti-LANA antibodies (IFA-LANA) produced at UCL and IMT had similar performances, with sensitivities of 61% (95% confidence interval [CI], 48% to 74%) and 72% (95% CI, 58% to 83%) and specificities of 99% (95% CI, 94% to 100%) and 100% (95% CI, 96% to 100%), respectively, and only the IMT IFA-LANA was included in LCA, together with the IMT IFA-lytic and four enzyme-linked immunosorbent assays (ELISAs). The LCA indicated that the IMT whole-virus ELISA performed best (sensitivity, 87% [95% CI, 81% to 91%]; and specificity, 100% [95% CI, 98% to 100%]), confirming the results obtained with the conventional statistical approach. Commercially available ELISA-based tests yielded the lowest specificities using a spectrum of serum samples. The evaluation of KSHV serological assays is warranted before planning serosurveys in various settings.
Rapid and Highly Sensitive Pathotyping of Avian Influenza A H5N1 Virus by Using Real-Time Reverse Transcription-PCR
Authors: Hoffmann Bernd , Harder Timm , Starick Elke , Depner Klaus , Werner Ortrud , Beer Martin 
Published: December 2006

ABSTRACT
Rapid typing of the pathogenicity of avian influenza A viruses (AIV) of subtypes H5 and H7 is crucial to initiate adequate protective measures preventing the spread of highly pathogenic AIV (HPAIV). Here, a new real-time reverse transcription-PCR assay which enables sensitive and specific detection and cleavage site analysis of HPAIV H5N1 of the Qinghai lineage is described.
Use of New T-Cell-Based Cell Lines Expressing Two Luciferase Reporters for Accurately Evaluating Susceptibility to Anti-Human Immunodeficiency Virus Type 1 Drugs
Authors: Chiba-Mizutani Tomoko , Miura Hideka , Matsuda Masakazu , Matsuda Zene , Yokomaku Yoshiyuki , Miyauchi Kosuke , Nishizawa Masako , Yamamoto Naoki , Sugiura Wataru 
Published: December 2006

ABSTRACTTwo new T-cell-based reporter cell lines were established to measure human immunodeficiency virus type 1 (HIV-1) infectivity. One cell line naturally expresses CD4 and CXCR4, making it susceptible to X4-tropic viruses, and the other cell line, in which a CCR5 expression vector was introduced, is susceptible to both X4- and R5-tropic viruses. Reporter cells were constructed by transfecting the human T-cell line HPB-Ma, which demonstrates high susceptibility to HIV-1, with genomes expressing two different luciferase reporters, HIV-1 long terminal repeat-driven firefly luciferase and cytomegalovirus promoter-driven renilla luciferase. Upon HIV infection, the cells expressed firefly luciferase at levels that were highly correlated (r2= 0.91 to 0.98) with the production of the capsid antigen p24. The cells also constitutively expressed renilla luciferase, which was used to monitor cell numbers and viability. The reliability of the cell lines for two in vitro applications, drug resistance phenotyping and drug screening, was confirmed. As HIV-1 efficiently replicated in these cells, they could be used for multiple-round replication assays as an alternative method to a single-cycle replication protocol. Coefficients of variation for drug susceptibility evaluated with the cell lines ranged from 17 to 41%. The new cell lines were beneficial for evaluating antiretroviral drug resistance. Firefly luciferase gave a wider dynamic range for evaluating virus infectivity, and the introduction of renilla luciferase improved assay reproducibility. The cell lines were also beneficial for screening new antiretroviral agents, as false inhibition caused by the cytotoxicity of test compounds was easily detected by monitoring renilla luciferase activity.
GeneChip Resequencing of the Smallpox Virus Genome Can Identify Novel Strains: a Biodefense Application
Authors: Sulaiman Irshad M. , Tang Kevin , Osborne John , Sammons Scott , Wohlhueter Robert M. 
Published: December 2006

ABSTRACT
We developed a set of seven resequencing GeneChips, based on the complete genome sequences of 24 strains of smallpox virus (variola virus), for rapid characterization of this human-pathogenic virus. Each GeneChip was designed to analyze a divergent segment of approximately 30,000 bases of the smallpox virus genome. This study includes the hybridization results of 14 smallpox virus strains. Of the 14 smallpox virus strains hybridized, only 7 had sequence information included in the design of the smallpox virus resequencing GeneChips; similar information for the remaining strains was not tiled as a reference in these GeneChips. By use of variola virus-specific primers and long-range PCR, 22 overlapping amplicons were amplified to cover nearly the complete genome and hybridized with the smallpox virus resequencing GeneChip set. These GeneChips were successful in generating nucleotide sequences for all 14 of the smallpox virus strains hybridized. Analysis of the data indicated that the GeneChip resequencing by hybridization was fast and reproducible and that the smallpox virus resequencing GeneChips could differentiate the 14 smallpox virus strains characterized. This study also suggests that high-density resequencing GeneChips have potential biodefense applications and may be used as an alternate tool for rapid identification of smallpox virus in the future.
Vibrio cholerae
Strain Typing and Phylogeny Study Based on Simple Sequence Repeats
Authors: Danin-Poleg Yael , Cohen Lyora A. , Gancz Hanan , Broza Yoav Y. , Goldshmidt Hanoh , Malul Elinor , Valinsky Lea , Lerner Larisa , Broza Meir , Kashi Yechezkel 
Published: December 2006

ABSTRACT

Vibrio cholerae
is the etiological agent of cholera. Its natural reservoir is the aquatic environment. To date, practical typing of
V. cholerae
is mainly serological and requires about 200 antisera. Simple sequence repeats (SSR), also termed VNTR (for variable number of tandem repeats), provide a source of high genomic polymorphism used in bacterial typing. Here we describe an SSR-based typing method that combines the variation in highly mutable SSR loci, with that of shorter, relatively more stable mononucleotide repeat (MNR) loci, for accurate and rapid typing of
V. cholerae
. In silico screening of the
V. cholerae
genome revealed thousands of perfect SSR tracts with an average frequency of one SSR every 152 bp. A panel of 32
V. cholerae
strains, representing both clinical and environmental isolates, was tested for polymorphism in SSR loci. Two strategies were applied to identify SSR variation: polymorphism of SSR tracts longer than 12 bp (L-SSR) assessed by capillary fragment-size analysis and MNR polymorphism assessed by sequencing. The nine L-SSR loci tested were all polymorphic, displaying 2 to 13 alleles per locus. Sequence analysis of eight MNR-containing loci (MNR-multilocus sequence typing [MLST]) provided information on both variations in the MNR tract itself, and single nucleotide polymorphism (SNP) in their flanking sequences. Phylogenetic analysis of the combined SSR data showed a clear discrimination between the clinical strains belonging to O1 and O139 serogroups, and the environmental isolates. Furthermore, discrimination between 27 strains of the 32 strains was achieved. SSR-based typing methods combining L-SSR and MNR-MLST were found to be efficient for
V. cholerae
typing.
Vancomycin-Resistant
Enterococcus faecalis
Endocarditis: Linezolid Failure and Strain Characterization of Virulence Factors
Authors: Tsigrelis Constantine , Singh Kavindra V. , Coutinho Thais D. , Murray Barbara E. , Baddour Larry M. 
Published: December 2006

ABSTRACT

Infective endocarditis due to vancomycin-resistant (VR)
Enterococcus faecalis
has only rarely been reported. We report a case of VR
E. faecalis
endocarditis that failed to respond to linezolid therapy, outline the virulence traits of the isolate, and review previously published cases of VR
E. faecalis
endocarditis.
Characterization of Primary and Memory CD8 T-Cell Responses against Ranavirus (FV3) inXenopus laevis
Authors: Morales Heidi D. , Robert Jacques 
Published: December 2006

ABSTRACTIn mammals, resistance to primary and secondary viral infections critically involves major histocompatibility complex class I-restricted cytotoxic CD8+T lymphocytes (CTLs). Although many gene homologues involved in CTL function have been identified in all vertebrate classes, antiviral CTL responses have been poorly characterized for ectothermic vertebrates. Because of the threat of emerging wildlife viral diseases to global biodiversity, fundamental research on comparative viral immunity has become crucial. Ranaviruses (familyIridoviridae) are double-stranded DNA viruses possibly implicated in the worldwide decline of amphibian populations. We used the frogXenopus laevisas a model to evaluate adaptive immune responses to the ranavirus frog virus 3 (FV3). FV3 infects the kidneys of adults but is cleared within 4 weeks, with faster clearance upon secondary infections. In vivo depletion of CD8+T cells markedly decreases the survival of adults after viral infection. To further investigate the involvement of anti-FV3 CD8+T-cell effectors in host resistance in vivo, we determined the proliferation kinetics of CD8+T cells in the spleen by bromodeoxyuridine incorporation and their infiltration of kidneys by immunohistology. Upon primary infection, CD8+T cells significantly proliferate in the spleen and accumulate in infected kidneys from day 6 onward, in parallel with virus clearance. Earlier proliferation and infiltration associated with faster viral clearance were observed during a secondary infection. These results provide in vivo evidence of protective antigen-dependent CD8+T-cell proliferation, recognition, and memory in fighting a natural pathogen inXenopus.