bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sensitive Detection of HBsAg Mutants by a Gap Ligase Chain Reaction Assay
Authors: Osiowy Carla 
Published: July 2002

ABSTRACT
Hepatitis B virus (HBV) surface gene variants have been associated with diagnostic escape and immune escape following vaccination. The most common mutation observed in these variants is a glycine-to-arginine substitution at amino acid 145 (G145R). In order to sensitively detect the presence of this mutant in serum, a new molecular detection system was developed; in this new system, a gap ligase chain reaction (gLCR) assay was coupled with electrochemiluminescence detection of reaction products. The gLCR assay could detect approximately 10 copies of mutant DNA and could discriminate low levels of mutant DNA in the presence of excess wild-type DNA. Detection of the G145R mutant in clinical specimens was evaluated by testing 56 suspect serum specimens. The G145R mutation was observed in 18 of 28 HBV-DNA-positive samples. The approximate percentage of mutant present in each specimen was calculated by comparison with a standard curve of an increasing ratio of mutant DNA to wild-type DNA. Most samples contained a very low percentage of mutant virus (approximately 5%), with an observed range of approximately 3 to 74%. The G145R mutation was most frequently observed in specimens producing a diagnostic anomaly or from transplant patients but was also observed in specimens from vaccinated individuals and specimens in which HBsAg diagnostic escape was suspected. Therefore, the gLCR assay is a sensitive and specific method for detection of G145R mutants, which could be modified to include the detection of other HBV mutants.
Genetic Diversity of the Outer Surface Protein C Gene of SouthernBorreliaIsolates and Its Possible Epidemiological, Clinical, and Pathogenetic Implications
Authors: Lin Tao , Oliver, James H. , Gao Lihui 
Published: July 2002

ABSTRACTTheospCgenes of 20 southernBorreliastrains were sequenced. The strains consisted ofB. burgdorferisensu stricto,B. andersonii,B. bissettii, one undescribed genospecies, MI-8, and one probably newBorreliaspecies, TXW-1. A high degree of similarity exists betweenB. burgdorferisensu stricto andB. bissettiiand betweenB. bissettiiandB. andersonii. Lateral transfers of theospCgene probably occurred betweenB. burgdorferisensu stricto andB. bissettiiand betweenB. bissettiiandB. andersonii. Internal gene recombination appears to occur among them. The highest degree of genetic diversity among them was observed in the two variable domains (V1 and V2), semivariable domain (SV), and the species-specific epitopes (between amino acids 28 and 31). Differences inospCsequences among southern strains reflect diversity at the strain and genospecies levels. MI-8, which was recognized as an undescribed genospecies in our previous reports, remains distinguishable in our current analysis ofospCgenes and is distinct fromB. burgdorferisensu stricto. Interestingly, another undescribed southern isolate, TXW-1, was not amplified under various PCR conditions. Compared to EuropeanB. burgdorferisensu stricto strains, AmericanB. burgdorferisensu stricto strains show greater genetic heterogeneity. SouthernB. burgdorferisensu stricto,B. andersonii, andB. bissettiiisolates were intermixed with each other in the phylogenetic trees. In the derived trees in our work, at least one southeastern strain ofB. burgdorferi, MI-2, most closely aligns with a so-called invasive cluster that possesses many proven human-invasive strains. Transmission experiments show that MI-2 and the strains in this group of southern spirochetes are able to infect mice and hamsters and that the typical vector of Lyme disease,Ixodes scapularis, can acquire the spirochetes from infected mammals. Currently, strain MI-2 appears to be the only southern isolate among the 20 we analyzed that clusters with an OspC invasive group and thus might be invasive for humans.
Accuracy of Results Obtained by Performing a Second Ligase Chain Reaction Assay and PCR Analysis on Urine Samples with Positive or Near-Cutoff Results in the LCx Test for
Chlamydia trachomatis
Authors: Castriciano S. , Luinstra K. , Jang D. , Patel J. , Mahony J. , Kapala J. , Chernesky M. 
Published: July 2002

ABSTRACT

Nucleic acid amplification assays such as the ligase chain reaction and PCR have encountered reproducibility problems. The initial extract and a newly extracted aliquot of urine specimens (
n
= 120) which had signal-to-cutoff (S/CO) ratios above 0.80 by the LCx Chlamydia assay were retested. Nucleic acid was extracted from an additional urine sample for testing by the AMPLICOR PCR Chlamydia assay. Fifteen percent (18 of 120) of the urine specimens were negative by all repeat tests (initial mean S/CO ratio by the LCx Chlamydia assay, 0.93; S/CO ratio range, 0.80 to 3.30). Repeat testing of the 102 specimens with possible positive results by the LCx Chlamydia assay by use of the initially extracted aliquot confirmed the results for 95 (93.1%) of the specimens; repeat testing of a newly extracted aliquot confirmed the results for 87 (85.3%) of the specimens. Twenty specimens had discordant results by the two repeat LCx Chlamydia assays. A total of 78 of 102 (76.5%) of the specimens were positive by the AMPLICOR PCR, and the AMPLICOR PCR confirmed the results for 82.1% (78 of 95) and 89.6% (78 of 87) of the specimens positive by the two repeat LCx Chlamydia assays, respectively. Some of the discrepancies observed by multiple repeat tests may have been due to specimen mislabeling or contamination during performance of the procedure rather than to the LCx Chlamydia assay. Both assays suffered from a lack of reproducibility on repeat testing with a small proportion of specimens, probably due to the presence of low levels of DNA, the presence of variable amounts of amplification inhibitors, and the loss of DNA during extraction.
Native Valve Endocarditis Due to
Enterococcus hirae
Authors: Poyart Claire , Lambert Thierry , Morand Philippe , Abassade Philippe , Quesne Gilles , Baudouy Yves , Trieu-Cuot Patrick 
Published: July 2002

ABSTRACT

Enterococcus hirae
is a rare isolate in clinical specimens. We describe a case of native aortic-valve endocarditis that was caused by
Enterococcus hirae
in a 72-year-old man. This is the first reported case of endocarditis due to this organism.
Fatal
Saccharomyces cerevisiae
Aortic Graft Infection
Authors: Smith Davey , Metzgar David , Wills Christopher , Fierer Joshua 
Published: July 2002

ABSTRACT

Saccharomyces cerevisiae
is a yeast commonly used in baking and a frequent colonizer of human mucosal surfaces. It is considered relatively nonpathogenic in immunocompetent adults (J. N. Aucott, J. Fayan, H. Grossnicklas, A. Morrissey, M. M. Lederman, and R. A. Salata, Rev. Infect. Dis. 12:406-411, 1990). We present a case of
S. cerevisiae
fungemia and aortic graft infection in an immunocompetent adult. This is the first reported case of
S. cerevisiae
fungemia where the identity of the pathogen was confirmed by rRNA sequencing.
Neutralizing Anti-F Glycoprotein and Anti-Substance P Antibody Treatment Effectively Reduces Infection and Inflammation Associated with Respiratory Syncytial Virus Infection
Authors: Haynes Lia M. , Tonkin Joelyn , Anderson Larry J. , Tripp Ralph A. 
Published: July 2002

ABSTRACTRespiratory syncytial virus (RSV) is the most important virus mediating lower respiratory tract illness in infants and young children. RSV infection is associated with pulmonary inflammation and increased levels of substance P (SP), making the airways and leukocytes that express SP receptors susceptible to the proinflammatory effects of this peptide. This study examines combining neutralizing anti-F glycoprotein and anti-SP antibody treatment of RSV-infected BALB/c mice to inhibit RSV replication and inflammation associated with infection. BALB/c mice were prophylactically treated with antibody prior to RSV infection or were therapeutically treated at day 2 or 6 post-RSV infection. Prophylactic or therapeutic treatment with anti-SP antibodies promptly reduced pulmonary inflammatory cell infiltration and decreased the number of cells expressing proinflammatory cytokines, while anti-F antibody treatment reduced virus titers. The results suggest that combined anti-viral and anti-SP antibody treatment may be effective in treating RSV disease.
Complementary Roles of Multiple Nuclear Targeting Signals in the Capsid Proteins of the Parvovirus Minute Virus of Mice during Assembly and Onset of Infection
Authors: Lombardo Eleuterio , Ramírez Juan C. , Garcia Javier , Almendral José M. 
Published: July 2002

ABSTRACT
This report describes the distribution of conventional nuclear localization sequences (NLS) and of a beta-stranded so-called nuclear localization motif (NLM) in the two proteins (VP1, 82 kDa; VP2, 63 kDa) forming the T=1 icosahedral capsid of the parvovirus minute virus of mice (MVM) and their functions in viral biogenesis and the onset of infection. The approximately 10 VP1 molecules assembled in the MVM particle harbor in its 142-amino-acid (aa) N-terminal-specific region four clusters of basic amino acids, here called BC1 (aa 6 to 10), BC2 (aa 87 to 90), BC3 (aa 109 to 115), and BC4 (aa 126 to 130), that fit consensus NLS and an NLM placed toward the opposite end of the polypeptide (aa 670 to 680) found to be necessary for VP2 nuclear uptake. Deletions and site-directed mutations constructed in an infectious MVM plasmid showed that BC1, BC2, and NLM are cooperative nuclear transport sequences in singly expressed VP1 subunits and that they conferred nuclear targeting competence on the VP1/VP2 oligomers arising in normal infection, while BC3 and BC4 did not display nuclear transport activity. Notably, VP1 proteins mutated at BC1 and -2, and particularly with BC1 to -4 sequences deleted, induced nuclear and cytoplasmic foci of colocalizing conjugated ubiquitin that could be rescued from the ubiquitin-proteasome degradation pathway by the coexpression of VP2 and NS2 isoforms. These results suggest a role for VP2 in viral morphogenesis by assisting cytoplasmic folding of VP1/VP2 subviral complexes, which is further supported by the capacity of NLM-bearing transport-competent VP2 subunits to recruit VP1 into the nuclear capsid assembly pathway regardless of the BC composition. Instead, all four BC sequences, which are located in the interior of the capsid, were absolutely required by the incoming infectious MVM particle for the onset of infection, suggesting either an important conformational change or a disassembly of the coat for nuclear entry of a VP1-associated viral genome. Therefore, the evolutionarily conserved BC sequences and NLM domains provide complementary nuclear transport functions to distinct supramolecular complexes of capsid proteins during the autonomous parvovirus life cycle.
Immune Control of the Number and Reactivation Phenotype of Cells Latently Infected with a Gammaherpesvirus
Authors: Tibbetts Scott A. , van Dyk Linda F. , Speck Samuel H. , Virgin Herbert W. 
Published: July 2002

ABSTRACTDespite active immune responses, gammaherpesviruses establish latency. In a related process, these viruses also persistently replicate by using a mechanism that requires different viral genes than acute-phase replication. Many questions remain about the role of immunity in chronic gammaherpesvirus infection, including whether the immune system controls latency by regulating latent cell numbers and/or other properties and what specific immune mediators control latency and persistent replication. We show here that CD8+T cells regulate both latency and persistent replication and demonstrate for the first time that CD8+T cells regulate both the number of latently infected cells and the efficiency with which infected cells reactivate from latency. Furthermore, we show that gamma interferon (IFN-γ) and perforin, which play no significant role during acute infection, are essential for immune control of latency and persistent replication. Surprisingly, the effects of perforin and IFN-γ are site specific, with IFN-γ being important in peritoneal cells while perforin is important in the spleen. Studies of the mechanisms of action of IFN-γ and perforin revealed that perforin acts primarily by controlling the number of latently infected cells while IFN-γ acts primarily by controlling reactivation efficiency. The immune system therefore controls chronic gammaherpesvirus infection by site-specific mechanisms that regulate both the number and reactivation phenotype of latently infected cells.
Importance of the Cytoplasmic Tails of the Measles Virus Glycoproteins for Fusogenic Activity and the Generation of Recombinant Measles Viruses
Authors: Moll Markus , Klenk Hans-Dieter , Maisner Andrea 
Published: July 2002

ABSTRACT
The generation of replication-competent measles virus (MV) depends on the incorporation of biologically active, fusogenic glycoprotein complexes, which are required for attachment and penetration into susceptible host cells and for direct virus spread by cell-to-cell fusion. Whereas multiple studies have analyzed the importance of the ectodomains of the MV glycoproteins hemagglutinin (H) and fusion protein (F), we have investigated the role of the cytoplasmic tails of the F and H proteins for the formation of fusogenic complexes. Deletions in the cytoplasmic tails of transiently expressed MV glycoproteins were found to have varying effects on receptor binding, fusion, or fusion promotion activity. F tail truncation to only three amino acids did not affect fusion capacity. In contrast, truncation of the H cytoplasmic tail was limited. H protein mutants with cytoplasmic tails of <14 residues no longer supported F-mediated cell fusion, predominantly due to a decrease in surface expression and receptor binding. This indicates that a minimal length of the H protein tail of 14 amino acids is required to ensure a threshold local density to have sufficient accumulation of fusogenic H-F complexes. By using reverse genetics, a recombinant MV with an F tail of three amino acids (rMV-FcΔ30), as well as an MV with an H tail of 14 residues (rMV-HcΔ20), could be rescued, whereas generation of viruses with shorter H tails failed. Thus, glycoprotein truncation does not interfere with the successful generation of recombinant MV if fusion competence is maintained.
CD4 T-Cell-Mediated Demyelination Is Increased in the Absence of Gamma Interferon in Mice Infected with Mouse Hepatitis Virus
Authors: Pewe Lecia , Haring Jodie , Perlman Stanley 
Published: July 2002

ABSTRACTMice infected with the murine coronavirus, mouse hepatitis virus, strain JHM (MHV) develop an immune-mediated demyelinating encephalomyelitis. Adoptive transfer of MHV-immune splenocytes depleted of either CD4 or CD8 T cells to infected mice deficient in recombination activation gene 1 resulted in demyelination. We showed previously that the process of CD8 T-cell-mediated demyelination was strongly dependent on the expression of gamma interferon (IFN-γ) by donor cells. In this report, we show, in contrast, that demyelination and lymphocyte infiltration were increased in recipients of IFN-γ−/−CD4 T cells when compared to levels in mice receiving C57BL/6 CD4 T cells.
Matrix Metalloproteinase Expression Correlates with Virulence following Neurotropic Mouse Hepatitis Virus Infection
Authors: Zhou Jiehao , Stohlman Stephen A. , Atkinson Roscoe , Hinton David R. , Marten Norman W. 
Published: July 2002

ABSTRACTThe relationship(s) between viral virulence and matrix metalloproteinase (MMP) expression in the central nervous system (CNS) of mice undergoing lethal and sublethal infections with neurotropic mouse hepatitis virus was investigated. Lethal infection induced increased levels of MMP-3 and MMP-12 mRNAs as well as that of tissue inhibitor of matrix metalloproteinases 1 (TIMP-1) compared to sublethal infection. Increased induction of MMP, TIMP, and chemokine expression correlated with increased virus replication but not with inflammatory cell infiltration. Infection of immunosuppressed mice suggested that expression of most MMP, TIMP, and chemokine mRNA was induced primarily in CNS-resident cells. By contrast, MMP-9 protein activity was associated with the infiltration of neutrophils into the CNS. These data indicate an association between the magnitude of inflammatory gene expression within the CNS and viral virulence.
Avian and Mammalian Hepadnaviruses Have Distinct Transcription Factor Requirements for Viral Replication
Authors: Tang Hong , McLachlan Alan 
Published: July 2002

ABSTRACT
Hepadnavirus replication occurs in hepatocytes in vivo and in hepatoma cell lines in cell culture. Hepatitis B virus (HBV) replication can occur in nonhepatoma cells when pregenomic RNA synthesis from viral DNA is activated by the expression of the nuclear hormone receptors hepatocyte nuclear factor 4 (HNF4) and the retinoid X receptor α (RXRα) plus peroxisome proliferator-activated receptor α (PPARα) heterodimer. Nuclear hormone receptor-dependent HBV replication is inhibited by hepatocyte nuclear factor 3 (HNF3). In contrast, HNF3 and HNF4 support duck hepatitis B virus (DHBV) replication in nonhepatoma cells, whereas the RXRα-PPARα heterodimer inhibits HNF4-dependent DHBV replication. HNF3 and HNF4 synergistically activate DHBV pregenomic RNA synthesis and viral replication. The conditions that support HBV or DHBV replication in nonhepatoma cells are not able to support woodchuck hepatitis virus replication. These observations indicate that avian and mammalian hepadnaviruses have distinct transcription factor requirements for viral replication.
Zinc Finger Domain of Murine Leukemia Virus Nucleocapsid Protein Enhances the Rate of Viral DNA Synthesis in Vivo
Authors: Zhang Wen-hui , Hwang Carey K. , Hu Wei-Shau , Gorelick Robert J. , Pathak Vinay K. 
Published: July 2002

ABSTRACT

In vitro studies have indicated that retroviral nucleocapsid (NC) protein facilitates both DNA synthesis by reverse transcriptase (RT) and annealing of the nascent DNA with acceptor template. Increasing the rate of DNA synthesis is expected to reduce the frequency of RT template switching, whereas annealing the nascent DNA with acceptor template promotes template switching. We performed a mutational analysis of the murine leukemia virus (MLV) NC zinc finger domain to study its effect on RT template switching in vivo and to explore the role of NC during reverse transcription. The effects of NC mutations on RT template switching were determined by using a previously described in vivo direct-repeat deletion assay. A
trans
-complementation assay was also developed in which replication-defective NC mutants were rescued by coexpression of replication-defective RT mutants that provided wild-type NC in
trans
. We found that mutations in the MLV NC zinc finger domain increased the frequency of template switching approximately twofold. When a predicted stem-loop RNA secondary structure was introduced into the template RNA, the template-switching frequency increased 5-fold for wild-type NC and further increased up to an additional 6-fold for NC zinc finger domain mutants, resulting in an overall increase of as much as 30-fold. Thus, wild-type NC increased the efficiency with which RT was able to reverse transcribe through regions of RNA secondary structure that might serve as RT pause sites. These results provide the first in vivo evidence that NC enhances the rate of DNA synthesis by RT in regions of the template possessing stable RNA secondary structure.
Reassessing the Role of Region A in Pit1-Mediated Viral Entry
Authors: Farrell Karen B. , Russ Jill L. , Murthy Ravi K. , Eiden Maribeth V. 
Published: July 2002

ABSTRACT
The mammalian gammaretroviruses gibbon ape leukemia virus (GALV) and feline leukemia virus subgroup B (FeLV-B) can use the same receptor, Pit1, to infect human cells. A highly polymorphic nine-residue sequence within Pit1, designated region A, has been proposed as the virus binding site, because mutations in this region abolish Pit1-mediated cellular infection by GALV and FeLV-B. However, a direct correlation between region A mutations deleterious for infection and loss of virus binding has not been established. We report that cells expressing a Pit1 protein harboring mutations in region A that abolish receptor function retain the ability to bind virus, indicating that Pit1 region A is not the virus binding site. Furthermore, we have now identified a second region in Pit1, comprising residues 232 to 260 (region B), that is required for both viral entry and virus binding. Epitope-tagged Pit1 proteins were used to demonstrate that mutations in region B result in improper orientation of Pit1 in the cell membrane. Compensatory mutations in region A can restore proper orientation and full receptor function to these region B mutants. Based on these results, we propose that region A of Pit1 confers competence for viral entry by influencing the topology of the authentic binding site in the membrane and hence its accessibility to a viral envelope protein. Based on glycosylation studies and results obtained by using N- and C-terminal epitope-tagged Pit1, region A and region B mutants, and the transmembrane helices predicted with the PHD PredictProtein algorithm, we propose a new Pit1 topology model.
Contribution of Protein p40 to Hypovirus-Mediated Modulation of Fungal Host Phenotype and Viral RNA Accumulation
Authors: Suzuki Nobuhiro , Nuss Donald L. 
Published: July 2002

ABSTRACT

The papain-like protease p29, derived from the N-terminal portion of the hypovirus CHV1-EP713-encoded open reading frame (ORF) A polyprotein, p69, was previously shown to contribute to reduced pigmentation and sporulation by the infected host, the chestnut blight fungus
Cryphonectria parasitica
, while being dispensable for virus replication and attenuation of fungal virulence (hypovirulence). We now report that deletion of the C-terminal portion of p69, which encodes the highly basic protein p40, resulted in replication-competent mutant viruses that were, however, significantly reduced in RNA accumulation. While the Δp40 mutants retained the ability to confer hypovirulence, Δp40-infected fungal strains produced more asexual spores than strains infected with either wild-type CHV1-EP713 or a Δp29 mutant virus. As observed for Δp29-infected colonies, pigment production was significantly increased in Δp40-infected fungal strains relative to that in CHV1-EP713-infected strains. Virus-mediated suppression of laccase production was not affected by p40 deletion. A gain-of-function analysis was employed to map the p40 symptom determinant to the N-terminal domain, encompassing p69 amino acid residues Thr(288) to Arg(312). Evidence that the gain of function was due to the encoded protein rather than the corresponding RNA sequence element was provided by introducing frameshift mutations on either side of the activity determinant domain. Moreover, restoration of symptoms correlated with increased accumulation of viral RNA. These results suggest that p40 indirectly contributes to virus-mediated suppression of fungal pigmentation and conidiation by providing an accessory function in hypovirus RNA amplification. A possible role for p40 in facilitating ORF B expression and the relationship between hypovirus RNA accumulation and symptom expression are discussed.