bluuebunny/crossref_metadata_2025_split huggingface dataset:
Integrated Human Papillomavirus Type 16 Is Frequently Found in Cervical Cancer Precursors as Demonstrated by a Novel Quantitative Real-Time PCR Technique
Authors: Peitsaro Panu , Johansson Bo , Syrjänen Stina 
Published: July 2002

ABSTRACT
In contrast to cervical cancer, integration of human papillomavirus (HPV) DNA into the host genome has been considered a rare event in cancer precursor lesions (cervical intraepithelial neoplasia [CIN]). With our new real-time PCR method, we demonstrated that integrated HPV type 16 (HPV16) is already present in CIN lesions. The physical state of HPV16 and the viral load were simultaneously detected. A unique region of the E2 open reading frame (ORF) that is most often deleted during HPV16 integration is targeted by one set of PCR primers and a probe, and another set targets the E6 ORF. In episomal form, both targets should be equivalent, while in integrated form, the copy numbers of E2 would be less than those of E6. The method was tested with DNAs from 31 cervical lesions (non-CIN to CINIII) from 24 women prospectively followed up for 10 years. This report presents viral load and integration results from the largest series of CIN lesions described to date. Only one sample contained exclusively episomal HPV16 DNA, and this lesion regressed spontaneously. Samples from another patient, with only integrated HPV16, rapidly progressed from CINI to CINIII in 2 years. In all other patients, episomal and integrated forms of HPV16 DNA were found to coexist. Rapid progression of the CIN lesions was closely associated with a heavy load of integrated HPV16. Thus, the method described here is a very sensitive tool with which to assess the physical state of HPV, which is useful in predicting disease progression.
Limited Genetic Diversity of Recent Invasive Isolates of Non-Serotype b Encapsulated
Haemophilus influenzae
Authors: Omikunle Adebomi , Takahashi Shinji , Ogilvie Chrishana L. , Wang Yan , Rodriguez Carina A. , St. Geme Joseph W. , Adderson Elisabeth E. 
Published: July 2002

ABSTRACT

Invasive infections caused by non-type b encapsulated
Haemophilus influenzae
have increased in frequency in the last decade. This change prompted us to characterize the genetic relationships of 48 recently isolated invasive
H. influenzae
type a (Hia), e (Hie), and f (Hif) strains by comparison of restriction digest patterns (RDPs). Recent Hia isolates exhibited moderate genetic diversity, with the majority segregating into two major clonotypes. Recent Hie and, especially, Hif strains displayed considerably restricted genetic diversity. In particular, all but one Hif strain segregated into a single clonotype, and half of these isolates had identical RDPs. These results are consistent with the hypothesis that the increased incidence of disease due to non-type b encapsulated
H. influenzae
reflects the emergence of hypervirulent clones, especially in the case of Hif. Alternatively, it is possible that non-type b encapsulated
H. influenzae
strains have limited overall genetic diversity.
Fluorescence In Situ Hybridization with Peptide Nucleic Acid Probes for Rapid Identification of
Candida albicans
Directly from Blood Culture Bottles
Authors: Rigby Susan , Procop Gary W. , Haase Gerhard , Wilson Deborah , Hall Geraldine , Kurtzman Cletus , Oliveira Kenneth , Von Oy Sabina , Hyldig-Nielsen Jens J. , Coull James , Stender Henrik 
Published: July 2002

ABSTRACT

A new fluorescence in situ hybridization (FISH) method that uses peptide nucleic acid (PNA) probes for identification of
Candida albicans
directly from positive-blood-culture bottles in which yeast was observed by Gram staining (herein referred to as yeast-positive blood culture bottles) is described. The test (the
C. albicans
PNA FISH method) is based on a fluorescein-labeled PNA probe that targets
C. albicans
26S rRNA. The PNA probe is added to smears made directly from the contents of the blood culture bottle and hybridized for 90 min at 55°C. Unhybridized PNA probe is removed by washing of the mixture (30 min), and the smears are examined by fluorescence microscopy. The specificity of the method was confirmed with 23 reference strains representing phylogenetically related yeast species and 148 clinical isolates covering the clinically most significant yeast species, including
C. albicans
(
n
= 72),
C. dubliniensis
(
n
= 58),
C. glabrata
(
n
= 5),
C. krusei
(
n
= 2),
C. parapsilosis
(
n
= 4), and
C. tropicalis
(
n
= 3). The performance of the
C. albicans
PNA FISH method as a diagnostic test was evaluated with 33 routine and 25 simulated yeast-positive blood culture bottles and showed 100% sensitivity and 100% specificity. It is concluded that this 2.5-h method for the definitive identification of
C. albicans
directly from yeast-positive blood culture bottles provides important information for optimal antifungal therapy and patient management.
Evidence for a Stable Interaction of gp41 with Pr55Gagin Immature Human Immunodeficiency Virus Type 1 Particles
Authors: Wyma Donald J. , Kotov Alexander , Aiken Christopher 
Published: July 2002

ABSTRACTAssembly of infectious human immunodeficiency virus type 1 (HIV-1) virions requires incorporation of the viral envelope glycoproteins gp41 and gp120. Several lines of evidence have suggested that the cytoplasmic tail of the transmembrane glycoprotein, gp41, associates with Pr55Gagin infected cells to facilitate the incorporation of HIV-1 envelope proteins into budding virions. However, direct evidence for an interaction between gp41 and Pr55Gagin HIV-1 particles has not been reported. To determine whether gp41 is associated with Pr55Gagin HIV-1 particles, viral cores were isolated from immature HIV-1 virions by sedimentation through detergent. The cores contained a major fraction of the gp41 that was present on untreated virions. Association of gp41 with cores required the presence of the gp41 cytoplasmic tail. In HIV-1 particles containing a functional protease, a mutation that prevents cleavage of Pr55Gagat the matrix-capsid junction was sufficient for the detergent-resistant association of gp41 with the isolated cores. In addition to gp41, a major fraction of virion-associated gp120 was also detected on immature HIV-1 cores. Isolation of cores under conditions known to disrupt lipid rafts resulted in the removal of a raft-associated protein incorporated into virions but not the HIV-1 envelope proteins. These results provide biochemical evidence for a stable interaction between Pr55Gagand the cytoplasmic tail of gp41 in immature HIV-1 particles. Moreover, findings in this study suggest that the interaction of Pr55Gagwith gp41 may regulate the function of the envelope proteins during HIV-1 maturation.
Role of the Pseudorabies Virus gI Cytoplasmic Domain in Neuroinvasion, Virulence, and Posttranslational N-Linked Glycosylation
Authors: Tirabassi R. S. , Enquist L. W. 
Published: July 2002

ABSTRACTThe glycoproteins I and E of pseudorabies virus are important mediators of cell-to-cell spread and virulence in all animal models tested. Although these two proteins form a complex with one another, ascribing any function to the individual proteins has been difficult. We have shown previously, using nonsense mutations, that the N-terminal ectodomain of the gE protein is sufficient for gE-mediated transsynaptic spread whereas the cytoplasmic domain of the protein is required for full expression of virulence. These same studies demonstrated that the cytoplasmic domain of gE is also required for endocytosis of the protein. In this report, we describe the construction of viruses with nonsense mutations in gI that allowed us to determine the contributions of the gI cytoplasmic domain to protein expression as well as virus neuroinvasion and virulence after infection of the rat eye. We also constructed double mutants with nonsense mutations in both gE and gI so that the contributions of both the gE and gI cytoplasmic domains could be determined. We observed that the gI cytoplasmic domain is required for efficient posttranslational modification of the gI protein. The gE cytoplasmic domain has no effect on gE posttranslational glycosylation. In addition, we found that infection of all gE-gI-dependent anterograde circuits projecting from the rat retina requires both ectodomains and at least one of the cytoplasmic domains of the proteins. The gI cytoplasmic domain promotes transsynaptic spread of virus better than the gE cytoplasmic domain. Interestingly, both gE and gI cytoplasmic tails are required for virulence; lack of either one or both results in an attenuated infection. These data suggest that gE and gI play differential roles in mediating directional neuroinvasion of the rat; however, the gE and gI cytoplasmic domains most likely function together to promote virulence.
Identification and Characterization of Novel Human Endogenous Retrovirus Families by Phylogenetic Screening of the Human Genome Mapping Project Database
Authors: Tristem Michael 
Published: July 2002

ABSTRACT

Human endogenous retroviruses (HERVs) were first identified almost 20 years ago, and since then numerous families have been described. It has, however, been difficult to obtain a good estimate of both the total number of independently derived families and their relationship to each other as well as to other members of the family
Retroviridae
. In this study, I used sequence data derived from over 150 novel HERVs, obtained from the Human Genome Mapping Project database, and a variety of recently identified nonhuman retroviruses to classify the HERVs into 22 independently acquired families. Of these, 17 families were loosely assigned to the class I HERVs, 3 to the class II HERVs and 2 to the class III HERVs. Many of these families have been identified previously, but six are described here for the first time and another four, for which only partial sequence information was previously available, were further characterized. Members of each of the 10 families are defective, and calculation of their integration dates suggested that most of them are likely to have been present within the human lineage since it diverged from the Old World monkeys more than 25 million years ago.
trans
-Acting Inhibition of Genomic RNA Dimerization by Rous Sarcoma Virus Matrix Mutants
Authors: Garbitt Rachel A. , Albert Jessica A. , Kessler Michelle D. , Parent Leslie J. 
Published: July 2002

ABSTRACT

The genomic RNA of retroviruses exists within the virion as a noncovalently linked dimer. Previously, we identified a mutant of the viral matrix (MA) protein of Rous sarcoma virus that disrupts viral RNA dimerization. This mutant, Myr1E, is modified at the N terminus of MA by the addition of 10 amino acids from the Src protein, resulting in the production of particles containing monomeric RNA. Dimerization is reestablished by a single amino acid substitution that abolishes myristylation (Myr1E−). To distinguish between
cis
and
trans
effects involving Myr1E, additional mutations were generated. In Myr1E.cc and Myr1E−.cc, different nucleotides were utilized to encode the same protein as Myr1E and Myr1E−, respectively. The alterations in RNA sequence did not change the properties of the viral mutants. Myr1E.ATG− was constructed so that translation began at the
gag
AUG, resulting in synthesis of the wild-type Gag protein but maintenance of the
src
RNA sequence. This mutant had normal infectivity and dimeric RNA, indicating that the
src
sequence did not prevent dimer formation. All of the
src
-containing RNA sequences formed dimers in vitro. Examination of MA-green fluorescent protein fusion proteins revealed that the wild-type and mutant MA proteins Myr1E.ATG−, Myr1E−, and Myr1E−.cc had distinctly different patterns of subcellular localization compared with Myr1E and Myr1E.cc MA proteins. This finding suggests that proper localization of the MA protein may be required for RNA dimer formation and infectivity. Taken together, these results provide compelling evidence that the genomic RNA dimerization defect is due to a
trans
-acting effect of the mutant MA proteins.
Elevated Levels of Interleukin-8 in Serum Are Associated with Hepatitis C Virus Infection and Resistance to Interferon Therapy
Authors: Polyak Stephen J. , Khabar Khalid S. A. , Rezeiq Mohammed , Gretch David R. 
Published: July 2002

ABSTRACT
Hepatitis C virus (HCV), a major cause of liver disease worldwide, is frequently resistant to the antiviral alpha interferon (IFN). We have recently found that the HCV NS5A protein induces expression of the proinflammatory chemokine IL-8 to partially inhibit the antiviral actions of IFN in vitro. To extend these observations, in the present study we examined the relationship between levels of IL-8 in serum, HCV infection, and biochemical response to IFN therapy. Levels of IL-8 were significantly elevated in 132 HCV-infected patients compared to levels in 32 normal healthy subjects and were also significantly higher in patients who did not respond to IFN therapy than in patients who did respond to therapy. This study suggests that HCV-induced changes in levels of chemokine and cytokine expression may be involved in HCV antiviral resistance, persistence, and pathogenesis.
Selective Regulation of Human Immunodeficiency Virus-Infected CD4+Lymphocytes by a Synthetic Immunomodulator Leads to Potent Virus Suppression In Vitro and in hu-PBL-SCID Mice
Authors: Bahr George M. , Darcissac Edith C. A. , Castéran Nathalie , Amiel Corinne , Cocude Cécile , Truong Marie-José , Dewulf Joëlle , Capron André , Mouton Yves 
Published: July 2002

ABSTRACTWe have previously observed that the synthetic immunomodulator Murabutide inhibits human immunodeficiency virus type 1 (HIV-1) replication at multiple levels in macrophages and dendritic cells. The present study was designed to profile the activity of Murabutide on CD8-depleted phytohemagglutinin-activated lymphocytes from HIV-1-infected subjects and on the outcome of HIV-1 infection in severe combined immunodeficiency mice reconstituted with human peripheral blood leukocytes (hu-PBL-SCID mice). Maintaining cultures of CD8-depleted blasts from 36 patients in the presence of Murabutide produced dramatically reduced levels of viral p24 protein in the supernatants. This activity correlated with reduced viral transcripts and proviral DNA, was evident in cultures harboring R5, X4-R5, or X4 HIV-1 isolates, was not linked to inhibition of cellular DNA synthesis, and did not correlate with β-chemokine release. Moreover, c-mycmRNA expression was down-regulated in Murabutide-treated cells, suggesting potential interference of the immunomodulator with the nuclear transport of viral preintegration complexes. On the other hand, daily treatment of HIV-1-infected hu-PBL-SCID mice with Murabutide significantly reduced the viral loads in plasma and the proviral DNA content in human peritoneal cells. These results are the first to demonstrate that a clinically acceptable synthetic immunomodulator with an ability to enhance the host's nonspecific immune defense mechanisms against infections can directly regulate cellular factors in infected lymphocytes, leading to controlled HIV-1 replication.
Genome of Lumpy Skin Disease Virus
Authors: Tulman E. R. , Afonso C. L. , Lu Z. , Zsak L. , Kutish G. F. , Rock D. L. 
Published: July 2002

ABSTRACT

Lumpy skin disease virus (LSDV), a member of the capripoxvirus genus of the
Poxviridae
, is the etiologic agent of an important disease of cattle in Africa. Here we report the genomic sequence of LSDV. The 151-kbp LSDV genome consists of a central coding region bounded by identical 2.4 kbp-inverted terminal repeats and contains 156 putative genes. Comparison of LSDV with chordopoxviruses of other genera reveals 146 conserved genes which encode proteins involved in transcription and mRNA biogenesis, nucleotide metabolism, DNA replication, protein processing, virion structure and assembly, and viral virulence and host range. In the central genomic region, LSDV genes share a high degree of colinearity and amino acid identity (average of 65%) with genes of other known mammalian poxviruses, particularly suipoxvirus, yatapoxvirus, and leporipoxviruses. In the terminal regions, colinearity is disrupted and poxvirus homologues are either absent or share a lower percentage of amino acid identity (average of 43%). Most of these differences involve genes and gene families with likely functions involving viral virulence and host range. Although LSDV resembles leporipoxviruses in gene content and organization, it also contains homologues of interleukin-10 (IL-10), IL-1 binding proteins, G protein-coupled CC chemokine receptor, and epidermal growth factor-like protein which are found in other poxvirus genera. These data show that although LSDV is closely related to other members of the
Chordopoxvirinae
, it contains a unique complement of genes responsible for viral host range and virulence.
CD8+Lymphocytes from Simian Immunodeficiency Virus-Infected Rhesus Macaques Recognize 14 Different Epitopes Bound by the Major Histocompatibility Complex Class I Molecule Mamu-A*01: Implications for Vaccine Design and Testing
Authors: Allen Todd M. , Mothé Bianca R. , Sidney John , Jing Peicheng , Dzuris John L. , Liebl Max E. , Vogel Thorsten U. , O'Connor David H. , Wang Xiaochi , Wussow Michael C. , Thomson James A. , Altman John D. , Watkins David I. , Sette Alessandro 
Published: July 2002

ABSTRACTIt is becoming increasingly clear that any human immunodeficiency virus (HIV) vaccine should induce a strong CD8+response. Additional desirable elements are multispecificity and a focus on conserved epitopes. The use of multiple conserved epitopes arranged in an artificial gene (or EpiGene) is a potential means to achieve these goals. To test this concept in a relevant disease model we sought to identify multiple simian immunodeficiency virus (SIV)-derived CD8+epitopes bound by a single nonhuman primate major histocompatibility complex (MHC) class I molecule. We had previously identified the peptide binding motif of Mamu-A*012, a common rhesus macaque MHC class I molecule that presents the immunodominant SIVgag-derived cytotoxic T lymphocyte (CTL) epitope Gag_CM9 (CTPYDINQM). Herein, we scanned SIV proteins for the presence of Mamu-A*01 motifs. The binding capacity of 221 motif-positive peptides was determined using purified Mamu-A*01 molecules. Thirty-seven peptides bound with apparentKdvalues of 500 nM or lower, with 21 peptides binding better than the Gag_CM9 peptide. Peripheral blood mononuclear cells from SIV-infected Mamu-A*01+macaques recognized 14 of these peptides in ELISPOT, CTL, or tetramer analyses. This study reveals an unprecedented complexity and diversity of anti-SIV CTL responses. Furthermore, it represents an important step toward the design of a multiepitope vaccine for SIV and HIV.
Cyclin T1 Expression Is Mediated by a Complex and Constitutively Active Promoter and Does Not Limit Human Immunodeficiency Virus Type 1 Tat Function in Unstimulated Primary Lymphocytes
Authors: Martin-Serrano Juan , Li Kelvin , Bieniasz Paul D. 
Published: July 2002

ABSTRACTCyclin T1 (CycT1), a component of positive-transcription-elongation factor-b (P-TEFb), is an essential cofactor for transcriptional activation by lentivirus Tat proteins. It is thought that low CycT1 expression levels restrict human immunodeficiency virus type 1 (HIV-1) expression levels and replication in resting CD4+lymphocytes. In this study, we undertook a functional analysis of thecycT1promoter to determine which, if any, promoter elements might be responsible for cellular activation state-dependent CycT1 expression. ThecycT1gene contains a complex promoter that exhibits an extreme degree of functional redundancy: five nonoverlapping fragments were found to exhibit significant promoter activity in immortalized cell lines, and these elements could interact in a synergistic or redundant manner to mediatecycT1transcription. Reporter gene expression, mediated by thecycT1promoter, was detectable in unstimulated transfected primary lymphocytes and multiple sites within the promoter could serve to initiate transcription. While utilization of these start sites was significantly altered by the application of exogenous stimuli to primary lymphocytes and two distinct promoter elements exhibited enhanced activity in the presence of phorbol ester, overallcycT1transcription was only modestly enhanced in response to cell activation. These observations prompted a reexamination of CycT1 protein expression in primary lymphocytes. In fact, steady-state CycT1 expression is only slightly lower in unstimulated lymphocytes compared to phorbol ester-treated cells or a panel of immortalized cell lines. Importantly, CycT1 is expressed at sufficient levels in unstimulated primary cells to support robust Tat activity. These results strongly suggest that CycT1 expression levels in unstimulated primary lymphocytes do not profoundly limit HIV-1 gene expression or provide an adequate mechanistic explanation for proviral latency in vivo.
Induction of CD8 T-Cell-Specific Systemic and Mucosal Immunity against Herpes Simplex Virus with CpG-Peptide Complexes
Authors: Gierynska Malgorzata , Kumaraguru Uday , Eo Seong-Kug , Lee Sujin , Krieg Arthur , Rouse Barry T. 
Published: July 2002

ABSTRACTOligodeoxynucleotides (ODN) containing unmethylated CpG motifs exert powerful adjuvant activity in vivo and in vitro. Administered with antigen they induce a population of antigen-specific CD8+T cells. In this study we immunized C57BL/6 mice with bioactive CpG ODN combined with an immunodominant epitope derived from herpes simplex virus (HSV) glycoprotein B (amino acids 498 to 505; SSIEFARL) and analyzed the magnitude and durability of the peptide-specific response. The effectiveness of the CD8+T-cell response as measured by peptide-specific tetramers, peptide-induced intracellular gamma interferon expression, and resistance to systemic and mucosal challenge during the acute and memory phases was compared with the response induced by immunization with recombinant vaccinia virus encoding SSIEFARL as a minigene (VvgB498-505). Confirming the reports of others, our results demonstrate that the CpG ODN-peptide approach generates an antigen-specific CD8+T-cell population, but the frequency of CD8+T cells is lower than that induced by VvgB498-505. Nevertheless, the protection level was comparable when mice were systemically and mucosally challenged during the acute phase. However, such responses by both groups waned with time and were functionally less effective. Still, our results indicate that the CpG ODN-peptide immunization system holds promise as a means of selectively inducing a CD8+T-cell response against HSV.
Crystal Structure of Bis(ethylenediphosphinetetraacetic acid)palladium(II) Bis(dihydrogenethylenediphosphinetetraacetato)palladate(II) Tetrahydrate Hydrobromide (1/1): An Unusual Case of Bromide Uncoordinated to Palladium(II)
Authors: Podlaha Jaroslav , Podlahová Jana 
Published: July 2002

Synthesis, crystal and molecular structure of the title compound as a representative of the complexes of phosphineacetic acids with soft metal ions is described. Crystal data: triclinic, P1, a = 10.809(1), b = 14.922(1), c = 10.617(1) Å, α = 108.871(7), β = 102.773(9), γ = 102.970(7)°, Z = 1, R = 0.031 for 4 545 observed reflections. The structure is built of layers composed of hydrogen-bonded ligands; the layers are joined, in alternate manner, by Pd(II) ions, water molecules and bromide anions. As an average, one proton of the tetraprotonated ligand is released during complexation even in protic solvent (aqueous acetic acid), resulting in the given ionic composition with non-equivalent Pd(II) centers. The carboxyl groups and, surprisingly, even the bromide ion are not coordinated to Pd. The proton assigned formally to hydrobromic acid was not located and is possibly disordered.