bluuebunny/crossref_metadata_2025_split huggingface dataset:
Serodiagnosis of Human Granulocytic Ehrlichiosis by Using Novel Combinations of Immunoreactive Recombinant Proteins
Authors: Lodes Michael J. , Mohamath Raodoh , Reynolds Lisa D. , McNeill Patricia , Kolbert Christopher P. , Bruinsma Elizabeth S. , Benson Darin R. , Hofmeister Eric , Reed Steven G. , Houghton Raymond L. , Persing David H. 
Published: July 2002

ABSTRACT

A panel of seven recombinant antigens, derived from
Ehrlichia phagocytophila
(the agent of human granulocytic ehrlichiosis), was evaluated by class-specific enzyme-linked immunosorbent assays (ELISAs) for utility in the diagnosis of the infection. Fourteen genomic fragments, obtained by serologic expression screening, contained open reading frames (ORFs) encoding 16 immunodominant antigens. Eleven of these antigens were members of the major surface protein (MSP) multigene family. Alignment of their predicted protein sequences revealed a pattern of conserved sequences, which contained short direct repeats, flanking a variable region. In addition, two genomic clones contained two and three MSP ORFs, respectively, indicating that these genes are clustered in tandem copies. The implications for this pattern of both genomic and protein arrangements in antigenic variations of MSPs and in their utilities in a diagnostic assay are discussed. In addition to two MSP recombinant antigens (rHGE-1 and -3) and a fusion protein of these antigens (rErf-1), five further recombinants were evaluated by ELISA. Two of these antigens (rHGE-14 and -15) were novel, while a third (rHGE-2), with no known function, has been described. The final two recombinant antigens (rHGE-9 and -17) represent overlapping segments of the ankyrin gene (
ank
). The addition of rHGE-9 ELISA data resulted in the detection of 78% (21 of 27) of acute-phase sera. When serologic data for all recombinants are combined, 96.2% (26 of 27) of convalescent-phase patient serum samples and 85.2% (23 of 27) of acute-phase patient serum samples are detected, indicating the potential of these antigens for use in the development of a rapid serologic assay for the detection of
E. phagocytophila
infection.
Efficacy of New Low-Cost Filtration Device for Recovering
Schistosoma haematobium
Eggs from Urine
Authors: Gyorkos Theresa W. , Ramsan Mahdi , Foum Ali , Khamis Iddi Simba 
Published: July 2002

ABSTRACT

A new, inexpensive filtration device for the diagnosis of urinary schistosomiasis was tested against the commonly used Millipore device. The experimental protocol was performed with 25 urine samples known to be positive for
Schistosoma haematobium
. The results suggest that the new device is as effective as the Millipore device for the diagnosis of urinary schistosomiasis. Its low cost will be attractive to schistosomiasis control programs.
Human Infections Due to
Mycobacterium lentiflavum
Authors: Tortoli Enrico , Bartoloni Alessandro , Erba Maria Luigia , Levrè Egle , Lombardi Natalia , Mantella Antonia , Mecocci Lorenzo 
Published: July 2002

ABSTRACT

Three cases of human disease due to
Mycobacterium lentiflavum
are reported. In the first, the mycobacterium was responsible for chronic pulmonary disease in an elderly woman; in the second, it gave rise to cervical lymphadenitis in a child; and in the third, it caused a liver abscess in a young AIDS patient.
Human Cytomegalovirus and Human Umbilical Vein Endothelial Cells: Restriction of Primary Isolation to Blood Samples and Susceptibilities of Clinical Isolates from Other Sources to Adaptation
Authors: Gerna Giuseppe , Percivalle Elena , Sarasini Antonella , Revello M. Grazia 
Published: July 2002

ABSTRACT

In immunocompromised patients with disseminated infection, human cytomegalovirus (HCMV) is widespread in the microvascular endothelium of multiple organs. Human umbilical vein endothelial cells (HUVEC) were used in parallel to human embryonic lung fibroblasts (HELF) to recover HCMV from blood samples of immunocompromised patients. Using the shell vial technique, comparable median numbers of p72-positive HUVEC and HELF cells were found with the 26 HCMV-positive buffy coat samples out of 150 examined. Analysis of other clinical samples inoculated as controls revealed, in the presence of highly infected HELF monolayers, either the presence of very few infected HUVEC with urine specimens (
n
= 10 samples) or the lack of infected HUVEC with throat washes (
n
= 3) or amniotic fluid samples (
n
= 2). Thus, peripheral blood leukocytes (PBL) appear essential for primary isolation of HCMV in HUVEC. In this respect, HCMV strains, recovered from clinical samples other than buffy coats in HELF only, could be readily adapted to growth in HUVEC by coculturing PBL from healthy blood donors with infected HELF and then inoculating infected PBL onto HUVEC. Recently elucidated mechanisms of interaction of leukocytes and HUVEC with bidirectional transfer of virus seem to provide the basis for the restriction of HCMV primary isolation in HUVEC to blood samples. However, virus strains recovered from only HELF could be adapted to growth in HUVEC when inoculated with HELF-derived (either cell-associated or cell-free) HCMV strains upon primary isolation. In conclusion, due to the in vitro selection of virus variants provided with both PBL tropism and HUVEC tropism, HCMV recovery in HUVEC is PBL mediated and substantially restricted to blood samples. Lack of HCMV recovery in HUVEC from clinical samples other than blood leads to the assumption that epithelial cells, such as urinary, amniotic, or pharyngeal cells, do not possess adequate adhesion molecules to establish close contacts with HUVEC.
Typing and Subtyping of 83 Clinical Isolates Purified from Surgically Implanted Silicone Feeding Tubes by Random Amplified Polymorphic DNA Amplification
Authors: Dautle Melanie P. , Ulrich Ricky L. , Hughes Thomas A. 
Published: July 2002

ABSTRACT

In this study, 83 clinical isolates purified from biofilms colonizing 18 silicone gastrostomy devices (12 “buttons” and six tubes converted to skin level devices) were selected for subtype characterization utilizing genetic analysis. The tubes, previously used for feeding, remained in place for 3 to 47 months (mean, 20.0 months) in children ranging in age from 6 months to 17 years. Classification of specific microbes using random amplified polymorphic DNA (RAPD) analysis revealed genetic similarities and differences among isolates belonging to the same genus. Both gram-positive and -negative bacteria were investigated, including 2 isolates of
Bacillus brevis,
4 isolates of
Bacillus licheniformis
, 2 isolates of
Bacillus pumilus
, 3 isolates of
Enterococcus durans
, 19 isolates of
Enterococcus faecalis
, 8 isolates of
Enterococcus faecium
, 2 isolates of
Enterococcus hirae
, 7 isolates of
Escherichia coli
, 8 isolates of
Lactobacillus plantarum
, 19 isolates of
Staphylococcus aureus
, 2 isolates of
Staphylococcus epidermidis
, and 7 isolates of
Staphylococcus saprophyticus.
Amplified DNA fragments (amplicons) provided species-specific fingerprints for comparison by agarose gel electrophoresis. A total of 62 distinct RAPD types were categorized from the five genera studied. Typing analysis suggested cross acquisition of
E. coli
,
E. faecalis
, and
S. aureus
in three patient pairs. Genomic polymorphism detection proved efficient and reliable for classifying bacterial subtypes isolated from biofilms adhering to various portions of commonly employed enteral access tubes.
Mitochondrial DNA Restriction Fragment Length Polymorphism (RFLP) and 18S Small-Subunit Ribosomal DNA PCR-RFLP Analyses ofAcanthamoebaIsolated from Contact Lens Storage Cases of Residents in Southwestern Korea
Authors: Kong Hyun-Hee , Shin Ji-Yeol , Yu Hak-Sun , Kim Jin , Hahn Tae-Won , Hahn Young-Ho , Chung Dong-Il 
Published: July 2002

ABSTRACTWe applied ribosomal DNA PCR-restriction fragment length polymorphism (RFLP) and mitochondrial DNA (mtDNA) RFLP analyses to 43Acanthamoebaenvironmental isolates (KA/LH1 to KA/LH43) from contact lens storage cases in southwestern Korea. These isolates were compared to American Type Culture Collection strains and clinical isolates (KA/E1 to KA/E12) from patients with keratitis. Seven riboprint patterns were seen. To identify the species of the isolates, a phylogenetic tree was constructed based on the comparison of riboprint patterns with reference strains. Four types accounted for 39 of the isolates belonging to theA. castellaniicomplex. The most predominant (48.8%) type wasA. castellaniiKA/LH2 type, which had identical riboprint and mtDNA RFLP patterns to those ofA. castellaniiCastellani, KA/E3 and KA/E8. The riboprint and mtDNA RFLP patterns of the KA/LH7 (20.9%) type were identical to those ofA. castellaniiMa, a corneal isolate from the United States. The riboprint and mtDNA RFLP patterns of the KA/LH1 (18.6%) type were the same as those ofA. lugdunensisL3a, KA/E2, and KA/E12. The prevalent pattern for each type ofAcanthamoebain southwestern Korea was very different from that from southeastern Korea and Seoul, Korea. It is noteworthy that 38 (88.4%) out of 43 isolates from contact lens storage cases of the residents in southwestern Korea revealed mtDNA RFLP and riboprint patterns identical to those found for clinical isolates in our area. This indicates that most isolates from contact lens storage cases in the surveyed area are potential keratopathogens. More attention should be paid to the disinfection of contact lens storage cases to prevent possible amoebic keratitis.
Comparison of an Automated Ribotyping System to Restriction Endonuclease Analysis and Pulsed-Field Gel Electrophoresis for Differentiating Vancomycin-Resistant
Enterococcus faecium
Isolates
Authors: Price Connie S. , Huynh Holly , Paule Suzanne , Hollis Richard J. , Noskin Gary A. , Pfaller Michael A. , Peterson Lance R. 
Published: July 2002

ABSTRACT

The RiboPrinter Microbial Characterization System was compared with pulsed-field gel electrophoresis (PFGE), restriction endonuclease analysis (REA), and epidemiological data for typing 45 vancomycin-resistant
Enterococcus faecium
(VRE) isolates. In 21 clinically related isolates, 90 to 100% were similar by PFGE and REA, but only 57% were similar by the RiboPrinter. In another eight clinically related isolates, three isolates similar by PFGE and REA were all unique by the RiboPrinter. In contrast, in 16 clinically unrelated isolates, the predominant RiboPrinter ribotype represented 50% of the strains, while the largest PFGE and REA clones represented less than 19% of the strains. These data suggest that the RiboPrinter is not reliable for VRE investigation.
Efficient DNA Transfection Mediated by the C-Terminal Domain of Human Immunodeficiency Virus Type 1 Viral Protein R
Authors: Kichler Antoine , Pages Jean-Christophe , Leborgne Christian , Druillennec Sabine , Lenoir Christine , Coulaud Dominique , Delain Etienne , Le Cam Eric , Roques Bernard P. , Danos Olivier 
Published: July 2002

ABSTRACT

Viral protein R (Vpr) of human immunodeficiency virus type 1 is produced late in the virus life cycle and is assembled into the virion through binding to the Gag protein. It is known to play a significant role early in the viral life cycle by facilitating the nuclear import of the preintegration complex in nondividing cells. Vpr is also able to interact with nucleic acids, and we show here that it induces condensation of plasmid DNA. We have explored the possibility of using these properties in DNA transfection experiments. We report that the C-terminal half of the protein (Vpr
52–96
) mediates DNA transfection in a variety of human and nonhuman cell lines with efficiencies comparable to those of the best-known transfection agents. Compared with polylysine, a standard polycationic transfection reagent, Vpr
52–96
was 10- to 1,000-fold more active. Vpr
52–96
-DNA complexes were able to reach the cell nucleus through a pH-independent mechanism. These observations possibly identify an alternate pathway for DNA transfection.
Cellular and Viral Specificities of Human Immunodeficiency Virus Type 1 Vif Protein
Authors: Madani Navid , Kabat David 
Published: July 2002

ABSTRACTThevifgene of human immunodeficiency virus type 1 (HIV-1) greatly enhances the infectivity of HIV-1 virions that are released from cells classified as nonpermissive (e.g., lymphocytes, macrophages, and H9 leukemic T cells) but is irrelevant in permissive cells (e.g., HeLa or COS cells). Recently, it was reported thatvifexpression in nonpermissive cells dramatically increases infectivity not only of HIV-1 but also of other enveloped viruses, including murine leukemia viruses (MLVs). This was surprising in part because MLVs and other murine retroviruses lackvifgenes yet replicate efficiently in T lymphocytes. To investigate these issues, we first developed improved methods for producing substantial quantities of HIV-1 virions withvifdeletions from healthy H9 cells. These virions had approximately the same amounts of major core proteins and envelope glycoproteins as the control wild-type virions but were only approximately 1% as infectious. We then produced H9 cells that contained wild-type orvifdeletion HIV-gptproviruses, which lack a functionalenvgene. After superinfection with either xenotropic or amphotropic MLVs, these cells released HIV-gptvirions pseudotyped with an MLV envelope plus replication-competent MLV. Interestingly, the pseudotyped HIV-gpt(vifdeletion) virions were noninfectious, whereas the MLV virions simultaneously released from the same H9 cells were fully infectious. These results strongly suggest that the Vif protein functions in a manner that is both cell specific and at least substantially specific for HIV-1 and related lentiviruses. In addition, these results confirm thatvifdeletion HIV-1 virions from nonpermissive cells are blocked at a postpenetration stage of the infection pathway.
Pseudotyping of Glycoprotein D-Deficient Herpes Simplex Virus Type 1 with Vesicular Stomatitis Virus Glycoprotein G Enables Mutant Virus Attachment and Entry
Authors: Anderson Dina B. , Laquerre Sylvie , Goins William F. , Cohen Justus B. , Glorioso Joseph C. 
Published: July 2002

ABSTRACT
The use of herpes simplex virus (HSV) vectors for in vivo gene therapy will require the targeting of vector infection to specific cell types in certain in vivo applications. Because HSV glycoprotein D (gD) imparts a broad host range for viral infection through recognition of ubiquitous host cell receptors, vector targeting will require the manipulation of gD to provide new cell recognition specificities in a manner designed to preserve gD's essential role in virus entry. In this study, we have determined whether an entry-incompetent HSV mutant with deletions of all Us glycoproteins, including gD, can be complemented by a foreign attachment/entry protein with a different receptor-binding specificity, the vesicular stomatitis virus glycoprotein G (VSV-G). The results showed that transiently expressed VSV-G was incorporated into gD-deficient HSV envelopes and that the resulting pseudotyped virus formed plaques on gD-expressing VD60 cells, albeit at a 50-fold-reduced level compared to that of wild-type gD. This reduction may be related to differences in the entry pathways used by VSV and HSV or to the observed lower rate of incorporation of VSV-G into virus envelopes than that of gD. The rate of VSV-G incorporation was greatly improved by using recombinant molecules in which the transmembrane domain of HSV glycoprotein B or D was substituted for that of VSV-G, but these recombinant molecules failed to promote virus entry. These results show that foreign glycoproteins can be incorporated into the HSV envelope during replication and that gD can be dispensed with on the condition that a suitable attachment/entry function is provided.
The Interaction of Bacteriophage P2 B Protein with
Escherichia coli
DnaB Helicase
Authors: Odegrip Richard , Schoen Stephan , Haggård-Ljungquist Elisabeth , Park Kyusung , Chattoraj Dhruba K. 
Published: July 2002

ABSTRACT

Bacteriophage P2 requires several host proteins for lytic replication, including helicase DnaB but not the helicase loader, DnaC. Some genetic studies have suggested that the loading is done by a phage-encoded protein, P2 B. However, a P2 minichromosome containing only the P2 initiator gene
A
and a marker gene can be established as a plasmid without requiring the P2
B
gene. Here we demonstrate that P2 B associates with DnaB. This was done by using the yeast two-hybrid system in vivo and was confirmed in vitro, where
35
S-labeled P2 B bound specifically to DnaB adsorbed to Q Sepharose beads and monoclonal antibodies directed against the His-tagged P2 B protein were shown to coprecipitate the DnaB protein. Finally, P2 B was shown to stabilize the opening of a reporter origin, a reaction that is facilitated by the inactivation of DnaB. In this respect, P2 B was comparable to λ P protein, which is known to be capable of binding and inactivating the helicase while acting as a helicase loader. Even though P2 B has little similarity to other known or predicted helicase loaders, we suggest that P2 B is required for efficient loading of DnaB and that this role, although dispensable for P2 plasmid replication, becomes essential for P2 lytic replication.
N-Glycans of F Protein Differentially Affect Fusion Activity of Human Respiratory Syncytial Virus
Authors: Zimmer Gert , Trotz Ina , Herrler Georg 
Published: July 2002

ABSTRACT

The human respiratory syncytial virus (Long strain) fusion protein contains six potential N-glycosylation sites: N27, N70, N116, N120, N126, and N500. Site-directed mutagenesis of these positions revealed that the mature fusion protein contains three N-linked oligosaccharides, attached to N27, N70, and N500. By introducing these mutations into the F gene in different combinations, four more mutants were generated. All mutants, including a triple mutant devoid of any N-linked oligosaccharide, were efficiently transported to the plasma membrane, as determined by flow cytometry and cell surface biotinylation. None of the glycosylation mutations interfered with proteolytic activation of the fusion protein. Despite similar levels of cell surface expression, the glycosylation mutants affected fusion activity in different ways. While the N27Q mutation did not have an effect on syncytium formation, loss of the N70-glycan caused a fusion activity increase of 40%. Elimination of both N-glycans (N27/70Q mutant) reduced the fusion activity by about 50%. A more pronounced reduction of the fusion activity of about 90% was observed with the mutants N500Q, N27/500Q, and N70/500Q. Almost no fusion activity was detected with the triple mutant N27/70/500Q. These data indicate that N-glycosylation of the F
2
subunit at N27 and N70 is of minor importance for the fusion activity of the F protein. The single N-glycan of the F
1
subunit attached to N500, however, is required for efficient syncytium formation.
Adaptation of Chimeric Retroviruses In Vitro and In Vivo: Isolation of Avian Retroviral Vectors with Extended Host Range
Authors: Barsov Eugene V. , Payne William S. , Hughes Stephen H. 
Published: July 2002

ABSTRACT

We have designed and characterized two new replication-competent avian sarcoma/leukosis virus-based retroviral vectors with amphotropic and ecotropic host ranges. The amphotropic vector RCASBP-M2C(797-8), was obtained by passaging the chimeric retroviral vector RCASBP-M2C(4070A) (6) in chicken embryos. The ecotropic vector, RCASBP(Eco), was created by replacing the
env
-coding region in the retroviral vector RCASBP(A) with the
env
region from an ecotropic murine leukemia virus. It replicates efficiently in avian DFJ8 cells that express murine ecotropic receptor. For both vectors, permanent cell lines that produce viral stocks with titers of about 5 × 10
6
CFU/ml on mammalian cells can be easily established by passaging transfected avian cells. Some chimeric viruses, for example, RCASBP(Eco), replicate efficiently without modifications. For those chimeric viruses that do require modification, adaptation by passage in vitro or in vivo is a general strategy. This strategy has been used to prepare vectors with altered host range and could potentially be used to develop vectors that would be useful for targeted gene delivery.
Hairpin Loop Structure in the 3′ Arm of the Influenza A Virus Virion RNA Promoter Is Required for Endonuclease Activity
Authors: Leahy Michael B. , Dobbyn Helen C. , Brownlee George G. 
Published: July 2002

ABSTRACT
Previous studies have shown that the 5′ arm of the influenza A virus virion RNA promoter requires a hairpin loop structure for efficient endonuclease activity of influenza virus RNA polymerase, an activity that is required for the cap-snatching activity of primers from host pre-mRNA. Here we examine whether a hairpin loop is also required in the 3′ arm of the viral RNA promoter. We study point mutations at each nucleotide position (1 to 12) within the 3′ arm of the promoter as well as complementary “rescue” mutations which restored base pairing in the stem of a potential hairpin loop. Our results suggest that endonuclease activity is absolutely dependent on the presence of a 3′ hairpin loop structure. This is the first direct evidence for RNA secondary structure within the 3′ arm being required for a specific stage, i.e., endonuclease cleavage, in the influenza virus replicative cycle.
Conserved CDR3 Regions in T-Cell Receptor (TCR) CD8+T Cells That Recognize the Tax11-19/HLA-A*0201 Complex in a Subject Infected with Human T-Cell Leukemia Virus Type 1: Relationship of T-Cell Fine Specificity and Major Histocompatibility Complex/Peptide/TCR Crystal Structure
Authors: Bourcier Katarzyna D. , Lim Dong-Gyun , Ding Yuan-Hua , Smith Kathrine J. , Wucherpfennig Kai , Hafler David A. 
Published: July 2002

ABSTRACTWe investigated the T-cell receptor (TCR) repertoire of CD8+T cells that recognize the Tax11-19 immunodominant epitope of Tax protein expressed by human T-cell leukemia virus (HTLV-1) that is implicated in the disease HTLV-1-associated myelopathy (HAM/TSP). A panel of Tax11-19-reactive CD8+T-cell clones was generated by single-cell cloning of Tax11-19/HLA-A*0201 tetramer-positive peripheral blood lymphocytes from an HTLV-1-infected individual. The analyses of TCR usage revealed that the combination of diverse TCR alpha and beta chains could be used for the recognition of Tax11-19 but the major population of T-cell clones (15 of 24 clones) expressed the TCR V beta 13S1 and V alpha 17 chain. We found striking similarities in CDR3 regions of TCR alpha and beta chains between our major group of CD8+T-cell clones and those originating from different subjects as previously reported, including TCRs with resolved crystal structures. A 3-amino-acid sequence (PG-G) in the CDR3 region of the V beta chain was conserved among all the Tax11-19-reactive T-cell clones expressing V beta 13S1 and V alpha 17 chains. Conserved amino acids in the CDR3 region do not directly contact the Tax11-19 peptide, as corroborated by the crystal structure of B7-TCR, a TCR that is almost identical to VB13S1 clones isolated in this study. Analysis of fine peptide specificity using altered peptide ligands (APL) of Tax11-19 revealed a similar recognition pattern among this panel of T-cell clones. These data suggest that the PG-G amino acids in the CDR3 beta loop provide a structural framework necessary for the maintenance of the tertiary TCR structure.