bluuebunny/crossref_metadata_2025_split huggingface dataset:
Comparative Evaluation of Three Human Immunodeficiency Virus Genotyping Systems: the HIV-GenotypR Method, the HIV PRT GeneChip Assay, and the HIV-1 RT Line Probe Assay
Authors: Wilson John W. , Bean Pamela , Robins Terry , Graziano Frank , Persing David H. 
Published: December 2019

ABSTRACT

Evaluation of drug resistance by human immunodeficiency virus (HIV) genotyping has proven to be useful for the selection of drug combinations with maximum antiretroviral activity. We compared three genotyping methods for identification of mutations known to confer drug resistance in the reverse transcriptase (RT) and protease genes of HIV type 1 (HIV-1). The HIV-GenotypR method (GenotypR; Specialty Laboratories, Inc., Santa Monica, Calif.) with the ABI 377 DNA sequencer (Applied Biosystems Inc.), the HIV PRT GeneChip assay (GeneChip; Affymetrix, Santa Clara, Calif.), and the HIV-1 RT Line Probe Assay (LiPA; Innogenetics, Alpharetta, Ga.) were used to genotype plasma samples from HIV-infected patients attending the University of Wisconsin Hospitals and Clinics and the Mayo Clinic. At the time of analysis, patients were failing combination therapy (
n
= 18) or were treatment naive (
n
= 6). Forty codons of the RT and protease genes were analyzed by GenotypR and GeneChip for resistance-associated mutations. LiPA analyzed seven RT codons for mutations. Each sample was genotyped by all three assays, and each assay was subjected to pairwise comparisons. At least 92% of the codons tested (by the three assays) in paired comparisons were concordant. GenotypR and GeneChip demonstrated 96.6% concordance over the 40 codons tested. GenotypR identified slightly more mutations than GeneChip and LiPA; GeneChip identified all primary mutations that corresponded to failing treatment regimens. Each assay identified at least 84% of the mutations identified by the other assays. Mutations that were discordant between the assays mainly comprised secondary mutations and natural polymorphisms. The assays had better concordance for mutations that corresponded to current failing regimens, present in the more predominant viral quasispecies. In the treatment-naive patients, GenotypR, GeneChip, and LiPA mainly identified wild-type virus. Only the LiPA identified K70R, a possible transmitted zidovudine resistance mutation, in the RT gene of a treatment-naive patient. We conclude that although discrepancies in results exist between assays, each assay showed a similar capacity to identify potentially clinically relevant mutations related to patient treatment regimens.
Duplex PCR for Differential Identification ofMycobacterium bovis,M. avium, andM. aviumsubsp.paratuberculosisin Formalin- Fixed Paraffin-Embedded Tissues from Cattle
Authors: Coetsier Christophe , Vannuffel Pascal , Blondeel Nathalie , Denef Jean-Francois , Cocito Carlo , Gala Jean-Luc 
Published: December 2019

ABSTRACTWe previously isolated and sequenced two genomic segments ofMycobacterium aviumsubsp.paratuberculosis, namely, f57, a species-specific sequence, and the p34 gene, coding for a 34-kDa antigenic protein. Comparison of sequences upstream of the p34 open reading frame (us-p34) fromM. aviumsubsp.paratuberculosisandM. tuberculosisshowed a 79-base deletion inM. tuberculosis. Sequence analysis of the p34 genes in another two species,M. bovis(strain BCG) andM. avium(strain D4), confirmed the differences observed between tuberculous and nontuberculous species. A duplex diagnostic PCR strategy based on coamplification of nonhomologous us-p34 and species-specific f57 sequences was therefore developed. Duplex PCR yielded three different patterns, specific either for tuberculous bacilli (M. tuberculosis,M. bovis, andM. africanum), for both nontuberculous mycobacteriaM. aviumandM. intracellulare, or forM. aviumsubsp.paratuberculosis. The specificity of this single-step DNA-based assay was assessed on DNA from cultured mycobacterial strains, as well as on a panel of formalin-fixed and paraffin-embedded tissues from cattle. Molecular assay results from tissular DNA were compared to conventional bacteriological and histological test results, including those obtained by Ziehl-Neelsen staining on tissue biopsy specimens. Molecular discrimination was successful and confirmed the value of duplex us-p34 and f57 sequence amplification for differential diagnosis of tuberculosis, paratuberculosis, or infections caused by other members of theM. aviumcomplex.
High-Resolution Genotyping of
Chlamydia trachomatis
from Recurrent Urogenital Infections
Authors: Pedersen Lisbeth Nørum , Kjær Holger Olaf , Møller Jens Kjølseth , Ørntoft Torben Falck , Ostergaard Lars 
Published: December 2019

ABSTRACT

A method for
Chlamydia trachomatis
restriction fragment length polymorphism (RFLP) analysis and complete sequencing of
omp1
was developed for use on samples collected at home, and results were compared. Genotyping by sequencing was superior to RFLP analysis. The
omp1
gene in 31 clinical strains harbored few mutations compared to the same gene in ATCC reference strains. Follow-up samples obtained during a 24-week period from 31 patients showed recurrence with the same genotype in five cases and a new genotype in one case.
Validation of the Abbreviated
Brucella
AMOS PCR as a Rapid Screening Method for Differentiation of
Brucella abortus
Field Strain Isolates and the Vaccine Strains, 19 and RB51
Authors: Ewalt Darla R. , Bricker Betsy J. 
Published: December 2019

ABSTRACT

The
Brucella
AMOS PCR assay was previously developed to identify and differentiate specific
Brucella
species. In this study, an abbreviated
Brucella
AMOS PCR test was evaluated to determine its accuracy in differentiating
Brucella abortus
into three categories: field strains, vaccine strain 19 (S19), and vaccine strain RB51/parent strain 2308 (S2308). Two hundred thirty-one isolates were identified and tested by the conventional biochemical tests and
Brucella
AMOS PCR. This included 120 isolates identified as
B. abortus
S19, 9 identified as
B. abortus
strain RB51, 57 identified as
B. abortus
biovar 1, 15 identified as
B. abortus
bv. 2, 1 identified as
B. abortus
bv. 2 (M antigen dominant), 7 identified as
B. abortus
bv. 4, and 22 identified as
B. abortus
S2308 and isolated from experimentally infected cattle. The
Brucella
AMOS PCR correctly identified each isolate as RB51/S2308, S19, or a field strain of
Brucella
.
Lactoferrin and Eosinophilic Cationic Protein in Nasal Secretions of Patients with Experimental Rhinovirus Colds, Natural Colds, and Presumed Acute Community-Acquired Bacterial Sinusitis
Authors: Niehaus Mark D. , Gwaltney Jack M. , Hendley J. Owen , Newman Mercy J. , Heymann Peter W. , Rakes Gary P. , Platts-Mills Thomas A. E. , Guerrant Richard L. 
Published: December 2019

ABSTRACT
To distinguish sinusitis from uncomplicated “colds,” we examined lactoferrin and eosinophilic cationic protein (ECP) in nasal secretions. Lactoferrin titers were ≥1:400 in 4% of persons with uncomplicated colds and controls but in 79% of persons with sinusitis or purulent sputa. ECP levels were >200 ng/ml in 61% of persons with colds and >3,000 ng/ml in 62% of persons with sinusitis. Nasal lactoferrin helps distinguish sinusitis from colds.
Recovery and Identification of West Nile Virus from a Hawk in Winter
Authors: Garmendia Antonio E. , Van Kruiningen Herbert J. , French Richard A. , Anderson John F. , Andreadis Theodore G. , Kumar Asok , West A. Brian 
Published: December 2019

ABSTRACT
West Nile virus was recovered from the brain of a red-tailed hawk that died in Westchester County, N.Y., in February 2000. Multiple foci of glial cells, lymphocytes, and a few pyknotic nuclei were observed in the brain. Three to 4 days after inoculation of Vero cells with brain homogenates, cytopathic changes were detected. The presence of West Nile virus antigen in fixed cells or cell lysates was revealed by fluorescent antibody testing or enzyme-linked immunosorbent assay, respectively. Furthermore, Reverse transcriptase-PCR with primers specific for the NS3 gene of West Nile virus resulted in an amplicon of the expected size (470 bp). Electron microscopy of thin sections of infected Vero cells revealed the presence of viral particles approximately 40 nm in diameter, within cytoplasmic vesicles. The demonstration of infection with the West Nile virus in the dead of the winter, long after mosquitoes ceased to be active, is significant in that it testifies to the survival of the virus in the region beyond mosquito season and suggests another route of transmission: in this case, prey to predator.
Utility of Commercial Systems for Identification of
Burkholderia cepacia
Complex from Cystic Fibrosis Sputum Culture
Authors: Shelly Deborah Blecker , Spilker Theodore , Gracely Edward J. , Coenye Tom , Vandamme Peter , LiPuma John J. 
Published: December 2019

ABSTRACT

Performances of several commercial test systems were reviewed to determine their relative levels of accuracy in identifying
Burkholderia cepacia
complex isolates recovered from cystic fibrosis sputum culture. Positive predictive values ranged from 71 to 98%; negative predictive values ranged from 50 to 82%. All systems misidentified
B. cepacia
complex. The species most frequently misidentified as
B. cepacia
was
Burkholderia gladioli
. These data support the results of previous studies that recommend confirmatory testing, including the use of DNA-based methods, for sputum culture isolates presumptively identified as
B. cepacia
.
Two Successive Hepatitis C Virus Infections in an Intravenous Drug User
Authors: Proust Barbara , Dubois Frederic , Bacq Yannick , Le Pogam Sophie , Rogez Sylvie , Levillain Romuald , Goudeau Alain 
Published: December 2019

ABSTRACT
We report the case of an occasional intravenous drug user who developed two successive hepatitis C virus (HCV) infections. The first infection led to seroconversion (anti-HCV antibodies detected) and the detection of HCV RNA in serum. After a spontaneous recovery (normalization of alanine aminotransferase levels and HCV RNA clearance), a second HCV infection was observed, with the recurrence of HCV viremia. Antibody directed against HCV serotype 1 was detected throughout the follow-up monitoring, but two different HCV strains were identified during the two infectious episodes: genotype 1a for the first and genotype 3a for the second. This observation shows that primary HCV infection does not confer protective immunity against subsequent infection with viruses of other genotypes. This may hamper the development of a vaccine. Conflicting results were obtained in genotyping and serotyping assays, suggesting that the serotyping method cannot be used to identify the HCV type in patients, such as intravenous drug users, who are exposed to successive HCV infections.
The Papillomavirus Minor Capsid Protein, L2, Induces Localization of the Major Capsid Protein, L1, and the Viral Transcription/Replication Protein, E2, to PML Oncogenic Domains
Authors: Day Patricia M. , Roden Richard B. S. , Lowy Douglas R. , Schiller John T. 
Published: December 2019

ABSTRACTWe have used immunofluorescent staining and confocal microscopy to examine the subcellular localization of structural and nonstructural bovine papillomavirus (BPV) proteins in cultured cells that produce infectious virions. When expressed separately, L1, the major capsid protein, showed a diffuse nuclear distribution while L2, the minor capsid protein, was found to localize to punctate nuclear regions identified as promonocytic leukemia protein (PML) oncogenic domains (PODs). Coexpression of L1 and L2 induced a relocation of L1 into the PODs, leading to the colocalization of L1 and L2. The effect of L2 expression on the distribution of the nonstructural viral proteins E1 and E2, which are required for maintenance of the genome and viral DNA synthesis, was also examined. The localization of the E1 protein was unaffected by L2 expression. However, the pattern of anti-E2 staining was dramatically altered in L2-expressing cells. Similar to L1, E2 was shifted from a dispersed nuclear locality into the PODs and colocalized with L2. The recruitment of full-length E2 by L2 occurred in the absence of other viral components. L2 was shown previously to be essential for the generation of infectious BPV. Our present results provide evidence for a role for L2 in the organization of virion components by recruiting them to a distinct nuclear domain. This L2-dependent colocalization probably serves as a mechanism to promote the assembly of papillomaviruses either by increasing the local concentration of virion constituents or by providing the physical architecture necessary for efficient packaging and assembly. The data also suggest a role for a nonstructural viral protein, E2, in virion assembly, specifically the recruitment of the viral genome to the sites of assembly, through its high-affinity interaction with specific sequences in the viral DNA.
The 131-Amino-Acid Repeat Region of the Essential 39-Kilodalton Core Protein of Fowlpox Virus FP9, Equivalent to Vaccinia Virus A4L Protein, Is Nonessential and Highly Immunogenic
Authors: Boulanger Denise , Green Philip , Smith Tim , Czerny Claus-Peter , Skinner Michael A. 
Published: December 2019

ABSTRACTThe immunodominant, 39,000-molecular weight core protein (39K protein) of fowlpox virus (FP9 strain), equivalent to the vaccinia virus A4L gene product, contains highly charged domains at each end of the protein and multiple copies of a 12-amino-acid serine-rich repeat sequence in the middle of the protein. Similar repeats were also detected in other fowlpox virus strains, suggesting that they might confer a selective advantage to the virus. The molloscum contagiosum virus homolog (MC107L) also contains repeats, unlike the vaccinia virus protein. The number of repeats in the fowlpox virus protein does not seem to be crucial, since some strains have a different number of repeats, as shown by the difference in the size of the protein in these strains. The repeat region could be deleted, indicating that it is not essential for replication in vitro. It was not possible to delete the entire 39K protein, indicating that it was essential (transcriptional control signals for the flanking genes were left intact). The repeat region is partly responsible for the immunodominance of the protein, but the C-terminal part of the protein also contains highly antigenic linear epitopes. A role for the 39K protein in immune system modulation is discussed.
The N Terminus of Rotavirus VP2 Is Necessary for Encapsidation of VP1 and VP3
Authors: Zeng Carl Q.-Y. , Estes Mary K. , Charpilienne Annie , Cohen Jean 
Published: December 2019

ABSTRACT
The innermost core of rotavirus is composed of VP2, which forms a protein layer that surrounds the two minor proteins VP1 and VP3, and the genome of 11 segments of double-stranded RNA. This inner core layer surrounded by VP6, the major capsid protein, constitutes double-layered particles that are transcriptionally active. Each gene encoding a structural protein of double-layered particles has been cloned into baculovirus recombinants and expressed in insect cells. Previously, we showed that coexpression of different combinations of the structural proteins of rotavirus double-layered particles results in the formation of virus-like particles (VLPs), and each VLP containing VP1, the presumed RNA-dependent RNA polymerase, possesses replicase activity as assayed in an in vitro template-dependent assay system (C. Q.-Y. Zeng, M. J. Wentz, J. Cohen, M. E. Estes, and R. F. Ramig, J. Virol. 70:2736–2742, 1996). This work reports construction and characterization of VLPs containing a truncated VP2 (VPΔ2, containing amino acids [aa] Met-93 to 880). Expression of VPΔ2 alone resulted in the formation of single-layered Δ2-VLPs. Coexpression of VPΔ2 with VP6 produced double-layered Δ2/6-VLPs. VLPs formed by coexpression of VPΔ2 and VP1 or VP3, or both VP1 and VP3, resulted in the formation of VLPs lacking both VP1 and VP3. The presence of VP6 with VPΔ2 did not result in encapsidation of VP1 and VP3. To determine the domain of VP2 required for binding VP1, far-Western blot analyses using a series of truncated VP2 constructs were performed to test their ability to bind VP1. These analyses showed that (i) full-length VP2 (aa 1 to 880) binds to VP1, (ii) any N-terminal truncation lacking aa 1 to 25 fails to bind VP1, and (iii) a C-terminal 296-aa truncated VP2 construct (aa 1 to 583) maintains the ability to bind VP1. These analyses indicate that the N terminus of rotavirus VP2 is necessary for the encapsidation of VP1 and VP3.
Cells with High Cyclophilin A Content Support Replication of Human Immunodeficiency Virus Type 1 Gag Mutants with Decreased Ability To Incorporate Cyclophilin A
Authors: Ackerson Bradley , Rey Osvaldo , Canon Jude , Krogstad Paul 
Published: December 2019

ABSTRACT
Gag polyprotein-mediated incorporation of cellular cyclophilin A (CyPA) into virions is essential for the formation of infectious human immunodeficiency virus type 1 (HIV-1) virions. Either a point mutation in Gag (P222A) or drugs which bind CyPA decrease virion incorporation of CyPA and interfere with HIV-1 replication. We have found that lymphoid cells varied greatly in their CyPA content and that cells with high CyPA content supported the replication of P222A HIV-1 Gag mutants. These experiments demonstrated that a higher cellular CyPA content of some cells was able to compensate for the decreased binding affinity of P222A mutant Gag for CyPA, allowing virus replication to occur.
Quantitative Disassembly and Reassembly of Human Papillomavirus Type 11 Viruslike Particles In Vitro
Authors: McCarthy Michael P. , White Wendy I. , Palmer-Hill Frances , Koenig Scott , Suzich Joann A. 
Published: December 2019

ABSTRACT
The human papillomavirus (HPV) capsid is primarily composed of a structural protein denoted L1, which forms both pentameric capsomeres and capsids composed of 72 capsomeres. The L1 protein alone is capable of self-assembly in vivo into capsidlike structures referred to as viruslike particles (VLPs). We have determined conditions for the quantitative disassembly of purified HPV-11 L1 VLPs to the level of capsomeres, demonstrating that disulfide bonds alone are essential to maintaining long-term HPV-11 L1 VLP structure at physiological ionic strength. The ionic strength of the disassembly reaction was also important, as increased NaCl concentrations inhibited disassembly. Conversely, chelation of cations had no effect on disassembly. Quantitative reassembly to a homogeneous population of 55-nm, 150S VLPs was reliably achieved by the re-formation of disulfide linkages following removal of reducing agent at near-neutral pH and moderate NaCl concentration. HPV-11 L1 VLPs could also be dissociated by treatment with carbonate buffer at pH 9.6, but VLPs could not be regenerated following carbonate treatment. When probed with conformationally sensitive and/or neutralizing monoclonal antibodies, both capsomeres generated by disulfide reduction of purified VLPs and reassembled VLPs formed from capsomeres upon removal of reducing agents exhibited epitopes found on the surface of authentic HPV-11 virions. Antisera raised against either purified VLP starting material or reassembled VLPs similarly neutralized infectious HPV-11 virions. The ability to disassemble and reassemble VLPs in vitro and in bulk allows basic features of capsid assembly to be studied and also opens the possibility of packaging selected exogenous compounds within the reassembled VLPs.
The Murine CAR Homolog Is a Receptor for Coxsackie B Viruses and Adenoviruses
Authors: Bergelson Jeffrey M. , Krithivas Anita , Celi Leo , Droguett Gustavo , Horwitz Marshall S. , Wickham Thomas , Crowell Richard L. , Finberg Robert W. 
Published: December 2019

ABSTRACTComplementary DNA clones encoding the murine homolog (mCAR) of the human coxsackievirus and adenovirus receptor (CAR) were isolated. Nonpermissive CHO cells transfected with mCAR cDNA became susceptible to infection by coxsackieviruses B3 and B4 and showed increased susceptibility to adenovirus-mediated gene transfer. These results indicate that the same receptor is responsible for virus interactions with both murine and human cells. Analysis of receptor expression in human and murine tissues should be useful in defining factors governing virus tropism in vivo.
Analysis of the Temporal Relationship between Human Immunodeficiency Virus Type 1 Quasispecies in Sequential Blood Samples and Various Organs Obtained at Autopsy
Authors: van’t Wout Angélique B. , Ran Leonie J. , Kuiken Carla L. , Kootstra Neeltje A. , Pals Steven T. , Schuitemaker Hanneke 
Published: December 2019

ABSTRACTWe studied the temporal relationship between human immunodeficiency type 1 (HIV-1) quasispecies in tissues and in peripheral blood mononuclear cells (PBMC) of infected individuals. Sequential PBMC and tissue samples from various organs obtained at autopsy from three patients who died of AIDS-related complications were available for analysis. Biological HIV-1 clones were isolated from PBMC samples, and cellular tropism and syncytium-inducing (SI) capacity were determined. Genomic DNA was isolated from 1 cm3of organ tissue, and proviral DNA was amplified by means of PCR and cloned with the PGEM-T vector system. A 185-bp region encompassing the third variable domain of the virus envelope, known to influence HIV-1 biological properties, was sequenced. HIV-1 could be amplified from all PBMC and organ samples, except from liver tissue for two patients. Both SI and non-syncytium-inducing (NSI) genotypes could be detected in the different tissues. Tissue-specific quasispecies were observed in brain, lung, and testis. Lymphoid tissues, such as bone marrow, lymph node, and spleen, harbored several different variants similar to those detected in blood in the last PBMC samples. In general, only tissues in which macrophages are likely to be the main target cell for HIV-1 harbored NSI HIV-1 sequences that clustered separately. Both SI and NSI sequences that clustered with sequences from late-stage PBMC were present in other tissues, which may indicate that the presence of HIV-1 in those tissues is secondary to lymphocyte infiltration rather than to tissue tropism of HIV-1 itself. These data suggest that the viral reservoir may be limited, which will have important implications for the success of HIV-1 eradication.
Role of Baculovirus IE2 and Its RING Finger in Cell Cycle Arrest
Authors: Prikhod’ko Elena A. , Miller Lois K. 
Published: December 2019

ABSTRACT

The
ie2
gene of
Autographa californica
nuclear polyhedrosis virus (Ac
M
NPV) is known to transactivate transient expression from viral promoters in a host cell-specific manner. We report that transfection of
Spodoptera frugiperda
(SF-21) cells with
ie2
was sufficient to arrest the cell cycle, resulting in the accumulation of enlarged cells with abnormally high DNA contents. By 72 h posttransfection, more than 50% of
ie2
-transfected cells had DNA contents greater than 4N. There was no evidence of mitotic spindle formation in these cells, and expression of
ie2
appeared to block cell cycle progression in S phase. Several
ie2
mutants were analyzed to further define the region of IE2 responsible for arresting the cell cycle. Analysis of these mutants showed that deletion of the RING finger motif eliminated the ability of IE2 to arrest the cell cycle but did not affect its ability to transactivate the
ie1
promoter. Moreover, mutation of a single conserved cysteine (C251) of the RING finger motif abolished the ability of IE2 to block cell cycle progression but had no apparent effect on its
trans
-regulatory activity. In contrast, a mutant of IE2 containing a deletion of residues 94 to 173 was able to block cell division but lacked
trans
-regulatory activity. Thus, the ability of IE2 to arrest the cell cycle depended on the integrity of the RING finger motif and was distinct from and independent of its ability to
trans
-activate the
ie1
promoter. IE2 also arrested the division of cells derived from other insect species,
Trichoplusia ni
(TN-368 and BTI-TN-5B1-4) and
Helicoverpa zea
(Hz-AM1).