bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genetic characterisation of Australian strains of porcine circovirus types 1 and 2
Authors: Muhling J , Raye WS , Buddle JR , Wilcox GE 
Published: December 2006

Objective  As post‐weaning multi‐systemic wasting syndrome (PMWS) has not been identified within Australia, to determine if the absence of disease was associated with genetic differences between the strains of porcine circovirus (PCV) present in Australia and those from countries in association with PMWS.Design  Pig tissues were obtained from weaned pigs found dead or presenting with clinical signs of illthrift and also from neonatal pigs with congenital tremors and used as a source of virus DNA for sequence analysis.Procedure  DNA was extracted from the tissues and PCV detected by polymerase chain reaction (PCR). PCR with PCV type‐specific primers was used to amplify the entire genome from selected tissues. The genomes of three strains of PCV1 and seven strains of PCV2 from three Australian states were sequenced and subjected to phylogenetic analysis using standard procedures.Results  The three Australian PCV1 strains had 98 to 99% nucleotide identity to strains in other countries and the seven Australian PCV2 strains had 94 to 99% identity to PCV2 strains in other countries where PMWS has occurred. Six of the seven Australian PCV2 strains were genetically similar to each other, while the seventh was more distantly related. There were no consistent differences in the predicted amino acid sequence of the Australian strains of PCV2 and strains associated with PMWS in other countries.Conclusion  There were no consistent differences between Australian strains of PCV and those that have been associated with PMWS in other countries and it appears likely that other factors are responsible for the absence of PMWS in Australia.
Detection and Differentiation of
Bordetella
spp. by Real-Time PCR
Authors: Koidl Christoph , Bozic Michael , Burmeister Anja , Hess Markus , Marth Egon , Kessler Harald H. 
Published: December 2006

ABSTRACT

Molecular detection of
Bordetella pertussis
DNA is a sensitive and specific method for the rapid diagnosis of pertussis. In this study, a new molecular assay for the detection and differentiation of
Bordetella
spp. based on automated DNA extraction and real-time PCR was evaluated. The analytical sensitivity of the new assay was determined by Probit analysis of serial dilutions of both cloned PCR products IS
481
and IS
1001
and cell suspensions of
B. pertussis
,
B. parapertussis
, and
B. bronchiseptica
. The specificity was analyzed by testing a number of pathogens producing respiratory infections. Moreover, a total of 92 clinical samples were investigated. The results were compared to those obtained by an in-house assay based on manual DNA extraction, followed by real-time PCR and detection of IS
481
. The analytical sensitivity of the new assay for the detection of IS
481
and IS
1001
was determined to be 2.2 and 1.2 genome equivalents/μl, respectively. The analytical sensitivity for the detection of
B. pertussis
,
B. parapertussis
, and
B. bronchiseptica
was determined to be 1.6, 1.0, and 2.7 genome equivalents/μl, respectively. When clinical specimens were tested with the new assay, 46 of 92 were found to be positive for
Bordetella
DNA. With the in-house assay, 45 samples tested positive. The new molecular assay proved to be suitable for the rapid diagnosis of pertussis in the routine diagnostic laboratory.
Isolation of
Trichophyton violaceum
and
Trichophyton soudanense
in Baltimore, Maryland
Authors: Magill Shelley S. , Manfredi Liliana , Swiderski Andrew , Cohen Bernard , Merz William G. 
Published: December 2006

ABSTRACT

Tinea capitis is of public health importance because of its transmissibility.
Trichophyton violaceum
and
Trichophyton soudanense
, which are common causes of tinea capitis in parts of Africa and West Asia, have only rarely been reported to cause dermatophytoses in the United States. We identified 24 patients with 25 positive cultures for
T. violaceum
or
T. soudanense
that were processed in a single hospital laboratory in Baltimore, Maryland, between 1 January 2000 and 30 June 2006. Most patients for whom clinical information was available had tinea capitis. There was a marked increase in the isolation of these organisms between the period from 2000 to 2002 and the period from 2003 to 2006, possibly associated with changes in immigration to the Baltimore metropolitan area. The changing epidemiology of this transmissible fungal infection not only is of public health interest as an example of the introduction of a “new” pathogen to an area where it traditionally was not endemic but also is of clinical and microbiological importance given reports suggesting an increasing incidence of tinea capitis in some areas and increasing clinical failure rates of current therapies.
Detection of
Treponema pallidum
subsp.
pallidum
from Skin Lesions, Serum, and Cerebrospinal Fluid in an Infant with Congenital Syphilis after Clindamycin Treatment of the Mother during Pregnancy
Authors: Woznicová Vladana , Šmajs David , Wechsler Dan , Matějková Petra , Flasarová Magdalena 
Published: December 2006

ABSTRACT

We report here a case of congenital syphilis in a newborn after clindamycin treatment in pregnancy. Using PCR detection of
tmpC
(TP0319) and DNA sequencing of the genes TP0136 and TP0548, DNA sequences identical to
Treponema pallidum
subsp.
pallidum
strain SS14 were detected in the infant's skin lesions, serum, and cerebrospinal fluid.
Absence or Overexpression of the Varicella-Zoster Virus (VZV) ORF29 Latency-Associated Protein Impairs Late Gene Expression and Reduces VZV Latency in a Rodent Model
Authors: Cohen Jeffrey I. , Krogmann Tammy , Pesnicak Lesley , Ali Mir A. 
Published: December 2006

ABSTRACT

Varicella-zoster virus (VZV) ORF29 encodes the viral single-stranded DNA binding protein and is expressed during latency in human ganglia. We constructed an ORF29 deletion mutant virus and showed that the virus could replicate only in cells expressing ORF29. An ORF29-repaired virus, in which ORF29 was driven by a cytomegalovirus promoter, grew to peak titers similar to those seen with the parental virus. The level of ORF29 protein in cells infected with the repaired virus was greater than that seen with parental virus. Infection of cells with either the ORF29 deletion or repaired virus resulted in similar levels of VZV immediate-early proteins but reduced levels of glycoprotein E compared to those observed with parental virus. Cotton rats infected with the ORF29 deletion mutant had a markedly reduced frequency of latent infection in dorsal root ganglia compared with those infected with parental virus (
P
< 0.00001). In contrast, infection of animals with the ORF29 deletion mutant resulted in a frequency of ganglionic infection at 3 days similar to that seen with the parental virus. Animals infected with the ORF29-repaired virus, which overexpresses ORF29, also had a reduced frequency of latent infection compared with those infected with parental virus (
P
= 0.0044). These studies indicate that regulation of ORF29 at appropriate levels is critical for VZV latency in a rodent model.
Novel Pathway for Induction of Latent Virus from Resting CD4+T Cells in the Simian Immunodeficiency Virus/Macaque Model of Human Immunodeficiency Virus Type 1 Latency
Authors: Shen Anding , Yang Hung-Chih , Zhou Yan , Chase Amanda J. , Boyer Jean D. , Zhang Hao , Margolick Joseph B. , Zink M. Christine , Clements Janice E. , Siliciano Robert F. 
Published: December 2006

ABSTRACTAlthough combination therapy allows the suppression of human immunodeficiency virus type 1 (HIV-1) viremia to undetectable levels, eradication has not been achieved because the virus persists in cellular reservoirs, particularly the latent reservoir in resting CD4+T lymphocytes. We previously established a simian immunodeficiency virus (SIV)/macaque model to study latency. We describe here a novel mechanism for the induction of SIV from latently infected resting CD4+T cells. Several human cell lines including CEMx174 and Epstein-Barr virus-transformed human B-lymphoblastoid cell lines mediated contact-dependent activation of resting macaque T cells and induction of latent SIV. Antibody-blocking assays showed that interactions between the costimulatory molecule CD2 and its ligand CD58 were involved, whereas soluble factors and interactions between T-cell receptors and major histocompatibility complex class II were not. Combinations of specific antibodies to CD2 also induced T-cell activation and virus induction in human resting CD4+T cells carrying latent HIV-1. This is the first demonstration that costimulatory signals can induce latent virus without the coengagement of the T-cell receptor, and this study might provide insights into potential pathways to target latent HIV-1.
Nonstructural Proteins of Respiratory Syncytial Virus Suppress Premature Apoptosis by an NF-κB-Dependent, Interferon-Independent Mechanism and Facilitate Virus Growth
Authors: Bitko Vira , Shulyayeva Olena , Mazumder Barsanjit , Musiyenko Alla , Ramaswamy Murali , Look Dwight C. , Barik Sailen 
Published: December 2006

ABSTRACT
The two nonstructural (NS) proteins NS1 and NS2 of respiratory syncytial virus (RSV) are abundantly expressed in the infected cell but are not packaged in mature progeny virions. We found that both proteins were expressed early in infection, whereas the infected cells underwent apoptosis much later. Coincident with NS protein expression, a number of cellular antiapoptotic factors were expressed or activated at early stages, which included NF-κB and phosphorylated forms of protein kinases AKT, phosphoinositide-dependent protein kinase, and glycogen synthase kinase. Using specific short interfering RNAs (siRNAs), we achieved significant knockdown of one or both NS proteins in the infected cell, which resulted in abrogation of the antiapoptotic functions and led to early apoptosis. NS-dependent suppression of apoptosis was observed in Vero cells that are naturally devoid of type I interferons (IFN). The siRNA-based results were confirmed by the use of NS-deleted RSV mutants. Early activation of epidermal growth factor receptor (EGFR) in the RSV-infected cell did not require NS proteins. Premature apoptosis triggered by the loss of NS or by apoptosis-promoting drugs caused a severe reduction of RSV growth. Finally, recombinantly expressed NS1 and NS2, individually and together, reduced apoptosis by tumor necrosis factor alpha, suggesting an intrinsic antiapoptotic property of both. We conclude that the early-expressed nonstructural proteins of RSV boost viral replication by delaying the apoptosis of the infected cell via a novel IFN- and EGFR-independent pathway.
Fitness Landscape of Human Immunodeficiency Virus Type 1 Protease Quasispecies
Authors: Fernàndez Guerau , Clotet Bonaventura , Martínez Miguel Angel 
Published: December 2006

ABSTRACT
Here we show, at a high resolution (1%), the human immunodeficiency virus
type 1 (HIV-1) protease gene quasispecies landscape from
three infected naïve individuals. A huge range of genetic
configurations was found (67%, 71%, and 80% of the nucleotide clones
from the three individuals, respectively, were different), and these
configurations created a dense net that linked different parts of the
viral population. Similarly, a vast diversity of different protease
activities was also found. Importantly, 65% of the analyzed enzymes had
detectable protease activity, and 11% of the minority individual
variants showed similar or better fitness than the master (most
abundant) enzyme, suggesting that the viral complexity in this genomic
region does not exclusively depend on the enzyme's catalytic
efficiency. Several high-fitness minority variants had only one
substitution compared to the master sequence, supporting the
possibility that the rugged HIV-1 protease quasispecies fitness
landscape may be formed by a continuous network that can be traversed
by single mutational steps without passing through defective or
less-adapted
proteins.
The Minimal Replicator Element of the Kaposi's Sarcoma-Associated Herpesvirus Terminal Repeat Supports Replication in a Semiconservative and Cell-Cycle-Dependent Manner
Authors: Verma Subhash C. , Choudhuri Tathagata , Robertson Erle S. 
Published: December 2006

ABSTRACT
Kaposi's
sarcoma-associated herpesvirus (KSHV) persists as episomes in infected
cells by circularizing at the terminal repeats (TRs). The KSHV episome
carries multiple reiterated copies of the terminal repeat, and each
copy is capable of supporting replication. Expression of the
latency-associated nuclear antigen (LANA) is critical for the
replication of TR-containing plasmids. A 32-bp sequence upstream of
LANA binding site 1 (LBS1), referred to as RE (replication element),
along with LANA binding sites 1 and 2 (RE-LBS1/2), is sufficient to
support replication (J. Hu and R. Renne, J. Virol.
79:2637-2642, 2005). In this report we demonstrate that the
minimal replicator element (RE-LBS1/2) replicates in synchrony with the
host cellular DNA, and only once, in a cell-cycle-dependent
manner. Overexpression of the mammalian replication inhibitor geminin
blocked replication of the plasmid containing the minimal replicator
element, confirming the involvement of the host cellular replication
control mechanism, and prevented rereplication of the plasmid in the
same cell cycle. Overexpression of Cdt1 also rescued the replicative
ability of the RE-LBS1/2-containing plasmids. A chromatin
immunoprecipitation assay performed using anti-origin recognition
complex 2 (α-ORC2) and α-LANA antibodies from cells
transfected with RE-LBS1/2, RE-LBS1, LBS1, or RE showed the association
of ORC2 with the RE region. Expression of LANA increased the number
of copies of chromatin-bound DNA of replication elements,
suggesting that LANA is important for the recruitment of ORCs and may
contribute to the stabilization of the replication protein complexes at
the RE
site.
Human Cytomegalovirus Disrupts both Ataxia Telangiectasia Mutated Protein (ATM)- and ATM-Rad3-Related Kinase-Mediated DNA Damage Responses during Lytic Infection
Authors: Luo Min Hua , Rosenke Kyle , Czornak Kamila , Fortunato Elizabeth A. 
Published: December 2006

ABSTRACT
Many viruses (herpes simplex virus type 1, polyomavirus, and human immunodeficiency virus type 1) require the activation of ataxia telangiectasia mutated protein (ATM) and/or Mre11 for a fully permissive infection. However, the longer life cycle of human cytomegalovirus (HCMV) may require more specific interactions with the DNA repair machinery to maximize viral replication. A prototypical damage response to the double-stranded ends of the incoming linear viral DNA was not observed in fibroblasts at early times postinfection (p.i.). Apparently, a constant low level of phosphorylated ATM was enough to phosphorylate its downstream targets, p53 and Nbs1. p53 was the only cellular protein observed to relocate at early times, forming foci in infected cell nuclei between 3.5 and 5.5 h p.i. Approximately half of these foci localized with input viral DNA, and all localized with viral UL112/113 prereplication site foci. No other DNA repair proteins localized with the virus or prereplication foci in the first 24 h p.i. When viral replication began in earnest, between 24 and 48 h p.i., there were large increases in steady-state levels and phosphorylation of many proteins involved in the damage response, presumably triggered by ATM-Rad3-related kinase activation. However, a sieving process occurred in which only certain proteins were specifically sequestered into viral replication centers and others were particularly excluded. In contrast to other viruses, activation of a damage response is neither necessary nor detrimental to infection, as neither ATM nor Mre11 was required for full virus replication and production. Thus, by preventing simultaneous relocalization of all the necessary repair components to the replication centers, HCMV subverts full activation and completion of both double-stranded break and S-phase checkpoints that should arrest all replication within the cell and likely lead to apoptosis.
Protective Role of Beta Interferon in Host Defense against Influenza A Virus
Authors: Koerner Iris , Kochs Georg , Kalinke Ulrich , Weiss Siegfried , Staeheli Peter 
Published: December 2006

ABSTRACT

Type I interferon (IFN), which includes the IFN-α and -β subtypes, plays an essential role in host defense against influenza A virus. However, the relative contribution of IFN-β remains unresolved. In mice, type I IFN is effective against influenza viruses only if the IFN-induced resistance factor Mx1 is present, though most inbred mouse strains, including the recently developed IFN-β-deficient mice, bear only defective
Mx1
alleles. We therefore generated IFN-β-deficient mice carrying functional
Mx1
alleles (designated Mx-BKO) and compared them to either wild-type mice bearing functional copies of both
IFN-β
and
Mx1
(designated Mx-wt) or mice carrying functional
Mx1
alleles but lacking functional type I IFN receptors (designated Mx-IFNAR). Influenza A virus strain SC35M (H7N7) grew to high titers and readily formed plaques in monolayers of Mx-BKO and Mx-IFNAR embryo fibroblasts which showed no spontaneous expression of
Mx1
. In contrast, Mx-wt embryo fibroblasts were found to constitutively express
Mx1
, most likely explaining why SC35M did not grow to high titers and formed no visible plaques in such cells. In vivo challenge experiments in which SC35M was applied via the intranasal route showed that the 50% lethal dose was about 20-fold lower in Mx-BKO mice than in Mx-wt mice and that virus titers in the lungs were increased in Mx-BKO mice. The resistance of Mx-BKO mice to influenza A virus strain PR/8/34 (H1N1) was also substantially reduced, demonstrating that IFN-β plays an important role in the defense against influenza A virus that cannot be compensated for by IFN-α.
Persistent Activation of STAT3 by Latent Kaposi's Sarcoma-Associated Herpesvirus Infection of Endothelial Cells
Authors: Punjabi Almira S. , Carroll Patrick A. , Chen Lei , Lagunoff Michael 
Published: December 2006

ABSTRACT
Kaposi's sarcoma-associated herpesvirus (KSHV) is the infectious cause of Kaposi's sarcoma, primary effusion lymphoma, and plasmablastic multicentric Castleman's disease. STAT3 has been shown to be important for the maintenance of primary effusion lymphoma cells in culture and is chronically activated in many tumor cell lines. However, little is known about the role of KSHV in the activation of STAT3 or the role of STAT3 in KS tumors. We demonstrate that STAT3 is activated by KSHV infection of endothelial cells, the KS tumor cell type, in a biphasic fashion. Viral binding and entry activate STAT3 in the first 2 h after infection, but this activation dissipates by 4 h postinfection. By 12 h after KSHV infection, concomitant with the expression of latent genes, STAT3 is once again activated, and this activation persists for as long as latent infection is maintained. Activated STAT3 translocates to the nucleus, where it can bind to STAT3-specific DNA elements and can activate STAT3-dependent promoter activity. Conditioned medium from KSHV-infected endothelial cells is able to transiently activate STAT3, indicating the involvement of a secreted factor and that a latency-associated factor in KSHV-infected cells is necessary for sustained activation. KSHV upregulates gp130 receptor expression, and both gp130 and JAK2 are required for the activation of STAT3. However, neither human nor viral interleukin-6 is required for STAT3 activation. Persistent activation of the oncogenic signal transducer, STAT3, by KSHV may play a critical role in the viral pathogenesis of Kaposi's sarcoma, as well as in primary effusion lymphomas.
Notch1 Augments Intracellular Trafficking of Adeno-Associated Virus Type 2
Authors: Ren Changchun , White April F. , Ponnazhagan Selvarangan 
Published: December 2006

ABSTRACT
We report here the significance of the Notch1 receptor in intracellular trafficking of recombinant adeno-associated virus type 2 (rAAV2). RNA profiling of human prostate cancer cell lines with various degrees of AAV transduction indicated a correlation of the amount of Notch1 with rAAV transgene expression. A definitive role of Notch1 in enhancing AAV transduction was confirmed by developing clonal derivatives of DU145 cells overexpressing either full-length or intracellular Notch1. To discern stages of AAV2 transduction influenced by Notch1, competitive binding with soluble heparin and Notch1 antibody, intracellular trafficking using Cy3-labeled rAAV2, and blocking assays for proteasome and dynamin pathways were performed. Results indicated that in the absence or low-level expression of Notch1, only binding of virus was found on the cell surface and internalization was impaired. However, increased Notch1 expression in these cells allowed efficient perinuclear accumulation of labeled capsids. Nuclear transport of the vector was evident by transgene expression and real-time PCR analyses. Dynamin levels were not found to be different among these cell lines, but blocking dynamin function abrogated AAV2 transduction in DU145 clones overexpressing full-length Notch1 but not in clones overexpressing intracellular Notch1. These studies provide evidence for the role of activated Notch1 in intracellular trafficking of AAV2, which may have implications in the optimal use of AAV2 in human gene therapy.
Recovery of Infectious Bluetongue Virus from RNA
Authors: Boyce Mark , Roy Polly 
Published: December 2006

ABSTRACT

Bluetongue virus (BTV) is an insect-vectored emerging pathogen of ruminants with the potential for devastating economic impact on European agriculture. BTV and many other members of the
Reoviridae
have remained stubbornly refractory to the development of methods for the rescue of infectious virus from cloned nucleic acid (reverse genetics). Partially disassembled virus particles are transcriptionally active, synthesizing viral transcripts in the cytoplasm of infected cells, in essence delivering viral nucleic acids in situ. With the goal of generating a reverse-genetics system for BTV, we examined the possibility of recovering infectious BTV by the transfection of BSR cells with BTV transcripts (single-stranded RNA [ssRNA]) synthesized in vitro using BTV core particles. Following transfection, viral-protein synthesis was detected by immunoblotting, and confocal examination of the cells showed a punctate cytoplasmic distribution of inclusion bodies similar to that seen in infected cells. Viral double-stranded RNA (dsRNA) was isolated from ssRNA-transfected cells, demonstrating that replication of the ssRNA had occurred. Additionally, infectious virus was present in the medium of transfected cells, as demonstrated by the passage of infectivity in BSR cells. Infectivity was sensitive to single-strand-specific RNase A, and cotransfection of genomic BTV dsRNA with transcribed ssRNA demonstrated that the ssRNA species, rather than dsRNA, were the active components. We conclude that it is possible to recover infectious BTV wholly from ssRNA, which suggests a means for establishing helper virus-independent reverse-genetics systems for members of the
Reoviridae
.