bluuebunny/crossref_metadata_2025_split huggingface dataset:
Improved Diagnosis of Periprosthetic Joint Infection by Multiplex PCR of Sonication Fluid from Removed Implants
Authors: Achermann Yvonne , Vogt Markus , Leunig Michael , Wüst Jürg , Trampuz Andrej 
Published: February 2010

ABSTRACT

The microbiological diagnosis of periprosthetic joint infection (PJI) is crucial for successful antimicrobial treatment. Cultures have limited sensitivity, especially in patients receiving antibiotics. We evaluated the value of multiplex PCR for detection of microbial DNA in sonication fluid from removed orthopedic prostheses. Cases of PJI in which the prosthesis (or part of it) was removed were prospectively included. The removed implant was sonicated, and the resulting sonication fluid was cultured and subjected to multiplex PCR. Of 37 PJI cases (17 hip prostheses, 14 knee prostheses, 4 shoulder prostheses, 1 elbow prosthesis, and 1 ankle prosthesis), pathogens were identified in periprosthetic tissue in 24 (65%) cases, in sonication fluid in 23 (62%) cases, and by multiplex PCR in 29 (78%) cases. The pathogen was detected in 5 cases in sonication fluid only (
Propionibacterium acnes
in all cases; none of these patients had previously received antibiotics) and in 11 cases by multiplex PCR only (all of these patients had previously received antibiotics). After exclusion of 8 cases caused by
P. acnes
or
Corynebacterium
species, which cannot be detected due to the absence of specific primers in the PCR kit, sonication cultures were positive in 17 cases and multiplex PCR sonication cultures were positive in 29 cases (59% versus 100%, respectively;
P
< 0.01). Among 19 cases (51%) receiving antibiotics, multiplex PCR was positive in all 19 (100%), whereas sonication cultures grew the organism in 8 (42%) (
P
< 0.01). Multiplex PCR of sonication fluid is a promising test for diagnosis of PJI, particularly in patients who previously received antibiotics. With modified primer sets, multiplex PCR has the potential for further improvement of the diagnosis of PJI.
Comparison of Traditional Phenotypic Identification Methods with Partial 5′ 16S rRNA Gene Sequencing for Species-Level Identification of Nonfermenting Gram-Negative Bacilli
Authors: Cloud Joann L. , Harmsen Dag , Iwen Peter C. , Dunn James J. , Hall Gerri , LaSala Paul Rocco , Hoggan Karen , Wilson Deborah , Woods Gail L. , Mellmann Alexander 
Published: February 2010

ABSTRACT
Correct identification of nonfermenting Gram-negative bacilli (NFB) is crucial for patient management. We compared phenotypic identifications of 96 clinical NFB isolates with identifications obtained by 5′ 16S rRNA gene sequencing. Sequencing identified 88 isolates (91.7%) with >99% similarity to a sequence from the assigned species; 61.5% of sequencing results were concordant with phenotypic results, indicating the usability of sequencing to identify NFB.
Performance of an Agar Dilution Method and a Vitek 2 Card for Detection of Inducible Clindamycin Resistance in
Staphylococcus
spp
Authors: Lavallée Christian , Rouleau Danielle , Gaudreau Christiane , Roger Michel , Tsimiklis Catherine , Locas Marie-Claude , Gagnon Simon , Delorme Jocelyn , Labbé Annie-Claude 
Published: February 2010

ABSTRACT

The D-zone test detects inducible clindamycin resistance in
Staphylococcus
spp. Two other methods not described by the Clinical and Laboratory Standards Institute (CLSI) are available to test for this resistance mechanism: an agar dilution method and new Vitek 2 cards. This study evaluated the performance of both methods in detecting inducible clindamycin resistance. Nonduplicate clinical strains of
Staphylococcus
spp. (111
Staphylococcus aureus
and 52 coagulase-negative staphylococcus strains), intermediate or resistant to erythromycin but susceptible to clindamycin, were obtained from three hospitals in Montreal, Quebec, Canada. Molecular analysis to detect resistance genes was conducted on all strains. A Mueller-Hinton agar containing 1 mg of erythromycin and 0.5 mg of clindamycin/liter was used for the dilution method, and two inocula were tested: 10
4
and 10
5
CFU per spot. Plates were read at 24 and 48 h. The Vitek 2 AST-P580 card was used according to the manufacturer's recommendations. The results were compared to those of the D-zone test. The D-zone test was positive in 134 of 163 (82%) strains. With the 10
4
CFU inoculum, the sensitivities were 84 and 99% at 24 and 48 h, respectively. The 10
5
CFU inoculum increased the sensitivities at 24 and 48 h to 91 and 100%, respectively. The specificity was 100% for the 10
4
CFU inoculum at 24 h and 97% for the other combinations. The sensitivity and specificity for the Vitek 2 card were 93 and 100%, respectively. The performance of both the agar dilution method and the Vitek 2 card was good, but these methods were not as sensitive as the D-zone test at 24 h.
Molecular Typing of Pneumococci for Investigation of Linked Cases of Invasive Pneumococcal Disease
Authors: Pichon Bruno , Moyce Laura , Sheppard Carmen , Slack Mary , Turbitt Deborah , Pebody Richard , Spencer David A. , Edwards Justin , Krahé Daniel , George Robert 
Published: February 2010

ABSTRACT
In winter 2007-2008, an outbreak of pediatric pneumonia caused by serotype 5 pneumococci was identified in a northeast London suburb. Variable number of tandem repeat analyses clustered these pneumococci from the other serotype 5 pneumococci in the United Kingdom, highlighting the importance of this discriminative typing method in supporting epidemiological investigations.
Development of Real-Time PCR Assays for Detection of the
Streptococcus milleri
Group from Cystic Fibrosis Clinical Specimens by Targeting the
cpn60
and 16S rRNA Genes
Authors: Olson A. B. , Sibley C. D. , Schmidt L. , Wilcox M. A. , Surette M. G. , Corbett C. R. 
Published: February 2010

ABSTRACT

Cystic fibrosis (CF) is a multiorgan disease, with the majority of mortalities resulting from pulmonary failure due to repeated pulmonary exacerbations. Recently, members of the
Streptococcus anginosus
group (
S. anginosus
,
S. constellatus
, and
S. intermedius
), herein referred to as the “
Streptococcus milleri
group” (SMG) have been implicated as important etiological pathogens contributing to pulmonary exacerbations in CF patients. This is partly due to better microbiological detection of the SMG species through the development of a novel specific medium termed “McKay agar.” McKay agar demonstrated that SMG has been an underreported respiratory pathogen contributing to lung exacerbations. Our aim was to develop a real-time PCR assay to expedite the detection of SMG within diagnostic samples. The
cpn60
gene was chosen as a target, with all three members amplified using a single hybridization probe set. SMG strain analysis showed that speciation based on melting curve analysis allowed for the majority of the
S. constellatus
(96%),
S. intermedius
(94%), and
S. anginosus
(60%) strains to be correctly identified. To increase specificity for
S. anginosus
, two 16S rRNA real-time PCR assays were developed targeting the 16S rRNA gene. The 16s_SA assay is specific for
S. anginosus
(100%), while the 16s_SCI assay is specific for
S. constellatus
and
S. intermedius
(100%). These assays can detect <10 genome equivalents in pure culture and >10
4
genome equivalents in sputum samples, making this a great tool for assessment of the presence of SMG in complex polymicrobial samples. Novel molecular methods were developed providing detection ability for SMG, an emerging opportunistic pathogen.
Results from the ARTEMIS DISK Global Antifungal Surveillance Study, 1997 to 2007: a 10.5-Year Analysis of Susceptibilities of
Candida
Species to Fluconazole and Voriconazole as Determined by CLSI Standardized Disk Diffusion
Authors: Pfaller M. A. , Diekema D. J. , Gibbs D. L. , Newell V. A. , Ellis D. , Tullio V. , Rodloff A. , Fu W. , Ling T. A. 
Published: February 2010

ABSTRACT

Fluconazole
in vitro
susceptibility test results for 256,882 isolates of
Candida
spp. were collected from 142 sites in 41 countries from June 1997 to December 2007. Data were collected for 197,619 isolates tested with voriconazole from 2001 to 2007. A total of 31 different species of
Candida
were isolated. Increased rates of isolation of the common non-
albicans
species
C. glabrata
(10.2% to 11.7%),
C. tropicalis
(5.4% to 8.0%), and
C. parapsilosis
(4.8% to 5.6%) were noted when the time periods 1997 to 2000 and 2005 to 2007 were compared. Investigators tested clinical isolates of
Candida
spp. by the CLSI M44-A disk diffusion method. Overall, 90.2% of
Candida
isolates tested were susceptible (S) to fluconazole; however, 13 of 31 species identified exhibited decreased susceptibility (<75% S), similar to that seen with the resistant (R) species
C. glabrata
and
C. krusei
. Among 197,619 isolates of
Candida
spp. tested against voriconazole, 95.0% were S and 3% were R. About 30% of fluconazole-R isolates of
C. albicans
,
C. glabrata
,
C. tropicalis
,
C. rugosa
,
C. lipolytica
,
C. pelliculosa
,
C. apicola
,
C. haemulonii
,
C. humicola
,
C. lambica
, and
C. ciferrii
remained S to voriconazole. An increase in fluconazole resistance over time was seen with
C. parapsilosis
,
C. guilliermondii
,
C. lusitaniae
,
C. sake
, and
C. pelliculosa
. Among the emerging fluconazole-R species were
C. guilliermondii
(11.4% R),
C. inconspicua
(53.2% R),
C. rugosa
(41.8% R), and
C. norvegensis
(40.7% R). The rates of isolation of
C. rugosa
,
C. inconspicua
, and
C. norvegensis
increased by 5- to 10-fold over the 10.5-year study period.
C. guilliermondii
and
C. rugosa
were most prominent in Latin America, whereas
C. inconspicua
and
C. norvegensis
were most common in Eastern European countries. This survey identifies several less-common species of
Candida
with decreased susceptibility to azoles. These organisms may pose a future threat to optimal antifungal therapy and underscore the importance of prompt and accurate species identification and antifungal susceptibility testing.
Recruitment of Herpes Simplex Virus Type 1 Immediate-Early Protein ICP0 to the Virus Particle
Authors: Maringer Kevin , Elliott Gillian 
Published: February 2010

ABSTRACT
Although the herpes simplex virus type 1 (HSV-1) tegument is comprised of a large number of viral and cellular proteins, how and where in the cell these proteins are recruited into the virus structure is poorly understood. We have shown previously that the immediate-early gene product ICP0 is packaged by a mechanism dependent on the major tegument protein VP22, while others have shown a requirement for ICP27. We now extend our studies to show that ICP0 packaging correlates directly with the ability of ICP0 to complex with VP22 in infected cells. ICP27 is not, however, present in this VP22-ICP0 complex but is packaged into the virion in a VP22- and ICP0-independent manner. Biochemical fractionation of virions indicated that ICP0 associates tightly with the virus capsid, but intranuclear capsids contained no detectable ICP0. The RING finger domain of ICP0 and the N terminus of VP22 were both shown to be essential but not sufficient for ICP0 packaging and complex formation. Strikingly, however, the N-terminal region of VP22, while unable to form a complex with ICP0, inhibited its translocation from the nucleus to the cytoplasm. PML degradation by ICP0 was efficient in cells infected with this VP22 mutant virus, confirming that ICP0 retains activity. Hence, we would suggest that VP22 is an important molecular partner of ICP0 that controls at least one of its activities: its assembly into the virion. Moreover, we propose that the pathway by which VP22 recruits ICP0 to the virion may begin in the nucleus prior to ICP0 translocation to its final site of assembly in the cytoplasm.
Human T-Lymphotropic Virus Type 1 Transcription and Chromatin-Remodeling Complexes
Authors: Easley Rebecca , Carpio Lawrence , Guendel Irene , Klase Zachary , Choi Soyun , Kehn-Hall Kylene , Brady John N. , Kashanchi Fatah 
Published: February 2010

ABSTRACT
Human T-lymphotropic virus type 1 (HTLV-1) encodes the viral protein Tax, which is believed to act as a viral transactivator through its interactions with a variety of transcription factors, including CREB and NF-κB. As is the case for all retroviruses, the provirus is inserted into the host DNA, where nucleosomes are deposited to ensure efficient packaging. Nucleosomes act as roadblocks in transcription, making it difficult for RNA polymerase II (Pol II) to proceed toward the 3′ end of the genome. Because of this, a variety of chromatin remodelers can act to modify nucleosomes, allowing for efficient transcription. While a number of covalent modifications are known to occur on histone tails in HTLV-1 infection (i.e., histone acetyltransferases [HATs], histone deacetylases [HDACs], and histone methyltransferases [HMTs]), evidence points to the use of chromatin remodelers that use energy from ATP hydrolysis to remodel nucleosomes. Here we confirm that BRG1, which is the core subunit of eight chromatin-remodeling complexes, is essential not only for Tax transactivation but also for viral replication. This is especially evident when wild-type infectious clones of HTLV-1 are used. BRG1 associates with Tax at the HTLV-1 long terminal repeat (LTR), and coexpression of BRG1 and Tax results in increased rates of transcription. The interaction of BRG1 with Tax additionally recruits the basal transcriptional machinery and removes some of the core histones from the nucleosome at the start site (Nuc 1). When using the BRG1-deficient cell lines SW13, C33A, and TSUPR1, we observed little viral transcription and no viral replication. Importantly, while these three cell lines do not express detectable levels of BRG1, much of the SWI/SNF complex remains assembled in the cells. Knockdown of BRG1 and associated SWI/SNF subunits suggests that the BRG1-utilizing SWI/SNF complex PBAF is responsible for HTLV-1 nucleosome remodeling. Finally, HTLV-1 infection of cell lines with a knockdown in BRG1 or the PBAF complex results in a significant reduction in viral production. Overall, we concluded that BRG1 is required for Tax transactivation and HTLV-1 viral production and that the PBAF complex appears to be responsible for nucleosome remodeling.
Identification and Characterization of Clevudine-Resistant Mutants of Hepatitis B Virus Isolated from Chronic Hepatitis B Patients
Authors: Kwon So Young , Park Yong Kwang , Ahn Sung Hyun , Cho Eun Sook , Choe Won Hyeok , Lee Chang Hong , Kim Byung Kook , Ko Soon Young , Choi Hyo Sun , Park Eun-Sook , Shin Gu Choul , Kim Kyun-Hwan 
Published: February 2010

ABSTRACT

Clevudine (CLV) is a nucleoside analog with potent antiviral activity against chronic hepatitis B virus (HBV) infection. Viral resistance to CLV in patients receiving CLV therapy has not been reported. The aim of this study was to characterize CLV-resistant HBV in patients with viral breakthrough (BT) during long-term CLV therapy. The gene encoding HBV reverse transcriptase (RT) was analyzed from chronic hepatitis B patients with viral BT during CLV therapy. Sera collected from the patients at baseline and at the time of viral BT were studied. To characterize the mutations of HBV isolated from the patients, we subjected the HBV mutants to
in vitro
drug susceptibility assays. Several conserved mutations were identified in the RT domain during viral BT, with M204I being the most common.
In vitro
phenotypic analysis showed that the mutation M204I was predominantly associated with CLV resistance, whereas L229V was a compensatory mutation for the impaired replication of the M204I mutant. A quadruple mutant (L129M, V173L, M204I, and H337N) was identified that conferred greater replicative ability and strong resistance to both CLV and lamivudine. All of the CLV-resistant clones were lamivudine resistant. They were susceptible to adefovir, entecavir, and tenofovir, except for one mutant clone. In conclusion, the mutation M204I in HBV RT plays a major role in CLV resistance and leads to viral BT during long-term CLV treatment. Several conserved mutations may have a compensatory role in replication. Drug susceptibility assays reveal that adefovir and tenofovir are the most effective compounds against CLV-resistant mutants. These data may provide additional therapeutic options for CLV-resistant patients.
Adenovirus Protein E4orf4 Induces Premature APC
Cdc20
Activation in
Saccharomyces cerevisiae
by a Protein Phosphatase 2A-Dependent Mechanism
Authors: Mui Melissa Z. , Roopchand Diana E. , Gentry Matthew S. , Hallberg Richard L. , Vogel Jackie , Branton Philip E. 
Published: February 2010

ABSTRACT

Protein phosphatase 2A (PP2A) has been implicated in cell cycle progression and mitosis; however, the complexity of PP2A regulation via multiple B subunits makes its functional characterization a significant challenge. The human adenovirus protein E4orf4 has been found to induce both high Cdk1 activity and the accumulation of cells in G
2
/M in both mammalian and yeast cells, effects which are largely dependent on the B55/Cdc55 regulatory subunit of PP2A. Thus, E4orf4 represents a unique means by which the function of a specific form of PP2A can be delineated
in vivo
. In
Saccharomyces cerevisiae
, only two PP2A regulatory subunits exist, Cdc55 and Rts1. Here, we show that E4orf4-induced toxicity depends on a functional interaction with Cdc55. E4orf4 expression correlates with the inappropriate reduction of Pds1 and Scc1 in S-phase-arrested cells. The unscheduled loss of these proteins suggests the involvement of PP2A
Cdc55
in the regulation of the Cdc20 form of the anaphase-promoting complex (APC). Contrastingly, activity of the Hct1 form of the APC is not induced by E4orf4, as demonstrated by the observed stability of its substrates. We propose that E4orf4, being a Cdc55-specific inhibitor of PP2A, demonstrates the role of PP2A
Cdc55
in regulating APC
Cdc20
activity.
The Interferon-Induced Gene ISG15 Blocks Retrovirus Release from Cells Late in the Budding Process
Authors: Pincetic Andrew , Kuang Zhizhou , Seo Eun Joo , Leis Jonathan 
Published: February 2010

ABSTRACT
The release of retroviruses from cells requires ubiquitination of Gag and recruitment of cellular proteins involved in endosome sorting, including the ESCRT-III proteins and the Vps4 ATPase. In response to infection, cells have evolved an interferon-induced mechanism to block virus replication through expression of the interferon-stimulated gene 15 (ISG15), a dimer homologue of ubiquitin, which interferes with ubiquitin pathways in cells. Previously, it has been reported that ISG15 expression inhibited the E3 ubiquitin ligase, Nedd4, and prevented association of the ESCRT-I protein Tsg101 with human immunodeficiency virus type 1 (HIV-1) Gag. The budding of avian sarcoma leukosis virus and HIV-1 Gag virus-like particles containing L-domain mutations can be rescued by fusion to ESCRT proteins, which cause entry into the budding pathway beyond these early steps. The release of these fusions from cells was susceptible to inhibition by ISG15, indicating that there was a block late in the budding process. We now demonstrate that the Vps4 protein does not associate with the avian sarcoma leukosis virus or the HIV-1 budding complexes when ISG15 is expressed. This is caused by a loss in interaction between Vps4 with its coactivator protein LIP5 needed to promote the formation of the ESCRT-III-Vps4 double-hexamer complex required for membrane scission and virus release. The inability of LIP5 to interact with Vps4 is the probable result of ISG15 conjugation to the ESCRT-III protein, CHMP5, which regulates the availability of LIP5. Thus, there appear to be multiple levels of ISG15-induced inhibition acting at different stages of the virus release process.
Achieving a Golden Mean: Mechanisms by Which Coronaviruses Ensure Synthesis of the Correct Stoichiometric Ratios of Viral Proteins
Authors: Plant Ewan P. , Rakauskaitė Rasa , Taylor Deborah R. , Dinman Jonathan D. 
Published: February 2010

ABSTRACT
In retroviruses and the double-stranded RNA totiviruses, the efficiency of programmed −1 ribosomal frameshifting is critical for ensuring the proper ratios of upstream-encoded capsid proteins to downstream-encoded replicase enzymes. The genomic organizations of many other frameshifting viruses, including the coronaviruses, are very different, in that their upstream open reading frames encode nonstructural proteins, the frameshift-dependent downstream open reading frames encode enzymes involved in transcription and replication, and their structural proteins are encoded by subgenomic mRNAs. The biological significance of frameshifting efficiency and how the relative ratios of proteins encoded by the upstream and downstream open reading frames affect virus propagation has not been explored before. Here, three different strategies were employed to test the hypothesis that the −1 PRF signals of coronaviruses have evolved to produce the correct ratios of upstream- to downstream-encoded proteins. Specifically, infectious clones of the severe acute respiratory syndrome (SARS)-associated coronavirus harboring mutations that lower frameshift efficiency decreased infectivity by >4 orders of magnitude. Second, a series of frameshift-promoting mRNA pseudoknot mutants was employed to demonstrate that the frameshift signals of the SARS-associated coronavirus and mouse hepatitis virus have evolved to promote optimal frameshift efficiencies. Finally, we show that a previously described frameshift attenuator element does not actually affect frameshifting per se but rather serves to limit the fraction of ribosomes available for frameshifting. The findings of these analyses all support a “golden mean” model in which viruses use both programmed ribosomal frameshifting and translational attenuation to control the relative ratios of their encoded proteins.
Two N-Linked Glycosylation Sites in the V2 and C2 Regions of Human Immunodeficiency Virus Type 1 CRF01_AE Envelope Glycoprotein gp120 Regulate Viral Neutralization Susceptibility to the Human Monoclonal Antibody Specific for the CD4 Binding Domain
Authors: Utachee Piraporn , Nakamura Shota , Isarangkura-na-ayuthaya Panasda , Tokunaga Kenzo , Sawanpanyalert Pathom , Ikuta Kazuyoshi , Auwanit Wattana , Kameoka Masanori 
Published: February 2010

ABSTRACT
A recombinant human monoclonal antibody, IgG1 b12 (b12), recognizes a conformational epitope on human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) gp120 that overlaps the CD4 binding domain. Although b12 is able to broadly neutralize HIV-1 subtype B, C, and D viruses, many HIV-1 CRF01_AE viruses are resistant to b12-mediated neutralization. In this report, we examined the molecular mechanisms underlying the low neutralization susceptibility of CRF01_AE viruses to b12, using recently established CRF01_AE Env recombinant viruses. Our results showed that two potential N-linked glycosylation (PNLG) sites in the V2 and C2 regions of Env gp120 played an important role in regulating the susceptibility of CRF01_AE Env to b12. The locations of these PNLG sites correspond to amino acid positions 186 and 197 in HXB2 Env gp120; thus, they are designated N186 and N197 in this study. Removal of N186 significantly conferred the b12 susceptibility of 2 resistant CRF01_AE Env clones, 65CC4 and 107CC2, while the introduction of N186 reduced the b12 susceptibility of a susceptible CRF01_AE Env clone, 65CC1. In addition, removal of both N186 and N197 conferred the b12 susceptibility of 3 resistant CRF01_AE Env clones, 45PB1, 62PL1, and 101PL1, whereas the removal of either N186 or N197 was not sufficient to confer the b12 susceptibility of these CRF01_AE Env clones. Finally, removal of N197 conferred the b12 susceptibility of 2 resistant CRF01_AE Env clones lacking N186, 55PL1 and 102CC2. Taken together, we propose that two PNLG sites, N186 and N197, in Env gp120 are important determinants of the b12 resistance of CRF01_AE viruses.