bluuebunny/crossref_metadata_2025_split huggingface dataset:
Clinical Application of Real-Time PCR to Screening Critically Ill and Emergency-Care Surgical Patients for Methicillin-Resistant
Staphylococcus aureus
: a Quantitative Analytical Study
Authors: Herdman M. Trent , Wyncoll Duncan , Halligan Eugene , Cliff Penelope R. , French Gary , Edgeworth Jonathan D. 
Published: October 2009

ABSTRACT

The clinical utility of real-time PCR screening assays for methicillin (meticillin)-resistant
Staphylococcus aureus
(MRSA) colonization is constrained by the predictive values of their results: as MRSA prevalence falls, the assay's positive predictive value (PPV) drops, and a rising proportion of positive PCR assays will not be confirmed by culture. We provide a quantitative analysis of universal PCR screening of critical care and emergency surgical patients using the BD GeneOhm MRSA PCR system, involving 3,294 assays over six months. A total of 248 PCR assays (7.7%) were positive; however, 88 failed to be confirmed by culture, giving a PPV of 65%. Multivariate analysis was performed to compare PCR-positive culture-positive (P+C+) and PCR-positive culture-negative (P+C−) assays. P+C− results were positively associated with a history of methicillin-sensitive
Staphylococcus aureus
infection or colonization (odds ratio [OR], 3.15; 95% confidence interval [CI], 1.32 to 7.54) and high PCR thresholds of signal intensity, indicative of a low concentration of target DNA (OR, 1.19 per cycle; 95% CI, 1.11 to 1.26). P+C− results were negatively associated with a history of MRSA infection or colonization (OR, 0.19; 95% CI, 0.09 to 0.42) and male sex (OR, 0.40; 95% CI, 0.20 to 0.81). P+C+ patients were significantly more likely to have subsequent positive MRSA culture assays and microbiological evidence of clinical MRSA infection. The risk of subsequent MRSA infection in P+C− patients was not significantly different from that in case-matched PCR-negative controls. We conclude that, given the low PPV and poor correlation between a PCR-positive assay and the clinical outcome, it would be prudent to await culture confirmation before altering infection control measures on the basis of a positive PCR result.
Incubation Alone Is Adequate as a Culturing Technique for Cardiac Rhythm Management Devices
Authors: Viola George M. , Mansouri Mohammad D. , Nasir Nadim , Darouiche Rabih O. 
Published: October 2009

ABSTRACT
There exist no standardized methods for culturing cardiac rhythm management devices. To identify the most optimal culturing method, we compared various techniques that comprise vortex, sonication, and incubation or combinations thereof. Incubation alone yielded bacterial colony counts similar to those of other culturing combinations and is the least labor-intensive.
Evaluation of a New Commercial TaqMan PCR Assay for Direct Detection of the
Clostridium difficile
Toxin B Gene in Clinical Stool Specimens
Authors: Stamper Paul D. , Babiker Wisal , Alcabasa Romina , Aird Deborah , Wehrlin Jennifer , Ikpeama Ijeoma , Gluck Linda , Carroll Karen C. 
Published: October 2009

ABSTRACT

The ProGastro Cd assay (Prodesse, Inc., Waukesha, WI) is a new commercial TaqMan PCR assay that detects
tcdB
. The ProGastro Cd assay was compared to the Wampole
Clostridium difficile
toxin B test (TOX-B test; TechLab, Blacksburg, VA), a cell culture cytotoxicity neutralization assay (CCCNA), and to anaerobic toxigenic bacterial culture, as the “gold standard,” for 285 clinical stool specimens. Assays were independently performed according to manufacturers' directions. A 1.0-ml sample was removed from the stool specimen, of which 20 μl was used for extraction on the NucliSENS easyMAG platform (bioMérieux, Inc., Durham, NC) for the Prodesse ProGastro Cd assay and 200 μl of the stool filtrate was used for the TOX-B CCCNA. Anaerobic toxigenic culture was done by heating an additional 1.0 ml of the stool sample to 80°C for 10 min before inoculation onto modified cycloserine, cefoxitin, and fructose agar with horse blood (Remel, Lenexa, KS) and into a prereduced chopped meat glucose broth (BBL, BD Diagnostics, Sparks, MD). The prevalence of toxin-producing strains of
C. difficile
was 15.7% (
n
= 44) as determined by anaerobic toxigenic culture. The sensitivity, specificity, and positive and negative predictive values of the Prodesse ProGastro Cd assay compared to the TOX-B test were 83.3%, 95.6%, 69.4%, and 98%, respectively. Compared to toxigenic culture, the sensitivity, specificity, and positive and negative predictive values of the Prodesse ProGastro Cd assay were 77.3%, 99.2%, 94.4%, and 95.9%, respectively, and those of the TOX-B test were 63.6%, 99.2%, 93.3%, and 93.6%, respectively. Although no statistical difference (Fisher's exact test) was detected (
P
= 0.242) between the sensitivities of the Prodesse ProGastro Cd assay and a standard CCCNA compared to anaerobic culture for the detection of toxigenic
C. difficile
, the Prodesse ProGastro Cd assay did detect more toxigenic
C. difficile
isolates than the CCCNA.
Comparison of PCR and Culture for Screening of Vancomycin-Resistant Enterococci: Highly Disparate Results for
vanA
and
vanB
Authors: Mak Anton , Miller Mark A. , Chong George , Monczak Yury 
Published: October 2009

ABSTRACT

We compared PCR to conventional culture for the detection of vancomycin-resistant enterococci (VRE) in 30,835 rectal samples over a 3-year period. The positive and negative predictive values of
vanB
PCR were 1.42% and 99.9%, respectively. A positive
vanB
result by PCR is poorly predictive and necessitates culture for differentiation of VRE-positive and -negative individuals.
Evaluation of Adult Chronic Chagas' Heart Disease Diagnosis by Molecular and Serological Methods
Authors: Ramírez Juan David , Guhl Felipe , Umezawa Eufrosina Setsu , Morillo Carlos A. , Rosas Fernando , Marin-Neto Jose A. , Restrepo Silvia 
Published: October 2009

ABSTRACT

Chagas' disease caused by
Trypanosoma cruzi
is endemic in Latin America.
T. cruzi
presents heterogeneous populations and comprises two main genetic lineages, named
T. cruzi
I and
T. cruzi
II. Diagnosis in the chronic phase is based on conventional serological tests, including indirect immunofluorescence (IIF) and enzyme-linked immunosorbent assay (ELISA), and diagnosis in the acute phase based on parasitological methods, including hemoculture. The objective of this study was to evaluate the diagnostic procedures of Chagas' disease in adult patients in the chronic phase by using a PCR assay and conventional serological tests, including TESA-blot as the gold standard. Samples were obtained from 240 clinical chronic chagasic patients. The sensitivities, compared to that of TESA-blot, were 70% for PCR using the kinetoplast region, 75% for PCR using the nuclear repetitive region, 99% for IIF, and 95% for ELISA. According to the serological tests results, we recommend that researchers assess the reliability and sensitivity of the commercial kit Chagatest ELISA recombinant, version 3.0 (Chagatest Rec v3.0; Wiener Lab, Rosario, Argentina), due to the lack of sensitivity. Based on our analysis, we concluded that PCR cannot be validated as a conventional diagnostic technique for Chagas' disease. These data have been corroborated by low levels of concordance with serology test results. It is recommended that PCR be used only for alternative diagnostic support. Using the nuclear repetitive region of
T. cruzi
, PCR could also be applicable for monitoring patients receiving etiologic treatment.
Differential Specificity and Immunogenicity of Adenovirus Type 5 Neutralizing Antibodies Elicited by Natural Infection or Immunization
Authors: Cheng Cheng , Gall Jason G. D. , Nason Martha , King C. Richter , Koup Richard A. , Roederer Mario , McElrath M. Juliana , Morgan Cecilia A. , Churchyard Gavin , Baden Lindsey R. , Duerr Ann C. , Keefer Michael C. , Graham Barney S. , Nabel Gary J. 
Published: October 2009

ABSTRACT
A recent clinical trial of a T-cell-based AIDS vaccine delivered with recombinant adenovirus type 5 (rAd5) vectors showed no efficacy in lowering viral load and was associated with increased risk of human immunodeficiency virus type 1 (HIV-1) infection. Preexisting immunity to Ad5 in humans could therefore affect both immunogenicity and vaccine efficacy. We hypothesized that vaccine-induced immunity is differentially affected, depending on whether subjects were exposed to Ad5 by natural infection or by vaccination. Serum samples from vaccine trial subjects receiving a DNA/rAd5 AIDS vaccine with or without prior immunity to Ad5 were examined for the specificity of their Ad5 neutralizing antibodies and their effect on HIV-1 immune responses. Here, we report that rAd5 neutralizing antibodies were directed to different components of the virion, depending on whether they were elicited by natural infection or vaccination in HIV vaccine trial subjects. Neutralizing antibodies elicited by natural infection were directed largely to the Ad5 fiber, while exposure to rAd5 through vaccination elicited antibodies primarily to capsid proteins other than fiber. Notably, preexisting immunity to Ad5 fiber from natural infection significantly reduced the CD4 and CD8 cell responses to HIV Gag after DNA/rAd5 vaccination. The specificity of Ad5 neutralizing antibodies therefore differs depending on the route of exposure, and natural Ad5 infection compromises Ad5 vaccine-induced immunity to weak immunogens, such as HIV-1 Gag. These results have implications for future AIDS vaccine trials and the design of next-generation gene-based vaccine vectors.
Andes Virus Recognition of Human and Syrian Hamster β
3
Integrins Is Determined by an L33P Substitution in the PSI Domain
Authors: Matthys Valery S. , Gorbunova Elena E. , Gavrilovskaya Irina N. , Mackow Erich R. 
Published: October 2009

ABSTRACT

Andes virus (ANDV) causes a fatal hantavirus pulmonary syndrome (HPS) in humans and Syrian hamsters. Human α
v
β
3
integrins are receptors for several pathogenic hantaviruses, and the function of α
v
β
3
integrins on endothelial cells suggests a role for α
v
β
3
in hantavirus directed vascular permeability. We determined here that ANDV infection of human endothelial cells or Syrian hamster-derived BHK-21 cells was selectively inhibited by the high-affinity α
v
β
3
integrin ligand vitronectin and by antibodies to α
v
β
3
integrins. Further, antibodies to the β
3
integrin PSI domain, as well as PSI domain polypeptides derived from human and Syrian hamster β
3
subunits, but not murine or bovine β
3
, inhibited ANDV infection of both BHK-21 and human endothelial cells. These findings suggest that ANDV interacts with β
3
subunits through PSI domain residues conserved in both Syrian hamster and human β
3
integrins. Sequencing the Syrian hamster β
3
integrin PSI domain revealed eight differences between Syrian hamster and human β
3
integrins. Analysis of residues within the PSI domains of human, Syrian hamster, murine, and bovine β
3
integrins identified unique proline substitutions at residues 32 and 33 of murine and bovine PSI domains that could determine ANDV recognition. Mutagenizing the human β
3
PSI domain to contain the L33P substitution present in bovine β
3
integrin abolished the ability of the PSI domain to inhibit ANDV infectivity. Conversely, mutagenizing either the bovine PSI domain, P33L, or the murine PSI domain, S32P, to the residue present human β
3
permitted PSI mutants to inhibit ANDV infection. Similarly, CHO cells transfected with the full-length bovine β
3
integrin containing the P33L mutation permitted infection by ANDV. These findings indicate that human and Syrian hamster α
v
β
3
integrins are key receptors for ANDV and that specific residues within the β
3
integrin PSI domain are required for ANDV infection. Since L33P is a naturally occurring human β
3
polymorphism, these findings further suggest the importance of specific β
3
integrin residues in hantavirus infection. These findings rationalize determining the role of β
3
integrins in hantavirus pathogenesis in the Syrian hamster model.
Modulation of Hepatitis C Virus RNA Abundance and the Isoprenoid Biosynthesis Pathway by MicroRNA miR-122 Involves Distinct Mechanisms
Authors: Norman Kara L. , Sarnow Peter 
Published: October 2009

ABSTRACTMicroRNA 122 (miR-122) promotes hepatitis C virus (HCV) RNA abundance through a direct interaction with the viral RNA and stimulates the mevalonate pathway in the animal liver. We found that overexpression of miR-122 enhanced viral RNA accumulation without affecting genes in the mevalonate pathway, such as the 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) gene. However, inhibition of miR-122 decreased both HCV RNA and HMGCR RNA with little effects on the rates of HCV and HMGCR RNA synthesis. Loss of HCV RNA could not be restored by isoprenoid intermediate metabolites. Overall, these findings suggest that miR-122 modulates viral RNA abundance independently of its effect on isoprenoid metabolism.
Hepatitis B Virus Requires Intact Caveolin-1 Function for Productive Infection in HepaRG Cells
Authors: Macovei Alina , Radulescu Cristina , Lazar Catalin , Petrescu Stefana , Durantel David , Dwek Raymond A. , Zitzmann Nicole , Nichita Norica Branza 
Published: October 2009

ABSTRACTInvestigation of the entry pathways of hepatitis B virus (HBV), a member of the familyHepadnaviridae, has been hampered by the lack of versatile in vitro infectivity models. Most concepts of hepadnaviral infection come from the more robust duck HBV system; however, whether the two viruses use the same mechanisms to invade target cells is still a matter of controversy. In this study, we investigate the role of an important plasma membrane component, caveolin-1 (Cav-1), in HBV infection. Caveolins are the main structural components of caveolae, plasma membrane microdomains enriched in cholesterol and sphingolipids, which are involved in the endocytosis of numerous ligands and complex signaling pathways within the cell. We used the HepaRG cell line permissive for HBV infection to stably express dominant-negative Cav-1 and dynamin-2, a GTPase involved in vesicle formation at the plasma membrane and other organelles. The endocytic properties of the newly established cell lines, designated HepaRGCav-1, HepaRGCav-1Δ1-81, HepaRGDyn-2, and HepaRGDyn-2K44A, were validated using specific markers for different entry routes. The cells maintained their properties during cell culture, supported differentiation, and were permissive for HBV infection. The levels of both HBV transcripts and antigens were significantly decreased in cells expressing the mutant proteins, while viral replication was not directly affected. Chemical inhibitors that specifically inhibit clathrin-mediated endocytosis had no effect on HBV infection. We concluded that HBV requires a Cav-1-mediated entry pathway to initiate productive infection in HepaRG cells.
Effect of Killer Immunoglobulin-Like Receptors in the Response to Combined Treatment in Patients with Chronic Hepatitis C Virus Infection
Authors: Vidal-Castiñeira J. R. , López-Vázquez A. , Díaz-Peña R. , Alonso-Arias R. , Martínez-Borra J. , Pérez R. , Fernández-Suárez J. , Melón S. , Prieto J. , Rodrigo L. , López-Larrea C. 
Published: October 2009

ABSTRACTKiller immunoglobulin-like receptors (KIRs) are related to the activation and inhibition of NK cells and may play an important role in the innate response against infection with viruses such as hepatitis C virus (HCV). We examined whether the different combinations of KIRs with their HLA class I ligands influenced the response to combined treatment (pegylated alpha interferon and ribavirin) of patients infected by HCV. A total of 186 consecutive patients diagnosed with chronic HCV infection were analyzed. Seventy-seven patients exhibited HCV RNA levels at 6 months posttreatment and were called nonresponders (NR), while 109 cleared viral RNA and were named sustained viral responders (SVR). Patients were typed for HLA-B, HLA-Cw, KIR genes, and HCV genotype. In our study, the frequency of the KIR2DL2 allele was significantly increased in NR (P< 0.001; odds ratio [OR] = 1.95), as was the frequency of the KIR2DL2/KIR2DL2 genotype (P< 0.005; OR = 2.52). In contrast, the frequencies of the KIR2DL3 genotype (P< 0.001) and KIR2DL3/KIR2DL3 genotype (P< 0.05; OR = 0.54) were significantly increased in the SVR. Different combinations of KIR2DL2 and KIR2DL3 alleles with their ligands were analyzed. The frequency of the KIR2DL2/KIR2DL2-HLA-C1C2 genotype was significantly increased in the NR (P< 0.01; OR = 3.15). Additionally, we found a higher frequency of the KIR2DL3/KIR2DL3-HLA-C1C1 genotype in the SVR group (P< 0.05; OR = 0.33). These results were not affected by the HCV genotype. In conclusion, patients who carried the KIR2DL2/KIR2DL2-HLA-C1C2 genotype were less prone to respond to treatment. However, the KIR2DL3/KIR2DL3-HLA-C1C1 genotype clearly correlated with a satisfactory response to treatment, defined by the clearance of HCV RNA.
Novel Approach to the Formulation of an Epstein-Barr Virus Antigen-Based Nasopharyngeal Carcinoma Vaccine
Authors: Lutzky Viviana P. , Corban Monika , Heslop Lea , Morrison Leanne E. , Crooks Pauline , Hall David F. , Coman William B. , Thomson Scott A. , Moss Denis J. 
Published: October 2009

ABSTRACTEpstein-Barr virus (EBV) is associated with several malignant diseases including nasopharyngeal carcinoma (NPC), a common neoplasm throughout southeast Asia. Radiotherapy and chemotherapy can achieve remission, but a reemergence of disease is not uncommon. Therefore, there is a need for specific therapies that target the tumor through the recognition of EBV antigens. In NPC, latent membrane protein 1 (LMP1) and LMP2 offer the best opportunity for specific targeting since they are typically expressed and T-cell determinants in each of these proteins have been defined. We have attempted to maximize the opportunity of incorporating every possible CD4 and CD8 determinant in a single formulation. We have achieved this by generating a scrambled protein incorporating random overlapping peptide sets from EBNA1, LMP1, and LMP2, which was then inserted into a replication-deficient strain of adenovirus (adenovirus scrambled antigen vaccine [Ad-SAVINE]). This report describes the construction of this Ad-SAVINE construct, its utility in generating LMP1 and LMP2 responses in healthy individuals as well as NPC patients, and its capacity to define new epitopes. This formulation could have a role in NPC immunotherapy for all ethnic groups since it has the potential to activate all possible CD4 and CD8 responses within EBNA1 and LMPs.
Effects of Phosphorylation of Herpes Simplex Virus 1 Envelope Glycoprotein B by Us3 Kinase In Vivo and In Vitro
Authors: Imai Takahiko , Sagou Ken , Arii Jun , Kawaguchi Yasushi 
Published: October 2009

ABSTRACTWe recently reported that the herpes simplex virus 1 (HSV-1) Us3 protein kinase phosphorylates threonine at position 887 (Thr-887) in the cytoplasmic tail of envelope glycoprotein B (gB) (A. Kato, J. Arii, I. Shiratori, H. Akashi, H. Arase, and Y. Kawaguchi, J. Virol. 83:250-261, 2009; T. Wisner, C. C. Wright, A. Kato, Y. Kawaguchi, F. Mou, J. D. Baines, R. J. Roller and D. C. Johnson, J. Virol. 83:3115-3126, 2009). In the studies reported here, we examined the effect(s) of this phosphorylation on viral replication and pathogenesis in vivo and present data showing that replacement of gB Thr-887 by alanine significantly reduced viral replication in the mouse cornea and development of herpes stroma keratitis and periocular skin disease in mice. The same effects have been reported for mice infected with a recombinant HSV-1 carrying a kinase-inactive mutant of Us3. These observations suggested that Us3 phosphorylation of gB Thr-887 played a critical role in viral replication in vivo and in HSV-1 pathogenesis. In addition, we generated a monoclonal antibody that specifically reacted with phosphorylated gB Thr-887 and used this antibody to show that Us3 phosphorylation of gB Thr-887 regulated subcellular localization of gB, particularly on the cell surface of infected cells.
Pathogenesis of Hepatitis C Virus Infection inTupaia belangeri
Authors: Amako Yutaka , Tsukiyama-Kohara Kyoko , Katsume Asao , Hirata Yuichi , Sekiguchi Satoshi , Tobita Yoshimi , Hayashi Yukiko , Hishima Tsunekazu , Funata Nobuaki , Yonekawa Hiromichi , Kohara Michinori 
Published: October 2009

ABSTRACTThe lack of a small-animal model has hampered the analysis of hepatitis C virus (HCV) pathogenesis. The tupaia (Tupaia belangeri), a tree shrew, has shown susceptibility to HCV infection and has been considered a possible candidate for a small experimental model of HCV infection. However, a longitudinal analysis of HCV-infected tupaias has yet to be described. Here, we provide an analysis of HCV pathogenesis during the course of infection in tupaias over a 3-year period. The animals were inoculated with hepatitis C patient serum HCR6 or viral particles reconstituted from full-length cDNA. In either case, inoculation caused mild hepatitis and intermittent viremia during the acute phase of infection. Histological analysis of infected livers revealed that HCV caused chronic hepatitis that worsened in a time-dependent manner. Liver steatosis, cirrhotic nodules, and accompanying tumorigenesis were also detected. To examine whether infectious virus particles were produced in tupaia livers, naive animals were inoculated with sera from HCV-infected tupaias, which had been confirmed positive for HCV RNA. As a result, the recipient animals also displayed mild hepatitis and intermittent viremia. Quasispecies were also observed in the NS5A region, signaling phylogenic lineage from the original inoculating sequence. Taken together, these data suggest that the tupaia is a practical animal model for experimental studies of HCV infection.
Role of a Putative gp41 Dimerization Domain in Human Immunodeficiency Virus Type 1 Membrane Fusion
Authors: Liu Jie , Deng Yiqun , Li Qunnu , Dey Antu K. , Moore John P. , Lu Min 
Published: October 2009

ABSTRACT
The entry of human immunodeficiency virus type 1 (HIV-1) into a target cell entails a series of conformational changes in the gp41 transmembrane glycoprotein that mediates the fusion of the viral and target cell membranes. A trimer-of-hairpins structure formed by the association of two heptad repeat (HR) regions of the gp41 ectodomain has been implicated in a late step of the fusion pathway. Earlier native and intermediate states of the protein are postulated to mediate the antiviral activity of the fusion inhibitor enfuvirtide and of broadly neutralizing monoclonal antibodies (NAbs), but the details of these structures remain unknown. Here, we report the identification and crystal structure of a dimerization domain in the C-terminal ectodomain of gp41 (residues 630 to 683, or C54). Two C54 monomers associate to form an asymmetric, antiparallel coiled coil with two distinct C-terminal α-helical overhangs. This dimer structure is conferred largely by interactions within a central core that corresponds to the sequence of enfuvirtide. The mutagenic alteration of the dimer interface severely impairs the infectivity of Env-pseudotyped viruses. Moreover, the C54 structure binds tightly to both the 2F5 and 4E10 NAbs and likely represents a potential intermediate conformation of gp41. These results should enhance our understanding of the molecular basis of the gp41 fusogenic structural transitions and thereby guide rational, structure-based efforts to design new fusion inhibitors and vaccine candidates intended to induce broadly neutralizing antibodies.