bluuebunny/crossref_metadata_2025 huggingface dataset:
Outbreak of Scrub Typhus in Southern India during the Cooler Months
Authors: MATHAI E. , ROLAIN J. M. , VERGHESE G. M. , ABRAHAM O. C. , MATHAI D. , MATHAI M. , RAOULT D.
Published: July 2009

Abstract: Orientia tsutsugamushi, the agent of scrub typhus, is a strict intracellular bacterium which is found in many parts of Asia including India. During the past few years, the number of patients with rickettsial infection and scrub typhus has increased, especially during the cooler months. We report in this study a recent outbreak of scrub typhus recorded during the cooler months (October 2001 to February 2002) in patients admitted to our hospital with acute febrile illness associated with diverse signs and symptoms. Overall, 28 patients were clinically and serologically confirmed to have scrub typhus. Fever for more than one week was the only common manifestation. Myalgias was the next most common feature (52%), and rash was observed in only 22% of the cases. Seventeen patients treated with doxycycline recovered in 1 to 3 days, as well as two patients who received chloramphenicol. In five patients who received ciprofloxacin, fever subsided only after five days. Finally three patients (10.7%) died, including one patient treated with doxycycline. These data indicate that scrub typhus is a reemerging infectious disease in India with a possibility of drug resistance. This reemergence emphasizes the need for further prospective studies to design effective control measures.
Cat Scratch Disease with Lymphadenitis, Vertebral Osteomyelitis, and Spleen Abscesses
Authors: ROLAIN J. M. , CHANET V. , LAURICHESSE H. , LEPIDI H. , BEYTOUT J. , RAOULT D.
Published: July 2009

Abstract: In this report we describe a 30‐year old male patient with vertebral osteomyelitis and spleen abscesses with cat scratch disease. The diagnosis was made on the basis of molecular detection of Bartonella henselae either on lymph node biopsies or on bone biopsy, histology of the lymph node, serology using either our in‐house microimmunofluorescence assay or a commercial kit (Focus Technologies). Immunofluorescent detection was also performed directly on slide appositions using a monoclonal antibody. Treatment consisted of administration of antibiotics with rapid clinical improvement and a stabilization of skeletal lesions on the magnetic resonance imaging performed three months later. Twenty two other cases of this unusual manifestation associated with cat scratch disease have been reported in the literature and are reviewed here. Our case represents the second case of osteomyelitis associated with cat scratch disease in which B. henselae has been specifically identified as the etiological agent using several direct and indirect methods.
Rickettsia rickettsii Infection in the Pine Vole, Microtus pinetorum
Authors: EREMEEVA MARINA E. , LIANG ZHONGXING , PADDOCK CHRISTOPHER , ZAKI SHERIF , VANDENBERGH JOHN G. , DASCH GREGORY A. , SILVERMAN DAVID J.
Published: July 2009

Abstract: The pine vole, Microtus pinetorum, was evaluated as a laboratory animal model for infection with Rickettsia rickettsii. Voles demonstrated signs of acute disease, and 45% of infected animals died following intraperitoneal infection with 3 × 106 plaque forming units of R. rickettsii. Spleen, liver, kidney, lung, brain, testes and blood were analyzed for rickettsial burden by a quantitative PCR assay. The distribution of rickettsiae in tissues during the course of infection was determined by immunohistochemical staining and pathological changes in tissues were correlated with the clinical severity of infection. Quantitative RT‐PCR assays were designed to measure the mRNA levels of the antioxidant enzyme genes for catalase, glutathione peroxidase, glutathione reductase, heme oxygenase, Cu‐Zn superoxide dismutase (SOD) and Mn‐SOD, and 2 housekeeping genes, actin and glyceraldehyde phosphate dehydrogenase. Tissues from acutely ill animals on days 2 to 6 of infection, convalescent animals, and uninfected control animals were studied. The number of transcripts of each enzyme gene was determined and compared to the degree of rickettsial infection present. These studies demonstrate that the pine vole is a valuable experimental model for studying infection with R. rickettsii. Our results provide the first experimental evidence that R. rickettsii causes alteration(s) of the anti‐oxidant system in vivo.
Development of Improved Vaccines for Heartwater
Authors: COLLINS NICOLA E. , PRETORIUS ALRI , VAN KLEEF MIRINDA , BRAYTON KELLY A. , ZWEYGARTH ERICH , ALLSOPP BASIL A.
Published: July 2009

Abstract: Heartwater is a tick‐borne disease of ruminants which causes major economic losses for domestic livestock owners throughout sub‐Saharan Africa and the Caribbean. It is caused by the intracellular rickettsia Ehrlichia (formerly Cowdria) ruminantium and the only commercially available vaccination procedure is over 50 years old. It involves infecting animals with cryopreserved sheep blood containing virulent E. ruminantium organisms, followed by antibiotic treatment when fever develops. Experimental attenuated, inactivated, and nucleic acid vaccine procedures have been investigated over the last half‐century, but none of them has yet been particularly successful. We have developed two new experimental vaccines, a live attenuated vaccine and a nucleic acid vaccine. The attenuated vaccine was developed by continuous passage of E. ruminantium organisms of the virulent Welgevonden isolate in a continuous canine macrophage‐monocyte cell line. After more than 50 passages the cultures produced no disease when inoculated into mice or sheep, and the inoculated animals were 100% immune to a subsequent lethal homologous needle challenge. The nucleic acid vaccine is based on four E. ruminantium genes from a genetic locus involved in nutrient transport. A cocktail of all four genes, cloned in a DNA vaccine vector and used to immunize sheep, engendered 100% protection against a subsequent lethal needle challenge with the homologous isolate and with each of five different virulent heterologous isolates. Sheep immunized with this cocktail were also exposed to a field challenge in a heartwater‐endemic area and few animals survived. This suggests that the local E. ruminantium genotypes were different from any which were administered by needle challenge, or that needle challenge is not a good model for tick challenge in the field.
Immunogenicity of a 40kDa Fragment of the 47kDa Recombinant Protein and DNA Vaccine from Karp Strain of Orientia tsutsugamushi
Authors: NIU DONGSHENG , CHEN WEIJUN , ZHANG XUEYING , CHEN MEILING , CUI HONG , WEI WENJIN , WEN BOHAI , CHEN XIANGRUI
Published: July 2009

Abstract: In this study, the fragment of 47 kDa gene (301 bp‐1428 bp) was cloned into a prokaryotic expression vector pBV220 to construct a recombinant plasmid pBV‐47. The E. coli cells were transformed with pBV‐47 and the transformants were induced to express the recombinant protein at 42°C. The expression product (40 kDa) was detected by SDS‐PAGE analysis and the 40kDa protein was recognized by mouse polyclonal antibodies against O. tsutsugamushi Karp strain in western blot analysis. The entire 47 kDa protein gene was inserted into an eukaryotic expression vector pcDNA3.1(+) to construct a recombinant plasmid pcDNA3.1/47 and Balb/c mice were immunized with recombinant pcDNA3.1/47, control vector pcDNA3.1, PBS buffer, 40 kDa protein, and recombinant pcDNA3.1/47 plus 40 kDa protein (pcDNA3.1/47/40), respectively. The results showed that spleen cells from pcDNA3.1/47/40‐immunized mice gave higher proliferation than other groups. A significant IgG rise was detected in mice immunized with 40 kDa protein, but it was less strong than that in mice immunized with pcDNA3.1/47/40. The results suggested that immunization with pcDNA3.1/47 and 40 kDa protein simultaneously could induce a strong immune response.
Experimental Infection of Human Erythrocytes from Alcoholic Patients with Bartonella quintana
Authors: ROLAIN JEAN‐MARC , ARNOUX D. , PARZY D. , SAMPOL J. , RAOULT DIDIER
Published: July 2009

Abstract: Bartonella spp. are found in the erythrocytes of their specific natural hosts and B. quintana bacteremia is associated epidemiologically with lice, alcoholism, and homelessness. The aim of our study was to compare the growth and the number of bacteria per erythrocyte in vitro in laboratory‐infected red blood cells from alcoholic patients versus normal blood donor erythrocytes. Enumeration of bacteria was performed either with plate counting or with a real‐time PCR quantitative assay. Number of bacteria per cell was determined using immunofluorescence assay and laser confocal microscopy. Although the number of bacteria after 4 days of incubation was similar in the two groups of erythrocytes, we found that the distribution of bacteria per erythrocyte in the two groups was different. Erythrocytes from alcoholics contain significantly more bacteria per cell than erythrocytes from blood donors. Our results suggest that there is a link between alcoholism and infections of B. quintana that may be due to the macrocytosis of erythrocytes.
An Experimental Model of Human Body Louse Infection with Rickettsia typhi
Authors: HOUHAMDI LINDA , FOURNIER PIERRE‐EDOUARD , FANG RONG , RAOULT DIDIER
Published: July 2009

Abstract: Murine (endemic) typhus caused by Rickettsia typhi, one of the most widely distributed arthropod‐borne diseases, is transmitted to humans mainly by the oriental rat flea. The human body louse, Pediculus humanus corporis, has been suspected to have a role in the transmission of R. typhi to humans. To evaluate the potential role of Pediculus humanus corporis as a vector of murine typhus, we used R. typhi in an experimental model of human body louse infection previously used with R. prowazekii. A rabbit was made bacteremic by inoculating it with 2 × 106 plaque‐forming units of R. typhi; it remained bacteremic for at least 59 hours. Two hundred body lice infected by feeding on the bacteremic rabbit were compared to 200 uninfected control lice. Each louse population was fed once a day on the abdomen of a seronegative rabbit. On day 8 post‐infection, as a result of disruption of the gut cells and leakage of the blood meal into the hemolymph, four infected lice became bright red and died within four hours. The life span of infected lice was 20 days less than that of the controls. Infected lice did not transmit R. typhi to their progeny (eggs and larvae) as demonstrated by PCR amplification and cell culture. With an immunofluorescence assay, R. typhi was detected in feces from day 7 post‐infection, and the organism remained viable in feces for up to 80 days as demonstrated by cell culture. From the 21st day post‐infection, the rabbit used to feed the R. typhi‐infected lice developed an immunoglobulin response with a titer of 1:50 increasing to 1:200 at the 42nd day post‐infection. It showed no clinical signs of infection. The rabbit that was used to feed uninfected lice remained seronegative. Although body lice are not clearly identified vectors of R. typhi, it seems that under certain circumstances they could transmit R. typhi.
Novel Immunoreactive Glycoprotein Orthologs of Ehrlichia spp.
Authors: McBRIDE JERE W. , COMER JASON E. , WALKER DAVID H.
Published: July 2009

Abstract: We previously identified a strongly immunoreactive 43 kDa protein (p43) of Ehrlichia canis. As an immunodiagnostic antigen, the p43 had a 96% accuracy as compared with IFA and provided species‐specific diagnosis of E. canis infections. Further investigation has revealed that the E. canis p43 represents the N‐terminal portion of the largest immunoreactive protein described in Ehrlichia spp. with a predicted molecular mass of 153 kDa. Analysis of the recombinant N‐terminal region (p43) of the p153 by protein gel electrophoresis demonstrated a larger than predicted molecular mass (∼30%), and presence of carbohydrate glycans, indicating that the p153 is a glycoprotein. A BLASTn search was performed on the E. chaffeensis genome sequence (95%), and the gene encoding the p153 ortholog was identified in E. chaffeensis. The E. canis p153 (4,263 bp) and E. chaffeensis p156 (4,389 bp) genes had similar chromosomal locations, downstream of the homologous (∼87%) deoxyguanosine‐triphosphate triphosphohydrolase genes, and homologous (∼90%) intergenic sequences preceding the open reading frames. Nucleic acid sequence homology (52%) observed between the glycoprotein genes supported previous findings with regard to genetic divergence of the p43 gene fragment, and the p153 and p156 proteins had amino acid similarity of 32%. A native E. canis protein with a molecular mass of 200 kDa reacted with antisera produced against the N‐terminal region (p43) of the p153, suggesting that the native protein was posttranslationally modified. Similarly, recombinant constructs of E. chaffeensis p156 migrated larger than predicted (∼200 kDa), and carbohydrate was detected on the recombinant proteins. The chromosomal location, amino acid homology, and biophysical properties support the conclusion that the p153 and p156 glycoproteins (designated gp200s) are species‐specific immunoreactive orthologs.
rRNA Intergenic Spacer Regions for Phylogenetic Analysis of Rickettsia Species
Authors: VITORINO LILIANA , ZÉ‐ZÉ LÍBIA , SOUSA ANA , BACELLAR FATIMA , TENREIRO ROGÉRIO
Published: July 2009

Abstract: Species of the genus Rickettsia are responsible for several human diseases, namely epidemic typhus, Rocky Mountain spotted fever, and tick‐borne typhus transmitted by arthropod vectors. The rrl‐rrf intergenic spacer region (rrl‐rrf ITS) was sequenced for 12 Rickettsia strains, including R. typhi, 6 untested species, R. aeschlimannii, Bar29, R. helvetica, R. honei, R. massilae, and R. slovaca as well as 5 Portuguese isolates. Phylogenetic trees inferred from rrl‐rrf spacer sequences using maximum‐parsimony and distance methods provided largely congruent tree topologies, supported by significant bootstrap values, enabling the identification of five distinct rickettsiae clusters.
Effect of Ethanol on Lipid‐Mediated Transfection of Primary Cortical Neurons
Authors: ANJI ANTJE , SHAIK KAMRAN A. , KUMARI MEENA
Published: July 2009

Abstract: Successful introduction of nucleic acids into mammalian neurons has revolutionized the analyses of gene regulation and cellular function. Various methods, including viral infection, have been developed to introduce plasmid DNA into primary neuronal cultures. However, transfection of primary cultures of neurons using the calcium phosphate precipitation method and electroporation have been comparatively inefficient. In this paper, we describe a method to successfully transfect cultured fetal cortical neurons using a cationic lipid reagent, lipofectamine. Cells were cultured in the absence and presence of 50 mM ethanol. To monitor transfection of neurons, we employed three mammalian expression vectors containing Renilla luciferase and/or firefly luciferase, or the beta‐galactosidase reporter gene. Fetal cortical neurons were isolated and cultured in the absence or presence of 50 mM ethanol, for two days. On day 3, neurons were washed, fed with serum‐free medium, and transfected with the DNA‐lipofectamine complex. After two hours, cells were washed, fed complete medium lacking or containing 50 mM ethanol and cultured for two additional days with a change of medium after 24 h. Cultures were terminated 48 h after transfection. Cells were either stained for beta‐galactosidase activity using X‐gal or lysed to prepare cell extracts to assay for luciferase activity using a luminometer. When neurons were cotransfected, Renilla luciferase was used as an internal control to normalize the expression of the firefly luciferase reporter gene. Analysis of results showed that expression of the reporter gene, firefly luciferase, was approximately 2.5 times greater in ethanol treated neuronal cultures than for neurons cultured in the absence of ethanol. An increased number of neurons expressing beta‐galactosidase was also observed in ethanol‐treated neurons. These data suggest that perhaps ethanol treatment of fetal cortical neurons improved the DNA uptake and/or increased the expression of the reporter genes.
Prediction of Experimental Data for an Independent Variable Using the Experimental Data Collected for Other Independent Variables in a Study of Skin Cancer Caused by Exposure to UV Radiation
Authors: HASHEMI RAY R. , BAHAR MAHMOOD , TANG NAN , TYLER ALEXANDER A. , HINSON WILLIAM
Published: July 2009

Abstract: In this study, two algorithms (ONE and TWO) are introduced to determine the position of the t‐distribution of variable Vi (with 95% confidence) in the treated group in reference to the t‐distribution of variable Vi (with 95% confidence) in the control group of an experimental study involving UV radiation exposure of a group of rodents. The outcome of applying the two algorithms is two discretized files. A reduct of each file is generated using the rough sets methodology and then the measurements for one independent variable are predicted using the measurements of the other independent variables in the same reduct. The rough sets methodology and the fuzzy‐rough classifier are used for this prediction. The results reveal that (1) algorithm TWO is the best, (2) the values for non‐core variables are predicted with minimum accuracy of 87%, and (3) the prediction of values for core variables is not successful.
Possible Mechanism for the Neuroprotective Effects of l‐Carnitine on Methamphetamine‐Evoked Neurotoxicity
Authors: VIRMANI ASHRAF , GAETANI FRANCO , IMAM SYED , BINIENDA ZBIGNIEW , ALI SYED
Published: July 2009

Abstract:Some of the damage to the CNS that is observed following amphetamine and methamphetamine (METH) administration is known to be linked to increased formation of free radicals. This increase could be, in part, related to mitochondrial dysfunction and/or cause damage to the mitochondria, thereby leading to a failure of cellular energy metabolism and an increase in secondary excitotoxicity. The actual neuronal damage that occurs with METH‐induced toxicity seems to affect dopaminergic cells in particular. METH‐induced toxicity is related to an increase in the generation of both reactive oxygen (hydroxyl, superoxide, peroxide) and nitrogen (nitric oxide) species. Peroxynitrite (ONOO−), which is a reaction product of either superoxide or nitric oxide, is the most damaging radical. It can be reduced by antioxidants such as selenium, melatonin, and the selective nNOS inhibitor, 7‐nitroindazole. METH‐induced toxicity has been previously shown to increase production of the peroxynitrite stress marker, 3‐nitrotyrosine (3‐NT),in vitro, in cultured PC12 cells, and alsoin vivo, in the striatum of adult male mice. Pre‐ and post‐treatment of mice with l‐carnitine (LC) significantly attenuated the production of 3‐NT in the striatum after METH exposure. LC is a mitochondriotropic compound in that it carries long‐chain fatty acyl groups into mitochondria for β‐oxidation. It was shown also to play a protective role against various mitochondrial toxins, such as 3‐nitropropionic acid. The protective effects of LC against METH‐induced toxicity could be related to its prevention of possible metabolic compromise produced by METH and the resulting energy deficits. In particular, LC may be maintaining the mitochondrial permeability transition (MPT) and modulating the activation of the mitochondrial permeability transition pores (mPTP), especially the cyclosporin‐dependent mPTP. The possible neuroprotective mechanism of LC against METH‐toxicity and the role of the mitochondrial respiratory chain and the generation of free radicals and their subsequent action on the MPT and mPTP are also being examined using anin vitromodel of NGF‐differentiated pheochromocytoma cells (PC12). In preliminary experiments, the pretreatment of PC12 cells with LC (5 mM), added 10 min before METH (500 μM), indicated that LC enhances METH‐induced DA depletion. The role of LC in attenuating METH‐evoked toxicity is still under investigation and promises to reveal information regarding the underlying mechanisms and role of mitochondria in the triggering of cell death.
Prostaglandins of J Series Control Heme Oxygenase Expression
Authors: ZHUANG HEAN , KIM YUN‐SOOK , NAMIRANIAN KHODADAD , DORÉ SYLVAIN
Published: July 2009

Abstract:Cyclopentenone prostaglandins (cyPGs) are a subfamily of prostaglandins that are characterized by the cyclopentenone ring in their structure. They exert their effect after active transportation into the cell, probably by interacting with cellular target proteins or DNA sequences. The cyPGs have anti‐inflammatory activities, especially important during the resolution of inflammation, anticancer, and cytoprotective properties. Here, we show that the cyPGs, especially the 15‐deoxy‐Δ12,14PGJ2, can specifically induce heme oxygenase 1 in mouse primary neuronal cells. Heme oxygenase is the enzyme responsible for the degradation of heme into biliverdin, ferrous iron, and carbon monoxide. This enzyme conveys protection to oxidative cellular injury by degrading the pro‐inflammatory heme; producing biliverdin and bilirubin, potent antioxidants; producing carbon monoxide, a neurotransmitter that also has anti‐inflammatory and vasodilatory properties; and assisting in keeping iron cellular homeostasis. CyPGs appear to possess a promising future in designing therapeutics for many neurologic diseases, such as Alzheimer's disease, vascular‐related dementia, multiple sclerosis, ischemic conditions, and many others in which inflammation is a part of the pathophysiology.
Differential Effects of Lipopolysaccharide on Lipid Peroxidation in F344N, SHR Rats and BALB/c Mice, and Protection of Melatonin and NAS against Its Toxicity
Authors: REQUINTINA PURA J. , OXENKRUG GREGORY F.
Published: July 2009

Abstract: The effect of bacterial lipopolysaccharide (LPS) injection on the lipid peroxidation process in Fischer (F344N) rats, spontaneously hypertensive (SHR) rats, and BALB/c mice was studied. Lipid peroxidation, as measured by malondialdehyde + 4‐hydroxyalkenals (MDA + HAE) levels, was decreased in brain, kidney, and liver homogenates of F344N rats injected with lower LPS doses of 0.5, 1.0, and 2.0 mg/kg, but was increased with the highest dose of 10 mg/kg body weight. The dose of 10 mg/kg LPS decreased the MDA + HAE levels in SHR brain homogenates and increased levels in the liver homogenates. MDA + HAE levels in the brain and liver but not kidney homogenates in BALB/c mice also increased after administration of LPS at the highest dose (10 mg/kg body weight). The effect of melatonin, N‐acetylserotonin (NAS), and GR‐135,531 (a melatonin ligand with high affinity for MT3 receptor) on the survival of BALB/c mice injected with lethal dose of LPS was also tested. A single dose of 5 mg/kg of melatonin or NAS simultaneously injected with LPS (25 mg/kg body weight) markedly protected mice from the lethal effect of LPS with survival rates of 90% and 95% for melatonin and NAS, respectively, and 59% for mice injected with just LPS after 24 hours; a survival rate of 50% for both melatonin and NAS, and 32% was obtained for mice injected with just LPS after five days. GR‐135,531 did not show protection against a lethal dose of LPS. Our results indicated that the effect of LPS on lipid peroxidation is dose‐, time‐, and species‐dependent, and that melatonin and NAS are equally effective in protecting mice from lethality caused by LPS.
Gene and Protein Expression Profiles of Anti‐ and Pro‐apoptotic Actions of Dopamine,R‐Apomorphine, Green Tea Polyphenol (−)‐Epigallocatechine‐3‐gallate, and Melatonin
Authors: WEINREB ORLY , MANDEL SILVIA , YOUDIM MOUSSA B.H.
Published: July 2009

Abstract:Significant evidence has been provided to support the hypothesis that oxidant stress may be responsible for degeneration of dopaminergic neurons in the substantia nigra pars compacta in Parkinson's disease. Dopamine (DA),R‐apomorphine (R‐APO), green tea polyphenol (−)‐epigallocatechine‐3‐gallate (EGCG), and melatonin are neuroprotective and radical scavenger compounds. The aim of this study was to establish the mechanism of the concentration‐dependent neuroprotective and pro‐apoptotic action of these drugs via gene expression and protein determination. cDNA microarrays provide new prospects to study and identify various mechanisms of drug action. We employed this technique for the study reported in this paper. Total RNA was extracted from SH‐SY5Y cells exposed to low neuroprotective and high toxic concentrations of the drugs, followed by synthesis of cDNA, and hybridization to a microarray membrane related to apoptosis, survival, and cell cycle pathways. We demonstrated a concentration and time‐dependent correlation betweenR‐APO, DA, EGCG, and melatonin in modulation of cell survival/cell death‐related gene pathways. The results were confirmed by quantitative real‐time PCR and protein profiles. Unlike the effects of low concentrations (1‐10 μM), where an antiapoptotic response was manifest, a proapoptotic pattern of gene expression was observed at high toxic concentrations (50‐500 μM) of the antioxidants (e.g., increase incaspases, fas,andgadd45). Our results have provided novel insights into the gene mechanisms involved in both the neuroprotective and proapoptotic activities of neuroprotective drugs. We have shown that DA,R‐APO, EGCG, and melatonin exhibit similar gene expression and protein profiles.
Correlation of routinely used coagulation parameters and presence of portal vein thrombosis in patients with liver cirrhosis
Authors: Weber Andreas , Krebs Stephanie , Lenhardt Cosima , Wagenpfeil Stefan , Schmid Roland M. , Schulte‐Frohlinde Ewert
Published: July 2009

Aim:  Portal vein thrombosis (PVT) is a serious complication in patients with liver cirrhosis. In patients with advanced stages of liver cirrhosis plasmatic coagulation and platelet count are often reduced. However, patients with normal coagulation status might carry a high risk for developing PVT. A correlation between coagulation status used in clinical routine and the incidence of PVT in patients with liver cirrhosis has been evaluated in the present retrospective analysis.

Methods:  88 patients with liver cirrhosis were identified by screening a database. Of these patients, 23 suffered from PVT. Patients were classified according to the Child–Pugh classification. Patients were subdivided into early stages (Child A) and advanced stages (Child B/C) of liver cirrhosis.

Results:  In patients with Child–Pugh A cirrhosis, there was no difference in activated partial thromboplastin time (apTT), international normalized ratio (INR), and platelet count between the PVT (n = 7) and the control group (n = 35). In contrast, the median apTT and INR were significantly lower in patients with Child B/C cirrhosis and PVT (n = 16) in comparison with patients without PVT (37 s vs 43 s [P = 0.017] and 1.25 vs 1.40 [P = 0.022]), respectively. Platelet count did not differ significantly in patients with advanced liver cirrhosis and PVT from those without PVT.

Conclusion:  Patients with advanced liver cirrhosis and PVT displayed lower apTT and INR compared with those without PVT. Therefore, patients with advanced liver cirrhosis and almost normal coagulation parameters might be at particular risk of developing PVT. The results suggest that regular monitoring using Doppler‐ultrasound should be carried out in these patients, especially when liver transplantation is intended.
Vascular Cell Adhesion Molecule-1 Expression in Endothelial Cells Exposed to Physiological Coronary Wall Shear Stresses
Authors: O'Keeffe Lucy M. , Muir Gordon , Piterina Anna V. , McGloughlin Tim
Published: July 2009

Atherosclerosis is consistently found in bifurcations and curved segments of the circulatory system, indicating disturbed hemodynamics may participate in disease development. In vivo and in vitro studies have shown that endothelial cells (ECs) alter their gene expression in response to their hemodynamic environment, in a manner that is highly dependent on the exact nature of the applied forces. This research exposes cultured ECs to flow patterns present in the coronary arterial network, in order to determine the role of hemodynamic forces in plaque initiation. Vascular cell adhesion molecule-1 (VCAM-1) was examined as an indicator of plaque growth, as it participates in monocyte adhesion, which is one of the initial steps in the formation of fatty lesions. The hemodynamics of a healthy right and left coronary artery were determined by reconstructing 3D models from cineangiograms and employing computational fluid dynamic models to establish physiological coronary flow patterns. Wall shear stress (WSS) profiles selected from these studies were applied to ECs in a cone and plate bioreactor. The cone and plate system was specifically designed to be capable of reproducing the high frequency harmonics present in physiological waveforms. The shear stresses chosen represent those from regions prone to disease development and healthier arterial segments. The levels of the transcriptional and cell surface anchored VCAM-1 were quantified by flow cytometry and real time RT-PCR over a number of timepoints to obtain a complete picture of the relationship between this adhesion molecule and the applied shear stress. The WSS profiles from regions consistently displaying a higher incidence of plaques in vivo, induced greater levels of VCAM-1, particularly at the earlier timepoints. Conversely, the WSS profile from a straight section of vessel with undisturbed flow indicated no upregulation in VCAM-1 and a significant downregulation after 24 h, when compared with static controls. Low shear stress from the outer wall of a bifurcation induced four times the levels of VCAM-1 messenger ribonucleic acid (mRNA) after four hours when compared with levels of mRNA induced by WSS from a straight arterial section. This shear profile also induced prolonged expression of the surface protein of this molecule. The current study has provided insight into the possible influences of coronary hemodynamics on plaque localization, with VCAM-1 only significantly induced by the WSS from disease prone regions.
Induction of Synaptic Long-Term Potentiation After Opioid Withdrawal
Authors: Drdla Ruth , Gassner Matthias , Gingl Ewald , Sandkühler Jürgen
Published: July 2009

Paradoxical Pain After Opioid Withdrawal

Opioids are widely used both by pain patients and substance abusers. Withdrawal from opioids can be complicated by opioid-induced hyperalgesia, a paradoxical amplification of pain by opioids.

Drdla
et al.

(p.
207
) detected a novel cellular opioid action that may help to explain this paradoxical pain: the induction of synaptic long-term potentiation (LTP) in pain pathways by opioids. In vitro and in vivo µ-opioid receptor agonist withdrawal induced robust LTP at synapses of nociceptive C fibers. This is in contrast to the presynaptic inhibition by opioids at these synapses. Because opioidergic LTP and acute synaptic depression are spatially and mechanistically distinct, it may be possible to treat opioid-induced hyperalgesia selectively without compromising opioid analgesia.
Caloric Restriction Delays Disease Onset and Mortality in Rhesus Monkeys
Authors: Colman Ricki J. , Anderson Rozalyn M. , Johnson Sterling C. , Kastman Erik K. , Kosmatka Kristopher J. , Beasley T. Mark , Allison David B. , Cruzen Christina , Simmons Heather A. , Kemnitz Joseph W. , Weindruch Richard
Published: July 2009

Starved to Life?

Caloric restriction—reducing the calories ingested by around 30% of that of a normal, fit individual—leads to substantial increases in life span in experimental animals. In an extensive study of caloric restriction in primates,

Colman
et al.

(p.
201
) report that rhesus monkeys, which were subjected to caloric restriction as adults and followed for the last 20 years, show decreased mortality and delayed onset of age-related diseases when compared to normally fed control animals. If compliance with such a diet were not so difficult, many humans would be strongly tempted to enjoy the decreased incidence of brain degeneration, cardiovascular disease, diabetes, and cancer apparent in this population of monkeys.
Quantum Walk in Position Space with Single Optically Trapped Atoms
Authors: Karski Michal , Förster Leonid , Choi Jai-Min , Steffen Andreas , Alt Wolfgang , Meschede Dieter , Widera Artur
Published: July 2009

Strolling Out on a Quantum Walk

In a random walk, a walker moves one step to the left or one step to the right depending on the outcome of a coin toss. The distribution between possible locations is well known and forms the basis for algorithms in information processing, describing diffusion processes in physics or biology, and has even been used as a model for stock market prices.

Karski
et al.

(p.
174
) use a single caesium atom trapped in a one-dimensional optical lattice to implement the quantum counterpart—a quantum walk. The coherence of a quantum system results in a departure from the classical picture, producing a distribution that is quite different that depends on the internal state of the atom. The results may have implications for search algorithms and quantum information processing protocols.
DOE's Push to Train a New Generation Falters in House
Authors: Mervis Jeffrey
Published: July 2009

Secretary of Energy Steven Chu has proposed a 10-year, $1.7 billion education program called RE-ENERGYSE to help the United States find the talent to fuel its clean-energy economy. But Congress may prefer to wait until next year.
An Inside/Outside View of U.S. Science
Authors: Mervis Jeffrey
Published: July 2009

Three surveys conducted this spring by the Pew Research Center offer some fresh insights into what both the U.S. public and scientists think about science and its impact on society.
Bringing Hominins Back to Life
Authors: Balter Michael
Published: July 2009

To reconstruct our ancestors, paleoartists weave art and science together in a sometimes uneasy marriage. The result is lifelike models that influence how both researchers and the public view ancient humans.
Genomic Clues to DNA Treasure Sometimes Lead Nowhere
Authors: Monroe Don
Published: July 2009

Geneticists used to think that important DNA sequences always reject mutations. Now they are not so sure what sequence conservation really means.
Take-Charge B Cells Create a Buzz
Authors: Leslie Mitch
Published: July 2009

Long regarded as the source of antibodies, B cells are gaining new respect because some appear to fine-tune the overall immune response.
Bicentric Evaluation of Six Anti-Toxoplasma Immunoglobulin G (IgG) Automated Immunoassays and Comparison to the Toxo II IgG Western Blot
Authors: Maudry Arnaud , Chene Gautier , Chatelain Rémi , Patural Hugues , Bellete Bahrie , Tisseur Bernard , Hafid Jamal , Raberin Hélène , Beretta Sophie , Tran Manh Sung Roger , Belot Georges , Flori Pierre
Published: July 2009

A comparative study of the Toxoplasma IgG
I
and IgG
II
Access (Access I and II, respectively; Beckman Coulter Inc.), AxSYM Toxo IgG (AxSYM; Abbott Diagnostics), Vidas Toxo IgG (Vidas; bioMerieux, Marcy l'Etoile, France), Immulite Toxo IgG (Immulite; Siemens Healthcare Diagnostics Inc.), and Modular Toxo IgG (Modular; Roche Diagnostics, Basel, Switzerland) tests was done with 406 consecutive serum samples. The Toxo II IgG Western blot (LDBio, Lyon, France) was used as a reference technique in the case of intertechnique discordance. Of the 406 serum samples tested, the results for 35 were discordant by the different techniques. Using the 175 serum samples with positive results, we evaluated the standardization of the titrations obtained (in IU/ml); the medians (second quartiles) obtained were 9.1 IU/ml for the AxSYM test, 21 IU/ml for the Access I test, 25.7 IU/ml for the Access II test, 32 IU/ml for the Vidas test, 34.6 IU/ml for the Immulite test, and 248 IU/ml for the Modular test. For all the immunoassays tested, the following relative sensitivity and specificity values were found: 89.7 to 100% for the Access II test, 89.7 to 99.6% for the Immulite test, 90.2 to 99.6% for the AxSYM test, 91.4 to 99.6% for the Vidas test, 94.8 to 99.6% for the Access I test, and 98.3 to 98.7% for the Modular test. Among the 406 serum samples, we did not find any false-positive values by two different tests for the same serum sample. Except for the Modular test, which prioritized sensitivity, it appears that the positive cutoff values suggested by the pharmaceutical companies are very high (either for economical or for safety reasons). This led to imperfect sensitivity, a large number of unnecessary serological follow-ups of pregnant women, and difficulty in determining the serological status of immunosuppressed individuals.
Prevalence of CTX-M β-Lactamases in Philadelphia, Pennsylvania
Authors: McGettigan Shannon E. , Hu Baofeng , Andreacchio Kathleen , Nachamkin Irving , Edelstein Paul H.
Published: July 2009

CTX-M β-lactamases were thought to be rare in the United States, but a recent study in Texas showed that up to 70% of extended-spectrum β-lactamase (ESBL)-containing members of the
Enterobacteriaceae
family were CTX-M positive (J. S. Lewis, M. Herrera, B. Wickes, J. E. Patterson, and J. H. Jorgensen, Antimicrob. Agents Chemother. 51:4015-4021, 2007). We used PCR to detect CTX-M in all 291 extended-spectrum cephalosporin-resistant gram-negative bacteria isolated in our laboratory during 2007. Thirty (48%)
Escherichia coli
isolates, 6 (3%)
Klebsiella
sp. isolates, and 7 (100%)
Proteus mirabilis
isolates tested were CTX-M positive, with 15% of all
Enterobacteriaceae
tested being positive. The
E. coli
CTX-M groups were I (57%), IV (37%), II (3%), and not groupable (3%); three of the group IV isolates were positive for CTX-M-18, and three of the group I isolates were positive for CTX-M-15. One of seven positive
P. mirabilis
isolates was in group II, with the remainder being positive for a CTX-M-25-like β-lactamase; and 33% of the
Klebsiella
sp. isolates were in group I or IV, with the remainder not being in groups I to IV. CTX-M-producing bacteria were isolated from urine (
n
= 13), blood (
n
= 13), wounds (
n
= 12), and the respiratory tract (
n
= 4). All 31 CTX-M-positive isolates tested for the presence of ESBL were confirmed to produce ESBLs by the use of tests recommended by the CLSI. Pulsed-field gel electrophoresis of the CTX-M-positive isolates showed that six
P. mirabilis
isolates were clonal and that there were seven different
E. coli
clusters. Five of seven
P. mirabilis
isolates were from blood cultures. The CLSI tests for the confirmation of ESBL production reliably detect these isolates if both cefotaxime and ceftazidime are tested, but only about half would be classified as a possible CTX-M producers on the basis of the antibiogram alone. A new panprimer set increases the ability to detect CTX-M-producing strains. CTX-M-positive bacteria are common in our geographic region, are often invasive, and, with the exception of
P. mirabilis
, are multiclonal.
Rapid and Highly Sensitive Neuraminidase Subtyping of Avian Influenza Viruses by Use of a Diagnostic DNA Microarray
Authors: Gall Astrid , Hoffmann Bernd , Harder Timm , Grund Christian , Ehricht Ralf , Beer Martin
Published: July 2009

Rapid neuraminidase subtyping of avian influenza viruses from diagnostic samples is crucial considering the existence of permanently emerging and evolving strains. Here we report an easy-to-use, low-cost microarray for neuraminidase subtyping following fragment amplification by a generic, neuraminidase-specific reverse transcription-PCR (RT-PCR). This method enables highly specific characterization with a sensitivity equal to that of matrix gene-specific real-time RT-PCR.
Naturally Occurring Hepatitis C Virus Subgenomic Deletion Mutants Replicate Efficiently in Huh-7 Cells and Are
trans
-Packaged In Vitro To Generate Infectious Defective Particles
Authors: Pacini Laura , Graziani Rita , Bartholomew Linda , De Francesco Raffaele , Paonessa Giacomo
Published: July 2009

Naturally occurring hepatitis C virus (HCV) subgenomic RNAs have been found in several HCV patients. These subgenomic deletion mutants, mostly lacking the genes encoding envelope glycoproteins, were found in both liver and serum, where their relatively high abundance suggests that they are capable of autonomous replication and can be packaged and secreted in viral particles, presumably harboring the envelope proteins from wild type virus coinfecting the same cell. We recapitulated some of these natural subgenomic deletions in the context of the isolate JFH-1 and confirmed these hypotheses in vitro. In Huh-7.5 cells, these deletion-containing genomes show robust replication and can be efficiently
trans
-packaged and infect naïve Huh-7.5 cells when cotransfected with the full-length wild-type J6/JFH genome. The genome structure of these natural subgenomic deletion mutants was dissected, and the maintenance of both core and NS2 regions was proven to be significant for efficient replication and
trans
-packaging. To further explore the requirements needed to achieve
trans
-complementation, we provided different combinations of structural proteins in
trans
. Optimal
trans
-complementation was obtained when fragments of the polyprotein encompassing core to p7 or E1 to NS2 were expressed. Finally, we generated a stable helper cell line, constitutively expressing the structural proteins from core to p7, which efficiently supports
trans
-complementation of a subgenomic deletion encompassing amino acids 284 to 732. This cell line can produce and be infected by defective particles, thus representing a powerful tool to investigate the life cycle and relevance of natural HCV subgenomic deletion mutants in vivo.
The Genome of Human Parvovirus B19 Can Replicate in Nonpermissive Cells with the Help of Adenovirus Genes and Produces Infectious Virus
Authors: Guan Wuxiang , Wong Susan , Zhi Ning , Qiu Jianming
Published: July 2009

Human parvovirus B19 (B19V) is a member of the genusErythrovirusin the familyParvoviridae. In vitro, autonomous B19V replication is limited to human erythroid progenitor cells and in a small number of erythropoietin-dependent human megakaryoblastoid and erythroid leukemic cell lines. Here we report that the failure of B19V DNA replication in nonpermissive 293 cells can be overcome by adenovirus infection. More specifically, the replication of B19V DNA in the 293 cells and the production of infectious progeny virus were made possible by the presence of the adenovirus E2a, E4orf6, and VA RNA genes that emerged during the transfection of the pHelper plasmid. Using this replication system, we identified the terminal resolution site and the nonstructural protein 1 (NS1) binding site on the right terminal palindrome of the viral genome, which is composed of a minimal origin of replication spanning 67 nucleotides. Plasmids or DNA fragments containing an NS1 expression cassette and this minimal origin were able to replicate in both pHelper-transfected 293 cells and B19V-semipermissive UT7/Epo-S1 cells. Our results have important implications for our understanding of native B19V infection.
Differential Incorporation of Cholesterol by Sindbis Virus Grown in Mammalian or Insect Cells
Authors: Hafer Amanda , Whittlesey Rebecca , Brown Dennis T. , Hernandez Raquel
Published: July 2009

Cholesterol has been shown to be essential for the fusion of alphaviruses with artificial membranes (liposomes). Cholesterol has also been implicated as playing an essential and critical role in the processes of entry and egress of alphaviruses in living cells. Paradoxically, insects, the alternate host for alphaviruses, are cholesterol auxotrophs and contain very low levels of this sterol. To further evaluate the role of cholesterol in the life cycle of alphaviruses, the cholesterol levels of the alphavirus Sindbis produced from three different mosquito (
Aedes albopictus
) cell lines; one other insect cell line, Sf21 from
Spodoptera frugiperda
; and BHK (mammalian) cells were measured. Sindbis virus was grown in insect cells under normal culture conditions and in cells depleted of cholesterol by growth in serum delipidated by using Cab-O-sil, medium treated with methyl-β-cyclodextrin, or serum-free medium. The levels of cholesterol incorporated into the membranes of the cells and into the virus produced from these cells were determined. Virus produced from these treated and untreated cells was compared to virus grown in BHK cells under standard conditions. The ability of insect cells to produce Sindbis virus after delipidation was found to be highly cell specific and not dependent on the level of cholesterol in the cell membrane. A very low level of cholesterol was required for the generation of wild-type levels of infectious Sindbis virus from delipidated cells. The data show that one role of the virus membrane is structural, providing the stability required for infectivity that may not be provided by the delipidated membranes in some cells. These data show that the amount of cholesterol in the host cell membrane in and of itself has no effect on the process of virus assembly or on the ability of virus to infect cells. Rather, these data suggest that the cholesterol dependence reported for infectivity and assembly of Sindbis virus is a reflection of differences in the insect cell lines used and the methods of delipidation.
The Immune Adaptor Molecule SARM Modulates Tumor Necrosis Factor Alpha Production and Microglia Activation in the Brainstem and Restricts West Nile Virus Pathogenesis
Authors: Szretter Kristy J. , Samuel Melanie A. , Gilfillan Susan , Fuchs Anja , Colonna Marco , Diamond Michael S.
Published: July 2009

Sterile alpha and HEAT/Armadillo motif (SARM) is a highly conserved Toll/interleukin-1 receptor (TIR)-containing adaptor protein that is believed to negatively regulate signaling of the pathogen recognition receptors Toll-like receptor 3 (TLR3) and TLR4. To test its physiological function in the context of a microbial infection, we generated SARM−/−mice and evaluated the impact of this deficiency on the pathogenesis of West Nile virus (WNV), a neurotropic flavivirus that requires TLR signaling to restrict infection. Although SARM was preferentially expressed in cells of the central nervous system (CNS), studies with primary macrophages, neurons, or astrocytes showed no difference in viral growth kinetics. In contrast, viral replication was increased specifically in the brainstem of SARM−/−mice, and this was associated with enhanced mortality after inoculation with a virulent WNV strain. A deficiency of SARM was also linked to reduced levels of tumor necrosis factor alpha (TNF-α), decreased microglia activation, and increased neuronal death in the brainstem after WNV infection. Thus, SARM appears to be unique among the TIR adaptor molecules, since it functions to restrict viral infection and neuronal injury in a brain region-specific manner, possibly by modulating the activation of resident CNS inflammatory cells.