bluuebunny/crossref_metadata_2025 huggingface dataset:
In
Vitro
Evaluation of Antibiotic Synergy for Polymyxin B-Resistant Carbapenemase-Producing
Klebsiella pneumoniae
Authors: Elemam Azza , Rahimian Joseph , Doymaz Mehmet
Published: August 2010

Since carbapenemase-producing
Klebsiella pneumoniae
strains were first reported in North Carolina, these highly resistant organisms have been isolated with increasing frequency, especially in the New York City area. Polymyxin B is one of the few antimicrobials that retain reliable activity against these organisms. However, polymyxin B MICs are elevated against
K. pneumoniae
isolates with increasing frequency, leaving clinicians with few therapeutic options. We investigated several antimicrobial agents for potential synergy with polymyxin B against 12 clinical strains of carbapenemase-producing
K. pneumoniae.
A broth microdilution assay using a 96-well plate was developed in which graded dilutions of polymyxin B and the study drug were incubated with resistant isolates in a checkerboard pattern. Polymyxin B was studied in combination with cefazolin, ceftriaxone, cefepime, imipenem, gentamicin, tigecycline, doxycycline, and rifampin. All
K. pneumoniae
strains tested positive for
K. pneumoniae
carbapenemase (KPC) genes by real-time PCR and had elevated polymyxin B MIC values ranging from 16 to 128 μg/ml. Synergy was observed with the combination of polymyxin B and rifampin as well as with polymyxin B and doxycycline, resulting in at least a 4-fold decrease in the polymyxin B MIC. For both combinations, this effect occurred at physiologically achievable concentrations. Less pronounced synergy was noted with tigecycline and polymyxin B. No synergy was observed at physiologic concentrations with the other antimicrobials studied. These results suggest that rifampin, doxycycline, and tigecycline may be useful additions to polymyxin B in the treatment of infections caused by highly resistant carbapenemase-producing
K. pneumoniae
. Further studies are warranted to determine if these
in vitro
findings translate into clinical efficacy.
Identification, Molecular Characterization, and Occurrence of Two Bovine Hemoplasma Species in Swiss Cattle and Development of Real-Time TaqMan Quantitative PCR Assays for Diagnosis of Bovine Hemoplasma Infections
Authors: Meli Marina L. , Willi Barbara , Dreher Ute M. , Cattori Valentino , Knubben-Schweizer Gabriela , Nuss Karl , Braun Ueli , Lutz Hans , Hofmann-Lehmann Regina
Published: August 2010

Concomitantly with an outbreak of fatal anaplasmosis in a cattle herd in Switzerland in 2002, we detected two bovine hemoplasma species in diseased animals:Mycoplasma wenyonii(formerlyEperythrozoon wenyonii) and a second, novel bovine hemoplasma species later designated “CandidatusMycoplasma haemobos” (synonym, “CandidatusMycoplasma haemobovis”). The second species was characterized by a shorter 16S rRNA gene. The aims of the present study were to provide a detailed molecular characterization of this species, to develop specific quantitative real-time PCR assays for the two bovine hemoplasma species, and to apply these assays in order to evaluate the prevalence and clinical significance of the hemoplasmas. Sequencing of the near-complete 16S rRNA gene of the second hemoplasma revealed that it was 94% identical to that ofMycoplasma haemofelis, an anemia-inducing feline hemoplasma species, but less than 85% identical to that of the bovine hemoplasmaM. wenyonii. Using the newly developed assays, a total of 159 animals from the anaplasmosis outbreak were reexamined. In addition, we tested 57 clinically ill and 61 healthy Swiss cattle, as well as 47 calves. Both hemoplasmas were highly prevalent in adult cattle but occurred rarely in calves. Animals from the herd with the fatal anemia outbreak were more frequently infected withM. wenyoniiand exhibited higherM. wenyoniiblood loads than animals with unrelated diseases and healthy animals. Coinfections may increase the pathogenicity and clinical significance of bovine hemoplasmosis.
Genome-Wide Detection and Characterization of Endogenous Retroviruses inBos taurus
Authors: Garcia-Etxebarria Koldo , Jugo Begoña Marina
Published: August 2010

Endogenous retroviruses (ERVs) are the proviral phase of exogenous retroviruses that become integrated into a host germ line. They can play an important role in the host genome. Bioinformatic tools have been used to detect ERVs in several vertebrates, primarily primates and rodents. Less information is available regarding ERVs in other mammalian groups, and the source of this information is basically experimental. We analyzed the genome of the cow (Bos taurus) using three different methods. A BLAST-based method detected 928 possible ERVs, LTR_STRUC detected 4,487 elements flanked by long terminal repeats (LTRs), and Retrotector detected 9,698 ERVs. The ERVs were not homogeneously distributed across chromosomes; the number of ERVs was positively correlated with chromosomal size and negatively correlated with chromosomal GC content. The bovine ERVs (BoERVs) were classified into 24 putative families, with 20 of them not previously described. One of these new families, BoERV1, was the most abundant family and appeared to be specific to ruminants. An analysis of representatives of ERV families from rodents, primates, and ruminants showed a phylogenetic relationship following their hosts' relationships. This study demonstrates the importance of using multiple methods when trying to identify new ERVs and shows that the number of bovine ERV families is not as limited as previously thought.
Simian Virus 40 Activates ATR-Δp53 Signaling To Override Cell Cycle and DNA Replication Control
Authors: Rohaly Gabor , Korf Katharina , Dehde Silke , Dornreiter Irena
Published: August 2010

During infection, simian virus 40 (SV40) attempts to take hold of the cell, while the host responds with various defense systems, including the ataxia-telangiectasia mutated/ATM-Rad3 related (ATM/ATR)-mediated DNA damage response pathways. Here we show that upon viral infection, ATR directly activates the p53 isoform Δp53, leading to upregulation of the Cdk inhibitor p21 and downregulation of cyclin A-Cdk2/1 (AK) activity, which force the host to stay in the replicative S phase. Moreover, downregulation of AK activity is a prerequisite for the generation of hypophosphorylated, origin-competent DNA polymerase α-primase (hypo-Polα), which is, unlike AK-phosphorylated Polα (P-Polα), recruited by SV40 large T antigen (T-Ag) to initiate viral DNA replication. Prevention of the downregulation of AK activity by inactivation of ATR-Δp53-p21 signaling significantly reduced the T-Ag-interacting hypo-Polα population and, accordingly, SV40 replication efficiency. Moreover, the ATR-Δp53 pathway facilitates the proteasomal degradation of the 180-kDa catalytic subunit of the non-T-Ag-interacting P-Polα, giving rise to T-Ag-interacting hypo-Polα. Thus, the purpose of activating the ATR-Δp53-p21-mediated intra-S checkpoint is to maintain the host in S phase, an optimal environment for SV40 replication, and to modulate the host DNA replicase, which is indispensable for viral amplification.
Variable Loop Glycan Dependency of the Broad and Potent HIV-1-Neutralizing Antibodies PG9 and PG16
Authors: Doores Katie J. , Burton Dennis R.
Published: August 2010

The HIV-1-specific antibodies PG9 and PG16 show marked cross-isolate neutralization breadth and potency. Antibody neutralization has been shown to be dependent on the presence of N-linked glycosylation at position 160 in gp120. We show here that (i) the loss of several key glycosylation sites in the V1, V2, and V3 loops; (ii) the generation of pseudoviruses in the presence of various glycosidase inhibitors; and (iii) the growth of pseudoviruses in a mutant cell line (GnT1
−/−
) that alters envelope glycosylation patterns all have significant effects on the sensitivity of virus to neutralization by PG9 and PG16. However, the interaction of antibody is not inhibited by sugar monosaccharides corresponding to those found in glycans on the HIV surface. We show that some of the glycosylation effects described are isolate dependent and others are universal and can be used as diagnostic for the presence of PG9 and PG16-like antibodies in the sera of HIV-1-infected patients. The results suggest that PG9 and PG16 recognize a conformational epitope that is dependent on glycosylation at specific variable loop N-linked sites. This information may be valuable for the design of immunogens to elicit PG9 and PG16-like antibodies, as well as constructs for cocrystallization studies.
Long-Term Restriction by APOBEC3F Selects Human Immunodeficiency Virus Type 1 Variants with Restored Vif Function
Authors: Albin John S. , Haché Guylaine , Hultquist Judd F. , Brown William L. , Harris Reuben S.
Published: August 2010

Tandem stop mutations K26X and H27X in human immunodeficiency virus type 1 (HIV-1)
vif
compromise virus replication in human T-cell lines that stably express APOBEC3F (A3F) or APOBEC3G (A3G). We previously reported that partial resistance to A3G could develop in these Vif-deficient viruses through a nucleotide A200-to-T/C transversion and a
vpr
null mutation, but these isolates were still susceptible to restriction by A3F. Here, long-term selection experiments were done to determine how these A3G-selected isolates might evolve to spread in the presence of A3F. We found that A3F, like A3G, is capable of potent, long-term restriction that eventually selects for heritable resistance. In all 7 instances, the selected isolates had restored Vif function to cope with A3F activity. In two isolates, Vif Q26-Q27 and Y26-Q27, the resistance phenotype recapitulated in molecular clones, but when the selected
vif
alleles were analyzed in the context of an otherwise wild-type viral background, a different outcome emerged. Although HIV-1 clones with Vif Q26-Q27 or Y26-Q27 were fully capable of overcoming A3F, they were now susceptible to restriction by A3G. Concordant with prior studies, a lysine at position 26 proved essential for A3G neutralization. These data combine to indicate that A3F and A3G exert at least partly distinct selective pressures and that Vif function may be essential for the virus to replicate in the presence of A3F.
Chromosomal Integration of Adenoviral Vector DNA
In Vivo
Authors: Stephen Sam Laurel , Montini Eugenio , Sivanandam Vijayshankar Ganesh , Al-Dhalimy Muhseen , Kestler Hans A. , Finegold Milton , Grompe Markus , Kochanek Stefan
Published: August 2010

So far there has been no report of any clinical or preclinical evidence for chromosomal vector integration following adenovirus (Ad) vector-mediated gene transfer
in vivo
. We used liver gene transfer with high-capacity Ad vectors in the FAH
Δexon5
mouse model to analyze homologous and heterologous recombination events between vector and chromosomal DNA. Intravenous injection of Ad vectors either expressing a fumarylacetoacetate hydrolase (FAH) cDNA or carrying part of the FAH genomic locus resulted in liver nodules of FAH-expressing hepatocytes, demonstrating chromosomal vector integration. Analysis of junctions between vector and chromosomal DNA following heterologous recombination indicated integration of the vector genome through its termini. Heterologous recombination occurred with a median frequency of 6.72 × 10
−5
per transduced hepatocyte, while homologous recombination occurred more rarely with a median frequency of 3.88 × 10
−7
. This study has established quantitative and qualitative data on recombination of adenoviral vector DNA with genomic DNA
in vivo
, contributing to a risk-benefit assessment of the biosafety of Ad vector-mediated gene transfer.
Convergence of Kaposi's Sarcoma-Associated Herpesvirus Reactivation with Epstein-Barr Virus Latency and Cellular Growth Mediated by the Notch Signaling Pathway in Coinfected Cells
Authors: Spadavecchia Sophia , Gonzalez-Lopez Olga , Carroll Kyla Driscoll , Palmeri Diana , Lukac David M.
Published: August 2010

Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiologic agent of primary effusion lymphoma (PEL). All PEL cell lines are infected with KSHV, and 70% are coinfected with Epstein-Barr virus (EBV). KSHV reactivation from latency requires promoter-specific transactivation by the KSHV Rta protein through interactions with RBP-Jk (CSL), the cellular DNA-binding component of the Notch signal transduction pathway. EBV transformation of primary B cells requires EBV nuclear antigen 2 (EBNA-2) to interact with RBP-Jk to direct the latent viral and cellular gene expression program. Although KSHV Rta and EBV EBNA-2 both require RBP-Jk for transactivation, previous studies have suggested that RBP-Jk-dependent transactivators do not function identically. We have found that the EBV latent protein LMP-1 is expressed in less than 5% of KSHV
+
/EBV
+
PEL cells but is induced in an Rta-dependent fashion when KSHV reactivates. KSHV Rta transactivates the EBV latency promoters in an RBP-Jk-dependent fashion and forms a ternary complex with RBP-Jk on the promoters. In B cells that are conditionally transformed by EBV alone, we show that KSHV Rta complements a short-term EBNA-2 growth deficiency in an autocrine/paracrine manner. Complementation of EBNA-2 deficiency by Rta depends on RBP-Jk and LMP-1, and Rta transactivation is required for optimal growth of KSHV
+
/EBV
+
PEL lines. Our data suggest that Rta can contribute to EBV-driven cellular growth by transactivating RBP-Jk-dependent EBV latency genes. However, our data also suggest that EBNA-2 and Rta induce distinct alterations in the cellular proteomes that contribute to the growth of infected cells.
Basic Residues within the Ebolavirus VP35 Protein Are Required for Its Viral Polymerase Cofactor Function
Authors: Prins Kathleen C. , Binning Jennifer M. , Shabman Reed S. , Leung Daisy W. , Amarasinghe Gaya K. , Basler Christopher F.
Published: August 2010

The ebolavirus (EBOV) VP35 protein binds to double-stranded RNA (dsRNA), inhibits host alpha/beta interferon (IFN-α/β) production, and is an essential component of the viral polymerase complex. Structural studies of the VP35 C-terminal IFN inhibitory domain (IID) identified specific structural features, including a central basic patch and a hydrophobic pocket, that are important for dsRNA binding and IFN inhibition. Several other conserved basic residues bordering the central basic patch and a separate cluster of basic residues, called the first basic patch, were also identified. Functional analysis of alanine substitution mutants indicates that basic residues outside the central basic patch are not required for dsRNA binding or for IFN inhibition. However, minigenome assays, which assess viral RNA polymerase complex function, identified these other basic residues to be critical for viral RNA synthesis. Of these, a subset located within the first basic patch is important for VP35-nucleoprotein (NP) interaction, as evidenced by the inability of alanine substitution mutants to coimmunoprecipitate with NP. Therefore, first basic patch residues are likely critical for replication complex formation through interactions with NP. Coimmunoprecipitation studies further demonstrate that the VP35 IID is sufficient to interact with NP and that dsRNA can modulate VP35 IID interactions with NP. Other basic residue mutations that disrupt the VP35 polymerase cofactor function do not affect interaction with NP or with the amino terminus of the viral polymerase. Collectively, these results highlight the importance of conserved basic residues from the EBOV VP35 C-terminal IID and validate the VP35 IID as a potential therapeutic target.
Chikungunya Virus Nonstructural Protein 2 Inhibits Type I/II Interferon-Stimulated JAK-STAT Signaling
Authors: Fros Jelke J. , Liu Wen Jun , Prow Natalie A. , Geertsema Corinne , Ligtenberg Maarten , Vanlandingham Dana L. , Schnettler Esther , Vlak Just M. , Suhrbier Andreas , Khromykh Alexander A. , Pijlman Gorben P.
Published: August 2010

Chikungunya virus (CHIKV) is an emerging human pathogen transmitted by mosquitoes. Like that of other alphaviruses, CHIKV replication causes general host shutoff, leading to severe cytopathicity in mammalian cells, and inhibits the ability of infected cells to respond to interferon (IFN). Recent research, however, suggests that alphaviruses may have additional mechanisms to circumvent the host's antiviral IFN response. Here we show that CHIKV replication is resistant to inhibition by interferon once RNA replication has been established and that CHIKV actively suppresses the antiviral IFN response by preventing IFN-induced gene expression. Both CHIKV infection and CHIKV replicon RNA replication efficiently blocked STAT1 phosphorylation and/or nuclear translocation in mammalian cells induced by either type I or type II IFN. Expression of individual CHIKV nonstructural proteins (nsPs) showed that nsP2 was a potent inhibitor of IFN-induced JAK-STAT signaling. In addition, mutations in CHIKV-nsP2 (P718S) and Sindbis virus (SINV)-nsP2 (P726S) that render alphavirus replicons noncytopathic significantly reduced JAK-STAT inhibition. This host shutoff-independent inhibition of IFN signaling by CHIKV is likely to have an important role in viral pathogenesis.
Mucosal Trafficking of Vector-Specific CD4
+
T Lymphocytes following Vaccination of Rhesus Monkeys with Adenovirus Serotype 5
Authors: Masek-Hammerman Katherine , Li Hualin , Liu Jinyan , Abbink Peter , La Porte Annalena , O'Brien Kara L. , Whitney James B. , Carville Angela , Mansfield Keith G. , Barouch Dan H.
Published: August 2010

Post hoc analysis of the phase 2b Step study evaluating a recombinant adenovirus serotype 5 (rAd5)-based HIV-1 vaccine candidate suggested a potential increased risk of HIV-1 acquisition in subjects who were baseline Ad5 seropositive and uncircumcised. These concerns had a profound impact on the HIV-1 vaccine development field, although the mechanism underlying this observation remains unknown. It has been hypothesized that rAd5 vaccination of baseline Ad5-seropositive individuals may have resulted in anamnestic, vector-specific CD4
+
T lymphocytes that could have trafficked to mucosal sites and served as increased targets for HIV-1 infection. Here we show that Ad5-specific CD4
+
T lymphocyte responses at mucosal sites following rAd5-Gag/Pol/Nef vaccination were comparable in rhesus monkeys with and without baseline Ad5 immunity. Moreover, the total cellular inflammatory infiltrates and the CD3
+
, CD4
+
, HLA-DR
+
, Ki67
+
, and langerin
+
cellular subpopulations in colorectal and foreskin mucosa were similar in both groups. Thus, no greater trafficking of Ad5-specific CD4
+
T lymphocytes to mucosal target sites was observed following rAd5 vaccination of rhesus monkeys with baseline Ad5 immunity. These findings from this nonhuman primate model provide evidence against the hypothesis that recruitment of vector-specific target cells to mucosal sites led to increased HIV-1 acquisition in Ad5-seropositive, uncircumcised vaccinees in the Step study.
A Single-Amino-Acid Substitution in Herpes Simplex Virus 1 Envelope Glycoprotein B at a Site Required for Binding to the Paired Immunoglobulin-Like Type 2 Receptor α (PILRα) Abrogates PILRα-Dependent Viral Entry and Reduces Pathogenesis
Authors: Arii Jun , Wang Jing , Morimoto Tomomi , Suenaga Tadahiro , Akashi Hiroomi , Arase Hisashi , Kawaguchi Yasushi
Published: August 2010

Paired immunoglobulin-like type 2 receptor α (PILRα) is a herpes simplex virus 1 (HSV-1) entry receptor that associates with O-glycans on HSV-1 envelope glycoprotein B (gB). Two threonine residues (Thr-53 and Thr-480) in gB, which are required for the addition of the principal gB O-glycans, are essential for binding to soluble PILRα. However, the role of the two threonines in PILRα-dependent viral entry remains to be elucidated. Therefore, we constructed a recombinant HSV-1 carrying an alanine replacement of gB Thr-53 alone (gB-T53A) or of both gB Thr-53 and Thr-480 (gB-T53/480A) and demonstrated that these mutations abrogated viral entry in CHO cells expressing PILRα. In contrast, the mutations had no effect on viral entry in CHO cells expressing known host cell receptors for HSV-1 gD, viral entry in HL60 cells expressing myelin-associated glycoprotein (MAG) (another HSV-1 gB receptor), viral attachment to heparan sulfate, and viral replication in PILRα-negative cells. These results support the hypothesis that gB Thr-53 and Thr-480 as well as gB O-glycosylation, probably at these sites, are critical for PILRα-dependent viral entry. Interestingly, following corneal inoculation in mice, the gB-T53A and gB-T53/480A mutations significantly reduced viral replication in the cornea, the development of herpes stroma keratitis, and neuroinvasiveness. The abilities of HSV-1 to enter cells in a PILRα-dependent manner and to acquire specific carbohydrates on gB are therefore linked to an increase in viral replication and virulence in the experimental murine model.
Translational Research on Trichoderma: From 'Omics to the Field
Authors: Lorito Matteo , Woo Sheridan L. , Harman, Gary E. , Monte Enrique
Published: August 2010

Structural and functional genomics investigations are making an important impact on the current understanding and application of microbial agents used for plant disease control. Here, we review the case of Trichoderma spp., the most widely applied biocontrol fungi, which have been extensively studied using a variety of research approaches, including genomics, transcriptomics, proteomics, metabolomics, etc. Known for almost a century for their beneficial effects on plants and the soil, these fungi are the subject of investigations that represent a successful case of translational research, in which 'omics-generated novel understanding is directly translated in to new or improved crop treatments and management methods. We present an overview of the latest discoveries on the Trichoderma expressome and metabolome, of the complex and diverse biotic interactions established in nature by these microbes, and of their proven or potential importance to agriculture and industry.
Potyviruses and the Digital Revolution
Authors: Gibbs Adrian , Ohshima Kazusato
Published: August 2010

The potyviruses are one of the two most speciose taxa of plant viruses. Our expanded knowledge of the breadth and depth of their diversity and its origins has depended greatly on the use of computing and the Internet in biological research and is reviewed here. We report a fully supported phylogeny based on gene sequence data for approximately half the named species. The phylogeny shows that the genus probably originated from a virus of monocotyledonous plants and that it first diverged approximately 7250 years ago in Southwest Eurasia or North Africa. The use of computer programs to better understand the structure and evolutionary trajectory of potyvirus populations is illustrated. The review concludes with recommendations for improving potyvirus nomenclature and the databasing of potyvirus information.
Go Where the Science Leads You
Authors: Hussey Richard S.
Published: August 2010

This article relates how my lifelong passion for nematology evolved and the philosophy that drove my research program, including maintaining a balance between applied and basic research, and key collaborations I have had with other researchers. Although the driving force behind my basic research was to advance our understanding of the molecular mechanisms of nematode parasitism of plants, the underlying theme was how to apply the new basic knowledge of nematode biology to provide better control of these economically important crop pathogens in grower fields. There are high expectations that new nematode control strategies will result from science-based solutions that can be delivered through biotechnology-derived crops and provide an unprecedented opportunity for limiting nematode damage to multiple crops.
Current Epidemiological Understanding of Citrus Huanglongbing
Authors: Gottwald Tim R.
Published: August 2010

Huanglongbing (HLB) is the most destructive citrus pathosystem worldwide. Previously known primarily from Asia and Africa, it was introduced into the Western Hemisphere in 2004. All infected commercial citrus industries continue to decline owing to inadequate current control methods. HLB increase and regional spatial spread, related to vector populations, are rapid compared with other arboreal pathosystems. Disease dynamics result from multiple simultaneous spatial processes, suggesting that psyllid vector transmission is a continuum from local area to very long distance. Evolutionarily, HLB appears to have originated as an insect endosymbiont that has moved into plants. Lack of exposure of citrus to the pathogen prior to approximately 100 years ago did not provide sufficient time for development of resistance. A prolonged incubation period and regional dispersal make eradication nonviable. Multiple asymptomatic infections per symptomatic tree, incomplete systemic distribution within trees, and prolonged incubation period make detection difficult and greatly complicate disease control.
Managing Nematodes Without Methyl Bromide
Authors: Zasada Inga A. , Halbrendt John M. , Kokalis-Burelle Nancy , LaMondia James , McKenry Michael V. , Noling Joe W.
Published: August 2010

Methyl bromide is an effective pre-plant soil fumigant used to control nematodes in many high-input, high-value crops in the United States, including vegetables, nursery plants, ornamentals, tree fruits, strawberries, and grapes. Because methyl bromide has provided a reliable return on investment for nematode control, many of these commodities have standardized their production practices based on the use of this chemical and will be negatively impacted if effective and economical alternatives are not identified. Alternative control measures based on other chemicals, genetic resistance, and cultural practices require a greater knowledge of nematode biology to achieve satisfactory results. Here, we provide an overview of nematode management practices that we believe will be relied upon heavily in U.S. high-value crop production systems in a world without methyl bromide. Included are case studies of U.S. high-value crop production systems to demonstrate how nematode management practices other than methyl bromide may be incorporated.
Engineering Pathogen Resistance in Crop Plants: Current Trends and Future Prospects
Authors: Collinge David B. , Jørgensen Hans J.L. , Lund Ole S. , Lyngkjær Michael F.
Published: August 2010

Transgenic crops are now grown commercially in 25 countries worldwide. Although pathogens represent major constraints for the growth of many crops, only a tiny proportion of these transgenic crops carry disease resistance traits. Nevertheless, transgenic disease-resistant plants represent approximately 10% of the total number of approved field trials in North America, a proportion that has remained constant for 15 years. In this review, we explore the socioeconomic and biological reasons for the paradox that although technically useful solutions now exist for providing transgenic disease resistance, very few new crops have been introduced to the global market. For bacteria and fungi, the majority of transgenic crops in trials express antimicrobial proteins. For viruses, three-quarters of the transgenics express coat protein (CP) genes. There is a notable trend toward more biologically sophisticated solutions involving components of signal transduction pathways regulating plant defenses. For viruses, RNA interference is increasingly being used.
Role of Small RNAs in Host-Microbe Interactions
Authors: Katiyar-Agarwal Surekha , Jin Hailing
Published: August 2010

Plant defense responses against pathogens are mediated by activation and repression of a large array of genes. Host endogenous small RNAs are essential in this gene expression reprogramming process. Here, we discuss recent findings on pathogen-regulated host microRNAs (miRNAs) and small interfering RNAs (siRNAs) and their roles in plant-microbe interaction. We further introduce small RNA pathway components, including Dicer-like proteins (DCLs), double-stranded RNA (dsRNA) binding protein, RNA-dependent RNA polymerases (RDRs), small RNA methyltransferase HEN1, and Argonaute (AGO) proteins, that contribute to plant immune responses. The strategies that pathogens have evolved to suppress host small RNA pathways are also discussed. Collectively, host small RNAs and RNA silencing machinery constitute a critical layer of defense in regulating the interaction of pathogens with plants.
Plant Proteins Involved in Agrobacterium-Mediated Genetic Transformation
Authors: Gelvin Stanton B.
Published: August 2010

Agrobacterium species genetically transform plants by transferring a region of plasmid DNA, T-DNA, into host plant cells. The bacteria also transfer several virulence effector proteins. T-DNA and virulence proteins presumably form T-complexes within the plant cell. Super-T-complexes likely also form by interaction of plant-encoded proteins with T-complexes. These protein-nucleic acid complexes traffic through the plant cytoplasm, enter the nucleus, and eventually deliver T-DNA to plant chromatin. Integration of T-DNA into the plant genome establishes a permanent transformation event, permitting stable expression of T-DNA-encoded transgenes. The transformation process is complex and requires participation of numerous plant proteins. This review discusses our current knowledge of plant proteins that contribute to Agrobacterium-mediated transformation, the roles these proteins play in the transformation process, and the modern technologies that have been employed to elucidate the cell biology of transformation.
Pathogen Refuge: A Key to Understanding Biological Control
Authors: Johnson Kenneth B.
Published: August 2010

Pathogen refuge is the idea that some potentially infectious pathogen propagules are not susceptible to the influence of an antagonistic microbial agent. The existence of a refuge can be attributable to one or more factors, including temporal, spatial, structural, and probabilistic, or to the pathogen's evolved ability to acquire antagonist-free space prior to ingress into a plant host. Within a specific pathosystem, refuge size can be estimated in experiments by measuring the proportion of pathogen propagules that remain infective as a function of the amount of antagonist introduced to the system. Refuge size is influenced by qualities of specific antagonists and by environment but less so by the quantity of antagonist. Consequently, most efforts to improve and optimize biological control are in essence efforts to reduce refuge size. Antagonist mixtures, optimal timing of antagonist introductions, integrated biological and chemical control, environmental optimization, and the utilization of disarmed pathogens as antagonists are strategies with potential to minimize a pathogen refuge.
Expression profile and protein translation of TMEM16A in murine smooth muscle
Authors: Davis Alison J. , Forrest Abigail S. , Jepps Thomas A. , Valencik Maria L. , Wiwchar Michael , Singer Cherie A. , Sones William R. , Greenwood Iain A. , Leblanc Normand
Published: August 2010

Recently, overexpression of the genes TMEM16A and TMEM16B has been shown to produce currents qualitatively similar to native Ca2+-activated Cl− currents ( IClCa) in vascular smooth muscle. However, there is no information about this new gene family in vascular smooth muscle, where Cl− channels are a major depolarizing mechanism. Qualitatively similar Cl− currents were evoked by a pipette solution containing 500 nM Ca2+ in smooth muscle cells isolated from BALB/c mouse portal vein, thoracic aorta, and carotid artery. Quantitative PCR using SYBR Green chemistry and primers specific for transmembrane protein (TMEM) 16A or the closely related TMEM16B showed TMEM16A expression as follows: portal vein > thoracic aorta > carotid artery > brain. In addition, several alternatively spliced variant transcripts of TMEM16A were detected. In contrast, TMEM16B expression was very low in smooth muscle. Western blot analysis with different antibodies directed against TMEM16A revealed a number of products with a consistent band at ∼120 kDa, except portal vein, where an 80-kDa band predominated. TMEM16A protein was identified in the smooth muscle layers of 4-μm-thick slices of portal vein, thoracic aorta, and carotid artery. In isolated myocytes, fluorescence specific to a TMEM16A antibody was detected diffusely throughout the cytoplasm, as well as near the membrane. The same antibody used in Western blot analysis of lysates from vascular tissues also recognized an ∼147-kDa mouse TMEM16A-green fluorescent protein (GFP) fusion protein expressed in HEK 293 cells, which correlated to a similar band detected by a GFP antibody. Patch-clamp experiments revealed that IClCa generated by transfection of TMEM16A-GFP in HEK 293 cells displayed remarkable similarities to IClCa recorded in vascular myocytes, including slow kinetics, steep outward rectification, and a response similar to the pharmacological agent niflumic acid. This study shows that TMEM16A expression is robust in murine vascular smooth muscle cells, consolidating the view that this gene is a viable candidate for the native Ca2+-activated Cl− channel in this cell type.
A new method for direct detection of the sites of actin polymerization in intact cells and its application to differentiated vascular smooth muscle
Authors: Kim Hak Rim , Leavis Paul C. , Graceffa Philip , Gallant Cynthia , Morgan Kathleen G.
Published: August 2010

Here we report and validate a new method, suitable broadly, for use in differentiated cells and tissues, for the direct visualization of actin polymerization under physiological conditions. We have designed and tested different versions of fluorescently labeled actin, reversibly attached to the protein transduction tag TAT, and have introduced this novel reagent into intact differentiated vascular smooth muscle cells (dVSMCs). A thiol-reactive version of the TAT peptide was synthesized by adding the amino acids glycine and cysteine to its NH2-terminus and forming a thionitrobenzoate adduct: viz. TAT-Cys-S-STNB. This peptide reacts readily with G-actin, and the complex is rapidly taken up by freshly enzymatically isolated dVSMC, as indicated by the fluorescence of a FITC tag on the TAT peptide. By comparing different versions of the construct, we determined that the optimal construct for biological applications is a nonfluorescently labeled TAT peptide conjugated to rhodamine-labeled actin. When TAT-Cys-S-STNB-tagged rhodamine actin (TSSAR) was added to live, freshly enzymatically isolated cells, we observed punctae of incorporated actin at the cortex of the cell. The punctae are indistinguishable from those we have previously reported to occur in the same cell type when rhodamine G-actin is added to permeabilized cells. Thus this new method allows the delivery of labeled G-actin into intact cells without disrupting the native state and will allow its further use to study the effect of physiological intracellular Ca2+ concentration transients and signal transduction on actin dynamics in intact cells.
Cystic fibrosis transmembrane conductance regulator is involved in airway epithelial wound repair
Authors: Schiller Katherine R. , Maniak Peter J. , O'Grady Scott M.
Published: August 2010

The role of cystic fibrosis (CF) transmembrane conductance regulator (CFTR) in airway epithelial wound repair was investigated using normal human bronchial epithelial (NHBE) cells and a human airway epithelial cell line (Calu-3) of serous gland origin. Measurements of wound repair were performed using continuous impedance sensing to determine the time course for wound closure. Control experiments showed that the increase in impedance corresponding to cell migration into the wound was blocked by treatment with the actin polymerization inhibitor, cytochalasin D. Time lapse imaging revealed that NHBE and Calu-3 cell wound closure was dependent on cell migration, and that movement occurred as a collective sheet of cells. Selective inhibition of CFTR activity with CFTRinh-172 or short hairpin RNA silencing of CFTR expression produced a significant delay in wound repair. The CF cell line UNCCF1T also exhibited significantly slower migration than comparable normal airway epithelial cells. Inhibition of CFTR-dependent anion transport by treatment with CFTRinh-172 slowed wound closure to the same extent as silencing CFTR protein expression, indicating that ion transport by CFTR plays a critical role in migration. Moreover, morphologic analysis of migrating cells revealed that CFTR inhibition or silencing significantly reduced lamellipodia protrusion. These findings support the conclusion that CFTR participates in airway epithelial wound repair by a mechanism involving anion transport that is coupled to the regulation of lamellipodia protrusion at the leading edge of the cell.
Rapid diagnosis and quantification of acute kidney injury using fluorescent ratio-metric determination of glomerular filtration rate in the rat
Authors: Wang Exing , Sandoval Ruben M. , Campos Silvia B. , Molitoris Bruce A.
Published: August 2010

The rapid diagnosis and quantification of acute kidney injury (AKI) severity remain high clinical priorities. By combining intravital fluorescent ratiometric two-photon kidney imaging and the two-compartment pharmacokinetics model, we demonstrate that rapid quantification of glomerular filtration rate (GFR) can be achieved in physiologic and AKI rat kidney models. Using a bolus infusion of a mixture of FITC-inulin and a 500-kDa Texas Red dextran, a full spectrum of GFR values, ranging from 0.17 to 1.12 ml·min−1·100 g−1, was obtained. The GFR values thus determined correlated well with values obtained by the standard 2-h inulin infusion clearance method with a Pearson's correlation coefficient of 0.85. In addition, postischemia deterioration was studied by measuring GFR using the two-photon approach during 24 h following a 45-min bilateral ischemia clamp model. The GFR was found to decline sharply during the initial 4 h followed by a nadir with little sign of rising over the ensuing 24-h period. Moreover, a FITC-labeled 5-kDa dextran was identified as having nearly identical filtration characteristics as FITC-inulin, but had markedly increased fluorescent intensity, thus minimizing the quantity needed for individual studies. The technique reported allows for very rapid GFR determinations, within 10–15 min, based on plasma clearance of a freely filtered fluorescence probe, instead of a prolonged one-compartment interstitial space reporter molecule clearance employed by other technologies.
Flexible Macroblock Ordering for Context-Aware Ultrasound Video Transmission over Mobile WiMAX
Authors: Martini Maria G. , Hewage Chaminda T. E. R.
Published: August 2010

The most recent network technologies are enabling a variety of new applications, thanks to the provision of increased bandwidth and better management of Quality of Service. Nevertheless, telemedical services involving multimedia data are still lagging behind, due to the concern of the end users, that is, clinicians and also patients, about the low quality provided. Indeed, emerging network technologies should be appropriately exploited by designing the transmission strategy focusing on quality provision for end users. Stemming from this principle, we propose here a context-aware transmission strategy for medical video transmission over WiMAX systems. Context, in terms of regions of interest (ROI) in a specific session, is taken into account for the identification of multiple regions of interest, and compression/transmission strategies are tailored to such context information. We present a methodology based on H.264 medical video compression and Flexible Macroblock Ordering (FMO) for ROI identification. Two different unequal error protection methodologies, providing higher protection to the most diagnostically relevant data, are presented.
Bridged Phthalocyanine Systems for Sensitization of Nanocrystalline TiO2Films
Authors: Zanotti Gloria , Angelini Nicola , Notarantonio Sara , Paoletti Anna Maria , Pennesi Giovanna , Rossi Gentilina , Lembo Angelo , Colonna Daniele , Di Carlo Aldo , Reale Andrea , Brown Thomas M. , Calogero Giuseppe
Published: August 2010

Phthalocyanines based-dyes represent attractive alternatives to the expensive and polluting pyridyl based Ru complexes because of their photochemical and thermal stability, they do show in fact intense absorption in the UV/blue (Soret band) and the red/near IR (Q band) spectral regions and appear very promising as sensitizer dyes for DSSC. In this contribution we review the state of the art and the recent progress in the application of these materials as dyes for DSSC and present three new dyes which are bridged derivatives of Iron phthalocyanine. Synthesis, optical properties, electrochemical characterization and device performances are discussed with regard to the different substitution degree of the macrocycle.
Epigenetic Regulatory Mechanisms Associated with Infertility
Authors: Minocherhomji Sheroy , Madon Prochi F. , Parikh Firuza R.
Published: August 2010

Infertility is a complex human condition and is known to be caused by numerous factors including genetic alterations and abnormalities. Increasing evidence from studies has associated perturbed epigenetic mechanisms with spermatogenesis and infertility. However, there has been no consensus on whether one or a collective of these altered states is responsible for the onset of infertility. Epigenetic alterations involve changes in factors that regulate gene expression without altering the physical sequence of DNA. Understanding these altered epigenetic states at the genomic level along with higher order organisation of chromatin in genes associated with infertility and pericentromeric regions of chromosomes, particularly 9 and Y, could further identify causes of idiopathic infertility. Determining the association between DNA methylation, chromatin state, and noncoding RNAs with the phenotype could further determine what possible mechanisms are involved. This paper reviews certain mechanisms of epigenetic regulation with particular emphasis on their possible role in infertility.
The Kinematics Analysis of a Novel 3-DOF Cable-Driven Wind Tunnel Mechanism
Authors: Yu Yao , Hongtao Wu
Published: August 2010

The kinematics analysis method of a novel 3-DOF wind tunnel mechanism based on cable-driven parallel mechanism is provided. Rodrigues' parameters are applied to express the transformation matrix of the wire-driven mechanism in the paper. The analytical forward kinematics model is described as three quadratic equations using three Rodridgues' parameters based on the fundamental theory of parallel mechanism. Elimination method is used to remove two of the variables, so that an eighth-order polynomial with one variable is derived. From the equation, the eight sets of Rodridgues' parameters and corresponding Euler angles for the forward kinematical problem can be obtained. In the end, numerical example of both forward and inverse kinematics is included to demonstrate the presented forward-kinematics solution method. The numerical results show that the method for the position analysis of this mechanism is effective.
Active Microstructured Optical Arrays of Grazing Incidence Reflectors
Authors: Willingale Richard , Feldman Charlotte , Michette Alan , Button Tim , Dunare Camelia , Folkard Melvyn , Hart David , McFaul Chris , Morrison Graeme R. , Parkes William , Pfauntsch Sławka , Powell A. Keith , Rodriguez-Sanmartin Daniel , Sahraei Shahin , Shand Matthew T. , Stevenson Tom , Vojnovic Boris , Zhang Dou
Published: August 2010

The UK Smart X-Ray Optics (SXO) programme is developing active/adaptive optics for terrestrial applications. One of the technologies proposed is microstructured optical arrays (MOAs), which focus X-rays using grazing incidence reflection through consecutive aligned arrays of microscopic channels. Although such arrays are similar in concept to polycapillary and microchannel plate optics, they can be bent and adjusted using piezoelectric actuators providing control over the focusing and inherent aberrations. Custom configurations can be designed, using ray tracing and finite element analysis, for applications from sub-keV to several-keV X-rays, and the channels of appropriate aspect ratios can be made using deep silicon etching. An exemplar application will be in the microprobing of biological cells and tissue samples using Ti Kα radiation (4.5 keV) in studies related to radiation-induced cancers. This paper discusses the optical design, modelling, and manufacture of such optics.
Opposing Effects of Omega-3 and Omega-6 Long Chain Polyunsaturated Fatty Acids on the Expression of Lipogenic Genes in Omental and Retroperitoneal Adipose Depots in the Rat
Authors: Muhlhausler B. S. , Cook-Johnson R. , James M. , Miljkovic D. , Duthoit E. , Gibson R.
Published: August 2010

This study aimed to determine the effect of varying dietary intake of the major n-3 PUFA in human diets,α-linolenic acid (ALA; 18 : 3n-3), on expression of lipogenic genes in adipose tissue. Rats were fed diets containing from 0.095%en to 6.3%en ALA and a constant n-6 PUFA level for 3 weeks. Samples from distinct adipose depots (omental and retroperitoneal) were collected and mRNA expression of the pro-lipogenic transcription factors Sterol-Retinoid-Element-Binding-Protein1c (SREBP1c) and Peroxisome Proliferator Activated Receptor-γ(PPARγ), lipogenic enzymes Sterol-coenzyme Desaturase1 (SCD-1), Fatty Acid Synthase (FAS), lipoprotein lipase (LPL) and glycerol-3-phosphate dehydrogenase (G3PDH) and adipokines leptin and adiponectin determined by qRT-PCR. Increasing dietary ALA content resulted in altered expression of SREBP1c, FAS and G3PDH mRNA in both adipose depots. SREBP1c mRNA expression was related directly to n-6 PUFA concentrations (omental,r2=.71;P<.001; Retroperitoneal,r2=.20;P<.002), and inversely to n-3 PUFA concentrations (omental,r2=.59;P<.001; Retroperitoneal,r2=.19;P<.005) independent of diet. The relationship between total n-6 PUFA and SREBP1c mRNA expression persisted when the effects of n-3 PUFA were controlled for. Altering red blood cell concentrations of n-3 PUFA is thus associated with altered expression of lipogenic genes in a depot-specific manner and this effect is modulated by prevailing n-6 PUFA concentrations.
Cognitive Deterioration Rates in Patients with Parkinson’s Disease from Northeastern China
Authors: Luo Xiao-Guang , Feng Yu , Liu Rong , Yu Hong-Mei , Wang Li , Wu Zhe , Ren Yan , He Zhi-Yi
Published: August 2010

Background/Aims: Cognitive impairment (CI) is common in Parkinson’s disease (PD), and the cognitive deterioration rate (CDR) is heterogeneous among PD patients. However, very few studies have reported on the association of PD features and risk factors with rapid CDR. The Montreal Cognitive Assessment (MoCA) is considered to be a sensitive and reliable approach to detect mild CI. In the present study, we sought to define and compare the cognitive profiles and clinical features of PD patients with slow or rapid CDRs, and then to identify the PD risk factors associated with rapid CDR. Methods: A cross-sectional study of cognitive rate was performed using the MoCA in a cohort of 73 PD patients and 41 controls matched for age, sex and education level. Results: The rapid CDR group was characterized by older age (58.8 years in slow CDR vs. 64.1 in rapid CDR; p = 0.02), later age at disease onset (52.7 vs. 61.7 years; p < 0.001), a faster deterioration rate of movement symptoms (UPDRS III increment of 12.8 vs. 5.9/year; p < 0.001), a higher rate in multiple-domain CI (38.9 vs. 10.8%), and generally lower MoCA subscores for the Clock Drawing Test, attention, verbal fluency and abstraction. According to the univariate logistic regression model, onset age, movement deterioration rate, multiple domains CI and executive function CI were risk factors for rapid CDR. However, only the movement deterioration rate (p = 0.01) and onset age (p = 0.05) remained independent predictors for rapid CDR according to the multivariate logistic regression model. Conclusions: The CI deterioration in a subset of PD patients appears to progress more rapidly. Identifying those PD patients may not only help to predict the development of PD dementia, but also facilitate therapeutic intervention at early disease stages.