bluuebunny/crossref_metadata_2025 huggingface dataset:
Molecular cloning, characterization, and heterologous expression analysis of heat shock protein genes (hsp70 and hsp90) of the invasive alien weed, Ageratina adenophora (Asteraceae)
Authors: GONG WEI NA , XIE BING YAN , WAN FANG HAO , GUO JIAN YING
Published: June 2010

An invasive alien weed, Ageratina adenophora, has become widespread in the southwest of China due to its high environmental adaptation. In the present study, in order to understand the molecular aspects of resistance to heat and cold stress, we investigated the characterization and expression of heat shock protein genes (designated as Aahsp70 and Aahsp90) in A. adenophora. The cDNA of A. adenophora Aahsp70 and Aahsp90 was cloned by the combination of homology cloning and rapid amplification of cDNA ends (RACE). The Basic Local Alignment Search Tool analyses revealed that Aahsp70 and Aahsp90 shared high levels of identity with corresponding proteins from other species. Hybridization was employed to examine the expression patterns of Aahsp70 and Aahsp90. The relative expression levels of the Aahsp70 and Aahsp90 transcripts were both up‐regulated and reached maximal levels at 2–4 h after heat shock. Then, Aahsp70 dropped progressively to the original level at ∼4 h after heat shock. The transcriptional changes of Aahsp70 and Aahsp90 both were more obvious under cold stress. Recombinant Aahsp70 and Aahsp90 improved the viability, in comparison with the control cultures, of Escherichia coli under stress conditions. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis analysis of the cell lysates suggested that the protective effect in vivo was related to increased thermo‐stability of the soluble cytosolic proteins. This study emphasizes that the relationship between, and regulation of, heat shock proteins might incorporate variable strengths to increase the adaptation of A. adenophora.
Ecological pest management and control by using allelopathic weeds (Ageratum conyzoides, Ambrosia trifida, and Lantana camara) and their allelochemicals in China
Authors: KONG CHUI HUA
Published: June 2010

The development of pest management and control is striving toward a future of sustainable agriculture. Weeds cause serious problems in agricultural ecosystems and attempts to control them have met with limited success. However, many weeds are allelopathic; that is, they can produce and release allelochemicals to interact with other plant competitors and to attack microbes or insect and other animal predators. These allelopathic weeds and their allelochemicals may be put into use for ecological pest management and control or employed for other uses. Currently, little attention has been paid to how allelopathic weeds and their allelochemicals potentially can be utilized as an important part of pest management and control in agricultural ecosystems. This review outlines recent research regarding the potential for pest management and control by allelopathic weeds and their allelochemicals by studying the cases of Ageratum conyzoides, Ambrosia trifida, and Lantana camara and provides examples of allelopathic weeds and their allelochemicals that have been incorporated into ecological pest management and control in China.
Demography of Digitaria sanguinalis: Effect of the emergence time on survival, reproduction, and biomass
Authors: GALLART MONTSERRAT , MAS M. TERESA , VERDÚ ANTONI M.C.
Published: June 2010

Knowledge of weed population dynamics is crucial for designing effective weed management practises. A field experiment was carried out at Torre Marimon (near Barcelona, north‐eastern Spain) in 2006 and 2007 in order to study the effects of the seedling emergence time on Digitaria sanguinalis fitness. Three and four cohorts were monitored in 2006 and 2007, respectively. In 2007, the seedling density was fourfold greater than in 2006. At the end of the growing season, no statistically significant differences in survival were detected among the cohorts in 2006 (the mean survival rate was 88%); in contrast, in 2007, individual survival varied depending on the cohort, ranging from 25–88%. The reproductive traits and seed production were significantly influenced by the emergence time. In the cohorts that emerged very close in time, the individuals of the first cohort on average had more than threefold the amount of seed production than those of the last cohort. However, not all the surviving individuals were able to reproduce because they were infected by the smut, Ustilago syntherismae.
Melatonin potentiates glycine currents through a PLC/PKC signalling pathway in rat retinal ganglion cells
Authors: Zhao Wen‐Jie , Zhang Min , Miao Yanying , Yang Xiong‐Li , Wang Zhongfeng
Published: June 2010

In vertebrate retina, melatonin regulates various physiological functions. In this work we investigated the mechanisms underlying melatonin‐induced potentiation of glycine currents in rat retinal ganglion cells (RGCs). Immunofluorescence double labelling showed that rat RGCs were solely immunoreactive to melatonin MT2receptors. Melatonin potentiated glycine currents of RGCs, which was reversed by the MT2receptor antagonist 4‐P‐PDOT. The melatonin effect was blocked by intracellular dialysis of GDP‐β‐S. Either preincubation with pertussis toxin or application of the phosphatidylcholine (PC)‐specific phospholipase C (PLC) inhibitor D609, but not the phosphatidylinositol (PI)‐PLC inhibitor U73122, blocked the melatonin effect. The protein kinase C (PKC) activator PMA potentiated the glycine currents and in the presence of PMA melatonin failed to cause further potentiation of the currents, whereas application of the PKC inhibitor bisindolylmaleimide IV abolished the melatonin‐induced potentiation. The melatonin effect persisted when [Ca2+]iwas chelated by BAPTA, and melatonin induced no increase in [Ca2+]i. Neither cAMP‐PKA nor cGMP‐PKG signalling pathways seemed to be involved because 8‐Br‐cAMP or 8‐Br‐cGMP failed to cause potentiation of the glycine currents and both the PKA inhibitor H‐89 and the PKG inhibitor KT5823 did not block the melatonin‐induced potentiation. In consequence, a distinct PC‐PLC/PKC signalling pathway, following the activation of Gi/o‐coupled MT2receptors, is most likely responsible for the melatonin‐induced potentiation of glycine currents of rat RGCs. Furthermore, in rat retinal slices melatonin potentiated light‐evoked glycine receptor‐mediated inhibitory postsynaptic currents in RGCs. These results suggest that melatonin, being at higher levels at night, may help animals to detect positive or negative contrast in night vision by modulating inhibitory signals largely mediated by glycinergic amacrine cells in the inner retina.
Atrial natriuretic peptide inhibits angiotensin II‐stimulated proliferation in fetal cardiomyocytes
Authors: O’Tierney P. F. , Chattergoon N. N. , Louey S. , Giraud G. D. , Thornburg K. L.
Published: June 2010

The role of atrial natriuretic peptide (ANP) in regulating fetal cardiac growth is poorly understood. Angiotensin II (Ang II) stimulates proliferation in fetal sheep cardiomyocytes when growth is dependent on the activity of the mitogen‐activated protein kinase (MAPK) and phosphoinositol‐3‐kinase (PI3K) pathways. We hypothesized that ANP would suppress near‐term fetal cardiomyocyte proliferation in vitro and inhibit both the MAPK and PI3K pathways. Forty‐eight hour 5‐bromodeoxyuridine (BrdU) uptake (used as an index of proliferation) was measured in cardiomyocytes isolated from fetal sheep (135 day gestational age) in response to 100 nm Ang II with or without ANP (0.003–100 nm) or 1 μm 8‐bromo‐cGMP. The effects of these compounds on the MAPK and PI3K pathways were assessed by measuring extracellular signal‐regulated kinase (ERK) and AKT phosphorylation following 10 min of treatment with Ang II, ANP or 8‐bromo‐cGMP. In right ventricular myocytes (RV), the lowest dose of ANP (0.003 nm) inhibited Ang II‐stimulated BrdU uptake by 68%. Similarly, 8‐bromo‐cGMP suppressed Ang II‐stimulated proliferation by 62%. The same effects were observed in left ventricular (LV) cardiomyoytes but the RV was more sensitive to the inhibitory effects of ANP than the LV (P < 0.0001). Intracellular cGMP was increased by 4‐fold in the presence of 100 nm ANP. Ang II‐stimulated ERK and Akt phosphorylation was inhibited by 100 nm ANP. The activity of ANP may in part be cGMP dependent, as 8‐bromo‐cGMP had similar effects on the cardiomyocytes.
A Modified Efficient Global Optimization Algorithm for Maximal Reliability in a Probabilistic Constrained Space
Authors: Huang Yen-Chih , Chan Kuei-Yuan
Published: June 2010

Design optimization problems under random uncertainties are commonly formulated with constraints in probabilistic forms. This formulation, also referred to as reliability-based design optimization (RBDO), has gained extensive attention in recent years. Most researchers assume that reliability levels are given based on past experiences or other design considerations without exploring the constrained space. Therefore, inappropriate target reliability levels might be assigned, which either result in a null probabilistic feasible space or performance underestimations. In this research, we investigate the maximal reliability within a probabilistic constrained space using modified efficient global optimization (EGO) algorithm. By constructing and improving Kriging models iteratively, EGO can obtain a global optimum of a possibly disconnected feasible space at high reliability levels. An infill sampling criterion (ISC) is proposed to enforce added samples on the constraint boundaries to improve the accuracy of probabilistic constraint evaluations via Monte Carlo simulations. This limit state ISC is combined with the existing ISC to form a heuristic approach that efficiently improves the Kriging models. For optimization problems with expensive functions and disconnected feasible space, such as the maximal reliability problems in RBDO, the efficiency of the proposed approach in finding the optimum is higher than those of existing gradient-based and direct search methods. Several examples are used to demonstrate the proposed methodology.
Modeling, Prototyping, and Testing of Helical Shape Memory Compression Springs With Hollow Cross Section
Authors: Spinella Igor , Dragoni Eugenio , Stortiero Francesco
Published: June 2010

Shape memory alloys (SMAs) are used in many applications as actuators. The main drawbacks that limit the use of the SMAs in the field of mechanical actuation are the low mechanical bandwidth (up to a few Hertzs) and the unsatisfactory stroke (several millimeters). This paper contributes to enhancing the performances of SMA actuators by proposing a new SMA helical spring with a hollow section. The hollow spring is modeled, then it is constructed, and finally it is tested in compression to compare its performances with those of a spring with a solid cross section of equal stiffness and strength. Emptied of the inefficient material from its center, the hollow spring features a lower mass (37% less) and an extremely lower cooling time (four times less) than its solid counterpart. These results demonstrate that helical springs with a hollow construction can be successfully exploited to build SMA actuators for higher operating frequencies and improved strokes.
The Thermodynamics of the Elusive HO
3
Radical
Authors: Le Picard Sébastien D. , Tizniti Meryem , Canosa André , Sims Ian R. , Smith Ian W. M.
Published: June 2010

The Weakness of HO
3

OH radicals play a critical role in the chemistry of Earth's atmosphere. Understanding atmospheric reaction networks thus requires an accurate knowledge of OH sources and sinks. One vexing question has been whether or not a significant pool of OH binds temporarily with oxygen to form HO
3
.

Le Picard
et al.

(p.
1258
) have succeeded in measuring the equilibrium constant for this reaction using sensitive fluorescent tracking of OH in a laboratory apparatus. This measurement was then used to quantify the strength of the O
2
–OH bond, which was found to be too weak for the complexation to play a major role in the atmosphere.
Meiotic Recombination Provokes Functional Activation of the p53 Regulatory Network
Authors: Lu Wan-Jin , Chapo Joseph , Roig Ignasi , Abrams John M.
Published: June 2010

Germline Quality Control

The p53 tumor suppressor protein plays a key role in protecting organisms from aberrant cancer cells. But during evolution, animals would rarely, if ever, have lived long enough to develop cancer and so need such a function of p53. What, then, were the original functions for which p53 was selected?

Lu
et al.

(p.
1278
) observed a pulse of p53 activation during
Drosophila
development in the female germ line. In cancer, p53 is activated in response to DNA damage. Similarly, in this study, breaks in DNA that occur normally during meiosis also caused p53 activation. In animals in which resolution of DNA breaks during crossing over was inhibited, activation of p53 was prolonged; furthermore, when p53 was also lacking oogenesis was abnormal. Exactly how activation of p53 contributes to the process of chromosome recombination during meiosis remains unclear, but it may provide quality control, only allowing survival of gametes that possess intact DNA.
Induction of Fear Extinction with Hippocampal-Infralimbic BDNF
Authors: Peters Jamie , Dieppa-Perea Laura M. , Melendez Loyda M. , Quirk Gregory J.
Published: June 2010

Pharmacological Forgetting

Extinction of conditioned fear forms a new memory in the infralimbic medial prefrontal cortex that is critical for subsequent retrieval of the extinction memory. Understanding the mechanisms that underlie this extinction-related plasticity could help in the treatment of anxiety disorders. By infusing BDNF (brain-derived neurotrophic factor) into the infralimbic cortex,

Peters
et al.

(p.
1288
) caused the extinction of conditioned fear, even without an extinction trial. In fact, BDNF infusion seemed to act as if an extinction training session had been given. Thus, the hippocampus is a likely source of the BDNF input to the infralimbic cortex, and individual differences in extinction memory may reflect variations in hippocampal BDNF content.
Natural and Sexual Selection in a Wild Insect Population
Authors: Rodríguez-Muñoz R. , Bretman A. , Slate J. , Walling C. A. , Tregenza T.
Published: June 2010

Insects in the Wild

Insects are of fundamental importance to terrestrial ecosystems and provide laboratory model systems for the study of physiology and genetics. Studies examining how natural and sexual selection operate to drive evolution in wild populations have often neglected invertebrates, resulting in a chasm between our understandings of how things work in the lab versus the natural environment.

Rodríguez-Muñoz
et al.

(p.
1269
; see the Perspective by

Zuk

) bridge this gap by comprehensively monitoring the life histories, behavior, and reproductive success of an entire population of field crickets. Adding genetic data allowed evaluation of how behavior impacts reproductive success and confirmed that male reproductive success varies more than that of females.
Dynamic Kinetic Resolution of Biaryl Atropisomers via Peptide-Catalyzed Asymmetric Bromination
Authors: Gustafson Jeffrey L. , Lim Daniel , Miller Scott J.
Published: June 2010

Selectively Spun

Biaryl compounds, in which two phenyl rings are linked by a single bond, exhibit an interesting sort of chirality, termed atropisomerism. If bulky substituents block the mutual rotation of the rings about the linking bond, then two isomers can be isolated that differ only in the direction one ring has swiveled away from the plane of the other. This feature is useful in ligand design for asymmetric catalysis and also appears in a number of polycyclic natural products. However, selective synthesis of a single isomer is difficult.

Gustafson
et al.

(p.
1251
) now show that a simple tripeptide derivative acts as an efficient catalyst for this purpose, trapping a freely rotating precursor in one orientation through selective bromination; the large bromine substituents then inhibit further swiveling.
Asymmetrical Distribution of the Second Messenger c-di-GMP upon Bacterial Cell Division
Authors: Christen Matthias , Kulasekara Hemantha D. , Christen Beat , Kulasekara Bridget R. , Hoffman Lucas R. , Miller Samuel I.
Published: June 2010

Keeping Tabs on Second-Messenger Localization

The prokaryotic second-messenger cyclic diguanosine monophosphate (c-di-GMP) is a global bacterial signaling molecule that controls the switch between motile or planktonic life-styles and sedentary or adhesive life-styles. Regulation by this second messenger has been associated with virulence traits, biofilm formation, and antibiotic tolerance.

Christen
et al.

(p.
1295
) engineered fluorescence resonance energy transfer–based sensors that enabled the visualization of c-di-GMP fluctuations in individual bacterial cells. Using the sensor, c-di-GMP distribution was found to change during the cell cycle in
Caulobacter crescentus
and
Pseudomonas aeruginosa
cells.
Top-Down Versus Bottom-Up
Authors: Nisbet Euan , Weiss Ray
Published: June 2010

Can national emissions inventories be verified through direct atmospheric measurements?
Along the Indus River, Saber Rattling Over Water Security
Authors: Bagla Pallava
Published: June 2010

Pakistan accuses India of contravening a treaty that governs Indus water; with climate change likely to make things tenser, better data sharing may forge mutual trust.
Importance of Antirabies Revaccination for an Adequate Antirabies Protection in Bovine Newborns
Authors: Filho O. A. , Megid J. , Geronutti L. , Ratti J. , Kataoka A. P. A. G. , Martorelli L. F. A.
Published: June 2010

The transfer of antirabies immunoglobulins in cows that were prime vaccinated and cows that were revaccinated against rabies correlated to the serum titers in their offspring was evaluated. The results demonstrated that revaccination against rabies during pregnancy induces neutralizing antibody titers at a protective level that are transferred directly to calves through colostrum and reinforce the importance of revaccination for improved colostral antibody transfer and offspring protection against rabies.
Antigen Specificity of the Humoral Immune Response toMycoplasma haemofelisInfection
Authors: Peters Iain R. , Helps Chris R. , Gruffydd-Jones Timothy J. , Day Michael J. , Tasker Séverine
Published: June 2010

The aim of the present study was to characterize the antigenic specificity of the humoral immune response made by cats infected with the feline hemoplasma,Mycoplasma haemofelis.A crudeM. haemofelisantigen preparation was prepared from red blood cells (RBCs) collected from a cat at the time of a high level of bacteremia. Plasma samples were collected from six cats before and after experimental infection withM. haemofelis, with regular sampling being performed from 15 to 149 or 153 days postinfection (dpi). Preinfection RBC membrane ghosts were prepared from these six cats and used to identify erythrocyte proteins that may have contaminated theM. haemofelisantigen preparation. TheM. haemofelisantigen preparation comprised 11 protein bands. The immunodominant bands on Western blotting with infected cat plasma had molecular masses of 78, 68, 60, 48, and 38 kDa. Most cats (n= 5) had plasma antibody that reacted with at least one band (always including the one of 68 kDa) at 15 dpi, and all cats were seroreactive by 29 dpi. The maximum number of antibodies from an individual animal specific for an antigen was identified in plasma collected from 57 to 99 dpi. Contamination of theM. haemofelisantigen preparation with RBC membrane proteins was observed. The contaminating RBC proteins had molecular masses of from 71 to 72 kDa (consistent with band 4.2) and 261 and 238 kDa (consistent with spectrin), and these were recognized by all plasma samples. A range ofM. haemofelisantigens is recognized by cats infected experimentally with the organism. These represent possible targets for immunoassays, but care must be taken to prevent false-positive results due to host protein contamination.
Resistance of
Acanthamoeba
Cysts to Disinfection Treatments Used in Health Care Settings
Authors: Coulon Céline , Collignon Anne , McDonnell Gerald , Thomas Vincent
Published: June 2010

Free-living amoebae that belong to the genus
Acanthamoeba
are widespread in the environment, including water. They are responsible for human infections and can host pathogenic microorganisms. Under unfavorable conditions, they form cysts with high levels of resistance to disinfection methods, thus potentially representing a threat to public health. In the present study we evaluated the efficacies of various biocides against trophozoites and cysts of several
Acanthamoeba
strains. We demonstrated that disinfectant efficacy varied depending on the strains tested, with environmental strains demonstrating greater resistance than collection strains. Trophozoites were inactivated by all treatments except those using glutaraldehyde as an active compound: for these treatments, we observed resistance even after 30 min exposure. Cysts resisted many treatments, including certain conditions with glutaraldehyde and other biocides. Moist heat at 55°C was not efficient against cysts, whereas exposure at 65°C was. Several chemical formulations containing peracetic acid, hydrogen peroxide, or
ortho
-phthalaldehyde presented greater efficacy than glutaraldehyde, as did ethanol and sodium hypochlorite; however, some of these treatments required relatively long incubation times to achieve cyst inactivation. Amoebal cysts can be highly resistant to some high-level disinfectants, which has implications for clinical practice. These results highlight the need to consider the effective disinfection of protozoa in their vegetative and resistant forms due to their intrinsic resistance. This is important not only to prevent the transmission of protozoa themselves but also due to the risks associated with a range of microbial pathogens that are found to be associated intracellularly with these microorganisms.
Meningitis Caused by
Escherichia coli
Producing TEM-52 Extended-Spectrum β-Lactamase within an Extensive Outbreak in a Neonatal Ward: Epidemiological Investigation and Characterization of the Strain
Authors: Moissenet Didier , Salauze Béatrice , Clermont Olivier , Bingen Edouard , Arlet Guillaume , Denamur Erick , Mérens Audrey , Mitanchez Delphine , Vu-Thien Hoang
Published: June 2010

Outbreaks caused by
Enterobacteriaceae
isolates producing extended-spectrum β-lactamases (ESBL) in neonatal wards can be difficult to control. We report here an extensive outbreak in a neonatal ward with a case of meningitis caused by an ESBL-producing
Escherichia coli
strain. Between 24 March and 29 April 2009, among the 59 neonates present in the ward, 26 neonates with ESBL-producing
E
.
coli
rectal colonization were detected (44%). One of the colonized neonates developed meningitis with a favorable outcome after treatment combining imipenem, gentamicin, and ciprofloxacin. Despite strict intensification of hygiene and isolation procedures for more than 1 month, ward closure to new admissions was necessary to control the outbreak. Randomly amplified polymorphic DNA and pulsed-field gel electrophoresis analysis performed on 31 isolates recovered from 26 neonates and two mother's milk samples showed a clonal strain. ESBL PCR assays indicated that the strain harbored a TEM-52 ESBL encoded by an IncI1 replicon. Phylogenetic analysis by multilocus sequence typing showed that the strain belonged to rare phylogenetic group C, which is closely related to group B1 but appears as group A by the triplex PCR phylogrouping method. The strain harbored the virulence genes
fuyA
,
aer
, and
iroN
and was virulent in a mouse model of septicemia. This work indicates the high potential of colonization, transmission, and virulence of some ESBL-producing
E
.
coli
clones.
Detection of “
Candidatus
Neoehrlichia mikurensis” in Two Patients with Severe Febrile Illnesses: Evidence for a European Sequence Variant
Authors: von Loewenich Friederike D. , Geißdörfer Walter , Disqué Claudia , Matten Jens , Schett Georg , Sakka Samir G. , Bogdan Christian
Published: June 2010

Recently, a new genus of
Anaplasmataceae
termed “
Candidatus
Neoehrlichia” was discovered in ticks and rodents. Here, we report on two patients who suffered from febrile bacteremia due to “
Candidatus
Neoehrlichia mikurensis” associated with thrombotic or hemorrhagic events. 16S rRNA and
groEL
gene sequencing provided evidence of three groups of sequence variants.
Identification ofPaecilomyces variotiiin Clinical Samples and Settings
Authors: Houbraken Jos , Verweij Paul E. , Rijs Anthonius J. M. M. , Borman Andrew M. , Samson Robert A.
Published: June 2010

Paecilomyces variotiiis a commonly occurring species in air and food, but it is also associated with many types of human infections and is among the emerging causative agents of opportunistic mycoses in immunocompromised hosts.Paecilomycescan cause hyalohyphomycosis, and two species,Paecilomyces lilacinusandP. variotii, are the most frequently encountered organisms. In the present study, a set of 34 clinical isolates morphologically identified asP. variotiiorP. lilacinuswere formally identified by sequencing intergenic transcribed spacer regions 1 and 2 (including 5.8S rDNA) and a part of the β-tubulin gene. Three isolates were identified asP. lilacinus, and five of the presumptiveP. variotiiisolates did not belong to the genusPaecilomycesbut were identified asTalaromyces eburneus(anamorph,Geosmithia argillacea) orHamigera avellanea(anamorph,Merimbla ingelheimense). Applying the most recent taxonomy, we found that the clinicalP. variotiiisolates could be identified asP. variotiisensu stricto (14 strains),P. formosus(11 strains), andP. dactylethromorphus(1 strain). These data indicate thatP. formosusoccurs in clinical samples as commonly asP. variotii. Susceptibility tests showed that the antifungal susceptibility profiles ofP. variotii,P. formosus, andP. dactylethromorphusare similar and that all strains tested were susceptible to amphotericin Bin vitro.P. lilanicus,T. eburneus, andH. avellaneahad different susceptibility profiles; and flucytosine and voriconazole were the least active of the antifungal drugs tested against these species. Our results indicate that correct species identification is important to help guide appropriate antifungal therapy.
Rapid Identification of Methicillin-Resistant
Staphylococcus aureus
Transmission in Hospitals by Use of Phage-Derived Open Reading Frame Typing Enhanced by Multiplex PCR and Reverse Line Blot Assay
Authors: O'Sullivan Matthew V. N. , Kong Fanrong , Sintchenko Vitali , Gilbert Gwendolyn L.
Published: June 2010

The relatively high-level clonality of methicillin-resistant
Staphylococcus aureus
(MRSA) and its frequent high-level endemicity in nosocomial settings complicate the development of methods for rapid subtyping of MRSA strains that are capable of identifying person-to-person transmission in hospitals. Phage-derived open reading frame (PDORF) typing is an MRSA typing method targeting mobile genetic elements that was recently described and evaluated using a geographically restricted set of isolates. The objective of this study was to develop a multiplex PCR-reverse line blot (mPCR/RLB) assay for PDORF typing and to test its applicability on a broad range of isolates and in an environment where MRSA is highly endemic. The 16 targets were identified using a 23-primer-pair mPCR/RLB assay with two probes for each target. The method was evaluated using 42 MRSA reference strains, including those representing major international clones, and 35 isolates from episodes of suspected nosocomial transmission.
In vivo
stability was explored using 81 isolate pairs. Pulsed-field gel electrophoresis (PFGE) and
spa
typing were performed for comparison. Among the 42 reference strains, there were 33 PFGE subtypes, 30 PDORF types, and 22
spa
types. Simpson's index of diversity was 0.987, 0.971, and 0.926 for PFGE subtyping, PDORF typing, and
spa
typing, respectively. Typing of clinical isolates by PDORF typing and PFGE demonstrated concordant results. mPCR/RLB-based PDORF typing has similar discriminatory power to that of PFGE, can assist in tracking MRSA transmission events in a setting of high-level endemicity, and has the advantage of being a high-throughput technique.
Subcutaneous
Mycoleptodiscus indicus
Infection in an Immunosuppressed Dog
Authors: Metry Catherine A. , Hoien-Dalen Patricia S. , Maddox Carol W. , Thompson Elizabeth H. , Sutton Deanna A. , Romanelli Anna M. , Wickes Brian L. , MacNeill Amy L.
Published: June 2010

An 8-year-old dog presented with several dermal excoriations. Lesion cytology revealed pyogranulomatous inflammation with branching, septate hyphae. A mold identified as
Mycoleptodiscus indicus
by morphology and sequencing was cultured from fine-needle aspirates. This is the first report of a
Mycoleptodiscus
species as an etiologic agent in a dog.
Recovery of Replication-Competent Residual HIV-1 from Plasma of a Patient Receiving Prolonged, Suppressive Highly Active Antiretroviral Therapy
Authors: Sahu Gautam K. , Sarria Juan C. , Cloyd Miles W.
Published: June 2010

The clinical significance of persistent residual viremia in patients on prolonged highly active antiretroviral therapy (HAART) is not clear. Moreover, it remains to be demonstrated whether residual viremia consists of viruses capable of spreading infection
in vivo
upon termination of therapy. Using residual viral RNAs (vRNAs) isolated from a HAART-treated patient's plasma, we cloned full-length viral genomes and found that most of them could produce infectious, replication-competent HIVs when transfected into TZM-bl cells, suggesting that residual viruses produced in the absence of therapy can initiate fresh cycles of infection and spread in host cells. The data further indicate that residual viremia may pose a major concern with regard to the emergence of drug-resistant HIVs during periods of low adherence to therapy.
Role of Noncovalent SUMO Binding by the Human Cytomegalovirus IE2 Transactivator in Lytic Growth
Authors: Kim Eui Tae , Kim Young-Eui , Huh Yong Ho , Ahn Jin-Hyun
Published: June 2010

The 86-kDa immediate-early 2 (IE2) protein of human cytomegalovirus (HCMV) is a promiscuous transactivator essential for viral gene expression. IE2 is covalently modified by SUMO at two lysine residues (K175 and K180) and also interacts noncovalently with SUMO. Although SUMOylation of IE2 has been shown to enhance its transactivation activity, the role of SUMO binding is not clear. Here we showed that SUMO binding by IE2 is necessary for its efficient transactivation function and for viral growth. IE2 bound physically to SUMO-1 through a SUMO-interacting motif (SIM). Mutations in SIM (mSIM) or in both SUMOylation sites and SIM (KR/mSIM), significantly reduced IE2 transactivation effects on viral early promoters. The replication of IE2 SIM mutant viruses (mSIM or KR/mSIM) was severely depressed in normal human fibroblasts. Analysis of viral growth curves revealed that the replication defect of the mSIM virus correlated with low-level accumulation of SUMO-modified IE2 and of viral early and late proteins. Importantly, both the formation of viral transcription domains and the association of IE2 with viral promoters in infected cells were significantly reduced in IE2 SIM mutant virus infection. Furthermore, IE2 was found to interact with the SUMO-modified form of TATA-binding protein (TBP)-associated factor 12 (TAF12), a component of the TFIID complex, in a SIM-dependent manner, and this interaction enhanced the transactivation activity of IE2. Our data demonstrate that the interaction of IE2 with SUMO-modified proteins plays an important role for the progression of the HCMV lytic cycle, and they suggest a novel viral mechanism utilizing the cellular SUMO system.
Limited Contribution of Mucosal IgA to Simian Immunodeficiency Virus (SIV)-Specific Neutralizing Antibody Response and Virus Envelope Evolution in Breast Milk of SIV-Infected, Lactating Rhesus Monkeys
Authors: Permar Sallie R. , Wilks Andrew B. , Ehlinger Elizabeth P. , Kang Helen H. , Mahlokozera Tatenda , Coffey Rory T. , Carville Angela , Letvin Norman L. , Seaman Michael S.
Published: June 2010

Breast milk transmission of human immunodeficiency virus (HIV) remains an important mode of infant HIV acquisition. Interestingly, the majority of infants remain uninfected during prolonged virus exposure via breastfeeding, raising the possibility that immune components in milk prevent mucosal virus transmission. HIV-specific antibody responses are detectable in the milk of HIV-infected women and simian immunodeficiency virus (SIV)-infected monkeys; however, the role of these humoral responses in virus neutralization and local virus quasispecies evolution has not been characterized. In this study, four lactating rhesus monkeys were inoculated with SIVmac251 and monitored for SIV envelope-specific humoral responses and virus evolution in milk and plasma throughout infection. While the kinetics and breadth of the SIV-specific IgG and IgA responses in milk were similar to those in plasma, the magnitude of the milk responses was considerably lower than that of the plasma responses. Furthermore, a neutralizing antibody response against the inoculation virus was not detected in milk samples at 1 year after infection, despite a measurable autologous neutralizing antibody response in plasma samples obtained from three of four monkeys. Interestingly, while IgA is the predominant immunoglobulin in milk, the milk SIV envelope-specific IgA response was lower in magnitude and demonstrated more limited neutralizing capacity against a T-cell line-adapted SIV compared to those of the milk IgG response. Finally, amino acid mutations in the envelope gene product of SIV variants in milk and plasma samples occurred in similar numbers and at similar positions, indicating that the humoral immune pressure in milk does not drive distinct virus evolution in the breast milk compartment.
A Nine-Segment Influenza A Virus Carrying Subtype H1 and H3 Hemagglutinins
Authors: Gao Qinshan , Lowen Anice C. , Wang Taia T. , Palese Peter
Published: June 2010

Influenza virus genomic RNAs possess segment-specific packaging signals that include both noncoding regions (NCRs) and adjacent terminal coding region sequences. Using reverse genetics, an A/Puerto Rico/8/34 (A/PR/8/34) virus was rescued that contained a modified PB1 gene such that the PB1 packaging sequences were exchanged for those of the neuraminidase (NA) gene segment. To accomplish this, the PB1 open reading frame, in which the terminal packaging signals were inactivated by serial synonymous mutations, was flanked by the NA segment-specific packaging sequences including the NCRs and the coding region packaging signals. Next, the ATGs located on the 3′ end of the NA packaging sequences of the resulting PB1 chimeric segment were mutated to allow for correct translation of the full-length PB1 protein. The virus containing this chimeric PB1 segment was viable and able to stably carry a ninth, green fluorescent protein (GFP), segment flanked by PB1 packaging signals. Utilizing this method, we successfully generated an influenza virus that contained the genes coding for both the H1 hemagglutinin (HA) from A/PR/8/34 and the H3 HA from A/Hong Kong/1/68 (A/HK/1/68); both subtypes of HA protein were also incorporated into the viral envelope. Immunization of mice with this recombinant virus conferred complete protection from lethal challenge with recombinant A/PR/8/34 virus and with X31 virus that expresses the A/HK/1/68 HA and NA. Using the described methodology, we show that a ninth segment can also be incorporated by manipulation of the PB2 or PA segment-specific packaging signals. This approach offers a means of generating a bivalent influenza virus vaccine.
The Localization of APOBEC3H Variants in HIV-1 Virions Determines Their Antiviral Activity
Authors: Ooms Marcel , Majdak Susan , Seibert Christopher W. , Harari Ariana , Simon Viviana
Published: June 2010

Several members of the human APOBEC3 family of cytidine deaminases can potently restrict retroviruses such as HIV-1. The single-domain APOBEC3H (A3H) is encoded by four haplotypes, of which only A3H haplotype II-RDD (hapII-RDD) restricts HIV-1 efficiently. The goal of this study was to elucidate the mechanisms underlying the differences in antiviral activity among A3H haplotypes. The naturally occurring A3H hapI-GKE and hapII-RDD variants differ at three amino acid positions. A panel of six site-directed mutants containing combinations of the three variable residues was used to determine A3H protein expression, requirements of A3H virion incorporation, and A3H-Gag interactions. The catalytic activity of each A3H protein was assessed directly by using an
Escherichia coli
mutator assay. We found that the incorporation efficiencies of A3H variants into HIV-1 virions were comparable despite major differences in cellular expression. An assessment of the enzymes' catalytic activities showed that the deaminase activity of each A3H variant correlated with protein expression, suggesting similar enzymatic efficiencies. Surprisingly, virion incorporation experiments using Gag deletion mutants demonstrated that A3H haplotypes interacted with different Gag regions. A3H hapII-RDD associated with nucleocapsid in an RNA-dependent manner, whereas A3H hapI-GKE associated with the C-terminal part of matrix and the N-terminal capsid domain. Our results show that the A3H hapII-RDD interaction with nucleocapsid is critical for its antiviral activity and that the inability of A3H hapI-GKE to interact with nucleocapsid underlies its limited antiviral potential. Thus, the antiviral activity of A3H haplotypes is determined by its incorporation into the viral core, in proximity to the reverse transcription complex.
A Stretch of 11 Amino Acids in the βB-βC Loop of the Coat Protein of
Grapevine Fanleaf Virus
Is Essential for Transmission by the Nematode
Xiphinema index
Authors: Schellenberger Pascale , Andret-Link Peggy , Schmitt-Keichinger Corinne , Bergdoll Marc , Marmonier Aurélie , Vigne Emmanuelle , Lemaire Olivier , Fuchs Marc , Demangeat Gérard , Ritzenthaler Christophe
Published: June 2010

Grapevine fanleaf virus
(GFLV) and
Arabis mosaic virus
(ArMV) from the genus
Nepovirus
, family
Secoviridae
, cause a severe degeneration of grapevines. GFLV and ArMV have a bipartite RNA genome and are transmitted specifically by the ectoparasitic nematodes
Xiphinema index
and
Xiphinema diversicaudatum
, respectively. The transmission specificity of both viruses maps to their respective RNA2-encoded coat protein (CP). To further delineate the GFLV CP determinants of transmission specificity, three-dimensional (3D) homology structure models of virions and CP subunits were constructed based on the crystal structure of
Tobacco ringspot virus
, the type member of the genus
Nepovirus
. The 3D models were examined to predict amino acids that are exposed at the external virion surface, highly conserved among GFLV isolates but divergent between GFLV and ArMV. Five short amino acid stretches that matched these topographical and sequence conservation criteria were selected and substituted in single and multiple combinations by their ArMV counterparts in a GFLV RNA2 cDNA clone. Among the 21 chimeric RNA2 molecules engineered, transcripts of only three of them induced systemic plant infection in the presence of GFLV RNA1. Nematode transmission assays of the three viable recombinant viruses showed that swapping a stretch of (i) 11 residues in the βB-βC loop near the icosahedral 3-fold axis abolished transmission by
X. index
but was insufficient to restore transmission by
X. diversicaudatum
and (ii) 7 residues in the βE-αB loop did not interfere with transmission by the two
Xiphinema
species. This study provides new insights into GFLV CP determinants of nematode transmission.
Possible effect of environmental exposure to asbestos on geographical variation in mesothelioma rates
Authors: Goldberg Stephen , Rey Grégoire , Luce Danièle , Gilg Soit Ilg Anabelle , Rolland Patrick , Brochard Patrick , Imbernon Ellen , Goldberg Marcel
Published: June 2010

Background

In population-based mesothelioma studies in industrialised countries, the incidence of mesothelioma without any identified asbestos exposure (IAE) is usually higher among women, while male incidence is mainly attributed to IAE. Through a comparison of the spatial distribution of male and female rates, and IAE and no IAE incidence, this study investigated whether mesotheliomas without IAE are in fact induced by non-recognised asbestos exposure, mostly from environmental sources.

Methods

We calculated mesothelioma mortality (SMR) and incidence (SIR) ratios by district in France, pooling 30 and 10 years of data, respectively. Using correlation coefficients, we compared geographical patterns of male and female mesothelioma ratios, and IAE and no IAE mesothelioma ratios.

Results

The raw numbers of male and female mesothelioma cases were equivalent. Mesothelioma SMR (0.76) and SIR (0.80) geographical correlations between men and women were strongly positive. SIR correlation between occupationally IAE and no IAE cases was also positive (0.69). Correlation between occupationally IAE and no IAE cases was positive among women but not among men.

Conclusions

Data analyses of mesothelioma mortality and incidence showed that female cases occur in the same geographical areas as male cases. Female mesotheliomas with no IAE occur in the same geographical areas as exposed cases, suggesting asbestos has a major influence on female mesothelioma, likely through environmental exposure.
Using live tissue laboratories to promote clinical reasoning in doctor of physical therapy students
Authors: Moore W. Allen , Noonan Ann Cassidy
Published: June 2010

Recently, the use of animal laboratories has decreased in medical and basic science programs due to lack of trained faculty members, student concerns about animal welfare, and the increased availability of inexpensive alternatives such as computer simulations and videos. Animal laboratories, however, have several advantages over alternative forms of instruction, including an increased sense of learner responsibility, long-term psychomotor competency, and improved ability to examine functional relationships. While the use of animal laboratories has been studied in medical programs, it has not been examined in physical therapy or other allied health programs. The purposes of this study were to examine the attitudes of student physical therapists toward clinically relevant animal laboratories and to determine if student physical therapists felt that the laboratory modules prepared them for future course work and clinical internships. Written surveys completed by the study participants indicated that most students enjoyed the labs and believed that they helped prepare them for course examinations. Additionally, the majority of students believed that the laboratory modules helped them prepare for future course work and clinical internships.
PTP-PEST controls motility, adherens junction assembly, and Rho GTPase activity in colon cancer cells
Authors: Espejo Rosario , Rengifo-Cam William , Schaller Michael D. , Evers B. Mark , Sastry Sarita K.
Published: June 2010

An important step in carcinoma progression is loss of cell-cell adhesion leading to increased invasion and metastasis. We show here that the protein tyrosine phosphatase, PTP-PEST, is a critical regulator of cell-cell junction integrity and epithelial cell motility. Using colon carcinoma cells, we show that the expression level of PTP-PEST regulates cell motility. Either transient small interfering RNA or stable short hairpin RNA knockdown of PTP-PEST enhances haptotactic and chemotactic migration of KM12C colon carcinoma cells. Furthermore, KM12C cells with stably knocked down PTP-PEST exhibit a mesenchymal-like phenotype with prominent membrane ruffles and lamellae. In contrast, ectopic expression of PTP-PEST in KM20 or DLD-1 cells, which lack detectable endogenous PTP-PEST expression, suppresses haptotactic migration. Importantly, we find that PTP-PEST localizes in adherens junctions. Concomitant with enhanced motility, stable knockdown of PTP-PEST causes a disruption of cell-cell junctions. These effects are due to a defect in junctional assembly and not to a loss of E-cadherin expression. Adherens junction assembly is impaired following calcium switch in KM12C cells with stably knocked down PTP-PEST and is accompanied by an increase in the activity of Rac1 and a suppression of RhoA activity in response to cadherin engagement. Taken together, these results suggest that PTP-PEST functions as a suppressor of epithelial cell motility by controlling Rho GTPase activity and the assembly of adherens junctions.
Stretch magnitude and frequency-dependent actin cytoskeleton remodeling in alveolar epithelia
Authors: DiPaolo Brian C. , Lenormand Guillaume , Fredberg Jeffrey J. , Margulies Susan S.
Published: June 2010

Alveolar epithelial cells (AEC) maintain integrity of the blood-gas barrier with gasket-like intercellular tight junctions (TJ) that are anchored internally to the actin cytoskeleton. We hypothesize that stretch rapidly reorganizes actin (<10 min) into a perijunctional actin ring (PJAR) in a manner that is dependent on magnitude and frequency of the stretch, accompanied by spontaneous movement of actin-anchored receptors at the plasma membrane. Primary AEC monolayers were stretched biaxially to create a change in surface area (ΔSA) of 12%, 25%, or 37% in a cyclic manner at 0.25 Hz for up to 60 min, or held tonic at 25% ΔSA for up to 60 min, or left unstretched. By 10 min of stretch PJARs were evident in 25% and 37% ΔSA at 0.25 Hz, but not for 12% ΔSA at 0.25 Hz, or at tonic 25% ΔSA, or with no stretch. Treatment with 1 μM jasplakinolide abolished stretch-induced PJAR formation, however. As a rough index of remodeling rate, we measured spontaneous motions of 5-μm microbeads bound to actin focal adhesion complexes on the apical membrane surfaces; within 1 min of exposure to ΔSA of 25% and 37%, these motions increased substantially, increased with increasing stretch frequency, and were consistent with our mechanistic hypothesis. With a tonic stretch, however, the spontaneous motion of microbeads attenuated back to unstretched levels, whereas PJAR remained unchanged. Stretch did not increase spontaneous microbead motion in human alveolar epithelial adenocarcinoma A549 monolayers, confirming that this actin remodeling response to stretch was a cell-type specific response. In summary, stretch of primary rat AEC monolayers forms PJARs and rapidly reorganized actin binding sites at the plasma membrane in a manner dependent on stretch magnitude and frequency.