bluuebunny/crossref_metadata_2025 huggingface dataset:
MRI urethrovesical junction mobility is associated with global pelvic floor laxity in female stress incontinence
Authors: ANSQUER YAN , FERNANDEZ PEDRO , AIMOT SABINE , BENNIS MALIKA , SALOMON LAURENCE , CARBONNE BRUNO
Published: September 2007

Objective. To assess in a population of stress incontinent patients without genital prolapse whether urethrovesical junction mobility is associated with global pelvic floor laxity. Methods. Dynamic MRI of 40 patients referred prior to surgery for urinary stress incontinence were reviewed retrospectively. The orientation of the urethrovesical junction was evaluated at rest and at straining in reference to the pubococcygeal line, and defined as the bladder neck pubococcygeal angle. The urethrovesical junction mobility was calculated as the difference between the bladder neck pubococcygeal angles at rest and at straining. Urethrovesical junction mobility and bladder neck pubococcygeal angles at rest and at straining were tested for correlations with the resting and straining position of different pelvic organs, urogenital hiatus size, levator plate angle, and anterior rectal bulging when straining. Results. Urethrovesical junction mobility was correlated with the position of the bladder neck (p<0.0001), bladder base (p<0.0001) and uterine cervix (p<0.0001) at straining, as well as the hiatus length (p = 0.0012) and width (p = 0.0002), and levator plate angle (p<0.0001). The bladder neck pubococcygeal angle at rest was correlated with the resting position of the bladder neck (p<0.0001), bladder base (p<0.0001), uterine cervix (p = 0.02), and the hiatus length (p = 0.0004) and width (p = 0.045) at rest, whereas the bladder neck pubococcygeal angle at straining was correlated with the straining position of the bladder neck (p<0.0001), bladder base (p<0.0001), uterine cervix (p<0.0001), and hiatus length (p = 0.0005) and width (p = 0.0004), and levator plate angle (p<0.0001) at straining. Conclusion. In a population of stress incontinent patients, the urethrovesical junction mobility was correlated with global pelvic floor laxity.
Use of recombinant activated factor VII for massive postpartum hemorrhage
Authors: HOSSAIN NAZLI , SHAMSI TAHIR , HAIDER SAEEDA , SOOMRO NARGIS , KHAN NUSRAT H. , UMER MEMON GHUFFRANA , FARZANA TASNEEM , ANSARI SAQIB , TRICHE ELIZABETH W. , KUCZYNSKI EDWARD , LOCKWOOD CHARLES J. , PAIDAS MICHAEL J.
Published: September 2007

Objective. We hypothesised that patients with massive postpartum hemorrhage (PPH), defined as blood loss >1,500 ml, may benefit from the use of activated recombinant factor VII (rFVIIa). Design. Retrospective cohort study. Setting. Department of Obstetrics & Gynaecology, Dow University of Health Sciences. Population. Thirty‐four women with a diagnosis of massive PPH. Methods. All patients with PPH who were admitted to the Department of Obstetrics & Gynecology and Surgical Intensive Care Unit of Civil Hospital Karachi, Pakistan, were included in the study. From March 2005 to October 2006, 34 patients fulfilled the criteria of massive PPH, of which 18 received rFVIIa to control bleeding, and 16 patients did not. Availability and cost of rFVIIa were the factors in drug allocation. Main outcome measures. Maternal mortality, correction of coagulopathy, the amount of blood products transfused and preservation of fertility. Results. Patients receiving rFVIIa had lower maternal mortality (5/18, 28% versus 8/16, 50%, OR: 0.04 (0.002, 0.83)), and received a lower number of packed red cell transfusions (4.0±4.46 versus 9.61±6.7, p value 0.007), against the comparison group. Patients receiving rFVIIa had lower activated partial thromboplastin (median: 13.0; 25–75th percentile: −25.0, −8.0, signed rank p<0.0001), and lower prothrombin times (median: −8.8; 25–75th percentile: −24.2, −4.8), after administration of drug. There was no significant difference in the rate of hysterectomy between the 2 groups (11/18 (61%) versus 6/16 (38%)). No adverse event attributable to rFVIIa was observed in the study. Conclusion. Activated recombinant factor VII can be a life‐saving drug in patients with massive PPH.
New Wine in Old Bottles? England's Parish and Town Councils and New Labour's Neighbourhood Experiment
Authors: Jones Andrew
Published: September 2007

This paper deals with English parish and town councils in the context of English local government reform. It explores their potential for community and local economic development and in fulfilling a role in neighbourhood government in the light of the current UK Government's declared intention of empowering neighbourhoods. There are some suggestions for the development of realistic ‘grass roots’ local government and examples of involvement in local area regeneration. It ends with some thoughts on legislation currently (June 2007) before Parliament, and in particular on their reception in London. Much of material on which this paper is based was gathered through project work undertaken for Defra, the Countryside Agency (now the Commission for Rural Communities) and the Society for Local Council Clerks, dealing, with variously, the capacity and democratic legitimacy of local councils and their relationship with principal authorities and other community and statutory agencies. Altogether, approximately 130 interviews were conducted with parish and town council councillors and clerks, officers and members of principal authorities, and officers of County Associations of Local Councils. A case study approach was adopted, covering a large number of English counties and the cities of Bradford and Milton Keynes.
CXCL12 and CCL20 Play a Significant Role in Mucosal T‐Lymphocyte Adherence to Intestinal Microvessels in Mice
Authors: OYAMA TOKUSHIGE , MIURA SOICHIRO , WATANABE CHIKAKO , HOKARI RYOTA , FUJIYAMA YOICHI , KOMOTO SHUNSUKE , TSUZUKI YOSHIKAZU , HOSOE NAOKI , NAGATA HIROSHI , HIBI TOSHIFUMI
Published: September 2007

Objective: Although it is known that the chemokines CXCL12 and CCL20 are expressed in the intestine, their contribution to lymphocyte homing has not been investigated in detail. The authors investigated whether the CXCL12‐CXCR4 and CCL20‐CCR6 systems are involved in T lymphocyte–endothelial interaction in microvessels of the small and large intestines.

Methods: Labeled lamina proprial lymphocytes (LPLs) were administered to mice, and their adhesion to microvessels of normal and TNF‐α –induced inflamed intestinal mucosa was observed under an intravital microscope. Antibodies against CXCL12, CCL‐20, or CCL‐25 were administered prior to lymphocyte administration, and in some experiments CXCR4 or CCR6 on LPLs was desensitized with an excess amount of chemokine.

Results: LPLs adhered to microvessels of the ileum and colon, and TNF‐α induced a significant accumulation at both sites. Blocking of the CXCL12‐CXCR4 system significantly inhibited the LPL adhesion in the ileum and colon under both normal and TNF‐α –treated conditions. However, blocking of the CCL20‐CCR6 system significantly attenuated LPL adhesion only under a TNF‐α –treated condition. There was an additive inhibitory effect on LPL adherence by CXCL12 and CCL20 blocking in TNF‐α –induced inflamed intestines. There was also an additive function of the CCL25‐CCR9 system in LPL accumulation in the small intestine.

Conclusion: Several chemokine systems may play significant roles cooperatively in vivo in LPL adherence to microvessels of intestinal mucosa.
Migration and coronary heart disease: A study of Finnish twins living in Sweden and their co-twins residing in Finland
Authors: Hedlund Ebba , Kaprio Jaakko , Lange Anders , Koskenvuo Markku , Jartti Laura , Rönnemaa Tapani , Hammar Niklas
Published: September 2007

Background: Finland and Sweden are neighbouring countries with a substantially higher incidence and mortality from coronary heart disease (CHD) in Finland. Migration from Finland to Sweden has resulted in a population of about 187,000 Finnish immigrants, with a higher risk of CHD than Swedes. The aim of the present study was to analyse the prevalence of CHD in migrants to Sweden compared with co-twins remaining in Finland. Methods: The study population consisted of twin pairs of the Finnish Twin Cohort Study where at least one twin had lived one year or more in Sweden, including 1,534 subjects and 251 complete twin pairs discordant regarding residency in Sweden. Emigrant twins were compared with non-migrant co-twins regarding prevalence of CHD in 1998. CHD prevalence was assessed by self-reported questionnaires validated using information from a clinical examination. Results: Self-reported CHD showed a good correspondence with clinical diagnosis. Differences in social and behavioural risk factors for CHD among men were small but emigrants were more physically active than non-migrants. Female emigrants had less overweight and better education, but were more often working class than non-migrants. Intra-pair comparisons restricted to migration discordant pairs showed a tendency towards a reduced prevalence of CHD in the migrant co-twins (0.6; 0.3—1.4). In analyses of all subjects disregarding pair status, emigrants showed a reduced prevalence of CHD compared with subjects always living in Finland (0.6; 0.4—0.9). Conclusion: Emigration from Finland to Sweden may be associated with a reduced prevalence of CHD. The causes are most likely multifactorial and may involve changes in dietary habits, physical activity, psychosocial factors, and inflammation.
Konrad Hochedlinger: The new kid of nuclear reprogramming
Authors: Williams Ruth
Published: September 2007

Hochedlinger pushes cells' developmental rewind buttons to examine their epigenetic history.
Inhibition of nonsense-mediated mRNA decay (NMD) by a new chemical molecule reveals the dynamic of NMD factors in P-bodies
Authors: Durand Sébastien , Cougot Nicolas , Mahuteau-Betzer Florence , Nguyen Chi-Hung , Grierson David S. , Bertrand Edouard , Tazi Jamal , Lejeune Fabrice
Published: September 2007

In mammals, nonsense-mediated mRNA decay (NMD) is a quality-control mechanism that degrades mRNA harboring a premature termination codon to prevent the synthesis of truncated proteins. To gain insight into the NMD mechanism, we identified NMD inhibitor 1 (NMDI 1) as a small molecule inhibitor of the NMD pathway. We characterized the mode of action of this compound and demonstrated that it acts upstream of hUPF1. NMDI 1 induced the loss of interactions between hSMG5 and hUPF1 and the stabilization of hyperphosphorylated isoforms of hUPF1. Incubation of cells with NMDI 1 allowed us to demonstrate that NMD factors and mRNAs subject to NMD transit through processing bodies (P-bodies), as is the case in yeast. The results suggest a model in which mRNA and NMD factors are sequentially recruited to P-bodies.
APC mutations lead to cytokinetic failures in vitro and tetraploid genotypes in Min mice
Authors: Caldwell Christine M. , Green Rebecca A. , Kaplan Kenneth B.
Published: September 2007

Previous research has proposed that genomic instability contributes to cancer progression, with its initiation linked to tetraploid cell formation (Duesberg, P., and R. Li. 2003. Cell Cycle. 2:202–210; Ganem, N.J., Z. Storchova, and D. Pellman. 2007. Curr. Opin. Genet. Dev. 17:157–162). However, there is little direct evidence linking cancer-causing mutations with such events, and it remains controversial whether genomic instability is a cause or an effect of cancer. In this study, we show that adenomatous polyposis coli (APC) mutations found in human colorectal cancers dominantly inhibit cytokinesis by preventing mitotic spindle anchoring at the anaphase cortex and, thus, blocking initiation of the cytokinetic furrow. We find that dividing crypt cells in the small intestines of APCMin/+ mice exhibit similar mitotic defects, including misoriented spindles and misaligned chromosomes. These defects are observed in normal crypt cells with wild-type levels of β-catenin and, importantly, are associated with tetraploid genotypes. We provide direct evidence that the dominant activity of APC mutants induces aneuploidy in vivo. Our data support a model whereby tetraploid cells represent a first step in the onset of genomic instability and colorectal cancer.
The type III effector EspF coordinates membrane trafficking by the spatiotemporal activation of two eukaryotic signaling pathways
Authors: Alto Neal M. , Weflen Andrew W. , Rardin Matthew J. , Yarar Defne , Lazar Cheri S. , Tonikian Raffi , Koller Antonius , Taylor Susan S. , Boone Charles , Sidhu Sachdev S. , Schmid Sandra L. , Hecht Gail A. , Dixon Jack E.
Published: September 2007

Bacterial toxins and effector proteins hijack eukaryotic enzymes that are spatially localized and display rapid signaling kinetics. However, the molecular mechanisms by which virulence factors engage highly dynamic substrates in the host cell environment are poorly understood. Here, we demonstrate that the enteropathogenic Escherichia coli (EPEC) type III effector protein EspF nucleates a multiprotein signaling complex composed of eukaryotic sorting nexin 9 (SNX9) and neuronal Wiskott-Aldrich syndrome protein (N-WASP). We demonstrate that a specific and high affinity association between EspF and SNX9 induces membrane remodeling in host cells. These membrane-remodeling events are directly coupled to N-WASP/Arp2/3–mediated actin nucleation. In addition to providing a biochemical mechanism of EspF function, we find that EspF dynamically localizes to membrane-trafficking organelles in a spatiotemporal pattern that correlates with SNX9 and N-WASP activity in living cells. Thus, our findings suggest that the EspF-dependent assembly of SNX9 and N-WASP represents a novel form of signaling mimicry used to promote EPEC pathogenesis and gastrointestinal disease.
Accessibility of Targeted DHPR Sites to Streptavidin and Functional Effects of Binding on EC Coupling
Authors: Lorenzon Nancy M. , Beam Kurt G.
Published: September 2007

In skeletal muscle, the dihydropyridine receptor (DHPR) in the plasma membrane (PM) serves as a Ca2+ channel and as the voltage sensor for excitation–contraction (EC coupling), triggering Ca2+ release via the type 1 ryanodine receptor (RyR1) in the sarcoplasmic reticulum (SR) membrane. In addition to being functionally linked, these two proteins are also structurally linked to one another, but the identity of these links remains unknown. As an approach to address this issue, we have expressed DHPR α1S or β1a subunits, with a biotin acceptor domain fused to targeted sites, in myotubes null for the corresponding, endogenous DHPR subunit. After saponin permeabilization, the ∼60-kD streptavidin molecule had access to the β1a N and C termini and to the α1S N terminus and proximal II–III loop (residues 671–686). Steptavidin also had access to these sites after injection into living myotubes. However, sites of the α1S C terminus were either inaccessible or conditionally accessible in saponin- permeabilized myotubes, suggesting that these C-terminal regions may exist in conformations that are occluded by other proteins in PM/SR junction (e.g., RyR1). The binding of injected streptavidin to the β1a N or C terminus, or to the α1S N terminus, had no effect on electrically evoked contractions. By contrast, binding of streptavidin to the proximal α1S II–III loop abolished such contractions, without affecting agonist-induced Ca2+ release via RyR1. Moreover, the block of EC coupling did not appear to result from global distortion of the DHPR and supports the hypothesis that conformational changes of the α1S II–III loop are necessary for EC coupling in skeletal muscle.
Regulation of TRPM2 by Extra- and Intracellular Calcium
Authors: Starkus John , Beck Andreas , Fleig Andrea , Penner Reinhold
Published: September 2007

TRPM2 is a calcium-permeable nonselective cation channel that is opened by the binding of ADP-ribose (ADPR) to a C-terminal nudix domain. Channel activity is further regulated by several cytosolic factors, including cyclic ADPR (cADPR), nicotinamide adenine dinucleotide phosphate (NAADP), Ca2+ and calmodulin (CaM), and adenosine monophosphate (AMP). In addition, intracellular ions typically used in patch-clamp experiments such as Cs+ or Na+ can alter ADPR sensitivity and voltage dependence, complicating the evaluation of the roles of the various modulators in a physiological context. We investigated the roles of extra- and intracellular Ca2+ as well as CaM as modulators of ADPR-induced TRPM2 currents under more physiological conditions, using K+-based internal saline in patch-clamp experiments performed on human TRPM2 expressed in HEK293 cells. Our results show that in the absence of Ca2+, both internally and externally, ADPR alone cannot induce cation currents. In the absence of extracellular Ca2+, a minimum of 30 nM internal Ca2+ is required to cause partial TRPM2 activation with ADPR. However, 200 μM external Ca2+ is as efficient as 1 mM Ca2+ in TRPM2 activation, indicating an external Ca2+ binding site important for proper channel function. Ca2+ facilitates ADPR gating with a half-maximal effective concentration of 50 nM and this is independent of extracellular Ca2+. Furthermore, TRPM2 currents inactivate if intracellular Ca2+ levels fall below 100 nM irrespective of extracellular Ca2+. The facilitatory effect of intracellular Ca2+ is not mimicked by Mg2+, Ba2+, or Zn2+. Only Sr2+ facilitates TRPM2 as effectively as Ca2+, but this is due to Sr2+-induced Ca2+ release from internal stores rather than a direct effect of Sr2+ itself. Together, these data demonstrate that cytosolic Ca2+ regulates TRPM2 channel activation. Its facilitatory action likely occurs via CaM, since the addition of 100 μM CaM to the patch pipette significantly enhances ADPR-induced TRPM2 currents at fixed [Ca2+]i and this can be counteracted by calmidazolium. We conclude that ADPR is responsible for TRPM2 gating and Ca2+ facilitates activation via calmodulin.
Understanding researchers: findings from a focus group
Authors: PITTS SiNae , STANLEY Adrian
Published: September 2007

In November 2006, a focus group of researchers was assembled at a conference hosted by the Society of Scholarly Publishers in Washington, DC. The panel addressed a series of prepared questions regarding current research practices, perceptions, and preferences, particularly as they applied to scientific and medical publications, in corporate, academic, and medical environments. This article highlights issues and views the panel discussed, with the aim of narrowing the gap of understanding between researchers and publishers. This information will help publishers to develop content and delivery methods that are closely aligned with the needs of researchers.
Nicholas Harold Lloyd Ridley. 10 July 1906 — 25 May 2001
Authors: Apple David J.
Published: September 2007

Sir Harold Ridley invented and refined the modern miracle of replacing lenses obscured by cataracts with plastic optical lenses, thus rendering a complete cataract cure. This operation, broadly termed the cataract–intraocular lens (IOL) operation, has since brought sight to many millions of people throughout the world, and continues to improve the quality of life of more than 10 million patients worldwide each year.
Modelling transdermal delivery of high molecular weight drugs from microneedle systems
Authors: Al-Qallaf Barrak , Das Diganta Bhusan , Mori Daisuke , Cui Zhanfeng
Published: September 2007

In the past few years, a number of microneedle designs have been proposed for transdermal drug delivery of high molecular weight drugs. However, most of them do not increase the drug permeability in skin significantly. In other cases, designs developed based on certain criteria (e.g. strength of the microneedles) have failed to meet other criteria (e.g. drug permeability in skin, throughputs of the drugs, etc.). It is obvious therefore that in order to determine the ‘optimum’ design of these microneedles, the effect of different factors (e.g. length of the microneedle, surface area of the patch, etc.) along with various transport properties of drug transport behaviour using microneedles should be determined accurately. Appropriate mathematical models for drug transport from these systems into skin have the potential to resolve some of these issues. To address this, a parametric analysis for transdermal delivery of a high molecular weight drug from a microneedle is presented in this paper. The simulations have allowed us to identify the significance of various factors that influence the drug delivery while designing microneedle arrays. A scaling analysis is also done which shows the functional dependence of drug concentration on other variables of skin and microneedle arrays.
The fractionation of spoken language understanding by measuring electrical and magnetic brain signals
Authors: Hagoort Peter
Published: September 2007

This paper focuses on what electrical and magnetic recordings of human brain activity reveal about spoken language understanding. Based on the high temporal resolution of these recordings, a fine-grained temporal profile of different aspects of spoken language comprehension can be obtained. Crucial aspects of speech comprehension are lexical access, selection and semantic integration. Results show that for words spoken in context, there is no ‘magic moment’ when lexical selection ends and semantic integration begins. Irrespective of whether words have early or late recognition points, semantic integration processing is initiated before words can be identified on the basis of the acoustic information alone. Moreover, for one particular event-related brain potential (ERP) component (the N400), equivalent impact of sentence- and discourse-semantic contexts is observed. This indicates that in comprehension, a spoken word is immediately evaluated relative to the widest interpretive domain available. In addition, this happens very quickly. Findings are discussed that show that often an unfolding word can be mapped onto discourse-level representations well before the end of the word. Overall, the time course of the ERP effects is compatible with the view that the different information types (lexical, syntactic, phonological, pragmatic) are processed in parallel and influence the interpretation process incrementally, that is as soon as the relevant pieces of information are available. This is referred to as the immediacy principle.
Speech perception at the interface of neurobiology and linguistics
Authors: Poeppel David , Idsardi William J , van Wassenhove Virginie
Published: September 2007

Speech perception consists of a set of computations that take continuously varying acoustic waveforms as input and generate discrete representations that make contact with the lexical representations stored in long-term memory as output. Because the perceptual objects that are recognized by the speech perception enter into subsequent linguistic computation, the format that is used for lexical representation and processing fundamentally constrains the speech perceptual processes. Consequently, theories of speech perception must, at some level, be tightly linked to theories of lexical representation. Minimally, speech perception must yield representations that smoothly and rapidly interface with stored lexical items. Adopting the perspective of Marr, we argue and provide neurobiological and psychophysical evidence for the following research programme. First, at the implementational level, speech perception is a multi-time resolution process, with perceptual analyses occurring concurrently on at least two time scales (approx. 20–80 ms, approx. 150–300 ms), commensurate with (sub)segmental and syllabic analyses, respectively. Second, at the algorithmic level, we suggest that perception proceeds on the basis of internal forward models, or uses an ‘analysis-by-synthesis’ approach. Third, at the computational level (in the sense of Marr), the theory of lexical representation that we adopt is principally informed by phonological research and assumes that words are represented in the mental lexicon in terms of sequences of discrete segments composed of distinctive features. One important goal of the research programme is to develop linking hypotheses between putative neurobiological primitives (e.g. temporal primitives) and those primitives derived from linguistic inquiry, to arrive ultimately at a biologically sensible and theoretically satisfying model of representation and computation in speech.
Neural specializations for speech and pitch: moving beyond the dichotomies
Authors: Zatorre Robert J , Gandour Jackson T
Published: September 2007

The idea that speech processing relies on unique, encapsulated, domain-specific mechanisms has been around for some time. Another well-known idea, often espoused as being in opposition to the first proposal, is that processing of speech sounds entails general-purpose neural mechanisms sensitive to the acoustic features that are present in speech. Here, we suggest that these dichotomous views need not be mutually exclusive. Specifically, there is now extensive evidence that spectral and temporal acoustical properties predict the relative specialization of right and left auditory cortices, and that this is a parsimonious way to account not only for the processing of speech sounds, but also for non-speech sounds such as musical tones. We also point out that there is equally compelling evidence that neural responses elicited by speech sounds can differ depending on more abstract, linguistically relevant properties of a stimulus (such as whether it forms part of one's language or not). Tonal languages provide a particularly valuable window to understand the interplay between these processes. The key to reconciling these phenomena probably lies in understanding the interactions between afferent pathways that carry stimulus information, with top-down processing mechanisms that modulate these processes. Although we are still far from the point of having a complete picture, we argue that moving forward will require us to abandon the dichotomy argument in favour of a more integrated approach.
Improved contamination control for a rapid phage-based rifampicin resistance test for Mycobacterium tuberculosis
Authors: Mole Richard , Trollip Andre , Abrahams Celeste , Bosman Marlein , Albert Heidi
Published: September 2007

A prospective study was conducted of the rapid FASTPlaque-Response test for determination of rifampicin resistance in Mycobacterium tuberculosis with and without the addition of an antimicrobial supplement containing nystatin, oxacillin and aztreonam (NOA) to control specimen-related contamination. A total of 631 smear-positive sputum specimens was tested. The age of specimens ranged from 0 to 21 days. The NOA antimicrobial was effective at controlling contamination, with 4.1 % of specimens contaminated when the NOA antimicrobial supplement was used compared with 13.9 % contamination without NOA. Overall levels of interpretability of the test with NOA were 87.8 % with specimens of ≤3 days and 79.0 % for all specimens. This compared with 70.1 and 73.8 % readable results, respectively, from conventional culture-based drug susceptibility testing (DST). Sensitivity, specificity and overall accuracy of the FASTPlaque-Response test for rifampicin resistance were 98.1, 96.3 and 96.6 %, respectively, for all specimens with NOA, and 93.2, 96.3 and 95.9 % without NOA, when compared with resolved conventional DST results. Inclusion of the NOA supplement reduced contamination, increased the number of interpretable results and did not adversely affect the performance of the FASTPlaque-Response test. Thus, the use of NOA improves the robustness of the test, facilitating its wider implementation.
Assessment of four serological techniques in the immunological diagnosis of farmers' lung disease
Authors: Reboux Gabriel , Piarroux Renaud , Roussel Sandrine , Millon Laurence , Bardonnet Karine , Dalphin Jean-Charles
Published: September 2007

Farmers' lung disease (FLD) is a pulmonary disease that results from repeated inhalation of antigens from mouldy hay or straw. The objective of this prospective study was to assess the reliability of four serological techniques in FLD diagnosis. Sera from 15 consecutive patients with FLD, 15 healthy control farmers and 30 urban controls were analysed using four serological techniques [electrosyneresis (ES), Ouchterlony double diffusion (DD), ELISA and Western blot (WB)] with four antigens (Absidia corymbifera, Eurotium amstelodami, Wallemia sebi and Saccharopolyspora rectivirgula). In the authors' region, ES on cellulose acetate with A. corymbifera antigen was the most relevant diagnostic tool for discriminating FLD patients from healthy exposed farmers (sensitivity 87 %, specificity 100 %). DD tests were in accordance with ES, but their discriminatory power was lower. No threshold indicating both good sensitivity and specificity could be established with ELISA. WB analysis failed to identify specific bands for FLD. This study demonstrates the efficacy of determining precipitin levels with an appropriate technique, using a panel of antigens consistent with the specific exposure of a given area.
Evolution of slime production by coagulase-negative staphylococci and enterotoxigenic characteristics of Staphylococcus aureus strains isolated from various human clinical specimens
Authors: Boynukara Banur , Gulhan Timur , Gurturk Kemal , Alisarli Mustafa , Ogun Erdal
Published: September 2007

The present study was designed to determine the slime production of coagulase-negative staphylococci (CoNS) and the enterotoxigenic properties of Staphylococcus aureus strains, and to evaluate the clinical importance of slime-producing CoNS and enterotoxigenic S. aureus strains isolated from various human clinical specimens. For this purpose, a total of 120 Staphylococcus strains were isolated and identified, and further characterized for their slime production and enterotoxigenicity. Of the clinical isolates, 55 (45.8 %) were found to be S. aureus, and the others (54.2 %) were identified as CoNS. Of the CoNS, 20 (16.7 %) were further identified as Staphylococcus hominis, 18 (15 %) as Staphylococcus epidermidis, six (5 %) as Staphylococcus xylosus, six (5 %) as Staphylococcus warneri, five (4.2 %) as Staphylococcus sciuri, four (3.3 %) as Staphylococcus haemolyticus, and two each (1.7 %) as Staphylococcus simulans, Staphylococcus capitis and Staphylococcus saprophyticus, respectively. Thirty-nine (60 %) of 65 CoNS were found to be slime producers. Slime production was observed in all CoNS, except S. capitis, mostly from blood (38.5 %), tracheal aspiration (20.5 %) and urine (12.8 %) specimens. In addition, of the 55 S. aureus isolates, 46 (83.6 %) were found to be enterotoxigenic, and of these S. aureus strains, 39 (84.7 %) were positive for staphylococcal enterotoxin (SE)A. The results of this study showed that the slime-producing CoNS were mostly found in clinical specimens of blood, tracheal aspirate and urine. SEA was the predominant enterotoxin type detected in S. aureus strains from human clinical specimens.
Multiple genus-specific markers in PCR assays improve the specificity and sensitivity of diagnosis of brucellosis in field animals
Authors: Mukherjee Falguni , Jain Jainendra , Patel Vipul , Nair Mrinalini
Published: September 2007

Brucella-specific nucleotide sequences encoding the BCSP 31 kDa protein, Omp2 and the 16S rRNA were employed in three independent diagnostic PCR assays. Results of the three PCR assays on six reference strains of Brucella were in complete agreement. The results of PCR assays based on bcsp and omp2 on 19 Indian field isolates (human, bovine and murine tissues) also agreed completely. However, when the 16S rRNA gene was employed as the diagnostic target in the PCR, only 14 out of these 19 isolates and 2 out of 7 bovine milk isolates were identified as the genus Brucella. The bovine blood samples were insensitive to 16S rRNA PCR. The antibody-detecting ELISA results of field samples (n=87) from a serologically positive herd in India were compared separately with omp2 and bcsp PCRs of blood (n=62). While the bcsp PCR was the most sensitive, the degree of association of ELISA with omp2 blood PCR (κ=0.37 at P <0.05) was similar to that with the bcsp blood PCR (κ =0.34 at P <0.05). An improvement in the correlation between ELISA and blood PCR was noticed (κ =0.5 at P <0.05) when a consensus result of omp2 and bcsp blood PCR was considered for comparison with ELISA. The use of more than one marker-based PCR gave increased sensitivity and higher specificity and appears to be a more reliable molecular diagnostic approach for screening of field animals.
Chronic meningitis caused by Erysipelothrix rhusiopathiae
Authors: Kim Suk Ran , Kwon Min Jung , Lee Jang Ho , Lee Nam Yong
Published: September 2007

A 47-year-old man presented with headache, nausea, vomiting and fever. Laboratory findings including analysis of cerebrospinal fluid suggested bacterial meningitis. Erysipelothrix rhusiopathiae was identified in cultures of cerebrospinal fluid. The patient recovered without any neurological sequelae after antimicrobial treatment. It is interesting that intracranial infection by E. rhusiopathiae reappeared after scores of years and that it presented with absence of an underlying cause or bacteraemia.
Genotypic analyses of uropathogenic Escherichia coli based on fimH single nucleotide polymorphisms (SNPs)
Authors: Tartof Sara Y. , Solberg Owen D. , Riley Lee W.
Published: September 2007

The application of genotyping techniques for subtyping uropathogenic Escherichia coli has contributed to better understanding of the epidemiology of community-acquired urinary tract infection (UTI). However, the current techniques are hampered by limited reproducibility, poor discriminatory power, labour-intensive performance or high cost. A screening test that is sequence-based would provide an inexpensive, reproducible way to subtype E. coli isolates. Such a test, if also discriminatory, would be highly useful for epidemiological studies. The discriminatory ability of 12 putative virulence genes (fimH, fliD, fliM, iha, motA, papA/H, kpsMTII, fepE, fimA, flgA, malG, purD) was evaluated based on single nucleotide polymorphisms (SNPs) in nine uropathogenic E. coli isolates, all previously found to belong to a single multilocus sequence type (MLST) complex (ST69). An additional 25 epidemiologically well-characterized E. coli isolates belonging to 12 distinct MLST clonal complexes were analysed for fimH SNP. None of the 12 genes except fimH were able to further discriminate the nine ST69-complex strains. Isolates belonging to the 12 non-ST69 MLST groups were separated into 10 fimH SNP subgroups. While fimH SNP analysis may not be an appropriate phylogenetic method, it offers discriminatory power similar to that of MLST and could be used as a simple, inexpensive screening test for epidemiological studies of uropathogenic E. coli.
Simplified detection of Mycobacterium tuberculosis in sputum using smear microscopy and PCR with molecular beacons
Authors: Haldar Sagarika , Chakravorty Soumitesh , Bhalla Manpreet , De Majumdar Shyamasree , Tyagi Jaya Sivaswami
Published: September 2007

The prompt diagnosis of smear-negative cases is a prerequisite to controlling tuberculosis (TB). Several new laboratory approaches, including nucleic acid amplification (NAA), are being evaluated in various disease settings to meet this challenge. However, NAA needs simplification before it is widely accepted. Furthermore, a supporting smear result improves confidence in and reliability of PCR. In this context, an asymmetric devR PCR assay using two molecular beacon probes for visual or fluorimetric end-point detection of Mycobacterium tuberculosis was developed. The assays reproducibly detected 25 fg M. tuberculosis DNA versus 100 fg by conventional gel electrophoresis (henceforth referred to as gel assay). The devR and IS6110 PCR assays were blindly evaluated on sputum specimens obtained from a directly observed-treatment short-course centre. Universal sample processing (USP) smear microscopy and culture were used as a supportive test and the ‘gold’ standard, respectively. Among the 148 specimens analysed, 120 were M. tuberculosis culture-positive. Amongst the 122 direct smear-negative samples, 96 were culture-positive, of which 61 were detected by USP smear microscopy. All 35 USP smear-negative samples were positive by three or more PCR methods. devR PCR had a sensitivity of 92.5 % in the fluorimetric assay versus 86.7 % by visual inspection and 90.8 % by the gel method. IS6110 PCR performed at almost equivalent levels. devR visual and fluorimetric assays considered together yielded an increased sensitivity of 95 % without compromising on a specificity of 92.9 %. The results suggest that the USP smear test is useful for diagnosing direct smear-negative TB and judiciously restricting PCR testing to only smear-negative samples. When used together, these tests can provide rapid diagnosis of smear-negative TB in a cost-effective manner.
Influence of Bifidobacterium longum BB536 intake on faecal microbiota in individuals with Japanese cedar pollinosis during the pollen season
Authors: Odamaki Toshitaka , Xiao Jin-Zhong , Iwabuchi Noriyuki , Sakamoto Mitsuo , Takahashi Noritoshi , Kondo Shizuki , Miyaji Kazuhiro , Iwatsuki Keiji , Togashi Hideo , Enomoto Tadao , Benno Yoshimi
Published: September 2007

It has been reported that intake of yogurt or powder supplemented with theBifidobacterium longumBB536 probiotic strain alleviated subjective symptoms and affected blood markers of allergy in individuals with Japanese cedar pollinosis (JCPsis) during the pollen seasons of 2004 and 2005, based on randomized, double-blind, placebo-controlled trials. Furthermore, the 2004 study found that intestinal bacteria such as theBacteroides fragilisgroup significantly fluctuated during the pollen season in JCPsis individuals and intake of BB536 yogurt tended to suppress these fluctuations. The present study investigated faecal microbiota to examine whether any changes occurred during the pollen season and whether any influence was exerted by intake of BB536 powder in the 2005 pollen season, which happened to be a heavy season, to confirm the 2004 findings and to evaluate the relationship of microbiota with symptom development. In a randomized, double-blind, placebo-controlled trial, 44 JCPsis subjects received BB536 or a placebo for 13 weeks during the pollen season. Another 14 Japanese cedar pollen (JCP)-specific IgE negative healthy subjects received placebo for the same period. Faecal samples were collected before (week 0), during (weeks 4, 8 and 13) and after (week 17) intervention, and out of JCP season (week 28). Faecal microbiota were analysed using terminal-RFLP (T-RFLP) and real-time PCR methods. Principal component analysis based on T-RFLP indicated distinct patterns of microbiota between healthy subjects and JCPsis subjects in the placebo group, but an intermediate pattern in the BB536 group at week 13, the last stage of the pollen season. The coordinate of principal component 1 at week 13 correlated with composite scores of JCPsis symptoms recorded during the pollen season.Faecalibacterium prausnitziiand theBacteroides fragilisgroup were identified as the main contributors to microbiotal fluctuations. Real-time PCR indicated that BB536 intake suppressed increases in theBacteroides fragilisgroup compared with the placebo group (P<0.05). These results suggest that faecal microbiota in JCPsis subjects, but not healthy subjects, fluctuate at the end of the pollen season and that BB536 intake plays a role in maintaining normal microbiota.
Establishment of a UK National Influenza H5 Laboratory Network
Authors: Curran Martin D. , Ellis Joanna S. , Wreghitt Tim G. , Zambon Maria C.
Published: September 2007

Avian (H5N1) influenza continues to pose a significant threat to human health, although it remains a zoonotic infection. Sensitive and robust surveillance measures are required to detect any evidence that the virus has acquired the ability to transmit between humans and emerge as the next pandemic strain. An integral part of the pandemic planning response in the UK was the creation in 2005 of the UK National H5 Laboratory Network, capable of rapidly and accurately identifying potential human H5N1 infections in all regions of the UK, and the Republic of Ireland. This review details the challenges that designing molecular detection methods for a rapidly evolving virus present, and the strategic decisions and choices required to ensure successful establishment of a functional national laboratory network, providing round the clock testing for H5N1. Laboratory partnerships have delivered improved real-time one-step multiplex PCR methodologies to ensure streamlined testing capable of not only detecting H5 but also a differential diagnosis of seasonal influenza A/B. A range of fully validated real-time PCR H5 confirmatory assays have been developed to run in parallel with a universal first-screening assay. Regular proficiency panels together with weekly surveillance runs, intermittent on-call testing for suspect cases of avian flu in returning travellers, and several outbreaks of avian influenza outbreaks in poultry that have occurred since 2005 in the UK have fully tested the network and the current diagnostic strategies for avian influenza. The network has clearly demonstrated its capability of delivering a confirmed H5N1 diagnosis within 3–4 h of receipt of a sample, an essential prerequisite for administration of the appropriate antiviral therapy, effective clinical management, disease containment and implementation of infection control measures. A functional network is an important means of enhancing laboratory capability and building diagnostic capacity for a newly emerging pandemic of influenza, and is an essential part of pandemic preparedness.
Structure of the Murray Valley encephalitis virus RNA helicase at 1.9 Å resolution
Authors: Mancini Erika J. , Assenberg Rene , Verma Anil , Walter Thomas S. , Tuma Roman , Grimes Jonathan M. , Owens Raymond J. , Stuart David I.
Published: September 2007

Murray Valley encephalitis virus (MVEV), a mosquito‐borne flavivirus endemic to Australia, is closely related to Japanese encephalitis virus and West Nile virus. Nonstructural protein 3 (NS3) is a multifunctional enzyme with serine protease and DEXH/D‐box helicase domains, whose activity is central to flavivirus replication and is therefore a possible target for anti‐flaviviral compounds. Cloning, purification, and crystal structure determination to 1.9 Å resolution of the NS3 helicase of MVEV and characterization of its enzymatic activity is reported. Comparison with the structures of helicases from related viruses supports a possible mechanism of ATP hydrolysis‐driven strand separation.
The Bacillus licheniformis BlaP β‐lactamase as a model protein scaffold to study the insertion of protein fragments
Authors: Vandevenne Marylène , Filee Patrice , Scarafone Natacha , Cloes Benoît , Gaspard Gilles , Yilmaz Nursel , Dumoulin Mireille , François Jean‐Marie , Frère Jean‐Marie , Galleni Moreno
Published: September 2007

Using genetic engineering technologies, the chitin‐binding domain (ChBD) of the human macrophage chitotriosidase has been inserted into the host protein BlaP, a class A β‐lactamase produced by Bacillus licheniformis. The product of this construction behaved as a soluble chimeric protein that conserves both the capacity to bind chitin and to hydrolyze β‐lactam moiety. Here we describe the biochemical and biophysical properties of this protein (BlaPChBD). This work contributes to a better understanding of the reciprocal structural and functional effects of the insertion on the host protein scaffold and the heterologous structured protein fragments. The use of BlaP as a protein carrier represents an efficient approach to the functional study of heterologous protein fragments.
Reduced Cβ statistical potentials can outperform all‐atom potentials in decoy identification
Authors: Fitzgerald James E. , Jha Abhishek K. , Colubri Andres , Sosnick Tobin R. , Freed Karl F.
Published: September 2007

We developed a series of statistical potentials to recognize the native protein from decoys, particularly when using only a reduced representation in which each side chain is treated as a single Cβ atom. Beginning with a highly successful all‐atom statistical potential, the Discrete Optimized Protein Energy function (DOPE), we considered the implications of including additional information in the all‐atom statistical potential and subsequently reducing to the Cβ representation. One of the potentials includes interaction energies conditional on backbone geometries. A second potential separates sequence local from sequence nonlocal interactions and introduces a novel reference state for the sequence local interactions. The resultant potentials perform better than the original DOPE statistical potential in decoy identification. Moreover, even upon passing to a reduced Cβ representation, these statistical potentials outscore the original (all‐atom) DOPE potential in identifying native states for sets of decoys. Interestingly, the backbone‐dependent statistical potential is shown to retain nearly all of the information content of the all‐atom representation in the Cβ representation. In addition, these new statistical potentials are combined with existing potentials to model hydrogen bonding, torsion energies, and solvation energies to produce even better performing potentials. The ability of the Cβ statistical potentials to accurately represent protein interactions bodes well for computational efficiency in protein folding calculations using reduced backbone representations, while the extensions to DOPE illustrate general principles for improving knowledge‐based potentials.
Solution structure and proposed domain–domain recognition interface of an acyl carrier protein domain from a modular polyketide synthase
Authors: Alekseyev Viktor Y. , Liu Corey W. , Cane David E. , Puglisi Joseph D. , Khosla Chaitan
Published: September 2007

Polyketides are a medicinally important class of natural products. The architecture of modular polyketide synthases (PKSs), composed of multiple covalently linked domains grouped into modules, provides an attractive framework for engineering novel polyketide‐producing assemblies. However, impaired domain–domain interactions can compromise the efficiency of engineered polyketide biosynthesis. To facilitate the study of these domain–domain interactions, we have used nuclear magnetic resonance (NMR) spectroscopy to determine the first solution structure of an acyl carrier protein (ACP) domain from a modular PKS, 6‐deoxyerythronolide B synthase (DEBS). The tertiary fold of this 10‐kD domain is a three‐helical bundle; an additional short helix in the second loop also contributes to the core helical packing. Superposition of residues 14–94 of the ensemble on the mean structure yields an average atomic RMSD of 0.64 ± 0.09 Å for the backbone atoms (1.21 ± 0.13 Å for all non‐hydrogen atoms). The three major helices superimpose with a backbone RMSD of 0.48 ± 0.10 Å (0.99 ± 0.11 Å for non‐hydrogen atoms). Based on this solution structure, homology models were constructed for five other DEBS ACP domains. Comparison of their steric and electrostatic surfaces at the putative interaction interface (centered on helix II) suggests a model for protein–protein recognition of ACP domains, consistent with the previously observed specificity. Site‐directed mutagenesis experiments indicate that two of the identified residues influence the specificity of ACP recognition.