bluuebunny/crossref_metadata_2025 huggingface dataset:
Application of a digital technique in evaluating the reliability of shade guides
Authors: Cal E. , Sonugelen M. , Guneri P. , Kesercioglu A. , Kose T.
Published: May 2004

summary  There appears to be a need for a reliable method for quantification of tooth colour and analysis of shade. Therefore, the primary objective of this study was to show the applicability of graphic software in colour analysis and secondly to investigate the reliability of commercial shade guides produced by the same manufacturer, using this digital technique. After confirming the reliability and reproducibility of the digital method by using self‐assessed coloured images, three shade guides of the same manufacturer were photographed in daylight and in studio environments with a digital camera and saved in tagged image file format (TIFF) format. Colour analysis of each photograph was performed using the Adobe Photoshop 4·0 graphic program. Luminosity, and red, green, blue (L and RGB) values of each shade tab of each shade guide were measured and the data were subjected to statistical analysis using the repeated measure Anova test. The L and RGB values of the images taken in daylight differed significantly from those of the images taken in studio environment (P < 0·05). In both environments, the luminosity and red values of the shade tabs were significantly different from each other (P < 0·05). It was concluded that, when the environmental conditions were kept constant, the Adobe Photoshop 4·0 colour analysis program could be used to analyse the colour of images. On the other hand, the results revealed that the accuracy of shade tabs widely being used in colour matching should be readdressed.
Bonding to and protecting worn palatal surfaces of teeth with dentine bonding agents
Authors: Sundaram G. , Bartlett D. , Watson T.
Published: May 2004

summary  Dentine bonding resins may play a role in protecting dentine against acids for patients with erosion. Therefore, it is important to try and understand what potential difficulties may ensue when bonding to worn dentine. This paper reviews the literature on the changes between normal and worn dentine and the effect of this on the bonding to composite resins. A medline search was undertaken using the terms erosion, tooth wear and dentine bonding up until the spring of 2003. Although there are changes on the surface of worn dentine, there is little evidence that this may affect the bond.
An in vitro study into the effect of a limited range of denture cleaners on surface roughness and removal of Candida albicans from conventional heat‐cured acrylic resin denture base material
Authors: Harrison Z. , Johnson A. , Douglas C. W. I.
Published: May 2004

summary  This study evaluated the abrasiveness of four denture cleaners on the surface of denture base material and assessed their ability to remove Candida albicans. Acrylic resin discs 20 mm diameter and 2 mm thick were identically produced and polished. Four cleaners were evaluated: conventional toothpaste; toothpaste with stain remover; denture cleaning paste and an immersion type cleaner, and water were used as control. These were used at dilutions of 1:1, 1:2 and 1:3 with water. An electric toothbrush was used, and the discs cleaned to simulate 1 years’ cleaning. The surface roughness of the discs were then measured, before and after cleaning, using a stylus profilometer, then inoculated with 1·2 × 106C. albicans cells. The effectiveness of the denture cleaners to remove C. albicans cells was assessed following a single cleaning event. The immersion cleaner was significantly less abrasive than paste cleaners (P < 0·05). There were no significant differences between any dilutions for any cleaner used (P > 0·05). Immersion and paste cleaners removed almost all recoverable C. albicans from the discs, as cleaning with water alone was less effective (P < 0·05). An immersion type cleaner was found to be the most suitable cleaner because of its low abrasivity and effective removal of organic debris.
Prediction of different mandibular activities by EMG signal levels
Authors: Acosta‐Ortiz R. , Schulte J. K. , Sparks B. , Marsh W.
Published: May 2004

summary  The purpose of this study was to make observations of electromyographic (EMG) signal levels from a group of individuals to describe different mandibular tasks and use them as predictors of these activities. The sample consisted of 22 volunteer subjects (12 women and 10 men) with mean values for age, weight and height of 29·3 years, 69 kg and 168 cm respectively. Three separate recording sessions were performed for each subject. During each session the subjects were asked to perform different voluntary mandibular tasks. The EMG activity for each task was recorded from the anterior temporalis (Ta) and masseter muscles (Mm) bilaterally. The EMG recordings were related to task, session, task–session interaction, gender, age, weight and height using SAS version 6·12. The EMG data from the four muscles were considered as covariates to classify tasks. Overall, 78% of the group tasks were correctly classified. When the masticatory activities were classified as high and low EMG signal level group, the overall classification rate improved with an accuracy of 88%, sensitivity of 86%, specificity of 88%, positive predictive value of 76% and negative predictive value of 94%. The tasks, sessions, task–session interactions, gender, age, weight and height did not have any significant effect on the EMG recordings. It can be concluded that distinguishing among different mandibular tasks of a subject can be achieved when the individual EMG signal levels were compared with the EMG signal levels of subjects that were used to describe different mandibular activities.
Effect of age on salivary flow obtained under feeding and non‐feeding conditions
Authors: Bourdiol P. , Mioche L. , Monier S.
Published: May 2004

summary  Age, health status and disease treatments are thought to influence salivary flow. In this study, age effect on salivation was compared in non‐feeding (at rest and during parafilm chewing) and feeding (during meat chewing) conditions for two groups of healthy subjects, 25 young subjects (mean age 27·4 years) and 20 old subjects (mean age 71·2 years). In non‐feeding conditions, parotid flow was assessed at rest (3 min) and during parafilm chewing (1 min) from the absorptive capacity of a cotton roll placed in front of the upper duct apertures. Remaining saliva emanating mainly from the submandibular/sublingual glands was determined at rest by a sublingual cotton roll. In order not to impede in the chewing process during parafilm chewing, no cotton roll was placed in the lower part of the mouth and the remaining saliva was simply spit out for evaluation. Assessments were made under feeding conditions during the mastication of meat of different textures. The saliva content of the bolus was evaluated at different stages of the chewing process by weighing the mouth contents after spitting. No direct age effect was found on the different salivary flow rates measured during different conditions of stimulation. However, a significant correlation was found between the salivary flow rates at rest and those obtained during meat chewing in the elderly group but not in the young group. In elderly adults, rest salivary flow rate appears as a good predictor of salivary flow during the consumption of food. Within each group, significant correlations were found between salivation elicited by meat and by parafilm chewing. These results confirm the lack of direct global age effect on salivary flow rate by chewing in the 3 min after the stimulation, although adaptations to the measurement conditions are different between both groups of subjects.
Efficacy of sodium hypochlorite and coconut soap used as disinfecting agents in the reduction of denture stomatitis, Streptococcus mutans and Candida albicans
Authors: Barnabé W. , de Mendonça Neto T. , Pimenta F. C. , Pegoraro L. F. , Scolaro J. M.
Published: May 2004

summary  This study evaluated the reduction of denture stomatitis and the antimicrobial activity of 0·05% sodium hypochlorite opposed to Candida albicans and Streptococcus mutans (SGM) when associated with brushing complete dentures with coconut soap. The mucosal characteristics were evaluated according to Newton's classification at baseline, after cleansing the dentures with coconut soap for 15 days in group 1 (nine patients). In the other group (19 patients) the analysis were made before and after cleansing the dentures with coconut soap and with disinfection in a soak solution of 0·05% sodium hypochlorite for 10 min during 15 days. Microbiological tests were used to isolate C. albicans and SGM. Mann–Whitney and Wilcoxon tests were used to compare the mucosal characteristics and Fisher test and McNemar test to compare C. albicans and SGM levels. Statistical analysis at the 95% confidence level (P < 0·05) showed that: (i) the association of coconut soap and 0·05% sodium hypochlorite significantly reduced clinical signs of denture stomatitis, (ii) C. albicans did not reduce in counts, (iii) SGM were reduced but not significantly and (iv) the association of coconut soap and 0·5% sodium hypochlorite was effective in controlling denture biofilm.
Morphology of the articular eminence in temporomandibular joints and condylar bone change
Authors: Yamada K. , Tsuruta A. , Hanada K. , Hayashi T.
Published: May 2004

summary  The purpose of the present study was to investigate the relationship between the inclination of the articular eminence and temporomandibular joint (TMJ) pathology in orthognathic surgery patients with signs and symptoms of TMJ disorders. Twenty‐one female orthognathic surgery patients with signs and symptoms of TMJ disorders were examined using pre‐treatment helical computed tomography scans. The slope of the eminence in the medial, central and lateral sections of the subjects with osteophyte formation was significantly less than in the subjects with no bone change, and the medial section of the subjects with osteophyte formation was also significantly less steep than in the subjects with erosion. The central and lateral sections in the subjects with anterior disc displacement with reduction were significantly steeper than in subjects with anterior disc displacement without reduction. These results suggest that eminence flattening might occur during changes from erosion to osteophyte formation and from anterior disc displacement with reduction to anterior disc displacement without reduction. This appears to represent adaptation of the condyle, articular disc and articular eminence to changes in loading.
Chewing side preference as a type of hemispheric laterality
Authors: Nissan J. , Gross M. D. , Shifman A. , Tzadok L. , Assif D.
Published: May 2004

summary  Chewing side preference is a factor that could effect prosthodontic treatment. The purpose of this study was to determine whether chewing side was another type of hemispheric lateralization comparable with footedness, handedness, eyedness and earedness. Chewing side preference was tested in 189 subjects of whom 84 were partially edentulous, 98 had a full compliment of dental units (81 included implant‐supported restoration restoring the missing teeth and 17 with fully intact dentitions), and seven were fully edentulous, restored with complete dentures. Laterality tests were carried out for the first cycle of mastication, handedness, footedness, earedness and eyedness and patient questionnaire. Most patients preferred chewing on the right side (78ḃ3%) and were right sided. Chewing side preference correlated with other tested hemispherical lateralities. Missing teeth, occlusion type, lateral guidance, gender, implant‐supported restorations and complete dentures do not affect the side preference for chewing. This presents a strong argument that chewing side preference is centrally controlled and provides food for thought regarding its significance in prosthodontics.
Edentulousness, denture wear and denture needs of Indian elderly – a community‐based study
Authors: Shah N. , Parkash H. , Sunderam K. R.
Published: May 2004

Summary  India has a large geriatric population (60+ years) of 77 millions, comprising 7·7% of its total population. One of the major handicaps in the elderly is loss of teeth, affecting their mastication, dietary intake and nutritional status. The present study was planned to assess the level of edentulousness, denture wearing and denture needs of the elderly in the community and to study the correlation between oral health parameters and socio‐demographic variables, diet and body mass index (BMI). A total of 1240 (716 urban and 524 rural) elderly people were studied. It was found that 60% were dissatisfied with their function of mastication because of loss of teeth and lack of dentures. The level of edentulousness was found to be high, more so in rural than in urban people and more so in advancing age, with no significant difference between male and females. The striking finding of this study was the very low level of denture wearing. Only <50% of those needing complete dentures and <13% of those needing partial dentures were wearing dentures. The denture needs of the rural elderly were higher than those of the urban elderly. Vegetarians had a higher level of edentulousness compared with non‐vegetarians. The BMI was not correlated with level of edentulousness.
Characteristics of the gum chewing occlusal phase in children with primary dentition
Authors: Saitoh I. , Hayasaki H. , Nakata S. , Iwase Y. , Nakata M.
Published: May 2004

Summary  Understanding of the growth and development of mandibular function is indispensable to the diagnosis of functional disturbances during childhood. The purpose of this study was to clarify the characteristics of the occlusal phase during gum chewing in children with primary dentition. Chewing motion at the working molar of 14 children with primary dentition and 28 female adults was recorded optoelectrically, and the frontal and sagittal angles of their closing and following opening strokes were measured and compared. In children the closing strokes were entered more vertically and anteriorly than in adults, and the opening strokes shifted to the non‐working side in adults but moved to the working side in children. The degree of variance also differed between the two groups; the variance of the frontal angle was larger than that of sagittal angle in adults, but the opposite was true in children. These results suggested that the chewing pattern in children during the occlusal phase is distinctly different from adults and the chewing movement in children is not always less stable than in adults.
The biomechanical relationship between incisor and condylar guidances in deep bite and normal cases
Authors: Darendeliler N. , Dinçer M. , Soylu R.
Published: May 2004

Summary  The purpose of this study was to investigate the biomechanical relationship between the condylar and anterior guidances in deep bite malocclusion and control groups. The subjects consisted of 18 deep bite patients and 14 normal occlusions. A four‐bar mechanism was described on the lateral cephalogram films and a computer program, for which the lengths of the links measured on the films were the inputs, was developed. The computer program was used to calculate the angles of rotation of the mandible and the condyle during contact of the mandibular anterior teeth with the lingual surface of the maxillary anterior teeth as the mandible moves forward. Comparison of the mean values of the angles of rotation of the mandible and condyle in the two groups has shown that the change in the angle of rotation of the condyle is statistically significant (P < 0.05). The angles of rotation of condyle in deep bite group were found larger than the angles of normal group. Correlation between the angles of rotation of the condyle and mandible has shown that they were significantly related. The cause of temporomandibular disorders may be attributed to the large angle of rotation of the condyle in deep incisal overbite.
Influence of occlusal contact on cervical muscle activity during submaximal clenching
Authors: So K. , Komiyama O. , Arai M. , Kawara M. , Kobayashi K.
Published: May 2004

Summary  The aim of this study was to clarify the relationship between changes in the occlusal contact area and cervical muscle activity. A decreases in the occlusal contact area using bite planes (stage 1: full contact bite plane, stage 2: bilateral molar removed from contact, and stage 3: bilateral molar and premolar removed from contact) was experimentally simulated in seven subjects (aged 23–25 years) with normal dentition, and muscle activity of the sternocleidomastoid muscle (SCM) and trapezius muscle (TRM) during 50 and 10% of maximum voluntary contraction (MVC) was measured by surface electromyography. The SCM activity during 50% MVC was stage 1: 31·2 ± 9·4 μV and stage 3: 35·3 ± 12·3 μV. The TRM activity during 50% MVC was stage 1: 15·2 ± 0·7 μV and stage 3: 18·3 ± 2·9 μV. At the 50% MVC, stage 3 showed significant differences in comparison with stage 1 (P < 0·05, anova). These findings suggested that the sternocleidomastoid and TRM play roles in the exertion of occlusal force, and decreases in the occlusal contact area influence the amount of SCM and TRM activity.
Influence of masticatory muscle pain on electromyographic activities of cervical muscles in patients with myogenous temporomandibular disorders
Authors: Pallegama R. W. , Ranasinghe A. W. , Weerasinghe V. S. , Sitheeque M. A. M.
Published: May 2004

Summary  The present study tested the hypothesis that the resting electromyographic (EMG) activities of sternocleidomastoid and trapezius muscles of myogenous temporomandibular disorder (TMD) patients would be significantly greater than healthy individuals. Eight masticatory muscle pain patients without disc interference disorders (DID) and 30 patients with DID along with 41 matched control subjects were included in the study. The resting EMG activities of sternocleidomastoid and trapezius muscles were recorded with a portable EMG machine. Pain intensities described over the muscles were recorded with a 100 mm visual analogue scale. The two groups of patients had significantly higher resting activities compared with control subjects. Patients (in both groups) who had pain in sternocleidomastoid and trapezius muscles reported significantly higher resting activities in the respective muscles than patients who were free of pain and the controls. The presence of pain over the examined muscles showed significant association with the myogenous TMDs without DID. The observed association of the cervical muscles with myogenous TMDs is expected to be a result of functional link between masticatory and cervical muscles. The present findings suggest that the myogenous TMDs without DID could be a distinct sub‐group that can be a part of broader regional or generalized diseased entity.
In vitro investigation of marginal accuracy of implant‐supported screw‐retained partial dentures
Authors: Koke U. , Wolf A. , Lenz P. , Gilde H.
Published: May 2004

Summary  Mismatch occurring during the fabrication of implant‐supported dentures may induce stress to the peri‐implant bone. The purpose of this study was to investigate the influence of two different alloys and the fabrication method on the marginal accuracy of cast partial dentures. Two laboratory implants were bonded into an aluminium block so that the distance between their longitudinal axes was 21 mm. Frameworks designed for screw‐retained partial dentures were cast either with pure titanium (rematitan) or with a CoCr‐alloy (remanium CD). Two groups of 10 frameworks were cast in a single piece. The first group was made of pure titanium, and the second group of a CoCr‐alloy (remanium CD). A third group of 10 was cast in two pieces and then laser‐welded onto a soldering model. This latter group was also made of the CoCr‐alloy. All the frameworks were screwed to the original model with defined torque. Using light microscopy, marginal accuracy was determined by measuring vertical gaps at eight defined points around each implant. Titanium frameworks cast in a single piece demonstrated mean vertical gaps of 40 μm (s.d. = 11 μm) compared with 72 μm (s.d. = 40 μm) for CoCr‐frameworks. These differences were not significant (U‐test, P = 0·124) because of a considerable variation of the values for CoCr‐frameworks (minimum: 8 μm and maximum: 216 μm). However, frameworks cast in two pieces and mated with a laser showed significantly better accuracy in comparison with the other experimental groups (mean: 17 μm ± 6; P < 0·01). (i) The fit of implant‐supported partial dentures cast with pure titanium in a single piece is preferable to that of those made with the CoCr‐alloy and (ii) the highest accuracy can be achieved by using a two‐piece casting technique combined with laser welding. Manufacturing the framework pieces separately and then welding them together provides the best marginal fit.
The effect of thermocycling and dentine pre‐treatment on the durability of the bond between composite resin and dentine
Authors: Huang M. S. , Li M. T. , Huang F. M. , Ding S. J.
Published: May 2004

Summary  The high bond strength between restorative resin and dentine plays an important role in long‐term performance of restorations in the oral environment. A variety of treatment techniques have been described to enhance the bond strength of composite resin to dentine. Unfortunately, few studies have reported available bond durability of adhesive resins to dentine. The purpose of this research was to study the shear bond strength of composite resin to dentine pre‐treated with phosphoric acid, self‐etching agent or Nd:YAP laser irradiation. The durability of bond strength between resin and dentine stored in the artificial saliva thermocycling between 5 and 55 °C was also evaluated. The scanning electron microscope was used to assess the treated‐dentine surfaces. The mean value of the shear bond strength in the acid‐etching group (18·2 ± 4·9 MPa) was the highest among the three dentine treatments (self‐etching system: 12·6 ±3·0 MPa, Nd:YAP laser: 13·4 ± 3·3 MPa) prior to thermocycling. After thermocycling, shear strength values of all treated dentines decreased with increasing number of the cycles. When subjected to 3000 thermocycles, the mean bond strengths of these pre‐treated samples to composite resin became 9·1 ± 1·4, 7·8 ± 1·8, and 8·1 ± 1·7 MPa for acid‐etching, self‐etching and laser‐irradiation, respectively, with a significant reduction of 38–50%.
Influence of different light sources on microleakage of class V composite resin restorations
Authors: Nilgun Ozturk A. , Usumez A. , Ozturk B. , Usumez S.
Published: May 2004

Summary  The aim of this study was to evaluate the effect of three different curing units on microleakage of class V composite restorations. Class V cavities were prepared on the buccal surfaces of 45 extracted premolar teeth. The teeth were randomly divided into three groups of 15 each. A conventional halogen curing unit (Hilux 350), a high intensity halogen curing unit (Optilux 501) and plasma arc curing unit (Power Pac) were used to polymerize composite resin (Vitalesence). After restoration, the teeth were thermocycled for 100 cycles between 5 ° and 55 °C using a dwell time of 30 s and exposed to a dye. Results showed that there was no significant difference among three different curing units (P > 0·05), however the microleakage at the dentin margins was greater than the enamel margins (P < 0·05). Clinical relevance: Plasma arc curing or fast halogen units cure composite materials at a faster rate than conventional curing units because of the high light intensity. High intensity halogen curing units and plasma arc curing units might be useful alternatives in composite polymerization. Therefore, these units are suggested for clinical use to save chair side time.
Genome‐wide analysis of transcriptional hierarchy and feedback regulation in the flagellar system of Helicobacter pylori
Authors: Niehus Eike , Gressmann Helga , Ye Fang , Schlapbach Ralph , Dehio Michaela , Dehio Christoph , Stack Allison , Meyer Thomas F. , Suerbaum Sebastian , Josenhans Christine
Published: May 2004

Summary The flagellar system of Helicobacter pylori, which comprises more than 40 mostly unclustered genes, is essential for colonization of the human stomach mucosa. In order to elucidate the complex transcriptional circuitry of flagellar biosynthesis in H. pylori and its link to other cell functions, mutants in regulatory genes governing flagellar biosynthesis (rpoN, flgR, flhA, flhF, HP0244) and whole‐genome microarray technology were used in this study. The regulon controlled by RpoN, its activator FlgR (FleR) and the cognate histidine kinase HP0244 (FleS) was characterized on a genome‐wide scale for the first time. Seven novel genes (HP1076, HP1233, HP1154/1155, HP0366/367, HP0869) were identified as belonging to RpoN‐associated flagellar regulons. The hydrogenase accessory gene HP0869 was the only annotated non‐flagellar gene in the RpoN regulon. Flagellar basal body components FlhA and FlhF were characterized as functional equivalents to master regulators in H. pylori, as their absence led to a general reduction of transcripts in the RpoN (class 2) and FliA (class 3) regulons, and of 24 genes newly attributed to intermediate regulons, under the control of two or more promoters. FlhA‐ and FlhF‐dependent regulons comprised flagellar and non‐flagellar genes. Transcriptome analysis revealed that negative feedback regulation of the FliA regulon was dependent on the antisigma factor FlgM. FlgM was also involved in FlhA‐ but not FlhF‐dependent feedback control of the RpoN regulon. In contrast to other bacteria, chemotaxis and flagellar motor genes were not controlled by FliA or RpoN. A true master regulator of flagellar biosynthesis is absent in H. pylori, consistent with the essential role of flagellar motility and chemotaxis for this organism.
Structural tolerance of bacterial autotransporters for folded passenger protein domains
Authors: Veiga Esteban , De Lorenzo Víctor , Fernández Luis Angel
Published: May 2004

In this report we investigate the capacity of bacterial autotransporters (AT) to translocate folded protein domains across the outer membrane (OM). Polypeptides belonging to the AT family contain a C‐terminal domain that supports the secretion of the N‐domain (the passenger) across the OM of Gram‐negative bacteria. Despite some controversial data, it has been widely accepted that N‐passenger domains of AT must be unfolded and devoid of disulphide bonds for efficient translocation. To address whether or not AT are able to translocate folded protein domains across the OM, we employed several types of recombinant antibodies as heterologous N‐passengers of the transporter C‐domain of IgA protease (C‐IgAP) of Neisseria gonorroheae. The N‐domains used were single chain Fv fragments (scFv) and variable mono‐domains derived from camel antibodies (VHH) selected on the basis of their distinct and defined folding properties (i.e. enhanced solubility, stability and presence or not of disulphide bonds). Expression of these hybrids in Escherichia coli shows that stable scFv and VHH domains are efficiently (>99%) translocated towards the bacterial surface regardless of the presence or not of disulphide bonds on their structure. Antigen‐binding assays demonstrate that surface‐exposed scFv and VHH domains are correctly folded and thus able to bind their cognate antigens. Expression of scFv‐ or VHH‐C‐IgAP hybrids in E. coli dsbA or fkpA mutant cells reveals that these periplasmic protein chaperones fold these N‐domains before their translocation across the OM. Furthermore, large N‐passengers composed of strings of VHH domains were secreted in a folded state by AT with no loss of efficacy (>99%) despite having multiple disulphide bonds. Thus AT can efficiently translocate toward the cell surface folded N‐passengers composed of one, two or three immunoglobulin (Ig) domains, each with a folded diameter between ∼2 nm and having disulphide bonds. This tolerance for folded protein domains of ∼2 nm fits with the diameter of the central hydrophilic channel proposed for the ring‐like oligomeric complex assembled by C‐IgAP in the OM.
Fur functions as an activator and as a repressor of putative virulence genes in Neisseria meningitidis
Authors: Delany Isabel , Rappuoli Rino , Scarlato Vincenzo
Published: May 2004

Fur is a well‐known iron‐responsive repressor of gene transcription, which is used by many bacteria to respond to the low‐iron environment that pathogens encounter during infection. Four promoters of Neisseria meningitidis predicted to have Fur‐binding boxes were selected to study the molecular interactions between Fur and the promoter regions of genes expected to play a central role in survival and pathogenesis. We demonstrate that Fur acts not only as a repressor, but also as an activator of gene expression both in vivo and in vitro. We report that Fur binds to operators located upstream of three promoters that are positively regulated in vivo by Fur and iron, whereas Fur binds to an operator overlapping the classically iron‐repressed tbp promoter. Deletion of the upstream operator in the norB promoter abolished activation of transcription in vivo in response to iron and in vitro in response to Fur. The role of such a dual mechanism of Fur regulation during infection is discussed.
The Bacillus subtilisσW anti‐sigma factor RsiW is degraded by intramembrane proteolysis through YluC
Authors: Schöbel Susanne , Zellmeier Stephan , Schumann Wolfgang , Wiegert Thomas
Published: May 2004

The Bacillus subtilisσW regulon is induced by different stresses such as alkaline shock, salt shock, phage infection and certain antibiotics that affect cell wall biosynthesis. The activity of the alternative, extracytoplasmic function (ECF) sigma factor σW is modulated by a specific anti‐sigma factor (RsiW or YbbM) encoded by the rsiW (ybbM) gene located immediately downstream of sigW. The RsiW membrane topology was determined, and a specific reporter system for RsiW function was constructed. Experiments using the yeast two‐hybrid system suggested a direct interaction of σW with the cytoplasmic part of RsiW. Analysis of truncated forms of the RsiW protein revealed that σW induction by alkaline shock is dependent on both the transmembrane and the extracytoplasmic domain of RsiW. Western blot and pulse–chase experiments demonstrated degradation of RsiW after an alkaline shock. A B. subtilis mutant strain deleted for the Escherichia coli yaeL orthologue yluC, encoding a transmembrane protease, was defective in inducing a σW‐controlled promoter after alkaline shock and accumulated a membrane‐bound truncated form of RsiW, suggesting that the activity of σW is controlled by the proteolysis of RsiW by at least two different proteolytic steps.
Characterization of an inducible vancomycin resistance system in Streptomyces coelicolor reveals a novel gene (vanK) required for drug resistance
Authors: Hong Hee‐Jeon , Hutchings Matthew I. , Neu John M. , Wright Gerard D. , Paget Mark S. B. , Buttner Mark J.
Published: May 2004

Vancomycin is the front‐line therapy for treating problematic infections caused by methicillin‐resistant Staphylococcus aureus (MRSA), and the spread of vancomycin resistance is an acute problem. Vancomycin blocks cross‐linking between peptidoglycan intermediates by binding to the d‐Ala‐d‐Ala termini of bacterial cell wall precursors, which are the substrate of transglycosylase/transpeptidase. We have characterized a cluster of seven genes (vanSRJKHAX) in Streptomyces coelicolor that confers inducible, high‐level vancomycin resistance. vanHAX are orthologous to genes found in vancomycin‐resistant enterococci that encode enzymes predicted to reprogramme peptidoglycan biosynthesis such that cell wall precursors terminate in d‐Ala‐d‐Lac rather than d‐Ala‐d‐Ala. vanR and vanS encode a two‐component signal transduction system that mediates transcriptional induction of the seven van genes. vanJ and vanK are novel genes that have no counterpart in previously characterized vancomycin resistance clusters from pathogens. VanK is a member of the Fem family of enzymes that add the cross‐bridge amino acids to the stem pentapeptide of cell wall precursors, and vanK is essential for vancomycin resistance. The van genes are organized into four transcription units, vanRS, vanJ, vanK and vanHAX, and these transcripts are induced by vancomycin in a vanR‐dependent manner. To develop a sensitive bioassay for inducers of the vancomycin resistance system, the promoter of vanJ was fused to a reporter gene conferring resistance to kanamycin. All the inducers identified were glycopeptide antibiotics, but teicoplanin, a membrane‐anchored glycopeptide, failed to act as an inducer. Analysis of mutants defective in the vanRS and cseBC cell envelope signal transduction systems revealed significant cross‐talk between the two pathways.
Differential recognition of members of the carcinoembryonic antigen family by Afa/Dr adhesins of diffusely adhering Escherichia coli (Afa/Dr DAEC)
Authors: Berger Cedric N. , Billker Oliver , Meyer Thomas F. , Servin Alain L. , Kansau Imad
Published: May 2004

Little is known about the molecular bases underlying the virulence of diffusely adhering Escherichia coli (DAEC) harbouring the Afa/Dr family of adhesins. These adhesins recognize as receptors the GPI‐anchored proteins CD55 (decay‐accelerating factor, DAF) and CD66e (carcinoembryonic antigen, CEA). CD66e is a member of the CEA‐related cell adhesion molecules (CEACAM) family, comprising seven members. We analysed the interactions of Afa/Dr DAEC with the CEACAMs using CEACAM‐expressing CHO and HeLa cells. The results demonstrate that only E. coli expressing a subfamily of Afa/Dr adhesins, named here Afa/Dr‐I, including Dr, F1845 and AfaE‐III adhesins, bound onto CHO cells expressing CEACAM1, CEA or CEACAM6. Whereas all the Afa/Dr adhesins elicit recruitment of CD55 around adhering bacteria, only the Afa/Dr‐I subfamily elicits the recruitment of CEACAM1, CEA and CEACAM6. In addition, although CEACAM3 is not recognized as a receptor by the subfamily of Afa/Dr adhesins, it is recruited around bacteria in HeLa cells. The recruited CEACAM1, CEA and CEACAM6 around adhering bacteria resist totally or in part a detergent extraction, whereas the recruited CEACAM3 does not. Finally, the results show that recognition of CEA and CEACAM6, but not CEACAM1, is accompanied by tight attachment to bacteria of cell surface microvilli‐like extensions, which are elongated. Moreover, recognition of CEA is accompanied by an activation of the Rho GTPase Cdc42 and by a phosphorylation of ERM, which in turn elicit the observed cell surface microvilli‐like extensions.
Analysis of the 5S rRNA gene promoter from Acanthamoeba castellanii
Authors: Peng Zhihua , Bateman Erik
Published: May 2004

The promoter region of the Acanthamoeba 5S rRNA gene was analysed by in vitro transcription of several 5′ and 3′ deletion and substitution mutants, as well as a series of linker scanning mutants. The promoter consists of three sequence regions contained entirely within the gene; two of these correspond to the A and C boxes that bind TFIIIA, found in the genes from other genera. In addition, a region immediately 3′ to the transcription start site has a strong effect on initiation efficiency. No strict requirement was found for specific sequences 5′ to the transcription start site. Substitution of a consensus TATA box at −29 had only a modest effect on transcription, and deletion or substitution of sequences between −15 and −10 as well as −34 and −21 was only modestly more active than the wild‐type template. Analysis of 3′ deletions sets the 3′ end‐point of the promoter between +79 and +97, and demonstrates the importance of a T‐rich region in transcription termination. Taken together, these results suggest that promoter elements within the Acanthamoeba 5S RNA gene are somewhat redundant, with the exception of a sequence between +50 and +60, which functions in binding TFIIIA. Remarkably, polymerase chain reaction product templates containing only non‐specific 5′ ends between −6 and +1 relative to the transcription start site are fully functional, demonstrating that no external DNA scaffold is needed for TFIIIB and RNA polymerase III binding, and that productive initiation can be mediated solely by protein–DNA interactions within the coding region of the 5S gene.
Dissecting the loci of low‐level quinine resistance in malaria parasites
Authors: Ferdig Michael T. , Cooper Roland A. , Mu Jianbing , Deng Bingbing , Joy Deirdre A. , Su Xin‐zhuan , Wellems Thomas E.
Published: May 2004

Quinine (QN) remains effective against Plasmodium falciparum, but its decreasing efficacy is documented from different continents. Multiple genes are likely to contribute to the evolution of QN resistance. To locate genes contributing to QN response variation, we have searched a P. falciparum genetic cross for quantitative trait loci (QTL). Results identify additive QTL in segments of chromosomes (Chrs) 13, 7 and 5, and pairwise effects from two additional loci of Chrs 9 and 6 that interact, respectively, with the QTL of Chrs 13 and 7. The mapped segments of Chrs 7 and 5 contain pfcrt, the determinant of chloroquine resistance (CQR), and pfmdr1, a gene known to affect QN responses. Association of pfcrt with a QTL of QN resistance supports anecdotal evidence for an evolutionary relationship between CQR and reduced QN sensitivity. The Chr 13 segment contains several candidate genes, one of which (pfnhe‐1) encodes a putative Na+/H+ exchanger. A repeat polymorphism in pfnhe‐1 shows significant association with low QN response in a collection of P. falciparum strains from Asia, Africa and Central and South America. Dissection of the genes and modifiers involved in QN response will require experimental strategies that can evaluate multiple genes from different chromosomes in combination.
Environmental signals and regulatory pathways that influence biofilm formation
Authors: Stanley Nicola R. , Lazazzera Beth A.
Published: May 2004

In nature, bacteria often exist as biofilms. Here, we discuss the environmental signals and regulatory proteins that affect both the initiation of bacterial biofilm formation and the nature of the mature biofilm structure. Current research suggests that the environmental signals regulating whether bacterial cells will initiate a biofilm differ from one bacterial species to another. This may allow each bacterial species to colonize its preferred environment efficiently. In contrast, some of the environmental signals that have currently been identified to regulate the structure of a mature biofilm are nutrient availability and quorum sensing, and are not species specific. These environmental signals evoke changes in the nature of the mature biofilm that may ensure optimal nutrient acquisition. Nutrient availability regulates the depth of the biofilm in such a way that the maximal number of cells in a biofilm appears to occur at suboptimal nutrient concentrations. At either extreme, nutrient‐rich or very nutrient‐poor conditions, greater numbers of cells are in the planktonic phase where they have greater access to the local nutrients or can be distributed to a new environment. Similarly, quorum‐sensing control of the formation of channels and pillar‐like structures may ensure efficient nutrient delivery to cells in a biofilm.
IS6110 functions as a mobile, monocyte‐activated promoter in Mycobacterium tuberculosis
Authors: Safi Hassan , Barnes Peter F. , Lakey David L. , Shams Homayoun , Samten Buka , Vankayalapati Ramakrishna , Howard Susan T.
Published: May 2004

The mobile insertion sequence, IS6110, is an important marker in tracking of Mycobacterium tuberculosis strains. Here, we demonstrate that IS6110 can upregulate downstream genes through an outward‐directed promoter in its 3′ end, thus adding to the significance of this element. Promoter activity was orientation dependent and was localized within a 110 bp fragment adjacent to the right terminal inverted repeat. Transcripts from this promoter, named OP6110, begin ≈ 85 bp upstream of the 3′ end of IS6110. Use of green fluorescent protein (GFP) expression constructs showed that OP6110 was upregulated in M. tuberculosis during growth in human monocytes and in late growth phases in broth. Analysis of natural insertion sites in M. tuberculosis showed that IS6110 upregulated expression of several downstream genes during growth in human monocytes, including Rv2280 in H37Rv and the PE‐PGRS gene, Rv1468c, in the clinical strain 210, which is a member of the Beijing family. Transcription between IS6110 and downstream genes was confirmed by reverse transcription polymerase chain reaction. The ability to activate genes during infection suggests that IS6110 has the potential to influence growth characteristics of different strains, and indicates another mechanism by which IS6110 can impact M. tuberculosis evolution.
Phage T4 early promoters are resistant to inhibition by the anti‐sigma factor AsiA
Authors: Orsini Gilbert , Igonet Sébastien , Pène Carole , Sclavi Bianca , Buckle Malcolm , Uzan Marc , Kolb Annie
Published: May 2004

Phage T4 early promoters are transcribed in vivo and in vitro by the Escherichia coli RNA polymerase holoenzyme Eσ70. We studied in vitro the effects of the T4 anti‐σ70 factor AsiA on the activity of several T4 early promoters. In single‐round transcription, promoters motB, denV, mrh.2, motA wild type and UP element‐deleted motA are strongly resistant to inhibition by AsiA. The α‐C‐terminal domain of Eσ70 is crucial to this resistance. DNase I footprinting of Eσ70 and Eσ70AsiA on motA and mrh.2 shows extended contacts between the holoenzyme with or without AsiA and upstream regions of these promoters. A TG → TC mutation of the extended −10 motif in the motA UP element‐deleted promoter strongly increases susceptibility to inhibition by AsiA, but has no effect on the motA wild‐type promoter: either the UP element or the extended −10 site confers resistance to AsiA. Potassium permanganate reactivity shows that the two structure elements are not equivalent: with AsiA, the motA UP element‐deleted promoter opens more slowly whereas the motA TC promoter opens like the wild type. Changes in UV laser photoreactivity at position +4 on variants of motA reveal an analogous distinction in the roles of the extended −10 and UP promoter elements.
Transcriptional fidelity and proofreading in Archaea and implications for the mechanism of TFS‐induced RNA cleavage
Authors: Lange Udo , Hausner Winfried
Published: May 2004

We have addressed the question whether TFS, a protein that stimulates the intrinsic cleavage activity of the archaeal RNA polymerase, is able to improve the fidelity  of  transcription  in  Methanococcus.  Using non‐specific transcription experiments, we could demonstrate that misincorporation of non‐templated nucleotides  is  reduced  in  the  presence  of  TFS.  A more detailed analysis revealed that elongation complexes containing a misincorporated nucleotide were arrested, but could be reactivated by TFS. RNase as well as exonuclease III footprinting experiments demonstrated that this arrest was not combined with extended backtracking. Analysis of paused elongation complexes demonstrated that TFS is able to induce a cleavage resynthesis cycle in such complexes, which resulted in the accumulation of dinucleotides corresponding to the last two nucleotides of the transcript. Further analysis of cleavage products revealed that, even under conditions that strongly promote misincorporation, still 50% of the released dinucleotides were correctly incorporated. Therefore, we assume that pausing of elongation complexes is an important determinant of TFS‐induced RNA cleavage from the 3′ end. As the incorporation rate of wrong nucleotides is about 700‐fold reduced, it is possible that this delay also provides an appropriate time window for cleavage induction in order to maintain transcriptional fidelity by preventing misincorporation.
Proteomic screening and identification of differentially distributed membrane proteins in Escherichia coli
Authors: Lai Erh‐Min , Nair Usha , Phadke Nikhil D. , Maddock Janine R.
Published: May 2004

Bacteria show asymmetric subcellular distribution of many proteins involved in diverse cellular processes such as chemotaxis, motility, actin polymerization, chromosome partitioning and cell division. In many cases, the specific subcellular localization of these proteins is critical for proper regulation and function. Although cellular organization of the bacterial cell clearly plays an important role in cell physiology, systematic studies to uncover asymmetrically distributed proteins have not been reported previously. In this study, we undertook a proteomics approach to uncover polar membrane proteins in Escherichia coli. We identified membrane proteins enriched in E. coli minicells using a combination of two‐dimensional electrophoresis and mass spectrometry. Among a total of 173 membrane protein spots that were consistently detected, 36 spots were enriched in minicell membranes, whereas 15 spots were more abundant in rod cell membranes. The minicell‐enriched proteins included the inner membrane proteins MCPs, AtpA, AtpB, YiaF and AcrA, the membrane‐associated FtsZ protein and the outer membrane proteins YbhC, OmpW, Tsx, Pal, FadL, OmpT and BtuB. We immunolocalized two of the minicell‐enriched proteins, OmpW and YiaF, and showed that OmpW is a bona fide polar protein whereas YiaF displays a patchy membrane distribution with a polar and septal bias.
Molecular analysis of the key cytokinetic components of cyanobacteria: FtsZ, ZipN and MinCDE
Authors: Mazouni Khalil , Domain Francis , Cassier‐Chauvat Corinne , Chauvat Franck
Published: May 2004

Using a bacterial two‐hybrid system and a combination of in vivo and in vitro assays that take advantage of the green fluorescent reporter protein (GFP), we have investigated the localization and the protein–protein interaction of several key components of the cytokinetic machinery of cyanobacteria (i.e. the progenitor of chloroplast). We demonstrate that (i) the ftsZ and zipN genes are essential for the viability of the model cyanobacterium Synechocystis sp. PCC 6803, whereas the minCDE cluster is dispensable for cell growth; (ii) the GTP‐binding domain of FtsZ is crucial to FtsZ assembly into the septal ring at mid‐cell; (iii) the Z‐ring of deeply constricted daughter cells is oriented perpendicularly to the mother Z‐ring, showing that Synechocystis divides in alternating perpendicular planes; (iv) the MinCDE system affects the morphology of the cell, as well as the position and the shape of FtsZ structures; and (v) MinD is targeted to cell membranes in a process involving its C‐terminal amphipathic helix, but not its ATP‐binding region. Finally, we have also characterized a novel Z‐interacting protein, ZipN, the N‐terminal DnaJ domain of which is critical to the decoration of the Z‐ring, and we report that this process is independent of MinCDE.