bluuebunny/crossref_metadata_2025 huggingface dataset:
Pretreatment verification of IMRT absolute dose distributions using a commercial EPID
Authors: Talamonti C. , Casati M. , Bucciolini M.
Published: November 2006

A commercial amorphous silicon electronic portal imaging device (EPID) has been studied to investigate its potential in the field of pretreatment verifications of step and shoot, intensity modulated radiation therapy (IMRT), photon beams. The EPID was calibrated to measure absolute exit dose in a water‐equivalent phantom at patient level, following an experimental approach, which does not require sophisticated calculation algorithms. The procedure presented was specifically intended to replace the time‐consuming in‐phantom film dosimetry. The dosimetric response was characterized on the central axis in terms of stability, linearity, and pulse repetition frequency dependence. The EPID demonstrated a good linearity with dose (within 2% from 1 monitor unit), which represent a prerequisite for the application in IMRT. A series of measurements, in which phantom thickness, air gap between the phantom and the EPID, field size and position of measurement of dose in the phantom (entrance or exit) varied, was performed to find the optimal calibration conditions, for which the field size dependence is minimized. In these conditions ( phantom thickness, air gap, exit dose measured at the isocenter), the introduction of a filter for the low‐energy scattered radiation allowed us to define a universal calibration factor, independent of field size. The off‐axis extension of the dose calibration was performed by applying a radial correction for the beam profile, distorted due to the standard flood field calibration of the device. For the acquisition of IMRT fields, it was necessary to employ home‐made software and a specific procedure. This method was applied for the measurement of the dose distributions for 15 clinical IMRT fields. The agreement between the dose distributions, quantified by the gamma index, was found, on average, in 97.6% and 98.3% of the analyzed points for EPID versus TPS and for EPID versus FILM, respectively, thus suggesting a great potential of this EPID for IMRT dosimetric applications.
Comparison of different commercial FFDM units by means of physical characterization and contrast‐detail analysis
Authors: Rivetti Stefano , Lanconelli Nico , Campanini Renato , Bertolini Marco , Borasi Gianni , Nitrosi Andrea , Danielli Claudio , Angelini Lidia , Maggi Stefania
Published: November 2006

The purpose of this study was to perform a complete evaluation of three pieces of clinical digital mammography equipment. Image quality was assessed by performing physical characterization and contrast‐detail (CD) analysis. We considered three different FFDM systems: a computed radiography unit (Fuji “FCR 5000 MA”) and two flat‐panel units, the indirect conversion a‐Si based GE “Senographe 2000D” and the direct conversion a‐Se based IMS “Giotto Image MD.” The physical characterization was estimated by measuring the MTF, NNPS, and DQE of the detectors with no antiscatter grid and over the clinical range of exposures. The CD analysis was performed using a CDMAM 3.4 phantom and custom software designed for automatic computation of the contrast‐detail curves. The physical characterization of the three digital systems confirms the excellent MTF properties of the direct conversion flat‐panel detector (FPD). We performed a relative standard deviation (RSD) analysis, for investigating the different components of the noise presented by the three systems. It turned out that the two FPDs show a significant additive component, whereas for the CR system the statistical noise is dominant. The multiplicative factor is a minor constituent for all the systems. The two FPDs demonstrate better DQE, with respect to the CR system, for exposures higher than . The CD analysis indicated that the three systems are not statistically different for detail objects with a diameter greater than . However, the IMS system showed a statistically significant different response for details smaller than . In this case, the poor response of the a‐Se detector could be attributed to its high‐frequency noise characteristics, since its MTF, NEQ, and DQE are not inferior to those of the other systems. The CD results were independent of exposure level, within the investigated clinical range. We observed slight variations in the CD results, due to the changes in the visualization parameters (window/level and magnification factor). This suggests that radiologists would benefit from viewing images using varied window/level and magnification.
Residual motion of lung tumors in end‐of‐inhale respiratory gated radiotherapy based on external surrogates
Authors: Berbeco Ross I. , Nishioka Seiko , Shirato Hiroki , Jiang Steve B.
Published: November 2006

It has been noted that some lung tumors exhibit large periodic motion due to respiration. To limit the amount of dose to healthy lung tissues, many clinics have begun gating radiotherapy treatment using externally placed surrogates. It has been observed by several institutions that the end‐of‐exhale (EOE) tumor position is more reproducible than other phases of the breathing cycle, so the gating window is often set there. From a treatment planning perspective, end‐of‐inhale (EOI) phase might be preferred for gating because the expanded lungs will further decrease the healthy tissue within the treatment field. We simulate gated treatment at the EOI phase, using a set of recently measured internal/external anatomy patient data. This paper attempts to answer three questions: (1) How much is the tumor residual motion when we use an external surrogate gating window at EOI? (2) How could we reduce the residual motion in the EOI gating window? (3) Is there a preference for amplitude‐ versus phase‐based gating at EOI? We found that under free breathing conditions the residual motion of the tumors is much larger for EOI phase than for EOE phase. The mean values of residual motion at EOI were found to be 2.2 and for amplitude‐ and phase‐based gating, respectively, and, at EOE, 1.0 and for amplitude‐ and phase‐based gating, respectively. However, we note that the residual motion in the EOI gating window is correlated well with the reproducibility of the external surface position in the EOI phase. Using the results of a published breath‐coaching study, we deduce that the residual motion of a lung tumor at EOI would approach that at EOE, with the same duty cycle (30%), under breath‐coaching conditions. Additionally, we found that under these same conditions, phase‐based gating approaches the same residual motion as amplitude‐based gating, going from a 28% difference to 11%, for the patient with the largest difference between the two gating modalities. We conclude that it is feasible to achieve the same reproducibility of tumor location at EOI as at EOE if breath coaching is implemented, enabling us to reap the benefits of the dosimetric advantage of EOI gating.
Improved quantitation for PET/CT image reconstruction with system modeling and anatomical priors
Authors: Alessio Adam M. , Kinahan Paul E.
Published: November 2006

Accurate quantitation of positron emission tomography (PET) tracer uptake levels in tumors is important for staging and monitoring response to treatment. Quantitative accuracy in PET is particularly poor for small tumors because of system partial volume errors and smoothing operations. This work proposes a reconstruction algorithm to reduce the quantitative errors due to limited system resolution and due to necessary image noise reduction. We propose a method for finding and using the detection system response in the projection matrix of a statistical reconstruction algorithm. In addition, we use aligned anatomical information, available in PET/CT scanners, to govern the penalty term applied during each image update. These improvements are combined with Fourier rebinning in a clinically feasible algorithm for reconstructing fully three‐dimensional PET data. Results from simulation and measured studies show improved quantitation of tumor values in terms of bias and variance across multiple tumor sizes and activity levels with the proposed method. At common clinical image noise levels for the detection task, the proposed method reduces the error in maximum tumor values by 11% compared to filtered back‐projection and 5% compared to conventional iterative methods.
Performance tests of two portable mini gamma cameras for medical applications
Authors: Sánchez F. , Fernández M. M. , Giménez M. , Benlloch J. M. , Rodríguez‐Alvarez M. J. , García de Quirós F. , Lerche Ch. W. , Pavón N. , Palazón J. A. , Martínez J. , Sebastiá A.
Published: November 2006

We have developed two prototypes of portable gamma cameras for medical applications based on a previous prototype designed and tested by our group. These cameras use a continuous scintillation crystal coupled to the new flat‐panel‐type multianode position‐sensitive photomultiplier tube, H8500 from Hamamatsu Photonics. One of the prototypes, mainly intended for intrasurgical use, has a field of view of , and weighs . Its intrinsic resolution is better than and its energy resolution is about 13% at . The second prototype, mainly intended for osteological, renal, mammary, and endocrine (thyroid, parathyroid, and suprarenal) scintigraphies, weighs a total of . Its average spatial resolution is ; it has a field of view of , with an energy resolution of about 15% at . The main advantages of these gamma camera prototypes with respect to those previously reported in the literature are high portability and low weight, with no significant loss of sensitivity and spatial resolution. All the electronic components are packed inside the mini gamma cameras, and no external electronic devices are required. The cameras are only connected through the universal serial bus port to a portable PC. In this paper, we present the design of the cameras and describe the procedures that have led us to choose their configuration together with the most important performance features of the cameras. For one of the prototypes, clinical tests on melanoma patients are presented and images are compared with those obtained with a conventional camera.
Reconstruction of dynamic gated cardiac SPECT
Authors: Jin Mingwu , Yang Yongyi , King Michael A.
Published: November 2006

In this paper we propose an image reconstruction procedure which aims to unify gated single photon emission computed tomography (SPECT) and dynamic SPECT into a single method. We divide the cardiac cycle into a number of gate intervals as in gated SPECT, but treat the tracer distribution for each gate as a time‐varying signal. By using both dynamic and motion‐compensated temporal regularization, our reconstruction procedure will produce an image sequence that shows both cardiac motion and time‐varying tracer distribution simultaneously. To demonstrate the proposed reconstruction method, we simulated gated cardiac perfusion imaging using the gated mathematical cardiac‐torso (gMCAT) phantom with Tc99m‐Teboroxime as the imaging agent. Our results show that the proposed method can produce more accurate reconstruction of gated dynamic images than independent reconstruction of individual gate frames with spatial smoothness alone. In particular, our results show that the former could improve the contrast to noise ratio of a simulated perfusion defect by as much as 100% when compared to the latter.
Testing of the analytical anisotropic algorithm for photon dose calculation
Authors: Van Esch Ann , Tillikainen Laura , Pyykkonen Jukka , Tenhunen Mikko , Helminen Hannu , Siljamäki Sami , Alakuijala Jyrki , Paiusco Marta , Iori Mauro , Huyskens Dominique P.
Published: November 2006

The analytical anisotropic algorithm (AAA) was implemented in the Eclipse (Varian Medical Systems) treatment planning system to replace the single pencil beam (SPB) algorithm for the calculation of dose distributions for photon beams. AAA was developed to improve the dose calculation accuracy, especially in heterogeneous media. The total dose deposition is calculated as the superposition of the dose deposited by two photon sources (primary and secondary) and by an electron contamination source. The photon dose is calculated as a three‐dimensional convolution of Monte‐Carlo precalculated scatter kernels, scaled according to the electron density matrix. For the configuration of AAA, an optimization algorithm determines the parameters characterizing the multiple source model by optimizing the agreement between the calculated and measured depth dose curves and profiles for the basic beam data. We have combined the acceptance tests obtained in three different departments for 6, 15, and photon beams. The accuracy of AAA was tested for different field sizes (symmetric and asymmetric) for open fields, wedged fields, and static and dynamic multileaf collimation fields. Depth dose behavior at different source‐to‐phantom distances was investigated. Measurements were performed on homogeneous, water equivalent phantoms, on simple phantoms containing cork inhomogeneities, and on the thorax of an anthropomorphic phantom. Comparisons were made among measurements, AAA, and SPB calculations. The optimization procedure for the configuration of the algorithm was successful in reproducing the basic beam data with an overall accuracy of 3%, in the build‐up region, and 1%, elsewhere. Testing of the algorithm in more clinical setups showed comparable results for depth dose curves, profiles, and monitor units of symmetric open and wedged beams below . The electron contamination model was found to be suboptimal to model the dose around , especially for physical wedges at smaller source to phantom distances. For the asymmetric field verification, absolute dose difference of up to 4% were observed for the most extreme asymmetries. Compared to the SPB, the penumbra modeling is considerably improved (1%, ). At the interface between solid water and cork, profiles show a better agreement with AAA. Depth dose curves in the cork are substantially better with AAA than with SPB. Improvements are more pronounced for than for . Point dose measurements in the thoracic phantom are mostly within 5%. In general, we can conclude that, compared to SPB, AAA improves the accuracy of dose calculations. Particular progress was made with respect to the penumbra and low dose regions. In heterogeneous materials, improvements are substantial and more pronounced for high than for low energies.
The correlation evaluation of a tumor tracking system using multiple external markers
Authors: Yan Hui , Yin Fang‐Fang , Zhu Guo‐Pei , Ajlouni Munther , Kim Jae Ho
Published: November 2006

The purpose of this study is to evaluate the correlations between external markers and internal targets for radiation therapy of lung cancer patients. Using an infrared camera system coupled with a clinical simulator, the simultaneous motions of multiple external markers and an internal target were obtained. The correlation between external and internal signals was analyzed using a cross‐covariance function. A linear regression model was employed to generate a composite signal from multiple external markers in order to predict the internal target motion. The external and internal signals, and their correlations, demonstrated a wide range of variation with respect to marker location, motion dimension, and breathing pattern. The performance of the composite signal indicates that when more external signals were taken into account, the mean correlation between the composite signal and internal signal was improved. This implies that a combination of multiple external signals might be an improved way to predict internal target motion. Also, since the characteristics of respiratory signals can vary significantly, certain methods of preprocessing and external signal combination are necessary.
Tumor delineation using PET in head and neck cancers: Threshold contouring and lesion volumes
Authors: Ford Eric C. , Kinahan Paul E. , Hanlon Lorraine , Alessio Adam , Rajendran Joseph , Schwartz David L. , Phillips Mark
Published: November 2006

Tumor boundary delineation using positron emission tomography (PET) is a promising tool for radiation therapy applications. In this study we quantify the uncertainties in tumor boundary delineation as a function of the reconstruction method, smoothing, and lesion size in head and neck cancer patients using FDG‐PET images and evaluate the dosimetric impact on radiotherapy plans. FDG‐PET images were acquired for eight patients with a GE Advance PET scanner. In addition, a diameter cylindrical phantom with six FDG‐filled spheres with volumes of was imaged. PET emission scans were reconstructed with the OSEM and FBP algorithms with different smoothing parameters. PET‐based tumor regions were delineated using an automatic contouring function set at progressively higher threshold contour levels and the resulting volumes were calculated. CT‐based tumor volumes were also contoured by a physician on coregistered PET∕CT patient images. The intensity value of the threshold contour level that returns 100% of the actual volume, , was measured. We generated intensity‐modulated radiotherapy (IMRT) plans for an example head and neck patient, treating to CT‐based gross disease and to nodal regions at risk, followed by a boost to the FDG‐PET‐based tumor. The volumes of PET‐based tumors are a sensitive function of threshold contour level for all patients and phantom datasets. A 5% change in threshold contour level can translate into a 200% increase in volume. Phantom data indicate that can be set as a fraction, , of the maximum measured uptake. Fractional threshold values in the cylindrical water phantom range from 0.23 to 0.51. Both the fractional threshold and the threshold‐volume curve are dependent on lesion size, with lesions smaller than approximately displaying a more pronounced sensitivity and larger fractional threshold values. The threshold‐volume curves and fractional threshold values also depend on the reconstruction algorithm and smoothing filter with more smoothing requiring a higher fractional threshold contour level. The threshold contour level affects the tumor size, and therefore the ultimate boost dose that is achievable with IMRT. In an example head and neck IMRT plan, the D95 of the planning target volume decreased from for 42% vs 55% contour threshold levels. PET‐based tumor volumes are strongly affected by the choice of threshold level. This can have a significant dosimetric impact. The appropriate threshold level depends on lesion size and image reconstruction parameters. These effects should be carefully considered when using PET contour and∕or volume information for radiotherapy applications.
Dose‐volume delivery guided proton therapy using beam on‐line PET system
Authors: Nishio Teiji , Ogino Takashi , Nomura Kazuhiro , Uchida Hiroshi
Published: November 2006

Proton therapy is one form of radiotherapy in which the irradiation can be concentrated on a tumor using a scanned or modulated Bragg peak. Therefore, it is very important to evaluate the proton‐irradiated volume accurately. The proton‐irradiated volume can be confirmed by detection of pair annihilation gamma rays from positron emitter nuclei generated by the target nuclear fragment reaction of irradiated proton nuclei and nuclei in the irradiation target using a positron emission tomography (PET) apparatus, and dose‐volume delivery guided proton therapy (DGPT) can thereby be achieved using PET images. In the proton treatment room, a beam ON‐LINE PET system (BOLPs) was constructed so that a PET apparatus of the planar‐type with a high spatial resolution of about was mounted with the field of view covering the isocenter of the beam irradiation system. The position and intensity of activity were measured using the BOLPs immediately after the proton irradiation of a gelatinous water target containing nuclei at different proton irradiation energy levels. The change of the activity‐distribution range against the change of the physical range was observed within . The experiments of proton irradiation to a rabbit and the imaging of the activity were performed. In addition, the proton beam energy used to irradiate the rabbit was changed. When the beam condition was changed, the difference between the two images acquired from the measurement of the BOLPs was confirmed to clearly identify the proton‐irradiated volume.
Accurate calibration of a stereo‐vision system in image‐guided radiotherapy
Authors: Liu Dezhi , Li Shidong
Published: November 2006

Image‐guided radiotherapy using a three‐dimensional (3D) camera as the on‐board surface imaging system requires precise and accurate registration of the 3D surface images in the treatment machine coordinate system. Two simple calibration methods, an analytical solution as three‐point matching and a least‐squares estimation method as multipoint registration, were introduced to correlate the stereo‐vision surface imaging frame with the machine coordinate system. Both types of calibrations utilized 3D surface images of a calibration template placed on the top of the treatment couch. Image transformational parameters were derived from corresponding 3D marked points on the surface images to their given coordinates in the treatment room coordinate system. Our experimental results demonstrated that both methods had provided the desired calibration accuracy of . The multipoint registration method is more robust particularly for noisy 3D surface images. Both calibration methods have been used as our weekly QA tools for a 3D image‐guided radiotherapy system.
The Lantibiotic Mersacidin Is an Autoinducing Peptide
Authors: Schmitz Stephanie , Hoffmann Anja , Szekat Christiane , Rudd Brian , Bierbaum Gabriele
Published: November 2006

The lantibiotic (lanthionine-containing antibiotic) mersacidin is an antimicrobial peptide consisting of 20 amino acids and is produced by
Bacillus
sp. strain HIL Y-85,54728. The structural gene (
mrsA
) and the genes for producer self-protection, modification enzymes, transport proteins, and regulator proteins are organized in a 12.3-kb biosynthetic gene cluster on the chromosome of the producer strain. Mersacidin is produced in stationary phase in a synthetic medium (K. Altena, A. Guder, C. Cramer, and G. Bierbaum, Appl. Environ. Microbiol. 66:2565-2571, 2000). To investigate the influence of the alternative sigma factor H on mersacidin biosynthesis, a SigH knockout was constructed. The knockout mutant was asporogenous, and a comparison to the wild-type strain indicated no significant differences concerning mersacidin production and immunity. Characterization of the
mrsA
promoter showed that the gene is transcribed by the housekeeping sigma factor A. The biosynthesis of some lantibiotic peptides like nisin or subtilin is regulated in a cell-density-dependent manner (M. Kleerebezem, Peptides 25:1405-1414, 2004). When mersacidin was added at a concentration of 2 mg/liter to an exponentially growing culture, an earlier production of antibacterial activity against
Micrococcus luteus
ATCC 4698 in comparison to that of the control culture was observed, suggesting that mersacidin itself functions as an autoinducer. In real-time PCR experiments, the expression of
mrsA
was remarkably increased in the induced culture compared to the control. In conclusion, mersacidin is yet another lantibiotic peptide whose biosynthesis can be regulated by an autoinducing mechanism.
Antimicrobial Susceptibilities of
Aeromonas
spp. Isolated from Environmental Sources
Authors: Huddleston Jennifer R. , Zak John C. , Jeter Randall M.
Published: November 2006

Aeromonas
spp. are ubiquitous aquatic bacteria that cause serious infections in both poikilothermic and endothermic animals, including humans. Clinical isolates have shown an increasing incidence of antibiotic and antimicrobial drug resistance since the widespread use of antibiotics began. A total of 282
Aeromonas
pure cultures were isolated from both urban and rural playa lakes in the vicinity of Lubbock, Texas, and several rivers in West Texas and New Mexico. Of these, at least 104 were subsequently confirmed to be independent isolates. The 104 isolates were identified by Biolog and belonged to 11 different species. The MICs of six metals, one metalloid, five antibiotics, and two antimicrobial drugs were determined. All aeromonads were sensitive to chromate, cobalt, copper, nickel, zinc, cefuroxime, kanamycin, nalidixic acid, ofloxacin, tetracycline, and sulfamethoxazole. Low incidences of trimethoprim resistance, mercury resistance, and arsenite resistance were found. Dual resistances were found in 5 of the 104
Aeromonas
isolates. Greater numbers of resistant isolates were obtained from samples taken in March versus July 2002 and from sediment versus water. Plasmids were isolated from selected strains of the arsenite- and mercury-resistant organisms and were transformed into
Escherichia coli
XL1-Blue MRF′. Acquisition of the resistance phenotypes by the new host showed that these resistance genes were carried on the plasmids. Mercury resistance was found to be encoded on a conjugative plasmid. Despite the low incidence of resistant isolates, the six playa lakes and three rivers that were sampled in this study can be considered a reservoir for antimicrobial resistance genes.
Diversity of Basidiomycetes in Michigan Agricultural Soils
Authors: Lynch Michael D. J. , Thorn R. Greg
Published: November 2006

We analyzed the communities of soil basidiomycetes in agroecosystems that differ in tillage history at the Kellogg Biological Station Long-Term Ecological Research site near Battle Creek, Michigan. The approach combined soil DNA extraction through a bead-beating method modified to increase recovery of fungal DNA, PCR amplification with basidiomycete-specific primers, cloning and restriction fragment length polymorphism screening of mixed PCR products, and sequencing of unique clones. Much greater diversity was detected than was anticipated in this habitat on the basis of culture-based methods or surveys of fruiting bodies. With “species” defined as organisms yielding PCR products with ≥99% identity in the 5′ 650 bases of the nuclear large-subunit ribosomal DNA, 241 “species” were detected among 409 unique basidiomycete sequences recovered. Almost all major clades of basidiomycetes from basidiomycetous yeasts and other heterobasidiomycetes through polypores and euagarics (gilled mushrooms and relatives) were represented, with a majority from the latter clade. Only 24 of 241 “species” had 99% or greater sequence similarity to named reference sequences in GenBank, and several clades with multiple “species” could not be identified at the genus level by phylogenetic comparisons with named sequences. The total estimated “species” richness for this 11.2-ha site was 367 “species” of basidiomycetes. Since >99% of the study area has not been sampled, the accuracy of our diversity estimate is uncertain. Replication in time and space is required to detect additional diversity and the underlying community structure.
Rapid In Situ Evolution of Nodulating Strains for
Biserrula pelecinus
L. through Lateral Transfer of a Symbiosis Island from the Original Mesorhizobial Inoculant
Authors: Nandasena Kemanthi G. , O'Hara Graham W. , Tiwari Ravi P. , Howieson John G.
Published: November 2006

Diverse rhizobia able to nodulate
Biserrula pelecinus
evolved following in situ transfer of
nodA
and
nifH
from an inoculant to soil bacteria. Transfer of these chromosomal genes and the presence of an identical integrase gene adjacent to a Phe tRNA gene in both the inoculant and recipients indicate that there was lateral transfer of a symbiosis island.
Role of the
ptsN
Gene Product in Catabolite Repression of the
Pseudomonas putida
TOL Toluene Degradation Pathway in Chemostat Cultures
▿
Authors: Aranda-Olmedo Isabel , Marín Patricia , Ramos Juan L. , Marqués Silvia
Published: November 2006

The
Pseudomonas putida
KT2440 TOL upper pathway is repressed under nonlimiting conditions in cells growing in chemostat with succinate as a carbon source. We show that the
ptsN
gene product IIA
Ntr
participates in this repression. Crc, involved in yeast extract-dependent repression in batch cultures, did not influence expression when cells were growing in a chemostat with succinate at maximum rate.
Global Gene Expression Analysis of Yeast Cells during Sake Brewing
Authors: Wu Hong , Zheng Xiaohong , Araki Yoshio , Sahara Hiroshi , Takagi Hiroshi , Shimoi Hitoshi
Published: November 2006

During the brewing of Japanese sake,
Saccharomyces cerevisiae
cells produce a high concentration of ethanol compared with other ethanol fermentation methods. We analyzed the gene expression profiles of yeast cells during sake brewing using DNA microarray analysis. This analysis revealed some characteristics of yeast gene expression during sake brewing and provided a scaffold for a molecular level understanding of the sake brewing process.
System for Determining the Relative Fitness of Multiple Bacterial Populations without Using Selective Markers
Authors: Ahn Hyo-Jin , La Hyun-Joon , Forney Larry J.
Published: November 2006

A device for simultaneously measuring the relative fitness of multiple bacterial populations was developed and evaluated. The new device eliminates the need to construct strains with selectively neutral markers so that strains can be readily distinguished, and it provides a means to perform multispecies competition experiments.
Rapid Chromatic Detection of Bacteria by Use of a New Biomimetic Polymer Sensor
Authors: Silbert Liron , Ben Shlush Izek , Israel Elena , Porgador Angel , Kolusheva Sofiya , Jelinek Raz
Published: November 2006

We present a new platform for visual and spectroscopic detection of bacteria. The detection scheme is based on the interaction of membrane-active compounds secreted by bacteria with agar-embedded nanoparticles comprising phospholipids and the chromatic polymer polydiacetylene (PDA). We demonstrate that PDA undergoes dramatic visible blue-to-red transformations together with an intense fluorescence emission that are induced by molecules released by multiplying bacteria. The chromatic transitions are easily identified by the naked eye and can also be recorded by conventional high-throughput screening instruments. Furthermore, the color and fluorescence changes generally occur in shorter times than the visual appearance of bacterial colonies on the agar. The chromatic technology is generic and simple, does not require identification a priori of specific bacterial recognition elements, and can be applied for detection of both gram-negative and gram-positive bacteria. We demonstrate applications of the new platform for reporting on bacterial contaminations in foods and for screening for bacterial antibiotic resistance.
Lactobacillus rhamnosus
Strain GG Modulates Intestinal Absorption, Fecal Excretion, and Toxicity of Aflatoxin B
1
in Rats
Authors: Gratz S. , Täubel M. , Juvonen R. O. , Viluksela M. , Turner P. C. , Mykkänen H. , El-Nezami H.
Published: November 2006

In this study, the modulation of aflatoxin B
1
(AFB
1
) uptake in rats by administration of the probiotic
Lactobacillus rhamnosus
GG was demonstrated. Fecal AFB
1
excretion in GG-treated rats was increased via bacterial AFB
1
binding. Furthermore, AFB
1
-associated growth faltering and liver injury were alleviated with GG treatment.
Toward a More Robust Assessment of IntraspeciesDiversity, Using Fewer GeneticMarkers
Authors: Konstantinidis Konstantinos T. , Ramette Alban , Tiedje James M.
Published: November 2006

Phylogenetic
sequence analysis of single or multiple genes has dominated the study
and census of the genetic diversity among closely related bacteria. It
remains unclear, however, how the results based on a few genes in the
genome correlate with whole-genome-based relatedness and what genes (if
any) best reflect whole-genome-level relatedness and hence should be
preferentially used to economize on cost and to improve accuracy. We
show here that phylogenies of closely related organisms based on the
average nucleotide identity (ANI) of their shared genes correspond
accurately to phylogenies based on state-of-the-art analysis of their
whole-genome sequences. We use ANI to evaluate the phylogenetic
robustness of every gene in the genome and show that almost all core
genes, regardless of their functions and positions in the genome, offer
robust phylogenetic reconstruction among strains that show 80 to 95%
ANI (16S rRNA identity, >98.5%). Lack of elapsed time and, to a
lesser extent, horizontal transfer and recombination make the selection
of genes more critical for applications that target the intraspecies
level, i.e., strains that show >95% ANI according to current
standards. A much more accurate phylogeny for the
Escherichia
coli
group was obtained based on just three best-performing genes
according to our analysis compared to the concatenated alignment of
eight genes that are commonly employed for phylogenetic purposes in
this group. Our results are reproducible within the
Salmonella
,
Burkholderia
, and
Shewanella
groups and therefore are expected to have general applicability for
microevolution studies, including metagenomic
surveys.
Application of Pseudomurein Endoisopeptidase to Fluorescence In Situ Hybridization of Methanogens within the Family
Methanobacteriaceae
Authors: Nakamura Kohei , Terada Takeshi , Sekiguchi Yuji , Shinzato Naoya , Meng Xian-Ying , Enoki Miho , Kamagata Yoichi
Published: November 2006

In situ detection of methanogens within the family
Methanobacteriaceae
is sometimes known to be unsuccessful due to the difficulty in permeability of oligonucleotide probes. Pseudomurein endoisopeptidase (Pei), a lytic enzyme that specifically acts on their cell walls, was applied prior to 16S rRNA-targeting fluorescence in situ hybridization (FISH). For this purpose, pure cultured methanogens within this family,
Methanobacterium bryantii
,
Methanobrevibacter ruminantium
,
Methanosphaera stadtmanae
, and
Methanothermobacter thermautotrophicus
together with a
Methanothermobacter thermautotrophicus
-containing syntrophic acetate-oxidizing coculture, endosymbiotic
Methanobrevibacter
methanogens within an anaerobic ciliate, and an upflow anaerobic sludge blanket (UASB) granule were examined. Even without the Pei treatment,
Methanobacterium bryantii
and
Methanothermobacter thermautotrophicus
cells are relatively well hybridized with oligonucleotide probes. However, almost none of the cells of
Methanobrevibacter ruminantium
,
Methanosphaera stadtmanae
, cocultured
Methanothermobacter thermautotrophicus
, and the endosymbiotic methanogens and the cells within UASB granule were hybridized. Pei treatment was able to increase the probe hybridization ratio in every specimen, particularly in the specimen that had shown little hybridization. Interestingly, the hybridizing signal intensity of
Methanothermobacter thermautotrophicus
cells in coculture with an acetate-oxidizing H
2
-producing syntroph was significantly improved by Pei pretreatment, whereas the probe was well hybridized with the cells of pure culture of the same strain. We found that the difference is attributed to the differences in cell wall thicknesses between the two culture conditions. These results indicate that Pei treatment is effective for FISH analysis of methanogens that show impermeability to the probe.
Complete Nucleotide Sequence of an Exogenously Isolated Plasmid, pLB1, Involved in γ-Hexachlorocyclohexane Degradation
Authors: Miyazaki Ryo , Sato Yukari , Ito Michihiro , Ohtsubo Yoshiyuki , Nagata Yuji , Tsuda Masataka
Published: November 2006

The α-proteobacterial strainSphingobium japonicumUT26 utilizes a highly chlorinated pesticide, γ-hexachlorocyclohexane (γ-HCH), as a sole source of carbon and energy, and haloalkane dehalogenase LinB catalyzes the second step of γ-HCH degradation in UT26. Functional complementation of alinBmutant of UT26, UT26DB, was performed by the exogenous plasmid isolation technique using HCH-contaminated soil, leading to our successful identification of a plasmid, pLB1, carrying thelinBgene. Complete sequencing analysis of pLB1, with a size of 65,998 bp, revealed that it carries (i) 50 totally annotated coding sequences, (ii) an IS6100composite transposon containing two copies oflinB, and (iii) potential genes for replication, maintenance, and conjugative transfer with low levels of similarity to other homologues. A minireplicon assay demonstrated that a 2-kb region containing the predictedrepAgene and its upstream region of pLB1 functions as an autonomously replicating unit in UT26. Furthermore, pLB1 was conjugally transferred from UT26DB to other α-proteobacterial strains but not to any of the β- or γ-proteobacterial strains examined to date. These results suggest that this exogenously isolated novel plasmid contributes to the dissemination of at least some genes for γ-HCH degradation in the natural environment. To the best of our knowledge, this is the first detailed report of a plasmid involved in γ-HCH degradation.
Survival of
Wolbachia pipientis
in Cell-Free Medium
Authors: Rasgon Jason L. , Gamston Courtney E. , Ren Xiaoxia
Published: November 2006

Wolbachia pipientis
is an obligate intracellular bacterium found in a wide range of invertebrate taxa. While over ecological timescales
Wolbachia
infections are maintained by strict maternal inheritance, horizontal transfer events are common over evolutionary time. To be horizontally transferred between organisms,
Wolbachia
bacteria must pass through and survive an extracellular phase. We used BacLight live-dead staining, PCR, and fluorescence in situ hybridization to assess the ability for purified
Wolbachia
bacteria to survive in cell-free media. We found that purified
Wolbachia
bacteria were able to survive extracellularly for up to 1 week with no decrease in viability. While no replication was observed in the extracellular phase, purified
Wolbachia
bacteria were able to reinvade cells and establish stable infections at all time points. The ability of
Wolbachia
bacteria to survive outside host cells may increase the probability of successful horizontal transfer and the exploitation of new ecological niches. Our development of methods to purify and maintain viable
Wolbachia
bacteria from cultured cells will be useful for other researchers studying
Wolbachia
biology.
Use of a Multiple Restriction Fragment Length Polymorphism Method for Detecting Vaccine-Derived Polioviruses in Clinical Samples
Authors: Romanenkova Natalia I. , Guillot Sophie , Rozaeva Nadejda R. , Crainic Radu , Bichurina Maina A. , Delpeyroux Francis
Published: November 2006

Since the announcement of the WHO program for the global eradication of poliomyelitis and the establishment of epidemiological and virological surveillance, the emergence and circulation of pathogenic vaccine-derived polioviruses (VDPV) presenting >1% nucleotide divergence from the sequence of the original vaccine strain have been demonstrated in certain regions. We developed and used a multiple restriction fragment length polymorphism (RFLP) method to investigate the frequency of these VDPV in a population with a high level of oral poliovirus vaccine coverage in northwestern Russia. Modified RFLP profiles were found to be strongly correlated with the presence of mutations and recombination events in vaccine strains. We found that a substantial proportion of vaccine strains had high percentages of nucleotide substitutions (>0.5%), including a type 3 VDPV with 1.4% nucleotide divergence. These findings indicate that VDPV or pre-VDPV strains are not rare in certain populations with high levels of vaccine coverage. The multiple RFLP method appears to be a simple and rapid tool for monitoring such strains, which could jeopardize the benefits of the eradication program.
Molecular Epidemiology of Hospital-Acquired Vancomycin-Resistant Enterococci
Authors: Abele-Horn Marianne , Vogel Ulrich , Klare Ingo , Konstabel Carola , Trabold Rene , Kurihara Risa , Witte Wolfgang , Kreth Wolfgang , Schlegel Paul-Gerhardt , Claus Heike
Published: November 2006

Vancomycin-resistant
Enterococcus faecium
strains are a significant cause of nosocomial infections in predisposed patients. Multiple-locus variable-number tandem repeat analysis (MLVA) has been validated recently by use of a global strain collection. In this report, we applied MLVA together with multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) to type 14 isolates from three clusters of patients colonized or infected with vancomycin-resistant
Enterococcus faecium
and another 10 epidemiologically unrelated isolates from the same hospital. The clusters could be distinguished by all three typing methods, which proved to be concordant. PFGE patterns provided the highest resolution. We observed seven sequence types (ST), six MLVA types (MT), and nine distinct ST/MT combinations. The combination of MLST and MLVA may be an alternative to PFGE in hospital epidemiology, providing the benefits of high accuracy, reproducibility, and portability.
Genotyping of Varicella-Zoster Virus and the Discrimination of Oka Vaccine Strains by TaqMan Real-Time PCR
Authors: Parker S. P. , Quinlivan M. , Taha Y. , Breuer J.
Published: November 2006

Single nucleotide polymorphisms (SNPs) in five genes have been used to identify four major subtypes of wild-type varicella-zoster virus (VZV) A, B, C, and J. Additional SNPs, located in the IE62 major transactivating gene can be used to differentiate the Oka vaccine strain (vOka) from wild-type VZV. Primer-probe sets for the detection of the five polymorphic loci were designed by Applied Biosystems for the ABI 7900HT platform. Probes for each allele were labeled with VIC or 6-carboxyfluorescein fluorogenic markers. Each primer-probe set was validated to establish assay sensitivity and specificity using VZV DNA of predetermined copy number and genotype. Further evaluation was carried out using DNA samples from the vesicle fluid or skin swab of the rash of adult patients with herpes zoster or rashes due to vOka. Assay sensitivity ranged from 10 and 10
8
copies/ml of VZV DNA (equivalent to 2 to 20 copies per reaction). Statistical analyses showed that for each genotype, a set of two probes clearly differentiated the nucleotide present (allele) at that locus (
P
< 0.0001). It was possible to determine the genotype of wild-type VZV using one of four SNP assays and also to differentiate wild type from vOka using a single SNP assay. The assay can be used for diagnostic and epidemiological studies of VZV, including the differentiation of vOka from wild-type strains, investigation of breakthrough infections, and varicella outbreaks following immunization.
Usefulness of Adult Bovine Serum for
Helicobacter pylori
Culture Media
Authors: Shibayama Keigo , Nagasawa Mitsuaki , Ando Takafumi , Minami Masaaki , Wachino Jun-ichi , Suzuki Satowa , Arakawa Yoshichika
Published: November 2006

Fetal bovine serum (FBS) and adult bovine serum (BS) exhibited bactericidal activity against
Helicobacter pylori
at various levels, which were higher in BS than in FBS. The bactericidal activity was inactivated by heat treatment at 56°C for 30 min. Our results demonstrated that heat-treated BS is a useful serum source of
H. pylori
culture medium.
Lymphogranuloma Venereum Prevalence in Sweden among Men Who Have Sex with Men and Characterization ofChlamydia trachomatis ompAGenotypes
Authors: Klint Markus , Löfdahl Margareta , Ek Carolina , Airell Åsa , Berglund Torsten , Herrmann Björn
Published: November 2006

An outbreak of lymphogranuloma venereum (LGV) infections has recently been reported from The Netherlands and other European countries. The Swedish surveillance system has identified three LGV cases since 2004, all with clinically suspected infection in men who have sex with men (MSM). In order to assess the prevalence of LGV in a high-risk group of MSM and include clinically atypical cases, retrospective analysis of 197Chlamydia trachomatis-infected men was performed. Sequencing of theompAgene showed a different serotype distribution compared to recent Swedish studies in heterosexual populations. The most common types were G (45%), D (27%), and J (26%), whereas the normally predominant type E accounted for only 4% of the chlamydia cases. Furthermore, certainompAgenotype variants of the dominant serotypes were highly prevalent among MSM, and the reason for this is discussed. No additional case of LGV was detected by retrospective analysis of the high-risk MSM population. This indicates that, thus far, LGV in Sweden is only a result of sporadic import from infected MSM clusters abroad.
Fatal Granulomatous
Acanthamoeba
Encephalitis Mimicking a Stroke, Diagnosed by Correlation of Results of Sequential Magnetic Resonance Imaging, Biopsy, In Vitro Culture, Immunofluorescence Analysis, and Molecular Analysis
Authors: McKellar Mehri S. , Mehta Lahar R. , Greenlee John E. , Hale Devon C. , Booton Gregory C. , Kelly Daryl J. , Fuerst Paul A. , Sriram Rama , Visvesvara Govinda S.
Published: November 2006

Amebic infections involving the central nervous system are rare and difficult to diagnose. Magnetic resonance imaging (MRI) at timed intervals may be helpful, where scans reveal enhancing lesions and increased signal. We report a unique case of granulomatous amebic encephalitis that was proven pathologically with progressive radiological findings on MRI.
Enterotoxigenic
Escherichia coli
with STh and STp Genotypes Is Associated with Diarrhea Both in Children in Areas of Endemicity and in Travelers
Authors: Bölin Ingrid , Wiklund Gudrun , Qadri Firdausi , Torres Olga , Bourgeois A. Louis , Savarino Stephen , Svennerholm Ann-Mari
Published: November 2006

Enterotoxigenic
Escherichia coli
(ETEC) is an important cause of diarrhea among children in developing countries and in travelers to areas of ETEC endemicity. ETEC strains isolated from humans may produce a heat-labile enterotoxin (LT) and two types of the heat-stable enterotoxin STa, called STh and STp, encoded by the
estA
gene. Two commonly used assay methods for the detection of STa, the infant mouse assay or different enzyme-linked immunosorbent assays, are unable to distinguish between the two subtypes of ST. Different genotypic methods, such as DNA probes or PCR assays, may, however, allow such discrimination. Using gene probes, it has recently been reported that ETEC strains producing STp as the only enterotoxin are not associated with diarrhea. In this study, we have used highly specific PCR methods, including newly designed primers for STh together with previously described STp primers, to compare the relative distribution of STh and STp in ETEC isolated from children with diarrhea in three different geographically distinct areas, i.e., Bangladesh, Egypt, and Guatemala, and from travelers to Mexico and Guatemala. It was found that ETEC strains producing STp were as commonly isolated from cases of diarrhea as strains producing STh both in Egypt and Guatemala, whereas STp strains were considerably less common in Bangladesh. No difference was found in the relative distribution of STh and STp in ETEC strains isolated from travelers with diarrhea and from asymptomatic carriers. Irrespective of ST genotype, the disease symptoms were also similar in both children and travelers.